Marine Biological Laboratory Library Woods Hole, Mass. Presented by the estate of Dr. Herbert W. Rand Jan, 9, 1964 i-n D a a m a THE PROTOZOA Columbia Sinibersttg Biological Before the modern system of classification was established, many of 1 The term " pseudopodia" was given by von Siebold to replace Dujardin's more descriptive phrase "changeable processes" {expansions variables), c 1 8 THE PROTOZOA the forms which are now recognized as Heliozoa or Radiolaria, were variously interpreted. Ehrenberg did great service in describing the skeletons of many Radiolaria, especially of the fossil forms, but he had no conception of their organization, and placed them with the Bryozoa, Rotifera arid Echinodermata as a special class (Tubulata). Under the name, ActinopJiryens, Dujardin grouped the Heliozoa, together with a modern subdivision of the Infusoria (Siictoria), as "forms with slowly contractile appendages." The structure of the Radiolaria was first made out by Huxley ('51), who recognized them as Protozoa, and correctly compared Tkalassicolla with the heliozoon Actinosphcerium. The pseudopodia, however, were not recognized, and he was inclined to regard these forms as higher in organization than a single cell, and placed them between the Protozoa and the Sponges. Johannes Miiller('55 '58) first saw the resemblance between the fine ray-like pseudopodia of the Radiolaria and of the Heliozoa, and his pupils, Claparede and Lachmann ('58), discovered the same granule-streaming in their pseudopodia that Schultze had observed in some of the Rhizopoda. With these data, M tiller included the Radiolaria and the Heliozoa in the class Rhizopoda of von Siebold, under the name RJiizopoda radiaria, which was modified into its modern form, Radiolaria, by another of his pupils, Ernst Haeckel ('62). Four years later Haeckel ('66) separated the Radiolaria from the similar fresh-water forms, to which he gave the name Heliozoa. The further subdivisions of the subclass Rhizopoda have been made upon two bases having almost equal value. In one system they are divided according to the nature of the pseudopodia into the orders Lobosa (Amcebcza of Ehrenberg) and the Reticulariida (Reti- cnlaria of Carpenter, '62). In the other they are subdivided accord- ing to the absence or presence of a shell, into the orders Amcebida (Ehbg.) and Testacea (M. Schultze, '54). 1 The former system is adopted by Delage and Herouard, by Lankester and the English zoologists generally ; the latter by Butschli. A third order under the name Mycetozoida, is usually included with the Amcebida and the Reticulariida. Although generally recognized in part at least, by zoologists as Protozoa, the taxonomic position of the organisms included in this order is in dispute. Under the name Myxomycetes they are included with the fungi by most botanists, while by the zool- ogists they are usually placed as a class of the Rhizopoda under the name Mycetozoa (de Bary, '59). The relation to the fungi is claimed on account of their saprophytic mode of life (terrestrial forms), and their mode of spore-formation in sporangia which are often compli- cated by the presence of stalks, columellae, and other plant-like 1 Thalamophora, R. Hertwig ('74); Foraminifera, d'Orbigny ('26). INTR OD UCTION structures such as elastic capillitia for dispersion of the spores. The relation to the Protozoa, on the other hand, is claimed on account of the unicellular nature, development of the swarm-spores, and occa- sional holozoic mode of nutrition. The spores leave the sporan- gium as amoeboid or flagellated organisms and may increase by simple division during the swarming stage. If flagellated, the spore after a time loses the flagellum and becomes amoeboid, in which con- dition division may again occur; finally numerous amoeboid indi- A Fig. 4. Flagellidia. [STEIN.] A. Chrysomonas (Chromulina) fiavicans. Ehr. with chromatophores and an engulfed diatom (d). B. The same encysted. C. Phacus longicaudus Duj. viduals group themselves together, forming a colony or plasmodium. In some cases the fusion is complete, in others the outlines of the individual Amoebae persist. In view of the questionable position which these forms occupy, there is some danger of their being neglected altogether, the botan- ists refusing them because of their animal characteristics, the zoolo- gists because of their plant-like features. No harm can be done by including them in both kingdoms, for on purely a priori grounds it is to be expected that some organisms should be on the boundary line between artificial groups such as the unicellular animals and plants. The present group and the Phytoflagellida among the Mastigophora appear to occupy such a position, and it is advisable to include them as provisional groups of the organisms with which they show the greatest number of common points. With our present knowledge, 20 THE PROTOZOA the majority of Mycetozoa undoubtedly resemble fungi more than they do Protozoa, and will not be further considered in the present work ; the Phytoflagellina have, on the other hand, so many obvious connec- tions with the animal flagellates that they cannot well be omitted. Fig. 5. Dinoflagellidia. [SCHUTT.] A. Gymnodinium ovum, Schiitt. B. Peridinium divergens Ehr. f, the transverse furrow. The flagellated organisms now included under Diesing's name, Mastigophora, fall naturally into three subclasses : (i) the Flagellidia (Fig. 4) (flagellates in a strict sense), recognized by Dujardin and n Fig. 6. Coccidiida in epithelial cells. [LABBE.] The coccidium, a species of the genus Myxinia, is supposed to have divided in one case (to the right), c, the sporozoon; n, the nucleus of an epithelial cell. named by Cohn ('53); (2) Dinoflagellidia (Biitschli) (Fig. 5), which were first seen by O. F. Miiller (1/73) and later fairly well described INTR OD UC TION 21 by Ehrenberg, but curiously misinterpreted as ciliated forms (a mis- take rectified only during the last twenty years), which led Clapa- rede and Lachmann ('58), R. S. Bergh ('84), and Saville Kent ('81) to regard these organisms, under the name Cilio-flagellata, as inter- mediate forms between the Ciliata and the Mastigophora; (3) Cysto- flagellidia (Haeckel), including two genera, Noctiluca and Leptodiscus, the former observed during the eighteenth century, the latter dis- covered by R. Hertwig ('77). Fig. 7. Two forms assumed by Leptotheca agilis, a myxospore. [DOFLEIN.] The history of the Sporozoa as a class dates from Kolliker's ('45- '48) and Stein's ('48) works, although the name Gregarine now used as the title of an order (Gregarinida) goes back to Leon Dufour ('28), and the first observation to Redi in the seventeenth century. 1 The different kinds of Sporozoa were first grouped together by Leuckart ('79) under the present name, and he subdivided the group into the Gregarinida (Fig. I, D) and the Coccidiida (Fig. 6), the former dwelling in cavities of various invertebrate hosts, the latter inside epithelial cells in, chiefly, vertebrate hosts. Under the term psorosperms (Joh. Miiller, '41), a number of fish parasites belong- ing to the Sporozoa were known early in the century, and these were grouped together by Biitschli under the term Myxospo- 1 Cf. Diesing, p. 183. 22 THE PROTOZOA ridia(Y'\g. 7), and in the present classification form a fourth order of the Sporozoa. A third order under the name H&mosporidiida ( Labbe, '94), includes the sporozoan parasites dwelling in blood-cells and plasm of different vertebrates (Fig. 8). The Infusoria finally have been variously classified since von Siebold restricted the term as given by Ledenmuller (i76o-'63) to its modern significance. Until their unicellular structure was defi- nitely established, the various types were placed among the higher animals, sometimes with the worms, sometimes with the Coelenterata. Even Perty ('52), who was the first to bring the ciliated forms together under their present name Ciliata, believed them to be combinations of Fig. 8. A blood parasite or Haemospore, Plasmodiummalarice. Amoeboid, spore-forming and sexual phases are shown. [WAS1ELEWSKY.] cells. He included the Suctoria and the Ciliata as subdivisions of the Infusoria. Stein ('57), who put the classification of the Ciliata upon its final modern basis, had previously confused the relations of Suctoria and Ciliata in his Acineta-theory. Claparede and Lachmann '('58-'6i ), after showing that Stein's interpretation was incorrect, raised those Infusoria which are provided with suctorial tentacles and are ciliated only during the embryonic phases, to the grade of a separate subclass to which they gave the modern name Suctoria (Fig. i, c, Ey Since Stein's work there have been but few important changes in the classification of the Infusoria. Biitschli ('83-'88) divided the Ciliata into two unequal groups distinguished by the nature of the mouth parts, the Gymnostomata and the Tricliostomata. Stein's sub- divisions based upon, the arrangement of the cilia are more simple, however, and the advantage of Biitschli's division is somewhat ques- tionable. C ANIMALS AND PLANTS In determining the boundaries of the subkingdom Protozoa, two very interesting controversies have arisen, one relating to the boun- dary between animals and plants, the other to the relations of Protozoa to Metazoa. The modern attitude toward the first of these problems is well expressed by Delage ('96), who says : "The question is not so important as it appears. From one point of view, and on purely !Cf. Stein, II ('67), p. 142. INTRODUCTION 23 theoretical grounds, it does not exist, while from another standpoint it is insoluble. If one be asked to divide living things into two distinct groups of which the one contains only animals, the other only plants, the question is meaningless, for plants and animals are concepts which have no objective reality, and in nature there are only indi- viduals. If, in considering those forms which we regard as true . animals and plants, we look for their phylogenetic history, and decide to place all of their allies in one or the other group, we are sure to reach no result ; such attempts have always been fruitless"." 1 No one at the present time denies the extremely close relation which Huxley ('76) has so clearly pointed out between the lower algae and some of the flagellates, and it is the general opinion that no one feature separates the lowest plants from the lowest animals, and the difficulty in many cases the impossibility of distinguishing between them is clearly recognized. Curiously enough, this modern idea was early expressed by Buffon at the time when Aristotle's view of the plant-like nature of some animals (Zoophyta) was still accepted in regard to the Coelenterata. Buffon wrote as follows in 1749: " From this investigation we are led to conclude that there is no absolute and essential distinction between the animal and vegetable kingdoms; but that Nature proceeds from the most perfect to the most imperfect animal, and from that to the vegetable." This state- ment might have been written in 1899, but Buffon unfortunately goes on to say : " Hence the fresh-water polypus (Hydra} may be regarded as the last of animals and the first of plants." 2 Ehrenberg included a large number of plant forms among his Infusoria, most of which Dujardin threw out, restricting the group, practically, to the Protozoa as known to-day. But the discovery of flagellated swarm-spores of algae cast doubt on the animal nature of the organisms which Dujardin had described as flagellates. Von Siebold ('45) was thus led to retain only the families Astasiidae and the Peridinidae in his zoology, removing the Mastigophora, as a group, to the botanical side. In this he was followed by Bergmann and Leuckart ('56), while Cienkowsky ('65) placed them as an intermediate group between animals and plants. Others went to the opposite extreme and actually excluded the algal swarm-spores from the plants, on the ground that they were merely flagellated parasites living on the plant-cells (Diesing, '65). Still others, noting that some of the flagel- lates are animal and some vegetable in their nature, undertook the impossible task of finding a single distinguishing character. The presence of green coloring matter or chlorophyl, upheld by Cohn ('76) and others as a characteristic vegetable feature, seemed to be a good 1 P- 5 J 8. 2 Edition 1812, p. 357. 24 THE PROTOZOA test; Oersted ('73), however, showed that in the lower plants there are forms differing only in the presence or absence of chlorophyl. These forms may be arranged in a series as follows : 1 With chlorophyl Without chlorophyl With chlorophyl Without chlorophyl Oscillaria. Beggiatoa. Spirulina. Spirochaeta. Leptothrix. Leptomitus. Palmellaceae. Chroococcaceae. Chlamydomonas. Chlamydomonas hyalina. Synedra. Synedra putrida. A similar series can be arranged among the Protozoa, including forms which cannot be genetically separated, though some contain chlorophyl, and some are colorless. In the first of these, nutrition is holopJiytic or of the green plant type, in the second saprophytic or of the fungus type. The chlorophyl differential, if used here, would separate closely allied and in other respects identical forms, always to be found among the Mastigophora, and would lead to confusion. Furthermore, the chlorophyl differential would cause confusion in the classification of the fungi, where colorless representatives of several families of the Phycomycetes reproduce by colorless swarm-spores. Again, some of the Mastigophora with chlorophyl are not dependent upon this substance for their nutriment, but may combine the plant type with the animal type of food-getting (e.g. Chromulina and some Dinoflagellidia, Fig. 4). Stein sought a differential in the presence of contractile vacuoles and of nuclei, which, he maintained, are not found in vegetable swarm- spores, but are characteristic of all animal cells. This view has not been supported by later discoveries, for not only have vegetable -spores been found to possess nuclei, but many of them are also provided with contractile vacuoles. Haeckel bases the classification of animals and plants upon nutri- tion, which differs but little from the earlier chlorophyl differential. All forms with the power of absorbing carbon dioxide, water, and nitrogen compounds, and of combining them into proteids, he calls plants, those without this power, animals, but he considers that this division, though logical, is at best only artificial, and gives no clue to the actual phylogenetic relations of Protozoa and Protophyta. As a single differential, however, the method of nutrition is probably as satisfac- tory as any, for there are only a few forms which combine the two modes of food-getting. If rigorously applied, however, it cannot fail to shock the prejudices of both botanists and zoologists in claiming for the animal kingdom forms which have usually been identified with the vegetable kingdom, and vice versa. Although Haeckel states that the dividing line is purely arbitrary and does not represent genetic affinity iSee Entz ('88). INTRODUCTION 2$ in the least, animal forms being derived from plants in a polyphyletic series, he does not hesitate to rank certain of the fungi, together with the Sporozoa and bacteria, as animal forms ; the majority of chlorophyl-bearing Protozoa, on the other hand, are placed with the plants. Another differential, which, perhaps, has been the most widely accepted, is the power of spontaneous motion. It is supported to-day *.',^ , Fig. 9. A sphaeroidal colony, Uroglena americana Calkins, consisting of monads embed- ded in a gelatinous matrix. as the most universal of the arbitrary differentials by Biitschli, Bergh, and Delage. Briefly stated, all forms, which are freely-motile in their adult life, are animals, while stationary forms are plants. This distinction is applied only to the lower forms, and not to the higher groups, but even as thus limited, this differential would neces- sitate some striking changes in existing schemes of classification. The freely-moving diatoms, which, since the time of Nitsch ('38) have been classed with the unicellular plants, would be included among the Protozoa, while the majority of Sporozoa, which are almost devoid of motion, would be excluded. The point of view which demands the strict separation of animals and plants has, however, little utility save perhaps to determine the limits of a text-book or monograph. Many observers, recognizing this truth, have included all forms in which the transition from plants to animals is shown, in a special group of the Protozoa, and usually with some heading which gives a clue to their position. This is first seen in Aristotle's Zoophyta (Coelenterata) ; again in a more modern form in Perty's Phytomastigoda t and in the Phytofiagellida of Delage. Haeckel ('66) made a group of equivocal forms large enough to include all of the Protozoa, and, under the name Protista, vainly attempted to establish a third kingdom between the animal and the plant. 26 THE PROTOZOA The arbitrary dividing line between the Metazoa and the Protozoa can be much more sharply drawn than that between animals and plants. The Protozoa are usually defined as single-celled animals, the Metazoa as many-celled ; but this definition is not strictly accurate, for many forms of Protozoa live in aggregates, or colonies in which specialization and division of labor have progressed to a considerable degree ( Volvox, Uroglcna, Magosphcera, etc. ; Fig. 9). As a rule, however, colonies do not form a distinct tissue of cells as in the bias- tula stage of Metazoa, while a still stronger point is that they never form a diblastic embryo. 1 D. GENERATION DE NOVO Leeuwenhoek's discovery of the Protozoa had a marked effect upon current thought, some speculative writers seeing in these minute organisms the hypothetical units of organic structure, which, from the time of Democritus to that of Descartes, had been a subject of philosophical discussion. The rapid and incomprehensible increase of Protozoa in standing water could apparently best be explained by a theory of spontaneous generation ; Leeuwenhoek, nevertheless, was convinced that their origin would be found in minute eggs or germs which are carried through the air as dust, or brought from place to place by birds, etc., thus showing his firm belief in Harvey's axiom, ex ovo omnia. He was supported in this view by Joblot (1718), whose experiments led him to the conclusion that the lower stratum of the air is filled with the germs of various kinds of animalcula, while Reaumur (1738) asserted that the dust of the air also contains dis- ease germs, which are the cause of epidemics. These men were, however, in the minority, and until the last fifty years only an occa- sional observer opposed the theory of spontaneous generation, as applied to these minute organisms. Even in Leeuwenhoek's time it was well known that dead organic matter of any kind, when left exposed in water, gradually decom- poses, while the water, at first clear, becomes murky, and minute organisms of various kinds develop in it. Adopting the view that higher organisms are composed of organic units, speculative writers inferred that the small animals discovered by Leeuwenhoek were the units which had again become freed from the aggregated condi- tion. This is the key-note of Buffon's (1749) famous theory of gen- eration, which, in one form or other, persisted well into the I9th century. Briefly stated, Buffon believed that all organisms are composed of an infinite number of organic particles. The In- 1 See Saville-Kent ('8l) for the obsolete theory that Sponges are colonial Protozoa. INTRODUCTION 2J fusoria he believed to be nothing but these particles become free. " The destruction of organized bodies is only a separation of the organic particles of which they are composed. These particles continue separate till they be again united by some active power. When, however, a man's body has nearly attained its full size, he does not require the same quantity of organic particles ; the surplus is, therefore, sent from all parts into reservoirs destined for their recep- tion. These reservoirs are the testes and seminal reservoirs " (page 397). " The different parts of the body are, however, built up of dif- ferent kinds of organic units, so that upon disintegration there are different forms of animalcula, which are in no respect different from the spermatozoa of the same animal. The freed units are therefore neither animals nor plants, but the formative elements of both. Arising as the disintegrated parts of dead organisms, or rather as elements which never die, they are organisms which pass from one living state into another." This view was carried further by Need- ham (1748), and as the Buffon-Needham hypothesis, was generally accepted. Thus the early advocates of the theory of spontaneous gen- eration did not maintain that living things arise from not-living sub- stances, but that all organisms are derived from parts of those living before, a sort of transmigration. Spallanzani, however, to whom so much credit is due for our early knowledge of the Protozoa, adhered to the view of Leeuwenhoek that -the Infusoria are not the units which constitute higher organisms, but distinct forms of life which, like other organisms, are derived from definite germs. Furthermore, he vigor- ously upheld their animal nature against Buffon and his school, basing his arguments upon their voluntary movements, changes of direction when moving, food taking, and upon their relations to moisture and dryness, warmth and cold, to which they reacted like higher animals. He found that Infusoria do not develop in a vacuum, and must, therefore, come from germs contained in the air. Seeing a Colpoda emerge from its cyst, he concluded that the cysts were eggs, mistaking the cyst-case for the egg-membrane. He separated the large from the small forms of Infusoria, a separation which was the first attempt to distinguish the Protozoa from bacteria, and which was destined to have great effect upon the theory of spontaneous generation, for it is a significant fact that the forms which have been supposed to arise by spontaneous generation have always been those approaching the limits of vision. 1 1 Spallanzani's work has hardly been sufficiently recognized by later writers. Never car- ried away by enthusiasm, but describing only what he saw, he placed himself outside the current of popular favor by opposing the tempting hypothesis of the nature-philosophers. He seems to have combined his power of observation with a remarkable breadth of view, which in some cases gave rise to daring conceptions. Thus, in 1776, he wrote : " Pour des 28 THE PROTOZOA The distinction which Spallanzani drew between large and small forms was also adopted by O. F. M tiller in his classification of the Protozoa. The latter maintained, however, that the lower of his two groups (Infusoria) were formed according to Buffon's view, from the disintegrated parts of higher organisms. These parts, after the disintegration, collect, forming a slimy scum on the surface of the in- fusion (Zooglcea), which formed a most important adjunct in all subsequent theories of spontaneous generation. Minute vesicles arise later from this scum, and these remain as living organisms in the form of " Infusoria," which included bacteria, spermatozoa, and the smallest forms of flagellates. This mode of origin he limited to those Protozoa which have no visible organs or means of locomotion. The other group (Bullaria), which included the larger of the animalcula (worms, ciliated Protozoa, rotifers, etc.), he maintained were formed, as in the higher animals, from eggs. Miiller also held that these units mix with the inorganic particles to form the solid and fluid portions of the body of higher forms, while alone, and without contact with foreign matter, they form the nerves and " soul." Oken (1805), another advocate of the theory of spontaneous gen- eration, held that all Protozoa arose in a similar manner. Since all plants and animals were built up of these Infusoria, he named the latter Urthiere, although, like Buffon, he held that they were neither plants nor animals. Infusions demonstrated to him that all plants and animals could disintegrate into Infusoria. Small Infusoria at first joined together to form larger ones, and out of their union arose the polyps and higher forms. While the outline of Oken's view would seem to indicate a prophecy of the cell-theory, it is quite evident from his book on creation that he had little real conception of what we now regard as the essence of that theory. 1 The majority of contemporary naturalists followed Buffon and Oken, either absolutely or with slight modifications. Among these Treviranus ('03), Goldfuss ('20), and Carus ('23) accepted Oken's views, while Lamarck ('15), Blainville ('22), and Bory de St. Vin- cent ('24) followed Miiller in restricting spontaneous generation to the simpler and smaller forms. Dallinger and Drysdale ('73 '75), taking turns over the microscope by day and night, followed out the life-histories of many of these simpler forms through the process of division and spore-formation, thus showing that the monads arise, as do the higher Protozoa, from animaux inferieurs, le changement de demeure, de dimat, de nourriture, doit produire pen a peu dans les individus, et ensuite dans Vespece, des modifications tres considerables qui deguiseni a nos yeux les formes primitives' 1 '' (cited by Dujardin ('41) from Spallanzani). 1 The name "Protozoa," given by Goldfuss ('20), meant the same as Oken's " Urthiere." It did not acquire its present significance until 1845, when von Siebold gave it a new meaning. INTRODUCTION 29 ancestors similar to themselves. They found that the spores which burst from the encysted forms were at first far beyond the limits of vision even with the high powers of the microscope at their command, but remained together in the form of a "glairy" mass in which minute specks soon appeared, and these specks were watched until they had become full-grown monads similar to the original form. In later years the theory of spontaneous generation has been limited almost exclusively to the bacteria, but even here it has been energeti- cally and successfully opposed by Pasteur, Tyndall, Milne-Edwards, Claude-Bernard, Quatrefages, and others, against a constantly de- creasing number of advocates. No one is in a position to assert, however, that it does not take place in some organisms, although such a view is highly improbable; nor can it be maintained that it never has taken place in the past. Many theories of "archigony " (Haeckel), or the first origin of life by spontaneous generation, have been held by modern naturalists ; but all such theories are of a purely inferential character and lack substantial foundation. Without attempting to discuss these l it may be pointed out that the eminent botanist Nageli has advocated an hypothesis which suggests that of Buffon. Assum- ing that protoplasm consists of minute structural units or " micellae," he suggests that such micellae were first formed from not-living matter and secondarily united into organisms. Nageli does not hesitate to say that the evolution of the simplest protozoon from inorganic compounds involved a far greater step than from the first organism to man, and in accordance with this idea Haeckel places the beginning of life in the oldest known geologic age and in the oldest period of that age, the Laurentian. This, again, is entirely specula- tive ; for if we except the questionable form Eozoon, the rocks of the Laurentian contain no recognizable records of past life. 2 The rocks of the period after the Laurentian, however, the Cam- brian, possess a great number of well-marked types, families, and genera, thus indicating, even at this time, a considerable antiquity. Haeckel and Nageli argue with Huxley, and the argument is of great 1 For a discussion of this topic the reader is referred to the essays of Huxley, Tyndall, and Haeckel, and to Verworn's Allgemeine Physiologic, pp. 298-319. Lee's translation, pp. 297-319. 2 The supposed genus Eozoon (" dawn of life ") was discovered by Logan, of the Geologic Survey of Canada in 1865, and the name was given by Dawson in the same year. There has been a lengthy dispute, however, in regard to this supposed fossil, some asserting that it is the earliest known foraminiferon, others that it is entirely inorganic. The former opinion was held by Carpenter ('65, '66, etc.) and Dawson ('65, '75, etc.). the latter by the majority of geologists and petrologists, beginning with King and Rowney ('66) and followed by Mobius and others. Biitschli, while admitting that Dawson and Carpenter had a certain amount of evidence, inclines to the opposite view, while petrologists maintain that the same structure as that of Eozoon has been frequently observed in minerals forming parts of rocks of undoubted igneous origin. 3 Fig. 13, C). Here the chitin which forms the shell is perfectly smooth ; but in other forms it may be ornamented in various ways by pits or pro- tuberances. Again, in many fresh-water Rhizopoda the shell-material is not secreted, but the test is composed of foreign particles, such as diatom shells, sand crystals, mud, or detritus of any kind, fused together and to a chitinous substratum by means of mucilaginous cement secreted by the organism. 3. Nuclei. Haeckel's claim ('68) that there are organisms without nuclei (Monera), although it rests upon negative evidence, cannot be rejected until all of the forms considered have been shown to possess them. On purely a priori grounds, it is possible to conceive such organisms, although the numerous experiments which have been performed dur- ing the last decade upon nucleated and non-nucleated parts of Pro- tozoa, show, in these cases at least, the absolute necessity of the nucleus for the life of the organism. These experiments make it probable that the so-called Monera have in reality some structure or structures which perform the functions of the nucleus, although a well-defined nucleus with membrane and other characteristic parts may be absent. In the majority of Protozoa there is but one nucleus (many Sar- codina, Mastigophora, Sporozoa), while in some forms two nuclei are the rule (some Rhizopoda). In others, again, there may be a great number of nuclei, the number varying with the age of the organism (examples occur in all groups of the Protozoa). In many of the Pro- tozoa, although not in all, the nucleus is provided with a membrane and contains two substances ; chroma tin, staining with certain basic dyes and consisting largely of nucleinic acid, and achromatin, a sub- stance which is not stained by the chromatin dyes, in the form of a 1 CL Schewiakoff ('88). Fig. 13. Shells and tests. [A, SCHEWIAKOFF; B, ORIGINAL; C, BUTSCHLI; A STEIN.] A. Euglypha alveolata Duj. The shell consists of oval siliceous plates glued together by a sili- ceous (?) cement. B. Cochliopodium digitatum, n. sp. The test is membranous and perforated for pseudopodia. C. Pseudochlamys patella Clp. and Lach. The test is membranous and shield- like. D. Ceratium tripos Nitsch. The shell consists of cellulose plates of diverse size and shape. THE PROTOZOA network, or of a homogeneous body of considerable size (Karyo- somes). Several different types of nuclei may be distinguished; some of the most important being : ( i ) The distributed nucleus, n N. ----- X E Fig. 14. Types of nuclei. [A. Calcituba polymorpha Roboz, from SCH AUDINN ; B. Colpidium colpoda, from a preparation ; C. Euglena viridis Ehr. from a preparation ; D. Tetramitus chilomo- nas, n. sp. ; E. Noctiluca miliaris Sur., from a preparation.] A single karyosome (A) becomes vesicular, and ultimately gives rise to several daughter-karyo- somes (so-called "fragmentation" Schaudinn). Several karyosomes in Noctiluca (E) hold the chromatin, the rest of the nucleus is filled with "achromatic " granules. In Tetramitus chilomonas (D) the chromatin is scattered throughout the cell; the lighter-colored body in the centre of the cell is the homologue of the deeply stained central body in Euglena (C). GENERAL SKETCH 43 consisting of innumerable chromatin granules distributed throughout the cell ( Trachelocerca, Chcenia teres, Holosticka flarta, H. scutellum, Tetramitus}. (2) The homogeneous nucletts consisting of a single mass of chromatin with a homogeneous structure throughout all stages, and with no trace of reticular substance (many Phytoflagel- lates). (3) Dimorphic nuclei, consisting of a large nucleus called the macronucleus, and a small one, the micronucleus, in the same individual. The former is generally regarded as functional chiefly in vegetation, the latter in conjugation. With the exception of Polykrikos among the Mastigophora, dimorphic nuclei are found only in the Infusoria (Fig. 14). The typical form of the nucleus is spherical, although it may be discoid or ellipsoid, or, in the case of the macronucleus, drawn out into various fantastic shapes, of which the horseshoe (Vorticellidae), the beaded (Stentor and Spirostomum, etc.), or branched (Acineta, Dendrosoma] are examples. 1 4. Organs of Locomotion. With very few exceptions, the Protozoa have the power of moving from place to place. The exceptions are found among the para- sitic Sporozoa, although even here there is, in some cases, a peculiar gliding motion. In no adult sporozoon is there a special organ of locomotion, yet the Gregarinida and Haemosporidiida actually move from place to place, although very slowly. In some cases, the motion is due to peculiar peristaltic waves of contraction ; in other cases to the contraction of muscle-like fibrils, the myonemes. An analogous movement is also known in certain flagellates (Euglenidae) and ciliates (Heterotrichida). In the majority of Protozoa, however, movement is accomplished by the activity of special motor organs, which may be either changeable processes, (pseudopodia) or permanent vibratile appendages (flagella and cilia). The changeable processes or pseudopodia, found chiefly in the Sar- codina, are sometimes numerous, sometimes few ; when few in number they are usually short, finger-formed, and quick to change in form and appearance by the flowing protoplasmic substance of which they are composed (Fig. I, A, and Fig. 15, A, B}; when numerous, they are fine-pointed, and often sticky, so that when two or more come in contact, they fuse or anastomose (Reticulariida, Fig. 15, C). Again, the pseudopodia may be fine and pointed, but rigid in structure and unchanging in form, a condition brought about by the presence of an axial filament of stiffened protoplasm, which runs down the centre of each pseudopodium (Heliozoa, Radiolaria, Fig. 15, D\ Unlike pseu- 1 Cf. Chapter VII. for further details concerning nuclei. 44 THE PROTOZOA dopodia, the protoplasmic filaments, known as flagella and cilia, are derived solely from the ectoplasm and are constant in their position, and, save for the occasional absorption within the body, for some rea- son or other, they are unchangeable. Flagella-motion, characterized by energetic contractions or undulations, or by rotary motions, differs -**-* X" Pig. 15. Types of pseudopodia. A. AmxialimicoIaRhmb. [RHUMBLER.] B, Amoeba blatt vac uole with external pore; N, macro- aboilt with the endoplasmiC nucleus ;, micronucleus. flow until, becoming heavier than the protoplasm, it remains stationary, while the rest of the endoplasm moves forward with the organism (many Rhizopoda). In this manner the vacuole, as it attains its full size, is gradually left at the posterior end of the moving organism, where it finally bursts. Again, as in some Mastigophora and Ciliata, the contractile vacuole is a stationary organ connected with the out- side by a definite pore. Here, too, are numerous accessory structures in the form of canals and reservoirs, the former apparently collecting the water and waste matters from all parts of the cell, and conducting them to the contractile vesicle, the latter receiving the fluid after con- traction of the vacuole, and conveying it to the outside, with which 54 THE PROTOZOA they are in open connection. The canal system, which some observers (e.g. Fabre Dumergue, '90) consider widely spread throughout the Ciliata, is often strikingly developed, as in Frontonia, where there is a complicated network traversing the entire cell (Fig. 21). While the excretory function of the contractile vacuole is generally accepted, there have been only a few satisfactory experiments to demonstrate it, and the possibility of other functions is not excluded. At the present time the balance of evidence is in favor of the view that the contractile vacuole has both excretory and respiratory func- tions, inasmuch as it regularly empties a fluid to the outside, which carries with it the products of destructive metabolism in the form of urea, and probably carbon dioxid, although the respiratory function has never been demonstrated. 1 Whatever may be the function of the contractile vacuole, it does not appear to be universally necessary for the life of the organism, for it is lacking in the Sporozoa and the majority of the Sarcodina (Reticulariida and Radiolaria). Furthermore, whatever the use of the vacuole, it is independent of the nucleus, non-nucleated fragments form- ing new vacuoles which pulsate rhyth- mically for some time. Hofer ('89) found that vacuoles in non-nucleated bits of Amceba proteus would contract for fourteen days. He also noted that whereas the regular period of pulsation c '*^tf^*^~%jS=*^ was seven minutes, the periods became J\ ^^W^^&K-^^ longer and longer, until at the end of the fourth day there was but one pulsation every two hours, and even then the con- tents were not completely expelled, a reaction which Penard ('90) formulated later by the statement that the activity of the contractile vacuole is directly proportional to that of the entire indi- vidual. Fig. 22. Division of Euplotes. [FROM A PREPARATION.] The daughter-cells are almost ready to separate; the daughter-micronuclei () are re-formed; the macronucleus (m) is not quite divided; the gastric vacuoles (/) are equally distributed in the two daughter-cells, one of which has generated the adoral zone (az). 3 . R eproduction . With the exception of the Sporozoa, simple division, or splitting into two parts, is the characteristic mode of re- production in all Protozoa. In the Sporozoa, and at times in most of the other Protozoa, division is replaced by Chapter IX, p. 28 J. GENERAL SKETCH 55 spore-formation or the breaking up of the body into many small particles, each the germ of a new organism. While the major- ity of the Protozoa reproduce asexually in these ways, reproduc- tion in some is bound up with complete sexual differentiation, and a series of forms may be selected which indicate the probable develop- ment of the sexual from the more primitive methods. In numerous cases the sexual phenomena include many of the preliminary matura- tion stages shown by the Metazoa, in the formation of polar bodies and reduction of the quantity of chromatin, etc. Simple division, the most common method of reproduction, is usually a separation of the body into two equal parts either longitu- A BCD Fig. 23. Division (budding) of Euglypha alveolata Duj. [SCHEWIAKOFF.] The shell-plates which were stored in the endoplasm about the nucleus pass out with the stream- ing protoplasm (A) to form the shell of the daughter-cell. The nucleus is shown in different stages of mitosis. dinally (Flagellidia) or transversely (Ciliata). It is invariably preceded by division of the nucleus, and is often accompanied by the equal division of certain of the internal structures of the cell, such as the chromatophores, pyrenoids, etc. It may take place either during active life or under the protection of a cyst. Ciliata in the process of division may be frequently seen swimming about actively, the con- necting-strand becoming narrower and narrower, until finally only a delicate strand of protoplasm separates the daughter-cells, and this, after a few energetic contortions, gives way and the young cells are THE PROTOZOA free (Fig. 22). The presence of a firm shell or coating complicates the process, especially if the shell is a secretion (Fig. 23). In many ./ Fig. 24. Microgromia sociahs Hert., a gregaloid colony. [HERTWIG.] cases the new shell-parts are secreted before the act of division be- gins, and when the protoplasm buds out of the original shell-mouth, they are carried with it, and the bud is covered with the newly formed A K iA*LMA/4 l X^ / v .-'" *V*V -^ & "'."' "ft/*?:-: A .'' -kM ^ftl^^ ^^^^:%^\^. ''.o^-jZ&r C) OY 0. ' o . *' t> ^-> '^* : fc* : " 'fe * S 'C- t: '!, * O ( Fig. 25. Uroglena americana Calkins, a sphaeroid colony. pieces, which are glued together by means of a chitinous or silicious secretion. GENERAL SKETCH 57 Simple division frequently leads to colony-formation through incom- plete separation of the daughter-individuals. Four general types of these colonies are met with among the Protozoa. Adopting Haeckel's terms, they may be designated according to their general structure as (i) gregaloid, (2) sphceroid, (3) arboroid, and (4) catenoid. A gregaloid colony is an aggregate of Protozoa having a round, ellipsoidal, or indefinite shape, and usually with a gelatinous basis in which the single individuals are variously distributed. The colonies may be formed by incomplete division of the individuals or by partial union of two or more adults (Fig. 24). A spheroid colony is a globu- Fig. 26. Codosiga cymosa Sav. K., an arboroid colony of Choanoflagellida. [KENT.] lar, ellipsoidal, or cylindrical aggregate in which the individual cells form a superficial layer in a common gelatinous matrix. When these superficial cells are closely packed together into an almost continuous layer as in Volvox^ Magosphcera, or Uroglena, they are extremely sug- gestive of certain stages in developing Metazoa (Fig. 25). An arbo- roid colony is a tree- or bush-like aggregate arising by the dendritic or dichotomous branching of a primary stalk or a gelatinous matrix. Such colonies are usually attached by the base to some foreign object and often resemble hydroids or Bryozoa (Fig. 26). They may, how- ever, as in Dinobryon, be free-swimming. A catenoid colony is fili- form or chain-shaped, arising from the union of cells end to end, or THE PROTOZOA side to side, or through the continuous division of the cells in one plane (Fig. 27). Many intermediate stages between simple binary division and spore- formation show that the latter method of reproduction was probably derived from the former. When simple division takes place within Fig. 27. Eiermocystis polymorpha Leg., a cate- noid colony of Gregarin- ida. [WASIELEWSKY.] Fig. 28. Exogenous budding in Ephelota Butschliana Ishik. [FROM A PREPARATION.] The macronucleus (N) is continued into the four daughter- cells, which appeared first as minute buds. a cyst, it not unfrequently happens that each of the daughter-cells divides again, thus forming four daughter-cells within the cyst (some Mastigophora and Sporozoa). Again, as many as eight or sixteen may be formed in the same way, and from this it is not a great step to the method of reproduction by spore-formation, where a great number of young individuals are produced at one time. Another modification of simple division is the process of budding GENERAL SKETCH 59 or gemmation which is common among the Suctoria and some of the Flagellidia, less frequently observed in the Ciliata, and is possibly allied to " spontaneous division " among Sporozoa. 1 A piece of the nucleus of the mother-animal is pinched off and becomes the nucleus of a much smaller daughter-cell, which usually arises from a certain definite place on the parent (Fig. 28). When numerous pieces are thus budded off, and each piece is surrounded by a bit of protoplasm, the process is again akin to spore-formation (Noctilnca). Spore-formation may thus be accomplished either by the repeated Fig. 29. Onychodromus grandis Stein. [MAUPAS.] A. Normal individual. B. Smaller form without micronuclei ; degenerate. C. A still more reduced and degenerate form. N, macronucleus ; n, micronucleus. division of the entire animal, repeated division of the nucleus, the products of which are later surrounded by minute bits of cytoplasm, or by fragmentation of the nucleus without the formality of regular division. In the latter case the body of the animal breaks up simul- taneously into many hundreds of small pieces., each surrounding one of the nuclear fragments and developing later into the parent form (Paramceba}. So-called sexual reproduction or some modification of this process is accomplished, either directly or indirectly, by the temporary or permanent union of two individuals of the same or of dissimilar size. iSee Chapter IV, p. 159. 6o THE PROTOZOA From an a priori point of view, there is no reason why the reproduction of Protozoa by simple division should not go on indefinitely. The mechanism of metabolism, growth, and reproduction is present, and the cell appears therefore to be self-sufficient. Nevertheless, Biitschli, Maupas, and many others have shown that, in many cases at least, these divisions can be maintained only for a certain period or number of generations, after which the in- dividuals become weaker, deformed, and finally die out, an exhausted race (Fig. 29). If two individuals conjugate while in this enfeebled condition, the result is a rejuve- nescence or renewal of youth in both cases, and each of the conju- gants enters upon a new cycle of cell-generations (Fig. 30). The union of two cells is accomplished in various ways. In some cases it is by the absolute and permanent fusion of two individuals ; in other cases there is a union for a short Fig. 30. - Onychodromus grandis St. in . f o l] owef l U v a senaration in .jugation. [MAUPAS.] m, micronuclei in ll DV a se P aratl ision. still other cases the entire cell does not conjugate, but develops a great number of swarm-spores, which again may be of equal or unequal size, and which conjugate usually by total fusion ; finally, sexual reproduction, almost as highly differentiated as in the Metazoa and Metaphyta, is found in some Sporozoa and in the complex colonies of Flagellidia. 1 * A very interesting controversy has arisen over the question whether Protozoa ever succumb to old age. Ehrenberg was apparently the first to suggest that because of their reproduction by division, a process in which no portion of the original organism is lost, the Protozoa are potentially immortal. This conception was greatly 'expanded by Weismann ('84) and formed the basis of a suggestive essay in which he maintained that the Protozoa are too simply organized to die a natural death and that death is first observed in multicellular animals and plants. Dujardin ('41), opposed to so many of Ehrenberg's theories, was equally opposed to this, and although he was unable to disprove the theory, he suggested that simple division cannot continue conjugation division. 