Cytochrome oxidase subunit 2 gene allows simultaneous detection and typing of Trypanosoma rangeli and Trypanosoma cruzi

Background: the parasites trypanosoma rangeli and trypanosoma cruzi share vectors and hosts over a wide geographical area in latin america. in this study,we propose a single molecular approach for simultaneous detection and typing of t. rangeli and t. cruzi. methods: a restriction fragment length polymorphism analysis of the mitochondrial cytochrome oxidase ii gene (coii-rflp) using enzyme alui and different amounts of dna from the major genetic groups of t. rangeli and t. cruzi (kp1+/kp1- and dtu-i/dtu-ii) was carried out. the same marker was tested on the other t. cruzi dtus (dtu-iii to dtu-vi) and on dna extracted from gut contents of experimentally infected triatomines. results: the coii pcr generates a ∼400 bp fragment,which after digestion with alui (coii-rflp) can be used to distinguish t. rangeli from t. cruzi and simultaneously differentiate the major genetic groups of t. rangeli (kp1+ and kp1-) and t. cruzi (dtu-i and dtu-ii). the coii-rflp generated bands of ∼120 bp and ∼280 bp for kp1+,whereas for kp1- no amplicon cleavage was observed. for t. cruzi,digestion of coii revealed a ∼300 bp band for dtu-i and a ∼250 bp band for dtu-ii. for dtu-iii to dtu-vi,coii-rflp generated bands ranging from ∼310 to ∼330 bp,but the differentiation of these dtus was not as clear as the separation between dtu-i and dtu-ii. after alui digestion,a species-specific fragment of ∼80 bp was observed for all dtus of t. cruzi. no cross-amplification was observed for leishmania spp.,t. vivax or t. evansi. conclusions: the coii-rflp allowed simultaneous detection and typing of t. rangeli and t. cruzi strains according to their major genetic groups (kp1+/kp1- and dtu-i/dtu-ii) in vitro and in vivo,providing a reliable and sensitive tool for epidemiological studies in areas where t. rangeli and t. cruzi coexist. © 2013 de sá et al.; licensee biomed central ltd.