Valmir Laurentino Silva

Amoebiasis distribution in the past: first steps using an immunoassay technique

The identification of parasites in ancient human faeces is compromised by differential preservation of identifiable parasite structures. However, protein molecules can survive the damage of the environment and can be detected even after centuries. In this paper it is shown that is possible to detect copro-antigen of Entamoeba histolytica in historic and prehistoric human faecal remains, using a commercially available enzyme immunoassay (ELISA) kit. The kit uses monoclonal antibody-peroxidase conjugate specific for E. histolytica adhesin. A total of 90 specimens of desiccated faeces found in mummies and ancient organic sediment from South America, North America, Africa, and Europe were examined. The ELISA detected 20 positive samples, dated to about 5300 years before present to the 19th Century ad. The positive samples are from archaeological sites in Argentina, USA, France, Belgium, and Switzerland. The detection of protozoan antigen using immunoassays is a reliable tool for the studies of intestinal parasites in the past.

Prevalência da infecção canina em áreas endêmicas de leishmaniose tegumentar americana, do município de Paracambi, Estado do Rio de Janeiro, no período entre 1992 e 1993

In the district of Paracambi, State of Rio de Janeiro an epidemiological survey for American tegumentary leishmaniasis in canine population was carried out in endemic localities. A total of 179 dogs was registered and 138 (77.1%) examined for their clinical aspects, development of delayed hypersensitivity (DHS) with Imunoleish® antigen and serological responses by indirect immunofluorescent reaction and enzyme-linked immunosorbent assay. In 9 (6.5%) dogs with active cutaneous lesions or suspect scars, 66.7% were caused by Leishmania sp; 44.4% produced infection in hamsters and showed growth in culture media, which was considered to be compatible with the species of Leishmania braziliensis complex. The molecular characterization (isoenzyme and KDNA restriction profiles) defined two strains with similar profiles for L. (Viannia) braziliensis. The prevalence of canine infection estimated by the cutaneous test, IFR and ELISA was 10.1%, 16.7% and 27.8%, respectively. The presence of clinical / sub-clinical form of ATL in canine population associated with human infections suggested that the dog can act as source of infection as well as for dissemination of the disease.

Canine tegumentar leishmaníasis in the town of Balneário Camboriú in the State of Santa Catarina

INTRODUCTION The increase in American tegumentary leishmaniasis (ATL) cases in 2005 was due to an outbreak that occurred in the towns of Itapema and Balneário Camboriú. This study aimed to determine the possible presence of infection by Leishmania sp in domestic dogs living in an endemic focus of ATL in the town of Balneário Camboriú and verify the existence of a correlation between the humoral and cellular immune responses, the presence of suggestive lesions and positivity in direct parasitological exams. METHODS A total of 275 dogs were evaluated, examined according to clinic aspects, the development of delayed hypersensitivity to Immunoleish antigens and serological responses to the indirect immunofluorescence reaction and immunoenzymatic assay. RESULTS Seven dogs presented suspect lesions, but Leishmania was not detected using the direct parasitological method. The serological result was 5.8% positivity using the IIFR technique, 6.2% by ELISA and 1.8% using the intradermal test. The total number of dogs positive for ATL using the serological exams and/or intradermal test was 24, indicating prevalence of 8.7% for ATL. CONCLUSIONS Further studies are required to evaluate the participation of dogs in the epidemiological chain of ATL in Balneário Camboriú.

Seroepidemiological aspects of Leishmania spp. in dogs in the Itaguai micro-region, Rio de Janeiro, Brazil.

