Infection rates and genotypes of Trypanosoma rangeli and T. cruzi infecting free-ranging Saguinus bicolor (Callitrichidae), a critically endangered primate of the Amazon Rainforest.

Parasites of wild primates are important for conservation biology and human health due to their high potential to infect humans. In the Amazon region, non-human primates are commonly infected by Trypanosoma cruzi and T. rangeli, which are also infective to man and several mammals. This is the first survey of trypanosomiasis in a critically endangered species of tamarin, Saguinus bicolor (Callitrichidae), from the Brazilian Amazon Rainforest. Of the 96 free-ranging specimens of S. bicolor examined 45 (46.8%) yielded blood smears positive for trypanosomes. T. rangeli was detected in blood smears of 38 monkeys (39.6%) whereas T. cruzi was never detected. Seven animals (7.3%) presented trypanosomes of the subgenus Megatrypanum. Hemocultures detected 84 positive tamarins (87.5%). Seventy-two of 84 (85.7%) were morphologically diagnosed as T. rangeli and 3 (3.1%) as T. cruzi. Nine tamarins (9.4%) yielded mixed cultures of these two species, which after successive passages generated six cultures exclusively of T. cruzi and two of T. rangeli, with only one culture remaining mixed. Of the 72 cultures positive for T. rangeli, 62 remained as established cultures and were genotyped: 8 were assigned to phylogenetic lineage A (12.9%) and 54 to lineage B (87.1%). Ten established cultures of T. cruzi were genotyped as TCI lineage (100%). Transmission of both trypanosome species, their potential risk to this endangered species and the role of wild primates as reservoirs for trypanosomes infective to humans are discussed.

Haemoproteus (Apicomplexa: Haemoproteidae) of tortoises and turtles.

It is the general opinion that the haemoproteid blood parasites of chelonians belong to the genus Haemoproteus. Different specific names have long been assigned to this parasite in birds, but some past authorities have accepted only a single species, H. metchnikovi, for all those haemoproteids recorded in a wide range of chelonian genera throughout the world. In the present study, a comparison of one such organism in the tortoise Geochelone denticulata with another in the river turtle Peltocephalus dumerilianus, from Amazonian Brazil, has revealed clear morphological differences. These distinguish the parasites from each other, H. metchnikovi and the other named species of chelonian Haemoproteus for which adequate descriptions are available. We have assigned to them the names Haemoproteus geochelonis n.sp. and Haemoproteus peltocephali n.sp.

Characterization and classification of leishmanial parasites from humans, wild mammals, and sand flies in the Amazon region of Brazil.

Ninety-four leishmanial isolates from the Brazilian Amazon Region (Amapá, Amazonas, Pará, and Rondônia) were identified and classified using specific monoclonal antibodies and an indirect radioimmunoassay (serodeme analysis); eighty-two were also characterized by enzyme electrophoresis (zymodeme analysis), the results of which were subjected to a numerical phenetic analysis. Six isolates from humans (3), Didelphis marsupialis (1), Lutzomyia olmeca nociva (1), and Lu, reducta (1) showed reactivity patterns and isoenzyme profiles similar to those obtained with the Leishmania amazonensis reference strains, and were identified as this species. Eighty-six stocks were classified as members of the L. braziliensis complex; of these, 61 were L. guyanensis or variants, which presented three serodeme subtypes, but whose isoenzyme profiles were all similar to the reference strain. A total of 15 isolates were distinguished as L. braziliensis or variants and were classified into five serodeme subtypes. The isolate from Psychodopugus davisi appeared, from the numerical analysis, to be a distinct parasite species. Ten isolates showed reactivity patterns and isoenzyme profiles similar to those obtained with the L. naiffi reference strain. A parasite isolated from Ps. claustrei appeared to be different from all reference strains by both techniques, and was classified as probably being a new species. The importance of these results with respect to the taxonomic status of the New World Leishmania, and their implications for both clinical and epidemiologic data are discussed.

Vertebrate hosts and vectors of Trypanosoma rangeli in the Amazon Basin of Brazil.

