Membrane fractions from Strongyloides venezuelensis in the immunodiagnosis of human strongyloidiasis / Frações de membrana de Strongyloides venezuelensis para o imunodiagnóstico da estrongiloidíase humana

Strongyloides venezuelensis is a parasitic nematode of rodents frequently used to obtain heterologous antigens for the immunological diagnosis of human strongyloidiasis. The aim of this study was to evaluate membrane fractions from S. venezuelensis for human strongyloidiasis immunodiagnosis. Soluble and membrane fractions were obtained in phosphate saline (SS and SM) and Tris-HCl (TS and TM) from filariform larvae of S. venezuelensis. Ninety-two serum samples (n = 92) were obtained from 20 strongyloidiasis patients (Group I), 32 from patients with other parasitic diseases (Group II), and 40 from healthy individuals (Group III), and were analyzed by enzyme-linked immunosorbent assay (ELISA). Soluble fractions (SS and TS) showed 90.0% sensitivity and 88.9% specificity, whereas the membrane fractions (SM and TM) showed 95.0% sensitivity and 94.4% specificity. The present results suggest the possible use of membrane fractions of S. venezuelensis as an alternative antigen for human strongyloidiasis immunodiagnosis.

Strongyloides venezuelensis é um nematódeo parasita de roedores, frequentemente usado como antígeno heterólogo para o diagnóstico imunológico da estrongiloidíase humana. O objetivo deste estudo foi avaliar frações de membrana de S. venezuelensis para o imunodiagnóstico da estrongiloidíase humana. Para tanto, frações solúveis e de membrana foram obtidas em solução salina fosfato (SS e MS) e Tris-HCl (ST e MT) de larvas filarioides de S. venezuelensis. Amostras de soro de 92 indivíduos, sendo 20 com estrongiloidíase (Grupo I); 32 com outras parasitoses (Grupo II), e 40 indivíduos saudáveis (Grupo III), foram analisadas pelo teste Imunoenzimático (ELISA). As frações solúveis (SS e ST) apresentaram 90,0% e 88,9%, enquanto que as frações de membrana (MS e MT) demonstraram 95,0% e 94,4%, de sensibilidade e especificidade, respectivamente. Os resultados obtidos permitem indicar as frações de membranas como antígeno alternativo para o diagnóstico da estrongiloidíase humana.

Lower production of IL-17A and increased susceptibility to Mycobacterium bovis in mice coinfected with Strongyloides venezuelensis

The presence of intestinal helminths can down-regulate the immune response required to control mycobacterial infection. BALB/c mice infected with Mycobacterium bovis following an infection with the intestinal helminth Strongyloides venezuelensis showed reduced interleukin-17A production by lung cells and increased bacterial burden. Also, small granulomas and a high accumulation of cells expressing the inhibitory molecule CTLA-4 were observed in the lung. These data suggest that intestinal helminth infection could have a detrimental effect on the control of tuberculosis (TB) and render coinfected individuals more susceptible to the development of TB.

Avaliação da resposta imune humoral frente a antígenos de Strongyloides venezuelensis / Evaluation of the humoral immune response to Strongyloides venezuelensis antigens

A estrongiloidíase afeta 30 milhões de pessoas em 70 países. Usualmente, o diagnóstico dessa enteroparasitose é realizado por testes parasitológicos baseados no hidro termotropismo das larvas eliminadas nas fezes, porém esses têm se mostrado pouco sensíveis. Neste trabalho, extratos antigênicos foram testados pelas técnicas de ELISA, Immunoblotting e IFI, utilizando larvas filarióides de Strongyloides venezuelensis, parasita de roedores, que mostram reação cruzada com epítopos de Strongyloides stercoralis. Sensibilidade de 89, 85, 57 por cento para a reação de ELISA e de 100, 100 e 96 por cento, para o Immunoblotting com os antígenos SAL, ZWIP e ZW, e especificidade de 90, 60 e 81 por cento para o ELISA e 96, 92 e 91 por cento para o Immunoblotting para os mesmos antígenos, foram encontradas nestes ensaios.

Strongyloidiasis affects 30 million people in 70 countries. This enteral parasitosis is usually diagnosed using parasitological tests based on hydrotropism or thermotropism of larvae eliminated in feces, but these tests have been shown to have low sensitivity. In this study, antigenic extracts were tested by means of ELISA, immunoblotting and IFI, using filariform larvae of Strongyloides venezuelensis, a parasite of rodents that shows cross-reactions with Strongyloides stercoralis epitopes. Sensitivity of 89, 85 and 57 percent for the ELISA reaction and 100, 100 and 96 percent for immunoblotting with the SAL, ZWIP and ZW antigens, and specificity of 90, 60 and 81 percent for ELISA and 96, 92 and 91 percent for immunoblotting with the same antigens, were found in these assays.