1 Cf. Chapter VII, p. 229. GENERAL SKETCH 6 1 indefinitely, but that periods of division recur at intervals. This sug- gestion was confirmed by Butschli ('76) and by Engelmann ('76), who demonstrated in connection with a number of Ciliata that after a certain number of divisions the resulting individuals become reduced in size and show other evidences of degeneration. Butschli regarded this as evidence of old age, and he observed that the normal size and the general vitality of the reduced organisms are restored by conju- gation ; and he was one of the first to demonstrate that the function of conjugation is not for purposes of reproduction, but for the renewal of vitality as expressed in his term Verjungung (rejuvenation). Maupas ('88), finally, has confirmed the latter conception of conjuga- tion, and in a series of brilliantly planned and carefully executed experiments has shown that Protozoa, contrary to Weismann's a priori assumption, may die of old age unless they be reinvigorated by conjugation. "Senescence," says Maupas, "appears to be a very general phenomenon, at least in the animal kingdom. ... It is inherent in the organism and comes from internal causes which act independently of the surrounding conditions. ... Its deleterious action is offset and annulled by sexual rejuvenescence or conjugation." l 4. Irritability. Unlike the Metazoa, where the phenomena which are characterized as manifestations of consciousness are expressed through special or- gans of the nervous system, the Protozoa in the simplest forms have nothing, so far as we know at present, but the undifferentiated proto- plasm which at the same time must be the seat of all functions. The sensory phenomena are, however, very little known. All Pro- tozoa are irritable, reacting in certain definite ways, although in dif- ferent degrees, toward various external stimuli. All are sensitive to electrical, mechanical, thermal, and chemical irritations, and many to light, while few or none are affected by acoustic vibrations. The reactions to these stimuli are usually expressed in motion of some sort, which may be either indefinite or definite, in the latter case, as a rule, either positive, i.e. toward the source of irritation, or nega- tive, i.e. away from it. In many Protozoa, particularly in the lower forms, there seems to be no portion of the cell more sensitive than others ; in the higher forms, however, there is a greater or less degree of sensory localization. Here, as a rule, the ectoplasm reacts ener- getically, and, like the cuticle of Metazoa, becomes a general sensory organ. The appendages frequently serve as special sensory organs of touch, as in the aboral cirri of the hypotrichous ciliates, while spe- cial organoids are frequently present in the form of "eye-spots," etc. i ('88), p. 272. 62 THE PROTOZOA C. SOME ECONOMIC ASPECTS OF THE PROTOZOA The Protozoa are frequently objectionable because of the appear- ances, odors, and tastes which they may impart to water. In the sea great areas may be colored orange, red, etc., by incalculable numbers of Noctiluca or Dinoflagellidia (Proroccntrum, Glenodinium\ while at night their presence is indicated by brilliant phosphorescence, the light being due to the rapid oxidization of a substance created by the organisms and thrown out by them upon irritation. In Puget Sound and in Alaska I have seen hundreds of acres of the sea surface colored orange by Noctilnca miliaris, although the single individuals are less than one-fiftieth part of an inch in diameter, and Haeckel ('90) graphically compares such masses to "tomato soup"! When Protozoa occur in great numbers in fresh water, and especially in drinking water, they may cause considerable annoyance ; for by the color, odor, and taste which they impart they render the water unfit to drink. The colors are due in the main to the Phytoflagellida, and only those forms which are capable of making their own food are able to live in pure drinking waters. The most frequent causes of trouble in this respect are Uroglena, Peridinium, or its allies, Euglena and other Euglenoids, and Synura, all of which are flagellates. The odors and tastes, however, are more offensive than the colors, and as they are frequently misunderstood and regarded as evidence of pollu- tion, an explanation may not be out of place. Ehrenberg noticed that certain flagellates (Chlawydomonas pulvisculus and Chlorogonium) impart a certain oily odor. Dunal ('38) and Joly ('40) described an odor like that of violets from the masses of Hczmatococcus which gave to a portion of the Mediterranean a distinct red color. The Massa- chusetts State Board of Health, dealing with this problem of the drinking waters, have obtained important results in this direction. They have shown that certain of these organisms may have definite and specific odors which, like the odors of flowers, can be recognized. An "oily odor" was traced to Synnra and Uroglena, an "Iceland moss" odor to Peridinium, a "violet odor" to certain Euglenoids, etc. 1 The cause of these odors has been the subject of a number of investigations, and it has been found that they are " living odors " due to disintegration of the cells rather than to their decomposition, a view first advanced, I believe, by Biitschli ('84), who described a highly characteristic " fishy odor " from Euglena sanguinea, while the cells were found to be disintegrated, although not decomposed. The matter was considered more extensively by the writer ('92), who found that in waters infested with colonies of Uroglena americana the odor was not developed until the organisms had passed through the water iSeeS. B. H. ('92). GENERAL SKETCH pipes, but after such passage the odor was extremely strong and repulsive, while no colonies could be found. It was suggested at this time that the odor was one of disintegratidn, and due to the libera- tion of minute drops of oil-like substance which become disseminated through the water, giving it the characteristic Uroglena smell. It was also suggested that these drops of oil are analogous to the perfume oils of the fragrant plants, like them having a certain individual odor often strong enough and characteristic enough to identify the organ- ism. Similar oil-like inclusions are found in the protoplasm of all Protozoa, but to be detected through the sense of smell, they must be present in great numbers. Far more serious noxious effects of the Protozoa are produced through their frequently parasitic mode of life. In all classes there B D Fig. 31. Internal parasites. [A, B, LEUCKART; C, GRASSI ; D, BiJTSCHLl .] A. Amaba colt Losch, a supposed cause of dysentery. D. Monocystis agilis Leuck., a grega- rine. C. Megastoma. entericum Grassi, a flagellate. D, Balantidium entozoon Ehr., a ciliate. are certain forms which live as parasites (Fig. 31), and which for con- venience may be separated into two groups, the intercellular and the intracellular forms. So far as known these parasites, with few excep- tions, do not produce noxious products like bacterial ptomaines, but whatever damage they may cause is due to the mechanical disturb- ances set up by their presence. The intercellular parasites infest the body cavities of various hosts, the cavities of blood-vessels, and ducts of various glands, or penetrate the spaces between muscle-fibres, while the intracellular parasites (which belong almost exclusively to the Sporozoa) bore into cells of epithelia (Gregarines, Coccidia), or the corpuscles of the blood (Haemosporidiida). From the wide distribu- tion of the intercellular parasites, it is quite possible that no animal is entirely free from Protozoa of some kind. Without entering upon a 64 THE PROTOZOA discussion of these forms, it may be stated that Rhizopoda, Flagellidia, Ciliata, and Sporozoa may be found in the various cavities and canals in man and the other vertebrates, where they usually give little or no trouble. One form, however, Amoeba coli (Fig. 31, A), has been long in dispute as the reputed cause of dysentery. If it is the specific cause of this disease, the animal occupies an interesting position amongst the intercellular parasites ; for, so far as known, none other of this kind of protozoan parasites exerts a deleterious effect upon the intestinal epithelia. Nor is it proven that Ainceba coli does this in the case of dysentery, although a belief to that effect is widespread. Briefly reviewing the history of this belief, it appears that Lambl ('60) 1 was the first to observe Amceba in the human intestine, although Lb'sch ('75)> who named it, was the first to consider it in connection with dysentery. Many subsequent observers (Kartulis, Mannaberg, Cohn, etc.) found Amoeba coli in the faeces of dysentery patients, Kartulis ('89) amongst others stating that he found them in no less than five hundred cases. The belief received a setback, however, by the observations of Cunningham ('81), Grassi ('82), and Calendruccio ('90), who found Amceba coli in the intestine of sound and healthy men as well as in dysenteric patients, while still other observers maintained the entire absence of such an enteric organism. Councilman ('91)5 i n a work which is certainly as reliable as any that has been undertaken upon this subject, partially harmonized these views by showing that there are at least three forms of dysentery, of which one, at least, is characterized by certain definite symptoms and by the presence of Amceba coli, although it was not demonstrated that the rhizopod was the cause. The entire matter received impartial and critical treatment by Laveran ('93) in France and by Schuberg ('93) in Germany, and both came to the conclusion that the cause of dysentery was not yet known, the former basing his opinion largely upon the absence of Amceba in all but one of ten cases, the latter upon numerous experiments and observations upon normal and diseased individuals. Schuberg not only found that Amceba coli is present in normal men, but also found that there is no specific difference in the various intestinal Amoebce which have been described by various observers as living freely in the intestine in the same way as the commensal ciliates and flagellates also found there and generally believed to be harmless. He pertinently says : " If the flagellates are harmless, it is certainly not impossible that Amoeba is also. The increased number of Amoebce in dysenteric patients is not necessarily evidence that they are the cause of this disease." 2 Both he and Laveran expressed the view that all experiments which had been made up to that time had not excluded the possibility of other .causes, e.g. bacteria. That dysentery is due to some specific cause had been early demonstrated by experiment, but in none of these experiments had it been possible to isolate the Protozoa from bacteria which invari- ably accompany them. The reverse experiment is, however, possible, and it is singular that it has not been made more frequently. Among the first to exclude the Protozoa were Celli and Fiocca ('95), who obtained cases of dysentery by inject- ing cats with material from faeces in which the Protozoa had been killed by heat ; the same result was also obtained by injecting material in which both Amceba and bacteria were absent, the cause evidently being in the poison of the sterilized matter and not in Amoeba coli. They concluded that the poison is the product of bacteria (Bacillus coli communis, together with typhus-like bacteria and a streptococcus, were suggested as possible causes). This view was supported by a number of observers, amongst whom may be mentioned Gasser ('95), Cassagrandi and Bar- bagello ('95), and Petridis ('98). The latter especially has shown that dysentery as observed in Egypt is due to a bacillus and not to Protozoa. He found that Strep- a Cf. Leuckart ('79). . 2 Page 701. GENERAL SKETCH 65 tococcus is the most numerous of the micro-organisms and the probable cause of the disease, for he was able to isolate the bacillus and to produce dysentery in cats by injecting them with the culture obtained from it. Thus, as the matter stands, Petridis's results, the most positive that have yet appeared, together with growing evidence from the bacteriological side, make it exceedingly probable, although not definitely established as yet, that bacilli and not Amoeba coli are the cause of this disease. While the majority of intercellular parasites are harmless, it is quite different with the intracellular forms. . These, by making their way into the interior of the cell and growing at the expense of the cell-contents, gradually cause degeneration of the tissues which may end in death of the host. These parasites belong almost exclusively to the class Sporozoa of which the Coccidiida and Hasmosporidiida are found in vertebrates, while the Gregarinida are confined to the inverte- brates, where they are widely distributed. The Coccidiida are found in nearly all of the tissues of the lower vertebrates although rarely in man, unless indeed, as many observ- ers believe, they are the cause of various tumors and cancers. That there is some reason for this belief is shown by the fact that in the lower vertebrates, especially in fishes, the presence of Sporozoa leads to ulcers and tumors and to the ultimate death of the fish. The sub- ject, however, as far as man is concerned, is in a very unsatisfactory state, and opinions differ widely as to the nature of certain elements found in cancerous growths. By some observers these are regarded as parasites, by others as disintegrated or pathological cells. Up to the present time no satisfactory evidence has appeared to prove the former view, and until such evidence is forthcoming the entire matter must rest in abeyance. From the pathogenic point of view, the most important protozoon is the malaria germ {Plasmodium malarias), a form belonging to the Haemosporidiida. These organisms, in the young stages, move about by amoeboid motion in the blood-vessels of men and birds. They penetrate the red blood corpuscles, which slowly hypertrophy, until in one type of the disease, at least, they attain a size three to four times that of the normal corpuscle, the parasite in the meantime growing at the expense of the haemoglobin and finally reproducing by spore- formation. In this form alone there appears to be a poisonous sub- stance analogous to bacterial ptomaines, which is produced by the organism and periodically discharged (at spore-formation) into the blood, thus causing the pyrexial attacks so characteristic of malaria. The recent successful results obtained by Ross, Manson, Koch, Grassi, and others, in locating the seat of the malaria germ when outside the human body, leads to the hope that some successful means of guard- ing against this disease may soon follow. 1 1 Vide infra, pp. 160-165. 66 THE PROTOZOA SPECIAL BIBLIOGRAPHY II Biitschli. 0. Protozoa. In Brands Klassen nnd Ordnungen des Thierreichs. Leipzig, 1883-1888. Delage et HSrouard. La cellule et les Infusoires. In Traiti de Zoologie concrete. Paris, 1896. Ehrenberg, C. G. Die Infusionsthierchen als vollkommene Organismen. Leipzig, 1838. Entz, G. Protistenstudien. Budapesth, 1888. Lankester, E. R. Protozoa. In Zoological Articles from the Encyclopedia Britan- nica. 1891. Stein, Fr. Der Organismus der Infusionsthiere. Leipzig, 1861, 1867, and 1878. CHAPTER III THE SARCODINA THE term Sarcodina, introduced by Biitschli ('83) as the class name of the most primitive of the Protozoa, includes all forms which, like the common fresh-water type Amceba proteus, move by the pro- trusion of protoplasm in the form of broad and finger-like, or sharp and ray-like, processes called pseudopodia. These forms fall naturally into three groups readily distinguished by clearly marked differences in structure, the Rhizopoda, Heliozoa, and Radiolaria. Among the Rhizopoda are included forms of Sarcodina with blunt, finger- form or lobose pseudopodia (Amcebidd) or with branching and anastomosing pseudopodia (Reticulariida). They may be naked (Gymnamcebina\ or shelled {Thecamcebina or Foraminifera). The pseudopodia may arise from all parts of the body or they may be limited to special regions ; in shelled forms they may pass through one common opening (Reticulariida impcrforina}, or through many finer openings {Reticulariida pcrforina}. The body form is typically globular, but may be variable in consequence of amoeboid changes, or drawn out into a monaxonic form. The material of the shell may be chitin, silica, foreign particles, or calcium carbonate. The Heliozoa are naked or shelled forms of Sarcodina; they are usually globular with fine ray-like pseudopodia arising from all parts of the body. The rays are, as a rule, stiffened by an axial filament formed of modified protoplasm which may be readily dissolved by the organism. The shells are less compact than those of the Rhizopoda, and are usually formed of more or less loosely joined silicious spicules. The Radiolaria are similar in form to the Heliozoa. As in the latter, the pseudopodia arise from all parts of the body and occasion- ally anastomose. The endoplasm is separated from the outer plasm by a firm, chitinous, perforated membrane, the central capsule. A test or skeleton, often of exquisite beauty, is usually present, consist- ing of isolated spicules of silica, or of a compact skeleton of acanthin or silica. One or more nuclei are invariably present within the central capsule. The finer structure of the rhizopod protoplasm has already been mentioned. In many cases, especially in the monothalamous forms, the plasm is divided into a number of clearly marked zones. Schewi- 67 68 THE PROTOZOA akoff ('88) describes three, Penard ('90) no less than four in EtiglypJia, and Rhumbler ('98) the latter number in Cyphoderia. Schewiakoff ('88), apparently on very good grounds, maintained that certain spe- cific functions characterize each of these zones, indicating, in a general way, a regional differentiation and division of physiological labor. To the outer zone, which corresponds to the ectoplasm of Amceba, he ascribed a locomotor function, this being the seat of pseudopodia for- mation ; to the second zone, which contains the nucleus, the function of assimilation, and to the third zone a reproductive function. Penard and Rhumbler separate Schewiakoff 's second zone into two on account of certain structural differences. According to these observers the Pig. 32. Actinopkrys sol Ehr. [BUTSCHLI after GRENACHER.] The axial filaments (a) extend through the. endoplasm to the membrane of the nucleus ; c, a contractile vacuole in the ectoplasm ; g, an ingested food particle in a gastric vacuole. outermost zone is distinctly vacuolated, the second contains food-par- ticles in the process of digestion, the third, granules which represent waste matter not determined, and the fourth, excretory granules. The appearance of the protoplasm in Heliozoa or Radiolaria is quite different from that of the Rhizopoda. Ectoplasm and endoplasm can be distinguished, but unlike the hyaline .ectoplasm of Amoeba, the outer plasm of Heliozoa is made up of vacuoles much larger than those of the endoplasm, the walls of these vacuoles being distinctly granular (Fig. 32). The extremely vacuolated appearance, however, seems to be largely dependent upon the medium in which the animal r -t ' lives. Griiber found that an ActinopJirys when transferred from fresh into sea water soon loses its vacuoles ; and, vice versa, when trans- ferred back to fresh water, again acquires its vesicular appearance. In general appearance a radiolarian resembles a heliozoon, but there is a considerable difference in the corresponding regions. A typical radiolarian can be conceived if we imagine a thick perforated chitinous membrane between the ectoplasm and endoplasm of a heliozoon. The intra-capsular plasm (Fig. 33, c) contains nuclei, fat particles, and plastids of one form or another, and is in communication with the extra-capsnlar plasm through the pores in the membrane, although, as shown by Verworn's experiments upon the isolated central capsule, it can live for a time independently. The outer or extra-capsular plasm is composed, according to Haeckel, of four parts. The outermost {g) is a zone of pseudopodia ; the latter, however, originate in the deeper fourth zone, forming a network through the other extra-capsular parts. The second zone is of net- like (alveolar ?) protoplasm, the sarcodictyum. A third zone, the calymma, is of jelly-like consistency and forms the bulk of the ecto- plasm. The fourth and most im- portant zone, the sarcomatrix, lies close against the central capsule, and is the go-between for the intra- and extra-capsular portions. The sarcomatrix is also the seat of di- gestion and assimilation, the food coming to it through the pseudo- podia and the network. As the means of communication between the central protoplasm and the sar- comatrix is of vital importance to the organism, the arrangement of the apertures in the central capsule offers a good character for the classifi- cation of the Radiolaria. Hertwig ('79), who first used this character, divided the group into four legions, as follows: (i) the Peripylea, in Fig. 33. The protoplasmic regions of a radiolarian ( Thalassicolla maculata) Haeck. [HAECKEL.] a, large alveoli forming part of the calymma in which foreign bodies (b) are enclosed, and which is penetrated by meshes constituting the sacrodictyum ; c, the central capsule and intra- capsular plasm ; f, retracted pseudopodia. The nucleus (n) contains a distinct nucleolus (/); the sarcomatrix is darkened by pigment masses (p). which the membrane of the central capsule is perforated by pores arranged regularly about the entire surface (Fig. 34, A) ; (2) the Acti- pylea, in which the pores are arranged in groups over the surface (B); (3) the Monopylea, in which there is but one such group of pores in the membrane. In these forms the perforated disk is con- nected with the centre of the central capsule by a conical mass of endoplasm, the podocomis (D), rich in food particles and gran- D Fig. 34. Central capsules of Radiolaria. [HAECKEL.] A. Thalassolampe maxima Haeck., one of the Peripylea. B. Acanthometron dolichoscion Haeck., one of the Actipylea. C. Aulographis candelabrum Haeck., one of the Monopylea. D. Triptero- calpis ogntoptera Haeck., one of the Cannopylea. c, central capsules ; , nuclei. ules ; (4) the Cannopylea, in which the membrane around the pores is drawn out into funnel-like projections termed astropyles (C). The central capsule is double in these forms. Haeckel has found that certain skeletal forms accompany the structure of the membranes, and he names the above legions respectively as follows : ( I ) Spumel- laria ; (2) Acantharia ; (3) Nassclaria, and (4) Pliczodaria. In each of the orders of the Sarcodina, and especially in the Radiolaria, there are some forms with symbiotic plant-cells. The THE SARCODINA 71 relationship between the symbionts was worked out by Cienkowsky, Brandt, Haeckel, and Entz, the latter noting that the plant-cells are invariably found just outside of the endoplasm, where they do not come in contact with endoplasm and its digestive fluids. According to the more recent observations of Le Dantec ('92), however, the digestive fluid of these animals is unable to dissolve the cellulose membranes of the plant-cells, and they remain uninjured in the endo- plasm, dividing there when the conditions are favorable. D A. SHELLS AND TESTS The ectoplasm of naked protoplasm shows a tendency to condense or stiffen when in contact with water, and a cuticle or membrane is the result. Amceba proteus, with its differentiation into endoplasm and ectoplasm, shows a primitive stage in the development of such mem- branes. Here the ectoplasm remains plastic enough to yield to the inner pressure of the organ- ism and to form the first part of every pseudopodium ; it is rapidly pushed aside, however, and the endoplasm becomes the advancing part. In Amoeba tentaculata the outer layer has become more firm and the pressure from within expends itself upon pseudopodia which are pro- truded through permanent holes (Fjg. 12, A). The membrane may become Still Fi 8- 35- Types of marine rhizopod shells (Reticula- r. ., , rlida. [CARPENTER.] more firm through the ^ Lateral B Vent L ra] view of a j monothalamous deposition Of Chitin, Until, shell (Cornuspira foliacea Phillips). C. A simple poly- as in the radiolarian central thaiamous shell capsule, it is an efficient means of protection. In addition to the chitin, certain Sarcodina secrete a silicious mucilaginous material, which, like the chiti- nous cement, is frequently the means of gluing together not only regular plates or disks which the organism also secretes, but foreign particles of various kinds. The tests thus made may be entirely of lime, as in the Reticulariida, or of silica, as in the Radiolaria and many of the Heliozoa, or of sand crystals, diatom-shells, or detritus of various kinds. In the lime-shells (Reticulariida or Foraminifera) the secretion of calcium carbonate, except for the invariable presence of a mouth- D. Verte- bralma sp., a fossil form. -72 THE PROTOZOA opening, forms an almost complete investment like a cyst. In many cases this opening is the only means of communication with the sur- rounding medium (Imperforina), but in other cases the entire shell is punctured by minute openings through which pseudopodia pass to the outside (Perforina). These two types of shell are further distin- guished by their appearance ; the Imperforina when seen by reflected light are opaque and like porcelain, while the shells of the Perforina are almost transparent (vitreous). Monothalamous or single-shelled Foraminifera may be either im- perforate (e.g- Sqnamnlina, Pilnlina, or Saccammina) or perforate (Lagena). In each group a graded series of shells can be arranged, varying in complexity from the simple monothalamous to the compli- B C - 36- Polythalamous shell types schematized. [CARPENTER.] A. Linear Nodosaria type. D. Frondicularia form of the Nodosaria type. C. Spiral form of the Nodosaria type. cated polythalamous forms (Polystomella, Calcarina). One of the simplest of these shells is that of Cornuspira, where the plasm, as it slowly grows, constantly secretes new shell material and is capable of unlimited extension (Fig. 35, A). It is never divided by septa into separate chambers as in the polythalamous shells. A further step, the simplest of the polythalamous types, is found in shells where the separate chambers adhere end to end as in Nodosaria (C). Here there may be only a slight septum between adjacent chambers, but enough to indicate that growth is periodic, and not constant as in Cornuspira. In these chamber-dwelling animals the plasm, as it grows, extends out of the primary shell-opening and reaches to a certain distance down the outside ; new shell material is then secreted, and the process is repeated until a chain of chambers is the result (Fig. 36, A). If THE SARCODINA 73 the plasm extends entirely around the shell, the new chamber almost incloses the older ones as in Nodosarina (B}. In other cases the plasm may extend over one side only of the old shell, and a curvi- linear axis of growth is the result (Fig. 35, A, B> and 36, C}. The spiral thus formed may be flat or coiled around a longitudinal axis as in the mollusc Trochns, giving an involute shell. This type, the most highly differentiated of all of the rhizopod shells, exhibits all grades of complexity (Fig. 37). In the highest forms each new chamber has a complete wall, so that the septa between the adjacent chambers consist of two lamellae, while between the lamellae there is fre- Fig. 37. A complex polythalamous shell (schematic) of Operculina. [CARPENTER.] The shell is represented as cut in different planes to show the distribution of the canals (a', a", a'") ; c, c, c, the outer chambers with double walls (d, d, d), one of which is shown in sec- tion (g). The chambers communicate by apertures at the inner ends of the septa (e), and by minute pores (/). The outside () of the shell is marked by the radial septa. quently a space filled with a calcareous deposit or what Carpenter ('62) calls the " intermediate skeleton." This inter-lamellar deposit is traversed by a complicated system of canals, and the deposit itself is frequently carried out into external processes and knobs (Calcarina). In the annular or discoid types a process of budding takes place around the entire circumference instead of at a localized area, and concentric circles of chambers are thus formed (Orbitolites). The character of the mouth-openings between adjacent chambers depends upon the nature of the outer coating. If the Ijme casing is perforated by numerous pores through which pseudopodia can be thrust to collect food, then each chamber is sufficient for itself, and the so-called mouth-opening is small ; but if the perforations are absent, the mouth-openings are large and allow a free communication 74 THE PROTOZOA between the youngest or external chambers and the oldest or internal. Hence there are morphological and physiological grounds for sepa- rating the Reticulariida into Perforina and Imperforina. It frequently happens that the central or original chamber varies in size in the same species, being large (inegalospheric) in some individuals, and small (microspkeric) in others (Fig. 38). While the relations of these two forms have been much discussed, no satisfactory conclusion has yet been reached. Lister ('95) regards the case as one of alternation of generations in which spores from individuals A conjugate and form individuals of the type B, while the latter develops spores which grow into the form A again. The conjugation of swarmers in these dimor- phic types is a matter of inference rather than of observation, for the process has never been seen. B Fig. 38. Megalospheric (A) and microspheric (B) shells of Biloculina depressa Lam. [SCHLUMBERGER.] The dimorphism is shown by the central chamber c. Among the Heliozoa and Radiolaria, shell formation is of a somewhat different type, consisting of the deposition of spicules and rays rather than a continuous layer of material forming a compact coating. Even naked forms of Heliozoa, such as Actino- spharium, secrete these spicules at certain times for the purpose of encystment, while others have them in greater or less numbers throughout life. Isolated spicules are usually retained by a gelat- inous mantle, which covers the entire animal (Nnclearia, Acti- nolophus, etc.). These spicules are usually curved or straight rods, THE SARCODINA 75 spindles, or blade-shaped plates, and may become firmly attached to one another, forming latticed skeletons, like those of Radiolaria {Clathrulina, Fig. 39). Intermediate stages are seen in such forms as Diplocystis, where the plates are very small and arranged without Pig. 39. Clathrulina elegans Cienk. [GREEFF.] any apparent order in the gelatinous mantle. In Raphidiophrys (Fig. 40) the silicious plates are much larger and more regularly arranged, while in Pinaciophora and Acanthocystis (B, C, D} they be- come so closely knit that they form an efficient shield. In Acantho- cystis, each plate is a small rectangular prism, laid tangential to the surface with sharper spicules arranged at intervals at right angles to 70 THE PROTOZOA . these, thus forming a bristling coat. Pinaciophora is very similar, but the spicules are not so prismatic. Similarly the Radiolaria may have either simple isolated spicules or compact and strong skeletons. In many cases the outer plasm (calymma) is free from spicules, but in other cases isolated spicules of sharp and needle-like, or tri- or tetra-radiate form, are present. The Fig. 40. Types of spicules in Heliozoa. [PENARD.] A. Raphidiophrys pallida F. E. Sch., with curved silicious rods. B. Pinaciophora rubiconda Hert. and Less. C. Acanthocystls turfacea Carter. D. Pinaciophora fluviatilis Greeff. substance of the skeleton of Radiolaria is either silica or acanthin, a horn-like modification of protoplasm. According to Haeckel, the deposition of silica in many cases occurs only at certain periods, and an entire skeleton may be laid down at one time (Dictyotic moment, Haeckel, or Lorication moment, Dreyer). Again, it may be formed during the entire period of life. The material of the shell is secreted from the sarcodictyum, and as the deposition of the silica THE SARCODINA 77 follows the outlines of the vesicles which form this zone of proto- plasm, the resulting skeleton forms a reticulum. Growth may take place more rapidly, however, at certain places, and spines, spicules, or protuberances of one kind or another are the result. The usual form of the network upon which the skeleton is deposited is an hexagonal mesh, but this may become modified in numerous ways, the apertures becoming either circular, polygonal, or elliptical (Fig. 41). When spines are formed, a secondary calymma may also be developed, carrying with it the sarcodictyum, and the latter, in turn, may give rise to a secondary skeleton outside of the first. This process may be repeated until there are as many as six or seven acces- sory skeletons. Fig. 41. Schematic figure illustrating the modifications of skeletons according to mechanical principles of deposition. [DREYER.] The secretion is supposed to collect in the interstices between alveoli as at (c), forming simple spicules, or tri- and tetra-radiate spicules (&). Collecting in the lines of union of six alveoli, the deposit takes the form of an hexagonal mesh (d ) , which, by the addition of more material, becomes changed as at (a), (e), (f), and (g). A very interesting set of phenomena are connected with the acanthin skeletons where the spicules are not deposited in the calymma, but are formed at the centre of the central capsule, growing out centrifugally into the extra-capsular plasm and resulting in a skeleton of radiating spines. With a few exceptions these spines are twenty in number, and are arranged in a certain geometrical order which has been characterized as the Miillerian law. The points of the spines fall in five circles parallel to the equator, and there are four spines to each circle. The spines are named, according to this scheme, polar, tropical, equatorial, sub-tropical, and sub-polar (Fig. 42). The form of the silicious skeleton is quite varied. In its least-dif- ferentiated form, as in most Heliozoa, it is a mere collection of loosely arranged spicules. In other forms a uniform covering of silica covers the meshes of the sarcodictyum. Such a generalized condition of the skeleton becomes modified in many ways, the main types being the " sagittal ring," consisting of a simple ring of silica, like a girdle THE PROTOZOA around the body of the organism. A second type consists of a basal tripod, the arms of which inclose the central capsule (Fig. 43). A third modification is the simple alteration of the spherical latticed shell Fig. 42. Lichnaspis giltochh Haeck., one of the Acanthana (Actipylea). [HAECKEL.] The spines are arranged in accord with the Miillerian Law as follows : a, a, a, a, northern polar spines ; 6, b, b, b, northern tropical spines ; c, c, c, , equatorial spines ; d, d, d, d, southern tropi- cal spines ; and e, e, e, , southern polar spines. into elliptical, ovate, or sub-spherical forms. Again, the skeleton may be discoid, or even bivalved, and, in still other types, there may be a combination of two or more of the above modifications. THE SARCODINA 79 B. PSEUDOPODIA A pseudopodium is a portion of the body-plasm temporarily pro- truded. It is most variable in form, and at any moment can be with- drawn into the body of the animal to be replaced by others. In the Rhizopoda, the pseudopodia are coarse, blunt, and finger-formed (Amoebida), or fine, and often forming a network through anastomosis (Reticulariida). In the Heliozoa and Radio- laria they are more rigid and radiate out from the body in all directions, forming a pro- tective coating, and from their ray-like appearance suggest- ing the common name " sun- animalcula." There is a difference in the texture as well as in the form of the lobose and reticulate pseudopodia of the Rhizop- oda. In the former the hya- line ectoplasm, which goes into the pseudopodia, is ap- parently homogeneous and structureless, although, upon critical examination, Butschli ('92) was able to make out a fibrous structure in some forms, and in many of them a reticular appearance was obtained upon retrac- tion. His observations led him to the conclusion that the hyaline appearance is due to the close approximation of the walls of the alveoli, and not to their absence. The outer plasm is certainly more dense and non-granular than the endoplasm, and protoplasmic stream- ing is confined to the latter. The outer plasm in the reticulate type, on the other hand, is granular, while the central portion is denser and more resisting. Streaming of the granules here takes place in the ectoplasm, instead of in the endoplasm, and when two or more pseu- dopodia come in contact, the viscid character of this outer plasm leads to fusion. The lobose forms, on the other hand, never coalesce. The resemblance between the central denser strand of protoplasm in the pseudopodia of the reticulate type and the axial filament of the pseudopodia of Heliozoa and Radiolaria was early recognized by M. Schultze ('63) and critically examined by Butschli ('92) and Schaudinn ( '93 )> an d is now generally recognized. Fig. 43. PI agio carp a procortina Haeck., with tripod-like skeleton. [HAECKEL.J 8o THE PROTOZOA The nature and the number of pseudopodia have frequently been used as a method of identification of certain species of Rhizopoda. Amceba polypodia, A. radiosa, and A. proteus have certain characteris- tic pseudopodial structures which are seemingly of diagnostic value, yet A. proteus under the influence of a constant electric current can be made to assume the forms characteristic of A. polypodia and then --"-" Fig. 44. Types of pseudopodia. A. Amoeba limicola'Khmh. [RHUMBLER.] B. Amoeba blatta: Butsch. [BUTSCHLI.] C. Lie- berkiihnia sp. [VERWORN.] D. Actinospharium Eich. Ehr. [ORIGINAL.] x, axial filament. of A. Umax. Conversely, A. Umax, when placed in an alkaline solu- tion of potassium hydrate, becomes transformed into A. proteus, and later, into A. radiosa (Verworn, '94). When a change in the surround- ing medium can so affect the protoplasm that the entire character of pseudopodia-formation is altered, the specific value of pseudopodia alone may well be questioned, and species based upon such a variable THE SAKCODINA 8l character can have but little value. Despite the uncertainty of the pseudopodia as a basis of classification, their structure among the Rhizopoda is frequently so characteristic that the identification of some species of Amoeba is comparatively easy. Thus Anmba proteus has large and blunt pseudopodia in the adult phase, while the young form (known as A. radiosa} 1 has sharper, stiffer, and hyaline pseudopodia. When a pseudopodium of A. proteus starts from the periphery, it continues as a stream until, as a rule, a long, lobose structure results. When, however, a pseudopodium of A. blatta Biitschli or of A. limicola Rhumbler starts from the periphery of the spherical body, it resembles a miniature eruption. A break is Fig. 45. Camptonema nutans. [SCHAUDINN.] The axial filaments extend throughout the endoplasm (A), taking their origin at the nuclear membrane (B). x, an axial filament highly magnified. made on the periphery, and through it the granular endoplasm flows down the sides of the spherical body instead of outward into elongate pseudopodia (Fig. 44, A, B}. In such cases the pseudopodia may be used to identify the organism. The pseudopodia of the Heliozoa and the Radiolaria are far more complicated than those of the Rhizopoda. They usually have dis- tinct axial filaments, consisting of some unknown substance, extending throughout the entire length, and even into the endoplasm, where they not infrequently abut against the membrane of the nucleus or meet at a common centre (Actinophrys, Acanthocystis\ The granular protoplasm which surrounds the axial filament is in constant but slow streaming motion. The point of interest is the axial filament, which is not strictly comparable with the skeletal parts, but is probably stif- i Cf. Scheel ('99). 82 THE PROTOZOA f ened protoplasm similar to the central plasm of the reticulate pseudo- podia. It is easily softened by the animal, and when the latter is irritated may be withdrawn into the body. That there is some con- nection between the axial filament and the nucleus would seem to be indicated by their invariable propinquity, the nucleus in some cases being actually surrounded by the substance that forms the filament and which Schaudinn ('96) thinks is a soft fluid at this time (Campto- nema nutans, Fig. 45). In other cases the filament appears to end in a peculiar crescent or spherical capsule which lies within the endo- plasm (DimorpJia, Fig. 46). In many instances the rays pass com- pletely across the animal's body and rest against the nucleus on the opposite side ; in others they are focussed in a central or " astral " C Fig. 46. Flagella (/) and axial filaments of the pseudopodia of Ciliophrys (Dimorpka?) Cienk. [BLOCHMANN.] In the Heliozoa stage (A) the ray-like pseudopodia (/) and the flagella (/) are present ; in the flagellate stage (B) the pseudopodia are absent. The axial filaments (.r) and flagella centre in the excentric nucleus (C). granule (Gymnosphara, Actinophrys, Splicerastrum, etc.), which in some cases has been seen to divide like a centrosome and to form an amphiaster, as in the early stages in cell-division of many cells of the Metazoa (Acanthocystis, SpJuzrastrum, etc.). 1 The axial filaments have not been made out in all forms classed among Heliozoa, and it is a question whether such forms should be considered as Heliozoa or as Rhizopoda. Vampyrella and Nuclearia (Fig. 56), for example, have fine, radiating pseudopodia which change like those of the Rhizopoda and, as in many Amcebida, are formed of hyaline ectoplasm. They are placed among the Rhizopoda by some (Delage) and among Heliozoa by others (Butschli). The pseud- opodia occasionally vary in other respects from the sharp radial forms, as in Actinolophus, where they end in knobs ; or in Camp- 1 Cf. Chapter VIII. THE SARCODINA 83 tonema, where there is an elbow or joint which can be bent at right angles. No entirely satisfactory explanation of pseudopodia formation and movement has yet appeared, although the subject has been attacked on many sides, and by almost all students of the Rhizopoda since the time of Dujardin. Like the early attempts to explain other phe- nomena in the Protozoa, the first explanations of pseudopodia-motion were based upon the analogy to higher forms. Protoplasmic contractility, the basis of locomotion in all higher animals, and probably in many Protozoa (Mastigophora and Infusoria), was early suggested as the cause of the protrusion of pseudopodia. The majority of casual observers were content with this general explana- tion ; others, more definite, conceived the seat of contraction to be in the cortical plasm or ectoplasm (Ecker, '49; Dujardin, '41), which they compared with the dermal musculature of worms, and which they supposed forces out the pseudopodia by backward peripheral con- traction, as water can be forced out of a rubber tube by pressure from behind. Others, again, imagined that in addition to the contractile cortex the entire mass of the amoeboid body is penetrated by a con- tractile substance (Cienkowsky, '63), the sarcous matter of Briicke ('61). Still others conceived a contractile motor apparatus of even greater complexity. Amongst these, Heitzmann ('73), in working out his well-known theory of the structure of protoplasm, and adapting Brucke's view to his own interpretation, maintained that the body of Amceba is composed of contractile fibres and an inter-fibral " non- contractile fluid." The protrusion of a pseudopodium, he argued, is due to the local contraction or stretching of this fibrous framework. Modifications of Heitzmann's view have frequently appeared in sub- sequent writings. In connection with the Metazoa it still makes its appearance in the numerous theories of contractile fibres, especially in explanation of mitosis (van Beneden, Boveri, Flemming, Reinke, and many others). In connection with the Rhizopoda, it found its most ardent advocate in R. Greeff ('91), who described radial, fibrillar, contractile structures in the ectoplasm of many so-called Earth Aincebce, and interpreted them as muscle-fibres whose outer ends are inserted in the ectoplasm with their inner ends attached to the protoplasmic framework of the endoplasm. Subsequent research has shown that the supposed muscle-fibres are bacteria (Bourne, '91 ; Israel, '94 ; Gould, '95). Contractility in a somewhat different form was also brought in to explain pseudopodium formation. In connection with the Protozoa, the most noteworthy advocate was Engelmann ('79), who conceived units of contractile substance built up of molecules of protoplasm. To these hypothetical units he gave the name inotogmata. During 8 4 rest, Engelmann assumed, each inotogma has an elongated form, becoming spherical upon contraction. If all contract at the same time, as upon a sudden shock, the animal assumes a spherical con- dition ; if the inotogmata contract in certain groups, a pseudopodium is started, although some pseudopodia, notably the fine, thread-like forms, are due to " relaxation " of rows of contracted units. A considerable uncertainty is attached to Engelmann's theory, especially when the attempt is made to explain special cases, and Biitschli ('92) shows in a very convincing manner that it does not justify the expec- tations of its originator. 1 Wallich ('63) early observed that the current in a progressive pseudopodium does not begin in the body of the Amoeba, but at the periphery; an observation which de Bary ('64) confirmed in Mycetozoa. Biitschli ('73) drew attention to the same fact soon after, and upon the strength of his observations appeared, even at this early period, as an opponent of the contractility hypothesis. As stated previously, Biitschli holds that protoplasm is essentially a mixture of liquids consisting of a fluid alveolar substance and an intra-alveolar fluid of different physical nature. According to this conception, which is widely accepted, a naked protoplasmic mass such as Amoeba must be subject to the same physical laws as other fluids. The rounding-out of drops of exuded protoplasm was early interpreted by Hofmeister ('67), and by Engelmann ('69) before he adopted the theory of inotogmata, as the same phenomenon that causes the rounding-out of any liquid substance, i.e. to surface tension. Of late years, especially since the appearance of Biitschli's masterly work on the structure of protoplasm, there has been a general tendency to abandon the older theory of contractility and to explain the move- ments of amoeboid bodies through the physical laws of liquids, and in particular, by the laws of surface tension. Weber ('55) compared the protoplasmic movements in plant-cells to the streaming, due to surface tension, in drops of liquid, and subsequently Berthold ('86), Biatschli ('92), Rhumbler ('98), and others, following the same line of investi- gation, have obtained fruitful results. An excellent account of the several interpretations along this line of reasoning may be seen in Biitschli's Protoplasma? and it will be sufficient here to give the most recent explanation as worked out by Rhumbler ('98) upon the basis of Biitschli's earlier view. Biitschli says: "The expla- nation of the processes of movement in Amoeba is to be found, therefore, to my mind, in correspondence with the interpretation of the phenomena of streaming movements in the drops of foam, in the fact that, by the bursting of some of the superficial alveoli, enchylema 1 Cf. Biitschli, p. 275. 2 pp. 172-212. THE SARCODINA [BUTSCHLI.] is poured out upon the free surface of the protoplasmic body, where it produces a local diminution of surface tension, and in this way sets up an extension centre together with forward movement." 1 The origin of a pseudopodium, according to this conception, is in the ectoplasm, and the rapidity of a pseudopodium-formation is increased by the peculiar " fountain currents " char- acteristic of most pseudopodia. As observed by Biitschli, an advancing stream of granules flows through the centre or axis of the growing pseudopo- dium, while near the tip back-running currents like the falling drops of water in a fountain surround the central stream (Fig. 47). " In the formation of a Fig. 47. Diagram finger-shaped pseudopodium of Amceba protcus." says f the movements of , ,. J , .