title: Evaluation of the mtDNA-COII Region Based Species Specific Assay for Identifying Members of the Anopheles culicifacies Species Complex.
abstract: 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.
  
  
  CONCLUSIONS
  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.
authors.affiliation: Vector Control Research Centre, Medical Complex, Indira Nagar, Puducherry, India.
authors.forename: Arulsamy Mary
authors.initials: AM
authors.lastname: Manonmani
authors.affiliation: Vector Control Research Centre, Medical Complex, Indira Nagar, Puducherry, India.
authors.forename: Ashok Kumar
authors.initials: AK
authors.lastname: Mathivanan
authors.affiliation: Vector Control Research Centre, Medical Complex, Indira Nagar, Puducherry, India.
authors.forename: Candassamy
authors.initials: C
authors.lastname: Sadanandane
authors.affiliation: Vector Control Research Centre, Medical Complex, Indira Nagar, Puducherry, India.
authors.forename: Purushothaman
authors.initials: P
authors.lastname: Jambulingam
keywords: Anopheles culicifacies; COII PCR assay; Sibling species; rDNA-ITS2 PCR assay