Mucosal mast cell-derived chondroitin sulphate levels in and worm expulsion from FcRgamma-knockout mice following oral challenge with Strongyloides venezuelensis

Mucosal mast cell-derived chondroitin sulphates (sulphated proteoglycans) were assayed in gut washings and homogenate of FcRgamma-knockout (KO) and wild-type (WT) C57BL/6 mice challenged with Strongyloides venezuelensis in order to assess their possible role in secondary immunity against enteric nematodes. Groups of immune KO and WT mice were challenged by oral gavage with 300 infective larvae (L3). Establishment of infection was assessed by daily faecal analysis to determine the number of eggs per gram of faeces (EPG) and by adult worm recovery on days 5 and 13 post challenge. Mucosal mast cell (MMC) counts were done on days 5 and 13 post challenge while MMC-derived chondroitin sulphates in gut washings (days 1 and 5) and homogenate (day 8) were assayed by high performance liquid chromatography (HPLC). Results showed that patent infection occurred in challenged KO but not WT mice despite significantly higher mastocytosis in jejunal sections of KO than WT mice (p<0.001). Similarly but against prediction, significantly higher concentration of MMC-derived chondroitin sulphates was observed in gut homogenate of KO than WT mice (p<0.05). In contrast, significantly higher concentration of chondroitin sulphates was observed in gut washings of WT than KO mice (p<0.05). These results suggest that MMC in KO mice failed to release sufficient amount of sulphated proteoglycans into the gut lumen as did the WT mice, which may have been part of the hostile environment that prevented the establishment in and eventual expulsion of adult S. venezuelensis from the gut of WT mice following challenge.

Enzyme-linked immunosorbent assay for diagnosis of human strongyloidiasis.

Crude antigen (CA) was prepared from Strongyloides stercoralis filariform larvae obtained from in vitro culture of the human feces containing rhabditiform larvae. The lyophilized filariform larvae were ground and ultrasonicated in distilled water then the soluble antigenic preparation was delipidized. The protein content of the crude soluble antigen was 20% of the original dried larvae. The CA was passed through a gel filtration chromatography column and yielded three different protein fractions namely F1, F2 and F3. CA and its fractions were used in the indirect enzyme-linked immunosorbent assay (ELISA) for detecting antibodies to S. stercoralis in serum samples of 5 groups of individuals. These were patients with parasitologically confirmed strongyloidiasis (group 1), patients with mixed S. stercoralis and other parasitic infections (group 2), non-strongyloidiasis patients with other worm infestation(s) (group 3), normal parasite-free Thais (group 4) and normal parasite-free Swedes (group 5). It was found that F2 was the best antigen in the ELISA. The sensitivity, specificity and positive and negative predictive values of the test using F2 as the antigen were 95.0%, 96.4%, 95.0% and 96.4%, respectively.

Diagnóstico sorológico da estrongiloidíase humana, através do método imunoenzimático / Immunoenzymatic serological diagnosis of human strongyloidiasis

O presente trabalho demonstra a eficácia do método imunoenzimático utilizando antígeno solúvel de larvas de Strongyloides stercoralis no diagnóstico da estrongiloidíase humana. Foram avalidados 27 pacientes com as diversas formas clínicas da paraditose, sendo demonstrados títulos significativos em 25 (92%) dos casos. Em 17 controles, com ou sem outras parasitose intestinais, títulos significativos estavam presentes em 3 (18%). A sensibilidade do teste foi de 92% e a especificidade de 82%. Säo também relatados 3 casos clínicos, nos quais o diagnóstico da doença foi feito inicialmente pela determinaçäo de anticorpos contra larvas de S. stercoralis, havendo posteriormente demonstrado parasitológica em 2 casos. É ressaltada a importância do teste no diagnóstico da estrongiloidíase aguda e em situaçöes onde eosinofilia näo esteja associada a outras condiçöes clínicas

Antígeno de Strongyloides cebus (DARLING,1911) no diagnóstico da estrongiloidíase humana / Antigen of Strongyloides cebus on the diagnosis ofthe human strongyloidiasis

A técnica de imunofluorescência indireta, usando como antígeno, larvas filarióides de S. cebus, em cortes de congelaçäo infectados por S. stercoralis. Como testemunha foram utilizados 50 soros de indivíduos infectados por ancilostomídeos, 50 por A. lumbricóides, 50 por Protozoários intestinais e 50 com exame coproscópico negativo. Foram considerados níveis significativos de anticorpos anti-Strongyloides, títulos superiores à diluiçäo 1:80 e negativos, diluiçöes iguais ou inferiores a esta. Houve 82 pôr cento de positividade em 50 casos de infecçäo por S. stercoralis. Entre os 200 soros empregados como testemunha todos se mostraram negativos exceto 2 casos de infecçäo por ancilostomídeos