Evaluation of the mtDNA-COII region based species specific assay for identifying members of the Anopheles culicifacies species complex

Background: anopheles culicifacies,a major malarial vector has been recognized as a complex of five sibling species,a,b,c,d and e. these sibling species exhibit varied vectorial capacity,host specificity and susceptibility to malarial parasites/ insecticides. in this study,a pcr assay developed earlier for distinguishing the five individual species was validated on samples of an. culicifacies collected from various parts of india. methods: the samples were initially screened using the rdna-its2 region based primers which categorised the samples into either a/d group or b/c/e group. a proportion of samples belonging to each group were subjected to the mtdna-coii pcr assay for identifying individual species. results: among the 615 samples analysed by rdna-its2 pcr assay,303 were found to belong to a/d group and 299 to b/c/e group while 13 turned negative. among 163 samples belonging to a/d group,only one sample displayed the profile characteristic of species a and among the 176 samples falling in the b/c/e group,51 were identified as species b,14 as species c and 41 as species e respectively by the mtdna-coii pcr assay. samples exhibiting products diagnostic of b/c/e,when subjected to pcr-rflp assay identified 15 samples as species e. conclusion: validation of the mtdna-coii pcr assay on large number of samples showed that this technique cannot be used universally to distinguish the 5 members of this species complex,as it has been designed based on minor/ single base differences observed in the coii region.