eeo Revised eO a tO i tr i ru g a m a A LABORATORY MANUAL OF INVERTEBRATE ZOOLOGY BY OILMAN A. DREW, PH.D. ASSISTANT DIRECTOR OF THE MARINE BIOLOGICAL LABORATORY, WOODS HOLE. MASSACHUSETTS WITH THE AID OF FORMER AND PRESENT MEMBERS OF THE ZOOLOGICAL STAFF OF INSTRUCTORS AT THE MARINE BIOLOGICAL LABORATORY WOODS HOLE, MASS. SECOND EDITION, REVISED PHILADELPHIA AND LONDON W. B. SAUNDERS COMPANY 1913 Copyright, 1907, by W. B. Saunders Company. Revised, reprinted, and recopyrighted July, 1913 Copyright, 1913, by W. B. Saunders Company PRINTED IN AMERICA PRESS O F W. B. SAUNDERS COMPAI PHII.ADEl.PMIA PREFACE TO THE SECOND EDITION THIS edition contains descriptions for the study of a few forms and a rather simple literature list that were not included in the first edition. There has been no attempt to make the literature list at all complete, but it seems desirable to refer students to some of the available papers, for by consulting them in connection with their laboratory work they become acquainted with methods of work and develop the spirit of research that is the beginning of real understanding. Certain text-books should be used freely for reference. These have not been mentioned under the special heads, as they apply to practically all groups. Among these may be men- tioned Parker and Harwell's "Text-book of Zoology," Macmil- lan; Lankester's "A Treatise on Zoology/' Black; Harmer and Shipley's "The Cambridge Natural History," Macmillan; Lang's "Lehrbuch der Vergleichenden Anatomic," Fischer; or the English translation, Macmillan; Korschelt and Heider's "Lehrbuch der Vergleichenden Entwicklungsgeschichte," Fis- cher; or the English translation, Macmillan; and Delage et Herouard's "Traite" de Zoologie Concrete," Schmidt. For the many suggestions and criticisms from zoological friends the author is deeply grateful. To Dr. Lorande L. Woodruff, who has given much attention to the revision of the Protozoa, and to Dr. Winterton C. Curtis and Dr. Caswell Grave, who, with the men associated with them as instructors in the Zoology Course at the Marine Biological Laboratory, have given much attention to corrections and additions through- out the manual, special acknowledgments are due, for they have not only saved the author much labor, but have added mate- rially to the value of the revision. THE AUTHOR. July, 1913. iii PREFACE THE present manual has for its basis a set of laboratory direc- tions prepared by members of the staff of instructors to meet the needs of the class in general zoology at the Marine Biolog- ical Laboratory of Woods Hole, Massachusetts. Those who were associated with me in the preparation of the first notes were Dr. Robert W. Hall, Dr. James H. McGregor, Mr. Robert A. Budington and Dr. Caswell Grave. Other members of the staff who have either aided me in modifying the original notes or who have added others are Dr. Winterton C. Curtis, Dr. D. H. Tennant, Dr. Otto C. Glaser, Dr. Grant Smith, Dr. John H. McClellan and Dr. Lorande L. Woodruff. Each year for the past six years the directions have been changed where experi- ences indicated changes should be made. Probably few instructors will find it desirable for their stu- dents to follow closely all that is given in this manual, but it has seemed better to arrange the matter in a logical order, and in some of the forms to call attention to only the important points of anatomy or adaptation, than to try to make the directions for each form complete in themselves. To make the directions for each form complete would necessarily add much labor for the student and would, by the repetition of well-known facts, tend to blunt some of the new and important points to be gained. The type method of laboratory study has for many years been the prevailing method, but care needs to be exercised to keep students from making everything conform to type, and in lead- ing them to see the wonderful adaptations that fit the different animals for their particular lives. The manual is not intended to lead students to a knowledge of comparative anatomy alone, but to an appreciation of adaptation as well. It has fallen on me year by year to see that desirable changes were made in the directions, and it has finally