Evaluation of the western blot in the confirmatory serologic diagnosis of Chagas' disease.

The Western blot method, using antigens from epimastigote forms of the Trypanosoma cruzi Y strain, was evaluated for the confirmatory diagnosis of Chagas' disease. Serum samples were obtained from 136 chagasic patients (Group I), 23 patients with inconclusive serologic results for Chagas' disease (Group II), 53 patients with other diseases (Group III), and 50 healthy individuals (Group IV). The Western blot results for Group I gave a confirmatory diagnosis of Chagas' diseases in 118 (86.80%), an indeterminate pattern of reactivity in 16 (11.76%), and a negative pattern in only two (1.47%). Of the samples from Groups II, III and IV, none had a positive result in the Western blot; 60.86%, 52.83%, and 16.0%, respectively, showed indeterminate results; and 39.13%, 47,16% and 84,0%, respectively, showed negative results. The Western blot method showed a sensitivity of 86.60%, a specificity of 100.0%, a positive predictive value of 100.0%, and a negative predictive value of 73.50%, and the concordance coefficient kappa was high (0.7789). The results suggest that the previous serologic results for Chagas' disease could be confirmed by Western blot for the detection of specific antibodies to T. cruzi antigenic fractions, which may reduce the medical, legal, and social consequences of an inconclusive serologic result for Chagas' disease and also underscore the need for additional studies for continued efforts in the development of an ideal standard confirmatory test for Chagas' disease.

Genetic variability of trypanosomatids isolated from phytophagous hemiptera defined by morphological, biochemical, and molecular taxonomic markers.

In the present study, we investigated the genetic variability among 49 new isolates of trypanosomatids from phytophagous Hemiptera by means of morphological characters, growth features, and biochemical (enzymes of ornithine-arginine cycle) and molecular markers (based on spliced-leader, and ribosomal genes). From 402 phytophagous insects dissected and examined for the presence of trypanosomatids, 228 species belonging to Pyrrhocoridae, Coreidae, Lygaeidae, and Pentatomidae families harbored trypanosomatids in their salivary glands, or digestive tubes. Among these insects, 211 carried promastigotes and only 17 had choanomastigote forms. The results show a strong association among morphology, growth features, and biochemical and molecular markers and reveal the genetic diversity of the isolates, which were assigned to Crithidia, Phytomonas, and Leptomonas; we found genetic polymorphism within all these genera, thus indicating high genetic variability among trypanosomatids from phytophagous insects.

Surface component characterization as taxonomic tools for Phytomonas spp identification.

The genus Phytomonas arbitrarily includes all protozoa of the family Trypanosomatidae isolated from plants, but its differentiation is a complex task. The phase separation technique using Triton X-114 was used to analyze hydrophobic and hydrophilic surface proteins in ten strains of Phytomonas isolated from various fruits. The iodination of surface proteins by the Iodo-Gen method was also used for Phytomonas isolates from tomatoes, corn and annatto, Herpetomonas samuelpessoai and Crithidia fasciculata. The distribution of protein-bound radioactivity in acrylamide gels was determined by autoradiograms and showed the presence of protein bands of 36-68 kDa in all strains of Phytomonas: there were two major bands at 88 kDa and 94 kDa, with minor bands at 36 kDa and 142 kDa in H. samuelpessoai; and there were three bands at 74, 86 and 94 kDa, with minor bands at 23 kDa and 105 kDa in C. fasciculata. The results demonstrated that samples of plant parasites can be clearly differentiated from H. samuelpessoai and C. fasciculata. These plant parasites were also submitted to polysaccharide analysis by gas-liquid chromatography of the corresponding alditol acetate. Arabinose, galactose, glucose and mannose, were the major monosaccharides found, while fucose, rhamnose and xylose were found in smaller amounts. The results of all these methods indicated that, after extension to a wider range of trypanosomatid strains, they may be useful in Phytomonas taxonomy.