Overlapping Toxoplasma gondii genotypes circulating in domestic animals and humans in Southeastern Brazil.

Although several Toxoplasma gondii genotyping studies have been performed in Brazil, studies of isolates from animals in the state of Minas Gerais are rare. The objective of this study was to conduct a genotypic characterization of T. gondii isolates obtained from dogs, free-range chickens, and humans in Minas Gerais and to verify whether the T. gondii genotypes circulating in domestic animals correspond to the genotypes detected in humans. Genetic variability was assessed by restricted fragment length polymorphism at 11 loci (SAG1, 5'+3'SAG2, SAG2 alt, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1, and Apico). Twelve different genotypes were identified among the 24 isolates studied, including 8 previously identified genotypes and 4 new genotypes. The genetic relationship of the 24 T. gondii isolates, together with the genotypes previously described from 24 human newborns with congenital toxoplasmosis, revealed a high degree of similarity among the genotypes circulating in humans and animals in Minas Gerais. The most common genotypes among these species were BrII, BrIII, ToxoDB #108, and ToxoDB #206. Restricted fragment length polymorphism at the CS3 locus of these 48 isolates showed that the majority of isolates presented alleles I (50%) or II (27%). Isolates harboring allele III at the CS3 locus presented low virulence for mice, whereas those harboring alleles I or II presented higher virulence. These results confirm the utility of marker CS3 for predicting the virulence of Brazilian isolates of T. gondii in mice. No association was found between the allele type and clinical manifestations of human congenital toxoplasmosis. This is the first report of T. gondii genotyping that verifies the overlapping genotypes of T. gondii from humans and animals in the same geographic region of Brazil. Our results suggest that there is a common source of infection to the species studied, most likely oocysts contaminating the environment.

IgG and IgM western blot assay for diagnosis of congenital toxoplasmosis.

The aim of this study was to evaluate the utility of western blot (WB) analysis as a diagnostic tool for congenital toxoplasmosis in 215 newborn infants. The children were submitted to clinical examinations to assess macular, neurological and hearing signals. The WB results obtained were compared to the persistence of IgG antibodies at the end of 12 months, which is regarded as the "gold standard" diagnosis of congenital toxoplasmosis. Association between the WB results and the clinical signs presented by the infants was also assessed. Of the 215 children, 177 had a confirmed congenital toxoplasmosis diagnosis and 38 were uninfected. IgG-WB showed a sensitivity of 73.5% and a specificity of 97.4%. IgM-WB showed a sensitivity of 54.8% and a specificity of 94.7%. The IgG-WB and IgM-WB combination increased the sensitivity to 86.5%. The IgM-WB-positive children had a 1.4-fold greater risk of presenting active macular lesions than did those that were IgM-WB-negative. This study showed that the WB assay is a useful tool to confirm a diagnosis of congenital toxoplasmosis and that the IgM-WB-positive results can indicate active macular lesions in newborn infants.

Genetic variability of Brazilian Toxoplasma gondii strains detected by random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) and simple sequence repeat anchored-PCR (SSR-PCR).

The genetic variability of 19 Toxoplasma gondii strains isolated from humans and animals in Brazil was detected by both random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) and simple sequence repeat anchored-PCR (SSR-PCR) for the first time. Two reference strains, RH (highly virulent) and ME49 (avirulent), were submitted to both assays. Besnoitia sp., Plasmodium falciparum and Babesia bigemina were used as outgroups. RAPD-PCR and SSR-PCR profiles were used for building phenetic trees by unweighted pair group method with arithmetic averages (UPGMA). Phenograms built with TREECONW software showed great similarity in the topology of the trees. Both phenograms presented two major clusters that grouped T. gondii strains according to their murine virulence. The strains AS28, BV and N, which are highly virulent for BALB/c mice, were clustered with the reference strain RH, the highly virulent strain of T. gondii that has been most commonly studied. The group formed by the cystogenic strains showed that the strains which presented a level of virulence more similar to that of ME49 strain (avirulent) also presented a closer genetic relationship. Genetic variation within each lineage was significantly lower (P < 0.05) than that between the lineages. Regarding outgroups, Besnoitia sp. presented the closest relationship to T. gondii while P. falciparum was the most distant. The results presented here demonstrate that intraspecific genetic variability separates Brazilian T. gondii strains into two groups which correlate with murine virulence phenotype.