Multiepitope synthetic peptide and recombinant protein for the detection of antibodies to Trypanosoma cruzi in patients with treated or untreated Chagas' disease.
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A tetrapeptide and a recombinant protein, each representing 4 immunodominant epitopes of Trypanosoma cruzi, were tested by use of ELISA for the detection of serum antibodies. Sera from individuals with Chagas' disease, including persons untreated and successfully or unsuccessfully treated, were tested. These assays detected antibody in 100% of the parasitemias. The antibody reactivity decreased based on the success of treatment. Higher sensitivity was observed for tetrapeptide/recombinant protein assays than for lysate-based ELISA, and specificity was improved, particularly with Leishmania sera. The results indicate that multiepitope antigens provide a more sensitive and specific alternative to lysate for detection of anti-T. cruzi antibodies, as required for developing blood screening assays. (+info)
Use of polymerase chain reaction to diagnose the fifth reported US case of autochthonous transmission of Trypanosoma cruzi, in Tennessee, 1998.
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In July 1998, the mother of an 18-month-old boy in rural Tennessee found a triatomine bug in his crib, which she saved because it resembled a bug shown on a television program about insects that prey on mammals. The gut contents of the Triatoma sanguisuga were found, by light microscopy and polymerase chain reaction (PCR), to be infected with Trypanosoma cruzi; PCR products hybridized with T. cruzi-specific oligonucleotide probes. Whole-blood specimens obtained from the child in July and August were negative by buffy-coat examination and hemoculture but positive by PCR and DNA hybridization, suggesting that he had low-level parasitemia. Specimens obtained after treatment with benznidazole were negative. He did not develop anti-T. cruzi antibody; 19 relatives and neighbors also were seronegative. Two of 3 raccoons trapped in the vicinity had positive hemocultures for T. cruzi. The child's case of T. cruzi infection-the fifth reported US autochthonous case-would have been missed without his mother's attentiveness and the availability of sensitive molecular techniques. (+info)
Total and segmental colonic transit time in constipated patients with Chagas' disease without megaesophagus or megacolon.
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Manometric and pharmacological tests have shown that motor abnormalities may occur in the non-dilated colons of chagasic patients. In order to investigate the presence of abnormalities of colonic function in constipated patients with Chagas' disease (ChC) without megaesophagus or megacolon, studies of total and segmental colonic transit time with radiopaque markers were performed on 15 ChC patients, 27 healthy volunteers and 17 patients with idiopathic constipation (IC). The values obtained for the control group were similar to those reported in the literature (total colonic time: 34. 1 +/- 15.6 h; right colon: 9.9 +/- 7.3 h; left colon: 10.8 +/- 10 h, and rectosigmoid: 12.6 +/- 9.9 h). Colonic transit time data permitted us to divide both IC and ChC patients into groups with normal transit and those with slow colonic transit. Colonic inertia was detected in 41% of IC patients and in 13% of ChC patients; left colon isolated stasis (hindgut dysfunction) was detected in 12% of IC patients and 7% of ChC patients, and outlet obstruction was detected in 6% of IC patients and 7% of ChC patients. There were no significant differences in total or segmental colonic transit times between slow transit IC and slow transit ChC patients. In conclusion, an impairment of colonic motility was detected in about 30% of constipated patients with Chagas' disease without megaesophagus or megacolon. This subgroup of patients presented no distinctive clinical feature or pattern of colonic dysmotility when compared to patients with slow transit idiopathic constipation. (+info)
IFN-gamma-independent IgG2a production in mice infected with viruses and parasites.
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After infection with some viruses and intracellular parasites, antibody production is restricted to IgG2a. We first observed that, whereas live viruses such as lactate dehydrogenase-elevating virus (LDV) or mouse adenovirus induced mostly an IgG2a response, a large proportion of antibodies produced against killed viruses were IgG1. This IgG1 antiviral response was suppressed when live virions were added to inactivated viral particles. These results indicate that the IgG2a preponderance is related to the infectious process itself rather than to the type of antigen involved. Since IFN-gamma is known to stimulate IgG2a production by activated B lymphocytes and to be secreted after infection, we examined the role of this cytokine in the antibody isotypic distribution caused by LDV. Most IgG2a responses were relatively unaffected in mice deficient for the IFN-gamma receptor or treated with anti-IFN-gamma antibody. A similar IFN-gamma-independent IgG2a secretion was observed after infection with the parasites Toxoplasma gondii and Trypanosoma cruzi. However, the IFN-gamma-independent IgG2a production triggered by infection still required the presence of functional T(h) lymphocytes. Therefore, signal(s) other than IFN-gamma secretion may explain the T(h)-dependent isotypic bias in antibody secretion triggered by viruses and parasites. (+info)
Serologic testing for Trypanosoma cruzi: comparison of radioimmunoprecipitation assay with commercially available indirect immunofluorescence assay, indirect hemagglutination assay, and enzyme-linked immunosorbent assay kits.