been my lot to put them into their present form, but much of the credit be- longs to the men who have been associated with me in the instruction work at the Marine Biological Laboratory. THE AUTHOR. CONTENTS. PAOB PROTOZOA 1 RHIZOPODA 3 Amoeba proteus 3 Foraminifera 5 Actinosphaerium or Actinophrys 6 MASTIGOPHORA 7 Euglena 7 Volvox 8 Ceratium 9 Noctiluca 9 SPOROZOA 10 Gregarina 10 INFUSORIA 11 Paramsecium 11 Spirostomum 12 Vorticella 13 Oxytricha 14 Euplotes 15 PORIFERA 17 Grantia 18 CCELENTERATA 22 HYDROZOA 24 Hydra (Fresh-water Polyp) 24 Obelia 26 Parypha 28 Gonionemus 29 Hydrocorallina 31 Siphonophora 31 SCYPHOZOA 32 Aurelia 32 ACTINOZOA 34 Metridium (Sea-Anemone) 34 CTENOPHORA 37 Mnemiopsis 37 PLATYHELMINTHES 39 TtJRBELLARIA 40 Planaria maculata 40 Bdelloura or Syncoslidium 41 TREMATOD A 44 Haematoloechus (Distomum) 44 CESTODA 46 Crossobothrium laciniatum 40 VU1 CONTENTS. PLATYHELMINTHES (Continued). PA.QZ NEMERTINEA 49 Tetrastemma 49 NEMATHELMINTHES. 51 Ascaris 51 Trichina 52 TROCHELMINTHES 54 ROTIFERA 54 Brachionus (A Rotifer) 54 MOLLUSCOIDA 56 POLYZOA 56 Bugula 56 Plumatella 58 BRACHIOPODA 58 Terebratulina 58 ECHINODERMATA 60 ASTEROIDEA 61 Asterias (Starfish) 61 OPHIUROIDEA 67 Ophiura (Serpent-Star) 67 ECHINOIDEA 68 Strongylocentrotus (Sea-Urchin) 68 HOLOTHUROIDEA 74 Thyone (Sea-Cucumber) 74 ANNELIDA 78 CELETOPODA 79 Nereis virens (Clam-Worm) 79 Lumbricus (Earthworm) 82 Autplytus cornutus 88 Lepidonotus squamatus 89 Diopatra cuprea 90 Chaetopterus 90 Amphitrite ornata 91 Cistenides gouldii 92 Clymenella torquata 92 Arenicola cristata 93 Sabella microphthalma 93 Hydroides 94 Spirorbis borealis 94 GEPHYREA 95 Phascolosoma 95 MOLLUSCA 97 LAMELLIBRANCHIATA 99 Venus mercenaria (Quahog) 99 Yoldia limatula 107 Mytilus or Modiola (Mussels) 109 Pecten irradians (Scallop) 110 Ostrea virginiana (Oyster) Ill Solenomya 112 Mya arenaria (Long Clam) 112 Ensis directus (Razor-shell Clam) 113 CONTENTS. IX MOLLUSCA (Continued). PAGE AMPHINEURA 115 Chaetopleura 115 GASTROPODA 116 Fulgur (Sycotypus) 116 CEPHALOPODA 124 Loligo pealii (Squid) 124 ARTHROPODA 133 CRUSTACEA 137 Homarus americanus (Lobster) 137 Callinectes hastatus (Blue Crab) 144 Eupagurus (Hermit Crab) 148 Hippa (Sand Mole) 148 Squilla 149 Mysis 151 Talorchestia (Beach-Flea) 151 Porcellio or Oniscus (Sow-Bug) 152 Caprella 153 Branchipus (Fairy Shrimp) 153 Daphnia 154 Cyclops (Water-Flea) 155 Argulus (Fish-Louse) 156 Lepas (Goose-Barnacle) 156 ARACHNOIDEA 158 Limulus (Horseshoe Crab) 158 Buthus (Scorpion) 159 Epeira (Round-Web Spider) 160 Phoxichilidium 162 M YRIAPODA 163 Lithobius (Centipede, Earwig) 163 Julus (Thousand-legs) 164 INSECTA 164 Acridium (Grasshopper) 164 Apis mellifica (Honey-Bee) 170 CHORDATA 174 UROCHORDA 175 Dolichoglossus (Balanoglossus) 175 Molgula manhattensis 176 Perophora 180 Botryllus 180 Amarcecium (Sea-Pork) 181 Salpa cordiformis 183 ACRANIA 185 Amphioxus lanceolatus 185 NOTES FOR GUIDANCE IN MAKING PERMANENT PREPA- RATIONS 187 GLOSSARY 193 INDEX.. . 207 INVERTEBRATE ZOOLOGY. PROTOZOA. Unicellular Animals. CLASS 1. Rhizopoda. With changeable pseudopodia during adult life. Reproduction by simple division and by spore- formation. Subclass 1. Amoebina. With lobose pseudopodia. (Amoeba, Arcella, Difflugia.) Subclass 2. Foraminifera. With fine branching and anastomosing pseudo- podia. Shells, when present, usually calcare- ous. (Lecythium, Globigerina.) Subclass 3. Heliozoa. Typically spherical in form. The pseudopodia, which radiate from the entire surface of the body, are ray-like, seldom changeable, and usually possess an axial filament. (Actino- phrys, Actinosphserium, Clathrulina.) Subclass 4. Radiolaria. With ray-like pseudopodia, and with a chitinous capsule inclosing the nuclei. The skeleton, when present, is formed of silica or acanthin. All are marine. (Thallassicolla.) CLASS 2. Mastigophora. Motile organs in the form of flagella. Repro- duction by longitudinal division. Colony for- mation is frequent. Subclass 1. Flagellidia. With a definite anterior end on which there are one or more flagella. The members of one order (Choanoflagellida) have one or more collar-like 1 1 2 PROTOZOA. processes about the base of the single flagellum. (Mastigamceba, Trypanosoma, Euglena, Pera- nema, Proterospongia.) Subclass 