A total of 46 Trypanosoma rangeli stocks were isolated from naturally infected mammals and triatomine vectors. Twenty-two stocks were from the common opossum (Didelphis marsupialis), one from the brown "4-eyed" opossum (Metachirus nudicaudatus), one from the anteater (Tamandua tetradactyla), one from the coati (Nasua nasua), seven from Rhodnius pictipes and 14 from Rhodnius robustus. Two stocks were also isolated from recently fed sandflies (Lutzomyia sp., Shannoni group). The stocks were identified as T. rangeli on the basis of natural or experimental salivary gland infections in Rhodnius, inoculative (anterior station) transmission to mice, morphological parameters in parasitemic mice and comparisons of isozyme profiles with a known stock of T. rangeli isolated from man. Three other trypanosome stocks from D. marsupialis, T. tetradactyla and the three-toed sloth (Bradypus tridactylus) were morphologically similar to T. rangeli in culture but had quite different isozyme profiles and were not identified. It is concluded that T. rangeli is widely distributed in Amazonas, Pará and Rondonia States of Brazil, and probably extends into other regions where R. pictipes and R. robustus are known to occur. R. pictipes is light-attracted into houses and occasionally transmits Chagas' disease to man. It is likely that T. rangeli is also occasionally transmitted to man in the Amazon basin.

Flagellate infections of Brazilian sand flies (Diptera: Psychodidae): isolation in vitro and biochemical identification of Endotrypanum and Leishmania.

Flagellate infections were found in 1,063 of 18,895 sand flies collected in the states of Amazonas, Pará, Rondonia and Acre, Brazil. Infection rates were 13.4% (species group Shannoni); 7.5% (subgenus Nyssomyia); 6.7% (subgenus Lutzomyia series Cruciata); 0.5% (genus Psychodopygus) and 3.1% for other sand flies (various subgenera). Leishmania braziliensis guyanensis and L. mexicana amazonensis were isolated, respectively, from the known vectors, Lutzomyia umbratilis and L. flaviscutellata. Single stocks of L. braziliensis-like and L. mexicana-like organisms were isolated, respectively, from L. whitmani and L. yuilli. Thirty-eight flagellate stocks, isolated by direct culture from sand flies were characterized in detail by morphology in culture, behavior in hamsters and mice and by enzyme profiles. Sixteen stocks from Lutzomyia sp. (Shannoni group) were identified as Endotrypanum schaudinni; 8 stocks from Lutzomyia sp. (Shannoni group) were identified as Endotrypanum sp.; 7 stocks from Psychodopygus ayrozai and P. paraensis were identified as Leishmania sp. previously isolated from the armadillo, Dasypus novemcinctus; 2 stocks of Trypanosoma rangeli were isolated from recently fed Lutzomyia sp. (Shannoni group) sand flies; the remaining 5 stocks from L. umbratilis and L. yuilli could not be identified. Observations suggested that Shannoni group sand flies were the natural vectors of Endotrypanum. Leishmania sp. infections in the man-biting flies P. ayrozai and P. paraensis were restricted to the midgut and associated with recent bloodmeals. Unidentified flagellates in L. umbratilis and L. yuilli were distributed throughout the digestive tract with no trace of bloodmeals.

Brazilian isolates of Trypanosoma cruzi from humans and triatomines classified into two lineages using mini-exon and ribosomal RNA sequences.

Traditional molecular and biochemical methods, such as schizodeme analysis, karyotyping, DNA fingerprinting, and enzyme electrophoretic profiles, have shown a large variability among Trypanosoma cruzi isolates. In contrast to those results, polymerase chain reaction (PCR) amplification of sequences from the 24S alpha ribosomal RNA gene and from the mini-exon gene nontranscribed spacer indicated a dimorphism among T. cruzi isolates, which enabled the definition of two major parasite lineages. In the present study, 86 T. cruzi field stocks (68 isolated from humans with defined presentations of Chagas' disease and 18 from triatomines) derived from four Brazilian geographic areas were typed by the PCR assay based on the DNA sequences of the mini-exon and 24S alpha rRNA genes. These stocks were ordered into the two major T. cruzi lineages. Lineage 1 was associated mainly with human isolates and lineage 2 with the sylvatic cycle of the parasite.

New records of Histoplasma capsulatum from wild animals in the Brazilian Amazon.

