PCR-based screening and lineage identification ofTrypanosoma cruzidirectly from faecal samples of triatomine bugs from northwestern Argentina

Abstract

This study applied improved DNA extraction and polymerase chain reaction strategies for screening and identification ofTrypanosoma cruzilineages directly from faeces of triatomines collected in a well-defined rural area in northwestern Argentina. Amplification of the variable regions of the kinetoplastid minicircle genome (kDNA-PCR) was performed in faecal lysates from 33 microscope (MO)-positive and 93 MO-negativeTriatoma infestans, 2 MO-positive and 38 MO-negativeTriatoma guasayanaand 2 MO-positive and 73 MO-negativeTriatoma garciabesi. kDNA-PCR detectedT. cruziin 91% MO-positive and 7·5% MO-negativeT. infestans, which were confirmed by amplification of the minicircle conserved region. In contrast, kDNA-PCR was negative in all faecal samples from the other triatomine species. A panel of PCR-based genomic markers (intergenic region of spliced-leader DNA, 24Sα and 18S rRNA genes and A10 sequence) was implemented to identify the parasite lineages directly in DNA lysates from faeces and culture isolates from 28 infected specimens. Two were found to be infected with TCI, 24 with TCIIe, 1 with TCIId and 1 revealed a mixed TCI+TCII infection in the faecal sample whose corresponding culture only showed TCII, providing evidence of the advantages of direct typing of biological samples. This study provides an upgrade in the current diagnosis and lineage identification ofT. cruziin field-collected triatomines and showsT. cruziII strains as predominant in the region.