Schizodeme analysis of Trypanosoma cruzi stocks from South and Central America by analysis of PCR-amplified minicircle variable region sequences

Abstract

Kinetoplast DNA (kDNA) was isolated from 56 stocks of Trypanosoma cruzi isolated from human patients, animals and insects from Brazil, Venezuela, Colombia and Costa Rica. Comparison of the patterns of digested kDNA on acrylamide gels led to the grouping of several stocks into two schizodemes. Schizodeme analysis was also performed using a set of 330-bp fragments representing all the variable regions of the minicircle DNA molecules, which were obtained by PCR amplification of the kDNA using conserved region primers. The results of this analysis were consistent with the analysis using total kDNA, but the more informative restriction profiles allowed the construction of additional schizodemes. In addition, two oligomers were generated from variable region sequences of cloned minicircles from a Y and a Cl strain, and these were used as schizodeme-specific probes to detect homologous sequences in the amplified minicircle DNAs. The results indicate that a combination of restriction enzyme fingerprinting and hybridization of amplified variable region minicircle DNA with schizodeme-specific probes can be used for both sensitive detection and classification of T. cruzi.

Mexican Trypanosoma cruzi stocks: analysis of minicircle kDNA homologies by cross-hybridization

“…The absence of strict correlation between hybridization patterns and sub-clustering contrasted with studies showing that kDNA probes identified distinct monophyletic groups (Brenière et al 1998a) but similar results were observed previously among Bolivian stocks isolated from T. sordida vector (Brenière et al 1998b). The HVRm are highly heterogeneous in sequence (Morel et al 1980, Avila et al 1990) and the mechanisms which generate this diversity are unknown. Additional qualitative and quantitative studies of the different minicircle sequence classes would be necessary to best investigate the diversity and evolution of kDNA minicircles, particularly in Mexico where the vector species participate in sylvatic and domestic cycles.…”

Trypanosoma cruzi: clonal structure of parasite strains and the importance of principal clones

“…T. cruzi strains represent subspecies, based on intrinsic characters as antigenic composition (Andrade et al 1981) susceptibility to chemotherapy (Andrade et al 1975, Brener et al 1976), isoenzyme patterns ( Miles et al 1980, Andrade et al 1983, Tibayrenc & Ayala 1988 and genomic profils of kDNA (Morel et al 1980, Avila et al 1990). Studies on the host-parasite relationships have disclosed marked differences in the determinism of tissue lesions, dependent on tropism, virulence and pathogenicity of T. cruzi strains (Andrade et al 1970.…”

Evolution and pathology in Chagas disease: a review

“…Minicircle kDNA integrations behave as mutagens in the host. kDNA has been used for the identification of subgroups of T. cruzi populations (Avila et al 1990). The relationships among distinct parasite populations have been characterized using largely nuclear markers into six discrete typing units (DTUs) (Brisse et al 2000a(Brisse et al ,b, 2001.…”