Twenty-eight isolates of Histoplasma capsulatum were obtained from eight species of forest mammals from the States of Amazonas, Pará and Rondônia in the Amazon Region of Brazil. Primary isolates were obtained by inoculating triturated liver and spleen tissue intradermally and intraperitoneally in hamsters. Mycological diagnosis in hamsters presenting lesions was confirmed by histopathology and culture on Sabouraud dextrose-agar. Infected hamsters developed signs of disease within two to nine months; all had disseminated visceral lesions and most also had skin lesions at the sites of inoculation. None of the hamsters inoculated with skin macerates of the original hosts developed histoplasmosis, and histopathological examination of the viscera of the wild hosts failed to reveal H. capsulatum. Prevalence of infection was considerably higher in females than in males both for the opossum Didelphis marsupialis and for total wild animals (479) examined. It is proposed that canopy-dwelling mammals may acquire the infection from conidia borne on convective currents in hollow trees with openings at ground-level.

Paracoccidioides brasiliensis. A mycologic and immunochemical study of a sample isolated from an armadillo (Dasipus novencinctus).

A sample of P. brasiliensis isolated from the spleen and the liver of an armadillo (Dasipus novencinctus) has been analysed under a mycological and immunochemical viewpoint. The armadillo was captured in an area of Tucuruí (State of Pará, Brazil), the animal being already established as an enzootic reservoir of P. brasiliensis at that region of the country. This sample maintained in the fungal collection of the Tropical Medicine Institute of São Paulo (Brazil) numbered 135, has got all the characteristics of P. brasiliensis, with a strong antigenic power and low virulence for guinea-pigs and Wistar rats. The specific exoantigen of P. brasiliensis--the glycoprotein with a molecular weight of 43 kDa--was easily demonstrated with double immunodiffusion, immunoelectrophoresis, SDS-PAGE and immunobloting techniques.

[Wild vectors of Chagas' disease in an urban area of Manaus (AM): flying activity during dry and rainy seasons]. / Vetores selváticos de doença de Chagas na área urbana de Manaus (AM): atividade de vôo nas estações secas e chuvosas.

Adults of Rhodnius prolixus, R. pictipes and Panstrongylus geniculatus found in houses in Manaus have high rates of Trypanosoma cruzi infection. Rhodnius spp. were found throughout the year, but males of P. geniculatus were significantly more frequent in the dry season.

On Eimeria bragancaensis n. sp. (Apicomplexa: Eimeriidae) and tissue-cysts of an unidentified protozoan in the bat Peropteryx macrotis (Chiroptera: Emballonuridae) from Amazonian Brazil.

A description is given of the mature oocysts and endogenous stages of Eimeria bragancaensis n. sp., from the small intestine of the bat Peropteryx macrotis from north Brazil. Oocysts are spherical to subspherical, 15.9 x 14.6 microns (range 14-17 x 14-17.7). The wall is of two layers: an outer, thicker one which is of a brownish-yellow colour and striated (pitted), and an inner one which is thin and colourless. The outer layer is frequently lost. No micropyle or oocyst residuum present, but the oocyst usually contains one or two polar bodies. Sporocysts 8.4 x 5.3 microns (range 6.25-9 x 4-6) with inconspicuous Stieda and substiedal bodies. Endogenous stages are intracytoplasmic in the epithelial cells of the small intestine, above the host-cell nucleus. Stages of merogony and gametogony are described. Abundant tissue-cysts of an unidentified protozoan, containing from 1-4 zoites, were found in the parenchyma cells of the liver and, less frequently, in the lamina propria of the small intestine. Their possible nature is discussed.

Garnia karyolytica n. sp. (Apicomplexa: Haemosporina: Garniidae), a blood parasite of the Brazilian lizard Thecodactylus rapicaudus (Squamata: Gekkonidae).

Development of meronts and gametocytes of Garnia karyolytica nov. sp., is described in erythrocytes of the neotropical forest gecko Thecodactylus rapicaudus from Pará State, north Brazil. Meronts are round to subspherical and predominantly polar in position: forms reaching 12.0 x 10.0 microns contain from 20-28 nuclei. Macrogametocytes and microgametocytes are predominantly elongate, lateral in the erythrocyte and average 16.6 x 6.3 microns and 15.25 x 6.24 microns respectively. Occasional spherical forms of both sexes occur in a polar or lateropolar position. All stages of development are devoid of malarial pigment. They have a progressively lytic effect on the host-cell nucleus, particularly the mature gametocytes, which enlarge and deform the erythrocyte. Possible vector(s) of garniid parasites are considered, and phlebotomine sandflies are high on the list of suspects.