2. Dinoflagellidia. Usually with two flagella, one encircling and the other directed away from the body. (Peri- dinium, Ceratium.) Subclass 3. Cystoflagellidia. With two flagella, one of which is modified into a "tentacle," while the other is short and con- tained within the gullet. (Noctiluca.) CLASS 3. Sporozoa. Without flagella or cilia in the adult period of the life-cycle. Reproduction is by spore-form- ation. All are endoparasites. Subclass 1. Telosporidia. Sporulation phase of the life-cycle is distinct from and follows the trophic phase. Order 1. Gregarinida. The young stages are intracellular parasites, while the adults are free and motile in the di- gestive tract or body-cavity of the host. Sporulation occurs within a cyst during the free period of the life-cycle. (Gregarina.) Order 2. Coccidiidia. Without a free and motile adult stage. Sporu- lation occurs within a cyst during the intra- cellular period of the life-cycle. (Coccidium.) Order 3. Hsempsporididia. Living chiefly in the blood-corpuscles of verte- brates. In many forms the entire sexual period of the life-cycle takes place in an inter- mediate host, as the mosquito. (Plasmodium.) Subclass 2. Neosporidia. Sporulation takes place during the trophic phase of the life-cycle. Order 1. Myxosporididia. The initial free stage occurs in the tissues or the cavities of the organs of the host. The adult form is amreboid. (Myxidium.) Order 2. Microsporidia. Amoeboid trophozoites. Spores very minute and with but one polar capsule. PROTOZOA. Order 3. Sarcosporidia. The initial stage of the life-cycle occurs in the muscle-cells of vertebrates. (Sarcocystis.) CLASS 4. Infusoria. With motile organs in the form of cilia during all or part of the life-cycle. Nucleus dimorphic (macronucleus and micronucleus). Reproduc- tion is by simple transverse division or by bud- ding. Subclass 1. Ciliata. With cilia throughout the life-history. Order 1. Holotrichida. The cilia are of approximately equal length and thickness and equally distributed over the body. Trichocysts are present. (Prorodon, Paramse- cium.) Order 2. Heterotrichida. With a uniform covering of cilia, together with an "adoral zone" formed of cilia fused into membranelles. (Spirostomum, Stentor, Halte- ria.) Order 3. Hypotrichida. The cilia are limited to the ventral surface of a dorso-ventrally flattened body. Cilia often fused into cirri, membranelles, etc. (Oxy- tricha, Pleurotricha, Euplotes, Stylonychia.) Order 4. Peritrichida. More or less bell-shaped in form. Cilia usually reduced to those constituting the adoral zone. (Vorticella, Zoothamnium, Lichnophora.) Subclass 2. Suctoria. Usually possessing cilia only during the embry- onic stages of the life-history. Tentacles adapted for piercing and sucking are present. (Podophrya, Ephelota, Acineta.) Blochmann: Die Mikroscopische Tierwelt des Siisswassers. Abt. 1. Pro- tozoa, 1895. Biitschli: Protozoa. Bronn's Thierreich, 1889. Calkins: Protozoa, 1901. Protozoology, 1909. Marine Protozoa of Woods Hole. Bui. U. S. Fish. Com., 1901. The Scope of Protozoology. Science, 1911. I Conn: Fresh Water Protozoa of Connecticut. Bui. State Nat. Hist.1 Surv., 1905. 4 PROTOZOA. Doflein: Lehrbuch der Protozoenkunde. 3d Auf., 1911. Edmondson: Protozoa of Iowa. Davenport Acad. Sci., 1906. Hartmann: Praktikum der Protozoologie, 1910. Jennings : Behavior of the Lower Organisms, 1906. : Old Age, Death, and Conjugation in the Light of Work on the Lower Organisms. (Harvey Lectures), Pop. Sci. Mo., 1912. Kent: Manual of the Infusoria, 1881. Lankester: Treatise on Zoology. 