-', the endoplasm gran- Biitschh, "it can be seen that the current which u ies in an advanc- traverses the axis of the pseudopodium and flows in s pseudopodium ,, . , , .. of Amceba proteus. away on all sides from its tip, comes to rest 'at a very short distance behind the tip, a circumstance which in any case is extremely favorable to the rapid outgrowth of the pseudopodium, in contradistinction to the relations that obtain in the drops of foam, since the protoplasm that has come to rest is heaped up and the pseudopodium grows in this way." 2 Rhumbler ('98) attempts to explain the formation of new ectoplasm and the increase in surface of an advancing pseudopodium through the hardening effect of water upori protoplasm, a fact which has long been recognized (Biitschli, Pfeffer). An advancing pseudopodium of Amoeba proteus, if properly fixed and stained, shows an advanced mass of endoplasm broken through the walls of ectoplasm. 3 The outer ectoplasm has a firm consistency, and, as Rhumbler dem- onstrated by treatment with diluted caustic potash (Fig. 48), may be isolated from the endoplasm. Nevertheless, it is converted into streaming endoplasm again. The Fig. 48. The ectoplasm (e) and gastric conversion of ectoplasm into endo- p' asra - which was earl y noted b y Engelmann ('79) and recently by Penard, Pfeffer, Verworn, Biitschli, and others, takes place accord- ing to Rhumbler at all times. It is particularly well shown in Amoeba limicola Rhumbler, or A. blattce Biitschli, where the eruptive pseudopodium incloses a definite portion of the old ectoplasm, which soon disappears and becomes lost in the endoplasm. Both Biitschli 1 Biitschli, loc. cit., English translation, pp. 310-311. 2 Loc. cit., p. 312. 8 Cf. Fig. 10, A, p. 36. 86 THE PROTOZOA and Rhumbler recognize that the longer the action of water is con- tinued upon the ectoplasm, the greater the stiffening; hence, Rhumbler argues, the new ectoplasm forming at the edge of the advancing pseudopodium is less resisting than elsewhere and the forward flow continues in one direction until the surface tension is equalized. New material for the advancing pseudopodium must be supplied from endoplasm, and this in turn from the posterior ectoplasm, so the assumption is made by Rhumbler that there is a continual change of Amoeba s protoplasm from ectoplasm into endoplasm, and from endoplasm into ectoplasm. Explanations of this nature, based upon purely physical laws of fluid substances, seem inadequate to explain all types of pseudopodia, the reticulate and long filamentous forms in particular. Up to the present time no satisfactory and comprehensive explanation has been made, and it should be recognized that the theories advanced still remain only working hypotheses. Hofer ('89) and Verworn ('91), and many others have demonstrated that an enucleated amoeboid mass soon comes to rest and assumes a spherical form. After a few days, movement recommences, and is interpreted by Hofer as an expression of the changes in surface tension. Such observations make it" prob- able that the chemical activity, which is constantly operating between the numerous substances which make up the protoplasm, plays an important part in pseudopodia-formation, and with our present imper- fect knowledge of these intra-cellular reactions, it is premature to settle upon any one cause, however suggestive and attractive it may appear, of this widely varied phenomenon. In many cases, especially among the Heliozoa, pseudopodia-motion approximates flagella-motion. In many of the shelled Rhizopoda (e.g. Arcella, or some species of Difflugid), the hyaline pseudopodia sway backward and forward like thick, slow-moving flagella, while in some Heliozoa (e.g. Artodiscns} this motion is much more energetic, causing the organism to dance about like a monad. The resemblance is more noteworthy and interesting from a theoretical point of view, because both the flagellum of Mastigophora and the axial filament of Heliozoa arise in the same manner in the endoplasm, and both are apparently connected with a "division centre," a central granule, which is analogous to the centrosome of metazoan cells. 1 C. THE NUCLEUS Nuclei are almost as varied in the different forms of Sarcodina as are the different types of the animals as a whole. In some cases, there is no well-defined nucleus, the chromatin being scattered in the l Cf. p. 271. THE SARCODINA 8/ form of granules throughout the entire cell, as in some of the Masti- gophora ; again, it is confined to a solid sphere without membrane or intra-nuclear vacuoles ; or, there may be a membrane and a single compact mass of chromatin which occupies the centre of the distinct nucleus, and is separated from the membrane by hyaline matter. In other cases, there may be two or more karyosomes or chromatin reservoirs, or there may be a great number of granules in the nucleus without the reservoirs \Amceba protens}. In some of the Rhizopoda (Englypha) and Heliozoa (Actinop/trys and Actinosph&rium\ the nucleus is strikingly similar to that of metazoan cells, consisting of chromatin in the form of a reticulum and a network of linin (Figs. 14 and 54). The number of nuclei is also quite variable, many forms having only one (Amoeba proteus, ActinopJirys, etc.) ; others, two (Amoeba bi~ nucleata, Arcella, etc.); while some have many, the giant Amoeba, Pelomyxa, having, according to Bourne ('91), about ten thousand, although, even with this large number, the proportion of nuclear substance to the total mass of the organism is about the same as in other cells (Bourne). In almost all of the shelled forms, a multiple number of nuclei is the rule, but in the many-chambered Reticulari- ida, every chamber does not possess a nucleus, the number of nuclei being smaller than the number of chambers, thus indicating that these forms are not colonies, but syncytia, or multinucleate cells. D. THE CONTRACTILE VACUOLE Contractile vacuoles are almost entirely absent in the marine forms (Reticulariida, Radiolaria), and in a few of the fresh-water forms of Sarcodina {Protamosba, Pelomyxa, Myxodictyum, Protogenes, etc.), but they are generally present in the Amoebida and Heliozoa, some- times two or three in one organism. The number of contractile vacuoles is quite variable. In most of the naked forms there is but one ; this, however, may be of large size, sometimes measuring one-quarter of the volume of the organism (Actinophrys, Actino- spharium). In the shelled forms, on the other hand, there are two or more (2-3 in Euglypha, 12 or more in Arcella). The position of the vacuole in the naked forms is also variable, but becomes fixed in the shelled forms and in the Heliozoa. In the shelled forms they sometimes lie in the middle zone about the edge of the granular region (Euglypha), sometimes around the periphery of the flattened body, while in other forms they are found now in one zone, now in another. In all cases, shortly before contraction, they come to lie close to the outer edge, and in some cases they form minute wart-like excrescences. 88 THE PROTOZOA Amoeba proteus, with its comparatively clear protoplasm and free- dom from pigments, is one of the most favorable objects for the study of the contractile vacuole. If a sufficiently high power is used, the formation and contraction of the vacuole and the expulsion of the con- tents to the exterior can be followed step by step. At first the vacuole lies near the nucleus, but as it grows, it becomes separated from the latter, and at the time of its contraction lies at the end of the body farthest from the advancing pseudopodia, at what is sometimes called the posterior end (Fig. 49, F). Its reappearance is always somewhere near its point of disappearance. While still small it is carried along by the streaming protoplasm back to a position near the nucleus, where it completes its development. The increasing weight of the growing vacuole causes it to lag behind the streaming granules and nucleus, until at its full growth it is widely separated from the latter organ. The vacuole may appear to move in the direction contrary to that of the protoplasmic streaming, although in reality it is quiescent; for while it remains in the field of the microscope, the main body of the animal moves well out of it, until the vacuole is surrounded only by the posterior end of the animal (G), which is reduced to a thin layer of granules and a hyaline layer of ectoplasm between the vacu- ole and the exterior. The granules later move away, passing around the vacuole, until finally there . is only a thin layer of hyaline plasm between the vesicle and the exterior. Shortly after this the vacuole bursts and disappears, in most cases a distinct bulge toward the outside preceding contraction. Contraction always begins on one side of the vacuole, and is carried across it toward the outer edge (ff). Stokes ('93) asserts that there is no bursting of the wall, but that minute pores are formed through which the contents of the vacuole are forced to the outside. In some instances the contents of the vacuole are not completely emptied, but as much as half may be left, the vacuole then rounding out to be carried back by the streaming plasm to the nucleus, where it completes its growth. In other cases the contents of the old vacuole may be entirely discharged, with the exception of a small quantity of liquid retained in exceedingly small vesicles, each of which may grow to some extent independently, although they ultimately fuse to form the new vacuole (_/). Thus the new vacuole does not necessarily re-form at the place of dis- appearance, but may be derived by the coalescence of a number of smaller ones which themselves are the remains of an old one. As many as six may unite in this manner to form the new vacuole (A, B, C.) These unite two by two in various parts of the plasm, and the last two may not fuse until in the neighborhood of the nucleus. THE SARCODINA 8 9 In addition to the contractile vacuoles the Sarcodina occasionally possess gas vacuoles, which were first made out in Arcella, but which GUI Fig. 49. Am-. ;mz> within the cyst are not amoeboid, but are provided with flagella. The swarmers soon lose their flagella, how- ever, becoming amoeboid, a condition in which they fuse together to form larger or smaller plasmodia. This fusion is characteristic of the Vampyrellidae and of My- cetozoa, but not of the Rhizomastigidae, where it has never been observed. There are many features in this theory of Klebs to recommend it. It affords a logical and satisfactory ex- planation of the relations of the Mycetozoa to the Sar- codina, and from the stand- point of the botanist points out the relation of this group to the colorless plants. The close connection of the Heliozoa with the Mastigophora is shown in other ways than by the transitional forms Dimorpha, Actinomonas, etc. The finer structure of the body of the sun-animalcula, the nucleus and archo- plasmic substances, show a degree of differentiation approached only by the Flagellidia and Metazoa, while the axial filaments are homologous with flagella. It may be pointed out, however, that Klebs' theory leaves unexplained the relatively simple nuclear struc- tures and nuclear processes of division in the Rhizopoda. This objec- tion is fatal to the view that the Rhizopoda are derived from the higher types of Heliozoa, and it must be admitted that they arose from much less specialized forms, perhaps from the Pseudosporeae or Fig. 58. Actinosph&rium Etch. Ehr. Section. m, membrane between ectoplasm and endoplasm ; nuclei ; x, axial filaments. THE SARCODINA 105 other flagellated forms, perhaps from forms like the Vampyrellidae. On the whole, there is no conclusive evidence to support the view that Rhizopoda are more primitive than Flagellidia, or vice versa. Their mutual affinities are very close, and together they stand as the most primitive forms of modern Protozoa. The relations of the Radiolaria to the Heliozoa are extremely close, and there is abundant evidence to show that the former were derived from the latter by the acquisition of a chitinous membrane between ectoplasm and endoplasm, and the retention of a gelatinous mantle like that of Splicemstrum (Haeckel). As first pointed out by Brandt ('85), the young Radiolaria pass through flagellated and amoeboid swarm stages, then through Heliozoa stages, until the definitive radiolarian structure is attained. Haeckel described the intermediate forms which are represented in this growth, as flagellate, amoeboid, ActinopJnys, Sphcerastrum, and Actissa, the last being the simplest of the Radiolaria. Although the external appearance of a radiolarian is strikingly similar to that of a heliozoon, there is no structure in Heliozoa to compare with the chitinous central capsule of the Radio- laria. Greeff('67, '71) described a membrane-like thickening between the endoplasm and the ectoplasm of Actinosph(Eriiim, and regarded it as homologous with the central capsule. Other observers, e.g. F. E. Schultze, Hertwig and Lesser, Biitschli, etc., have not seen it, and the latter, especially, considers Greeff's contribution of little value. However incorrect his interpretation may have been that Actinosphce- rium is a fresh-water radiolarian belonging to the Acantharia, Greeff was not mistaken in his observation, for an occasional specimen is found which shows such a membrane (Fig. 58). CLASSIFICATION CLASS I. SARCODINA. Naked or shelled Protozoa, characterized by the possession during adult life of movable or changeable processes of protoplasm, the pseudo- podia, which may be finger-form, reticulate, or ray-like, and which may or may not have axial filaments. Reproduction is brought about by simple division and by spore-formation. Subclass I. RHIZOPODA. Naked or shelled Sarcodina having pseudopodia of the lobose (finger-formed) or reticulate (anastomosing) type. The adult form is amoeboid ; the young forms are amoeboid or flagellated, and are produced by spontaneous division of the cell during active phases or during encystment. The adults in some cases fuse to form plasmodia. Order i. AMCEBIDA. Rhizopoda provided with lobose pseudopodia, with or with- out a shell, with one or more nuclei, and usually with a contractile vacuole. Suborder i. GYMNAMCEBINA. Naked forms of Amcebida having lobose pseudo- podia, and with or without nucleus and contractile vacuoles. Family i. AMOEBID-ZE. The pseudopodia are lobose, occasionally sharp pointed and branched. Genera: Amoeba, marine and fresh water; Paramceba Schau- dinn ('96); Protaviceba Haeckel ('66), marine and fresh water; Gringa IO6 THE PROTOZOA Frenzel ('92), lagoons; Gloidium Sorokin ('78). fresh water; Chcetoproteus, (Dinamceba Leidy) Stein ('57), fresh water; Trichosphcerittm Schneider; Pachymyxa Gruber; Hyalodiscus Hertwig and Lesser ('74), fresh water.; Plakopus F. E. Schultze (75), fresh water; Dactylosphara Hert. & Less. (74), fresh water: Chroinatella Frenzel ('92), fresh water; Stylamceba Frenzel ('92), fresh water; Saltonella Frenzel ("92), fresh water; Eikenia Frenzel ('92), fresh water; Pelomyxa GreefF ('74), fresh water; Amphizonella Greeff ('66), fresh water; Podostoma Clap. & Lach. ('58), fresh water; Arcnothrix Hallez ('85), cultures of Ascaris megalocephala. Suborder 2. THECAMCEBINA. Amrebida provided with a shell, with lobose pseudopodia, which may be sharp pointed and branched, and with one or more nuclei and contractile vacuoles. Family 2. Arcellidae. The shell is more or less membranous. Contractile vacuoles are numerous; the nucleus is single or multiple. Genera: Arcella Ehbg. ('38), fresh water, common ; Cochliopodium Hert. & Less. ('74), fresh water, common ; Pyzidicida Ehbg. ('38), fresh water; Pseiidochlamys Clap. & Lach. ('58), fresh water; Hyalosphenia Stein ('57). fresh water; Qitadrula F. E. Schultze (75), fresh water; Difflugia Leclerc ('15), fresh water, common; Lecquereusia Schlumberger, fresh water. Family 3. Euglyphidae. The shell is formed of regular plates of chitin, or of silica, and is often provided with spines. The pseudopodia are sharp pointed and often branching, but do not anastomose. Genera: Euglypha Dujardin ('41), fresh water; Trinema Duj. ('36), fresh water; Cyphoderia Schlumberger ('45), marine and fresh water; Campascus Leidy (77), fresh water; Nadinella Penard ('99). Order 2. RETICDLARIIDA. Rhizopoda with fine branching and anastomosing, or reticulate pseudopodia, forming an irregular network around the body, which may or may not have a shell. Shells, when present, are calcareous (rarely silicious) and provided with many pores (Perforina), or without pores (Imper- forina), and consisting of one chamber (Monothalamous), or of many chambers (Polythalamous). Suborder i. NUDA. Shell absent; the pseudopodia are reticulate, and the cell- body, in many cases, is apparently without a nucleus ; marine. Genera : Gymnophrys Cienkowski ('76) ; Protamyxa Haeckel ('68) ; Myxodictyum Haeckel ('68); Protogenes Haeckel ('64) ; Pontomyxa Topsent ('93). Suborder 2. IMPERFORINA. [With but few modifications, the following classifica- tion of the Reticulariida is taken from Brady ('84) and Lankester ('85), after Carpenter ('69)]. Shell-bearing forms; the shells are calcareous, solid, and without minute apertures, or they are made up of foreign particles cemented upon a chitinous base. They may have one, two, or many mouth openings, and are either monothalamous or polythalamous. Family i. Gromidae. The shell is membranous and in the form of a simple sac, with a pseudopodial aperture either at one extremity or at each end. The pseudopodia are long, branching, and anastomosing ; marine and fresh-water fqrms. Subfamily i. Monostomince. The shell has but one aperture. Genera: Gromia Duj. ('35); Lieberkuhnia Clap. & Lach. ('58). both found in fresh water, the former also marine ; Microgromia R. Hertwig ('74), fresh water : Platoutn F. E. Schultze ('75); Plectophrys Entz ('77); Pseiidodifflugia Schlumberger ('45), brackish and fresh water. Subfamily 2. Amphistotnince. With an aperture at each end of the shell. Genera : Diplophrys Barker ; Ditrema Archer (70) ; Amphitrema Archer ('70) ; Shep- heardella Siddall ('80). THE SARCODINA IO/ Family 2. Miliolidae. The shell is mono- or polythalamous, usually calcareous and porcellanous, but may be covered with sand. The polythalamous forms may be linear, spiral, or a combination of the two. Subfamily i . Nubecularince. The shell has an irregular and asymmetrical form, with the aperture or apertures variously placed. Genera : Calcituba Roboz ; Sqttammulina Schultze ('54) ; Nubecularia Dufrance. Subfamily 2. Miliolince. The shell is coiled, either symmetrically or asymmetri- cally, on an elongated axis, with usually two chambers to each convolution. During growth the shell-mouth is alternately at each end of the shell. Genera : Spiroloculina d'Orb. ('26); Biloculina d'Orb. ('26); Fabularia Dufrance; Miliolina Williamson ('58). Subfamily 3. Hauert nines. The shells are varied, the chambers being partly milio- line in their arrangement, partly spiral or linear. Genera : Hatter ina d'Orb. ('46); Articttlina d'Orb. ('46). Subfamily 4. PeneropliditKE. The shells are piano-spiral or cyclical, and bilaterally symmetrical. Genera: Peneroplis Montfort ('10) ; Orbiiolites Lamarck (1801) ; Orbiculina Lamarck ( ? i6) ; Cornuspira M. Schultze ('54). Subfamily 5. Alveolinince. The shell is spiral and elongated in the axis of the convolution ; the chambers are subdivided into secondary chambers. Genera : Alveolina d'Orb ('26). Subfamily 6. Keramospharince. The shell is spherical with the chambers in con- centric layers. Genera: Keratnosphcera Brady ('84). Family 3. Astrorhizidae. The shell is invariably composite, consisting of foreign particles, such as diatom-cases, spicules, sand grains, etc. It is usually large and single chambered, frequently branched or even radiate, with usually a single pseudopodial aperture at the end of each branch. Subfamily i. Astrorhizina. The shells have thick walls, consisting of sand or mud, lightly cemented together. Genera: Astrorhiza Sandahl ('57); Den- drophrya Wright ('61); Syringammina Brady ('84); Pelositia Brady ('79). Subfamily 2. Pilulinince. The shell consists of one chamber, the walls being thick and composed of felted spicules and fine sand. Genera : Pilulina Carpenter ('70) ; Bathy siphon Sars (71). Subfamily 3. Saccamminince. The chambers are nearly spherical, with thin walls composed of closely cemented sand grains. Genera: Saccammina Sars ('65) ; Psammosphcera Schultze ('75) ; Sorosphcera Brady ('79). Subfamily 4. Rhabdaniininince. The shell is composed of sand grains, firmly cemented together, and often with sponge spicules intermixed. They are tubular, straight, radiate, branched or irregular, but rarely segmented. Genera : Jaculella Brady ('79) ? Botellina Carpenter ('70) ; Haliphysema Bowerbank ('62) ; Marsipella Norman ('78) ; Rhabdammina Sars ('65) ; Aschemonella Brady ('79) i Rhizammina Brady ('79) ; Sagenella Brady ('79). Family 4. Lituolidae. The shell is arenaceous, and the septa which imperfectly mark the chambers are often incomplete or absent. Subfamily i . Litnolince. The shell is composed of coarse sand grains, is rough externally, and often labyrinthic. Genera: Rheophax Montfort ('08); Haplo- phragmium Reuss ('60) ; Coskinolina Stache ; Haplostiche Reuss ('61) ; Lituola Lamarck (1801) ; Bdelloidina Carter ('77). Subfamily 2. Trochatnminince . The shell is thin, and consists of a chitinous basis in which are embedded minute sand grains. The outside of the shell is smooth and often polished ; the interior is smooth or occasionally reticulate, but never labyrinthic. Genera : Thurammina Brady ('79) ; Ammodiscus Reuss ; Tro- chammina Parker and Jones ('59) ; Webbina d'Orb. ('39) ; Carterina Brady ('79); Hippocrepina Parker; Hormosina Brady ('79)- IO8 THE PROTOZOA Subfamily 3. Endothyrintz . The shell is more calcareous and less sandy than in the other Lituolidce, and the septa between the chambers are distinct. Genera : Nodosinella Brady ; Endothyra Phillips ('46) ; Polyphragma Reuss ; Bradyina Moll. ('78); Stacheia Brady ('76). Subfamily 4. Loftusince. The shell is large, lenticular, spherical, or fusiform, and deposited either in concentric layers or spirally. The chambers are occupied to a large extent by an excessive enlargement of the arenaceous cancellated wall. Genera : Cyclammina Brady ('76) ; Loftusia Brady ('69) ; Parkeria Carpen- ter ('69) . Suborder 3. PERFORINA. The shell wall is perforated by numerous minute open- ings through which the pseudopodia can pass as well as through the main openings. Family 5. Textularidae . The shells of the larger species are arenaceous, either with or without a calcareous matrix ; the smaller forms are hyaline and conspicuously perforated. The chambers are arranged in alternating series, spirally or without apparent order. Subfamily i. Textularituz . The shells are typically bi- or tri-serial, and are often dimorphous. Genera: Textularia Dufrance ('28); Bigenerina d'Orb. ('26); Verneuilina d'Orb. ; Cuneolina d'Orb. ('39) ; Pavonina d'Orb. ('26) ; Valvulina d'Orb. ('26) ; Chrysalidina d'Orb. ('46) : Tritaxia Reuss ; Claimlina d'Orb. Subfamily 2. Buliminitue. The shells are typically spiral, the weaker forms are more or less bi-serial. The main aperture is not round, but elliptical, comma- shaped, etc. Genera : Virgulina d'Orb. ('26) ; Bulimina d'Orb. ('26) ; Bolivina d'Orb. ; Bifarinia Parker and Jones. Subfamily 3. Cassidulinece. The shell consists of a series of alternating segments more or less coiled. Genera: Cassidulina d'Orb. ('26) ; Ehrenbergina Reuss. Family 6. Chilostomellidae. The shell is calcareous, finely perforate, and polythala- mous. The segments follow each other from the same end of the long axis, or alternately from the two ends, or in cycles of three, which are more or less em- bracing. The aperture is a curved slit at the extremity of the final segment. Genera : Ellipsoidina Seguenza ; Chilostomella Reuss ; Allomorphina Reuss. Family 7. Lagenidae. The shell is calcareous and very finely perforated; it is monothalamous or polythalamous. In the latter the chambers may be joined together in a straight, curved, spiral, or branching series. The aperture is ter- minal, and may be simple or radiate. The shell is not complicated by inter- septal skeletons or by canal systems. Subfamily i. Lagenince. Shell monothalamous. Genera: Lagena Walker and Boys (1784) ; Nodosaria Lamarck ('16) ; Lingulina d'Orb. ('26) ; Vaginulina d'Orb. ('26); Rimulina d'Orb. ('26); Frondicularia Defrance ; Margimilina d'Orb. ('26), etc. Subfamily 2. Polymorphinince. The segments composing the shell are arranged spirally or irregularly around the long axis ; they are rarely biserial and alter- nate. Genera : Polymorphina d'Orb. ('26) ; Uvigerina d'Orb. ('26) ; Sagrina Parker and Jones. Subfamily 3. Ramulinince. The branching shell is composed of long tubulariform tubes. Genera : Ramulina Rupert Jones. Family 8. Globigerinidae. The shell is free, calcareous, and perforated. The con- spicuous shell-aperture may be single or multiple. There is no supplementary skeleton or canal system. The animals are normally pelagic in habit. Genera : Globigerina d'Orb. ('26) ; Orbicnlina Lam. ; Hastigerina Thompson ('76) ; Can- deina d'Orb. ('26); Pullenia Park. & Jones ('62); Spharoidina d'Orb. ('26). Family 9. Rotalidae. The shell is calcareous, perforated, free, or adherent ; it is typically spiral in form, but irregular forms may be outspread or flaring, acervu- THE SARCODINA 1 09 line or irregular. Some of the higher types have double walls, with supple- mental skeleton and a canal system. Subfamily i. SpirillincE. The shell is a flat spiral, without septa; it may be free or attached. Genera: Spirillina Ehbg ('41)- Subfamily 2. Rotalince. The shell is spiral, rotaliform, and rarely evolute or ir- regular. Genera : Discorbina Lamarck ('04) : Planorbulina d'Orb. ('26) ; Truncatulina d'Orb. ('26) ; Anomalina d'Orb. ('26) ; Rotalia Lamarck (1801) ; Calcarina d'Orb. ('26) ; Patellina Williamson ('58) ; Carpenteria Gray ('58) ; etc. Suborder 4. TINOPORIN.3L. The shell consists of irregularly heaped chambers, usually with a more or less spiral primordial portion ; a main pseudopodial aperture is usually absent. Genera: Tinoporus Carpenter ('57); Polytrema Risso ('26) ; Gypsina Carter ; Thalamopora Roemer ; Aphrosina Carter. Family 10. Nummulinidae. The shell is calcareous and finely tubulated ; it is typically polythalamous, free, and symmetrically spiral. The higher forms pos- sess a supplementary skeleton and a well-developed canal system. Subfamily i. FusuliHtnce. The shell is bilaterally symmetrical, with chambers extending from pole to pole, so that each convolution completely incloses the preceding whorl. The septa between the chambers are single as a rule. Genera: Fusulina Fischer ('29); Schwagerina Mbller ('77). Subfamily 2. Polystoniellince. The shell is bilaterally symmetrical and nautiloid. The simpler forms are without supplemental skeleton ; the more complex forms have a skeleton, and canals leading to the outside at regular intervals along the external septal depressions. Genera : Polystomella Lamarck ('22) ; Nonionina d'Orb. ('26). Subfamily 3. Nummulitina. The shell is lens-shaped or flattened. Genera: Archeodiscus Brady; Amphistegina d'Orb. ('26); Operculina d'Orb. ('26); Nummulites Lamarck ( 1 80 1 ) ; Heterostegina d'Orb. ('26). Subfamily 4. Cycloclypeina. The shell is flat, with a thickened centre, or lens- shaped, and consists of a disc of chambers arranged in concentric annuli with peripheral thickenings. The septa are double, and furnished with a system of interseptal canals. Genera: Cycloclypeics Carpenter ('56) ; Orbitoides d'Orb. Subclass II. HELIOZOA. These are naked or shelled forms of Sarcodina of typi- cally spherical form, with but little tendency to change form by amoeboid mo- tion. The pseudopodia, radiating from all parts of the body, are fine and ray-like, rarely changeable, and usually provided with an axial filament. Order i. APHROTHORACIDA. Heliozoa, without a skeleton, but provided with a more or less developed power of amoeboid motion, and with plastic (myxopo- dia) or stiff (axopodia) pseudopodia, the latter possessing axial filaments. Genera: Vampyrella Cienk. ('65); Nuclearia Cienk. ('65); Monobia A. Schneider ('78) ; Myxastrum Haeck. ('70) ; Actinophrys Ehr. ('30) ; Actino- spharium Stein ('57) ; Actinolophns F. E. Schultze ('74). Order 2. CHLAMYDOPHORIDA. Heliozoa, with a soft gelatinous or felted fibrous covering. Genera: Heterophrys Archer ('69); Spharastrum Greeff ('73); Astrodisculus Greeff ('69). Order 3. CHALARATHORACIDA. Heliozoa, with a silicious coating composed of .separate and loosely-jointed spicules. Genera: Pompholyxophrys Archer ('69) ; Raphidiophrys Archer ('70) ; Pinacocystis Hert. & Less. ('74) ; Pi- naciophora Greeff (^73) ; Acanthocystis Carter ('63) ; Diplocystis Pdnard ('90) ; Cienkowskya Schaudinn; Wagner ella Mereschkowsky ('81). Older 4. DESMOTHORACIDA. Heliozoa, with a shell of one piece perforated by numerous openings. Stalked or unstalked forms. Genera: Orbulinella Entz ('77) ; Clathrulina Cienk. ('67). HO THE PROTOZOA Subclass III. RADIOLARIA. Marine forms of Sarcodina. similar to Heliozoa in having ray-like pseudopodia (axopodia and myxopodia), but provided with a chitinous capsule which incloses the nuclei. They may or may not have. a skeleton ; when present the skeleton is formed of acanthin or of silica. The group is subdivided into 4 legions, 20 orders, 85 families, 739 genera, and 4318 species. (Haeckel, 1885.) Legion i. SPUMELLAPIA (or PERIPYLEA) . The central capsule is perforated by numerous fine pores. A skeleton may or may not be present. Order i. COLLOIDIDA. Without skeleton. Families: Thalassicollidae (solitary forms) ; Collozoidae (colonial). Order 2. BELOIDIDA. The skeleton consists of loose siliciolis needles. Families : Thalassosphaeridae (single); and Sphaerozoidae (colonial). Order 3. SPH7EROIDIDA. The skeleton consists of from one to many concentric globular shells. Families : Liosphaeridae (single) ; Collosphaeridae (colonial) ; Stylosphaeridae (single) ; Staurosphaeridae (single) ; Cubosphaeridae (single) ; Astrosphaeridae (single). Order 4. PRUNOIDIDA. With ellipsoidal to cylindrical latticed shells and similar central capsule. Families : Ellipsidae ; Druppulidae ; Sponguridae ; Artiscidae ; Cyphinidae ; Panartidae ; Zygartidse. Order 5. DISCOIDIDA. Shell and central capsule are discoidal or lenticular. Families : Cenodiscidae ; Phacodiscidae ; Coccodiscidae : Porodiscidae ; Pylodis- cidae ; Spongodiscidae. Order 6. LARCOIDIDA. The skeleton is irregularly lenticular or discoid. Families: Larcaridae ; Larnacidae ; Pylonidae ; Tholonidae ; Zonaridae ; Lithelidae ; Streb- lonidae ; Phorticidae ; Soreumidae. Legion 2. ACANTHARIA (or ACTIPYLEA). The skeleton is formed of acanthin arranged in radiating spines, usually twenty in number. Order 7. ACTINELIDA. The spines are more than twenty in number. FamilLes : Astrolophidae ; Litholophidae ; Chiastolidae . Order 8. ACANTHONIDA. With twenty spines arranged according to Miiller's law (four equatorial, eight tropical, and eight polar) . Families : Astrolonchidae ; Quadrilonchidae ; Amphilonchidae. Order 9. SPH^ROPHRACTIDA. With twenty equal quadrangular spines and a complete, fenestrated shell. Families : Sphaerocapsidae ; Dorataspidae ; Phrac- topeltidae. Order 10. PRUNOPHRACTIDA. With ellipsoidal, flat, or double-coned shell, through which twenty spines radiate according to Muller's law. Families : Belonaspidae ; Hexalaspidae ; Diploconidae. Legion 3. NASSELLARIA (or MONOPYLEA) . The skeleton is silicious and rarely absent. The central capsule has a single, limited, perforated area at one pole ; the extracapsular plasm has no pigment. Order 1 1. NASSOIDIDA. Monopylaria without a skeleton. Families: Nasselidae. Order 12. PLECTOIDIDA. A complete latticed shell is never formed, but the skele- ton consists of three or more spines radiating from one point below the central capsule, or from a central rod. Families : Plagonidae ; Plectanidae. Order 13. STEPHOIDIDA. The skeleton consists of one or two fused rings which may be connected by a loose network. Families : Stephanidae ; Semantidae ; Coronidae ; Tympanidae. Order 14. SPYROIDIDA. The skeleton consists of a single sagittal ring and a latticed shell which is furrowed in the sagittal plane. Families : Zygospyridae ; Tholospyridae ; Phormospyridae ; Androspy ridge. Order 15. BOTRYOIDIDA. The skeletons are similar to the preceding, but orna- mented by one or more wing-like processes. Families : Cannobotryidae ; Litho- botryidae ; Pylobotryidae. THE SARCODINA III Order 16. CYRTOIDIDA. Similar to the preceding, but without the sagittal fur- row. The skeleton is helmet-shaped. Families : Tripocalpidae ; Phaenocalpidae ; Cyrtocalpidae ; Tripocyrtidae ; Anthocyrtidas ; Sethocyrtidae ; Podocyrtidas ; Phormocyrtidae ; Theocyrtidas ; Podocampidae ; Phormocampidae ; Lithocampidas. Legion 4. PHJEODARIA (or CANNOPYLEA). The central capsule has a double membrane, with a spout-like main opening at one pole, and frequently with accessory openings on each side of the main axis at the opposite pole. The central capsule may be multiple in number. There is always a pigmented mass on the outside of the central capsule (the phceodium) and covering the main opening. The skeleton, which is rarely absent, is silicious and always outside of the central capsule. Order 17. PH^OCYSTINIDA. Skeletal structures may or may not be present; the central capsule is the centre of the spherical body. Families : Phaeodinidae ; Cannoraphidae ; Aulacanthidae. Order 18. PH^OSPHERIDA. The skeleton is a simple or a double latticed cover- ing; the central capsule is in the centre of the shell. Families: Orosphaeridse ; Sagosphseridae ; Aulosphaeridae ; Cannosphaeridae. Order 19. PH-ffiOGROMIDA. Radiolaria provided with a simple latticed shell, having a mouth opening at one (the main) pole. The central capsule is in the aboral half of the shell. Families: Challengeridae ; Medusettidae ; Castanel- lidae ; Cercoporidae : Tuscaroridae. Order 20. PELffiOCONCHIDA. The shell consists of two latticed valves, one dorsal, the other ventral (right and left according to Butschli). Families: Con- charidae ; Cselodendridae ; Caslographidse. SPECIAL BIBLIOGRAPHY III Brady, H. B. Report on the Foraminifera dredged by H..M. S. Challenger: Chal- lenger Reports, Zool., IX., 1882-4. Brandt, K. Koloniebildenden Radiolarien (Sphaerozreen) des Golfes von Neapel : Fauna and Flora, 1885. Butschli, 0. Protozoa; Sarcodina. In Bronn's Klassen und Ordnungen des Thierreichs. Leipzig. 1883. Carpenter, W. B. Introduction to the Study of the Foraminifera: Ray Society, London, 1862. Haeckel, E. The Radiolaria: Challenger Reports, Zool., XVII. and XVIII., 1888. Hertwig, R. Der Organismus der Radiolarien. Denkschrift. Jenaisch. Akad., II., 1879- Hertwig und Lesser. Ueber Rhizopoden und denselben nahe stehende Organismen : Arch. f. mik. Anat. Bd. X., Suppl., 1874. Leidy, Jos. Fresh-water Rhizopods of North America. Washington, 1879. Pnard, Eug. FJudes sur les rhizopodes d'eau douce : Mem.phys. hist. nat. Geneve, XXXI., 1890. * S CHAPTER IV THE MASTIGOPHORA THE Mastigophora are provided with a motile apparatus in the form of flagella, which may vary in number from one to many. In the majority of cases, the body is of well-defined and constant form, and covered with a cuticle, menibrane, or shell. They abound in infusions, in stagnant pools, in clear water, and in the sea, while many of them are found as parasites in higher animals, where they live in the cavities and cells of the body. In this class are found many diverse types of unicellular organisms, including, at one extreme, primitive forms whose allies are undoubt- edly among the bacteria and the lowest plants (monads), at the other extreme, colonial forms, which in the complexity of their structure and functions are little lower than some of the Metazoa and Metaphyta. It includes forms whose bodies are naked ; others that are clothed with complex membranes, or incased in chitinous, silicious, or cellu- lose shells. It includes organisms with very different methods of food-taking : in some forms the food, like that of the green plants, consists of products made from simple compounds by the organism itself ; in others, the food, like that of the fungi, consists of dissolved organic matters ; and in still others, the food, as in the higher animals, consists of solid particles of proteid and other matters. Notwithstanding these many structural and functional differences, there are some well-defined structural characteristics according to which the Mastigophora may be subdivided into a number of more or less homogeneous groups. These groups are the Flagellidia, Dinoflagellidia, and Cystoflagellidia. The first comprises the least homogeneous forms ; they consist usually of minute cells with a simple naked body, which may become more or less amoeboid, and with one, two, or several flagella. In some cases, there is a compli- cated cell-membrane, in others a shell, while colony-formation is fre- quently seen. The Dinoflagellidia are distinguished by the presence of one or two furrows, in which the flagella find their origin, one to pass around the organism transversely, the other to vibrate freely in the surrounding water. The majority are covered by a cellulose shell, consisting frequently of several plates. The Cystoflagellidia, a group consisting of only two genera, Noctiluca and Leptodiscus, are THE MASTIGOPHORA distinguished by the peculiar parenchymatous structure and by the presence of a tentacle and a collar. A. PROTOPLASMIC STRUCTURE The alveoli, forming the structural basis of the protoplasm, vary in size from minute and scarcely visible spaces to large vacuoles. In the majority of forms, they are arranged in a typical outer layer (Rin- denschichf) of small-sized alveoli, surrounding an inner mass of larger ones (e.g. Chilomanas}. The protoplasm is not equally dense in all Fi g- 59- Proter ospongia H&ckeli S. K. [S. KENT.] cases, but, as in the Rhizopoda, may be of variable consistency. It may be so soft and flexible that, as in Amoeba, the periphery will give way, and pseudopodia may be formed at any point in response to local changes in the surface tension (Euglenoids and forms of Asta- sia). There is but little tendency to the differentiation into zones, so frequently seen in Rhizopoda, and only rarely is there a differen- tiation into ectoplasm and endoplasm (Mastigamceba). Klebs ('92) distinguishes two types of peripheral structures, the periplasts and outer coats, stalks being included with the latter. The periplasts include all cuticular differentiations which are a living 114 THE PROTOZOA part of the organism, and even diverse modifications of protoplasm, such as the fine peripheral layer of alveoli {Pcllicula of Biitschli), and the complex membranes of Englena and Astasia (cf. Fig. 10, B\ The outer coatings as in all Protozoa, serving probably for the pur- pose of protection, include houses and tests of all kinds which are not a living part of the animal. In many cases they are simply jelly- like coverings, which in many colony-forms also serve to keep the individuals together (Uroglena, many Choanoflagellida, Fig. 59; see also Fig. 25, p. 56). In other cases, the gelatinous mantle becomes a tube, into which the organism can completely withdraw (some Choanoflagellida). In still other cases, the jelly is apparently hardened into a well-defined goblet or beaker-shaped cup with the consistency of c\\\\m(Codonceca, Epipyxis, Dinobjyon, Salpingceca, etc.). The relations of the firm case to the gelatinous mantle are shown in forms like Codonceca, where the chitin-like urn-shaped cup may become gelatinous (Fig. 60). The organisms are attached to the bottoms of such goblet-shaped cups by a protoplasmic process, and in no case does the cup fit the organism as tightly as a membrane. Colony- forms also are frequent in these types, arising, in the simplest cases, by a young individual attaching itself to the edge of the parent test and there secret- ing its own covering (Dinobiyon, Fig. 61). The majority of these colonies are attached, but Dinob- ryon is a free-swimming form, usually found in the clearest waters. Shells are distinguished from tests or houses by the fact that they completely inclose the animal, the so-called mouth-opening where the flagellum is in- serted being the only aperture. Both tests and shells are usually transparent and colorless, although tne y ma y ke colored by the presence of iron, as [JAMES-CLARK.] in Trackelomonas, Rhipidodendron, etc., where the shells, when present in any quantity, give a distinctly red color to the water. The simplest shells are the cellulose cover- ings of many Phytoflagellida, which, although lifeless, have the same general appearance as membranes. The shell, which is frequently protected by sharp spines {Trachelomonas\ may be separated from the plasm by a considerable space. It is bivalved in Phacotus, the two parts being easily separated (Fig. 62). In one form only, Dis- tephanus speculum Stohr, there is a silicious skeleton which recalls the latticed skeletons of Radiolaria (Fig. 63). The most highly differentiated of these outer coatings are found in the Dinoflagellidia, where the cellulose shells are often composed of separate plates fitted together with the greatest nicety and often Fig. 61. Dinobryon sertulana Ehr. [STEIN.] , chromatophore ; e, eye-spot or stigma. THE PROTOZOA complicated by the presence of spines, wing-like processes, and other appendages, or they may be pitted by minute depressions or pores. After the death of the animal the plates can, as a rule, be separated by gentle pressure. The substance of the shell is not true plant cel- lulose, but a modification, the exact nature of which has not been definitely determined. The furrows in the shells of the Dinoflagellidia, in which the two flagella lie, are perhaps the most characteristic feature of these forms. B Fig. 62. Phacotus lenticularis Ehr. [BiJTSCHLl.] A. Individual within its bivalved shell. B. Spore-forming individual. One runs across the organism, while the other, which may often, how- ever, be obliterated, is at right angles to this, usually in the direction of the longitudinal axis. There may be one, two, or many transverse furrows, the number determining the family to which the organism belongs. In the genus Hemidinium, the single transverse furrow begins on the ventral side and runs as far as the middle of the dorsal side, where it disappears. In the genera Gymnodininm, Glenodinium, and Peridinium, it runs completely around the organism ; while in Ceratium it may be* broken in its course (Fig. 64). The longitudinal furrow, on the other hand, is invariably confined to the ventral side, usually to the lower half, but in some cases (Glenodinium, Peridinium} it traverses the cross furrow and stretches some distance along the THE MASTIGOPHORA 117 upper half of the shell. The two flagella which lie in these grooves pass from the body-plasm to the outside through a distinct aperture in the shell, which Stein ('78) called the " mouth-opening " ; but as it serves no purpose in food-taking, Biitschli has substituted the better term of fiagellum fissure. The protoplasm of the Mastigophora usually contains chromato- phores in which one or more deeply staining bodies the pyrenoids may be found, and these are frequently covered by a shell of amylum or starch. Paramylum, a food product allied to starch, and various particles of oil-like substance are widely distributed. The latter are frequently so nu- merous that the cell is fairly filled with them. Upon dif- fluence, these oil-like bodies run together, forming glob- ules of large size; or they become finely divided, giving to the surrounding liquid the appearance of an emulsion. Not infrequently the oils have a characteristic odor and taste, Comparable to the SCent Of Oils Fig. 63. Distephanus speculum Stohr. [BORGERT.] Of plants' (Urogleiia ameri- A - Lateral view of skeleton. B. Surface view. cana, Symira uvella, Fig. 65). 1 Chromatophores are widely distributed among the various Flagel- lidia. They consist of clearly defined, thickened bodies, usually of definite size and shape and of different shades of green, yellow, and brown. Clear green chromatophores, colored by chlorophyl as in the plants, occur in Euglenidae, Peranemidae, Chlamydomonadidae, and Volyocina. Yellow chromatophores (colored by diatomin, as in Diatomaceae) occur in Chrysomonadidae, Cryptomonadidae, among the Flagellidia, and possibly in some Dinoflagellidia; but the yellow color, when present in the latter group, frequently shades off into brown. Bergh ('81), Klebs ('84), and others regarded the coloring matter of the Dinoflagellidia as pure or slightly mixed diatomin, which supported the popular view that the Diatomaceae and the Dinoflagellidia are closely related. Schutt ('90), however, who has made the most complete study of the coloring matter in these B Cf. p. 62. THE PROTOZOA forms, disproved this view by showing that the coloring matter is quite distinct from diatomin and is peculiar to the Dinoflagellidia. He suc- ceeded in extracting three substances : ( I ) pJiycopyrrin, similar to the brownish red coloring matter of the Florideae and Phaeophycaceae among the plants, and like this, soluble in clear water ; (2) peridinin, like chlorophyl soluble in alcohol, but of quite different spectrum ; and (3) cJiloropJiyllin, a substance more like chlorophyl, but difficult to isolate. The shape and size of the chromatophores vary considerably in different species, but are fairly constant for the same species. They increase by simple division. The pyrenoids, which seem to be the centre of starch formation, are sometimes quite naked (Euglena\ Fig. 64. A. Gymnodinium ovum Schiitt. B. Peridinium divergens Efir. f t transverse furrow with (A) flagellum. [SCHUTT.J sometimes covered by a shell of paramylum, which apparently differs from starch only in its reaction to iodine. The paramylum granules are round, rod-like, or ring-form bodies. Pure starch is also recorded as a product of non-colored, saprophytic forms (Chilomonas, Poly- toma, etc.). In many of the Mastigophora, especially in those holding chromatophores, there may be an intense red coloring matter, in the form of fine drops, scattered throughout the protoplasm. These consist of oil particles impregnated with a deep red pigment, hcemato chrome, and the same substance is found in the so-called "eye-spots," or stigmata, which are supposed to be more sensitive to light than other parts of the protoplasm, although Engelmann's ('82) results show that, in Eiiglena at least, the clear plasm just in front of the stigma is more definitely involved. In many cases the structures accompanying the stigmata are so strikingly analogous to the visual organs of higher THE MASTJGOPHORA 119 forms that there is apparently good reason for supposing them to play a similar physiological role. In the green flagellates there are often distinct concretions, regarded by some observers as lenses ; and if Pouchet ('86) is correct, a still more striking differentiation is found among the Dinoflagellidia. The so-called eye of Gymnodinium con- sists of a transparent, highly refracting lens, rounded at its free extremity, and always directed forward (Pouchet). The inner sur- face is embedded in a hemispherical, cap- like mass of red or black pigment, which Pouchet considered achoroid. The lenses develop by the union of from six to eight refringent corpus- cles, while the organ- ism is still encysted or while undergoing fission. The choroid likewise results from the union of several of the pigment gran- ules. Considerable doubt has, however, been thrown upon these observations subsequent i ob- Fig. 65. Synura uvella Ehr. Each individual of the colony is surrounded by a gelatinous membrane, and possesses two chromatophores (c) and a nucleus by servers. Other inclusions of ' interest are the thread-like structures which are common among holotrichous ciliates, and which occur sporadically in other Protozoa. Among the Mastigophora they are found in only two cases (Gony- ostomum Blochmann and Polykrikos Biitschli). In the former they are trichocysts similar to those of the ciliate Paramcecium and allied forms, but in the latter they are true nematocysts, comparable to those of the Coelenterata. B. THE FLAGELLA The most characteristic part of a flagellate is its motile organ, the flagellum. This consists of a vibratile filament usually tapering to a fine point, although in some cases (e.g. in all Choanoflagellida) it is iCf. Penard ('88). I2O THE PROTOZOA of one thickness throughout. In either case the base is inserted in the vicinity of a pharyngeal depression and usually at the end of the body. There is good reason to believe with Klebs, Frenzel, Bloch- mann, and others, that like the axial filaments of the pseudopodia in Heliozoa and Radiolaria, the flagellum originates at or near the nuclear membrane, and does not consist exclusively of the outer or peripheral plasm. Dallinger ('78) asserts that the newly forming flagella are smooth and uniform, arising in or near the nucleus. Fischer ('94), however, claims that many of them are provided with branches like the cilia of a typhoid germ. He finds others which are not vibratile throughout their entire length, but are rigid and uniform for a certain distance and then taper to the extremity. Such flagella resemble a whip stock and its lash, the relative proportion of stock and lash varying in different flagella, the stock sometimes running nearly to the end, and again only a short distance from the body. Other forms, especially in the Dinoflagellidia, have spirally rolled flagella of various kinds, while some have flattened or band forms (Pcridiniitm tabnlatum and P. divergens}. A difference of opinion exists as to the ability of the organism to absorb or retract its flagellum into the body-protoplasm. Most observers agree with the early observation of Dujardin that there is a close relation between pseudopodia and flagella, numerous observa- tions having been recorded of cases where, under certain conditions, the pseudopodia change into swinging flagella, and flagella into pseu- dopodia. There is no doubt that flagella can be absorbed after changing to pseudopodia. Whether the fully formed flagella can be changed over into plastic material and then withdrawn, is still a sub- ject of dispute; Fischer ('94) holds that they are invariably discarded upon irritation, and Schiitt ('95) shows that the longitudinal flagellum in the Dinoflagellidia is thrown off upon irritation, while the horizontal flagellum is flattened into a band form. A general rule, therefore, cannot be formulated in regard to the disposal of flagella. In some cases they are absorbed ; in others, thrown off. The action of the flagella varies with the type of structure. In the simple, straight, or tapering forms the tip moves in a circle while waves pass from the base to the extremity. In the whip-like flagella the basal portion moves back and forth or in a circle, while the distal region vibrates or undulates like the snapper of a whip. The band- formed flagella move by simple undulations. The position of the flagella is extremely variable. When there is but one, it is found at the anterior end of the cell, that is, the end which is directed forward when in motion. When there are two fla- gella, they may both be directed forward (Chilomonas, Cryptomonas, etc.), and may be of equal (Cryptomonas, etc.) or unequal length THE MASTIGOPHORA 121 (Uroglena, etc.). Again, they may be of equal length but turned in opposite directions (Bodo, etc.). When there are numerous flagella, they may be distributed about the body, regularly or irregularly or aggregated at certain points (JMulticilia^ Tetramitus, etc.). In the Dinoflagellidia, the longitudinal flagellum, a long, fine thread, is invariably directed backward or forward, while the other, the trans- verse, lies around the body in an equatorial groove. This flagellum has a simple undulating motion resembling a row of moving cilia, for which it was at first mistaken. 