The Raccoon As Intermediate Host of Three Sarcocystis Species in Europe

RESEARCH NOTES 145

"usual" range for M. moniliformis. These ob- (Bremser 1811) Travassos 1915 and Moliniformis servations confirm the wide range of morpho- clarki (Ward 1917) Chandler 1921. Journal of Par- logical variations reported for moniliformid asitology 61:996-998. 1975b. Host specificity and lack of hybrid- acanthocephalans. These size variations could be ization of Moliniformis clarki (Ward 1917) Chan- a cause of some of the confusion on the taxo- dler 1921 and Moliniformis moliniformis (Brem- nomic status of M. clarki and M. moniliformis ser 1811) Travassos 1915. Journal of Parasitology in North America. Populations of these 2 acan- 61:991-995. Chandler, A. C. 1921. Notes on the occurrence of thocephalan species exhibit considerable vari- Moliniformis sp. in rats in Texas. Journal of Par- ability, depending on host and geographical dis- asitology 7:179-183. tribution; see Chandler (1921,1941), Van Cleave . 1941. The specific status of Moliniformis (1924, 1953), Petrochenko (1958), and Buckner (Acanthocephala) of Texas rats, and a review of the species of this genus in the western hemisphere. and Nickol (1975a, b). This is the first morpho- Journal of Parasitology 27:241-244. logical study of M clarki from G. bursarius mis- Petrochenko, V. I. 1958. Acanthocephala of domestic souriensis. and wild animals. Vol. 2. Izdatel'stvo Akademii SPECIMENS: Three male and 4 female M. clar- Nauk S.S.S.R. Moscow. (English translation by ki on 7 slides in the University of Nebraska State Israel Program for Scientific Translations Ltd., 1971.478 pp.) Museum, Harold W. Manter Laboratory Coll. Singleton, J., D. J. Richardson, and J. M. Lockhart. 38227. 1993. Severe moniliformiasis (Acanthocephala: HOSTS. Skulls in the Museum of High Plains, Moniliformidae) in a gray squirrel, Sciurus caro- Fort Hays, Kansas Coll. nos. 31075, 31077, linensis, from Arkansas, USA. Journal of Wildlife Diseases 29:165-168. 31108, 31122, 31126, 31135, 31141 (one gopher Van Cleave, H. J. 1924. A critical study of the Acan- was not infected). thocephala described and identified by Joseph Lei- dy. Proceedings of the Academy of Natural Sci- Literature Cited ences of Philadelphia 76:279-334. Amin, O. M. 1985. Classification. Pages 27-72 in D. . 1953. Acanthocephala of North American W. T. Crompton and B. B. Nickol eds. Biology of Mammals. Illinois Biological Monographs 23.179 the Acanthocephala. Cambridge University Press, pp. London. Ward, H. B. 1917. Echinorhynchus moniliformis in Buckner, S. C., and B. B. Nickol. 1975a. Morpho- North America. Journal of Parasitology 3:141- logical variation of Moliniformis moliniformis 182.

J. Helminthol. Soc. Wash. 63(1), 1996, pp. 145-149

Research Note The Raccoon as Intermediate Host of Three Sarcocystis Species in Europe

M. STOLTE, K. ODENTNG, G. WALTER, AND I. BOCKHARDT Institute for Zoo Biology and Wildlife Research, PF 1103, D-10252 Berlin, Germany

ABSTRACT: One out of 12 raccoons from German Zoos Free-ranging raccoons, Procyon lotor L., in was found to possess musculature infected by sarco- North America are known as intermediate hosts cysts of 2 distinct Sarcocystis species (S. sp. 1 and S. sp. 2). The cyst wall of S. sp. 1 had fingerlike, and that of Sarcocystis kirkpatricki Snyder et al., 1990. In of S. sp. 2 hairlike, villar protrusions. Two out of 45 Europe raccoons occur not only in zoos but also raccoons from a free-ranging population in Germany free in certain areas. Since we are unaware of showed infection of the muscle by a third Sarcocystis reports of Sarcocystis spp. in raccoons in Europe, species (S. cf. sebeki, without villar protrusions of the we decided to investigate whether the raccoons cyst wall). None of the 3 species is identical with Sar- cocystis kirkpatricki, described from raccoons in North in North America and Europe are parasitized by America. the same or different Sarcocystis species. KEY WORDS: raccoon, Procyon lotor, Sarcocystis. Seven raccoons originated from the Leipzig

