Isolation and antigenic characterization of human immunodeficiency virus (HIV) in Brazil

A retrovirus infecting a Brazilian AIDS patient was isolated and characterized in terms of its reactivity with sera from individuals infected with human immunodeficiency viruses 1 and 2 (HIV-1 and HIV-2). The Western blot analysis revealed that the Brazilian isolate is very similar to the well characterized HIV-1 strain. The serum of the patient from whom the virus was isolated did not react with the 140 kDa envelope glycoprotein specific for HIV-2.

Characterization of protective and non-protective surface membrane carbohydrate epitopes of Schistosoma mansoni

We have produced a number of monoclonal antibodies, protective and non-protective, which recognize a complex of schistosomula antigens, including the 38 kDa antigen. Eight different protective and non-protective monoclonal antibodies, varying in isotypes, were used in the binding assays. Lectin inhibition studies suggested that the monoclonal antibodies probably recognized carbohydrate epitopes on the antigen(s). Immunoprecipitation studies showed that at least two of the monoclonal antibodies recognized different epitopes on the same molecule. Additionally, we tested for monoclonal antibody binding after the antigens were treated with; 1) proteases, 2) periodate, 3) various exo- and endoglycosidases, 4) mild acid hydrolysis. We also tested for binding of the antibodies to keyhole limpet hemocyanin (KLH). Using the 8 monoclonal antibodies as probes, we were able to define at least 4 different carbohydrate epitopes related to the protective monoclonal antibodies, and at least one epitope which is seen by the non-protective antibodies. The epitope seen by the non-protective antibodies was shown to be cross-reactive with epitopes on KLH. These results demonstrate the importance of epitope mapping studies for any defined vaccine.

Characterization of plasma menbrane polypeptides of trypanosoma from bats

Cell surface proteins of Trypanosoma dionisii, Trypanosoma vespertilionis and Trypanosoma sp. (M238) were radiodinated and their distribution both in the detergent-poor (DPP) and dertergent-enriched phase (DRP) was studied using a phase separation technique in Triton X-114 as well as polyacrylamide gel electrophoresis in sodium dodecyl sulphate (SDS-PAGE). Significant differences were observed in the proteins present in the DRP when the three species of trypanosoma were compared. Two major bands with 88 and 70 KDa were observed in T. sp. (M238) but were not detectable in T. dionisii and T. vespertilionis. Three polypeptides whith 96, 77 and 60 KDa were identified in the DRP of T. vespertilionis. Three major bands with 84, 72 and 60 KDa were observed in the DRP of T. dionisii. Two polypeptides with 34-36 KDa present in the DPP, were observed in the three Trypanosome species analyzed. Our observations show that T. sp. (M238) has characteristic surface polypeptides not found in T. vespertilionis.

Characterization of a Trypanosoma cruzi genomic fragment complementary to several species-specific mRNAs but different from the spliced leader sequence

We have isolated a clone of Trypanosoma cruzi genimic DNA, lambda 3b2-5, which contains sequences that are reiterated in the genome. Northtern blot analysis showed that clone 3b2-5 hybridizes to 1,200-5,000 bases different mRNA species. The number of mRNAs species hybridized to clone 3b2-5 exceeds its coding capacity showing that this clone carries sequences that are common to several mRNAs species and conserved in the poly A(+) RNA. These sequences are not homologous to the T. cruzi spliced leader sequence, since clone 3b2-5 hybridize to a synthetic 20 nucleotice complementary to the spliced leader sequence. Clone 3b2-5 does not hybridize to DNA and RNA from several genera of Trypanosomatidae and other Trypanosoma species indicating that it carries T. cruzi species-specific sequences.

Toxoplasma gondii: a rapid method for the isolation of pure tachyzoites: preliminary characterization of its genome

A rapid and simple technique for the purification of Toxoplasma gondii tachyzoites was developed. Highly purified parasites were obtained from the peritoneal exudates of infected mice by means of two consecutive discontinous sucrose gradients run at low speed (10,000xg, 30 min). Parasites obtained by this method conserved its biological activity. Hybridizations tudies with DNA from healthy mice and from purified tachyzoites preparations demonstrated that Toxoplasma gondii tachyzoites DNA could be obtained with better than 90 per cents purity. Preliminary studies with DNA endonucleases showed the presence in the tachyzoites genome of highly repetitives sequences.

