Molecular variation and phylogeny of the Anopheles minimus complex (Diptera: Culicidae) inhabiting Southeast Asian countries, based on ribosomal DNA internal transcribed spacers, ITS1 and 2, and the 28S D3 sequences.

Anopheles minimus (Theobald) is one of the most important vectors of human malaria in Southeast Asia. Morphological studies now have revealed five sibling species as its complex, designated as species A to E. The present study investigated the genetic divergence among An. minimus populations from four countries (Japan, China, Thailand and Indonesia), based on the DNA sequences data of the D3 (the third domain of the 28S ribosomal gene) and ITS2 (the second internal transcribed spacer of the ribosomal gene) is reported. The D3 and ITS2 phylogenetic trees, and the electrophoretic profile of ITS1 (the first internal transcribed spacer of the ribosomal gene) indicated that our An. minimus populations are comprised of three groups: the Japanese population as group I, the population from Guangxi Province of China (GX population) as group II, and others, as group III. The results showed the morphological similarity of group III and GX with the species complex A and B, respectively. It is interesting that both two species A (YN population) and species B (GX) occur in China, and that both species, An. minimus species A (LB-95 population) and the closer population An. flavirostris (Ludlow) (LB-00 population) appeared to be present on the Lombok Island of Indonesia, although in far separated localities. Moreover, this molecular evidence confirms the previous suggestion that the population from the Ishigaki Island of Japan should be classified as a new genetic status species E.