1. Protozoa. Leidy: Fresh Water Rhizopods of North America, 1879. Minchin: Protozoa, 1912. Prowazek: Einfiihrung in die Physiologie der Einzellgen (Protozoen), 1910. : Taschenbuch der Mikroskopischen der Protistenuntersuchen, 1907. Stokes: Contribution Toward a History of the Fresh Water Infusoria of the United States. Jour. Trenton Nat. Hist. Soc., 1, 1888. Whipple: Microscopy of Drinking Water, 2d ed., 1910. Woodruff: Observations on the Origin and Sequence of the Protozoan Fauna of Hay Infusions. Jour. Exp. Zool., 12, 1912. RHIZOPODA. AMOEBA PROTEUS. Amoebae are usually easily discernible under the low power of the microscope as irregular, semi-transparent, granular bodies. Find a specimen in the material provided, which is known to con- tain amoebae, and determine the following points: 1. With the high power observe the peculiar method of loco- motion, the constant but slow change in the shape of the body by means of projections, pseudopodia, or "false feet." Make sketches at intervals of one or two minutes to show the changes in the form of the body. 2. Observe the peripheral zone of hyaline protoplasm, the ectoplasm, and compare this with the inner protoplasm, the endo- plasm. Observe in detail the formation of a pseudopodium. Does the endoplasm extend into the pseudopodium? Can you explain how the movement is caused ? 3. Find a clear space which appears and disappears at inter- vals; this is the contractile vacuole. Determine the length of time between successive contractions. Are the intervals regu- lar? When the vacuole contracts what becomes of the con- tents? What is its function? 4. Note the oval or rounded nucleus moving with the flowing endoplasm. What is its structure? THE FOBAMINIFEBA. 5 5. Food materials in process of digestion are readily seen. Of what do they consist? They are contained in gastric vacu- oles. By careful watching, it is often possible to observe the man- ner in which food is ingested and the manner in which the undi- gested matter is egested. Make a careful drawing of an Amoeba. Amoebae of various kinds represent in many respects the simplest type of protozoan, and are therefore placed in the first class of these animals, the Sarcodina. The individuals of this class all possess pseudopodia, but many are quite unlike those of Amceba. Look over the figures of various Rhizopoda. If time and material permit, study Amoeba verrucosa, Arcella, and Difflugia, and compare them with Amceba proteus. Do you understand how the shells of the last two genera are made, and of what service they are? Why are not shells good for all forms? Drawings of these forms are desirable. Calkins: The Fertilization of A. proteus. Biol. Bui., 13, 1907. Bellinger : Locomotion of Amoeba and Allied Forms. Jour. Ex. Zool., 3, 1906. Metcalf : Amoeba Studies. Jour. Ex. Zool., 9, 1910. Mast: Reactions in Amceba to Light. Jour. Ex. Zool., 9, 1910. Popoff : Ueber den Entwicklungs cyclus von A. minuta. Arch. f. Protist., 22, 1911. THE FORAMINIFERA. With very few exceptions Foraminifera are marine and pro- vided with shells. Empty shells from deep-sea dredgings or from the sand beaches of such islands as the Bermudas may be had for study. Examine them with a low power by reflected light. 1. Carefully examine various shells, compare them with each other and with figures. Notice the great variety in size and shape and determine how the chambers must have been added during growth. 2. Observe a single opening in a shell, and determine whether the general surface has any finer perforations. Be sure to under- stand the relation of the live animal to the shell. Make drawings of several types of shells. Farmer: Foraminifera, pp. 133-139, Lankester's Treatise. Flint: Recent Foraminifera. Rep. U. S. Nat. Mus., 1897. PROTOZOA. ACTINOSPHAERIUM OR ACTINOPHRYS. Find, as usual, with the low power, and increase the magni- fication as occasion demands. 1. Note the many fine radiating pseudopodia. These are quite stiff compared with those of Amceba and for a considerable time show little change, not being pushed out and retracted constantly as in Amoeba. Is the animal flat or spherical ? 2. Both ectoplasm and endoplasm are so filled with vacuoles that they present a frothy appearance characteristic of most Heliozoa. The endoplasm of all Protozoa is alveolar in struc- ture, but in Actinosphserium the vacuoles are exceptionally large, though not as large as those in the ectoplasm. In Ac- tinophrys the endoplasm is not so sharply separated from the ectoplasm. 3. The nucleus of Actinophrys is present in the center of the organism, but it is somewhat difficult to demonstrate in the live animal. In Actinosphserium there are many nuclei. 4. At some point near the periphery, the contractile vacuole can usually be seen. When it is found notice its action, and immediately after it has contracted look among the pseudopodia of that region for indications of its extruded contents. Draw a specimen, indicating all of the points observed. 