Thomas C. Orihel

Parasites in Human Tissues

Since there are myriad books in the field of parasitology and tropical medicine, the question one is likely to ask when first confronted with the title of this book is whether it is yet another parasitology text. Parasites in Human Tissues is neither a general parasitology text nor a tropical medicine text. For example, in the malaria selection the authors describe the malarial parasite within vessels and in tissues such as the liver. If one is in need of aid in the identification of the blood stages of the various malarial parasites, one should seek out standard parasitology texts or atlases. Similarly, this volume is not a reference for the morphology of intestinal protozoa or helminths. There is a companion Atlas of Parasitology that can be used for such purposes. Parasites in Human Tissues, however, is a superb reference text for the histological diagnosis of parasites. The book is divided into the familiar, albeit arbitrary, divisions that have become part of any parasitology text, namely, protozoa, nematodes, trematodes, and arthropods; in addition there is a valuable section on artifacts. Series of helpful colorful charts in the beginning of the book assist the reader in understanding both the usual and the unusual locations of parasites within host tissues. For each parasite, there is a short summary of the biology and life

Morphology of the larval stages of Dirofilaria immitis in the dog.

filaria conjulnctivae (Addario, 1885). It is assumed that they are zoonotic forms. Little is known of the development of filarial species in the definitive host, the larval and adolescent stages having been described only for Conispiculum flavescens in lizards (Menon, Ramamurti, and Rao, 1944), Litomosoides carinii in cotton rats (Cross and Scott, 1945, 1947; Scott, 1945; Scott and Cross, 1946; Scott et al, 1951) and Dipetalonema blanci in rodents (Chabaud, 1954). Edeson and Buckley (1959) described the rate of growth and the migration of Wuchereria malayi in experimentally-infected cats.

NECATOR AMERICANUS INFECTION IN PRIMATES

Laboratory-bred and feral chimpanzees (Pan troglodytes) and young, laboratory-bred, patas monkeys (Erythrocebus patas) exposed to infective larvae of Necator americanus cultured from human feces developed patent infections in 42 to 54 days; adult, feral patas monkeys were refractory to infec- tion. Patent infections persisted for 16 months in the chimpanzee and for 10 mos. in the monkey with- out any indication of waning. Variation in egg output among animals exposed to the same number of larvae suggested that there was considerable variation in individual susceptibility. It was also evident that large numbers of worms, in some cases, will produce severe anemia in the host.

GASTRIC PAPILLOMATA IN MACACA SPECIOSA INDUCED BY NOCHTIA NOCHTI (NEMATODA: TRICHOSTRONGYLOIDEA)*

Papillomata in association with the trichostrongyle, Nochtia nochti Travassos and Vogelsang, 1929 were encountered in the stomachs of 16 of 48 stumptail macaques (Macaca speciosa) from Thailand examined at necropsy. The gross and histological features of the papillomata are described. The tumors were found in the prepyloric region of the stomach, showed no evidence of malignancy, and appeared to represent a reaction of the gastric mucosa to the presence of either the worms or their eggs. The tumors appear to be more common in male monkeys than in females. In experiments on the toxicity of pesticides in stumptail macaques (Macaca speciosa) im- ported from Thailand, incidental papillomata in association with a parasitic nematode were encountered in the stomachs of a large propor- tion of the animals examined at autopsy. Of 48 monkeys examined (24 males, 24 females), 16 (33%) possessed gastric papillomata (13 males, 3 females). Gross findings: The tumors were situated in

DEVELOPMENT AND LARVAL MORPHOLOGY OF LOA LOA IN EXPERIMENTAL PRIMATE HOSTS

Experimental infections with Loa loa were established in laboratory primates by inoculation of third-stage larvae recovered from either Chrysops silacea infected after engorging on a human volunteer with loiasis, or Chrysops atlanticus infected after blood feeding on experimentally infected primates. The third molt occurred at 16 to 20 days postinoculation of infective larvae, and the fourth at about 50 days. Early larval development was characterized by rapid growth and marked differentiation of the reproductive systems. Differences in the bodys growth rate between the sexes were seen as early as 30 days and was especially marked by 60 days. Females grew more rapidly, and for a longer time than males. Males reach maximum size by 120 days, females not until about 300 days. Female worms were inseminated prior to day 90 developing eggs filled the uteri by 120 days, and microfilariae appeared in the peripheral blood at approximately 150 days. Morphological features of the larval and adult stages of human Loa loa are described.