This study evaluated factors associated with the frequency of Leishmania spp. antibodies in dogs residing in the Itaguai micro-region, State of Rio de Janeiro, Brazil. Blood samples were collected from 524 dogs. The serum samples were submitted to indirect fluorescent antibody test (IFAT) and enzyme-linked immunosorbent assay (ELISA) for Leishmania spp. The frequency of seropositive dogs was 28.24% (n = 148) in the micro-region, and among the three municipalities within that region, the highest frequency (p < 0.05) was observed in Seropedica (59.46%), followed by Itaguai (29.05%) and Mangaratiba (11.49%). Regarding factors associated with the host, mongrel dogs and those over the age of two presented higher frequency of antibodies to Leishmania spp. (p < 0.05). Concerning factors related to the environment and habits of the animal, dogs residing in rural areas (FR = 1.67, p = 0.0002), living outside the residence (FR = 1.42, p = 0.0197), with access to forest, streams and pastures (FR = 2.81, p = 0.0007), remaining loose (FR = 1.66, p = 0.0073), and those that had no shelter (FR = 2.16, p < 0.0001) were more likely to be seropositive. Canine leishmaniasis is a disease with high occurrence in the Itaguai micro-region, and aspects such as the definition of breed, age, habits and care by owners showed significant association in this micro-region.

Estudo de validação comparativo entre as técnicas Elisa e RIFI para diagnosticar Leishmania sp em cães errantes apreendidos no município de Campos dos Goytacazes, Estado do Rio de Janeiro

A survey was carried out aiming to verify the ELISA test effectiveness for detecting antibodies against Leishmania sp in dogs, comparing with RIFI human pattern and for investigating sorological zoonosis situation in the microregion. An accordance about 97.6% considered strong was reported.

IDENTIFICATION OF CANINE VISCERAL LEISHMANIASIS IN A PREVIOUSLY UNAFFECTED AREA BY CONVENTIONAL DIAGNOSTIC TECHNIQUES AND CELL-BLOCK FIXATION.

After the report of a second case of canine visceral leishmaniasis (CVL) in São Bento da Lagoa, Itaipuaçu, in the municipality of Maricá, Rio de Janeiro State, an epidemiological survey was carried out, through active search, totaling 145 dogs. Indirect immunofluorescence assay (IFA), enzyme-linked immunosorbent assay (ELISA), and rapid chromatographic immunoassay based on dual-path platform (DPP(r)) were used to perform the serological examinations. The parasitological diagnosis of cutaneous fragments was performed by parasitological culture, histopathology, and immunohistochemistry. In the serological assessment, 21 dogs were seropositive by IFA, 17 by ELISA, and 11 by DPP(r), with sensitivity of 66.7%, 66.7% and 50%, and specificity of 87.2%, 90.2% and 94%, respectively for each technique. The immunohistochemistry of bone marrow using the cell-block technique presented the best results, with six positive dogs found, three of which tested negative by the other parasitological techniques. Leishmania sp. was isolated by parasitological culture in three dogs. The detection of autochthonous Leishmania infantum in Itaipuaçu, and the high prevalence of seropositive dogs confirm the circulation of this parasite in the study area and alert for the risk of expansion in the State of Rio de Janeiro.

Parasitological and immunological diagnoses from feces of captive-bred snakes at Vital Brazil Institute

Fecal samples from 56 snakes at the Vital Brazil Institute, in the city of Niterói, Rio de Janeiro, were tested using the sedimentation and flotation techniques to investigate the evolutionary forms of parasites such as helminths and protozoa, and using enzyme immunoassay techniques to detect antigens of Cryptosporidium sp. and Giardia sp. Among the animals tested, 80.3% were positive for parasites. Out of these, there were 16 Bothrops jararaca, 16 B. jararacussu and 13 Crotalus durissus. The prevalence of parasitic nematodes was 41.1%, and nematodes were found in all three snake species. Among these, the most frequent finding was eggs of Kalicephalus sp., which were diagnosed in 25% of the snakes. The positivity for protozoa detected using parasite concentration techniques was 75%, including oocysts of Caryospora sp. in 75%, cysts with morphology similar to Giardia sp. 3.6%, amoeboid cysts in 41.1% and unsporulated coccidia oocysts in 8.9%. Immunoassays for Cryptosporidium sp. antigens produced positive findings in 60.7%. Pseudoparasites were detected in 64.3%. These results show that there is a need to improve the sanitary handling of captive-bred snakes, and also for the animal house that supplies rodents to feed them. The results also highlight that diagnostic tests should be performed periodically on stool specimens from captive-bred snakes.

PREVALENCE OF AMERICAN TRYPANOSOMIASIS AND LEISHMANIASES IN DOMESTIC DOGS IN A RURAL AREA OF THE MUNICIPALITY OF SÃO JOÃO DO PIAUÍ, PIAUÍ STATE, BRAZIL.

