title: A multi-locus approach to barcoding in the Anopheles strodei subgroup (Diptera: Culicidae).
abstract: BACKGROUND
  The ability to successfully identify and incriminate pathogen vectors is fundamental to effective pathogen control and management. This task is confounded by the existence of cryptic species complexes. Molecular markers can offer a highly effective means of species identification in such complexes and are routinely employed in the study of medical entomology. Here we evaluate a multi-locus system for the identification of potential malaria vectors in the Anopheles strodei subgroup.
  
  
  METHODS
  Larvae, pupae and adult mosquitoes (n = 61) from the An. strodei subgroup were collected from 21 localities in nine Brazilian states and sequenced for the COI, ITS2 and white gene. A Bayesian phylogenetic approach was used to describe the relationships in the Strodei Subgroup and the utility of COI and ITS2 barcodes was assessed using the neighbor joining tree and "best close match" approaches.
  
  
  RESULTS
  Bayesian phylogenetic analysis of the COI, ITS2 and white gene found support for seven clades in the An. strodei subgroup. The COI and ITS2 barcodes were individually unsuccessful at resolving and identifying some species in the Subgroup. The COI barcode failed to resolve An. albertoi and An. strodei but successfully identified approximately 92% of all species queries, while the ITS2 barcode failed to resolve An. arthuri and successfully identified approximately 60% of all species queries. A multi-locus COI-ITS2 barcode, however, resolved all species in a neighbor joining tree and successfully identified all species queries using the "best close match" approach.
  
  
  CONCLUSIONS
  Our study corroborates the existence of An. albertoi, An. CP Form and An. strodei in the An. strodei subgroup and identifies four species under An. arthuri informally named A-D herein. The use of a multi-locus barcode is proposed for species identification, which has potentially important utility for vector incrimination. Individuals previously found naturally infected with Plasmodium vivax in the southern Amazon basin and reported as An. strodei are likely to have been from An. arthuri C identified in this study.
authors.affiliation: Departamento de Epidemiologia, Faculdade de Saúde Pública, Universidade de São Paulo, São Paulo-SP, Brazil. bpbourke@usp.br
authors.forename: Brian Patrick
authors.initials: BP
authors.lastname: Bourke
authors.forename: Tatiane Porangaba
authors.initials: TP
authors.lastname: Oliveira
authors.forename: Lincoln
authors.initials: L
authors.lastname: Suesdek
authors.forename: Eduardo Sterlino
authors.initials: ES
authors.lastname: Bergo
authors.forename: Maria Anice Mureb
authors.initials: MA
authors.lastname: Sallum
mesh_terms: D000818:Animals; D000852:Anopheles / Q000145:classification* / Q000235:genetics*; D001938:Brazil; D016000:Cluster Analysis; D058893:DNA Barcoding, Taxonomic / Q000379:methods*; D004772:Entomology / Q000379:methods*; D005260:Female; D006801:Humans; D008297:Male; D008969:Molecular Sequence Data; D058885:Multilocus Sequence Typing; D010802:Phylogeny; D017422:Sequence Analysis, DNA