Anatrichosomiasis in African monkeys.

Approximately 29% of 298 African monkeys belonging to 5 species (Erythrocebus patas, Cercopithecus talapoin, C. aethiops, Cercocebus galeritus, and Papio sp.) obtained from various parts of Africa were found to be infected with species of Anatrichosoma. The adult worms were found in the nasal epithelium. The embryonated eggs which accumulate on the epithelial surface were easily removed with a cotton-tipped applicator stick, nasopharyngeal swab, or similar implement. Eggs were rarely found in the feces by routine diagnostic methods. Swabbing of the nasal mucosa was found to be the most reliable and accurate procedure for diagnosis of infections in living animals. Although specific determinations were not made, it appeared that the parasites from the African monkeys were different from A. cutaneum and A. cynamolgi described previously from Asian monkeys. The anatrichosomes are relatively rare and poorly known trichuroids which, to date, have been found in the skin and nasal mucosa of Asian monkeys (Swift et al., 1922; Smith and Chitwood, 1954; Chitwood and Smith, 1958; Allen, 1960) and on two occasions in man (Morishita and Tani, 1960; Le-Van-Hoa et al., 1963). Infections have been recognized more frequently as a result of recovery of adult worms at necropsy than from finding the eggs of the parasite. The latter have been found in scrapings of the nasal mucosa and occasionally in feces (Allen, 1960; Reardon and Rininger, 1968). Reports concerned with this parasite are so few that our knowledge of the species, their host range, and geographical distribution is very limited. Recently, in the course of histological examination of nasal tissues from an African patas monkey (Erythrocebus patas), Dr. Herman Seibold of the Pathology Laboratories, Delta Regional Primate Research Center, observed sections of a nematode in the mucosa and referred the slides to the author for identification. Examination of the sections of worms revealed that they were trichuroids. Their location and the morphology of the fully embryonated eggs found in the tissues indicated that they were a species of Anatrichosoma. Examination of intact worms subsequently recovered from animals at necropsy confirmed the tentative diagReceived for publication 14 January 1970. * Presented at the Forty-fourth Annual Meeting of the American Society of Parasitologists in Washington, D. C. 3-7 November 1969. This investigation was supported by U. S. Public Health Service grants FR 00164, DRFR and Al 06828, NIAID, NIH. nosis. Because there are no existing reports of anatrichosomes in African primates and because routine stool examinations in the past have failed to demonstrate the presence of the parasite in colony animals except rarely, studies were undertaken to ascertain the prevalence of this parasite in African primates and to determine the most suitable method for its diagnosis. MATERIALS AND METHODS The monkeys utilized in the study were maintained in the Delta Primate Centers animal colony or, in some cases, at other institutions. The nasal passages of each animal were swabbed with a dampened, cotton-tipped applicator stick or commercially prepared, sterile, nasopharyngeal swab (Calgiswab?), or rectal swab (SWUBE?), depending on the size of the nostrils. The swabs were examined under a binocular dissecting microscope with incident light to determine whether or not eggs were present. Stool examinations were made by direct-smear, formalin-ether, and zinc sulfate flotation techniques. At necropsy, portions of the nose and upper lip were removed from infected monkeys and fixed in Bouins solution or 10% formalin. These tissues were processed by routine methods, sectioned at 7 ,i, and stained with hematoxylin and eosin. Worms removed from the tissues in toto were fixed in glacial acetic acid, transferred to 70% alcohol to which glycerin was added gradually, and were brought to pure glycerin by evaporation for microscopic study. RESULTS AND DISCUSSION Histological examination of tissues from infected animals revealed that adult worms usually are located in that portion of the nasal cavity lined with stratified squamous epithelium (Fig. 1). The female worms make extensive burrows in the epithelium as they wander through the tissues, depositing eggs in their wake. The eggs remain in these burrows or

Dirofilaria corynodes (von Linstow, 1899 ): morphology and life history.