5. When the contractile vacuole discharges, or when any foreign body touches the ends of the pseudopodia, notice the way in which this type of pseudopodium is moved. What does this indicate in regard to its structure? How far do the pseudo- podia extend? They may be seen to contain minute granules when studied with the high power and best light. 6. If possible, observe the process of catching food with the tips of the pseudopodia and the manner in which it is drawn toward the body. Note any motion on the surface of the body as the food is drawn closer, and also the manner in which the food is finally ingested. Are there any indications that the pseudopodia extend as still finer filaments beyond the point to which it is possible to trace them with the highest magnifica- EUGLENA. 7 tion at hand? If the capturing of food is observed, make a series of diagrams to illustrate the process. If possible, observe a specimen undergoing division. Draw. It is desirable to examine Clathrulina, noting the stalk and skeleton. Look over figures. R. Hertwig: Ueber die Kernteilung, Richtungskorperbildung und Befruch- tung bei Actinosphserium. Abt. d. Math. Phys. Kl. d. Ak. d. Wiss., Munchen, 19, 1898. HASTIGOPHORA: EUGLENA. Understand its habitat and with what forms it is usually associated. 1. Observe the free-swimming movements of the organism, and the euglenoid changes in the form of the body. Make drawings showing the changes in the shape of a single in- dividual. 2. Distinguish anterior and posterior ends. Is there any dorso- ventral differentiation? Note the motile organ, the flagel- lum. Where is it attached? What relation does it bear to the gullet? How is it directed during locomotion of the organism. Does it serve any other purpose besides locomotion? (See Doflein, pp. 604, 33 and 207.) 3. The green color of Euglena is due to chlorophyl, and this enables it to live in clear water, being nourished like a typical green plant. (See Minchin, p. 14.) 4. Note the absence of color near the anterior and posterior ends of the organism. Near the anterior end also notice the red pigment spot, or stigma. What is its probable function? 5. Stain a specimen with iodin and look for the nucleus. It is obscured by the chlorophyl. 6. Observe specimens in the resting stage. Make a drawing showing all of the points observed. Look through the stock cultures for other forms of Masti- gophora, such as Trachelomonas, Peranema, Phacus, etc. It is desirable to make drawings of the different forms. 8 PROTOZOA. Klebs: Ueber die Organisation einiger Flagellatengruppen und ihre Be- ziehungen zu Algen und Infusorien. Unt. Bot. Inst. Tubingen, 1, 1883. : Flagellatenstudien. Zeit f. Wise. Zool., 55, 1893. Wager: On the Effect of Gravity upon the Movements and Aggregation of Euglena viridis and Other Micro-organisms. Phil. Trans. Roy. Soc., London, Series B, 201, 1911. VOLVOX. Volvox globator is better for study than V. aurens. It may be distinguished from the latter by the larger size of the colony, the greater number of cells that compose it (about 15,000), the angular shape of the individual cells, and the stout connecting processes of protoplasm, into which chromatophores may enter. Observe the movements of colonies in a watch-glass of water, with the naked eye and with a low power of the microscope. 1. Do the colonies tend to collect toward a particular side of the dish? What reason is there for the reaction? 2. Place a number of colonies on a slide with enough water to allow them to be covered without crushing them. Study first with the low and then with the high power and determine the species. Understand the relation of the individual cells to the colony. (See Doflein, p. 222.) Draw a figure showing several cells and their protoplasmic con- nections. 3. Compare in detail an individual cell with Euglena. 4. Observe, if possible, certain cells, called parthenogonidia, which are specialized for asexual reproduction. These divide and form the daughter colonies, which become detached and swim in the interior of the parent colony. They are finally liberated by the rupture of the wall of the parent colony. Make a figure of a parent colony that incloses several daughter colonies of different sizes. 