The Hyrcanus group of Anopheles consists of many related species, of which An. sinensis, An. lesteri, and An. anthropophagus are known as malaria vector species. It is not possible to identify these species morphologically in the adult and larval stages. Nucleotide sequence alignment of 2nd internal transcribed spacer (ITS2) regions from 4 specimens of An. lesteri collected in Japan, 2 specimens of An. anthropophagus collected in China, and 1 specimen of An. sinensis collected in Korea were sequenced and compared. Sequences of ITS2 regions varied only at 4 sites between An. lesteri and An. anthropophagus, and individual variations among each An. lesteri and An. anthropophagus were found at 5 and 7 sites, respectively, whereas sequences varied at 161 sites between An. lesteri and An. sinensis. This molecular evidence strongly supports that An. lesteri from Japan and An. anthropophagus from China are the same species.