Characterization of a candidate Trypanosoma rangeli small nucleolar RNA gene and its application in a PCR-based parasite detection

In this study, we report the isolation and characterization of a candidate Trypanosoma rangeli small nucleolar RNA (snoRNA) gene, and the development of a PCR assay for detection of the parasite based on its nucleotide sequence. This gene, isolated from a T. rangeli genomic sub-library, was named snoRNA-c11 and is encoded by a multi-copy gene of 801 bp in length. Computer sequence analysis of snoRNA-c11 showed the presence of two sequence motifs, box C and box D, as well as of two long stretches that perfectly complement the universal core region of the mature rRNA 28S, suggesting that c11 encodes for a Box C/D snoRNA from the parasite. Hybridization analysis using c11 as probe, showed a weak hybridization signal with Trypanosoma cruzi DNA, demonstrating the existence of differences in this locus between these two species. Two oligonucleotide primers from this gene, which specifically amplified a 620-bp fragment in KP1 (+) and KP1 (-) strains of T rangeli, were used in a PCR assay. The amplification allowed the detection of 1 pg of DNA in the presence of heterologous DNA and no amplification was observed with different T cruzi strains (groups I and 11). In addition, the PCR assay reported here is able to detect T rangeli in the presence of T cruzi DNA, and is useful for detection of the parasite in samples from infected vectors. (C) 2003 Elsevier Science (USA). All rights reserved.