[Besnoitia (Protozoa: Toxoplasmatinae) isolated from opossums Didelphis marsupilis in the Amazon region, Brazil]. / Besnoitia (Protozoa: Toxoplasmatinae) isolado de mucuras Didelphis marsupialis na região Amazõnica, Brasil.

From a total of 224 Didelphis marsupialis examined, in 15 were found cysts of Besnoitia in muscles and viscera. It's the first time that this protozoan is isolated from naturally infected animals in Brazil. The experimental transmission to laboratory animals was done by the inoculum of tissue and cysts triturated.

Eimeria peltocephali n. sp., (Apicomplexa:Eimeriidae) from the freshwater turtle Peltocephalus dumerilianus (Chelonia:Pelomusidae) and Eimeria molossi n. sp., from the bat, Molossus ater (Mammalia:Chiroptera).

The oocyst is described of Eimeria peltocephali n.sp. from faeces of the freshwater turtle Peltocephalus dumerilianus from Barcelos, State of Amazonas, Brazil. Sporulation is exogenous and fully developed oocysts are elongate, ellipsoidal or cylindrical, frequently curved to a banana-shape, 54.4 x 19.1 (37.5-68.7 x 18.7-20.0 microns), shape-index 2.8 (1.8-3.9). The oocyst wall is a single thin, colourless layer about 1 micron thick, with no micropyle. There is a bulky oocyst residuum, at first spherical to ellipsoidal, 19 x 16 (16.2-26.2 x 16-21.5 microns), but becoming dispersed on maturation. There are no polar bodies. The sporocysts, 19.1 x 6.8 (17.5-21.2 x 6.2-7.5 microns), shape-index 2.8 (2.3-3.2), are usually disposed in pairs at each end of the oocyst, and bear an inconspicuous Stieda body in the form of a flat cap. The sporozoites are elongate and slightly curved around the residuum. No refractile bodies were seen. Eimeria molossi n.sp., is described from the molossid bat Molossus ater. Sporulation is exogenous and the mature oocysts are predominantly broadly ellipsoidal, 23.4 x 17.5 (18-30 x 15-22.5 microns), shape-index 1.3 (1-1.6). The oocyst wall is about 2 microns thick, and of three layers: an inner thin, colourless one and two outer layers which are thicker, yellowish-brown, prominently striated and in close apposition. There is no micropyle or oocyst residuum, but one and occasionally two polar bodies are usually present. Sporocysts are ellipsoidal, 10.2 x 7.5 (10-12.5 x 7.5 microns), shape-index 1.4 (1.3-1.7) with an inconspicuous Stieda body. Endogenous stages are described in the epithelial cells of the small intestine.

Trypanosomes of the subgenus Megatrypanum from armadillos (Xenarthra:Dasypodidae).

A new species of trypanosome, Trypanosoma (Megatrypanum) peba, is described from the peripheral blood of the armadillo Euphractus sexcinctus setosus from Bahia State, Brazil. Ten out of 29 specimens of the armadillo Dasypus novemcinctus from Pará State were found to have trypanosomes, including epimastigote forms, in impression smears of subcutaneous lymph nodes. The trypanosomes from D. novemcinctus are illustrated and were identified as belonging to the subgenus Megatrypanum on the basis of their general appearance, although they failed to multiply in blood-agar culture medium and no bloodstream forms were seen. This is the first published record of trypanosomes of this subgenus from armadillos and the first demonstration of epimastigote trypanosomes in the mammalian host other than in the bloodstream, or in the anal glands of opossums.

Distribution of Histoplasma capsulatum in Amazonian wildlife.

In a survey of 296 sylvatic animals captured from virgin forests in the north-eastern and south-western Amazon of Brazil, Histoplasma capsulatum was isolated, via the indirect hamster inoculation method, from the liver and spleen of four common opossums Didelphis marsupialis and two pacas Agouti paca. The infected animals did not show any clinical symptoms or histopathology. The known Amazonian mammalian species with natural histoplasmosis now total five, the previously reported species being the spiny rat Proechimys guyannensis, the two-toed sloth Choloepus didactylus and the nine-banded armadillo Dasypus novemcinctus.