1 So strictly does the transverse flagel- lum adhere to the usual direction of motion, that even when the groove is absent, as in Prorocentrum, where the flagellum no longer surrounds the body, the motion is retained, the flagellum being directed outward from the end of the body for a short distance and then turned at right angles to form a circle, with the customary undulatory motion, as though still encircling the body (Fig. 66). With the exception of the Choano- flagellida, which swim like a spermatozoon with the flagellum behind (James-Clark, '66), the Mastigophora swim with the flagellum in advance. The forward move- ment of most flagellated organisms is, therefore, exceedingly difficult to inter- pret. It is very curious to see the com- U O paratively large body of Peranema, for Fig- 66. Primitive forms of Dino- example, drawn steadily forward by the ^nidia. [BUTSCH^J > J a, Prorocentrum micans Ehr. d, minute tip of its rather long flagellum. Exuviaiia lima Ehr. No satisfactory mathematical demon- stration of the application of the force necessary to produce this motion has been given. Lankester ('91) compared it to the force produced by a man's arm and hand when swimming upon his side ; Biitschli ('83) offered a simple and apparently reasonable explanation, showing that the resistance, which is directed at right angles to the advancing undulation, can be reduced, through the parallelogram of forces, to a force of rotation and one of translation, but Delage ('96) holds that while this explanation is perfectly con- sistent with the mechanism of certain mechanical contrivances, it is incompatible with the structure of the flagellate body, and that the explanation is much more complicated. Delage's interpretation involves the principles of conic sections, the resisting force being 1 Hence the name of the group, Cilioflagellata, tively recent date. which was in use until a compara- 122 THE PROTOZOA applied in a very indirect manner, and he calls the resulting move- ment " conical translation." l Pig. 67. Phalansterium digitatum St. [S. KENT.] / Collared cells. A peculiar pseudopodial process, the collar, is found in one order of the Flagellidia, the Choanoflagellida. This collar, which forms a cup around the base of the flagellum, is extremely thin, delicate, and transparent, and like a pseudopodium can be altered in shape and 1 Cf. Delage ('96), p. 305, for a very full discussion. THE MASTIGOPHORA 123 either wholly or partly withdrawn into the protoplasm of the cell. It is occasionally so small and inconsiderable that, as in Phalansterium, it can have little or no use (Fig. 67). Again, it may be fully or even twice as long as the body. In shape it is either like a bowl or else like a truncated cone (Fig. 68, A, C). There may be two of the collars, one within the other (Diplosiga Frenzel, B). Butschli described a vacuole which appeared to him to move rapidly around the base of the collar and to disappear for a short time when a food particle A Fig. 68. Types of collars. A. Codosiga pulcherrimus Jas. Cl. [J.CLARK.] B. Dip- losiga socialis Frenz. [FRENZEL.] C. Salpingasca marinus Jas. Cl. [J. CLARK.] Fig. 69. A Choanoflagellate type. [FRANCE.] c, collar; m, swinging mem- brane. is taken in. Entz ('83) and France ('94) claim, however, that this " mouth-opening " is not a vacuole at all, but the edge of a swinging membrane. According to their view the collar is not a continuous structure with an unbroken wall, but is like a conical roll of paper with a free edge capable of motion (Fig. 69). 124 THE PROTOZOA C. THE NUCLEUS Inclosed in the protoplasm in all Mastig'ophora is a more or less clearly denned nucleus. It is variable in position, and although a multiple number may occur, is generally single. The structure also k ' B C E D Fig. 70. Nuclear division in Noctiluca miliaris. A. The first changes of the chromatin from the large karyosome condition; concentration of the substance of the divi- centre (s). B. Further disintegration of the chromatin and arrangement of granules to form the chromosomes (cK). imphiaster and completed chromosomes. D. The central spindle in the hollow of the nucleus; the nuclear plate of mosomes is thus wrapped around the division-centre. E. A section through the centre of the long axis of the division- e before division of the chromosomes. F. A section through the dividing chromosomes, c, the centrosome with iting mantle-fibres. is extremely variable. The chromatin may be either in the form of granules distributed throughout the cell and not confined by a distinct nuclear membrane ( Tetramitus\ or the granules may be aggregated without a membrane (Ckilomonas}, or again, the chromatin may be THE MASTIGOPHORA 12$ inclosed by a definite membrane (euglenoids and the majority of Mastigophora ; cf. Figs. 14, C, D, E, and 10, B). Again, it may be in the form of a homogeneous mass in which no granular structure can be seen (many Phytoflagellida), or, as in many Rhizopoda, it may be massed in several such aggregates (Noctiluca). Still another arrange- ment is seen in the Dinoflagellidia, where the chromatin is arranged in the form of a twisted thread or threads. Finally, in some forms the resting nucleus closely resembles that of the Metazoa in having a linin network in which the chromatin granules are suspended. An integral part of the nucleus is the so-called " nucleolus," which, however, is not analogous to the nucleolus of the Metazoa, but functions as a sphere or the division centre during mitosis. 1 Nuclear division in all forms of Mastigophora may be regarded as more or less simplified mitosis, or indirect division. In the simplest types the chromatin masses merely draw out and divide into equal parts, but in the more complicated types, the process closely resembles that in the Metazoa, the complete mitotic figure consisting, as in the higher forms, of chromosomes, mantle-fibres, centrosomes, and spheres (Fig. 70). D. FOOD-TAKING Closely dependent upon the mode of living is the manner of taking food. Some forms, which live in foul water, are saprophytic like the colorless plants, and absorb, through the body walls, the substances which are dissolved out of decaying vegetable matter. Those which live in pure and clear water generally have chromatophores, colored by chlorophyl, diatomin, or some allied substances, and have the power of manufacturing their food from carbon dioxide, water, and salts; like the green plants, their nutrition is holophytic. Parasitic forms live upon the juices of other living organisms, which are absorbed through any part of the body wall (Fig. 71). Finally, some take in solid food, which is acted upon and digested by the fluids of their inner plasm, the indigestible portions being excreted as in the higher animals. In the holophytic forms there is frequently an unbroken shell about the animal which makes it impossible for solid food to enter (Hcemato- coccus ; many Dinoflagellidia). Many of the holozoic forms have a distinct mouth and oesophagus. In its simplest form the mouth is merely a softened area about the base of the flagellum, against which the solid food particles strike (Oikomonas, see Fig. 18, -B). Others have a distinct mouth-opening leading into a gullet, which in turn opens into the fluid endoplasm (Perancma ; Petalomonas, see Fig. i, B). 1 See infra, p. 258. 126 THE PROTOZOA Where the flagella are in separate groups, there is a mouth at the base of each group. The collar of the Choanoflagellida is especially adapted for the collection and direction of the food particles into the interior. Perty ('52), Kent ('81), Stein ('67), Entz ('88), and others have described a number of types which they claim have both kinds of nutrition, and are intermediate between the holozoic and holophytic forms ; but Biitschli, although he admits that food-taking may be either holozoic or holophytic in one form at least (Chromulina flavi- cans\ which lives equally well when one or the other mode of nutrition is prevented, is inclined to doubt the wide distribution of this double function. Meyer ('97) main- tains that in one form (OcJiromonas gmnulosa} the organism may be either holozoic, saprophytic, or holophytic in nutrition. In many of the holophytic forms, there is a distinct gullet, which Perty and Kent regarded as a food- taking organ and, therefore, evidence of holozoic nutrition. Biitschli, how- ever, maintained that it is a part of the excretory system and connected with the contractile vacuole (Ettglcna, Cryp- tonionas, etc.). A close connection between holozoic Fig. 71. Megastoma enter icum Grassi. Ventral and side views. [GRASSI.] and holophytic forms is found, not only in Flagellidia, but in Dinoflagellidia as well. Possessing chromatophores and a coating of modified cellulose, these organisms were for a long time regarded as plants, but some forms among them are known to move about like animals and to ingulf solid food. Such forms may be either naked, as in Gymnodinium (Fig. 64, A) and Polykrikos, where food-taking has been actually seen by a number of observers (Schmarda, Stein, Bergh, Schilling, Dangeard), or shell-bearing, as in Glenodinium edax (Schilling). It is probable that they are much more closely related to the animal Flagellidia than to the Diatomaceae (as Warming maintains), or other plants, although no hard and fast line can be drawn about any of these groups. The food of the holozoic flagellates consists of bacteria and minute bits of disintegrated proteid matter. These in the Rhizomastigidae, as in the Rhizopoda, are surrounded by pseudopodia, and are subse- quently drawn into the body. In other Mastigophora, the flagellum is the chief factor in alimentation, and by its vibrations a current is created toward the base, where the mouth or its equivalent is THE MASTIGOPHORA I2/ situated. The particles of food brought with the current find their way into the body-plasm, where an indefinite cyclosis carries them hither and thither until the digestible portions are separated from the indigestible, and the latter are finally thrown out. James-Clark ('66), Kent ('81), and most observers have maintained that, in the Choanoflagellida, the food particles strike against the collar, subse- quently working down on the inside to the mouth, but Entz ('83) and France ('93, '97) claim that the mouth is not within the collar, but that the so-called vacuole described by Biitschli ('84) is a soft ingest- ing area at the base of the overlapping edge of the collar (see Fig. 69). In Noctilnca, the flagellum brings a current of food toward the collar, while the tentacle, which constantly beats down into the bottom of the collar area, drives it into the mouth situated at the bottom of the pharyngeal groove. The particles are then received into a gastric vacuole, which, in the vicinity of the relatively large nucleus, per- forms its function of digestion. E. VACUOLES Some of the vacuoles which make up the protoplasm of the Masti- gophora are gastric, while others are contractile. The former are formed about the food particles, which are probably digested in the same way as in the Sarcodina, although in this group no experiments have been made to test the digestive fluids. The contractile vacuoles, acting possibly as respiratory and excre- tory organs, pulsate rhythmically and at definite rates, varying from one or two pulsations per hour to five or six a minute, according to the temperature and nature of the surrounding medium. They are typically small, single or double in number (multiple in Chlorogonium\ and are situated at either end of the body, or near the centre, while in some cases they move with the granules in cyclosis. In some of the more complicated types of Euglenidae, the vacuole is connected with the so-called gullet by a minute canal. This canal in some cases receives its supply of waste matter from a reservoir, which is the receptacle for the contents of numerous small vacuoles surrounding it, and which pulsate at regular intervals. F. REPRODUCTION Binary fission, the typical method of reproduction among the Mastigophora, and the simplest of all modes of increase, is invariably preceded by division of the nucleus. When chromatophores, eye- spots, and pyrenoids are present, they also may be halved and 128 THE PROTOZOA equally represented in the daughter-cells, or they may remain whole, going to that daughter-cell to which they are nearest, the other cell forming a new set. The flagellum, in some cases, is also divided throughout the entire length, although in other cases it is thrown off before division takes place, new ones being formed by the daughter- cells. In many cases new flagella, as well as all of the important structures, are pre-formed before division. Such divisions may take place while the organisms are moving freely about in the water, or Fig. 72. Gonium pectorale O. F. M. [STEIN.] while they are quiescent and inclosed in a firm cyst. As a rule, division is longitudinal, but cases are well known where it is trans- verse (in most of the Dinoflagellidia, in Epipyxis, Sty lochry salts, Oxyrrhis, etc.). Some forms (e.g. Trachelomonas} reproduce by simple division while still within the shell, one half making its way out through the neck or flagella-opening, leaving the other in possession of the original home. Colony-formation is closely connected with the process of simple division, and nowhere among the Protozoa does it reach such high grades of differentiation as in this class. The colonies of this group THE MASTIGOPHORA 129 are composed, for the most part, of descendants of one ancestor and are formed by incomplete division or by subsequent attachment. In these colonies, which are wonderfully varied, the individual monads in some cases are embedded in a transparent cellulose jelly secreted by the cells, and in which they lie freely, or attached to one another Fig. 73. Division of Gonium pectorale O. F. M. a, b, and e, undivided cells ; c, d, f, k, and /, 4-celled stages ; h, i, j, n, o, 8-celled stages ; g, m, and/, i2-i6-celled stages. by stalks, while in other cases there is no surrounding matrix, the individuals remaining connected through incomplete division or by attachment subsequent to division. Gonium (Fig. 72), Pandorina, Uroglena, Proterospongia, Volvox, etc., have the cellulose jelly, while Dinobryon, Anthophysa, etc., are formed by attachment subsequent to division. In some of the more complicated colony-forms, especially in the Phytoflagellida, the adult condition is attained through cleavage stages as regular as in any metazoan egg. The formation of such a 130 THE PROTOZOA colony never varies, and the number of individuals is constant. (In Goninm sociale there are 4 individuals ; in G. pectorale 16, while in Eudorina there are from 16 to 32, and in Pandorina 32.) A Goninui colony lies in one plane, but this arrangement is brought about by a secondary shifting of the cells (Fig. 73), while a Eudorina colony retains the spherical form. Pandorina, a similar compact and defi- nite colony, is derived as in Eudorina, by the regular cleavage of a single cell. In some colonies the individuals are connected in the centre by protoplasmic strands, as in Synnra, while in one genus (Uroglena) connecting strands may or may not be present. Ehrenberg ('38) de- scribed U. volvox as a colony form whose peripheral individuals are connected in the centre by tail-like processes which, except for a much greater length, are similar to those of Synnm. He was confirmed in this by Zacharias ('95) and Kent ('81). Biatschli, however, regarded this central attachment as extremely doubtful. In one form, U. vol- vox, this connection does actually exist, but in another, U. americana, the posterior ends of the cells are rounded and have no trace of a central filament. The genus Uroglena may afford, therefore, a clue to the phylogenetic relations of the relatively huge gelatinous colonies which, save for the surrounding matrix, have no means of connection. 1 Proterospongia, in its general form and structure, agrees with U. americana. In both cases, as far as known, there is an indefinite number of individuals and no typical method of increase, as in Pan- dorina, Eudorina, and Goninm. The most highly differentiated colonial forms are the genera Volvox and Magosphcera, which should perhaps be considered simple multi- cellular forms rather than Protozoa. In Volvox the monads (often as many as 12,000 in a single colony) are arranged as in Uroglena, around the periphery of a gelatinous mass, and no organized connections with the centre of the cell can be traced, although they are connected with one another by definite protoplasmic strands. In Magosplicera the individuals are connected not only by the jelly matrix, but also, as in Symira, by protoplasmic stalks, and they are in close contact at the periphery. In both Magosphcera and Volvox the appearance of the peripheral cells is strikingly similar to a pave- ment epithelium, and the comparison which is so often made between such colonies and the blastula stage in the development of Metazoa is certainly justifiable. Stalked colonies have an entirely different mode of origin, being formed by repeated longitudinal division, the daughter-cells remain- l Cf. Calkins ('91). THE MASTIGOPHORA 131 ing attached to the stalk by their basal ends. In Dinobryon, a free- swimming colony of variable size, each monad occupies a small cup of cellulose (see Fig. 61). They increase by simple longitudinal division, one daughter-cell remaining in the original house, while the other moves out to the edge of the parent cup, where it attaches itself by the posterior end. A cellulose cup is then secreted about the daughter-cell, remaining firmly attached, however, to the parent cup. The mother-cell may divide again and again, the daughter-cells at- taching themselves to the edge of the cup already formed until there are three or more individuals around the edge of the original one. At the same time the daughter-cells may be dividing in a similar manner, and a much-branched bush-like colony is the result. Other forms have stalks which in some cases are much longer than the in- dividuals themselves (Codonocladium, Dendronwnas, Codonosiga, etc.). Still another type of colony-formation is found among the Dino- flagellidia, where from two to eight individuals are connected end to end by their shell processes (Ceratium}. The significance of this chain-formation (catenation) is not clearly established, many regard- ing it as the result of incomplete division (Pouchet, '85), others as preparatory to conjugation (Biitschli, '83). The colonies as well as the individuals may increase by division, a purely mechanical process, however, and probably due to the un- wieldy size of the overgrown aggregate. Zacharias ('95) and others have seen large colonies of Uroglena break into two portions through the asynchronous action of flagella in different regions. If the flagella of one half of the colony vibrate in one direction while those of the other half vibrate in an opposite direction, the result is a twist- ing of the entire mass which must ultimately give way. Such division cannot be regarded as reproduction in a strict sense. Closely allied to simple division is the formation of swarm-spores or microgonidia. This may occur either in the free motile condition as in Polytoma or Chlorogoninm, or in the encysted and protected state, as in many Monadida. The simplest form is seen in such cases as Polytoma, where, instead of dividing into two portions, the organism divides into four, eight, or, according to Dallinger and Drysdale, into sixteen smaller forms. These develop new flagella, make their way through the parent membrane, and grow to full size. The formation of similar gametes has been observed in most of the Mastigophora, either in their resting or in their encysted stages. The flagella are drawn in, a mantle or cyst is secreted, within which the protoplasm divides into a number of spores. In some cases swarm-spores, like those of the Radiolaria, are of different sizes (macro- and micro- gametes), and these may conjugate. So far as known, the formation of gametes is not accompanied by 132 THE PROTOZOA complex nuclear changes. As in the Reticulariida and some Sporo- zoa, the chromatin is reduced to minute granules which are spread throughout the cell, but in the Flagellidia they are so small that their further history is not known. Not all forms, however, are of this primitive type ; some, as for example Noctiluca miliaris, and some of the Dinoflagellidia, undergo a complicated mitotic process which in Noctiluca is repeated until five or six hundred spores are formed (Fig. 74)- The formation of spores or gametes may or may not be preceded by the conjugation of individuals. In those species of Mastigophora in which spore-formation is preceded by conjugation, a very interest- ing series of forms may be selected, showing the grad- ual development of sex from types in which there is a union of individuals of sim- ilar form, size, and, appar- ently, of condition, to the union of specially developed male and female reproduc- tive elements. Cienkowsky ('56) was the first to observe the fusion of similar monads, but the most complete obser- vations are those of Dallin- ger and Drysdale ('73), who watched the fusion of several ,,. individuals of Rodo (Cerco- Fig. 74. Noctiluca milians Sur. Spore-formation. [ROBIN.] monas) crassicanda, the en- cystment of the fused mass, and the subsequent divisions of the plasm up to the formation of an immense number of minute spores (Fig. 75). In another form (Oikpmonas Dallingeri} similar spores are formed, but without the preliminary fusion of two or more small individuals. The gametes move about until they come in contact with the adult individuals with which they fuse. The fused mass then encysts and finally breaks up into minute spores. An advance toward sexual differentiation is seen in Pandorina (Pringsheim, '69), where, after a long period of asexual reproduction resulting in numerous colonies, the cells separate and begin to form swarm-spores which may be of the same or of different size. These spores then swim about until two of them meet and fuse by the color- less ends into a common body (Fig. 76). Fusion may take place between two small gametes, or between a large and a small one. THE MASTIGOPHORA 133 Pringsheim regards the larger ones as females, while the smaller ones may be either male or female. Fig. 75. Cercomonas crassicauda Duj. [DALLINGER and DRYSOALE.] A. Ordinary forms. B. Division stage. C, Conjugation of two individuals in amosboid con- dition. D-E. The copula. F. Sporulation. The fused mass (zygospore) encysts and dries, the color changing from green to red. When remoistened, the contents again turn green and break open the cyst, usually as a single swarm-spore, although 134 THE PROTOZOA -Pandorina morum Ehr. Conjugation. [PRINGSHEIM.] A. i6-celled colony. B. Macrogamete. C and E. Fusion of macrogamete with microgamete. D and F. Fusion of micro- gametes. G. Copula. two or more may be formed. These gametes soon divide and form the typical sixteen-cell Pandorina colony. Thus in Pandorina, each of the cells forms both sexual elements, but an advance in differentia- tion is seen in Eudorina elegans, where, according to the rather incom- plete observations of Carter ('58), the thirty-two cells forming the colony have a different fate when the conju- gation period comes around. Four of the thirty -two cells situated at the end of the colony form gametes by re- peated divisions in one plane, while the other twenty-eight cells merely develop more amylum granules and turn darker. The ga- metes which are formed from the upper four cells are elongate and spindle-shaped, with two flagella, a red eye-spot, and a long tail. The fate of the different cells was not made out, but there seems reason to believe that if the observations were correct, the gametes represent male elements, the other cells female. A still more decided advance is shown by the colonies of Volvox. Volvox can scarcely be regarded as a unicellular organism, for differ- entiation has gone so far that the cells if separated, with the excep- tion of the reproductive elements, cantiot live. The individuals form- ing the peripheral layer (in Volvox globator about 12,000, Cohn, '75) form the sterile vegetative or somatic cells of the aggregate. A few of these cells, which reproduce asexually, are found upon the inside of the peripheral layer, which protects them like a mantle. Stein finds eight of these asexual cells or partJienogonidia in V. globator, and Cohn, one to nine in V. minor. The parthenogonidia, by repeated division, form daughter-colonies from one-quarter to two-fifths the size of the parent colony, which finally make their escape from the latter by rupture of its walls. After a considerable period of such asexual reproduction, sexual elements are formed. These are at first similar to the parthenogo- nidial cells, but are more numerous. Later they can be distinguished as male (androgonidia, Cohn) and female (gynogonidia, Cohn). In Volvox globator there may be from twenty to forty gynogonidia and THE MASTIGOPHORA 135 from two to five androgonidia, but there may be one hundred andro- gonidia in V. minor while there are only eight gynogonidia. Each female cell becomes at first flask-shaped, then withdraws inside of the colony and becomes a mature egg. The male cell, at the beginning about three times the size of the sterile cells, soon begins to divide in one plane until a bundle of from sixty-four to one hundred and twenty- eight flagellated, spindle-formed elements results. These, the sperma- tozoids, gather around the egg-cells, which are fertilized exactly as in higher animals or plants. Fertilization takes place while the egg is still in the parent colony ; the copula forms two membranes about itself, while the color changes from green to orange. After a consid- erable resting period, the egg undergoes regular cleavage, forming the adult colony, in which, even before the embryo leaves the egg, the cells are differentiated into somatic cells and parthenogonidia. After these cleavage stages the outer cyst wall is ruptured and the young colony swims out. G. INTER-RELATIONS OF THE MASTIGOPHORA Transitional forms between the Mastigophora and the Sarcodina show how closely the two classes are related. The development, both in Sarcodina and in Mastigophora, throws little or no light upon the question as to the more primitive nature of one or the other. While many Sarcodina have flagellated swarm-spores, many Flagel- lidia have amoeboid spores, and even in the same species both amoe- boid and flagellated swarm-spores are formed at the same time (Acanthocystis Schaudinn, see Fig. 53, F\ The most primitive forms of Flagellidia suggest the long-disputed question over the boundary-line between animals and plants. 1 Un- questionably, the most primitive flagellates are those forms which, while actively motile, possess chromatophores and chlorophyl, and are able to make their own food, or which, like the bacteria, can sup- port themselves upon simpler substances than proteid. The living flagellates which come closest to these primitive types are the monads, while the Choanoflagellida are probably not far removed. Klebs ('93) takes the ground that the collar-like process of the Choanoflagellida is not a sufficient taxonomic differential save for ordinal distinctions, and recent zoologists are inclined to accept this view. Stein, Biitschli, and Bergh have been the most active in formulat- ing views as to the origin of the Dinoflagellidia, although none of these is wholly satisfactory. Some authors derive them from the Phytoflagellida directly (Haeckel), others place them as a group of the 1 Cf. p. 22. 136 THE PROTOZOA Diatomaceae (Warming). The majority of observers are agreed, how- ever, that a connecting link with the Flagellidia is seen in certain species of the family Prorocentridae and represented among living forms by Exuvicella and Prorocentrum (Fig. 66). Btitschli ('83) and Klebs ('93) agree that these might be called true Flagellidia; for, as Delage happily expresses it, they are little more than a chromomonad in the shell of Phacotus. The shell is bivalve, and perforated by minute apertures characteristic of the Dinoflagellidia, and there is an entire absence of longitudinal and transverse furrows, while the flagella are directed outward from the anterior end. Butschli, Bergh, and Klebs derive them from forms like Cryptomonas, where the two flagella are pointed in the same direction and the chromatophores are yellow. The transition from these primitive forms of Dinoflagellidia to the more complex types with a shell composed of nicely articulated plates, is much more difficult than the connection between the main groups. Stein maintained that the simplest form is the unshelled Gymno- dininm, but Butschli showed that this view is not in harmony with the other forms of Dinoflagellidia, it being much more obvious to con- sider the shell of the Peridinidae, for example, as arising by the split- ting up of the bivalve shell of the primitive type, than by the loss of this shell and the subsequent formation of the articulated forms. Evidence in Biitschli's favor is seen in the forms where there are but few plates {e.g. Ceratocorys}, although we are inclined to agree with Klebs that a polyphyletic origin of the group is possible and that Gymnodininm might have been derived from the Rhizomastigidae. The origin of the Cystoflagellidia, composed of Noctilnca and Leptodiscus, is to-day generally conceded to be from the Dino- flagellidia, and is supported by direct evidence in the development of Noctiluca, where the swarm-spores are strikingly similar to Pcri- dininm. The relationship to the Dinoflagellidia, as first pointed out by Allman, was based upon superficial resemblances only, and the first conclusive observations must be credited to Pouchet('83) and to Stein ('83), while Butschli ('85) first applied the theory on the basis of the swarm-spore as described by Cienkowsky ('73) and Robin ('78). The interesting form which Pouchet later described ('92) as Peridininm psendonoctiluca is now considered a young stage of Noctiluca. CLASSIFICATION CLASS II. MASTIGOPHORA. Protozoa of definite or indefinite form; naked, or provided with a well-defined membrane. The nutrition is holozoic, parasitic, holophytic, or saprophytic. The motile organs are flagella, which may vary in number from one to many. Mouth, contractile vacuole, and nucleus are usually present. They are usually small forms with a widespread tendency to colony- formation. THE MASTIGOPHORA 137 Subclass I. FLAGELLIDIA. These are small organisms possessing usually a sharply defined, mononucleate body with a definite anterior end in which are inserted one or more flagella. They are actively motile during the greater period of life, but all have the power of encystment. Reproduction occurs by longitudinal division, usually during the flagellated stage, although it may take place during resting phases. Nutrition is holophytic, holozoic, parasitic, or saprophytic. Order i. MONADIDA. Small forms of Flagellidia having a simple structure. The body is frequently amoeboid, with one or two flagella at the anterior end. There is no distinct mouth-opening, but a localized area about the base of the flagella serves for the ingestion of food particles. Family i. Rhizomastigidae. Simple, mouthless forms with one or two flagella and an amoeboid body capable of putting out lobose pseudopodia like a rhizopod, or stiff radial pseudopodia like a heliozoon. The contractile vacuole is frequently at the posterior end. Food particles may be ingested at any part of the body by the aid of the pseudopodia. Genera: Mastigamceba F. E. Schultze ('75) ; Ciliophrys Cienk. ('76); Dimorpha Gruber ('81); Actinomonas Kent ('80); Trypanosoma Gruby ('43) ; Mastigophrys Frenzel ('91). Family 2. Cercomonadidae. Oval or elongated forms which are frequently amoeboid or changeable, but unable to form pseudopodia. There is one large fiagellum with a mouth area at its base. The family includes small forms, saprophytic, or holozoic, or sometimes parasitic in nutrition. Genera: Cercomonas Dujardin ('41); Herpetomonas Kent ('80), parasitic. Oikomonas Kent ('80); Ancyro- monas Kent ('80) ; Phyllomonas Klebs ('93). Family 3. Codoncecidae. Small colorless monads which secrete and remain in a gelatinous or membranous cup. Genera: Codonasca James-Clark ('66) ; Platy- theca Stein ('78). Family 4. Bikoecidae. Small monads of peculiar form. They are provided with a cup, to which they are attached by a slender thread. The basal portion is broader than the upper part, which bears a curious tentacle-like process. Nutri- tion is holozoic ; the individuals are single or colony-forming. Genera : Bicosceca James-Clark ('67) : Poteriodendron Stein ('78). Family 5. Heteromonadidae . Small colorless monads which have, in addition to the chief flagellum. one or two accessory flagella. They frequently form colonies upon a common stalk. Increase of the individuals is by longitudinal division. Genera: Monas Stein ('78); Dendronionas Stein (78); CepJialothamninin Stein ('78) ; Anthophysa Bory d. St. Vincent ('24) ; Epipynis Ehr. ('38) ; Amphimonas Kent ('81) ; Spongowonas Stein ('78) ; Cladomonas Stein (78) ; Rhipidodendron Stein ('78) ; Diplomita Kent ('80). Order 2. CHOANOFLAGELLIDA. Flagellidia with one or more collar-like pro- cesses about the base of the single flagellum. Family i. Phalansteridae. Colony-forming Choanoflagellida. Each individual is situated in a granular gelatinous tube. The gelatinous tubes form either a dis- coid colony in which the single tubes are arranged radially, or a dichotomously branched aggregate. Genera: Phalansterinm Cienk. ('70). Family 2. Craspedomonadidae. Solitary or colonial forms. The individuals are naked, or lie in an incomplete cup, or in a gelatinous mass. Genera : ftlono- siga Kent ('80) ; Codosiga, James-Clark ('67) ; Codonodadium Stein ('78) ; Hirmidium Perty ('52) ; Proterospongia Kent ('81) : Sphceraeca Lauterb. ('99) ; Salpingaeca James-Clark ('67) ; Polyaeca Kent ('81) ; Diplosiga Frenzel ('91). Order 3. HETEROMASTIGIDA. A small group with various kinds of flagellated organisms, which are sometimes naked and amoeboid, sometimes provided with a complex membrane. The essential character is the possession of two or more 138 THE PROTOZOA flagella, one being directed forward and used in locomotion, the others directed backward and trailed after the organism. Nutrition is holozoic. and all of the forms included are colorless. Family i. Bodonidae. Small naked forms in which there is only a slight difference, if any, between the flagella. Genera: Bodo Stein ('78); Phyllomitus Stein ('78) ; Colponema Stein ('78) ; Oxyrrhis Dujardin ('41). Family 2. Trimastigidae. With two accessory flagella. Genera: Dallingeria Kent ('8 1) ; Trimastix Kent ('Si). Order 4. POLYMASTIGIDA. The body is invariably without a shell, and is provided with a delicate membrane, which allows more or less amoeboid movement. The number of flagella varies from three to many, and the number of mouth open- ings, or food-taking areas, likewise varies. Nutrition is holozoic. They increase by longitudinal division. Tribe I. Astomea. Polymastigida with many flagella and without a mouth opening. Genera: Multicilia Cienk. ('81) ; Grassia Fisch ('85). Tribe 2. Monostomea. The anterior part is provided with a large mouth opening at the base of the four or six flagella. Genera: Collodictyon Carter ('65) ; Tetra- mitus Perty ('52) ; Monocercomonas Grassi ('82) ; Trichomonas Donne" ('37) ; Megastoma Grassi ('81). Tribe 3. Distomea. The flagella are separated into two symmetrical groups, with a mouth area at the base of each group. Genera : Trigonomonas Klebs ('93) ; Hexamitus Dujardin ('38) ; Trepomonas Dujardin ('39) ; Spironema Klebs ('93) ; Urophagus Klebs ('93). Tribe 4. Trichonymphinea. Polymastigida, of unknown affinities, provided with numerous flagella. They are parasites in the rectum of various hosts (Termites) . Genera: Lophomonas Stein ('78); Leidyonella Frenzel (^91); Trichonympha Leidy ('77) ; Jcenia Grassi ("85) ; Pyrsonympha Leidy ('77). Order 5. EUGLENIDA. Large forms, having one or two flagella. a contractile or firm body-wall, a mouth and pharynx at the base of the flagellum, and with a con- tractile vacuole opening into the pharynx. They frequently form colonies and are usually provided with chromatophores. Nutrition is holozoic, holophytic, or saprophytic. Family i. Euglenidse. Elongate forms, with a more or less pointed end and usually with one flagellum. The cuticle is marked with spiral stripings ; the contractile vacuole, or vacuoles, open into a reservoir, which in turn opens into the pharynx. A red eye-spot, or stigma, and green chromatophores, are usually present. Within the body there are discoid, or, occasionally, band-formed chromatophores. Paramylum granules are always present. Genera : Euglena Ehr. ('30) ; Colacimir Ehr. ('33) ; Eutreptia Perty ('52) ; Ascoglena Stein ('78) ; Trachelomonas Ehr. ('33) \ Lepocinclis Perty ('49) ; Phacus Nitsch ("16) ; Cryptoglena Ehr. ('31). Family 2. Astasiidae. The body is elongate and usually has a striped membrane. The anterior end is similar to that of Euglena, but there is no eye-spot. The body is invariably colorless. Nutrition is saprophytic. Genera: Astasia Ehr. ('38) ; Distigma Ehr. ('31) ; Rhabdomonas Fres. ('58) ; Menoidium Perty ('52) ; Atractonema St. ('78) ; Sphenomonas Stein ('78). Family 3. Peranemidae. The body is either stiff or plastic, and usually symmetrical. The anterior end bears either one or two dissimilar flagella, which are more or less deeply sunk in the body. A distinct mouth is found at the base of the flagella. Nutrition is holozoic. Genera : A. With plastic body and one flagel- lum : Euglenopsis Klebs ('93) ; Peranema Dujardin ('41); Urceolus Meresch- kowsky ('77)- B. With a plastic body and two flagella: Heteronema Dujardin ('41); Dinema Perty ('76); Zygoselmis Duj. ('41). C. With a constant body form and one flagellum : Scytomonas Stein ('78) ; Petalotnonas THE MASTIGOPHORA 139 Stein ('59). D. With a constant body form and two dissimilar flagella : Tro- pidoscyphus Stein ('78) ; Anisonema Duj. ('41) ; Entosiphoii Stein ('78) ; Thaumatomastix Lauterb. ('99). Order 6. PHYTOFLAGELLIDA. Flagellated unicellular organisms with chlorophyl and holophytic nutrition, or without chlorophyl, and saprophytic in nutrition. They are sometimes classified as plants, sometimes as animals. Suborder i. CHLOROMONADINA. The body is somewhat plastic and without a dis- tinct membrane ; with numerous discoid chromatophores but without stigmata. Genera : Vacuolaria Cienk. ('7) \ Coelomonas Stein ('78). Suborder 2. CHROMOMONADINA. Small forms with strong tendency to colony- formation. They are often inclosed in a gelatinous mass, or occupy cups. They may or may not have chromatophores, which, if present, are yellow or yellowish brown in color. Nutrition is usually holophytic, but holozoic and saprophytic forms are occasionally present. There may be one or two flagella, which are invariably directed forward. Family i. Chrysomonadidae. The body is rarely naked, but usually covered by a gelatinous mass or by a hyaline cup. With one or two flagella at the anterior end and with or without stigmata. One or two yellowish chromatophores are invariably present. Nutrition is holophytic or holozoic, sometimes both. Genera: A. With naked body which may be inclosed during resting stages in a gelatinous mass. Nutrition either holozoic or holophytic. Chrysamaeba Klebs ('90) ; Chromulina Cienk. ('7) ; Ochromonas Wysotzki ('87) ; Stylo- chrysalis Stein ('78). B. With a shell or lorica in which the individuals are attached. Nutrition is holophytic. Chrysococcus Klebs ('92) ; Dinobryon Ehr. ('38) ; Chrysopyxis Stein ('78) ; Nephroselmis St. ('78) ; Hyalobryon Lauterb. ('99). C. Individuals protected by a close-fitting membrane. Hy- menomonas Stein ('78) ; Microglena Ehr. ('31) ; Mallomonas Perty ('76) ; Synura Ehr. ('33) ; Syncrypta Ehr. ('33) ; Uroglena Ehr. ('33) ; Chryso- sphcerella Lauterb. ('99). Family 2. Cryptomonadidae. The body has a firm cuticle and is never amoeboid. There are two similar flagella, a peculiar oesophagus-like canal, and a contractile vacuole in the anterior end. Two chromatophores of variable color may or may not be present. Nutrition is holophytic or saprophytic. Genera: Cryptomonas Ehr. ('31); C/rilomonasE\\r. ('31); Cyathomonas Fromentel ("74)- Suborder 3. CHLAMYDOMONADINA. Body-form more or less changeable. Color usually green, and due to the presence of a large, sjngle chromatophore contain- ing chlorophyl. A firm shell is usually present. The body has two or four flagella, one or two contractile vacuoles, and a stigma at the anterior end. Re- production takes place by continued division within the shell either during active or resting phases. Macro- and micro-gametes may be formed. Family i. Chlamydomonadidae. With a stiff coating perforated only by minute apertures for the flagella. Genera : Chlamydomonas Ehr. ('33) ; Chlorogonium Ehr. ('35) ; Polytoma Ehr. ('38) ; Hcematococcus Agardh ('28) ; Carteria Diesing ('66) ; Spondylomorum Ehr. ('48) ; Chlorangium Stein ('78). Family 2. Phacotidae. The body of the flagellate corresponds to that of Hcemato- coccus, and is surrounded by a thick shell membrane which the body does not fill. The shell is frequently bivalved. Genera : Coccomonas Stein ('78) ; Meso- stigtna Lauterb. ('94) ; Phacotus Perty ('52) ; Tetratoma Butschli ('85) ; Pyra- mimonas Schmarda ('50) ; Chlor aster Ehr. ('48). Suborder 4. VOLVOCINA. Colony forms. The individuals possess two flagella and chlorophyl-bearing chromatophores. The number of individuals composing the colony may be constant or variable ; when constant, the colony is formed by regular cleavage, as in the eggs of Metazoa. Reproduction asexual by division 140 THE PROtUZOA or sexual. Genera: Gonium O. F. Miiller (1773); Stephanosphara Cohn ('53); Pandorina Bory de St. Vincent ('24); Eudorina Ehr. ('31); Volvox Leeuw. Ehr. ('38) ; Plaodortna Shaw ('94) ; Platydonna Kofoid ('99). Order 7. SILICOFLAGELLIDA. A single genus, Distephanus Stohr ('8 1 ), character- ized by the presence of a silicious latticed skeleton like that of the Radiolaria. There is no mouth nor modifications of the plasm whatsoever, but the animal is colored yellow by (probably) diatomin. Parasitic on Radiolaria. Subclass II. DINOFLAGELLIDIA. Naked or shelled Mastigophora. There are usually two flagella, of which one is directed away from the body, the other around the body ; the shell usually has two furrows, one running transversely around the body, the other vertically. Marine and fresh-water forms. The nutrition is holophytic or holozoic. Order i . ADINIDA. The transverse furrow is absent, and the two flagella arise from the anterior end of the body. The shell may be bivalved. Family i . Prorocentridae. With the characters of the order. Genera : Ewvialla Cienk ('82) ; Prorocentrum Ehr. ('33). Order 2. DINIFERIDA. Dinoflagellidia with two transverse furrows. Family i. Peridinidae. The cross furrow is near the middle of the body, which may be with or without a shell. The form is extremely variable. Genera: Podolampas Stein ('83); Blepharocysta Ehr. ('73); Diplopsalis Bergh ('82); Peridinium Ehr. ('32) ; Goniodoma Stein ('83) ; Gonyaitlax Diesing ('66) ; Ceratium Schrank (1793); Amphidoma Stein ('83) ; Oxytoxum Stein ('83) ; Pyrophacus Stein ('83) ; Ptychodiscus Stein ('83) ; Protoceratium Bergh ('82) ; Glenodinium Ehr. ('35) ; Gytnnodinium Stein ('78) ; Hemidinium Stein ('78) ; Steiniella Schiitt ('95) ; Monaster Schiitt ('95); Amphitholits Schiitt ('95). Family 2. Dinophysidse. The cross furrow is above the middle of the body, and its edges are raised into characteristic ledges. Marine. Genera: Phalacroma Stein ('83) ; Dinophysis Ehr. ('39) ; Amphisolenia Stein ('83) ; Citharistes Stein ('83) ; Histioneis Stein ('83) ; Ornithocercus Stein (''83) ; Amphidinium Clap, and Lach. ('59); Ceratocorys Stein ('83). Order 3. POLYDINIDA. The order consists of the single genus Polykrikos But- schli ('73), which is characterized by a naked body, by several transverse furrows and flagella, by macro- and micro-nuclei, and by nematocysts. Nutrition is holozoic. Subclass III. CYSTOFLAGELLIDIA. Mastigophora of considerable size, with a single nucleus, parenchymatous protoplasm, and a firm membrane. Nutrition is holozoic. Marine. Genera: Noctiluca Suriray ('36); Leptodiscus R. Hertwig ('77). SPECIAL BIBLIOGRAPHY IV Biitschli, 0. Protozoa, Mastigophora. In Bronn's Klassen und Ordnungen des Thierreichs. Leipzig, 1883-1887. FrancS, R. H. Der Organismus der Craspedomonaden. Budapesth, 1897. Kent, W. Saville. A Manual of the Infusoria. London, 1881. Klebs, G. Flagellatenstudien i. Zeit.f. wiss. Zool. Bd. LV., pp. 265-445. Schiitt, F. Die Peridineen der Plankton-Expedition. Th. I. Kiel and Leipzig, 1895. Senn, G. Flagellata. In A. Engler\s Die naturlichen Pflanzenfamilien. 