Resumen del taller sobre el uso de la reacción en cadena de la polimerasa (PCR) para distinguir entre Trypanosoma cruzi y tripanosoma rangeli

Abstract

This workshop on the use of the polymerase chain reaction (PCR) to detect and differentiate T. cruziand T. rangeli had as its main purpose the transfer of this technology to laboratories in Colombia. To demonstrate this technique, we used clinical samples or isolates obtained from endemic areas of Colombia. In order to evaluate the possible effect of different culture media components on the sensitivity of PCR, analyses were performed on trypanosomes that were grown in different culture media. The DNA was extracted from these cells by boiling, Geneclean, and hypotonic Iysis. The DNA was amplified using a conserved 22 synthetic oligonucleotide sequence within the tandemly repeated mini-exon gene from T. cruzi and T. rangel;. The two organisms were distinguished by the electrophoretic mobilities of their respective amplification products. The confirmation of the identity of the PCR products was obtained using species-specific probes from intergenic regions. Hybridisation was visualised by the NBT colour reaction. From a total of 28 samples analysed, 17 identifications were in agreement with their prior identification. From 5 unknown samples, three were identified as T. rangeli and two as mixed infections. We obtained only two ambiguous identificationscaused by contamination in samples or PCR reaction. Only two samples could not be identified by PCR because of DNA extraction problems. The results from this workshop support the potential of the PCR technique as a usefuI additional tool for the detection and diagnosis of Chagas' disease in Colombia.