SUMMARY Chagas disease and the leishmaniases are endemic zoonoses of great importance to public health in the state of Piauí, Brazil. The domestic dog (Canis familiaris) is a major reservoir, host of Trypanosoma cruzi and Leishmania spp. in both urban and rural areas, playing an important role in the transmission of these parasites. The present study evaluated the prevalence of both infectious diseases in dogs of a rural area in the municipality of São João do Piauí, Piauí State. One hundred twenty-nine blood samples were collected for serological assessment: for the leishmaniases, 49 (38%) animals tested positive by the Dual-Path Platform technology (DPP), nine (6%) by the Enzyme-linked Immunosorbent Assay (ELISA), and 19 (14.7%) by the Indirect Fluorescent Antibody test (IFA); while for American Trypanosomiasis, 36 (28%) dogs were reagent by ELISA and 21 by IFA. Of the 129 dogs sampled, 76 were submitted to xenodiagnosis, bone marrow aspiration and skin biopsy to perform parasitological tests whose results showed only one (2.3%) positive skin sample for Trypanosoma caninum and one positive xenodiagnosis for T. cruzi, both results confirmed by molecular assays. Three hundred triatomines of the species Triatoma brasiliensis and 552 phlebotomines - 509 (97%) of the species Lutzomyia longipalpis, were also captured.

CANINE VISCERAL LEISHMANIASIS CASE INVESTIGATION IN THE JACARE REGION OF NITEROI, RIO DE JANEIRO, BRAZIL

SUMMARY American visceral leishmaniasis is a vector-borne zoonosis in expansion in Brazil. Dogs are the main urban reservoir. Departing from a case of canine visceral leishmaniasis (CVL) in Jacaré, Niterói, Rio de Janeiro State, an epidemiological canine and entomological study was performed to assess the extension of the disease at the location. Sample was collected around the case and the dogs identified by serological tests (rapid double platform immunochromatographic exams, immunoenzymatic assay/ELISA, indirect immunofluorescence/IFAT). The parasitological diagnosis was performed in animals positive in at least one of these tests. The entomological study was carried out by using light traps and manual collection. The associations between canine variables and outcome (ELISA and IFAT reagents) were assessed by the chi-square test and adjusted by multivariate logistic regression for those associations with p < 0.1 in the bivariate analysis. Seventeen cases of CVL were detected among 110 evaluated dogs (prevalence of 15.5%). Presence of ectoparasites (OR 6.5; 95% CI 1.1-37.4), animals with clinical signs (OR 9.5; 95% CI 1.2-76.6), and previous cases of CVL in the same house (OR 17.9; 95% CI 2.2-147.1) were associated with the outcome. Lutzomyia longipalpiswas not detected. Our results are indicative of an ongoing transmission in the area.

Paleoparasitological analysis of the extinct Myotragus balearicus Bate 1909 (Artiodactyla, Caprinae) from Mallorca (Balearic Islands, Western Mediterranean)

Myotragus balearicus (Artiodactyla, Caprinae) is an extinct caprine endemic of the Eastern Balearic Islands or Gymnesics (i.e., Mallorca, Menorca and surrounding islets, Western Mediterranean Sea). In spite of its small size, c. 50cm height at the shoulder, it was the largest mammal inhabiting these islands until the human arrival, and it had peculiar short legs and frontal vision. It disappeared between 2830 and 2210calBCE. The coprolites here studied were recovered from Cova Estreta, in Pollença, Mallorca. The samples were subjected to microscopic examination and enzyme-linked immunosorbent assays (ELISA) for E. histolytica/E. dispar, Giardia intestinalis and Cryptosporidium parvum. This study provides new paleoparasitological data from an extinct animal species of the Holocene period. The microscopy revealed one sample containing uninucleated-cyst of Entamoeba sp., whereas ELISA detected nine positive samples for Cryptosporidium sp. The finding of these protozoans can help in the discussion of its extinction cause and demonstrates the antiquity and the evolutionary history of host-parasite relationships between protozoa and caprines since the Messinian.