202 and 203 Lieferung. Leipzig, 1900. Stein, Fr. Der Organismus der Infusionsthiere. Leipzig, 1878. CHAPTER V THE SPOROZOA THE Sporozoa are unicellular animal parasites living in the cells, tissues, and cavities of various hosts and, as the name indicates, char- acterized by reproduction through spore-formation. If we except the bacteria, they are the most widely distributed of all parasites, and are found in every class of animals, frequently in Vermes, Arthrop- oda, Mollusca, and Vertebrata, rarely in Protozoa, Coelenterata, and Echinodermata. They may infest the alimentary tract, and all Fig. 77. The vegetative phase in the life-history of a gregarine (schematic). [WASIELEWSKY.J The young sporozoite (A), liberated in the intestine, enters an epithelial cell (B), where as an intra-cellular parasite (p) it grows at the expense of the cell-contents, often forcing the nucleus (n) to a corner of the cell. It finally grows through the cell-wall (C) and ultimately drops into the lumen of the organ as a sporont (D). of the connecting organs and ducts ; the kidneys and their ducts ; the blood-vessels and the blood ; the muscles and connective tissues ; while even the skin is not exempted. In most instances they are harm- less, but they may produce morbid and even fatal results, either in- directly, by increasing to such numbers that the lymph-spaces and cavities are filled with them, thus preventing nutrition of the cells and tissues, or directly, by causing atrophy and death of the cells in which they live. They are usually taken into the system in the spore-stage with the food of their host, although infection may take place through the gills or lungs, or even by inoculation from insects. The spore- membranes are soon dissolved by the fluids of the host, and one or more germs are thus liberated. These germs, the sporozoites, then 141 142 THE PROTOZOA bore into the epithelial cells, where they grow (Fig. 77). All forms, apparently, begin life as intra-cellular parasites, where, at first, they do> little harm, but as they grow by the absorption of fluids contained within their cell-hosts, the latter are improperly nourished and, unless the parasites leave them, they degenerate and die (Fig. 78). The duration of intra-cellular life varies in different kinds of Spo- p rozoa : some are permanently intra-cellular {monopJiagoiis forms, so-called Cytosporidia,&,?)\ others are intra-cellular only in the young or immature phases (Gregarinida); while still others pass different phases of their life-history in dif- ---n Fig. 78. Coccidia in the epithelial cells of Triton cristatus. n, nuclei of the tissue cells; /, the intra-cellular parasite Pfeifferia tritonis. ferent cells (polyphagons forms). The mature parasites finally may leave the cell-host and sporulate in the digestive cavity or coslom, and the spores are then carried to the outside with the fasces or other excreta. A. PROTOPLASMIC STRUCTURE A typical sporozoon consists of protoplasm and one nucleus. It has no mouth, anus, excretory pore, or other openings. It has neither gastric nor contractile vacuoles, and has at most a sluggish movement in the adult stage, although the young forms may be amoeboid or flagellate. Owing to the number of cytoplasmic granules which make up the bulk of the animal, the protoplasmic structure of adult forms can be made out only with the greatest difficulty. Apart from these granules, however, which are regarded as reserve nutri- ment, it is probable that, as in all Protozoa, the protoplasm is alveolar. This is certainly the case in Coccidiida (intra-cellular Sporozoa), espe- cially in the young forms, where Labbe ('96) describes the cytoplasm as alveolar; in some forms of Gregarinida (Fig. 87), and in Myxo- sporidiida as described by Thelohan ('95) and Doflein ('98) (Fig. 79). The granules, which are so characteristic of the group, completely fill the alveolar network, and give to the protoplasm its peculiarly dense appearance. They differ somewhat in size and shape, and apparently in chemical composition, and are generally regarded as food substances reserved for use during the spore-producing period. THE SPOROZOA Wasielewsky ('96) enumerates the following kinds: (i) Paraglyco- These form the bulk of the granules in the Gregarinida ; they gen. are distinct refringent granules of variable size and are usually oval or spherical in form, consisting of a peculiar amyloid substance which Biitschli ('84) regarded as similar to amidon or glycogen. They give characteristic reactions, staining brown to violet with dilute sulphuric acid, and dissolving in potassium carbonate and strong mineral acids. (2) CarminopJiilous granules. These granules, which were first made out by Schneider ('75), are less numerous than the paraglycogen granules, but like them variable in size and strongly refractive. Fig. 79. Leptotheca agilis Dof., one of the Myxosporidiida. [DOFLEIN.] They are easily soluble in ammonia, but are not destroyed by alcohol, embedding in paraffine, etc., and are easily stained by carmine and many aniline colors, but not at all by haematoxylin. They consist, apparently, of albumen. (3) Fat. These granules are widely dis- tributed throughout the entire group, and have about the same appearance in all types, although they are colored differently in different species. They are soluble in alcohol, ether, and chloroform, and are stained black by osmic acid. In addition to the above gran- ules, which are found in most Sporozoa, there are others which have been found hitherto only in certain subdivisions. In the Gregarinida, pyxinine granules and protein crystals have been observed in certain species, the former by Frenzel ('85) in Pyxinia, where they appar- 144 THE PROTOZOA ently take the place of the paraglycogen granules, although of similar chemical nature, but slightly different in their reactions. The latter are also rather questionable inclusions in certain DidymopJiyes. In the Coccidiida all adult forms are characterized by the presence of so-called plastic grannies. These are globular, strongly refractive granules of slightly variable size which react differently from the glycogen granules, remaining unchanged in sulphuric acid and stain- ing yellow with iodine. Here also are found the so-called cJiromatoid granules, which are distinguished by their affinity for haematoxylin, and are probably albuminoid in nature. In the Haemosporidiida or blood- infesting Sporozoa, the effect of the intra-corpuscular life is shown by the presence of pigmented granules (melanin) of black, yellow ochre, or red color resulting from the disintegration of haemoglobin. In the majority of the Sporozoa the cell-body consists of a more or less sharply differentiated ectoplasm and endoplasm, while even a third layer, mesoplasm, is said to have been observed in some forms (Cohn, '96). It is possible that these different zones are function- ally specialized, a supposition first made by Labbe ('96) in connection with the Coccidiida, and repeated by Doflein ('98) in connection with the Myxosporidiida. As in the Sarcodina and Mastigophora, the ectoplasm may be plastic, yielding to the pressure from within and thus giving rise to pseudopodia (Monocystis ascidia, Siedlecki, '99, Myxosporidiida), or it may be modified into a hard and tough cuticle, which offers a good protection for the cell-body within. Again, it may be modified into a complex membrane, plastic and capable of various kinds of motion and similar to that of the higher types of Flagellidia. The most highly differentiated ectoplasm is found in the Gregarinida, where it forms a dense cortical layer about the body, while its outermost part is transformed into a complex membrane. In some cases the inner cortical layer of the ectoplasm is carried across the cell, forming a partition dividing the organism into two portions which are known as the protomerite and the deutomerite, the nucleus being in the latter. The non-nucleated portion is often further differentiated into an apparatus called the epimerite, which usu- ally develops hooks or anchors used for attaching the animal to its cell-host. The organism thus appears to be multi-chambered, and the presence or absence of such chambers was formerly regarded as a good basis for classification ; but it has been shown, especially by Leger ('92), that the partitions vary considerably in the same species, and even in the same individual, at different times, and in the recent systems of classification this feature has been discarded in determining the limits of the larger divisions. The epimerite, which is so important in holding the lumen-dwelling parasites in place, may be simple or branched ; plain, like a knob or a rod ; branched with filiform, or flat THE SPOROZOA 145 with digitiform, appendages. It may be closely attached to the pro- tomerite, or carried on a long neck, while variations in all types are numerous (Fig. 12, E, F, G, p. 39). Under certain conditions, prior to reproduction, the animal throws off the epimerite which may be left in the cell-host, and drops into the lumen of the organ in which it lives. Here it encysts, the protomerite and deutomerite forming one spore-producing individual. As the attached and the detached stages in the life-history of the Gregarine are each important, they have received special names, the former being known as a cephalont, the latter as a sporont. Between the cortical ectoplasm and the inner endoplasm there is a layer of myonemes, or muscular fibrils similar in all respects to those of the Ciliata (Fig. 80). These are occasionally found in the Haemosporidiida, but are much more characteristic of the Gre- garinida, where, except in the epimerite, they form a network about the entire animal. On the outside of this net- work, according to Schewiakoff ('94), there is, at times, a layer of gelatinous matter apparently secreted by the ecto- plasm, and this, in turn, is covered by the membrane proper. The mem- brane is longitudinally striated by rib- like projections, while the canals or furrows between them are filled with jelly from the gelatinous layer below (see Fig. 82, H}. Schewiakoff believes that the active secretion of jelly in these furrows accounts for the peculiar gliding motion of certain kinds of Gregarinida. In the region of the epimerite, the membrane is plain, the ribs and furrows stopping with the protomerite. The hooks or spines are formed from the cortical plasm. In one group of Sporozoa, the Sarcosporidiida, the protoplasmic body is inclosed in a peculiar pouch which appears to be a secretion from the protoplasm rather than a true cellular membrane. The mass slowly enlarges by regular growth until it reaches a considerable length, in some cases several millimetres. It then undergoes spore- formation. These organisms, known as Rainey's Tubes, are parasites of sheep, swine, deer, horses, rats, etc., where they infest the muscle- tissues, causing morbid symptoms, similar to those in trichinosis. Fig. 80. Schematic figure of the myonemes of Clepsidrina munieri ; ni, the myonemes. [SCHNEIDER.] 146 . THE PROTOZOA B. THE NUCLEUS With the exception of the multinucleate Myxosporidiida, the Sporo- zoa are mononucleate. Schneider ('81) abandoned the attempt to compare the nuclei in Sporozoa with those of ordinary animal and plant cells, because of their peculiar structure. In most cases they consist of a firm and resisting membrane containing a single large chromatin reservoir or karyosome, and are apparently without a linin reticulum, such as is found in the nuclei of Metazoa. In some forms the nucleus is similar to that of the Sarcodina and Mastigophora, consisting of membrane, reticulum, and one or more chromatin reservoirs. Recent observers have found that the different appear- ances of the nucleus are characteristic of different stages of nuclear activity, and that the reticulum, and even the nuclear membrane, are derived from the karyosome, which in the sporozoite appears as a solid homogeneous sphere of chromatin. 1 In the active phases the nuclei of the Sporozoa differ widely from those in other Protozoa, the most striking point of difference being the disappearance of the nuclear membrane during division. The chromatin reservoirs may divide directly, thus simulating the entire nucleus, or they may break down into small chromatin granules, resembling the first stages of chromo- some-formation in the flagellate Noctiluca. In the former there is no distinct spindle, in the latter the completed spindle-figure has two sets of fibres, although, according to Wolters's ('91) description, the fibres seem to have a different function from those in the mitotic figures of the Metazoa, since there is no connection between them and the chromatin. 2 C. FOOD-TAKING Like all endoparasites, the Sporozoa absorb fluid food through the body-wall, even when, as in the Myxosporidiida, pseudopodia are present. There is probably no specialized area devoted to food- taking, but all parts are equally receptive. It is believed that in some cases, notably in the Gregarinida and Myxosporidiida, minute pores -perforate the membrane between the outer markings. Although the taking of food has never been observed, the indirect effects are seen in the rapid growth of the parasite when in a suitable medium. Thus a young gregarine, when it penetrates an epithelial cell (Fig. 77, A\ is a minute ball of protoplasm; but it rapidly grows until it occupies the greater part of its host, often forcing the nucleus to one side. As it continues to grow, the front wall of the cell is pushed outward until it finally breaks, and the lower portion of the parasite 1 Cf. infra, p. 253. 2 See infra, p. 259. THE SPOROZOA is left exposed in the lumen of the digestive organ (Fig. 77, C}. If it is a polycystic or multi-chambered form, the exposed portion becomes differentiated into protomerite and deutomerite, while the intra- cellular portion remains as the epimerite. After growth, the surplus food is stored in the endoplasm in the form of granules as described A B Fig. 81. Lymphosporidium truttce Calkins. A. The young sporozoite and its development. B. Older forms in the muscle-bundles surround- ing the intestine. C. Still older amoeboid form prior to, and during, spore-formation. above, to be used during the process of spore-formation and encyst- ment. In Sarcosporidiida and other muscle-infesting Sporozoa, growth takes place at the expense of the muscle-cells, although the organisms are not intra-cellular parasites. Thus, Lymphosporidium trutta begins to grow in the lymph surrounding the intestine. The sporozoite develops into a small amoeboid form which penetrates the muscle- 148 THE PROTOZOA bundles, and there grows to adult size by absorbing food destined for the muscles. When mature, it leaves the muscle-bundles and returns, to the lymph-spaces, where it sporulates (Fig. 81). D. MOTION In addition to the amoeboid motion which has already been men- tioned, there are various movements, due to the contraction of the myonemes or of the entire ectoplasm. Among these may be men- . " ' -; m ?* "Hi : - 'Sf ,-5-z' ^-*^-^*^~^ ^ * *< - Pig. 82. Cortical modifications and movement of a gregarine. [SCHEWIAKOFF.] A. Moving gregarine with paths of excreted granules. B and D. The same, more highly mag- nified. Cand E. Details of structure, c, cortical plasm ; /, jelly-layer ; m, myoneme ; s t secretion. THE SPOROZOA 149 tioned the peristaltic contraction of certain Gregarinida, or the ener- getic jerking motion sometimes observed in the same forms. None of these movements, however, brings about a regular translation from place to place, and the Sporozoa are regarded as the most sluggish of the Protozoa. Food-seeking, which in free-living animals is the main occasion for locomotion, is here unnecessary ; for the adult animals, placed in the chyle of the host, or in the spaces between cells and tissues, or in the cells themselves, have little occasion for movement, save that which, in young forms, is necessary to reach the host, to maintain their positions, and to prevent displacement. There is, however, in certain forms of Gregarinida, a peculiar gliding motion on the part of the adult organism. This is accomplished without apparent exertion of any kind by the animal, and for a long time was a puzzle to students of the group. Schewiakoff ('94) offered an explanation, based upon actual observation and experiment, and although very improbable at first sight, it is the only one thus far that fits the case (Fig. 82). These observations have been confirmed re- cently by Siedlecki (') wno accepts Schewiakoffs interpretation, while Lauterborn also gives a similar interpretation of the movement in diatoms. According to Schewiakoff, the forward, gliding motion is the result of the active secretion of the gelatinous substance from the ectoplasm, which accumulates below the membrane to form a gelatinous layer. The membrane of the cell, as described above, is marked externally by clear longitudinal grooves, and the gelatinous substance after filling these grooves, instead of spreading over the surface of the membrane, flows down and backward in the grooves to the posterior end of the body, where the secretion from different furrows unites to form larger currents, and these, in turn, form still larger streams, which, like a spider's web, solidify upon leaving the body (/?). Thus, a solid cylinder is formed behind the animal, the pos- terior end of which fits into the basin-like depression like a cast in its mould. The addition of new jelly by active secretion in the ectoplasm, and the resistance of the solidified portion, causes a forward move- ment of the animal. The movement, Schewiakoff further observes, is only periodic, for the flowing of the jelly is more rapid than the secretion a fact which explains the occasional absence of the external gelatinous layer. E. REPRODUCTION The most characteristic phenomena connected with the Sporozoa are those of reproduction and development. The many methods occurring in the other forms of Protozoa are here limited to spore- formation, although Labbe" describes rather questionable simple divi- Fig. 83. Types of spores. [WASIELEWSKY ; A. SCHNEIDER ; THELOHAN, etc.] A. Eimeria nepce. /?, C. Barroussia ornata. D. Tailed-spore of Gregarine. E, F. Ophryocystis Biitschlii, with multiple epispores. G. Ceratomyxa sphterulosa. H. Klossia helicis. I. Crystallo- spora Thelohani. J. Leptotheca agilis. K. Myxobolus ellipsoides. L. Crystallospora crystalloides. M. Goussia clupearum. N. Adelea ovata. Coiled threads are shown in G and J, the extruded thread in K. THE SPOROZOA 151 sion in Coccidiida (Fig. 6, p. 20). Spore-formation is almost invariably preceded by encystment, an exception being found in the Gymno- sporea and Myxosporidiida. In general, it may be stated that the entire organism takes part in the formation of archispores (or sporoblasts\ each archispore gives rise to spores, and each spore to sporozoites, either .directly or indi- rectly. Each spore, containing from one to many sporozoites, is coated by either a single or a double membrane. When double, the inner membrane is called the endospore, and the outer the epispore (Fig. 83, F). The spores may be of similar or dissimilar size (inacrosporcs and tnicrospores\ they may be ovoid, spherical, biconvex, cylindrical, crystalline, discoid, etc., in form, and may be provided with diverse . kinds of appendages, ridges, spines, etc., or with polar capsules con- taining protrusible filaments (Myxosporidiida). In some cases there is a special apparatus for the dissemination of the spores (sporodncts, Fig. 85) ; in other cases, the spores are liberated by the simple bursting of the outer envelope, or by the rupture of the walls through swelling of a residual protoplasmic mass termed a pscudocyst. The process of spore-formation in the gregarine of an ascidian Monocystis ascidice may be given as an example of a type common to all Sporozoa, although in the several orders the details are vari- ously modified. Two animals come together and form a common cyst (Fig. 84, A). The nucleus of each divides by repeated mitoses into a great number of daughter-nuclei, which soon arrange themselves about the periphery (B, C) like the nuclei of a centrolecithal egg of some Metazoa. A portion of the endoplasm is then budded off about each of the daughter-nuclei, the buds thus formed becoming conju- gating gametes. The bulk of the original cells is not used in this pro- cess, a considerable portion which Labbe ('96) regards as a reserve store of nutriment J remaining unused ( Theilungskorper, Cystenrest, Reliquat de segmentation}. During this process the ectoplasm and the membrane in each cell disappear, leaving the gametes and the cen- tral residual masses within the cyst (D). The gametes now conjugate two by two (E) to form the spores (sporocysts). Each of the spores, which from their peculiar shape are known as pseudonavicelke t now in its turn secretes two distinct membranes (epispore and endospore), and within these the nucleus, with its surrounding plasm, divides into eight parts which are disposed quite regularly in the spore (F\ As in the formation of the archispores, a portion of the plasm is usually left unused (Sporenrest, Restkb'rperchen, Reliquat de differentiation}. Each of these parts is a sporozoite, which, after a developmental period, reproduces an adult gregarine. When mature, the spores or pseudo- navicellae are liberated by the bursting of the outer cyst-walls, brought 1 See, however, Thelohan, '95. 152 THE PROTOZOA about either by the simple rupture of the wall or by the swelling of the central mass of useless material. The spores are thus freed, but not the sporozoites ; the latter are still confined within their double walls, and cannot be liberated until they are swallowed by some host, where, in the digestive tract, the two coatings are dissolved off by the digestive fluids, and the sporozoites emerge in the form of minute elliptical bits of protoplasm, each containing a nucleus. Fig. 84. Scheme of speculation in gregarinida. A. Union of two individuals in a common cyst. B and C. The formation of gametes of similar size. D. Union of the amoeboid gametes. E and F. Formation of sporozoites in the fused gametes. The process of spore-formation in the many-chambered Gregarinida is more complicated. Thus in Clepsidrina, a frequent parasite of insects, the organism when mature throws off the epimerite by which it is attached to an epithelial cell of its own host and, as a sporont, secretes its cysts and undergoes nuclear division as in Monocystis. The encysted animal, however, is carried to the exterior with the faeces of the host, and sporulation is outside of the host or exogenous, as opposed to the endogenous sporulation of Monocystis. In these excreted cysts, according to Schneider ('75) and Biitschli ('84), the archispores, instead of, as in Monocystis, forming a peripheral layer THE SPOROZOA 153 about a central residual mass, lie in the centre, the unused portion of the original protoplasm forming a thick layer about them. At the same time, a third and very delicate membrane, probably com- posed of the residual peripheral mass, is formed inside of the cyst and against the second or inner coating. Six to eight radial thicken- ings can be seen later in this residual portion, and each of these develops a distinct lumen, thus becoming tubular and extending through the residual mass of protoplasm to the new internal mem- brane. Each tube expands at the extremity into a disc-like cup, while the inner part of the tube is lost in the central mass of spores. In some unexplained way the walls of the primary cyst open, leaving the protoplasm and the spores inclosed only by the third membrane. The tubes already formed then evag- inate, and the cylindrical portion of the tube is thrown to the outside. The tubes act as spore- ducts for the inner archi- spores, each of which contains the definite num- ber of sporozoites (Fig. 85). Sporulation of the Coc- cidiida is strikingly similar to that of the Gregarinida. Here, as a rule, only one membrane (capsule) is formed around the spheri- cal animal ; and the nu- cleus, in addition to division through mitosis, frequently fragments into as many pieces as there are to be archispores (fragmentation). Before the nucleus divides, a certain amount of the chromatin is given off, as in the Gregarinida, to form what Labbe calls the equivalent of the " polar body " of the Metazoa. Again, as in the Gregarinida, the archispores or sporo- cysts are arranged around the periphery, and a residual mass occu- pies the centre. The archispores which are liberated by simple rupture of the walls of the cyst, form a definite number of sporozoites, varying from one (monozoic)ov two (dizoic} to many (polyzoic). In some forms of Coccidiida either sporozoites or archispores may be formed directly. The number of spores formed is usually small, as in Coccidium, where the nucleus divides only twice, producing only four archispores, each Fig. 85. Spore-ducts of Gamocystis tenax. [A. SCHNEIDER.] d, spore ducts ; s, spores in an external gelatinous mantle. 154 THE PROTOZOA of which gives rise to two sporozoites. In other cases (viz. Pfeiffcria Labbe), a great number of nuclear divisions may take place, and the final daughter-nuclei with their surrounding protoplasm form sporozo- ites directly and without an intervening archispore stage. A similar direct sporozoite-formation takes place among the Haemosporidiida, the sporozoites being frequently of two kinds, macrosporozoites and microsporozoites. While not established, it is probable that in all forms this dimorphism in the spores has a sexual significance, the same individual giving rise to only one form. One peculiarity of these sporozoites is that the nucleus is apparently never provided with a nuclear membrane, the chromatin, as in some flagellates, lying freely in the plasm. Sporulation in the tribe Gymnosporea takes place without the pro- tection of a cyst. The parasite rounds out, but does not secrete a membrane. The nucleus divides into a great number of parts, which migrate to the periphery as in other forms, and there divide. Sporozoites are formed directly without preliminary spore-stages. An entirely different mode of sporulation occurs in the Myxospori- diida, where the process is somewhat similar to the internal budding of some of the Ciliata. In the genus Myxobolus, for example, one of the numerous nuclei of the amoeboid form is surrounded by a thick- ened mass of protoplasm, so that it can be distinguished from the remainder of the animal. The thickened plasm soon forms a mantle about the nucleus, which then divides by mitosis until there are ten or a dozen daughter-nuclei within the specialized protoplasmic region (Thelohan, '95 ; Gurley, '93). This mass, the sporoblast, which, how- ever, does not quite correspond to the archispores of preceding types, now divides into two equal parts, both of which remain inside of the original protoplasmic mass. Each is an archispore, and each con- tains three of the ten nuclei. The other four nuclei are left in the free plasm within the membrane and soon degenerate and disap- pear, corresponding, apparently, to the residual mass of chromatin (polar body) of other forms. Each archispore next divides into three cells (Biitschli, Balbiani for Myxobolus\ two of which are destined to form peculiar thread-bearing capsules known as the polar capsules. The other is much larger and represents the definitive spore. Each sporoblast thus contains one spore, whose nucleus soon divides to form the two nuclei which characterize the young myxospore. The formation of the thread in the polar capsules according to Thelohan ('95) seems to be the same in all species; a vacuole appears in each of the smaller cells of the sporoblast (Fig. 86), then a small knob-like projection grows up from one side of the vacuole, whose outer walls harden until a distinct capsule is formed. The bud of protoplasm within the vacuole now elongates and winds around until a spirally THE SPOROZOA 155 wound filament is the result. The nuclei of the two polar capsules soon degenerate and disappear, leaving only the capsules with their threads, which show a striking similarity to those of the nematocysts of the Coelenterata. The archispore in many cases develops a bivalve shell, and in this condition can remain for some time within the original spore-forming body (pan-sporoblast); or the original membrane may be thrown off, leaving the encapsuled spores sus- pended freely in the endoplasm of the parent organism. In most cases there is no means of exit for the spores from the body of the host until the latter dies. The archispores thus accumulate until great cysts, sometimes as large as 30 mm. in diameter (Zschokke, '98, E H Fig. 86. Myxobolus ; capsule-formation. [THELOHAN.] A-D. Division of the sporoblast nucleus. F. The sporoblast is divided into two " sporogenous masses " each containing three nuclei. G. Sporogenous mass with protoplasm of the spore and two masses which are destined to develop into capsules and filaments. H. The threads are first seen as buds in the vacuole. Myxobolus bicaudatus), are formed within the tissues of their host. When taken into a new host, the shell of the archispore under suitable excitant, either chemical or physical, soon opens, and the filaments contained within the capsules are thrown out, according to Balbiani, through special apertures, and by the pressure of the capsular walls (Biitschli). These filaments, several times the length of the spore, remain attached, their free ends being swayed about by the currents until they come in contact with and penetrate some cell of the mucous membrane of a new host. The parasite thus anchored retains its position in the lumen until its bivalve shell is thrown off and it can move for itself (Leuckart, Biitschli, Gurley 1 ). In the Myxosporidiida, therefore, sporulation is not the final act of a cell-parasite, but takes place while the animal is performing other 1 For discussion of various views see Gurley ('93). 156 THE PROTOZOA normal vegetative functions. It is a case of cellular division of labor in which possibly some of the multiple nuclei are specially differ- entiated for reproduction. The number of archispores formed in the pan-sporoblasts varies in the different species and genera. In some cases there is but one (Chloromyxiim}, in others a large number (Gluged). The polar capsules, which are particularly characteristic of this type of Sporozoa, also vary in number and in position. They may both be at one end of the spore (anterior), as in Myxobolus (Fig. 83, K\ at the two ends, as in Myxidium, or in the centre, as in Ceratomyxa (Fig. 83, G). The Sarcosporidiida resemble the Myxosporidiida in forming spores throughout life. The peculiar pouch, which corresponds to the amoe- boid body of the Myxosporidiida and which may grow to a consider- able length (up to 16 mm. in sheep), is filled with masses of nucleated protoplasm which may be called pan-sporoblasts. Those in the centre of the pouch become coated by a membrane and divide into a number of germs or sporozoites known as Raineys Corpuscles, which in some cases appear to have polar thread-bearing capsules similar to those of the Myxosporidiida. The life-history and mode of infection of new hosts is unknown. Conjugation is a well-authenticated phenomenon in at least three orders : Haemosporidiida, Gregarinida, and Coccidiida, although the observations have not been numerous enough to warrant further generalizations. Among the Haemosporidiida, where the intra-cellular parasites frequently leave their cell-hosts, there is a shorter or longer period of free life. During this period two individuals, upon meeting, fuse together, forming one individual (Labbe). The nuclei also fuse, forming a single nucleus. It is an instance of total conjugation, similar to the total fusion in some Monadida, but, unfortunately, the significance of the process and the bearing upon the life-history of the individuals are entirely unknown. The union of two individuals within a common cyst is not infre- quently observed among the Gregarinida, and has been a long-known phenomenon. Two or more individuals may join end to end, pro- tomerite to deutomerite, or side to side, and so form aggregates (Fig. 27, p. 58). If the individuals thus associated happen to be mature at the same time, they may develop a common cyst and so give the appearance of conjugation. Such pseudoconjugation frequently leads to the formation of catenoid colonies, where the protomerite of one (satellite} becomes attached to the deutomerite of another (primite). We are indebted to Wolters ('91), Siedlecki ('96, '98, '99), and Schaudinn ('96, '99) for more complete accounts of conjugation among the Gregarinida and Coccidiida. According to the former, two gregarines (Monocystis agilis} place themselves end to end, but THE SPOJKOZOA 157 without fusing. The nuclei of the two cells then divide by mitosis, and in each case one of the daughter-nuclei is thrown off as a useless moiety in the same way as a polar globule. The other two daughter- nuclei move toward the partition wall which separates the two individuals, and meet each other in an opening of this wall. They fuse, and this fused mass divides by mitosis, one of the daughter- halves going to each of the conjugants. The nuclei then divide repeatedly, and spores are formed in the usual manner. This method if correctly observed, in contradistinction to pseudoconjugation among the Gregarinida and Haemosporidiida, is nuclear conjugation as seen in its highest development among the Infusoria ; but, unfortu- nately, there are no observations similar to those of Biitschli, Engel- '" f ; Fig. 87. Monocystis ascidice Lankest. [SlEDLECKI.] A. Fusion of two individuals. B. Formation of gametes (cf. Fig. 84). C. Nuclear division after the fusion of gametes, and sporozoite formation. mann, Maupas, and others on Infusoria to indicate the significance, and the facts themselves rest upon the observation of a single observer (VVolters). In a closely related form (Monocystis ascidia), Siedlecki ('99) describes an entirely different process. Here two individuals come together in a single cyst, within which each forms a number of merozoites or gametes. The gametes fuse together, and thus affect the conjugation of the two original individuals (Fig. 87). The greatest advances in our knowledge of the reproduction in Sporozoa, during the last ten years, have been in connection with the Coccidiida, and modern research has shown that the life-history of these forms is bound up with a complicated alternation of generations, the product of the union of sex-cells being the permanent spores by which the infection is carried from one organism to another, while the products of asexual increase lead to auto-infection within the same host. Up to the last five years the usual description of the life-history of 158 THE PROTOZOA Coccidiida followed that of Leuckart ('79) in the case of Coccidinm oviformis, a parasite of the rabbit. According to this view, the adult Coccidinm, which consists of a globular or oval mononucleate parasite, living in the epithelial cells of the digestive tract and the related organs, encysts and falls into the lumen of the digestive tract, from which it is defecated with the faeces. Inside of the cyst the plasm divides into several parts (in Coccidium four), and these parts, after the formation of a firm, resisting membrane, form the permanent spores. Each spore divides into two parts in Coccidinm, and these two parts constitute the end product of reproduction, according to the older view. Each of the parts forms a germ or sporozoite, which penetrates a new cell-host and develops again to the adult organism. This cycle, while perfectly logical, left unexplained the immense multitudes of parasites found in the epithelial cells of every Ceccidium-intected rabbit. The first attempt to explain its wide distribution was made by R. Pfeiffer ('92), who insisted that, in addition to this exogenous spore-formation, there exists an internal reproduction as well, which leads to further infection in the same host, or, as he called it, to auto-infection. In contrast with the first method, this was called the endogenous sporozoite-formation. This view was based upon the discovery by Pfeiffer of spore-forming cells in the tissues of the host, in addition to those in the lumen of the digestive tract. The majority of investigators along this line have accepted the latter view. There are two notable exceptions, how- ever, one of whom is Labbe, who holds that these smaller forms are only poorly fed individuals and not sporozoites, and explains the undeniable auto-infection through simple division of the parasites. A large number of papers soon followed, some on Coccidium, others on related forms. Mingazzini ('92) followed out the multiple division of the nucleus in the formation of the endogenous sporozoites. Podwyssozki ('94) made the discovery that there are two kinds of these endogenous forms, which were accordingly named microsporo- zoites and macrosporozoites. It was Schuberg ('95), however, who first suggested, although he did not confirm the suggestion, that these two forms of sporozoites conjugate and thus lead to sexual reproduction. Labbe strongly opposed the latter view, and held that the larger types of supposed dimorphic spores belong to some un- known species of Coccidiida, and that the smaller forms are degen- eration types. The way was thus prepared for the discovery of conjugation among the cells of the Coccidiida, a discovery made first by Schaudinn and Siedlecki ('97). In two different species, Coccidium Schneidcri and Adelca ovata, it was found that a large cell, an egg, is fertilized by a THE SPOROZOA 159 small one, which has all of the characteristics of a spermatozoon. In the same year Simond worked out again the life-history of the Coc- cidium of the rabbit and described a true copulation between the microsporozoites. Schaudinn regards this, however, as an error, holding that copulation takes place between one of the smaller forms and an enlarged ordinary individual. Since then, the fact of fertilization, with the resulting formation of sporozoites through spores, has been safely established for a number of species by several different observers, the details alone differing in the several cases. The microsporozoites thus are not true sporo- zoites, but gametes having a sexual function. According to these various observations the life-history of the Coc- cidiida may now be described as follows : the permanent cysts contain spores, each of which contains sporozoites which are taken into the digestive tract with the food. Here the cyst membrane bursts or is dissolved, and the sporozoites are liberated. They penetrate the epithelial cells and grow to the normal size of the adult. They then undergo repeated nuclear division by a process which resembles frag- mentation rather than mitosis (Schaudinn), or (possibly) in some cases by binary division also (Labbe), and the nuclear parts wander out to the periphery, where small portions of the cytoplasm form around them and they are pinched off as minute germs which Simond called merozoites. These differ in several important respects from the sporozoites, but like them are capable of developing directly into new adult parasites. This process, which Schaudinn calls schizogony, leads to the increase of parasites within the host (Fig. 88, a-c). Dur- ing development, some of these merozoites store up reserve nutriment and form large ovoid cells, while others form the mother-cells of the microsporozoites or spermatozoids, without storing up a reserved food supply. The small forms, in some cases, are provided with flagella, which were first made out by Leger and by Wasielewski. Fertiliza- tion takes place in a manner almost identical with that of the Metazoa (d-j}. In some cases a micropyle is formed in the egg through which a spermatozoon can enter, and in all cases after one has entered, a hard membrane corresponding to the vitelline membrane is at once formed (). Complete fusion takes place between the nuclei, and the cleavage nucleus divides by repeated mitosis to form spores. It is quite probable that the other cases of dimorphism, which have been recorded from time to time, are instances of similar sex-differen- tiation. The motile forms especially, which numerous observers have recorded, will probably be found to be similar in function. Laveran ('98), Bosc ('98), Sjobring ('97), Wasielewsky ('98), Sied- lecki ('98), and others have recently described them in different Coccidiida (Fig. 89). i6o THE PROTOZOA Conjugation in the malaria-causing organism (Plasmodium malarice} is bound up with a change of hosts, thus giving a complicated life- history, which may, and probably does, occur in other kinds of Sporozoa as well, although the phenomenon has been only recently made known. Fig. 88. Life-history of a Coccidium. [SCHAUDINN.] a, b, c, schizonts and asexual reproduction (schizogony). The merozoites at c repeat the cycle or pass on to the following stages, d, e,f, development of the female or macrogamete. h, i,j, development of the male flagellated gametes ; g, copulation of the male and female gametes ; k and /, stages in the formation of the four spores and sporozoites. The sporozoon which is now positively known to be the cause of "malarial disease," lives in the human blood under various forms, which may possibly be distinct species differing from one another in the number of spores produced and in the pathogenic effects. Sev- eral varieties at least have been described and specially named on account of minute differences, but it is probable that these can be THE SPOROZOA 161 reduced to three principal types : Plasmodium malaria, P. vivax, and Laverania malaria. These all agree in having an intra-corpuscular, ^aasv ^BB&SV"-"',... - T-T j~t Fig. 89. Conjugation and sporulation in Klossia hehcina, Labbe. [SlEDLECKi.j A-E. Formation of microgametes. F. Conjugation. G. Union of the male and female nuclei. H. Formation of spores. amoeboid stage during which the body-plasm becomes stored with melanin granules or metamorphosed haemoglobin. They agree also in having an intra-corpuscular spore-forming stage (schizogony, Fig. 90, F, G, L, M), the spores bringing about auto-infection by pene- 1 62 THE PROTOZOA trating new blood-corpuscles and there repeating the cycle ; and under certain conditions they all produce flagellated bodies or spores {Polymitus form, R\ They differ in the number of spores that are formed, although in the same variety the number appears to be incon- stant, so that mere difference in number cannot be considered a good specific character. A much more satisfactory means of distinguishing them lies in the regular periodicity of spore-formation, which accom- panies well-marked morbid symptoms in the patient. Thus, in Plasmodium vivax, spore-formation occurs every forty-eight hours (ap- proximately) ; in P. malaria, every seventy-two hours. The pyrexial attacks in all cases consist of similar symptoms, a stage of fever following one of chill and followed by a stage of perspiring. Spore- formation is the signal for a chill in the patient. The pigment granules (melanin) become aggregated in the centre of the cell, while the protoplasm, breaks up into spores about them. The blood-corpuscle which contains the parasite then disintegrates, and the spores and melanin are liberated. It is supposed that this dis- ruption of the corpuscle, by liberating a toxin (melanin) created and stored up by the parasite, is the direct cause of the attack. If this hypothesis, which is certainly based upon considerable evidence, should be verified by future investigation, the malaria-organism will be the only protozoon known to produce poisonous growth-products. Quinine in the system is apparently fatal to the parasite by preventing its growth and sporulation. The spores of the malaria organism are not covered by a protective coating as in the majority of Telosporidia, and are, therefore, unsuited for an exposed life outside of their host. It was early recognized, however, that there must be an extra-corporeal period in the life- history of the parasite in order that the species should be perpetuated. That there actually is such a period is shown by the spread of the disease throughout a community. 