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Figures 1-3. Sarcocystis sp. 1 from a zoo raccoon. Arrowheads point to the wall protrusions. 1. Light microscope photomicrograph of the sarcocyst in longitudinal section. Bar = 10 tan. 2, 3. Transmission electron micrographs of the cyst wall and/or its villar protrusions. 2. Bar = 1.1 jwn. 3. Bar = 0.4 fan.

Zoo, 5 from other East German zoos, and 45 found to be infected with 2 Sarcocystis species, animals were taken from a free-ranging popu- and only 2 of 45 free-ranging raccoons were in- lation of the Buckow region (east of Berlin). For fected by a third species. the history of this population see Grummt (1965), and for the integration into a parasitological re- Sarcocystis sp. 1 search project see Lux and Priemer (1995). Mus- (Figs. 1-3) cle tissues were removed from esophagus and ORIGIN: Procyon lotor, Leipzig Zoo, born in loin of the zoo animals and from tongue, esoph- the zoo. agus, heart, chest, ribs, diaphragm, and thigh of LOCALIZATION: Loin musculature. the wild animals, and examined for the presence DESCRIPTION: A single sarcocyst detected in of sarcocysts by picking them to small pieces by histological sections, 99.5 wide by light micros- means of fine needles and forceps under a dis- copy; cyst wall thick, palisadelike, 4.6-6.9 wide secting microscope at 6.3-fold magnification. For (x = 5.9, SD = 0.5, TV = 50) by light microscopy; histological examination, infected tissues were ground substance plus primary cyst wall 0.48- fixed in 4% formaldehyde, sectioned at 3 to 5 0.78 wide (Jc = 0.62, SD = 0.09, N = 20) by /on, and stained with hematoxylin and eosin. electron microscopy, 0.46-1.20 (x = 0.74, SD = The length of the bradyzoites was determined by 0.23, N = 50) with light microscopy. Fingerlike measuring the more or less bent median line step villar protrusions arise from cyst wall, closely by step from pole to pole, and the width was packed, 5.8-7.4 long (Jc = 6.1, SD = 0.4, TV = measured at the widest diameter. For transmis- 50) by light microscopy, with basal width of 1.12- sion electron microscopy (TEM), portions of 1.38 (Jc = 1.23, SD = 0.10, N = 5) in ultrathin muscle from the single infected wild animal were sections; core of villar protrusions interwoven fixed according to Pospischil and von Bomhard with microfilaments; surface of protrusions with (1979). In the material from the zoo animal, a small invaginations at short distances; invagi- second Sarcocystis species was detected during nations 0.054-0.072 wide (x = 0.059, SD = 0.005, the histological investigation. Therefore, this N = 10) by electron microscopy; diameter of the species could only be prepared for TEM inves- compartments (created by septal invaginations) tigation according to the re-embedding method near cyst wall 9.2-19.9 (Jc = 14.5, SD = 3.0, TV described by Bergmann and Kinder (1987). For = 20) at light microscopic level; depth in direc- this purpose, the position of the sarcocyst within tion of center of sarcocyst 12.2-41.3 (Jc = 27.2, the musculature was marked on the back of the SD = 6.9, N = 20). slide and the cover glass was removed. The section was first dipped into absolute ethanol, then Sarcocystis sp. 2 into a mixture of ethanol and propylene oxide. (Figs. 4, 5) The section was then covered with a mixture of ORIGIN: Procyon lotor, Leipzig Zoo, born in glycid ether and propylene oxide, and a gelatin the zoo. case filled with glycid ether was put on the marked LOCALIZATION: Wall of the esophagus, loin place. After polymerization for 30 min at 105°C, musculature. the case was broken off and trimmed. Ultrathin DESCRIPTION: Two sarcocysts found, 1.8 and sections were cut and stained with uranyl acetate 1.9 mm long, 118-132 wide in the fresh state, and lead citrate. All measurements are in mi- extracted from muscle fibers; cyst wall thin with crometers unless stated otherwise. Abbrevia- light microscopy; ground substance plus primary tions: Jc = mean value, SD = standard deviation, cyst wall 0.30-0.54 wide (Jc = 0.38, SD = 0.06, TV = number of elements measured. TV = 20) in ultrathin sections; numerous, pre- Of the 12 raccoons from zoos, only one was dominantly diagonal and cross-cuts of hairlike

