title: Functional constraints and evolutionary dynamics of the repeats in the rDNA internal transcribed spacer 2 of members of the Anopheles barbirostris group.
abstract: BACKGROUND
  The Anopheles barbirostris group is widely distributed in Southeast Asia. Although seven species have been formally described, a molecular analysis of the rDNA ITS2 and the mitochondrial cytochrome oxidase I gene suggests that the group includes species that are morphologically very similar or identical.We have previously shown that species in the Anopheles barbirostris Subgroup have an exceptionally large ITS2 (>1.5 kb), greater than in any other Anopheline group. However, the molecular processes responsible for generating such a large ITS2 have not previously been explored.
  
  
  METHODS
  To determine the processes by which this large ITS2 is generated, we examined the sequence and secondary structure of the ITS2 of 51 specimens from five species of the Anopheles barbirostris Subgroup. These include the anthropophilic species An. campestris and three morphospecies of the Barbirostris Complex: An. vanderwulpi, An. barbirostris I and III, together with a previously undescribed member of this group (Clade IV).
  
  
  CONCLUSIONS
  All the specimens were found to have an ITS2 greater than 1.5 kb in length. The possibility that the spacer sequences amplified were pseudogenes was examined and discarded. The large size of ITS2 in the species studied is due to the presence of internal repeats of approximately 110 bp in length, confined to the central region of the spacer. Repeats varied markedly between the species examined, with respect to their organization, number and sequence similarity. The nucleotide diversity increased in direct relation to size variation and the presence of non-repeated elements.A secondary structure analysis showed that the repeats form hairpin structures with a wide range of free energy values. These hairpin structures are known to facilitate the subsequent processing of mature rRNA. An analysis of the repeats from the different species suggests they originate from a common ancestor, with the repeats appearing before speciation of the Barbirostris Group.
authors.forename: Claudia Caterina
authors.initials: CC
authors.lastname: Paredes-Esquivel
authors.affiliation: Department of Vector Biology, Liverpool School of Tropical Medicine, Pembroke Place, Liverpool, L3 5QA United Kingdom. htownson@liverpool.ac.uk.
authors.forename: Harold
authors.initials: H
authors.lastname: Townson
mesh_terms: D000818:Animals; D000852:Anopheles / Q000145:classification* / Q000235:genetics; D001210:Asia, Southeastern; D001483:Base Sequence; D004272:DNA, Mitochondrial / Q000737:chemistry / Q000235:genetics; D021903:DNA, Ribosomal Spacer / Q000737:chemistry / Q000235:genetics; D003576:Electron Transport Complex IV / Q000235:genetics; D019143:Evolution, Molecular; D014644:Genetic Variation; D008969:Molecular Sequence Data; D010802:Phylogeny; D012091:Repetitive Sequences, Nucleic Acid / Q000235:genetics; D016415:Sequence Alignment / Q000662:veterinary; D017422:Sequence Analysis, DNA / Q000662:veterinary
chemical_list: D004272:DNA, Mitochondrial; D021903:DNA, Ribosomal Spacer; D003576:Electron Transport Complex IV