1 Nevertheless, the whereabouts of the organism and its form during the extra-corporeal existence have remained a mystery until within the last few years. The key to the puzzle has been given by the flagellated body or Polymitus form. The blood when examined fresh from a malaria patient shows the ordinary form of the parasite, but after a short period of exposure to the air (10 to 30 minutes, Manson, '98), the parasite develops long flagelliform processes, which vibrate with great vigor and not infrequently break away from the body of the cell to swim about like Spirilla in the plasm. Danilewsky ('91) believed this stage to be an independent flagellated parasite of a special nature, and he named it Polymitns. Lave"ran, Metsch- 1 In Italy it is estimated that 2,000,000 people are ill every year with malaria (Santori, 1900). THE SPOROZOA 163 nikoff, Manson, and others regarded it as an essential develop- mental stage of the malaria organism, and believed that the free- Fig. 90. Life-history of Malaria, causing Sporozoa. [Ross and FiELDlNG-OULD.] A-F. Stages in the development of merozoites. A- 1. The sporozoite. B, C, D, and jf, K, L. The growing sporozoite in blood-corpuscles. . Asexual reproduction (schizogony). f, G, and M. Liberation of merozoites and melanin granules. O- W. Stages in the development of sexual individuals. R. Polymitus form. S. Fully developed microgametes. T- W. Development of the female individual (macrogamete). X. Fertilization of a macrogamete by a microgamete. Y. The fertilized cell copula, with its vitelline membrane, a-e. The copula in the stomach of the mosquito {Anopheles sp.). b. The copula penetrating the epithelium which lines the stomach of Anopheles, b, c, d, e. Growth of the copula in the body-cavity of Anopheles. The small spheerules at V, W, and X are supposed to be analogous to polar bodies of metazoan eggs. f. Sporulation in the body- cavity of Anopheles, g. Liberation of the sporozoites. h. Salivary gland (in section) , with sporo- zoites in the lumen, in the cells, and penetrating the membrane. swimming detached processes are germs which reproduce the adult. Many others (Grassi, Felletti, Celli, Sanfelice, Sacharoff, Labbe, etc.), however, regarded these forms as degenerating parasites induced by the abnormal conditions of exposure and not present, normally, in 1 64 THE PROTOZOA the blood. Manson ('96) was one of the first to call attention to the fact that the formation of these so-called degeneration forms, which occurs only at the time when the blood is exposed to the air, is evidence of the beginning of extra-corporeal life. He suggested a theory that the Polymitns forms are flagellated spores, the extra- corporeal homologue of the intra-corporeal spores. He further suggested that, since the parasite is normally incased within a blood- corpuscle, it is unable to leave the host by its own efforts and must be removed by some blood-eating animal, probably a suctorial insect, such as a mosquito, common in swampy, malarial regions. At the same time ('96) Laveran, in France, proposed an identical hypothesis. Subsequent investigation has given the complete confirmation of this hypothesis. The work of Major Ross, in India, of Koch, Grassi, and others, elsewhere, has established beyond a doubt, that extra-corporeal life of the parasite is spent in the mosquito, and that the disease is spread by these insects through inoculation. About eight or ten days after drawing blood from a malaria patient, the insects are able to transmit the germs to new hosts by inoculation through the proboscis. After a number of experiments, Ross and Grassi found that certain genera of mosquitoes, e.g. Culex s/>., are incapable of fostering the human parasite, while all species of the genus Anopheles are particu- larly susceptible. The history of the parasite in the mosquito has been variously in- terpreted. Manson ('96, '98), on a priori grounds, suggested that the Polymitns form is developed after the blood is taken from the host into the colder digestive tract of the insect, the change of medium acting upon the parasite in the same way that the air does. Here, he argued, it penetrates an epithelial cell and repeats the life-history of an ordinary form, sporulating and increasing by auto-infection. Mac- Callum ('97), however, described a true conjugation between a Poly- mitus form and a free pigmented parasite in a very similar organism (Halteridium Labbe), which is parasitic in the blood of the Ameri- can crow. In this case the impregnated Halteridium slowly changes form, becoming elongated and more or less worm-like, and moves about in the blood-plasm. Thus in this case the Polymitns form corresponds to a spermatozoon, and the ordinary individual, as in Coccidium, to an egg. Ross and Grassi, finally, have demonstrated the same relation in Plasmodium malaria ; the Polymitus form fuses with an ordinary individual in the intestine of the mosquito, and as in Halteridium^ the copula becomes a motile individual which, after a short period, penetrates the epithelial cells lining the digestive tract (Fig. 90, a-e). Here it resembles one of the Coccidiida, growing at the expense of the cell-host and finally sporulating. The spores do not form protective coatings, but divide at once into sporozoites (/, g). THE SPOROZOA 165 These make their way into the body cavity, or lymph spaces, of the mosquito, and ultimately find their way to the salivary glands, from which they may be deposited, together with the salivary fluid, in the blood of man (//). The life-history of a malaria organism thus involves a complete change of hosts, one phase being in the warm blood of man, the other a Coccidia-like stage in the Insecta, a group which above all others is noted for the frequency and number of sporozoan parasites. - It is not improbable that a similar change of hosts occurs in the parasites belonging to this same group of Haemosporidiida, which have been observed in birds (Laveran, Labbe, '94); in reptiles (Labbe, '94; Langmann, '99); and amphibia (Labbe, '94; Lang mann, '99); also it is possible that many of the uncertain forms, such as Scrumsporidinm of the Crustacea (Pfeiffer, '95) or Lymphospo- ridinm of the trout (Calkins, '99), have a similar complicated life- history. Apart from their pathogenic effects in man, the Sporozoa are frequently a pest in the lower animals. The Sarcosporidiida have already been mentioned as producing morbid symptoms resembling Trichinosis, in the domestic animals often leading to death. The Myxosporidiida occasion great loss to fish culturists by causing ulcers which ultimately result in the death of the fish, and to silkworm cul- turists on account of costly and extensive epidemics produced by them among the silkworms. These organisms (Glugea bombycis] were so disastrous to the silk industry during the years 1854-1867, that a loss was estimated of at least i ,000,000,000 francs (about $190,000,000). In regard to this epidemic Huxley ('70) writes: "In the years following 1853 this malady broke out with such extreme violence that, in 1858, the silk crop was reduced to a third of the amount which it had reached in 1853 ; and, up till within the last year or two, it has never attained half the yield of 1853. This means not only that the great number of people engaged in silk growing are some 30 millions sterling poorer than they might have been ; it means not only that high prices have had to be paid for imported silkworm eggs, and that, after investing his money in them, in paying for mul- berry leaves and for attendance, the cultivator has constantly seen his silkworms perish and himself plunged in ruin ; but it means that the looms of Lyons have lacked employment, and that, for years, enforced idleness and misery have been the portion of a vast popula- tion which, in former days, was industrious and well to do." The caterpillars, although infested by the parasites which were frequently so numerous that all of the organs of the body swarmed with them, were nevertheless able to produce the moth. The latter, though stunted and undeveloped, could lay eggs which them- selves contained spores of the organism, and these spread the disease. 1 66 THE PROTOZOA The disease was checked only by careful examination of the food of the caterpillar, and by microscopic examination of all eggs and rejec- tion of the infected ones. No remedy is known for the many other diseases due to Sporozoa, especially among domestic animals, or fresh-water fish, and careful prophylactic measures analogous to those employed in stopping the silkworm epidemic may be the only means of checking them. Such measures have already been successfully applied to prevent the spread of malaria, and the experiments which are now going on in all parts of the world justify the hope that this disease will be ultimately stamped out. F. INTER-RELATIONSHIPS OF THE SPOROZOA i The Sporozoa, modified beyond doubt by adaptation to a parasitic mode of life, have ever been a puzzle to systematists. Kolliker ('48) early suggested that they are single cells, and included them in his Protozoa. Stein ('48) agreed with him as to their primitive structure, but was loath to regard them as single animal cells, and compromised by calling them Symphyta, a group of the Protozoa. Another view, developed by Henle ('45) and Bruch ('50) and taken up by Leydig ('51) and Leuckart ('52), was based upon the superficial resemblance of the Gregarinida to Nematode worms. It found little support, how- ever, against Kolliker's view. Still another theory of the origin of the Sporozoa has been held by those who, following Gabriel ('75, '80), regard these forms as plants, placing them with the Mycetozoa, among the Fungi. Biitschli at first 1 favored the view that the Sporozoa are derived from the Rhizopoda, basing his belief upon the method of reproduction, general morphology, and physiology. Later, however, 2 he considered their relationship to the Flagellidia as much more close, not to the simplest forms, but to the higher types with a well-differentiated cuticle. The flagellum and mouth parts, he as- sumed, became lost with gradual adaptation to the intra-cellular mode of life, while the methods of reproduction became specialized in response to the requirements of a new environment. This view is strengthened by the close agreement in finer structures of the Grega- rinida and the Flagellidia, especially as regards the differentiations of the cuticle and the presence of muscular elements. Their move- ments, too, recall those of the Flagellidia, especially certain species of Astasia, where, in the non-flagellated condition, the plasm moves forward by a peculiar peristalsis, while the secretion of a jelly from the sub-cuticular or cortical plasm is identical in the two groups. The nuclei show perhaps a closer resemblance to those of the Rhizopoda 1 ('83), P- 479- 2 ('84), p- 807. THE SPOROZOA 167 than to the Flagellidia, but the conjugation processes are much more like those of the Flagellidia. Haeckel follows Biitschli in regarding the Sporozoa in this light, and derives them from the Phytoflagellida through adaptation, first to a saprophytic and then to a parasitic mode of life. Wasielewsky also favors the flagellate origin, basing his opin- ion, however, upon the uncertain ground of flagellated swarm-stages of certain Sporozoa as well as upon the general resemblance to the Astasiidae. In general, however, it must be admitted that there is very little support for any one of these theories, and all attempts to trace the origin of the Sporozoa upon the mere basis of their present degenerate condition are highly speculative. CLASSIFICATION CLASS III. SPOROZOA. The Sporozoa are Protozoa which are never provided with flagella or cilia in the adult state. They are always endoparasites in cells, tissues, or cavities of other animals, and food is taken in by osmosis. Repro- duction is always by spore-formation, and germs {sporozoites) are produced either directly from the parent, or indirectly through spores. Subclass I. TELOSPORIDIA. Sporozoa in which spore-formation ends the indi- vidual life, the entire cell then forming spores. Order i. GREGARINIDA. Telosporidia possessing a distinct membrane, with myonemes during adult life, locomotion being accomplished mainly by their contraction. The young stages alone (cephalonts) are intra-cellular parasites, the adults (sporonts) being found in the digestive tract or the body cavities. Sporulation takes place after or without conjugation, but within a cyst which is never formed while the parasite is intra-cellular. Suborder i. CEPHALINA. Gregarinida possessing an organ for attachment (epimerite), and with or without septa dividing the cell into chambers. Tribe i. Gymnosporea. The adults are solitary or associated ; the sporozoites are formed directly from the adult without encystment. Family i. Aggregatidae. Colonies consisting of two or more individuals. Several residual protoplasmic masses are found during sporulation in each cyst. Genera: Aggregata Frenzel ('85). Family 2. Porosporidae . The individuals are usually solitary. The sporozoites are arranged in groups around a central residual mass. Genera : Porospora A. Schn. ('75). Tribe 2. Angiosporea. Cephalina with well-developed spores, which are provided with spore-membranes (epispores and endospores) . Family i. Didymophyidae . Chain-forming aggregates, two individuals being so closely joined as to appear like one with three chambers. Genera : Didymophyes Stein ('48). Family 2. Gregarinidae . The individuals are solitary or associated. The epimerite is simple and regular. The cysts may or may not have spore-ducts. Genera : Gregarina Dufour ('28) ; Gamocystis A. Schn. ('75) i Hirmocystis Le"ger ('92) ; Hyalospora A. Schn. ('75) ; Euspora A. Schn. ('75) ; Sph&rocystis Le*ger ('92) ; Cnemidiophora A. Schn. ('82) ; Stenophora Labbe ('99). Family 3. Dactylophoridae . The epimerite is asymmetrical and irregular. Genera : Rhopalonia Leger ('93) ; Echinomera Labbe ('99) ; Trichorhynchus A. Schn. ('82) ; Pterocephalus A. Schn. ('87) ; Dactylophorus Balbiani ('89). Family 4. Actinocephalidae . The individuals are always single. The epimerite is 1 68 THE PROTOZOA simple or lobed and symmetrical. The cysts open by simple dehiscence. The spores are boat-shaped, bi-conical, or cylindro-conical. Genera: Sciadiophora Labbe" ('99) ; Antlwrhynchus Labbe" ('99) ; Pileocephalits A. Schn. ('75) ; Amphoroides Labbe" ('99); Discorhynchus Labbe ('99); Stictospora Le"ger ('93) ; Schneideria Le"ger ('92) ; Asterophora Le"ger ('92) ; Stephanophora L(?ger ('92) ; Bothriopsis A. Schn. ('75) Coleorhynchus Labbe" ('99) ; Actino- cephalns Stein ('48) ; Pyxinia Hammerschmidt ('38) ; Legeria Labbe" ('99) ; Phialoides Labbe" ('99) ; Beloides Labbe" ('99). Family 5. Acanthosporidae. Solitary. The spores are provided with equatorial or polar spines. Genera: Corycella Le"ger ('92); Acanthospora Le"ger ('92); Ancyrophora Leger ('92); Cometoides Labbe" ('99). Family 6. Menosporidae. Solitary. The epimerite is on a long neck. The spores are crescent-shaped. Genera: Menospora Le"ger ('92); Hoplorhynchus Carus ('63). Family 7. Stylorhynchidze. The spores are formed in chains, and the cysts have a double envelope. Genera : Lophocephalus Labbe ("99) ; Cystocephalus A. Schn. ('86) ; Oocephalus A. Schn. ('86) ; Sphcerorhynchus Labbe" ('99) ; Stylorhyn- chus Stein ('48). Family 8. Doliocystidae. Cephalina without septa dividing the cell into protomerite and deutomerite, but consisting of a single chamber with epimerite. Genera : Doliocystis Le"ger ('93). Suborder 2. ACEPHALINA. Gregarinida consisting of a single chamber, and with- out epimerite. They are parasites in the body cavity or cavities of the various organs of different animals. Genera : Monocystis Stein ('48) ; Zygocystis Stein ('48) ; Zygosoma Labb ('99) ; Pterospora Racovitza and Labbe ('96) ; Cystobia Mingazzini ('91); Lilhocystis Giard ('76); Ceratospora Le"ger ('92) ; Urospora A. Schn. ('75); Gonospora A. Schn. ('75); Syncystis A. Schn. 086). Order 2. COCCIDIIDA. Telosporidia having a spherical or oval form, without a free and motile adult stage, and never amoeboid. Sporulation takes place within cysts formed while the organism is an intra-cellular parasite. Family i. Disporocystidae . The cell forms two sporocysts, each sporocyst forming two or four sporozoites. Genera: Cyclospora A. Schn. ('81), with two sporo- zoites ; Isospora A. Schn. ('81), and Diplospora Labbd ('93), with four or more sporozoites. Family 2. Tetrasporocystidae. Each organism forms four sporocysts, each of which produces two sporozoites. Genera : Coccidium Leuckart ('79) (including Gous- sia Labbe") ; Crystallospora Labbe" ('96). Family 3. Polysporocystidae. Each organism produces an indefinite number of sporocysts, each of which produces one sporozoite, \_Barrouxia A. Schn. ('85),. Diaspora L^ger ('99)], two sporozoites, [Adelea A. Schn. ('75), and some species of Hyaloklossia Labbe" ('96)], three sporozoites, \_Benedenia A. Schn. ('75), or four, Genus Klossia A. Schn. ('75)]. Order 3. H.3LMOSPORIDIIDA. Sporozoa of small size living in the blood-corpuscles or plasm of vertebrates. The adult form is mobile, and in some cases is pro- vided with myonemes. They reproduce by endogenous or asexual spore- formation while in the host, and by exogenous spore-formation after conjugation. Genera : Lankesterella Labbe" ('99) ; Caryolysus Labbe" ('94) ; Hcemogregarina Danilewsky ('85); Caryophagus Steinhaus ('89); Halteridium Labbe* (""94); Hamoproteus Kruse ('90); Plasmodiitm Marchiafava & Celli ('85); Laverania Grassi & Feletti ('92) ; Cytamceba LabW ('94). Subclass II. NEOSPORIDIA. Sporozoa which form sporocysts throughout life; the entire cell is not used in the formation of spores. THE SPOROZOA 169 Order i . MYXOSPORIDIIDA. Neosporidia of amoeboid or spherical shape ; multi- nuclear. The initial free stage is passed in the cavities of the organs, or in the tissues of the host. In sporulation a definite or an indefinite number of sporo- blasts is formed, each of which gives rise to one or several spores ; the latter are provided with one or several polar capsules, which contain coiled threads like a nematocyst. Each spore gives rise to one amoeboid sporozoite. Suborder i. PIL3SNOCYSTINA. Spores with polar capsules distinctly visible when fresh. Family i. Myxidiidae. Myxosporidiida' forming two or more spores at the same time. Spores variable in form inclosing two polar capsules. Genera : Spharospora Thelohan ('92) ; Leptotheca The'l. ('95) ; CeratomyxaT\\&\. ('92) ; Myxidium Blitschli ('82) ; Sphceromyxa Thel. ('92) ; Cystodiscns Lutz ('89) : Myxosoma Thel. ('92). Family 2. Chloromyxidse . The spore has four polar capsules. Genera: Chloro- myxum Mingazzini ('90). Family 3. Myxobolidae. Adult stages very rare, ordinarily found encysted in the tissues ; usually polysporous. The spores have one or two polar capsules. Genera: Myxobolus Biitschli ('82) ; Hemieguya Thel. ('92). Suborder 2. MICROSPORIDIINA. Myxosporidiida in which the spores have but one polar capsule, which is invisible in the fresh state without the use of reagents. Family i. Nosematidce. With a bivalve spore. Genera: Nosema Na'geli ('57) (Glugea Thelohan, '92) ; Plistophora Gurley ('93) ; Thelohania Henneguy l ('9 2 )- Order 2. SARCOSPORIDIIDA. Sporozoa in which the initial stage is passed in muscle-cells of vertebrates. The form is usually elongate, tubular or oval, or sometimes spherical. It forms cysts with a double membrane, in which are formed kidney-shaped or falciform sporozoites, or else spores (?), provided with a polar capsule and projectile thread. Genera: Sarcocystis Lankester ('82). SPOROZOA INCERT^E SEDIS Amcebosporidia. Sporozoa possessing an amoeboid body, and reproducing either by division or by spore-formation after conjugation. Gene'ra : Ophryocyslis A. Schn. Serumsporidia. Sporozoa which reproduce by division (?) or by spore-formation, the sporozoites being minute oval or spherical bodies They are found in the cavities or ccelomic fluids of Invertebrates and Vertebrates. Genera: Serum- sporiditiin L. Pfeiffer ('95) ; Blanchardina Labbe ('99) ; Lymphosporidium Calkins (1900). SPECIAL BIBLIOGRAPHY V Gurley, R. R. The Myxosporidia, or Psorosperms of Fishes, and the Epidemics pro- duced by them. Bull. U. S. Fish Comm. XL, 1893. Labbe, A. Recherches zoologiques, cytologiques et biologiques sur les Coccidies. Arch. d. zool. exper. et gen. (3) IV., pp. 517-654, 1896. LabbS, A. Sporozoa. In Das Tierreich, Berlin, 1899. Leg6r, L. Recherches sur les Gregarines. Tablettes zoologiqrtes , III., pp. 1-182, 1892. Leuckart R. Die Parasiten des Menschen. Leipzig, 1879.' Ross. D. On Some Peculiar Pigmented Cells found in Two Mosquitoes fed on Mala- rial Blood. Brit. Med. and Surg. Jour., 1897, pp. 1786-1788. Schneider, A. Sur les psorospermies oviformes ou Coccidies, especes nouvelles ou peu connues. Arch. d. zool. exper. et gen. (i) IX., pp. 387-404, 1881. Siedlecki. M. Etude cytologique et cycle evolutif de la coccidie de la seiche. Ann. d. Vlnst. Pasteur, XII., 170 THE PROTOZOA Siedlecki, M. Ueber die geschlechliche Vermehrung der Monocystis ascidiae R. Lank. Bull. d. FAcad. d. Set. d. Cracovie, 1899. Thelohan. P. Recherches sur les Myxosporidies. Bull. Sci. de la France et de la Belgique, XXVI, pp. 101-394, 1895. Wasielewsky, Von. Sporozoenkunde, ein Leitfaden fiir Aerzte Tierarzte und Zoologen. Jena, 1896. CHAPTER VI THE INFUSORIA "Die Infusionsthiere gehoren in den Kreis der Protozoen. Innerhalb desselben bilden sie eine eigene und zwar die am hochsten stehende Klasse." STEIN. 1 As was long since clearly recognized by Stein, the Infusoria are the most highly differentiated of all Protozoa and often attain a .degree of complexity which is perhaps greater than in any other cells. Their form varies considerably in the several divisions, but all are characterized by certain structural features by which they can be distinguished at a glance. All are provided with cilia which may be retained throughout life (Ciliata), or may be replaced in the adult phases by suctorial tentacles, cilia being present only during the embryonic phases (Suctoria); they possess mouth parts which are adapted for swallowing, for simple ingestion, for sucking, or which may be entirely degenerate through parasitism ; and they are provided with two kinds of nuclei, known as macronuclei and micro- nuclei. They reproduce by simple division and by budding, or rarely by spore-formation. I. THE CILIATA Among the Ciliata the arrangement of the cilia upon the body affords a character which was first used by Stein ('59), and is still retained as a means of distinguishing the subdivisions of this group. In the first and probably the most primitive type, HolotricJiida, the cilia are arranged uniformly over the entire body of the animal and show no regional differentiations (Fig. 91, A). In the second type, Heterotrichida, the cilia are uniform over the main portion of the body, while a specialized set fused into a curved series of firm vibra- tory plates, or membranelles, are found in an adoral zone about the mouth (B). In the third type, Hypotrichida, the body is flattened dorso-ventrally and the dorsal side is entirely free from cilia, while on the ventral side the cilia are frequently fused together into stiff seta- like organs, the cirri, and as in the Heterotrichida, they may form a curved line of membranelles around the mouth (C). Finally, in the Peritrichida, the highest type of this class, the cilia are reduced to one or two bands or girdles in addition to the adoral zone (D). Although, with the exception of the motile organs, no single item of structure is found here which is not occasionally met with in other 1 ('59), P- 54- 171 1/2 THE PROTOZOA classes of Protozoa, yet in no other class are they all present in a single cell. Each of the different elements thus brought together has a definite function to play in the life of the organism, and intra-cellular division of labor is developed to a high degree. Leading an active life and forced to seek food in all sorts of places, from the clearest waters to the internal fluids of various hosts, the Ciliata have acquired a very great diversity of form. The simplest and probably the most primitive forms are monaxonic, the mouth being anterior and the anus posterior (Fig. 91, A). Symmetry, how- ever, is the exception and asymmetry the rule, the latter condition arising by the gradual shifting of the mouth to a more or less well- Fig. 91. Types of Ciliata. [BuTSCHLl.] A. Prorodon teres Ehr. ; an holotrichous form. B. Climacostomum virens Ehr. ; an hetero- trichous form. C. Pleurotricha grandis St. ; an hypotrichous form. D. Vorticella umbellaria ; a peritrichous form. z, adoral zone. defined ventral side, while the anus becomes more or less dorsal. The functional anterior end may thus be either ventral, or superior to the mouth, when the latter becomes sub-terminal. The simple monaxonic ground-type is subject to other minor variations among the Holotrichida, which point the way toward the more striking deviations among the Heterotrichida and the Hypotrichida. A fre- quent modification is the anterior prolongation of what might be con- sidered the upper lip, as in Dileptus or Lionottts (Fig. 92), where bilaterality and asymmetry are well established. The mouth becomes more and more ventral in the family Trachelinidae (Holotrichida), while in the order Hypotrichida it is always ventral and the original monaxonic structure is replaced by dorso-ventral differentiation and complete asymmetry. THE INFUSORIA 173 A. PROTOPLASMIC STRUCTURE As in all other Protozoa, the endoplasm consists of alveoli of vary- ing size and arrangement, the network being built up of plasm of greater or less density. Within the endoplasm there is a constant streaming of the granules, which varies greatly in the different species and even in the same individual under different conditions. In some / x. . 1/ N--4 Fig. 92. Dileptus anser O. F. M. [BUTSCHLI.] m, mouth ; N, macronucleus ; t, trichocysts on the tentacle-like end. cases the course of the streaming recalls cyclosis in some plant-cells, and frequently follows a well-defined and unvarying route (Colpoda cucullus, or Paramoscium burs aria). In all cases the current seems to start from the mouth, passing backward around the cell either to the right or left, then forward and back to the mouth (Butschli). In this stream, circulating through the body of the animal, are 174 THE PROTOZOA carried food-products in various stage's of digestion and assimilation, as well as excretory products in the form of granules similar in all respects to those found in other Protozoa. Engelmann ('83) found that in one of the Vorticellidae ( Vorticella campanula) the animal is colored by diffuse green pigment, which he took to be chlorophyl, and he further showed that oxygen is generated and that the animal can assimilate like a plant, but that it is not limited to this kind of nutrition, since it also takes in solid food through the mouth. Other colored particles which are found in various kinds of Ciliata, espe- cially in those forms which subsist upon plant food, are presumably due to the coloring matter contained in the food. Schewiakoff ('89) has shown that the colored balls which appear in some cases are merely fluid drops colored with the pigment contained in Oscillaria and other vegetable cells. In many cases, also, green algal cells live as sym- bionts within the endoplasm. Le Dantec ('92) found that these cells (Zoochlorella) are apparently taken in as food and become inclosed within gastric vacuoles, the fluids of which have no effect upon them. Soon the vacuoles disappear and the algae are left free in the plasm, where they live and multiply. There are also various fats and excretory products either crystalline or granular in form. The crystals, according to Schewiakoff ('93), are granules of calcium phosphate. 1 Among the pigmented inclusions of the cell must be noted the so-called " eye-spots " or stigmata, which, in some instances, are accompanied by lenticular differentiations of various kinds. These pigmented spots are, as a rule, mere heaps of granules colored either red, brown, black, or orange, and are probably the same in function as the similar products in Mastigophora. The protoplasm becomes more dense toward the periphery, and, as in the Sarcodina, it finally becomes too compact for the granules to penetrate. This outer portion is, therefore, comparable to the ectoplasm of the less differentiated Protozoa. The importance of this layer is seen in the fact that nearly all of the organs which characterize the Ciliata, including the myonemes, cilia, membranes and membranelles, the trichocysts, nematocysts, and the complex membranes and tests, are modifications of, or are produced by, the ectoplasm. In some forms the thickened plasm immediately adjoin- ing the endoplasm is distinctly marked off from the more external portions, forming a continuous layer around the entire body. This layer, which is in reality neither ectoplasm nor endoplasm, but inter- mediate between the two, is called the cortical plasm (Rindenparen- chyma, Stein), and is characterized by the reduced size and number of its vacuoles, by the absence of granules and streaming motion, and 1 Cf. p. 286, infra. THE INFUSORIA 175 by its fixity in the cell. It is occasionally thickened to form the denser ends of the body, as in the tail of Stentgr. In some cases, also, processes from the cortical plasm invade the endoplasm to surround the nucleus and hold it in a fixed position in the body (Dasytricha). The cortical plasm, furthermore, is the seat of the peculiar and characteristic offensive and defensive trichocysts. In some forms, probably representing the primitive condition, these are distributed about the body (Paramcecium\ but, even more than the cilia, they have been subject to reduction in most parts of the body until, in the majority of forms, they are restricted to a limited area, while in the Hypotrichida and Peritrichida they occur only sporadically. In Prorodon (Holotrichida), the trichocysts are found in the anterior end only, and in the family Trachelinida they are found only on the ventral side, while in Trachelius, Dileptns, etc., they are on the ventral side of the anterior process (Fig. 92). In Lionottts, they are reduced to a single line along the ventral side of the anterior process. The trichocysts are so minute that their finer structure has not been definitely made out, although a few different types have been studied (Fig. 12, C, p. 39). Rod-like forms have been seen in LoxopJiyl- Inm, Lionotus, and Strombidium, and spindle forms in Paramceciwn, Frontonia and Nassula. When protruded from the body they are, for the most part, apparently of the same size and shape as when within the ectoplasm. Occasionally when protruded, however, they have small hooks or swellings on the end (Maupas). They vary in size from three to twelve microns when within the body, but when protruded they measure from thirty to sixty microns. The cause of the protrusion is unknown ; certain reagents act as irritants and cause them to explode and throw out the long threads. Their func- tion, too, is purely conjectural, although it is generally supposed that they serve as defensive weapons. In some cases they appear to serve as weapons of offence as well, especially in those ciliates where they are limited in number to a comparatively few large ones. According to Maupas ('83), these Infusoria chase their prey and launch their trichocyst darts, which penetrate the outer coating of the victim and paralyze it, possibly through the action of some noxious fluid. Forms much larger than the hunters are frequently brought down in this way, to be swallowed either whole or piecemeal. The attack is not necessarily fatal, for the larger forms frequently revive. 1 The position of the trichocysts is primarily in the cortical plasm, but they are rarely entirely immersed, being much more frequently suspended in the streaming endoplasm. They are occasionally drawn 1 See ante, p. 50. 176 THE PROTOZOA out from the cortical plasm and are then carried about in the stream. In addition to the trichocysts, some of the Ciliata carry still more effective weapons in the form of nematocysts. In Vorticclla umbel- laria (Clap. & Lach.), Engelmann described from twelve to twenty pairs of capsules, each of which contained a coiled thread which, as in the Coelenterata, could be thrown out upon irritation. The cortical plasm may be considered the inner portion of the ectoplasm, the outer portion of which forms the covering of the animal, the membrane, cuticle, or pellicle. The latter is extremely variable in thickness and in complexity. It is apparently homolo- gous with the ex- ternal membrane of ordinary animal cells, and, accord- ing to Biitschli, is formed by conden- sation of the pro- toplasmic ground substance, and, as Stein ('59) first maintained, is in no sense a secre- tion. 1 Biitschli ('88), Schuberg ('87), and many others regarded it as the agglutina- tion of the outer c Fig. 93. Coleps hirtus Ehr. [MAUPAS.] A. Side. B. One of the is rows of plates composing the test. , . . c. Division-phase. thickened lamellae of the external alveoli into a continuous membrane. In forms where the cortical plasm is absent, as in Hypotrichida, the cuticle, or, as Biitschli prefers to call it, the pellicle, forms a thin coating to the cell and lies directly upon the endoplasm. In many cases the outer protoplasm either becomes changed into, or else secretes, an external casing or house, which may be either loose or tight-fitting. This covering may be of jelly (e.g. Ophrydium\ or of chitin (e.g. Folli- culina), or of a horny product without any mineral elements (e.g. Colef>s\ In Coleps hirtus (Ehr.) the horn-like covering is tight-fitting, and composed of separate pieces, which form four girdles about the body (Fig. 93). Each girdle is composed of separate pieces, each of which is straight on one edge and serrated upon the other in such a 1 Cf. Stein ('59), p. 56. THE INFUSORIA 177 manner that, when they are put together; the serrations slightly over- lap the straight edge of the next adjacent piece, thus leaving open- ings for the protrusion of the cilia. The lower end is covered by separate pieces, which open centrally for excretion. On the anterior end, the mouth, crowned by a ring of large cilia, is protected by a set of plates with teeth-like projections, which act somewhat like the similarly arranged teeth of a sea-urchin. In a great many cases the outer body wall is marked by striations of various kinds showing the lines of insertion of the cilia. A typical example is shown in the membrane of Holophrya, one of the Holotrichida(Fig. 91, A, and 97, g}. Here the cilia are inserted in regular lines which run from the anterior to the posterior end. In other cases, as in Lem- badion (Holotrichida), the cilia are inserted on minute papillae, which lie in rows upon the cuticle with more or less distinct furrows be- tween them, thus forming secondary, but very distinct, markings in addition to the primary lines formed by the insertion points of the cilia. That the striation is due to the ciliation can be easily seen in cases where the cilia are absent from one portion of the body and pres- ent in others, as in many of the Holotrichida. Fig.$>4. Lacrymana coronata The rows are not always straight, as in Holo- cu anc ? , Lach ' [ BUTSC LI : 1 ' 3 ' m, spirally wound rows of cilia. p/uya, but are variously changed through the alteration of the axial relations. The most frequent variation from the primitive condition is the spiral arrange- ment (e.g. Lacrymaria coronata, Fig. 94), where the course of the cilia has become changed by the alteration in the position of the mouth. A very curious type of striation is seen in DasytricJia ruminantum (Schuberg, '88) (Holotrichida), where the striations do not converge at the mouth as they do in the majority of forms, but in a line above it. The exception is significant, however, as showing the line of the shifting of the mouth, the path being marked by the meeting points of the converging striae (Fig. 95). The external markings were early recognized by Ehrenberg, who interpreted them as the insertion points of the cilia as described above. Stein, however, held that the markings are invariably due to the pres- ence of myonemes which form the insertion base of the cilia. Both observers were right in part, for striations in some of the Heterotri- chida and Peritrichida are due to the presence of myonemes, but in the Holotrichida, where, with one or two exceptions (Holophrya, Prorodon\ myonemes do not occur, the markings are unmistakably due to the cilia. THE PROTOZOA The myonemes are ectoplasmic differentiations which are contrac- tile in nature and are formed, according to Biitschli and Schewiakoff, from the walls of the alveoli which make up the sub-cuticular layer of the membrane. Although probably arising in the peripheral alveolar region, these threads occasionally become separated from this position and are then found in the cortical plasm or even in the endoplasm. Myonemes are most highly differentiated and are best known in the Vorticellidae, where the sudden contraction of the bell, or the instan- taneous rolling-up of the stalk, are due to their action. While the most conspicuous myonemes in Vorticclla run from the centre of the disk to the very base of the stalk, Entz ('91) has described additional fibrils which have a similar but less important function. According A B C D Fig. 95. Supposed change of position of the mouth in Ciliata. [BUTSCHLI.J A. Original position (as in Holophrya), B. The mouth has become elongated (as in Enchelys or Spa.thid.ium). C. Similar stage from the ventral side. D. The mouth has become closed be- hind, leaving the opening away from the body extremity. The markings on the membrane now meet in the line represented by the original mouth-slit (e.g. in Glaucoma). to this observer there are two sets of myonemes, one internal, the other external. Each set includes two groups of myonemes, one cir- cular in its course, the other longitudinal. The external layer, observed by Lachmann ('56) and Stein ('59, '67) but denied by many, is formed of a large, single fibre composed of fibrillae, which winds spirally about the bell from the junction of the peduncle to the centre of the disk. It is this myoneme, Entz maintains, which gives the annulate appearance to the bell, and, like a muscle-fibre, it is charac- terized by fine transverse striations. A second circular set is formed by another single fibre, which, however, is confined to the peristome disk, and is located deep in the ectoplasm (Fig. 91, D}. This fibre takes only a few spiral turns at the base of the elevated disk and around the edge of the collar, and functions as a sphincter-muscle to close over the disk. Two sets of longitudinal myonemes complete the muscular system. Of these, the external set, lying between the two circular myonemes, consists of fine fibres running from the peduncle to the THE INFUSORIA 179 disk where their course is radial. The largest and most important of all of the myonemes are those forming the fourth set. These are longitudinal muscle-fibres of considerable thickness running from the centre of the disk radially toward the periphery, then continuing down the sides of the bell as far as the ciliary girdle ( Wimperring), where they leave the wall of the body and come together to form the thick muscle-strand of the stalk. The latter highly contractile organ consists of a wall and of the central, contractile strands which are bathed with a fluid contained within the walls of the stalk. The wall itself, according to Entz, but contrary to Biatschli, is a continuation of the living wall of the bell, in which membrane and underlying mus- A Fig. 96. ?,oothamnium arbuscula Ehr. [ENTZ.] A. Lower portion of main trunk. B. One ot the branches of the main trunk, a, axoneme ; /, spasmoneme ; s, spironeme. cular structures can be distinguished, as in the main portion of the body. Biitschli, on a less substantial basis, described the stalk as a secretion similar to the stalks of the Mastigophora and Sarcodina, and chitinous in composition. The main strand within the stalk is formed by the collection of the strands of the inner longitudinal myonemes, and is covered by a delicate sheath which separates it from the fluid or gelatinous matter surrounding it. Biitschli regards this sheath as a continuous coat from the alveolar layer of the bell. The strand has three threads which Entz calls spasmoneme, spironeme, and axoneme (Fig. 96). The fibres of the first run to the base of the stalk. The other two are closely connected, and both are made up of microsomes, which Entz described as nucleus-like granules (karyophans} surrounded by an ovoid matrix (cytopkan}. These granules, so conspicuous in the stalks of Vorticella, evidently correspond to the Elementar-Granula (Greeff, '71) or cyto-microsomes. Entz figured them as arranged in i8o THE PROTOZOA rows like a string of beads. Without going deeply into the subject, which is far from settled, it will suffice here to state that two views are now held as to the seat of contractility in the stalks of Vorticella. One set of observers hold that the outer membrane of the stalk is the contractile portion, and that the contained thread merely counteracts the force of the membrane, which tends to contract and roll up the stalk. In other words, the myonemes of the spasmoneme are regarded as elastic and not as contractile fibrils, at rest when the stalk is coiled, active when the bell is extended (Cohn, '62 ; Metschmkoff , '63 ; Rouget, d a Fig. 97. Myonemes and cilia. [METSCHNIKOFF, BiJTSCHLl, and JOHNSON.] a, b, d, e. Cuticle and myonemes of Stentor cceruleus. d. More highly magnified piece of myoneme. g. Optical section through the body wall of Holophrya discolor. '61 ; Schaaffhausen, '68; Entz, '91). Entz described the membrane of the stalk and the spasmoneme as antagonistic elements. The former, which stretches out while at rest and contracts when irritated, opposes the latter, which acts in the reversed manner. The axoneme he re- garded as a sort of nerve-centre. The opponents of this view hold that the rolling of the stalk is accomplished by the contraction of the muscle-like spasmoneme (Stein '67, Clap. & Lachmann '58, Engelmann '76, Butschli '88, etc.). The myonemes lie in minute canals according to Butschli ('88) and Schewiakoff ('89) ; in direct contact with the plasm according to John- son ('93) and Entz ('91), and they probably vary in position in differ- ent forms. The structure and position of a myoneme in the ectoplasm can be more easily seen from the accompanying figure than from a description (Fig. 97). THE INFUSORIA l8l Other contractile elements are occasionally found : the most remarkable, perhaps, is the peculiar muscular band which surrounds the peristome of Bursaria trnncatella. This highly differentiated muscular organ, which functions as a sphincter, is, like the myonemes, derived from the alveolar layer immediately below the pellicle. The ectoplasm appears throughout to be the seat of motion. Not only are the contractile myonemes differentiations of this important layer, but the cilia and all of their modifications are likewise derived from it. The cilia themselves appear to be mere prolongations of the alveolar layer. They are minute, probably of similar diameter throughout, and except for regional differentiation in the vicinity of the mouth, are of uniform length. As a rule, they are inserted upon minute elevations or papillae on the cuticle and appear to be connected by minute fibrils with the myonemes. The finer structure of the cilia has not been satisfactorily made out, but the present results tend to the view that they are simple, firm threads without differentiations. Unlike flagella, they act in unison, and their motion is that of a paddle rather than a lash, as in flagella. Jensen ('93) has figured the absolute lifting power of the ciliary apparatus of Paramcecium at 0.00158 milli- grammes, or nine times the weight of the animal. The cilia are grouped together in various ways, forming more or less complex motile organs. These are rarely seen in Holotrichida, but in the other orders they may be pointed aggregates (cirri), plate-like vibratile organs (mem- branelles), or broad, undulating membranes. All of these modifica- tions are found in the Hypotrichida, where the motile apparatus is especially characteristic. The arched dorsal surface is without cilia, but occasionally holds a varying number of bristles which have, possibly, a sensory function. On the flat, ventral side of the most primitive forms of this order, the cilia are very generally distributed (Peritromus, Fig. 1 1 3, J5), but in the more differentiated forms they are reduced in number, and modified into cirri, membranes, and mem- branelles. In many forms they may be entirely absent, the only motile apparatus being the membranelles on the ventral side about the mouth. These form the adoral zone, which stretches from the mouth forward on the left side of the peristome, and as far as the dorsal anterior region. In some cases a row of cilia stretches along the floor of the peristome parallel with the membranelles, a single cilium opposite each mem- branelle. The right border of the peristome (Fig. 98) is continued into a vibratile membrane, and close to the left of this and running parallel with it is another row of cilia (praeoral cilia poc\ In the centre of the peristome is a second undulating membrane, the endoral membrane (em}, which passes downward and into the pharynx, and this, also, is sometimes accompanied by a row of cilia even into the pharynx (endoral cilia eo). 182 THE PROTOZOA Cirri, membranelles, and membranes are each striated, and when treated with certain reagents (e.g. gold chloride, Maupas), can be reduced to separate fibres which are similar to cilia. The simplest of these aggregations are the cirri. These bundles of threads are usually pointed, and either curved or straight, forming the Griffeln und Hakcn of Ehrenberg. A simple condition is seen in the tail-like process of Urocentrum, which has a distinctly fibrillar structure and can be readily reduced to a brush of very fine hairs (Fig. 99). Although the striated appear- ance and the reduction to the component fibrils make it prob- able that cirri arise by the fusion of cilia, an objection is met in the fact that their de- ^__ -n MA \ * velopment, after division, shows \-~em no such origin. On the con- }_ o trary, they arise from the ecto- plasm as cirri and not as cilia. This objection, however, seems 2 hardly sufficient to counter- \'1)0 C balance the evidence in favor of the concrescence theory, evidence which is strengthened by the position of the cirri along the lines of the cilia- markings. The membranelles are flat plates of striated appearance usually in the form of tri- angles, squares, or parallelo- grams. Each membranelle is inserted in a furrow below Fig. 98. Schematic hypotrichous ciliate. az.adoral zone; c, ventral cirri ; <>z, endoral mem- which IS a basal Stripe of brane; eo, endoral cilia; pm, praoral membrane ; po, thickened protoplasm COntinU- paroral cilia ; /, praeoral alia. ^ with ^ longitudinal dl _ iary markings (Heterotrichida). Like the cirri, they can be readily reduced to component fila- ments resembling cilia, and there is, therefore, every reason to suppose that the membranelles which form such a characteristic differential for all orders save the Holotrichida, are merely the differ- entiated portions of the ciliary rows. 1 The basal stripes of the membranelles, which are spirally arranged upon the peristome, are in turn inserted, in some cases at least, in a thick fibrous strand which 1 Johnson ('93) alone regards the membranelles in Stentor as endoplasmic in origin. THE INFUSORIA 183 runs around the peristome connecting the series, and possibly form- ing a nervous organ (Delage, '96; Moore, '93). The undulating membranes, finally, which are almost always con- fined to the oral region, and like the membranelles chiefly concerned with food-taking, have probably a similar origin, although the con- nection with the cilia is less apparent. They are frequently, as in the Hypotrichida, placed deep in the vestibule, but in many forms they are confined to the pharynx itself, as in many of the Holo- trichida. In addition to cilia, membranelles, and membranes, the ectoplasm has other modifications, such as pseudopodia (e.g. Stentor} and tenta- cles. The pseudopodia are used for anchoring the animal, and are produced at the posterior end by the so-called foot-disk (Johnson, '93). The cortical plasm gives rise to these processes, and also to the peculiar tentacle-like appendages found in some forms. In Actinobo- lus (Fig. 100) these pseudopodial tentacles are particularly well known through the complete study made by Entz ('82). Here the threads pass out between the cilia and not infrequently reach a length of twice or even three times the !