Figures 4, 5. Sarcocystis sp. 2 from a zoo raccoon. Transmission electron micrographs of the cyst wall. G = ground substance. Arrowheads point to the cuts of the hairlike villar protrusions. 4. Protrusions cut mainly crosswise. Bar = 0.4 /im. 5. Protrusions cut mainly diagonally. Bar = 0.6 jim. Figures 6,7. Sarcocystis cf. sebeki from a free-ranging raccoon. Transmission electron micrographs. 6. Section of the interior of a sarcocyst with bradyzoites and a septum (arrowhead). Bar = 1.7 jim. 7. Section of the cyst wall consisting of the ground substance (G) and the membrane of the parasitophorous vacuole plus underlying osmiophilic layer with small invaginations (arrowhead). Bar = 0.4 fim.

Copyright © 2011, The Helminthological Society of Washington 148 JOURNAL OF THE HELMINTHOLOGICAL SOCIETY OF WASHINGTON, 63(1), JAN 1996 villar protrusions of cyst wall visible by electron to type 10 of the classification by Dubey et al. microscopy; layer of these cut villar protrusions (1989). Some features are recognizable (fingerlike 0.50-1.26 wide (x = 0.89, SD = 0.25, TV = 20); outline of the villar protrusions and microfila- villar protrusions 0.18-0.50 in diameter (x = 0.35, ments in the core), although the TEM pictures SD = 0.08, ./V = 85), about 0.6 at bases; fine are not optimal because of the re-embedding granulation visible in the interior of villar pro- method applied. Thus it appears that the re-em- trusions by electron microscopy; surface of cyst bedding method of Bergmann and Kinder (1987) wall with small invaginations at short distances, allows a specific follow-up check of a paraffin 0.090-0.108 deep (x = 0.101, SD = 0.011, N = section by means of TEM and allows ultrastruc- 10) and 0.06-0.12 wide (x = 0.088, SD = 0.019, tural characterization of the cyst wall at least in N = 10) by electron microscopy; compartments some cases. It is possible to get diagnostically 21.4-51.8 (x = 29.9, SD = 9.5, N = 10) wide usable ultrathin sections by means of this meth- near cyst wall by light microscopy, depth in di- od that, however, cannot be compared with the rection of center 19.5-48.8 (x = 33.7, SD = 8.8, sarcocysts embedded especially for TEM inves- N= 10); bradyzoites 14.0-17.1 long (x = 15, SD tigation in regard to the shrinkage and preser- = 0.9, TV - 30) and 3.4-3.9 wide (x = 3.6, SD = vation state of the tissue (Figs. 2, 3). Sarcocystis 0.19, TV = 30) by light microscopy, squashed out sp. 1 is similar to Sarcocystis cf. hofmanni, a of extracted sarcocyst. species occurring frequently in roe deer (Capreo- lus capreolus) in central Europe (see Sedlaczek Sarcocystis cf. sebeki and Wesemeier, 1995), which is morphologically (Tadros and Laarman, 1976) indistinguishable from Sarcocystis hofmanni (Figs. 6, 7) Odening, Stolte, Walter, and Bockhardt, 1994, ORIGIN: Procyon lotor, from the free-ranging found in