358 J. PROTOZOOL.20(3), 358-362 (1973). Coccidia of Brazilian Snakes: Isospora deciPiens, Eimeria micruri, E. liophi and E. leimadoPhi Spp. n., with Redescriptions of Caryospora brasiliensis Carini, 1932 and Eimeria poecilogyri Carini, 1933* RALPH LAINSON and JEFFREY J. SHA Wt The Wellcome Parasitology Unit, Instituto Evandro Chagas, Caixa Postal 3, Belém 66.000, Pará State, Brazil 'SYNOPSIS. Caryospora brasiliensis Carini, 1932 is redescribed in the snake Philodryas olfersi, from Barcarena, Pará State, north Brazil, with some additional observations on the endogenous stages. lsospora decipiens sp. n., is described from the same snake: it develops to maturity in the subepithelial tissues of the small intestine. The oocyst wall is extremely delicate, and only free sporocysts could be found in the feces. Diagnosis depended on histologic sections. Oocysts are esti- mated at -18 X 9 IJm: there is no micropyle, oocyst residuum, or polar bodies. The sporocysts are oval and average 10.03 X 8;15 IJm; they have no Stieda body, and the residuum is formed of only a few granules. It is suggested that some previous reports of Cryptosporidium in reptiles mar have been based on the separated sporocysts of Isospora species, and that histologic confirmation is required to establish their identity. Eimeria micruri sp. n. was found in the cQral snake, Micrurus filiformis, Eram Belém, Pará. Oocysts are of an asymmetrical egg-shape, average 20.20 X 14.90 IJm, and have a very thin, colorless wall of 1 layer. There is no micropyle, oocyst residuum, or polar bodies. Sporocysts are ellipsoid with a small Stieda body, and average 12.80 X 6.60 IJm; there is a small sporocyst residuum. Eimeria liophi sp. n. is recorded in tlie colubrid snake Liophis cobella, also from Belém. The oocysts are spherical to subspherical, averaging 14.53 X 13.83 IJm; there is no micropyle, oocyst residuum or polar bodies. Sporocysts are ellipsoid and average 10.20 X 5.50 IJm. They have a delicate Stieda body and the sporocyst residuum is composed of fine scattered granules. A redescription is made of Eimeria poecilogyri Carini, 1933 Eram the snake Leimadophis poecilogyrus Eram Serra dos Carajás, Pará. An addi- tional new species, Eimeria leimadophi, also is reported Eram the same snake. Oocysts are spherical to subspherical, averaging 17.09 X 16.64 IJm, with a colorless wall of a single layer -0.5 IJm thick. There is no micropyle, and the oocyst has a large, spherical residuum. Sporocysts are pear-shaped, with a prominent Stieda body, and average 12.10 X 5.20 IJm; the sporocyst residuum is composed of a few small granules. lndex Key Words: Caryospora brasiliensis; Isospora decipiens sp. n.; sporulated oocysts, endogenous stages; Eimeriamicruri sp. n.; Eimeria liophi ~p. n.; Eimeria leimadophi sp. n.; Eimeria poecilogyri; Brazilian snakes, from; sporulated oocysts; taxonomy; Cryptosporidium, confusion with isosporan sporocysts. A SPECIMEN of Philodryas olfersi (Lichtenstein) (Colu- parasite were measured,using an ocular micrometer, X 8 eye- bridae) was obtained, alive, from Barcarena, Pará State, piece,and X iOO Neofluar objective. North Brazil, and examined for coccidia. Fecal samples showed When possible, the animaIs were killed and the entire gut 2 distinctly different parasites which we consider to represent removed into a dish of 0.9% (w/v) aqueous solution of NaCI Caryospora brasiliensis Carini, 1932 arid a new species of (saline). Scrapings ;lnd squash preparations of intestinal epi- Isospora. thelium were then examined at intervals along the gut to Three specimensof Leimadophis poeéilogyrus (Weid) (Colu- locate the sites of endogenous development; a similar check bridae) from Serra dos Carajás, Pará State, alI were found was made for parasites in the biIe duct epithelium and gall to be passing abundant oocysts of 2 types. One is identified bIadder. These fresh preparations were se~rched using dark as Eimeria poecilogyri Carini, 1933, and the other as a new phase-contrastoptics. Portions of tissues for histologic study speciesof Eimeria. were fixed in 10% (v/v) solution of formalin in saline, ór Two other previously undescribed species of Eimeria were Camoy's fluid, and sections stained with hematoxylin and encouniéred in single examples of Micrurus filiformis (Günther) eosin. Drawings of the endogenous stages are composites ~f (Elapidae) and Liophis cobella (Linnaeus) (Colubridae), from numerous sections examined. Where possible they have been arranged in sequence of development. AlI measurements are Belém, Pará State. in p.m. MA TERIALS AND METHODS RESULTS AND DISCUSSION The snakeswere kept alive in clean glass tanks, the bottoms Caryosporabrasiliensis Carini, 1932 of which had been moistened to prevent the drying of any (Figs. 1-13) feces passed. Fecal material was triturated gently in 2.0% (wjv) aqueous K2Cr207 solution and placed, in thin layers, Diagnosis in covered petri dishes. kept at roam temperature (22-26 C). Oocysts.