__ TI j.1 i r i ' body diameter. The threads are of nearly uni- form thickness, with blunt or slightly knobbed ends (Entz). These tentacles, while occasion- ally stiff and unyielding, can be shortened or lengthened, or drawn into the body in a manner surprisingly suggestive of pseudo- . ' ... . r . Fig. 99. Urocentrum turbo podia, while the protective and offensive func- o. F. M. [BUTSCHLI.] tion is shown by the presence of trichocysts at their extremities. Similar tentacles are found in Mesodinium and Ileonema (see Fig. 115). While the ectoplasm is devoted to the functions of motion and irri- tability, the endoplasm is charged with digestion and reproduction. Thus the membranelles and membranes are important in creating the current which brings the food particles ; the trichocysts are occasion- ally developed as food-killing organs, and these, with the mouth, vestibule, and pharynx, are ectoplasmic in origin. While all these special modifications are developed for the pur- pose of food-getting, the endoplasm, with its digestive processes, shows but little advance, so far as can be made out, over the already complicated endoplasm in the less highly organized forms. Simi- lar food substances are treated in similar gastric vacuoles, and the products of assimilation are carried about in the plasm by similar cyclosis, while indigestible remains are excreted in the same way. The Protozoa thus offer in the most striking manner an example of 1 84 THE PROTOZOA how species may have originated through structural adaptations of the parts (ectoplasm) that are in direct contact with the environment. The mouth parts, which are functionally the beginnings of the diges- tive system, are formed by the invagination of a limited portion of Fig. 100. Actinobolus radians St. the ectoplasm in a manner analogous to the formation of the stomo- daeum of Metazoa. The peristome is the beginning of the mouth depression, becoming more and more deeply depressed as the mouth region is reached. It is not present in all forms, the mouth in its original position probably being anterior and terminal in the monaxonic THE INFUSORIA 185 body, and leading by a mere passage into the endoplasm below. Mouthless forms are known, but these have degenerated through parasitism and are not primitive (Opalinidae). In the majority of forms the mouth is displaced from the original terminal position and has become ventral and central. Biitschli maintains that this change in the position of the mouth is brought about by the gradual shifting from the anterior end, as shown by the meeting point of the lines of ciliary markings. As previously indicated, the original course of these lines is from the anterior to the posterior end, but in numerous transitional forms in which the mouth has a more or less ventral position, the markings become curved to agree with the changed position, while the course which the mouth is supposed to have taken is shown by the converging lines (Fig. 95). In almost all cases the mouth is not in direct communication with the endoplasm, but is separated from the latter by a longer or shorter pharynx, oesophagus, or gullet, which frequently bears cilia, mem- branes, or membranelles. The oesophagus is likewise an ectoplasmic invagination, as is also a second oesophageal apparatus, found in some forms (Vorticellidae), where the mouth leads into a comparatively large ciliated or membraned space, known as the vestibule (Fig. 101, C, ./?), and this leads into the oesophagus or gullet proper, which, in turn, communicates with the endoplasm. This space begins as a wide tube and gradually narrows down to a more or less narrow aperture or constriction at the oesophagus. The anus and the contractile vacuole, in some forms, open to the exterior through the vestibule. In some of the Holotrichida, the region about the pharynx is strengthened by accessory apparatus developed in the cortical layer, which in this region is greatly thickened and which in some cases contains secretions in the form of bars arranged in a peculiar basket structure (A, B}. The membranelles which surround the mouth are usually in motion, as are the membranes and cilia which extend into the vestibule or oesophagus. Even while the animal is lying quiet, the membra- nelles continue their active vibrations, keeping a constant current of water toward the mouth. This current brings a supply of bacteria, diatoms, algae of various kinds, rotifers of small size, or parts of animals undergoing disintegration, flagellates, etc. A distinction can be made here between herbivorous and carnivorous forms, although the differences can hardly extend to structural adaptations, unless it be, perhaps, in some carnivorous forms, where special weapons of offence (the trichocysts) are found. Probably all forms are more or less omnivorous and make little or no selection of food. The food particles are thrown by the current of the membranelles into the peristomial depression and thence into the vestibule or 1 86 THE PROTOZOA oesophagus, until they come in contact with the endoplasm at the base of the latter. Here they are readily absorbed by the endoplasm, in which, together with a small amount of water, they are confined in a small gastric vacuole. The vacuole enlarges by the constant addition of new material, until it is caught up in the current of the endoplasm and dragged away. In this improvised " stomach " it is slowly digested, a new drop being formed in the meantime at the mouth opening. If food is abundant, the animal may become filled with these gastric vacuoles. The liquid of the vacuole is, at first, simply water, like the surrounding medium, but gradually becomes acid through osmosis in the plasm, and the digestible substances are D Fig. 101. Buccal apparatus. [BUTSCHLl.] A, B. Nassula aurea Ehr. A. From the ventral side. B. The buccal apparatus strongly magnified. C. Urostyla grandis Ehr. D. Vorticella nebulifera O. F. M. slowly dissolved, the residue being cast to the exterior through the anus. Unlike the mouth, the anus, as a rule, is a simple opening in the outer wall (Maupas, '83 ; Butschli, '88), although Stein ('67) described an anal tube in certain forms (Nyctotherus). In the Heterotrichida it is sub-terminal in position. In the Hypotrichida it is never ter- minal, but usually dorsal, and toward the left edge. In the Peritri- chida, especially in the Vorticellidae, the anus opens into the vestibule. B. CONTRACTILE VACUOLES A certain amount of water is taken in with the food through the mouth, and at the same time (as in those cases where the mouth is THE INFUSORIA I8 7 absent) by absorption through the body wall, and it is the function of the contractile vacuole to get rid of the surplus. This organ is variously complicated by the development of a more or less extensive series of canals, which empty in a common excretory vacuole. Always situated in the cortical plasm, the con- tractile vacuoles are fixed in position and com- municate with the exterior at systole by a permanent aperture, which, however, becomes covered internally during filling or diastole. They vary in number from one to a hundred, or even more, and are absent, apparently, in only one form (Ofalina), although Vejdovsky ('92) describes contractile vacuoles in a closely allied form, Monodontophrya longissima, while even in Opalina the reminiscence of the vacu- ole is seen in the remnants of the feeding canals (Delage, '96). In its simplest form the vacuole is single and terminal, a condition which may be found in each of the four orders. When there are more than one, they are grouped around the original vacuole in a ter- minal position, or arranged along one or more lines upon the dorsal side. In Discophrya and //tf//z/0//^77#(Holotrichida) there is no regular vesicle, but a long contracting canal which runs the length of the body. Spirostomum (Heterotrichida) has a terminal vesicle, with one long feeding canal, and from this the canal system is developed in a variety of ways. Thus there is a vesicle with two feeding canals in Climacostomum (Fig. 91, B), one terminal vesicle and four feeding canals in Urocentrnm. In Stentor there is a single vesicle near the peristome, with two feeding canals, one of which runs to the end of the body, while the ole () emptying through a ,, i i i T> i l n l canal into the vestibule ; other runs around the peristome edge. Fabre- Oi the oesophagus ; , the nu- Dumergue ('89) holds that canals, for the most cieus. part invisible, are present in all ciliates. This is certainly true in Frontonia, where there are one or two vesicles on one side and an immense number of feeding canals, which anasto- mose and branch to form a complicated network, involving the entire body. In some forms the vesicle communicates with the exterior directly, but it may be complicated by the formation of ducts or reservoirs. In the holotrichous form, Lembadion, the vesicle lies Fig. 102. Anterior end ot Ehn c , the reservoir of the vacu- 1 88 THE PROTOZOA dorsally in the middle of the body, but is connected with the terminal aperture by a long canal. In the Peritrichida there are one or two vesicles, which empty by contraction into special reservoirs, and these, in turn, empty into the oesophagus (Fig. 102). According to Delage ('96) the contraction of the vesicle is brought about by the contractility of the surrounding cortical plasm. Rhum- bler ('98) has shown, however, that the contained water may so affect the plasm that it becomes differentiated like ectoplasm, and this gives some substantial basis for the view that a special membrane surrounds the vacuole. There is no evidence, however, to show that this modi- fied protoplasm acts like a sphincter. Biatschli holds that the vesicle contracts through a mechanical force exerted upon the thin plasmic layer between the vesicle and the opening of the excretory pore by the pressure of the filling vacuole and the turgor of the cell. At the completion of the diastole the pressure becomes too great for the lamella, and the latter is ruptured, allowing the contained fluid to pass to the exterior. C. THE NUCLEUS The nuclei of the Infusoria show some of the most striking structural characteristics connected with the Protozoa. Here there is a differentiation of the nuclear material into two forms, a larger macronucleus, and a very much smaller micronucleus. With the single exception of Polykrikos (Dinoflagellidia), this differentiation of the nuclei is found nowhere outside of the present group. The functions of the two kinds of nuclei are supposed to be respectively vegetative and reproductive (Butschli), but this distinction is, perhaps, too sweeping. Julin ('93) held that the macronucleus stands not only for nutrition, movement, sensation, and regeneration, but for asexual division as well, in fact is a " somatic nucleus," while the micronucleus functions only as a sexual nucleus. There may be one or many of each kind in each cell. The macronucleus, which is invariably present, recalls the nucleus of tissue cells. It is usually single, and, lying in the endoplasm, it may be carried about with the flowing granules, or maintained in a permanent position in the cortical plasm, or by processes from this plasm (e.g. Isotricha). Its form is quite variable and has little significance for systematic work, for in the same species under certain conditions it may even become amoeboid (Loeb & Hardesty, '95). The usual form is spherical, but it may be elongated into an oval, or into a flattened rod which may be curved or straight, or it may be divided into small pieces resembling a string of beads, connected by a membrane (Fig. 103). It is always provided with a membrane (Maupas), but the chromatin contained within it is vari- THE INFUSORIA 189 ously distributed. The vesicular structure, in which the nuclear substance is so distributed as to leave more or less space filled with "nuclear sap," is almost never seen, the macronucleus appearing solid and completely filled with chromatin. Biitschli described the finer structure as almost invariably alveolar, the meshes correspond- ing to those of the surrounding plasm. The entire network stains deeply with the nuclear dyes, but at certain stages, especially during division, distinct fine lines can be made out connecting the D JVL F Fig. 103. Types of macronuclei. [SAVILLE KENT.] A. Macro- and micronucleus of Loxodes rostrum O. F. M. B. Of Nyctotherus cordiformis Leidy. C. Macronucleus of Plagiotoma lumbrici Duj. D. Dendrosoma radians S. K., a young nucleus. E. Dendrosoma radians S. K. F. Stentor polymorphus Ehr. G. Stylonychia mytilus O. F. M. H. The same in division. N, the macronucleus ; n, the micronucleus. chromatin granules, and corresponding to the linin reticulum of most nuclei. In some cases (e.g. Loxophylhim} a permanent spireme is present, as in the nucleus of cells frdm the salivary gland of Chironomus larvae, and is transversely striated, indicating disks which Balbiani ('90) thought are alternately chromatin and linin (Fig. 104). In many cases there are internal modifications of the nuclear material forming so-called "nucleoli," although there is a possibility that these structures are similar to the intranuclear bodies found in Mastigophora. In many macronuclei a peculiar division of the organ is made by a igo THE PRO7OZOA split, which the Germans call a Kernspalt and the French a fente. While this peculiar feature of the nucleus has not been explained, what may be an important light has been thrown upon it by Bal- biani ('95), who described the appearance as due to the presence of two materials within the nuclei, one of which is chromatin, the other, achromatic material or archoplasm. This interpretation, however, cannot be accepted as final. While the macronuclei are, as a rule, single in number, the micro- nuclei are often multiple. Probably all Ciliata have at -least one micro- nucleus, although the small size and the extreme difficulty in staining sometimes render it hard to find. In one case at least (Opalina rana- rum) there is only one kind of nu- cleus. The number of micronuclei is usually greater where the macro- nucleus is elongate, and especially where it is beaded (Stcntor, Spiro- stomum, etc.). As a rule the micro- nuclei are closely attached to the membrane of the macronucleus, occupying a minute indentation in the latter, but in some cases they are well separated. In form they are round, ellipsoidal, or spindle- shaped, but the form varies with the nuclear activity, and does not mean much in itself. Their longest axis measures from i /u to 10 /x, and like the macronuclei, they are covered with a distinct membrane, while the chromatin is usually massed at . LoxophyllummeleagrisQ>.f.^A. Some part of the nucleus. In Cer- D Fig. [BALBIANI.] tain cases the appearance is like A. Vegetative nuclei with chromatin in the ., c 4-u i vu *u form of a permanent spireme. B.C,D. The that f the macronudeUS With the same in division. chromatin in the form of a densely packed reticulum, giving to it a mas- sive appearance. Here two distinct portions, the chromatin and achromatin, can be made out (Fig. 105). Division of the nuclei takes place by mitosis in the micronuclei, and, THE INFUSORIA IQI as a rule, by amitosis in the macronuclei. The latter is the simpler ; in many spherical or elliptical nuclei the structure merely draws out and segments into two equal parts. It is more or less complicated, however, in different macronuclei, until well-developed mitosis replaces simple division {e.g. SpirocJiond}. There is reason to regard the sim- ple division of the larger nuclei as the mere degeneration of mitosis, by a process in which the various stages have gradually disappeared until only the preliminary stages of such division are to be found. These preliminary stages are seen in the transformation of the reticu- lum of chromatin into thread-like masses, which recall the spireme of D Fig. 105. Mitotic division of the nuclei of Spirochona and Paramcecium. [R. HERTWIG.] A-C. Macronucleus of Spirochona with well-developed pole plates. D-H. Different stages in division of the micronucleus of Paramascium. higher cells ; the threads are then divided across into equal parts. In Spirochona, however, the process of division is strikingly similar to mitosis in Metazoa (Fig. 105, B, C}. Division of the micronuclei is accompanied by the formation of polar attraction-spheres, and by the rearrangement of chromatin into chromosomes. Before division, the micronucleus swells to nearly twice its size during resting stages (Hertwig, '77), while the granu- lar chromatin begins to collect in lines at first equally thick, but later concentrated in the equatorial region (Fig. 105, D-H}. Divi- sion then takes place through the centre. I 92 THE PROTOZOA D. ENCYSTMENT The phenomenon of encystment may be seen in the Ciliata as in all other groups of the Protozoa. It occurs when the animal is in danger of drying, in some cases before division, in others, for the purpose of digesting a full meal. The cilia are drawn in, the mouth and peri- stome disappear, the contained body-granules are voided, and a gelati- nous secretion is poured out from the ectoplasm. The secretion soon hardens, becoming chitinous. The vacuole continues to pulsate for some time, and the secretion forms a liquid layer about the animal under the cyst. The cysts are variously diversified with spines and processes of different kinds, and are occasionally multiple, the spaces between the cysts being filled with water (Fig. 17, B, C, F, p. 47). E. REPRODUCTION Reproduction among the Ciliata takes place almost exclusively by simple division or fission. It is practically the same for all forms, the variations being of minor importance. The nuclei first divide, new mouth parts are developed in the posterior half, and then the cell divides. The first indication of division in Stentor, for example, is a rift in one side of the animal below the adoral zone. This rift rapidly develops motile organs (membranelles), and acquires the full length of the lower daughter-individual. The nuclei in the meantime divide, and the original animal draws out, leaving a slender foot for the upper or anterior cell, and a swollen portion for the pharyngeal region of the second individual. The new adoral zone is completely formed before actual division; the steps in the process are shown in the accompany- ing diagram from Johnson ('93) (Fig. 106). The new contractile vacu- ole, according to him, arises de novo in .S. cceruleus, and by a dilatation of the longitudinal canal in .S". rceselii. The new vacuole thus formed remains in connection with the longitudinal canal, the upper part of which becomes drawn out with the torsion of the adoral zone to form the much-discussed ring-canal discovered by Lachmann. In some forms, as in Spirochona, division simulates budding, un- equal division giving rise to mother- and daughter-cells. When division takes place within the cyst, the various mouth parts may or may not first be absorbed, but in all cases the vacuole still continues to pulsate. Here, as a rule, division is double, resulting in broods of four which escape as embryos, and gradually grow into the parent form. This condition closely simulates spore-formation, which results when, as in Ofialina, the number of divisions within the cyst reaches three, four, five, or six. Nowhere among the Protozoa has the process of conjugation been so thoroughly studied in connection with the life-history of the organ- THE INFUSORIA 193 ism as in the Ciliata. Worked out first by Biitschli and Engelmann in 1876, it has since been carefully studied by numerous observers, and the conditions preliminary to conjugation, during, and subsequent to it have been made known in a great variety of forms representing all divisions of the class. Biitschli and Engelmann early recognized that conjugation is necessary for continued life activity of the organ- ism, and came to the conclusion, which has been fully confirmed by subsequent observers, that conjugation is a process of rejuvenation or a renewal of vitality, the need of which is shown by the reduced size Fig. 106. Diagrams to illustrate the division of Stentor rceselh. [JOHNSON.] y, the vacuole ; A", the ring canal. and general degenerate condition of the organism prior to conjuga- tion. Maupas ('89), in his classical work on conjugation among Infu- soria, found that the number of generations which may be formed from one conjugating period to the next varies with the species, but is usually between three hundred and four hundred and fifty. He found, furthermore, that certain conditions are necessary for conjugation. These conditions are : (i) maturity of the organisms, i.e. forms which have just conjugated will not again conjugate until after a certain number of generations ; (2) partial lack of food, i.e. if plenty of food is present, conjugation will not take place even though the individuals are well along in degeneration ; (3) diverse ancestry, i.e. the conju- gants must come from different ancestral conjugants. 194 THE PROTOZOA The two conjugants fuse either temporarily or permanently, and the external structures, such as the membranelles, are absorbed. If the fusion is temporary, as in the majority of forms, the two ecto- plasms fuse at or near the mouth parts, and a protoplasmic bridge is formed between the two organisms. The two organisms then become sluggish, and rest for a considerable time upon the bottom without movement of any kind. They ultimately separate and begin to divide. The micronuclei play the most important part in conjugation. Each divides two or more times to form four or more daughter-nuclei, some of which de- generate, while one divides again, one half to fuse with a similarly derived nu- cleus of the other organism, while the other half remains as the receptive nu- cleus, or the female pronnclcns. The two conjugating nu- clei cannot be distin- guished from those which degenerate, but are apparently only those which lie nearest the bridge joining the two organisms. Mean- while the macro- nucleus undergoes complete degenera- tion, breaking up into a number of pieces, which are gradually absorbed by the proto- plasm. The new macronucleus is formed by the enlargement of a daughter-micronucleus derived from the fusion nucleus. Hoyer('99), however, asserts that in Colpidinm colpoda it forms by the union of two daughter-nuclei. When there are two or more micronuclei in each conjugant, the process is repeated for each of them, although it is not known whether this holds when, as in Stcntor, the number reaches sixty or seventy. In a few cases (Vorticellidae) the conjugants are of diverse size Fig. 107. Conjugation in Epistylis umbellaria Greeff. [GREEFF.] M, macrogamele ; m, microgametes. THE INFUSORIA 195 The larger form or macrogamete is usually a normal-sized individual, although in some cases it is somewhat larger than the ordinary cells (Zoothammum). The microgametes, on the other hand, are con- siderably smaller, and from four to eight are formed by each cell. These never develop a stalk, but leave the parent colony, and swim about by means of the ring of cilia around the lower pole. They finally come in contact with the macrogamete and fuse with it, the union taking place at the lower end of the attached organism, and near the insertion point of the stalk (Fig. 107). II. THE SUCTORIA The Suctoria differ decidedly from the Ciliata, from which they have undoubtedly sprung. With the exception of Hypocoma (Fig. 115, C ), which remains ciliated throughout life, the Suctoria possess cilia only during the embryonic stages. They are, for the most part, sedentary forms, and grow upon a chitinous peduncle, which is at- tached at the lower end to some foreign object. The upper end of the peduncle is hollowed out into a bowl, within which the animal lies. Owing to its attached mode of life, and to the equal pressure on all sides, the general form of the animal is spherical or radially symmetrical. In some cases there is a well-defined membrane, but the various students of the group are not agreed as to its structure. It is never striated, as in the Ciliata, and there is no cortical plasm. The endoplasm shows no differentiations other than the usual food granules or assimilation products common to all Protozoa. An essential point of difference from the ciliate structure is the presence of tentacles, which, in the majority of Suctoria, are the only motile appendages of the adult. In many respects they are similar to the tentacles of Actinobolus, Ilconema, and Mesodinium (Fig. 115), but differ in the very important fact that they are hollow, while the extremities bear the mouth openings. There are two general types of tentacles : one, according to Bu't- schli, captures prey, while the other devours it. Of the latter forms, there are also two types. One is long and broad, and, like a thorn, pointed at the extremity ; the other is nearly uniform in diameter and flattened at the top, or hollowed out into a cup-like sucking organ (Fig. 1 08). These are distinguished as the styliform and capitate tentacles (Delage). Both sets of tentacles are hollow, and their lumena open at the ends. There is a difference of opinion, however, in regard to their inner structure and function. Biitschli holds that some of them are solid, and others hollow. He maintains that, in the solid forms, the internal portion is formed of endoplasm, which is continuous with the inner plasm of the cell. Delage claims that they 196 are all hollow. The function of the endoplasm, according to Biitschli, differs in the styliform and the capitate tentacles. In the latter, the prey is retained by the sucking disk at the extremity while the endo- plasm within the tube meantime works up and down like a pump- piston, and a vacuum being thus formed, the cuticle of the prey is burst, and the fluid endoplasm flows down the tentacle canal to the endoplasm of the captor, where it is digested. Such an explanation of the action of these tentacles is regarded by most observers as extremely doubtful. Delage finds no motion in the endoplasm during feeding, save in the rhythmic pulsations of the contractile vacuole, an organoid which Eismond ('90) believed is the cause of the suction. The excretion of water from the vacuole, he argued, creates a semi-vacuum in the protoplasm, and the pressure Fig. 108. Tentacles of Suctoria. [R. HERTWIG.] A. Different types of styliform or piercing tentacles. B. Capitate and piercing tentacles. from without forces food or loose particles, etc., through the tentacle openings to the endoplasm. This explanation, although somewhat fanciful, is certainly as plausible as the principle of the pump, but the matter must remain for the present as one of the many unsolved problems connected with this group. In some cases (Trichophrya angulata) the tentacles are apparently unnecessary for food-taking, as Dangeard ('90) found that particles are occasionally engulfed, as in the Rhizopoda, at any point of the naked body. In the styliform tentacles, on the other hand, the sharp points pierce the membrane of the prey, while the endoplasm contained within the tentacle possibly flows into the prey, whose endoplasm is digested in situ. In some cases they appear to have a paralyzing effect upon other forms, and ciliates coming in contact with them have been seen to stop their movements as though stunned. THE INFUSORIA 197 In all cases the tentacles are remarkably like pseudopodia, and may change their form and their position, and may even be entirely with- drawn into the body, to reappear, possibly, at some other place. Some forms have the power of withdrawing their tentacles and developing cilia, which may be retained for a longer or shorter Fig. 109. Dendrosoma radians Ehr. [SAVILLE KENT.] n, nucleus. period. In some cases the tentacles distinctly originate in the endo- plasm, and penetrate the membrane (Hertwig, '76; Ishikawa, '96). The nuclei, like those of the Ciliata, are of two kinds, macro- and micronuclei. The former are little different from the macronuclei of the more generalized Ciliata, while very little is known about the latter. In the colony-form Dendrosoma, where the many branches suggest a hydroid colony, the macronucleus extends through all the 198 THE PROTOZOA branches and trunks, penetrating the entire system, like the coenosarc of a hydroid (Fig. 109). The contractile vacuole never becomes so complicated as in some Ciliata, but consists, usually, of a single vesicle, which may be surrounded by a circle of small vacuoles emptying into it. In some cases there is a short excretory duct leading from the vesicle to the excretory pore in the membrane, which, as in the Ciliata, is a per- manent opening. These animals en- cyst only for protec- tion, never, apparently, for reproduction. As in the Ciliata, the pro- cess consists of the secretion of a chitinous mantle about the cell, the tentacles being withdrawn into the body. Reproduction is almost invariably by simple division, which may be either equal or partial (budding). In the simplest cases the upper portion of the cell is constricted off, and moves away from the lower portion, which remains upon its stalk (/W0/>/rrj'rt, Spir- ophtya, Urnu/a, etc.). Fig. no. Exogenous budding in Ephelota BtitsMiatta Tshi. The detached part de- N, nucleus. velops cilia and, after a longer or shorter free- swimming period, settles down, loses its cilia, and secretes a stalk. Partial division, or budding, may be either endogenous or exogenous. The latter is the simpler; an individual prepares as for division, but instead of dividing into two equal portions, a number of papillae appear at the outer surface, each becomes a bud, receiving a portion of the nucleus (Fig. no). Endogenous division arises by the in vagi- nation of such a -budding area, while the walls surrounding it grow THE INFUSORIA 199 together above the developing buds, which, when ripe, break through the birth-opening left in the covering membrane (Fig. 1 1 1). In some cases the buds are multiple, again single, and a number may develop at the same time within the brood-sac (Acineta, OpJiryodendron}. The embryos thus formed are variously ciliated in the different genera. In some they are holotrichous, in others hypotrichous, and in others peritrichous (c, d\ Fig. in. Endogenous budding in Suctoria. [BuxsCHLi.] A-B. Two stages in the formation of the bud in Tokophrya quadripartita Cl. and Lach. c. The swarm-spore liberated. C. Buds in Acineta tuberosa Ehr. d. A swarm-spore liberated. Conjugation occurs here as in the Ciliata, but the process rests upon the single observations of Maupas, who shows, however, that it differs in no essential features from that already described. III. INTER-RELATIONS OF THE INFUSORIA In searching for the origin of the Ciliata, the naturalists of thirty years ago had an apparent advantage, in that the supposed ciliated girdle of the Dinoflagellidia offered a direct transition to the peritrichous Ciliata, which, accordingly, were regarded as coming from the flagellate stem at a comparatively late date. Unfortunately for the theory, however, it was ascertained by Biitschli ('85) and others that the girdle of cilia is only a vibrating flagellum in the transverse groove. In other directions the search for the origin of these forms has been almost equally vain. The singularly con- servative structure which the ciliate body presents leaves but little clue to their ancestry. The universal presence of macro- and micro- nuclei is paralleled by only one other known case, the almost universal reproduction by transverse division is met with elsewhere but rarely. The sole possibility which presents itself is that the 200 THE PROTOZOA inf usorian stem was derived from the flagellate at a very early period, and that the side branch became progressively differentiated until the well-marked characteristics of to-day distinguish the Infusoria as an entirely independent group. The first forms to diverge from the flagellate stem may have been like the type described by Cienkowsky, under the generic name of Multicilia (Fig. 112, A), a form with a number of long flagella. It is thought by Butschli that the Ciliata might have been derived from such generalized forms by progressive increase, with shortening of the motile elements, until cilia were the Fig. 112. A. Multicilia lacustris Lauterb. [LAUTERBORN.] B. Pyrsonympha vertens Leidy. [PORTER.] x, the vibrating band in the inner plasm. outcome. There is no close connection, however, between cilia and flagella, such as exists between the flagella and the pseudopodia. Other forms, more or less similar to Multicilia, have been described by various observers, so that the hypothesis of Butschli is not without warrant. Among these forms are Grassia, TrichonympJia, Leidyonella, Lophomonas, Pyrsonympha, etc., which are placed by some among the Flagellidia (Delage), by others among the Ciliata (Butschli). Another point of view has been based upon the relations of the Ciliata to the Suctoria, and through them to the Sarcodina THE INFUSORIA 201 (Entz, Maupas). This view will be more appropriately examined in connection with the Suctoria. The Holotrichida appear to be the most generalized of the entire group of Infusoria, but a few forms among them have a slight regional differentiation of cilia suggesting the characteristics of the Heterotri- chida (Z>7#^7/.s-, Pleuronema, Ophryoglena, etc.). In fact, there appears to be no sharp line between the two divisions, although the presence of an adoral band of cilia in the Heterotrichida is a sufficient differen- Pig. 113. Illustrating Butschli's hypothesis of the origin of the Hypotrichida. [BiJTSCHl.l.] A. Stephanopogon colpoda Entz. B. Peritromus emmce St. C. Onychodromus grandis St. c, cirri. tial. In some forms the uniform coating of cilia is broken in certain regions, giving characteristic girdled forms, which are included as a separate order apart from the Holotrichida by some writers (Haeckel). In the Holotrichida, also, there are a few forms which show a distinct tendency toward bilateral symmetry, due primarily to a bending of the body, and followed by a reduction of the cilia upon the arched side (Stephanopogon, Fig. 113, A). Biitschli derives the Hypotri- chida from the Heterotrichida by the supposition of the loss of cilia upon the arched dorsal side and incomplete closure of the adoral ring 2O2 THE PROTOZOA of cilia, which are here fused to form the characteristic membranelles; the mouth, as in Heterotrichida, remaining on the ventral side. In the most generalized forms, such as Peritromus or Oxytricha (B\ the cilia are well distributed over the ventral surface, but in most of the other Hypotrichida they are reduced, and many are obliterated or fused into character- istic cirri. The cirri in the Oxytrichinae are primitively ar- ranged in six rows, but in the various genera the number becomes reduced, and frequently only iso- lated cirri mark the original position of the row (C). The Peritrichida, finally, show the most far-reaching devia- tions from the holo- trichous type, from which they are prob- ably derived through the Heterotrichida and the Hypotrichida. In. all members of this group the adoral zone is continued into * E a spiral, which may Fig. 114. Illustrating Biitschli's view of the origin of the Vor- have as manv as five ticellidae. [BUTSCHLI.] The Trichodina form C is supposed to have arisen from the Complete tl^ns ( Cam- Lichnophora-Vtian. form A by the outgrowth of the lower ciliated panella). The chief area, first forming an intermediate stage B. This ring of cilia ,*,--,--<;<- rrmr^rnincr becomes lost in the Vorticellidge, appearing only when the indi- viduals are free-swimming. D, E. Side and front of Lichnophora this adoral Zone is ^""Ciap. that in some forms the spiral is turned to the left, similar to that of all of the other groups of Ciliata, while in other forms, belonging to the great family of the Vorticel- lidae, the spiral is turned to the right. The sinistral type of the Peritrichida originates, according to Butschli, from an hypotrichous form, becoming attached- at the posterior half of the ventral surface, THE INFUSORIA 2O3 with loss of the posterior girdle of cilia, and elevation of the anterior region bearing the adoral zone, which, as in the other groups of Ciliata, is turned toward the left. The key to the other group of Peritrichida is seen, Biitschli maintains, in the family Lichnophoridae, where the individuals closely resemble hypotrichous forms (Fig. 114), being oval, flattened ventrally, and arched dorsally. The cilia, as in the Hypotrichida, are limited to the ventral surface, and an adoral zone is present, which runs from the mouth near the middle of the left body edge, entirely around the anterior region of the body, to form an incomplete arc which terminates in the line of the mouth, but on the right side. Another closed ring of cilia is present in the pos- terior half of the ventral surface. The anterior and posterior rings of cilia are separated in LichnopJiora by a stretch of plasm in such a way as quite to divide that surface into a posterior and an anterior division (E). The posterior part becomes modified to form an attaching organ upon which the animal creeps about upon its host ; the anterior region at the same time is elevated, and held in a posi- tion at right angles to the plane of attachment, the apparent stalk which supports it being in reality the intermediate plasm between the posterior and the anterior regions of the ventral surface (-D). Thus the curious anomaly arises of an animal whose anterior and posterior ends represent parts of the same ventral surface. Butschli derives the' entire family of the Vorticellidae from this primitive type, through forms like the Urceolarinae, where the attaching disk, primarily, is not so far removed from the peristome, nor so stalk-like, as it is in the present-day Liehnophoridae (D, E\ He argues that the Vorticella-\.y\)Q, is derived from the Urceolaria- type by the attaching part of the ventral surface, i.e. the pos- terior part being carried outward from the remainder of the ventral surface, and thus borne upon a platform so that the two portions of the same surface are no longer in the same plane (B}. The anterior ring of cilia is then supposed to have grown around the base of the elevated portion until the original adoral zone of cilia now forms a ring about the entire ventral surface. The new arm of this line of cilia grows on past the mouth-opening and forms a spiral, which, looked at from the ventral side, turns to the left, as in all other Ciliata seen from the same surface. Looked at from the other side, however, i.e. dorsally, the spiral turns to the right (C). This condition is practi- cally represented by the genus Trichodina, which moves about on the skin of various Invertebrata*by means of the ciliated or attach- ment disk, in reality the posterior part of the original primitive ventral surface ; while the other portion is now carried dorsally and parallel to the attachment disk, the mouth being on the left side of this anterior part. In the Vorticellidae this posterior or attaching 204 THE PROTOZOA part becomes drawn out into the long contractile stalk, while the ciliated condition, as represented by Trichodina, is again brought about in Vorticella, when the latter breaks away from its stalk, develops a ciliated band in the posterior region, and swims freely about. The ciliated band is homologous with the posterior ring of Liclinophora and the attaching disk of Trichodina, while the adoral zone of cilia conforms to the typical left-handed spiral of the remain- ing ciliates, when looked at from the same morphological point of view. In Gerda, the peristomial region has degenerated, while the ciliated disk remains as the organ of locomotion. Returning now to the origin of the Ciliata as a group, quite another view has been maintained by a number of observers, the essential point of which is that the Ciliata are connected with the Sarcodina through the Suctoria, the tentacles in the latter being regarded as modified pseudopodia. This view was apparently first suggested by Stein ('54) when he included the Heliozoa and the simpler forms of Suctoria in the genus ActinopJirys. The assumption was taken up seriously by Maupas ('81), who held that through the Suctoria, the Ciliata were derived from the Sarcodina, and Penard ('90) accepted the same view in regarding Actinolophus capitatus as a connecting link between the two groups. Claparede and Lachmann were the first to deny the connection of Ciliata and Rhizopoda, but made the even more improbable assertion that the Suctoria are derived from the Flagellidia through forms like Syncrypta volvox. The close rela- tion of the Suctoria and the Ciliata was brought into prominence through Stein's famous, though erroneous, Acineta-theory, in which the Suctoria were supposed to be young forms of Ciliata. The con- nection between the two was, however, first put on a substantial basis by the discovery of the ciliated embryos of the Suctoria, a connection which was early accepted by students of the Protozoa, and which was greatly emphasized by the discovery that, like the Ciliata, the Suctoria have macro- and micronuclei. At the present time it is almost universally held that the Suctoria are offshoots of the Ciliata, although the opposite view is maintained by some observers, who, with Entz ('79, '82), consider the Ciliata as permanent forms of the ciliated embryos of Suctoria. Entz himself regards the matter as insoluble, and believes that the evidence is about equally balanced. A number of cases certainly gives strength to Entz's position, for many of the Enchelinidae, in addition to their cilia, have distinct tentacular processes (Ileonema, Mesodinium, Actinobolus, etc., Fig. 115). Actinobolus, discovered by Stein, and more recently examined by Entz, has long tentacle-like threads evenly distributed about and between the cilia (Fig. 100). They can be lengthened or shortened THE INFUSORIA 205 or entirely drawn 'in by the animal. In Mesodinium, there are only four of these tentacles, which are arranged about the mouth (Fig. 1 1 5, B). Ileonema has only one (A). These processes were considered so important from the phylogenetic standpoint that Mereschowsky ('82) formed a special group, the Suctociliata, for their reception. Neither Entz, nor Stein, nor Mereschowsky, however, regarded the tentacles as food-taking organs like the tentacles of the Suctoria ; the former, at best, could assign to them no other function than that of assisting in the capture of aliments. Maupas regarded them simply as pseudo- podia, and upon them as a basis formulated his view connecting the Ciliata with the Sarcodina. Biitschli strongly opposed Entz's view as to the origin of Suctoria and Ciliata, and believed that there is no Fig. 115. Ciliata with tentacles. A. Ileonema dispar Stokes. [STOKES.] B. Mesodinium pulex Clap, and Lach. [ENTZ.] C. Hypocoma parasitica Grub. [ENGELMANN.] /, tentacles. direct connection between the tentacles of the two groups, but regarded them as independent adaptations. The hypothesis advanced by Butschli is that primitive forms of Suctoria (such as Hypocoma (C), which has but one suctorial tentacle, and which retains its cilia throughout life, the cilia being upon the ventral side only, as in hypotrichous forms of Ciliata) were derived from hypotrichous ciliates by the mouth portion becoming progressively drawn out into a tentacle. Haeckel ('96), adopting Biitschli's view, compared the simple, single, and terminal mouth-tube of a primitive suctorian with the long, proboscis-like oral region of certain holotrichous ciliates, such as Lacrymaria olar or L. phcenicopterus. In the closely allied forms, Didinium and Mesodinium, the oral tube is not ciliated and is contractile, so that when food is taken in, the tube widens into more or less of a disk similar to many suctorian tentacles. In Mesodinium, 2O6 THE PROTOZOA this tube is not only retractile, but is also surrounded by four tentacle- like processes which simulate some kinds of tentacles in the Suctoria. As the majority of the larvae of the Suctoria are ciliated in girdles, Haeckel holds that this division of the Holotrichida represents the nearest allies of the Suctoria, and that the loss of cilia in the adult is already foreshadowed by the regional loss of cilia in these girdled forms. The entire matter, finally, of the origin of Infusoria from more generalized forms of Protozoa remains unsolved ; the various hypotheses are interesting possibilities, but no more can be said for them. This problem, like that of the origin of the Protozoa, may never come nearer settlement ; for, without the assistance of palaeonto- logical and embryological evidence, which in other great groups of the animal kingdom are of inestimable value in tracing ancestors, the possibility of tracing their origin is reduced to a minimum. CLASSIFICATION CLASS V. INFUSORIA. Protozoa in which the motor apparatus is in the form of cilia, either simple or united into membranes, membranelles, or cirri. The cilia may be permanent or limited to the embryonic stages. With two kinds of nuclei, macronucleus and micronucleus. Reproduction is effected by simple transverse division or by budding. Nutrition is holozoic or parasitic. Subclass I. CILIATA. Infusoria provided with cilia during adult as well as embryonic life. Reproduction is brought about typically by simple transverse division. Mouth and anus are usually present. The contractile vacuole is often connected with a complicated canal system. Order i. HOLOTRICHIDA. Ciliata in which the cilia are similar and distributed all over the body, with, however, a tendency to lengthen in the vicinity of the mouth. Trichocysts are always present, either distributed about the body or limited to a special region. Suborder i. GYMNOSTOMINA. Holotrichida without an undulating membrane about the mouth, which remains closed except during food-taking intervals. Family i. Enchelinidae. The mouth is always terminal or sub-terminal, and is usually round or oval in outline. Food-taking is usually a process of swal- lowing. Genera: Holophrya Ehr. ('31); Urotricha Clap. & Lach. ('58); Enchelys Hill (1752), Ehr. ('38) ; Spathidium Duj. ('41) ; Chcenia Quennerstadt ('68) ; Prorodoti Ehr. ('33); Dinophrya BUtschli ('88); Lacrytnaria Ehr. ('30); Trachelocerca Ehr. ('33); Actinobohts Stein ('67); Ileonema Stokes ('84); Plagiopogon Stein ('59); Coleps Nitsch ('27); Tiarina Bergh ('79); Stepha- nopogon Entz ('84) ; Didinium Stein ('59); Mesodinium Stein ('62); Butschlia Schuberg f86). Family 2. Trachelinidae. The body is distinctly bilateral or asymmetrical, with one side, the dorsal, slightly arched. The mouth may be terminal or sub- terminal, or the entire mouth-region may be drawn out into a long proboscis. An resophagus or gullet may or may not be present ; when present, it is usually supported by a specialized framework. Genera: Amphileptus Ehr. ('30); Lionotus Wrzesniowski ('7); Loxophylliim Duj. ('41); Trachelius Schrank ('03) ; Dileptus Duj. ('41) ; Loxodes Ehr. ('30). THE INFUSORIA 2O/ Family 3. Chlamydodontidse. The general form is oval or kidney-shaped. The mouth is almost always in the posterior region. The pharynx is supported by a rod-apparatus or a smooth, firm tube. Subfamily i. Nassulince. Ciliation is complete. Genera: Nassula Ehr. ('33). Subfamily 2. CkHodontiiue. The body is generally flattened, and the cilia are stronger on the dorsal side, or are confined to that region. Genera : Orthodon Gruber ('84) ; Chilodon Ehr. ('33) ; Chlamydodon Ehr. ('35) ; Opisthodon Stein ('59) ; Phascolodon Stein ('57); Scaphidiodon Stein ('57). Subfamily 3. Erviliince. The cilia are confined to the ventral surface or to a por- tion of it. The posterior end invariably possesses a movable style arising from the posterior ventral surface. Genera: sEgyria Clap. & Lach. ('58); Onychodactylus Entz. ('84) ; Trochilia Duj. ('41) ; Dysteria Huxley ('57). Suborder 2. TRICHOSTOMINA. In addition to the general coating of cilia there is an undulating membrane or membranes at the edge of the mouth or in the pharynx. The mouth is always open. Family i. Chiliferidse. The mouth is in the anterior half of the body or close to the middle The pharynx when present is short. The so-called "peristome area" leading to the mouth is absent or only slightly developed. Genera: Leucophrys Ehr. ('30) ; Glaucoma Ehr. ('30) ; Dallasia Stokes ('86) ; Fron- tonia Ehr. ('38); Ophryoglena Ehr. ('31 ); Colpidiunt Stein ('60); Chasmato- stoma Engelmann ('62) ; Uroncma Duj. ('41) ; Urozona Schewiakoff (BUtschli) ('88) ; Loxocephahis Kent ('81) ; Colpoda Miiller (1773). Family 2. Urccentridag. The mouth, with a long, tubular pharynx, is in the centre of the ventral side. The cilia are confined to two broad zones around the body at each end. Genera: Urocentrum Nitsch ('27). Family 3. Microthoracidae. Small asymmetrical forms, with the mouth invariably in the hinder portion. The cilia are always more or less dispersed, sometimes limited to the oral region. There may be one or two undulating membranes. Genera: Cinetochilum Perty ('49); Microthorax Engelmann ("62); Ptychosto- mum Stein ('60) ; Ancistrum Maupas ('83) ; Drepanomonas Fresenius ('58). Family 4. Paramcecidae. The mouth is sometimes in the anterior, sometimes in the posterior, half of the body, and is accompanied by a large, triangular "peri- stome area" running from the left anterior edge of the body to the mouth. Genera: Paramcecium Stein ('60). Family 5. Pleuronemidae. The mouth is at the end of a long peristome which runs along the ventral side : the body is dorso-ventrally or laterally compressed. The entire left edge of the peristome is provided with an undulating membrane which occasionally runs around the posterior end of the peristome to form a pocket leading to the mouth. The right edge of the peristome is pro- vided with a less developed membrane. There may or may not be a well- developed pharynx. Genera: Lembadion Perty ('49) ; Pleuronema Duj. ('41); Cyclidium Ehr. ('38), a sub-genus of the preceding; Calyptotricha Phillips ('82); Lembus Cohn ('66). Family 6. Isotrichidae. The body is more or less plastic, but not contractile. The cuticle is thick and provided with evenly distributed cilia. The mouth is posterior and accompanied by a distinct pharynx. They are parasites in the digestive tract of ruminants. Genera : Isotricha Stein ('59) : Dasytricha Schu- berg ('88). Family 7 . Opalinidae. The form is oval, and the body may be short or drawn out to resemble a worm. They are characterized mainly by the absence of mouth and pharynx. Genera: Atioplophrya Stein ('60); Hoplitophrya Stein ('60); Discophrya Stein ('60); Opalinopsts Foettinger ('81); Opalina Purkinje and Valentin ('35) ; Monodontophrya Vejdowsky ('92). 