Dirofilaria corynodes (von Linstow, 1899) was found in 12 of 211 patas monkeys, Erythrocebus patas, imported from northern Nigeria. The microfilariae are unsheathed and circulate in the peripheral blood with a marked nocturnal periodicity. Microfilariae develop to the third-stage in the fat body of Aedes aegypti in about 14 days when the latter are maintained at a temperature of 28 C and 60% to 70% relative humidity. In the course of their development in the mosquito, the larvae molt twice. The first molt occurs on about the 9th day and the second on approximately the 14th day of development. Patas and rhesus monkeys were infected in the laboratory by the injection of third-stage larvae and by surgical implantation of adolescent worms. Prepatent development required approximately 280 days. The adult worms in histological sections displayed the characteristic morphological features of Dirofilaria (Nochtiella) spp. In the course of routine blood surveys undertaken to determine the prevalence of filariasis in monkeys admitted to the Delta Primate Centers primate colony, a species of microfilaria was found in 12 of 211 patas monkeys (Erythrocebus patas) from northern Nigeria. Necropsy examinations of infected animals which died of other causes revealed a species of Dirofilaria in the subcutaneous tissues of each. Morphological and biological studies were undertaken to determine the specific identity of the parasite and to evaluate its potential usefulness as a model system for projected studies. The report that follows presents a brief description of the adult worms and the microfilaria as well as observations on the development of the parasite in intermediate and definitive hosts in the laboratory.

Dirofilaria lutrae sp. n. (Nematoda:Filarioidea) from otters in the southeast United States.

Dirofilaria lutrae sp. n. is described from the subcutaneous tissues of the American otter in Louisiana and Florida. The species is characterized by long and slender form, absence of longitudinal cuticular ridges (except on the tail of the male), extremely large left and relatively short right spicules, and an unusually large, unsheathed microfilaria. Morphologically, the species is most similar to Dirofilaria incrassata which parasitizes certain Central and South American sloths. In the course of a survey of filariae of animals in southern Louisiana in 1959 and 1960, a species of Dirofilaria producing a very large, unsheathed microfilaria was found in the subcutaneous tissues of the otter, Lutra canadensis texensis Goldman. Similar surveys were undertaken in south Florida in 1960-61 and the same species of Dirofilaria was recovered from the otter, Lutra canadensis vaga Bangs, in Dade and Collier counties. Preliminary studies indicated that the species was morphologically distinct from others of the genus. Additional specimens from the same host were obtained from Dr. J. F. Schacher who made collections at the Rockefeller Refuge in Louisiana and from Dr. W. H. Leigh whose material was collected in the Everglades region of Florida. Dirofilaria spp. have not previously been reported from the otter in North America. However, two species have been found in the South American otter, Pteronura brasiliensis. Vogelsang (1940) reported D. immitis in the heart and pulmonary vessels of an otter in Venezuela. Freitas and Lent (1949) described a new species, D. spectans, from the heart and pulmonary arteries of otters in Brazil. The species description presented in this report is based on the examination of specimens recovered from an otter captured in Terrebonne Parish, Louisiana. Measurements are in millimeters unless otherwise indicated. Range is followed by average in parentheses. Received for publication 29 May 1964. * Supported by Grant EF-10,217 from NIH, U. S. Public Health Service. t Present address: Delta Regional Primate Research Center, Covington, Louisiana. Dirofilaria lutrae sp. n. (Figs. 1-9) Description Onchocercidae (Leiper, 1911); Dirofilariinae (Sandground, 1921); Dirofilaria Railliet and Henry, 1911. Body long, slender, uniform in diameter except at the tapered extremities. Cuticle finely striated transversely; longitudinal ridges lacking except on ventral surface of male tail. Oral opening small, circular (Fig. 6), surrounded by four pairs of submedian papillae in addition to a pair of amphids. Esophagus distinctly divided into muscular and glandular portions (Figs. 1, 7, 8). Microfilariae circulate in peripheral blood. Male (15 specimens): Body length 49 to 73 (55); width 0.20 to 0.28 (0.23) at nerve ring, 0.20 to 0.31 (0.26) at mid-body, 0.07 to 0.10 (0.09) at level of cloaca. Esophagus length 0.87 to 1.00 (0.92). Nerve ring 0.26 to 0.29 (0.27) from anterior end. Cloaca 72 to 84 t from posterior end of body. Preanal papillae large, elongate, pedunculated, usually six or seven on left side, five to seven on right; posteriormost pair set inward toward midline (Fig. 2). Single pair of papillae immediately behind cloaca in midventral line. Paired postanal papillae on line with preanals, thre or four in number, usually four; first pair large, second, third, and fourth tall, slender, closely grouped (Fig. 2). Phasmids ventrolaterally placed near tip of tail. Spicules unequal and dissimilar; left 0.55 to 0.70 (0.60) long, right 0.15 to 0.20 (0.17); mean left : right spicular ratio, 3.5: 1. Left spicule with slender tubular shaft and lamina composed of two laterally oriented elements, one short, the other very long and filamentous (Fig. 3). Right spicule short, relatively narrow, boat-shaped (Fig. 3). Tail spirally coiled, usually in two turns. Caudal alae broad. Female (15 specimens): Body length 130 to 180 (151), width at nerve ring 0.33 to 0.36 (0.34), at vulva 0.32 to 0.40 (0.37), at mid-body 0.32 to 0.37 (0.36), at anus 0.09 to 0.11 (0.10). Esophagus 1.15 to 1.33 (1.28) long; nerve ring 0.31 to 0.37 (0.36) from anterior end. Vulva posterior to esophageal-intestinal junction, 1.7 to 2.9 (2.1) from anterior end. Vagina vera muscular, bulbous, ovejector-like; remainder of vagina, usually di-