5. V. globator is monoecious. Look for eggs and bundles of spermatozoa. Figure them. 6. Be sure to recognize the significance of the fact that the cells of Volvox are differentiated into somatic and germ cells, CERATIUM. NOCTILUCA. 9 and to understand the resulting physiological division of labor. (See Calkins, Protozoa, p. 232.) 7. Consider the reasons for and against regarding Volvox and allied organisms as animals rather than plants. Meyer: Ueber den Bau von V. aurens und V. globator. Bot. Cent., 63, 1895. CERATIUM. 1. Examine this form with a high power, and in a favorable specimen notice the sculptured outer surface of the cellulose test. The living animals are green or brown owing to the pres- ence of chromatophores in the protoplasm. 2. Note the furrow encircling the body. Does it extend completely around it? Is there a short furrow on one side at right angles to the first, or a depression of considerable size? Understand the position of the flagella. Draw the animal, showing the points observed. Look for examples of the earlier stages of division, and of later stages, which appear as chains of fully formed individuals attached together. Kofoid: Mutations in Ceratium. Bui. Mus. Comp. Zool., 52, 1909. NOCTILUCA. If living specimens are not to be had for study, material preserved in alcohol, after suitable fixation, can be used. Spec- imens are best examined in a cell-slide under a cover-glass. 1. Observe the nearly globular shape, and on one side a groove from which arises a large flagellum or " tentacle." Is there a deep groove near it ? At the bottom of this groove it is possible to see the mouth in a living specimen. Another smaller flagellum is visible in living specimens inserted at the bottom of the mouth, but in preserving the organism it is usually destroyed. 2. Note the appearance of the preserved protoplasm. The endoplasm appears parenchymatous. At one point a more com- 10 PROTOZOA. pact mass is seen, from which strands appear to radiate. This has been found to contain the nucleus. Noctiluca is phosphorescent, and frequently causes very bril- liant displays. Make a drawing. Calkins: Nuclear Division in Noctiluca. Jour. Morph., 15, 1899. SPOROZOA. GREGARINA. Remove the head and posterior end of a larval or adult meal beetle and pull out the digestive tract with a pair of for- ceps. Place the digestive tract on a slide, split it open length- wise with a sharp scalpel, and then spread it out, with the inner wall exposed, and cover. The operation should be per- formed rapidly to prevent the material from drying. If the beetle is infected, numerous gregarines will be visible under the microscope. Study with low and high powers. 1. Does the animal move? A great number of refractive granules are present in the protoplasm. They are regarded as reserve nourishment. They can be removed with acid. 2. Note that the body is covered with a membrane, and is divided into a dense superficial layer, the ectoplasm, and a cen- tral, more fluid mass, the endoplasm. 3. The endoplasm is separated into two parts by a portion of the ectoplasm. The anterior part is termed the protomerite, and the posterior part the deutomerite. In which is the nucleus situated? 4. Is it possible to distinguish a layer of myonemes just ex- ternal to the endoplasm? 5. Is there another section of the body just anterior to the protomerite? If so, this is the epimerite. 6. Note that occasionally two (or more) individuals are united. These aggregations are termed syzygies. Before reproduction Gregarina throws off the [epimerite, leaves it in the cell-host, and falls into the lumen of the digestive PARAMjECIUM. 11 tract. It then encysts, and the protomerite and the deutome- rite form one spore-producing individual. The attached stage in the life-history of Gregarina is termed the cephalont, and the detached stage, the sporont. (See Calkins' Protozoa, Fig. 77.) Make a drawing. Berndt : Beitrage zu Kenntnis der im Darme der Larve von Tenebrio moli- tor lebenden Gregarinen. Arch f. Protist., 1, 1902. Minchin: Sporozoa, pp. 177-179, Lankester's Treatise. INFUSORIA. PARAMAECIUM. Place a drop of the culture on a slide, cover, and examine with the low power. 