the European badger (Meles meles, population in the area of Bucko w (east of Berlin). Mustelidae) in Germany. Therefore, it is possible LOCALIZATION: Muscle of loin, ribs, thorax, that the species usually occurring in roe deer spo- thighs, tongue, diaphragm, and wall of the esoph- radically infects badger and also raccoon. agus. Sarcocystis sp. 2 is morphologically indistin- DESCRIPTION: Sarcocysts 4-18 mm long and guishable from other species with hairlike villar 27.5-112 (in histological sections 60.4-95.5) wide protrusions of the cyst wall (type 7 of the clas- in the fresh state, extracted from muscle fibers; sification by Dubey et al., 1989). From the zoo cyst wall thin and smooth at the light micro- area and its environment, the following species scopic level, with no villar protrusions by elec- would be in consideration: Sarcocystis capreo- tron microscopy, 1.06-1.71 wide (x = 1.3, SD = licanis (roe deer, see Sedlaczek and Wesemeier, 0.17, N = 30); surface of cyst wall with shortly 1995), S. cruzi (cattle), S. arieticanis (sheep), and spaced small invaginations, 0.072-0.126 deep (x S. hircicanis (goats) (see Dubey et al., 1989). It = 0.094, SD = 0.016, TV = 20) and 0.060-0.126 is interesting that again a species from roe deer wide (jc = 0.088, SD = 0.017, N = 20) by electron comes into question among them. microscopy; compartments 6.9-16.2 wide near Sarcocystis sebeki was described from the cyst wall (x = 10.6, SD = 2.9, N = 10) and 8.5- musculature of Apodemus sylvaticus and Mus 14.6 (x = 10.4, SD = 2.2, TV = 10) deep in di- musculus. The definitive host is the tawny owl. rection of center with light microscopy; brady- Tadros and Laarman (1979) found similar sar- zoites 6.6-8.5 long (x = 7.4, SD = 0.3, TV = 50) cocysts in a European weasel and fed them to a and 1.3-1.8 wide (x = 1.5, SD = 0.2, TV = 50) tawny owl, in which a weak infection was ob- by light microscopy, squashed out of extracted tained. Odening et al. (1994) described similar sarcocyst. sarcocysts in European badgers (also mustelids), Sarcocystis kirkpatricki is the only known Sar- designating them as "S. cf. sebeki." Because the cocystis species from raccoons in the U.S.A. (Se- sarcocysts found in wild European raccoons are neviratna et al., 1975; Kirkpatrick et al., 1987). morphologically indistinguishable from those of Its cyst wall is similar to type 11 (or lies between weasel and badger, we use the same designation types 9 and 11) of the classification by Dubey et "S. cf. sebeki." al. (1989) and differs distinctly in form and struc- The occurrence of specific Sarcocystis species ture from all 3 species found by us in European in the raccoon introduced into Europe is scarcely raccoons. imaginable, because stable predator-prey rela- The cyst wall of our Sarcocystis sp. 1 is similar tionships would not last with the raccoon as a