Identification and characterization of sex-linked proteins of Schistosoma mansoni

The proteins of adults worms (male and female) of two isolates (BH and RJ) of Shistosoma mansoni were extracted using Triton X-114 phase separation. The SDS-polyacrilamide gel electrophoresis profiles of the three phases (detergent, aqueous and insoluble proteins) obtained were compared after Coomassie blue and silver staining, surface radioiodination and Western blotting. No major differences were detected between the 2 isolates. Of the 25 or more proteins which partitioned into the detergent phase, only about 8 proteins could be surface radiodinated on live adult worms. A comparison was also made between the profiles of mael and females worms, isolated from bisexually infected mice. Two major female-specific and one male-specific band were detected by silver and/or Coomassie staining. The female bands, 32 KDa and 18 KDa, partitioned into the detergent and aqueous phase, respectively. The male-specific band of 42 KDa remained in the insoluble phase. Antigenic differences between male and females protins were detected by Western vlotting using a sera from infected Nectomys squamipes.

Biological characterization of Trypanosoma cruzi strains from different zymodemes and schizodemes

The development in C3H mice of thirteen strains of Trypanosoma cruzi belonging to different zymodemes ans schizodemes was studied. Host mortality, virulence, histiotropism, parasitemia and polymorphism of the parasites were recorded. The strains were grouped into: a) high virulence - causing 100% mortality and characterized by predominance of bery broad trypomastigotes in the bloodstream at the end of infection; b) medium virulence - causing no mortality and with a predominance of broad trypomastigotes; c) low virulence - causing no mortality with blood forms not described due to the very low parasitemia. During 18 months maintenance the parasitemia curves were kept constant for all strains except one. A direct correlation between either zymodeme or schizodeme and experimental biological properties of T. cruzi strains was not found. However, the parasitemia was subpatent and patent for strains from zymodeme C and the others respectively. Furthermore the high virulence seems to be related to one of two shizodemes found within zymodeme B strains. All strains presenting patent parasitemia independent of shizodeme and ymodeme showed a myotropism towards heart and skeletal muscle with varible inflammatory intensity. The present study confirmed the heterogeneity found by isoenzyme and K-DNA patterns among the strains of T. cruzi isolated from chagasic patients in Bambuí, Minas Gerais State, Brasil.

Characterization of a Plasmodium falciparum mutant that has deleted the majority of the gametocyte-specific Pf11-1 locus

We identified a gametocyte-specific protein of Plasmodium falciparum called Pf11-1 and provide experimental evidence that this molecule is involved in the emergence of gametes of the infected erythrocyte (gametogenesis). A mutant parasite clone, which has deleted over 90% of the PF11-1 gene locus, was an important control to establish the gametocyte-specific expression of the Pf11-1. Molecular analysis of the Pf11-1 deletion indicates that it is presumably due a chromosome breakage with subsequent "healing" by the addition of telomeric heptanucleotides. Moreover, similar DNA rearrangements are observed in most of the laboratory isolates during asexual propagation in vitro.

Morphological characterization of the hemocytes of the pulmonate snail Biomphalaria tenagophila

The blood cells of the pulmonate snail Biomphalaria tenagophila, an important transmiter of the trematode Schistosoma mansoni in Brazil, were examined by ligth and transmission electron microscopy (TEM). Two hemocyte types were identified: hyalinocytes and granulocytes. Hyalinocytes are small young (immature), poorly spreading cells, which have a high nucleocytoplasmic ratio and are especially rich in free ribosomes. They do not appear to contain lysosome-like bodies and represent less than 10% of the circulating hemocytes. Granulocytes are larger hemocytes which readily spread on glass surface and which strongly react to the Gomori substrate, indicating the enzyme acid phosphatase usually found in lysosomes. Ultra-structurally, they contain a well-developed rough endoplasmic reticulum, dictyosomes and some some lysosome-like dense bodies. Granulocytes do not exhibit a characteristic granular aspect and the few granules observed in the cytoplasm should correspond to a lysosome system. They were named granulocytes instead of amoebocytes to use the same terminology adopted for Biomphalaria glabrata in order to make easier comparative studies. This is a preface study for more specific investigations on the functional activities of the blood cells of B. tenagophila and their interactions with the trematode parasite.