1. In an animal not closely confined note the shape and movements. Is it possible to distinguish an anterior and a posterior end? A forward and backward movement? Is one side of the animal kept constantly uppermost ? Is there a dorsal and ventral surface? Do the animals change their shape either permanently or temporarily? Individuals tend to collect about air-bubbles and at the edge of the cover-glass. Why? Indicate by a sketch all the points which can be determined with the low power. 2. Draw off all superfluous - water by means of filter-paper, add a trace of powdered carmine, and then find a specimen which is narrowly confined and examine it with the high power. The particles of carmine are taken into the body. Deter- mine how and where. Note that the carmine collects in gastric vacuoks. What do you think is probably the nature of the fluid in the vacuoles? In watching them do you notice any definite movement of the protoplasm? Try to see the undi- gested material ejected. 3. Determine the arrangement of the cilia, and the nature of their motion. Is there a reversal of the direction of the stroke, etc.?1 1 It is possible to decrease the rate of movement of both animal and cilia by placing it in a solution of gum arabic. Specimens so treated remain alive for some time. 12 PROTOZOA. 4. Observe the contractile vacuoles. How many are there? Is their position constant? What is their action? In com- pressed specimens the contractile vacuoles and their reservoirs are usually conspicuous. Note the order of appearance and disappearance of the vacuoles and reservoirs. 5. Focus carefully on the margin of the body and note a very thin outer cuticle. A thick layer, the ectoplasm, devoid of gran- ules but containing radially arranged, minute, oval bodies, the trichocysts, is just internal to the cuticle. The inner mass of protoplasm, containing the contractile and gastric vacuoles, and small granules, is the endoplasm. 6. If possible distinguish the clear, centrally located nucleus (macronucleus'). Make a sketch shouting all of the above points. 7. Kill the animal by running a drop of methyl-green under the cover-glass. What happens to the cilia? To the trichocysts? Sketch the trichocysts with the threads protruded, and also note and sketch the macronucleus and the micronucleus. 8. Observe, if possible, animals dividing and conjugating. 9. Study demonstrations of permanently stained specimens for finer structure. Calkins and Cull: Conjugation of P. caudatum. Arch. f. Protist., 10, 1907. Jennings and Hargitt: Characteristics of the Diverse Races of Paramsecium. Jour. Morph., 21, 1910. Metalinkow: Contributions & l'e"tude de la digestion. Arch d. Zool. Exp. et Gen., 9, 1912. Schaeffer: Selection of Food in Stentor cseruleus. Jour. Ex. Zool., 8, 1909. Woodruff: Paramsecium aurelia and Paramsecium caudatum. Jour. Morph., 21, 1910. : A Five Year (3000 generations) Pedigreed Race of Paramsecium without Conjugation. Proc. Soc. Ex. Biol. and Med., 1912 (also Biol. Centr., 33, 1913). SPIROSTOMUM. 1. Compare Spirostomum with Paramsecium, noting the method of locomotion, the shape of the body, the ciliation, the buccal groove and mouth, and the large excretory reservoir, fill- ing the posterior end of the body and in communication with the anterior end of the body by a canal. VOBTICELLA. 13 2. Note the highly refractive, long, band-like (moniliform) macronucleus. In another species of Spirostomum the macro- nucleus is similar to that of Paramsecium. It is desirable to examine stained specimens of the two species of Spirostomum. 3. Note the sudden contractions of the body. When these occur spiral lines appear on the surface. Can you distinguish these lines when the animal is extended? These are primitive structures (myonemes) functioning as muscles. Make a drawing of the extended animal and a diagram show- ing the form when contracted. (See Doflein, p. 968.) VORTICELLA. Place a number of individuals on a slide and cover loosely to avoid crushing. As usual, study first with the low power and then with the high. 1. Notice that the body of Vorticella has the general shape of an inverted bell. The covering of the body is a very thin transparent layer, the cuticle, underneath which is the periphe- ral layer of ectoplasm enveloping the more fluid and granular endoplasm. 