-The oocysts are passed in the undifferentiated The oocysts were in alI cases relatively abundant, and there stage but, unfortunately, the sporulation time was not de- was no need to concentrate them. Fifty specimens of each termined. There is no significant difference in the size of the undifferentiated and fully developed oocysts. They are sph~rical * This study was made undet the auspices of the Wellcome to oval, averaging 21.7 (19-24) X 20.8 (18-24.). The oocyst Trust, London, ~nd the Instituto Evandro Chagas, Belém, Pará, wall, 1-2 thick, is smooth and colorless. It appears to be com- Brazil. posed of 3 layers, the middIe one of which is prominently * We thank Dr. William E. Duellman, Curat~r, Division of Herpetology, Museum of Natural History, University of Kansas, striated. Within the undeveIoped oocyst the sporont measures for identification of the snakes. -15 in dia meter, and it becomes oval before formation of the COCCillIA OF BRAZILIAN SNAKES 359 .. 1°f'm Figs. 1-4. [Caryospora brasiliensis Carini, from the snake Philodryas olfersi. Oocysts sporulated in 10% (w/v) aqueous K.Cr.O7 at 22-26 C.] 1. Newly passed, undifferentiated oocyst. 2. Oocyst with oval sporont, just before sporocyst formation. 3, 4. Mature oocysts in lengthwise and end-on views. single sporocysto The mature oocyst has no oocyst residuum, gametocyte development. The type of microgametocyte seen micropyle, or polar bodieso The sporocyst is oval, with 1 end here, in Caryospora brasiliensis,he refers to as "monocentric," drawn out into a short neck with a conspicuous, caplike Stieda there being only a single cytoplasmic residual body after the body. Average measurements for the sporocyst were 16.12 (16- shedding of the gametes from the surface; many species of 19) X 12.19 (12-18). The 8 sporozoites usually are arranged Eimeria have such microgametocytes. The other type is re- in a circle, surrounding a bulky central sporocyst residuumo ferred to as "polycentric," where cytomeres or microgametoblasts This is clearly seen in end-on views of the sporocysts, but the (16) form separate centers of gamete production and leave arrangement is less evident in other planes. several residual bodies; this is seen in other species of Eimeria Endogenous stages.-As shown by Carini (2), schizogony and is particularly pronounced in Isospora felis Wenyon (18). and gametogony of Caryospora brasiliensis takes place in the The type of microgametogony would thus arrear to be of epithelial cells of the small intestine and, in the present study, taxonomic value only at species leveI. In this respect it is there was no evidence of involvement of other tissues; both interesting to note that Kheisin (8) quotes Reichenow as de- sexual and asexual stages develop on the lumen side of the scribing "polycentric" microgametocytes for species of Caryo- host-cell nucleuso Only 2 schizonts were seen. One measured spora, whereas those figured here for C. brasiliensisare clearly 605 in diameter and had 8 nuclei (Figo 9); the other, algo im- of the "monocentric" formo mature, measured 800 and contained -16 chromatin pieces Carini (2) first described C. brasiliensis in the snake Philo- (Figo 10). No merozoites were seen in either fresh or fixed dryas aestivus Duméril Bibron and Duméril (syn. Chlorosoma material. aestivum). Later (4) he mentioned seeing the same parasite in The smallest macrogametocyte recognizable measured 4.5 X LeimadoPhis poecilogyrus schotti (.Schleger) (syn. Philodryas 205 (Figo 5), with a nucleus containing a large nucleolus. With schotti), Philodryas nattareri Steíndachner,and P. olfersi. Caryo- growth, a few dark-staining granules arrear in the cytoplasm spora brasiliensis ís distinguished from Caryospora jararacae (Figo 6) and later (Fig. 7) there appears a peripheral layer of Cariní, 1939 (4) by its much larger size, the latter measuring smaller granules, with additional larger and col~rless globules only 13-14. Anderson et aI. (1) described Caryospora lampropel- (vacuoles ?) scattered in a more random mannero Closer to tis Anderson, Duszynski & Marquardt in the north Amerícan maturity, the small dark granules enlarge, or merge, to form kingsnake, Lampropeltis C. calligaster (Harlan), and differen- a peripheral layer of intensely eosinophilic globules which mar tiated it from C. brasiliensisby differences in structure and dis- be up to 2 in diameter (Figo 8). These inclusions clearly position of the sporozoites,and wide geographíc separation. One correspond with the "chromatoid" and "plastinoid" granules difference they díd not stress is the presence of a prominent of earlier workers, which generally are regarded as playing a polar body ín the oocysts of C. lampropeltis whích was not de- part in the formation of the oocyst wall. Kheisin (8) has scríbedin C. brasiliensis. discussed them a"t some length in his section dealing with the Type host.-Philodryas aestivus Duméril, Bibron & Duméril. cytochemistry of the developing macrogametocyte, and it is Additional hosts.-Philodryas olfersi (Líchtensteín) j P. nat- inappropriate to dwell further on this complex subject here, tareri Steindachner; and LeimadoPhis poecilogyrus schotti except to say that the disposition and appearance of the (Colubridae).