Brugia guyanen-sis sp.n. (Nematoda: Filarioidea) from the coatimundi (Nasua nasua vittata) in British Guiana.

A new species of filaria, Brugia guyanensis, is described from the lymphatic system of the coatimundi (Nasua nasua vittata) in the Rupununi District of British Guiana. This finding constitutes a new host, host family, and geographic locality for the genus Brugia. As compared with other Brugia species, B. guyanensis is relatively small, its left spicule long and hooked at the tip. In November 1962, a microfilaria of the Brugia type was found in a coatimundi (Nasua nasua vittata) from the Rupununi District of British Guiana. Subsequently, adult worms belonging to the genus Brugia were recovered from the lymphatic system of a coatimundi from the same area (Orihel, 1963). To date, 19 of these animals, including some very young ones 4 to 6 months old, have been examined for filarial infections, and in 17 of them the Brugia parasite was found; adult worms were recovered from the lymphatic system of nine. Although the genus Brugia is widely distributed in the Eastern Hemisphere, principally in Southeast Asia, and occurs in a wide range of hosts (Buckley, 1960; Laing, Edeson, and Wharton, 1960), it has not previously been reported from the Western Hemisphere, nor has it been reported in mammals of the family Procyonidae. The specific name selected for the new species is derived from the geographical area in which the parasite was first found. Brugia guyanensis sp. n.

DEVELOPMENT OF DIPETALONEMA GRACILE AND D. CAUDISPINA TO THE INFECTIVE STAGE IN CULICOIDES HOLLENSIS

The microfilariae of 2 species of Dipetalonema (i.e., D. gracile and D. caudispina), common parasites of South American monkeys, develop to the third (infective) stage in the biting midge, Culicoides hollensis. Development of both species occurs in the abdominal fat body; D. gracile requires 13 days and D. caudispina 9 days. Despite similarities in the patterns of development of both species, the third-stage larvae of each is morphologically distinct.