Characterization of Trypanosoma evansi type B

A distinctive feature of Trypanosoma evansi is the possession of a kinetoplast that contains homogeneous DNA minicircles, but lacks DNA maxicircles. Two major sequence variants of the minicircle have been described and here we have sequenced the type B variant and designed a specific PCR test to distinguish it from type A. Further a test based on maxicircles to distinguish T. brucei brucei from T. evansi was designed and evaluated. Using the designed PCR tests, we detected three type B isolates from camel blood samples collected in northern Kenya, more than 20 years after the first isolation of type B. Comparison of minicircle sequences from all four type B isolates shows >96% identity within the group, and 50-60% identity to type A minicircles. Phylogenetic analysis based on minicircle sequences reveals two clusters, one comprising isolates of type A and one of type B, while random amplification of polymorphic DNA show slight polymorphic bands within type B. Most T. evansi isolates analysed were heterozygous at a repetitive coding locus (MORF2). All type B isolates had one genotype designated 3/5 based on the alleles present. Three camel isolates, which had homogenous type A minicircles, lacked the RoTat 1.2 gene, while another five isolates were T. b. brucei, based on the heterogeneity of their minicircles and presence of maxicircles as demonstrated by PCR amplification of the gene for cytochrome oxidase subunit 1. Our results confirm the existence of T. evansi type B isolates, T. b. brucei and existence of T. evansi type A without RoTat 1.2 gene in Kenyan isolates.