Copyright © 2011, The Helminthological Society of Washington RESEARCH NOTES 149 prey item in the absence of a predator regularly Lux, E., and J. Priemer. 1995. Zur Parasitierung eating it. Thus it is possible that all 3 species wildlebender Waschbaren unter dem Aspekt ihrer found by us in European raccoons refer to "let- nordamerikanischen Herkunft. Verhandlungsber- icht des 37. Internationalen Symposiums iiber die ters delivered to wrong address" which normally Erkrankungen der Zoo- und Wildtiere 37:429-434. occur in other mammals. Voucher specimens are Odening, K., M. Stolte, G. Walter, and I. Bockhardt. available from Institute of Zoo Biology and 1994. The European badger (Carnivora: Mustel- Wildlife Research, PF 1103, D-10252 Berlin, idae) as intermediate host of further three Sarco- cystis species (Sporozoa). Parasite 1:23-30. Germany: Collection of Protozoa, No. kT 687 Pospischil, A., and D. von Bombard. 1979. Der Ein- 60-W 1115/92 (histological sections of loin with satz des Elektronenmikroskops in der Diagnostik S. sp. 1), and No. kT 66/57-W 24 (histological des Veterinarpathologen. Zentralblatt fur Veteri- sections of tongue with S. cf. sebeki). narmedizin A 26:643-651. Sedlaczek, J., and H.-H. Wesemeier. 1995. On the diagnostics and nomenclature of Sarcocystis spe- Literature Cited cies (Sporozoa) in roe deer (Capreolus capreolus). Bergmann, V., and E. Kinder. 1987. Einsatz der Ul- Applied Parasitology 36:73-82. trahistologie in der veterinarmedizinischen Diag- Seneviratna, P., A. G. Edward, and D. L. DeGiusti. nostik—Methoden und Erfahrungen. Archiv fur 1975. Frequency of Sarcocystis spp. in Detroit, Experimentelle Veterinarmedizin 41:598-612. metropolitan area, Michigan. American Journal Dubey, J. P., C. A. Speer, and R. Payer. 1989. Sar- of Veterinary Research 36:337-339. cocystosis of Animals and Man. CRC Press, Boca Snyder, D. E., G. C. Sanderson, M. Toivio-Kinnucan, Raton, Florida. 215 pp. and B. L. Blagburn. 1990. Sarcocystis kirkpa- Grummt, W. 1965. Das Vorkommen des Waschbar- tricki n. sp. (Apicomplexa: Sarcocystidae) in mus- en im Berliner Raum. Beitrage zur Jagd- und Wild- cles of raccoons (Procyon lotor) from Illinois. Jour- forschung4:179-180. nal of Parasitology 76:495-500. Kirkpatrick, C. E., A. N. Namir, J. P. Dubey, and C. Tadros, W., and J. J. Laarman. 1979. Muscular sar- E. Rupprecht. 1987. Sarcocystis in muscles of cosporidiosis in the common European weasel, raccoons (Procyon lotor L.). Journal of Protozo- Mustela nivalis. Zeitschrift fur Parasitenkunde 58: ology 34:445-447. 195-200.

J. Helminthol. Soc. Wash. 63(1), 1996, pp. 149-152

Research Note Helminths of Cetaceans on the Southeastern Coast of Brazil

C. P. Santos,1 K. Rohde,2 R. Ramos,3 A. P. Di Beneditto,3 and L. Capistrano3 1 Dept. Biologia Animal, Univ. Santa Ursula, CP 16086, RJ, 22221-020, Brazil, e-mail: [email protected] 2 Dept. of Zoology, Univ. of New England, Armidale, NSW 2351, Australia, and 3 Projeto Cetaceos, CP 122903, SJ. Barra, 28200-000, Brazil

ABSTRACT: Seventy cetaceans accidentally captured in fecting cetaceans. Lack of parasites in 42 Pontoporia fishing nets in Rio de Janeiro State (Brazil) were dis- blainvillei (Pontoporiidae) within our study area was sected for parasites. Sotalia fluviatilis (Delphinidae) probably related to the age of the hosts and differences harbored Braunina cordiformis, Halocercus bras- in food habits between young and adults. iliensis, and Anisakis typica. Tursiops truncatus (Del- KEY WORDS: marine mammals, Cetacea, parasites, phinidae) was parasitized by Nasitrema sp. and B. cor- Brazil, Braunina cordiformis, Anisakis typica, Halo- diformis. Steno bredanensis (Delphinidae) had only B. cercus brasiliensis, Nasitrema. cordiformis. Sotalia fluviatilis represents a new host record for Braunina cordiformis that is reported for the first time from Brazil. In an attempt to correlate these Despite the long Brazilian coastline (about cetaceans' parasite infections with their food habits, a 8,700 km), the parasite fauna of marine mam- survey was made on fish of 20 species and Loligo san- paulensis (Cephalopoda) from the same area. Only mals in this part of the Neotropical region is Bagre bagre, Macrodon ancylodon, and Nebris microps largely unknown. Previous studies in Brazil are contained Anisakis sp. larvae, a parasite species in- restricted to only 2 reports: Halocercus bras-