2O8 THE PROTOZOA Order 2. HETEROTRICHIDA. Ciliata characterized by the possession of a uniform covering of cilia and an adoral zone, consisting of short cilia fused together into membranelles. Suborder I. POLYTRICHINA. Heterotrichous ciliates provided with a uniform coating of cilia. Family i. Plagiotomidae . The peristome is a narrow furrow, which begins, as a rule, close to the anterior end, and runs backward along the ventral side to the mouth, which is usually placed between the middle of the body and the posterior end. A well-developed adoral zone stretches along the left side of the peristome, and it is usually straight. Genera: Conchophthirits Stein ('61 ); Plagiotoma Duj. ('41); Nyctotherus Leidy ('49), a sub-genus; Blepharisma Perty ('49); Metopus Clap. & Lach. ('58) ; Spirostonntm Ehr. ("35). Family 2. Bursaridae. The body is usually short and pocket-like, but may be elongate. The chief characteristic is the peristome, which is not a furrow, but a broad triangular area, deeply insunk, and ending in a point at the mouth. The adoral zone is usually confined to the left peristome edge, or it may cross over to the right anterior edge. Genera : Balantidiutn Stein ('67) ; Balantidiopsis Biitschli ("88); Condylostoma Duj. ('41); Bitrsaria O. F. Mliller (1773); Thylakidium Schewiakoff ('92). Family 3. Stentoridae. The peristome is relatively short and limited to the front end of the animal, so that its plane is nearly at right angles to that of the longi- tudinal axis of the body. The adoral zone of cilia either passes entirely around the peristome edge, or ends at the right-hand edge. The surface of the peri- stome is spirally striated and provided with cilia. Undulating membranes are absent. Genera: Ctimacostomui\ia(?yf)\ Stentor Oken ('15) ; Folliculina Lamarck ('16). Genera incerta sedis: Canomorpha (Gyrocorys Stein) Perty ('52) ; Maryna Gruber ('79). Suborder 2. OLIGOTRICHINA. Heterotrichous ciliates characterized by the re- duced cilia, which are limited to certain localized areas. Family I. Lieberkiihnidae. This name was given by Butschli for certain little-known forms, which were at first considered young Stentors. Family 2. Halteriidae. The peristome has no cilia, and only a few scattered ones can be found on the ventral and dorsal surfaces. Genera : Strombidium Clap. & Lach. ('58) ; Halteria Duj. ('41). Family 3. Tintinnidae. The body is attached by a stalk to a theca. Inside of the adoral zone of membranelles is a ring of cilia (par-oral cilia). Genera : Tintin- nus Fol. ('89) ; Tintinnidium Kent ('81) ; Tintinnopsis Stein ('67) ; Codonella Haeckel ('73) ; Dictyocysta Ehr. ('54). Family 4. Ophryoscolecidae. Heterotrichous ciliates characterized by a thick cuticle and deep funnel-like peristome. The posterior end is provided with distinct spine-like processes, while the terminal anus is provided with a well-defined anal tube. Genera: Ophryocolex Stein ('59); Entodinium Stein ('59) ; Diplodinium Schuberg ('88). Order 3. HYPOTRICHIDA. Ciliata in which the cilia are limited to the ventral surface of a dorso-ventrally flattened body ; they are frequently fused to form larger appendages, the cirri, and an adoral zone of membranelles. The dorsal surface is frequently provided with bristles. A pharynx may be absent or but slightly developed. Family i. Peritromidae. The peristome is but slightly marked off from the remain- ing frontal area. The cilia on the ventral surface are uniform in size and arrangement, and are not differentiated into cirri. Genera : Peritromus Stein ('62). THE INFUSORIA 2OQ Family 2. Oxytrichidae. The peristome is not always distinctly marked off from the frontal area. In the most primitive forms the ciliation on the ventral surface is similar to that of the preceding family. Almost invariably in these primitive forms some of the anterior and some of the posterior cilia are fused into large and more powerful appendages, the cirri, which are distinguished as the frontal and anal cirri, respectively. In the majority of forms all of the cilia are thus dif- ferentiated ; strong marginal cirri are formed in perfect rows, and ventral cirri in imperfect rows. In addition to the adoral zone of membranelles, there is an undulating membrane on the right side of the peristome, and, in some cases, a row of cilia between the membrane and the adoral zone. These are the par- oral cilia, and they form the par-oral zone. Genera: Trichogaster Sterki ('78) ; Urostyla Ehr. ('30) ; Kerona Ehr. ('38) ; Epiclintes Stein ('62) ; Stichotricha Perty ('49) ; Strongylidium Sterki ('78) ; Amphtsia Sterki ('78) ; Uroleptus Stein ('59) ; Sparotricha Entz ('79) ; Onychodromus Stein ('59) ; Pleurotricha Stein ('59) ; Gastrostyla Engelmann ('62) ; Gonostomwn Sterki ('78) ; Urosoma Kowalewsky ('82) ; Oxytncha Ehr. ('30) ; Stylonychia Stein ('59) ; Actinotricha Cohn ('66) ; Balladina Kowalewsky ('82) ; Psilotricha Stein ("59) ; Tetrastyla Schewiakoff ('92) ; Holosticha Wrzesniowski ('77). Family 3. Euplotidae. Hypotrichous ciliates, which are characterized mainly by the considerable reduction of the cilia, as well as the frontal, marginal, and ventral cirri ; the anal cirri, on the other hand, are always present. The macronucleus is band-formed. Genera : Euplotes Stein ("59) ; Certesia Fabre-Dumergue ('85); Diophrys Duj. ('41) ; Ifronychia Stein ('57); Aspidisca Ehr. ('30). Order 4. PERITRICHIDA. Ciliata usually of cylindrical or cup-like form, in which the cilia are reduced, as a rule, to those which form the adoral zone, but sec- ondary rings of cilia may be present. Family i. Spirochonidae. Peritrichous ciliates in which the peristome is drawn out into a curious funnel-like process, either simple or rolled. They are parasitic forms in which reproduction by budding is characteristic. Genera: Spirochona Stein ('51) ; Kentrochona Rompel ('94) ; Kentrochonopsis Dorlein ('97)- Family 2. Lichnophoridae. In addition to the adoral zone, there is a secondary circlet of cilia around the opposite end. The adoral zone is a left-wound spiral. A single genus, Lichnophora, Claparede ('67), which is parasitic on various marine arthropods. Family 3. Vorticellidae. Attached or unattached forms of peritrichous ciliates, in which the adoral zone, seen from above, forms a right-wound spiral (dexiotropic). A secondary circlet of cilia around the under end may be present either perma- nently or periodically. Subfamily i . Urceolarince. Vorticellidae having a permanent secondary circlet of cilia which incloses an adhesive disk, and without a peristome fold. Genera: Trichodina Stein ('54) ; Cyclochceta Jackson ('75) 5 Trichodinopsis Clap. & Lach. ('58). Subfamily 2. Vorticellidince. Peritrichous forms without a permanent secondary circlet of cilia, and provided with a peristome fold which can be contracted sphincter-like to inclose the peristome. Genera : Scyphidia Lachmann ('56) ; Gerda Clap. & Lach. ('58); Astylozoon Engelmann ('62); Vorticetta Ehr. ('38) ; Carchesium Ehr. ('30) ; Zoothamniitm Stein ('54) ; Glossatella Biitschli ('88) ; Epistylis Ehr. ('30) ; Rhabdostyla Kent ('82) ; Opercularia Stein ('54) ; Ophrydium Ehr. ('38) ; Cothurnia Clap. & Lach. ('58) ; Vaginicola Clap. & Lach. ('58); Lagenophrys Stein ('51). Subclass II. SUCTORIA. Infusoria having no cilia during the adult stages, but provided with them during the embryonic period. In a few cases the cilia are retained. They have tentacles of various kinds, some adapted for sucking, some for piercing. 210 THE PROTOZOA Family i. Hypocomidae. These are unattached forms of Suctoria with a perma- nently ciliated ventral surface, and with one suctorial tentacle. Reproduction is effected by cross-division. A single genus, Hypocotna Gruber ('84). Family 2. Urnulidae. A family of small attached forms, with or without a cup or theca; with one or two, rarely more, simple tentacles. Swarm-spores holo- trichous. Genera: Rhyncheta Zenker ('66) ; Urnula Clap. & Lach. ('58). Family 3- Metacinetidae. Thecate forms ; the base of the cup is drawn out into a long stalk, and the walls are perforated for the exit of the tentacles. A single genus, Metacineta Biitschli ('88). Family 4. Podophryidae. Stalked or unstalked forms of more or less globular shape. The tentacles are numerous and distributed about the entire surface or limited to the apical region ; some of them are knobbed, others pointed and have a prehensile function. Genera: Sphcrrophrya Clap. & Lach. ('58); Endospluzra Engelmann ('76) ; Podophrya Ehr. ("38) ; Ephelota Str. Wright ('58); Podocyathus Kent f8l). Family 5. Acinetidae. The individuals are naked and stalked, or thecate and stalked or unstalked. The tentacles are numerous, usually knobbed and all alike. Reproduction is effected by inner or endogenous budding, which may be simple or multiple. The swarm-spores are usually peritrichous, but may be holotrichous or hypotrichous. Genera: Tokophrya Biitschli ('88); Acineta Ehr. ('33) ; Solenophrya Clap. & Lach. ('58) ; Suctorella Frenzel ('91). Family 6. Dendrosomidae. Suctoria without stalks or theca. The tentacles are numerous, all alike, and knobbed and grouped in distinct tufts ; they may be simple or branched. Reproduction by endogenous division ; the swarm-spores are peritrichous. Genera: Trichophrya Clap. & Lach. ('58); Dendrosoma Ehr. ('38) ; Staurophrya Zacharias ('93). Family 7. Dendrocometidae. Sessile Suctoria resting upon the entire basal surface or upon a portion of it raised as a stalk. The numerous tentacles are short and knobbed, and distributed over the entire apical surface or localized upon branched arms. Spore-formation is endogenous ; the swarm-spores peritrichous. Genera : Dendrocometes Stein ('67) ; Stylocometes Stein ('67). Family 8. Ophryodendridae. Stalked or sessile forms possessing numerous long, rarely knobbed tentacles, which are supported upon proboscis-like processes of the apical side. Reproduction is brought about by endogenous budding. The swarm-spores are peritrichous. Genera: Ophryodendron Clap. & Lach. ('58). SPECIAL BIBLIOGRAPHY VI Biitschli, 0. Protozoa, Infusoria. In Bronn's Klassen und Ordnungen des Thier- reichs. Leipzig, 1886-88. Entz, G. Protistenstudien. Budapesth, 1888. Kent, W. Saville. A Manual of the Infusoria. London, 1881. Schewiakoff, W. Beitrage zur Kenntniss der Holotrichen Ciliaten. Bibliotheca Zoologica, Heft 5, 1889. Stein, Fr. Der Organismus der Infusionsthiere. Leipzig, 1863 and 1867. CHAPTER VII SEXUAL PHENOMENA IN THE PROTOZOA " Die Bedeutung des Konjugationsaktes ist ein Verjiingung der ihn begehenden Tiere. Durch diese Verjiingung erscheinen uns die aus der Konjugation hervorgehenden Individuen sehr geeignet, zu den Stammvatern einer Reihe durch Theilung sich fortpflanzender Gene- rationen zu werden, im Laufe welcher allmahlich ein Sinken der Lebensenergiesicheinstellt. Letzter Umstand findet seinen Ausdruck darin, das die Grosse der Individuen mehr und mehr sinkt sodass schliesslich eine minimalgrdsse erreicht wird, worauf eine neue Konjuga- tionsepoche eintritt." BL'TSCHLi. 1 THE power of the animal or plant to reproduce its kind from a por- tion of its own body is bound up, in the higher forms of life, with sexual processes and, in its more familiar forms, accordingly, is characterized as sexual reproduction. It involves the union of two cells having quite different characteristics ; the spermatozoon or male cell, being minute and active, the ovum or female cell, larger and quiescent. Reproduc- tion of the individual without sexual processes is, however, possible, even in the higher organism, as we daily witness in plant " cuttings," and as is almost equally well known in the higher forms of inverte- brates such as insects, where, without . fertilization, an ovum may develop into an adult form. In the lower forms of Invertebrata, such as the worms and the Ccelenterata, so-called " asexual reproduction " by division or by budding is widespread, and in the Protozoa this method of reproduction is the usual form. The phenomena of parthenogenesis, or development from the egg without fertilization, and reproduction by simple division or by bud- ding as seen in the Protozoa and the lower Metazoa, have recently led to the pertinent query : With what right do we distinguish the phe- nomena of reproduction as "sexual " and " asexual " ? (Hertwig, '99). In two recent publications R. Hertwig ('98, '99) has discussed this question in a very interesting and convincing manner, and he main- tains that, in order to speak of " sexual " reproduction, it must be shown that in the Protozoa, for example, fertilization has some direct effect upon division or that a certain specific form of division results from fertilization, neither of which, he says, is true in the majority of cases. All observers are apparently agreed that asexual reproduction as seen in the processes of binary fission and spore-formation is a result of growth, usually expressed by the statement that increase 1 ('76), P . 421. 211 212 THE PROTOZOA in volume continually tends to outrun that of surface, for the former increases as the cube of the diameter, the latter only as the square. The mass of living protoplasm must, therefore, increase more rapidly than the surface which serves to keep it alive, and the isolated cell tends to come first to a physiological standstill, and second to a period of decline, since the surface of nutrition, respiration, and excretion is incommensurate with the bulk of protoplasm. Cell-division, there- fore, which Spencer characterized as an indication of the limit of growth, becomes an apparent necessity. Growth, or the preponderance of constructive processes, leads thus indirectly to increase of surface by division of the cell, but in conju- gation or fertilization the very opposite phenomenon occurs, vis. the increase in bulk of the cell with a consequent relative decrease in sur- face because of the union of two cells. Geddes and Thompson ('90), like Spencer, emphasize the connection between division and the advent of preponderating katabolism within the cells, and in their interesting work on the Evolution of Sex, interpret male and female organisms in terms of relative metabolism, the male being regarded as relatively katabolic, the female as anabolic. These authors interpret fertilization as a " katabolic stimulus to an anabolic cell, and on the other side, of course, as an anabolic renewal to a katabolic cell, as well as the union of opposed hereditary characteristics." 1 If, as Minot ('79) suggested, every newly formed organism be re- garded as having a certain initial potential energy which is gradually used up in its life-activities to be restored by conjugation, then the union of two cells may be interpreted as a renewal of vigor or a " re- juvenescence " (Maupas), a view of fertilization first expressed by Biitschli ('76), Engelmann ('76), and Minot ('77, '79), and apparently confirmed later by Maupas ('88, '89) through his admirable observa- tions on the life-cycle of Infusoria. O. Hertwig ('76) also, in con- nection with fertilization of the metazoan egg, arrived at a similar dynamic view of fertilization, maintaining that protoplasm gradually tends toward a state of stable equilibrium expressed by decreased activity, etc., and that it is restored to a more unstable or more labile condition by conjugation or fertilization. The force of these views as to the need of conjugation for different species of Infusoria, at least, can hardly be questioned ; for, as repeatedly stated in the pre- ceding chapters, reproduction by simple division may go on for a certain number of generations, but cannot continue indefinitely, unless at certain intervals, which Maupas has shown to be more or less definite, two individuals unite in conjugation. This union, in some wholly unexplained way, imparts to each of the conjugants 1 Loc. cit., p. 232. SEXUAL PHENOMENA IN THE PROTOZOA a renewed vitality, or in Biitschli's words ('76) a renewal of youth ( Verjiingung}, expressed by increased activity in movements and reproduction. Conjugation thus, as R. Hertwig insists, is not the beginning of a series of reproductive acts, but occurs at or near the end of such a series. Maupas's results seem to offer conclusive evi- dence that the absence of conjugation involves a cumulative degener- ative process which ultimately ends in death. The phenomena of so-called sexual reproduction and of sex-differentiation have, in all probability, grown out of this apparently fundamental requirement of living protoplasm, namely, the periodic union of two cells ; and I believe with Butschli, Engelmann, Maupas, Hertwig, and many others, that it cannot in itself be regarded as primarily a repro- ductive act. None of the facts that have been determined show that the morphological distinction of the sexes is a primary attribute or property of living organisms, nor do any of the dynamic views of fertilization afford an explanation of sex-differences any more than does the statement of Geddes and Thompson cited above. Even though the views of these authors be accepted, we should still have to admit that no explanation of fertilization can be wholly satisfactory unless it is equally applicable to forms which cannot be distinguished as more anabolic or katabolic, i.e. to the conjugation of equal-sized Infusoria, Mastigophora, or Sporozoa, as well as the union of differ- entiated male and female cells. If then we define sex as the condi- tion by which single-celled or many-celled organisms are differentiated into male and female, we must admit that the origin of sex is only a part of the problem, for fertilization occurs between individuals in which there is no apparent sex-difference, as well as between those possessing it. In the present chapter I have brought together some of the evi- dence which bears upon these several points. A number of phe- nomena which accompany reproduction in the higher animals, and which have attracted so much attention among biologists of all times, are seen in simpler forms in Protozoa. Among these the phenomena of sex-differentiation, of maturation or preparation for fertilization, and fertilization itself are of paramount interest. The explanation of sex-differentiation is not as yet made more easy by the study of Protozoa, and here as in higher animals it must remain entirely hypothetical until future research throws more light upon the problem. The conditions accompanying conjugation, how- ever, have been carefully studied and analyzed in relation to other vital functions of the cell, and the evidence thus acquired gives a clue, I believe, to an ultimate explanation of fertilization. An important underlying principle was first made out by Butschli ('76) and Engel- mann ('76) upon degenerating Infusoria, and was expressed by Minot 214 THE PROTOZOA three years later in the sentence, " the exhaustion of the rejuvenating power (i.e. the exhaustion of the initial potential of vitality) becomes the stimulus for the formation of the sexual products." 1 A. PHENOMENA OF CONJUGATION With our present knowledge it is impossible to say that conjuga- tion is absent in any group of Protozoa, and until the life-cycle of every genus is fully known, the conservative but logical view which Biitschli expressed in regard to flagellates, is the most acceptable. He says : " I, personally, am inclined to the view that the significance of this process in the life of these organisms is so general and deep- reaching that the failure to observe it in certain groups up to the present time is no reason for considering it absent." 2 Nevertheless, the observations which have been recorded show the greatest diversity in the process, the variations passing from extremely simple fusion, which only by stretching the meaning of the term can be called sexual, to the highly differentiated male and female organisms where, as in the Metazoa, fertilization is followed by cleavage. Stages in the development of so-called sexual reproduction may be considered as follows : 1. The permanent or temporary union of similar adult cells {Isogamy} (Sarcodina, Sporozoa, Mastigophora, Ciliata). 2. The union of individuals apparently similar in all respects save size (Anisogamy} (Sarcodina, Mastigophora, Ciliata). 3. The union of reduced individuals. Swarm-spores (Isogamy or Anisogamy} (Sarcodina, Mastigophora). 4. The union of specialized individuals male and female cells (Spermatozoa and eggs) (Sporozoa, Flagellidia). i. The permanent or temporary union of similar adult individuals (Isogamy). Thanks to the unbroken observations of Dallinger and Drysdale, the conjugation and full life-history of some of the lowest forms of Pro- tozoa (monads) have been made out. All of the forms examined reproduce by simple division for a few days, and then conjugate. In Cercomonas longicauda Duj. (typica Kent), one of the Monadida, reproduction by ordinary fission continues for two to four days, when the offspring, without losing their flagella, become amoeboid and con- jugate two by two (Fig. 116). The union begins with the fusion of the pseudopodial processes, and, as it progresses, the flagella are withdrawn. The nuclei finally unite (D\ and the product of the union, or the zygote, forms a thin-skinned globular cyst (). After a 1 ('79), p- 199- 2 ('83), p- 778- SEXUAL PHENOMENA IN THE PROTOZOA 215 short resting period the cyst breaks and an innumerable quantity of fine spores pour out (F). D Fig. 116. Conjugation of Cercomonas. [DALLINGER and DRYSDALE.] The vegetative cells increase by transverse division (A. B) . When sexually mature they become amoeboid, and then fuse (C, D). The result is a zygote (), which ultimately bursts and liberates masses of spores (F). In Tetramitus, one of the Polymastigida, a similar period of repro- duction by longitudinal division finally results in amoeboid forms which conjugate, forming cysts and spores (Fig. 117). The process 2l6 THE PROTOZOA of conjugation is somewhat modified in certain Bodos of the group Heteromastigida, where, after a period of binary fission, as in Cerco- monas, the individuals become amoeboid, two or three fusing while in D Fig. 117. Conjugation in Tetramitus rostratus Perty. [STEIN.] A, B. Individuals from the front and side. C. Amoeboid form. D. Conjugation of amoeboid forms. E. Cyst F, G, H. Development of the spores. this condition to form a common mass. After a resting period, the encysted mass breaks up into an immense number of small individuals, or spores, differing from those in Cercomonas, in that each one is similar to the parent organism. This union, however, appears to be purely facultative, for the same process of encystment and spore- SEXUAL PHENOMENA IN THE PROTOZOA 217 formation may take place in an isolated individual, and in this case, at least, reproduction cannot be dependent upon sexual union or conjugation, although it does not signify that conjugation is not nec- essary for the continued power of reproducing. Kent ('81), who has confirmed Cienkowsky's ('65) observations upon conjugation of Bodo augustatus, states that a difference exists in the number of spores that are formed in the fusion and in the solitary cysts, only four spores arising from the latter. A significant feature in the conjugation of these forms is that the individuals lose their customary outline and become amoeboid prior to Fig. 118. Conjugation in Rhizopoda. [RHUMBLER.] A, B, C. Difflugia lobostoma Duj. D. Aggregated condition of Amceba verrucosa Ehr. fusion, thus showing that some change has taken place in the con- sistency of the plasm. A great number of observations have been made among the Rhi- zopoda upon so-called conjugation phenomena between similar indi- viduals, the process varying in complexity from simple contiguity to the more complicated fusion of cell-bodies. While many of the earlier observations probably dealt with division phases rather than with conjugating individuals, a possibility of error first pointed out by Claparede and Lachmann ('56), conjugation of shelled forms has been safely established through the observations of Biitschli ('74), Jickeli ('84), Blockmann ('88), Penard ('90), Rhumbler ('98), and others, and of unshelled forms by Schultze (Gromia, '75), Holman ('86), and Kiihne ('64). The phenomena of cytotropy, or the mutual attraction of two or more cells,, among the Sarcodina at least, if not in all forms, are probably closely connected with conjugation and may possibly be the 2l8 THE PROTOZOA first stages in the development of sexual reproduction. 1 It is certainly reasonable to argue that the mutual attraction of two previously separated blastomeres of the frog's egg (Roux, '94), or the reunion of an amputated pseudopodium with the main body of Difflugia (Verworn, '88; Rhumbler, '98), the union of numerous naked Amoeba verrucosa into a common aggregate (Rhumbler, '98), and the union of two conjugating individuals, are all phenomena of the same order. The phenomenon in Amoeba appears to have no bearing upon the function of reproduction, for, according to Holman's and Kiihne's accounts, conjugation is not followed by reproduction, while true conjugation phases may yet be found in other stages of the life- history of Amoeba (Fig. 118, D}. The discovery by Schaudinn of swarm-spores in the allied form Paramceba, and of the union of swarm-spores in the more or less closely allied rhizopod Hyalopus, makes it not at all improbable that the same thing may occur in Amoeba verrucosa. Thus cytotropy, leading first to contiguity, may result in plastogamy, or the fusion of cell- plasms. The protoplasm, however, must be in the proper plastic condition for such a union. Some forms/ such as the Mycetozoa and some Heliozoa, are apparently always in this condition, and contact results in fusion. In many such cases plastogamy leads to noth- Pig. up. ^Conjugation in in g further, nuclear fusion (karyogamy) not ia vuigaris Ehr. occurring. Many instances of such union are found amon g the Sarcodina, and up to the present time they have been seen nowhere else. These unions take place not only in viscous forms such as the plasmodia of Mycetozoa, Actinophrys or Actinosphcerium, but also in the denser types of Rhizopoda. Thus, in Arcella, two or more individuals may fuse together (Fig. 119), and in the shelled rhizopod Difflugia lobostoma, Rhumbler ('98) has shown that two, three, or four individuals may be found in plastogamic union in from six to ten per cent of all cases. He also made the interesting and signifi- cant observation that this union cannot be induced at will, and concluded that plastogamy takes place here only under certain conditions of the plasm. 2 Such plastogamic union in the cases cited has apparently no effect upon the united organisms; both Johnson and Schaudinn found no changes in the nuclei in Actinophrys and Actinosphcerium, and 1 Cf. Cuenot ('97) ; Rhumbler ('98). 2 Cf. union during the amoeboid stage of Cercomonas and other flagellates. SEXUAL PHENOMENA IN THE PROTOZOA 2 19 * Rhumbler reached similar results in the fused Difflugias. The latter, however, calls attention to the fact that two organisms thus united are subject to the interchange of substances through osmosis, and he maintains that such an interchange must take place between them. This interchange may even extend to the substances of the nuclei, which are constantly renewed from, and given off to, the cytoplasm. Although fusion of the nuclei of the forms just mentioned does not take place, the stimulus of the cytoplasmic interchange in some simi- lar cases is apparently sufficient to bring about reproduction. Thus, in certain Reticulariida(/'Vr/V//<'# corrugata and Discorbina globularis> Schaudinn, '95), two, three, four, or even five individuals may fuse and form embryos without a previous nuclear union. In these cases plastogamy alone is apparently a sufficient stimulus for- reproduction. It is by no means a fanciful assumption to postulate the union of the two nuclei, or karyogamy, through conditions similar to those which lead to the union of two organisms through cytotropy, i.e. mutual attraction and consequent fusion when the nuclear plasm is in the right condition, a condition denned by Dangeard ('99) as "sexual hunger." Almost all cases of karyogamy are complicated by nuclear pro- cesses analogous to maturation in Metazoa, a few doubtful cases among the Mastigophora and Rhizopoda alone indicating that fusion may take place without a preliminary loss of a portion of the nucleus. Thus Jickeli ('84) and Rhumbler ('98) observed two individuals of Difflugia globulosa with but a single nucleus in conjugation, and similar observations by Penard ('90) upon a number of different species indicate that the phenomenon is widespread (Fig. 118, A). In D. lobostoma, Rhumbler occasionally found two mouth openings in one shell, and interpreted it as a case of fusion of shells as well as of protoplasm (B, C). Blochmann ('88) observed the fusion of two EuglypJias and the formation of a large double shell. In both cases there was but one functional nucleus observed, although in the latter form the peculiar behavior of the nucleus in one animal was very suggestive of maturation (vide infra}. The union of nuclei in temporary conjugants among the Flagel- lidia has been observed in at least one case, Noctiluca miliaris (Cienkowsky, '73, and Ishikawa, '91). Two individuals fuse, their nuclei come together, but do not fuse, and then separately undergo mitosis, which results in four daughter-nuclei. These separate and then fuse in the daughter-cells, two by two ; thus nuclear fusion takes place some time after conjugation. In the simpler flagellates the nuclei fuse before spore-formation (Dallinger and Drysdale, '73). 220 THE PROTOZOA * Karyogamy is widely spread throughout the Infusoria, where conjugation in the different species is characterized by very similar features. Two individuals unite, usually by the anterior ends, with the mouth openings apposed. A protoplasmic bridge is formed between the two, through which there is an interchange of micro- nuclei. This interchange is followed by final separation of the conjugants, each of which regenerates the parts lost during the period of conjugation (oral cilia, macronucleus, etc.). After careful observations upon many different species of Infusoria, Maupas ('88, '89) found that certain conditions are apparently necessary in order that conjugation between two individuals can take place and lead to fertile results. These conditions are: (i) Sexual maturity, that is, the individuals must be removed by some generations from the last conjugating pair. Maupas established the fact that, in LeucopJirys patula, only individuals of the three hundredth to the four hun- dred and fiftieth generation could be reinvigorated by conjugation; in OnycJiodromns only individuals between the one hundred and fortieth and the two hundred and thirtieth generation ; and in Stylo- nychia pustulata only individuals between the one hundred and thirtieth and the one hundred and eightieth generations. If these individuals, when thus mature, are restrained from pairing, they become over-mature, after which, if they conjugate, the union is without result, and the individuals finally succumb to what Maupas calls " senile degeneration." (2) A second condition is scarcity of food. Maupas also shows that an over-abundance of food causes the individuals to die from senile degeneration without developing the "sexual hunger." (3) A third condition is diverse ancestry. Maupas arrived at the conclusion that two individuals from the same ancestor would not conjugate. " In many pure cultures of nearly related individuals," he says, " the fast to which I subjected them resulted either in their becoming encysted, or in their dying of hunger." " It was not until after senile degeneration had already begun to make inroads in the culture that I noticed that the conjugation of nearly related indi- viduals occurred in the experimental cultivations. However, all such conjugations ended with the death of the Infusoria which had paired, but which were unable to develop further, or to reorganize themselves after they had fused. Such pairings are, therefore, pathological phenomena due to senile degeneration." J The latter conclusion, drawn by Maupas, has not been entirely sustained. Biitschli was one of the first to question it, 2 and recently Joukowsky ('98) observed fertile conjugations among descendants of 1 Loc. cit., p. 411. 2 ('88), p. 1638. SEXUAL PHENOMENA OF THE PROTOZOA 221 the same individual. Maupas's conclusion, therefore, that cross- fertilization is necessary for Infusoria, as for Metazoa, appears to have been somewhat premature, although in view of the extreme care with which his observations and experiments were made, the objections which have been brought against it are not entirely conclusive. It is evident from the foregoing review that, with the exception of sex-differentiation, all of the essential features which characterize fertilization are present in those forms of Protozoa where conjugation takes place between similar adult individuals. Here, also, a hint as to the significance of fertilization is seen in the fact that the form of the conjugating individuals is altered, thus indicating some change in the density of the protoplasm. Thus some Mastigophora and Sarco- dina become viscous, and some Infusoria show unmistakable signs of exhaustion. Under these changed conditions the fusion of the cell-body is possible (plastogamy). This fusion may be partial (Cystoflagellidia, Gregarinida, Infusoria), or total (Monadida, Heliozoa, Rhizopoda), and it may or may not be accompanied by nuclear fusion (karyogamy). The same, stages may be conceived for the union of the nuclei as for the union of the cell-bodies, the evidence appearing to show that as plastogamy is the outcome of cytotropy, or positive chemotaxis, so karyogamy is the outcome of karyotropy or nuclear attraction, and is made possible by plastogamy. 2. The union of similar but different-sized individuals, Sexual differentiation is established when the conjugating organ- isms are of different size. No sharp line, however, can be drawn between conjugation in isogamous and anisogamous forms, but a number of instances might be cited in which the union of different- sized individuals is purely facultative, and the same result is accom- plished either by the union of similar or of dissimilar forms. Thus, of two conjugating Bodos (Heteromita\ one, which is formed by transverse division, is motile and becomes attached to a stationary form resulting from a longitudinal division, and anchored by one of its flagella. With the exception of these differences, which certainly indicate some internal difference in the gametes, the conjugants are identical. Here, for the first time, a distinction can be made between the more quiescent and the more motile conjugant, although it is not marked by difference in size. The latter condition, however, exists in Polytoma, where, according to Krassilstschik ('82) and Dallinger and Drysdale, normal free-swimming forms unite with smaller ones. Here, however, the process is purely facultative, for conjugation between similar-sized individuals also takes place. Biitschli does not 222 THE PROTOZOA consider this an indication of sex-difference, but merely the chance fusion of two Polytomas of different age. Nevertheless, the phenom- enon is significant, and indicates that the smaller or less-grown individual has the capacity, whatever that may be, of conjugation, and may be regarded as an intermediate stage, at least, in the devel- opment of sexually differentiated forms (Fig. 120). A somewhat analogous process takes place in Codosiga botrytis, one of the Choano- flagellida, where, as first observed by Stein, an attached form con- jugates with a free-swimming and somewhat smaller animal. A similar but more definite sex-difference is seen in the peritri- chous Ciliata, where the individuals are of dissimilar size. In all of these, with the exception of the genus Zootkamnium, a normal-sized individual fuses with a smaller one. Engelmann ('76) made the A B c D E Fig. 120. Conjugation of Polytoma uvella Ehr. [DALLINGER and DRYSDALE.] interesting discovery that the larger form, or macrogamete, is always one whose sister-buds have given rise by division to smaller forms, or microgametes, which would certainly suggest that a particular condi- tion of the plasm accompanies conjugation. In the genus Zootham- nium, alone, the macrogametes are considerably larger than the normal individuals (Trembley 1747, Ehrenberg, Greeff, Engelmann, and others). The microgametes, which may arise by budding, as in Lagenophrys ampulla (Fig. 121), or by repeated divisions, as in Epistylis (Y\g. 122), swim about freely until they come in contact with macrogametes, to which they finally adhere. Upon fusing, the microgametes gradually lose their definite structure, until finally they are absorbed. Engelmann ('76), watching the process of sexual union in the ciliate, Vorticella, records the following interesting observations : " The buds, at the beginning, swarmed about with constant and considerable rapidity, rotating the while on their axes, but moving more or less in a straight line through the drop. This lasted from five to ten minutes SEXUAL PHENOMENA IN THE PROTOZOA 223 or even longer without any special occurrence. Then the scene sud- denly changed. Happening all at once in the vicinity of an attached Vorticella, a bud quickly changed its direction with a jerk, and approached the larger form, fluttering about it like a butterfly over a flower, and gliding over its surface here and there as though tasting. After this play, repeated upon several individuals, had gone on for some minutes, the bud finally became firmly attached." Again : " I observed another performance still more remarkable from its physio- logical and particularly from its psycho-physiological significance. A free-swimming bud crossed the path of a large Vorticella which had become free from its stalk in the usual manner and which was roaming about with great activity. At the instant of the meeting there was no trace of a pause the bud suddenly changed its direction and followed the Vorticella with great rapidity. It developed into a regular chase which lasted about five seconds, during which time 5 /-& B Fig. 121. Conjugation in Lagenophrys ampulla St. [BUTSCHLI.] s. Microgamete attached to normal cell (A). B. Fusion of macro- and microgametes. the bud remained about one-fifteenth of a millimeter behind the Vor- ticella, although it did not become attached, for it was lost by a sudden side movement of the larger form. The bud then continued its way as before. These processes are remarkable, since they demonstrate a fine and rapid perception, a rapid and safe will determination, and finely divided motor innervation. They show to what astonishing height and multiplicity physiological differentiation in animals can go, even within a single cell." 1 The phenomena which Engelmann observed and regarded as evidence of psychic activity, have been shown to owe their origin for the most part to chemical and physical stimuli. But the sex-differ- entiation indicated by the diversity in size and activity of the gametes, and the fusion of the two cells which he described, are typical of fer- 1 Loc. cit , p. 583. 224 THE PROTOZOA tilization, or of so-called sexual reproduction, throughout both animal and vegetable kingdoms. 3. The union of swarm-spores (Isogamy and Anisogamy). It is but a short step from the primitive condition described above, where an ordinary individual unites with a smaller one, to the condi- tion where both conjugants are reduced indeed, both conditions may be present in the same organism. Thus the genus Polytoma, as de- scribed above, shows a facultative union between two normal-sized individuals, or between one normal-sized individual and a microgamete, or between similar microgametes. Other members of the group to which Polytoma be- longs the Chlamy- domonadina show similar indefinite- ness, and in no case can it be positively stated that the union between a larger (ovoid) and a smaller (spermatoid) micro- gamete is obliga- tory. Goroschankin ('75 ) maintained that in Chlorogoniu mpul- visculus the sperma- toid microgametes arise by an eight division and con- jugate with the ovoid macrogametes which arise by a two or four division (Fig. 123), but Reinhardt ('76), on the other hand, observed conjugation between microgametes of the smaller size, although he noted a frequent difference in size. In Polytoma, while there is a facultative conjugation between individuals of diverse size, there is also, apparently, a tendency toward an obligatory union of microgametes. The differentiation goes a step further in Phacotus lenticularis, where, according to Carter ('58), an ovoid cell arising by two or four division in the normal manner, unites with a minute form which is the product of a sixty-four division. The obligatory conjugation of microgametes is, however, safely Fig. 122. Epistyhs umbellaria Leeuw. [GREEFF.] Macrogametes (M) and microgametes (m). SEXUAL PHENOMENA IN THE PROTOZOA 225 established in a great many Protozoa, especially among the colonial forms of Mastigophora, and, to a less extent, in Sarcodina and in some Sporozoa. In the Sarcodina, macro- and microgametes are formed by many marine types, including Reticulariida and Radiolaria, and Brady, Brandt, and Haeckel do not hesitate to say that sexual reproduction is brought about by their union. In only one case, how- ever (Hyalopus], has conjugation been actually observed. Here the cell-body spontaneously fragments into isogamous microgametes which swim away from the shell and conjugate (Schaudinn). In the Sporozoa, also, the recent results obtained by Siedlecki ('99) show an D Fig. 123. Conjugation in Chlorogonium euchlorum Ehr. [STEIN.] A. Adult individual. B. Macro- and C. microgamete formation. D. Conjugation. analogous phenomenon, and at the same time they throw considerable doubt upon Wolters's ('91) conclusions that the nuclei of two conju- gating Gregarines unite. 1 According to Siedlecki, two similar in- dividuals of Monocystis ascidice come together and secrete a common cyst within which they sporulate, each individual by itself. There is no union of nuclei as in Actinophrys, nor interchange of parts of nuclei as in the Infusoria, but the nuclei rapidly divide, and the subdivisions ultimately become the nuclei of minute cells resem- bling spores. These have been repeatedly observed in Gregarinida, iCf.p. 157. '226 THE PROTOZOA and by most authors are called sporoblasts. The so-called sporoblasts become motile and move about with considerable freedom, a hitherto unrecognized phenomenon. But more remarkable still, the sporo- blasts finally unite two by two, and after complete fusion of nuclei and cell-plasm, each double cell or copula divides into eight parts, the sporozoites (Fig. 124). These observations, which are the most conclusive that have yet appeared, place the Gregarinida in line with the Reticulariida and Radiolaria, and Siedlecki with Mesnil ('oo) sees in this isogamous union a feature which distinguishes the Gregarinida from the Coccidiida. The obligatory fusion of microgonidia is widely distributed in the colonial forms of Mastigophora, especially in the Phytoflagellida. It is to be regretted that we do not know the full life-history of the Fig. 124. Conjugation of Monocystis acidice Lank. [SIEDLECKI.] The two gregarines unite (A). The nuclei divide repeatedly, and many gametes are formed (B). These unite two by two, forming spores. Each spore divides to form eight sporo- zoites (c). simpler and more indefinite colonies such as Dinobryon, AnthopJiysa, or Synura, where the aggregate arises through continued binary division, for in the higher types, the fertilized egg, as in Metazoa, passes by a regular cleavage into the adult form, the cells becoming separated only in the later stages. Nevertheless, in these more differentiated types some stages are unquestionably more primitive than others. In Gonium pectorals, a colony consisting of sixteen in- dividuals, the colonies reproduce asexually by simultaneous division of all of the cells, four successive longitudinal divisions in each cell resulting in sixteen groups of sixteen cells each, and these groups form independent colonies (O. F. Miiller, Cohn, Stein Fig. I25). 1 1 The camera drawing from a permanent preparation shown in Fig. 125 throws consider- able doubt upon this interpretation of the first three division planes as described by Stein. According to this one preparation the third division is horizontal, giving four cells above and four below. The plate form is assumed in the early sixteen-cell stage. SEXUAL PHENOMENA IN THE PROTOZOA 227 Under certain conditions some of the adult cells become separated from the colony and pass into a resting state, during which they divide into eight biflagellated microgametes, and these, as soon as liberated, conjugate in pairs (Hieronymous, Rostafinski, '75). There is no size differentiation between the conjugating microgametes. Nor is there Fig. 125. Gonium pectorale O. F. M., in division. The third cleavage results in two layers of four cells (i, n, o). The flattening occurs in the !2-i6-cell stage (g, m,p). in the somewhat better-known form, Pandorina morum. In the latter, which is also a sixteen-cell colony, after a certain number of asexual generations, a generation appears in which each of the sixteen cells divides, not into sixteen parts for a new colony, but as in Gonium into eight gametes, which ultimately become free, conjugate, and pass into a resting zygote condition. After a longer or shorter period, either the cyst bursts, and a naked individual emerges, which by division 228 THE PROTOZOA D forms the complete colony ; or the zygote may first divide into two or three individuals, each of which forms a sixteen-cell colony. Pringsheim ('69) stated that a dimorphism exists in the gametes formed by large colonies and by small ones, and maintained that the larger gametes never conjugate amongst themselves, while the smaller ones can unite with each other or with the larger ones (Fig. 126). An incipient sexual difference in the colony is thus indicated, although, as in chlamydomonads, the size difference in gametes is apparently facultative. The sexual difference is somewhat better marked in the genus Endorina, although the most reliable authorities differ as to the details. According to Goroschankin ('76) and Dangeard ('89), the sexually mature colonies are easily distin- guished as male and female, the latter resembling the ordinary colo- nies save for a slightly larger size. The male colonies are at first quite similar to the ordinary colony, but each of the sixteen cells divides to form a six- teen or thirty-two celled plate, and each of these cells gradually becomes long and spindle-formed and de- velops flagella at the pointed end. They ultimately become free and unite with larger ovoid cells. Carter's ('58) description differs so much from this that Butschli doubts if he had the same species. He found that the colonies are hermaphrodite and divided into male and female por- tions. Four cells at one pole of the oval colony develop into sperma- tozoids, while the remaining twenty-eight cells become enlarged, and as ovoid cells, are probably fertilized by the spermatozoids. These observations, although conflicting, at l