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The radioimmunoprecipitation assay (RIPA) has been used as a confirmatory test in several ongoing and published studies of Trypanosoma cruzi in blood donors in the United States. Despite its use as a confirmatory test, few studies are available comparing RIPA to commercially available serologic test methods. Thus, we compared RIPA with two indirect hemagglutination assays (Biolab Diagnostica SA, Sao Paulo, Brazil; Hemagen Diagnostics, Inc., Waltham, Mass.) and four different enzyme-linked immunosorbent assays (Abbott Laboratories, Abbott Park, Ill.; Embrabio, Sao Paulo, Brazil; Organon Teknika, Sao Paulo, Brazil; and Gull Laboratories, Salt Lake City, Utah) using a panel of 220 serum specimens from Brazilian blood donors with a range of T. cruzi antibody titers as determined by indirect immunofluorescence assay (IFA). A titer of 1:20 was used as the baseline for seropositivity. All IFA-negative serum specimens (n = 19) were nonreactive on all tests. At a titer of 1:20 (n = 9), reactivity rates varied considerably among the tests, with only the RIPA and the Organon and Gull assays identifying reactive specimens. For specimens at a 1:40 titer (n = 35), most assays identified at least 32 of 35 (91%) specimens as reactive, but the Biolab assay only identified 24 (69%). At higher titers (1:80, n = 56; 1:160, n = 101) the assays were comparable, with the exception of the Biolab assay, demonstrating rates of agreement with IFA of >/=98%. Overall, when compared with several other test formats, RIPA demonstrated equivalent or superior rates of agreement with IFA-positive specimens across all titers examined. In particular, at titers of >1:40, the RIPA compared favorably with other test methods currently in use, supporting its application as a confirmatory test, particularly in a research setting. (+info)
Serological confirmation of Chagas' disease by a recombinant and peptide antigen line immunoassay: INNO-LIA chagas.
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Although screening for Trypanosoma cruzi antibodies is mandatory in most South American countries, current tests are insensitive and have poor specificity. A recently optimized line immunoassay (the INNO-LIA Chagas assay) for the serological confirmation of Chagas' disease was evaluated at a large blood bank in Sao Paulo, Brazil. Sera from blood donors who reacted in at least one of three serological screening assays (n = 1,604) and who returned for a follow-up were retested, and the donors were interviewed to assess their epidemiological risk. The results obtained by the confirmatory assay evaluated in this study were compared to those obtained by the three different screening assays. Upon consideration of the consensus results obtained by the three different screening assays as a "gold standard," the INNO-LIA Chagas assay showed a sensitivity of 99.4% (95% confidence interval [CI], 98.3 to 99.9) and a specificity of 98.1% (95% CI, 96.6 to 99.0) for positive (n = 503) and negative (n = 577) sera. The INNO-LIA Chagas assay confirmed the results for significantly larger numbers of positive samples of at-risk individuals independent of the number of positive screening tests (P = 0.017, Mantel-Haenszel test). In conclusion, the INNO-LIA Chagas assay reliably confirmed the presence of antibodies to T. cruzi and can be implemented as a confirmatory assay for Chagas' disease serology. (+info)
Genetic structure and phylogenetic relationships of Colombian Trypanosoma cruzi populations as determined by schizodeme and isoenzyme markers.
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Twenty-four Trypanosoma cruzi stocks isolated from vectors and from human and Didelphis marsupialis hosts from highly separated sylvatic localities in Colombia were characterized by isoenzyme and schizodeme analyses. The stocks were collected primarily from sylvatic ecotopes representing areas of low, moderate, and high endemicity for Chagas' diseases in Colombia. Parasites were characterized mainly by schizodeme analysis with the restriction enzyme Eco RI and the isoenzyme analysis was performed at 10 genetic loci. These analyses demonstrated an agreement between the classifications based on the isoenzyme analysis and on the restriction fragment length polymorphism patterns obtained with the Colombian stocks. There is clear evidence of demic subdivision between the eastern (E) and western (W) stocks separated by the Andean Mountains and Magdalena River, which is likely due to the geographic isolation generated by these topographic features. Heterozygosity estimates indicate that the E group could be more ancient than the W group. As was postulated in a previous study, these results are also compatible with the existence of a clonal population structure in Colombian sylvatic T. cruzi. Evidence presented here failed to demonstrate a correlation between the degree of endemy and genetic clustering. Finally, schizodeme and isoenzymatic analyses comparing Colombian T. cruzi stocks with others from Chile confirm that Colombian isolates are genetically related to zymodeme 1 and distant from zymodeme 2. (+info)
A role for extracellular amastigotes in the immunopathology of Chagas disease.
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In spite of the growing knowledge obtained about immune control of Trypanosoma cruzi infection, the mechanisms responsible for the variable clinico-pathological expression of Chagas disease remain unknown. In a twist from previous concepts, recent studies indicated that tissue parasitism is a pre-requisite for the development of chronic myocarditis. This fundamental concept, together with the realization that T. cruzi organisms consist of genetically heterogeneous clones, offers a new framework for studies of molecular pathogenesis. In the present article, we will discuss in general terms the possible implications of genetic variability of T. cruzi antigens and proteases to immunopathology. Peptide epitopes from a highly polymorphic subfamily of trans-sialidase (TS) antigens were recently identified as targets of killer T cell (CTL) responses, both in mice and humans. While some class I MHC restricted CTL recognize epitopes derived from amastigote-specific TS-related antigens (TSRA), others are targeted to peptide epitopes originating from trypomastigote-specific TSRA. A mechanistic hypothesis is proposed to explain how the functional activity and specificity of class I MHC restricted killer T cells may control the extent to which tissue are exposed to prematurely released amastigotes. Chronic immunopathology may be exacerbated due the progressive accumulation of amastigote-derived antigens and pro-inflammatory molecules (eg. GPI-mucins and kinin-releasing proteases) in dead macrophage bodies. (+info)