2. The peristome is the rounded rim about the base of the bell. 3. The elevated and inclined area included within the peri- stome, and ciliated around the edge, is the disk. It is some- what convex. 4. The marked depression between the disk and the peri- stome is the vestibule. It is also lined with cilia. The vestibule defines the ventral surface of the animal. 5. The gullet, a slender canal, leads from the vestibule toward the center of the body. 6. The anus occurs at the side of the vestibule. It is a tem- porary opening from which the undigested products are passed into the vestibule and so to the exterior. 7. Within the endoplasm are situated the clear contractile vacuole, several gastric vacuoles, the long U-shaped macronucleus, and the small round micronucleus. The macronucleus may be made more distinct by treating with methyl-green. 14 PROTOZOA. 8. The stalk is composed of a sheath, which is continuous with the cuticle of the body, and, within the sheath, the contractile axis or myoneme, which is continuous with the body ectoplasm. Notice that this myoneme is situated within the sheath hi a very loose spiral, and that the stalk quickly contracts into a close spiral when the animal is stimulated. Observe also the manner in which the peristome folds over simultaneously with the contraction of the stalk. What purpose does the contrac- tion of the stalk serve? Vorticella is distinguished from its allied genera by its sim- ple unbranched stalk and also by the spiral form assumed by the contracted stalk. In which order of the Ciliata does the cilia- tion of Vorticella place it? Compare with Zoothamnium. Make a drawing of an expanded individual and a sketch to show the condition when contracted. (See Doflein, Fig. 816, p. 867.) 9. Study, by means of finely powdered carmine, the vortex currents set up by the cilia. Note how the particles are collected in the gullet, and at intervals are forced in rounded masses into the endoplasm to form gastric vacuoles. Is there a definite circulation in the endoplasm? 10. Endeavor to find several stages of reproduction by divi- sion. Large fresh-water species of Vorticella are preferable for study, but marine species may be substitute^! when necessary. If time and material permit, study Lichnophora, a marine peri- trichous form parasitic on Crepidula. (See Calkins' Protozoa, p. 203.) Schroder: Beitrage zur Kenntnis von V. monilata. Arch f. Protist., 7, 1906. OXYTRICHA. Infusoria belonging to the genus Oxytricha, or the genera Stylonychia, Pleurotricha, Euplotes, etc. (see Doflein, Fig. 136, p. 138), may be used for the following study. These forms belong to the order Hypotrichida. Hypotrichous forms are EUPLOTES. 15 among the most highly organized of the class Infusoria, as well as of the entire phylum of Protozoa, and present a complexity of structure and function which is not found probably within the limits of a single cell elsewhere in the animal series. 1. In an animal which is becoming quiet, note the mode of locomotion, the shape of the body, the buccal groove, the con- tractile vacuole, etc., as in other forms studied. Compare the dilation with that of other forms. Refer to Calkins' Protozoa, Fig. 98, and understand the relation of cirri, membranelles, etc., to cilia. Draw, showing the structure in detail. 2. Run some methyl-green under the cover-glass. What is the shape of the macronucleus? The shape varies considera- bly in the different genera. Is it possible to distinguish the micronudeus? 3. Prepare a fresh slide and observe in detail the character- istic movements and manner of creeping over various objects. As the animal turns sidewise, note the marked dorso-ventral compression of the body. Represent this diagrammatically beside the previous drawing. It is desirable to examine permanently stained preparations for division stages, finer details of the nuclei, etc. Wallengren: Zur Kenntnis des Neubildungs und Resorptionsprocess bei den Teilung der Hypotrichen Infusorien. Zool. Jahrb., 15, 1901. EtfpLoTik. \ 0