海南岛按蚊的鉴定和迷走按蚊的进化研究

Identification of anopheline mosquitoes and evolution of Anopheles vagus （Diptera： Culicidae） in Hainan Island

目的阐明海南岛某些按蚊的分类地位，探讨迷走按蚊与其近缘种的亲缘关系。方法对采自海南岛8个地点的按蚊依据形态和分子特征进行鉴别研究，应用的分子特征包括核糖体DNA（ribosomalDNA，rDNA）第2内转录间隔区（second internal transcribed spacer，1TS2）和28S第3编码区（thirddomain，D3）序列，综合分析迷走按蚊及其近缘种的ITS2和D3序列，构建系统发育树。结果本研究共鉴定按蚊407只。其中84．9％的成蚊分子鉴定与依据形态鉴定的结果一致。在228条迷走按蚊的ITS2序列中，部分个体在2个位点存在单碱基套峰（G／A），出现的频率分别为57．9％、64．8％，同时有两个套峰的占样本数的57．0％，其他位点保守。迷走按蚊的D3序列，在种内无碱基差异。结合本研究和文献的迷走按蚊及其近缘种ITS2序列构建的ML树，显示迷走按蚊与浅色按蚊A／C／D亲缘关系近，与另一支的浅色按蚊B、圣代克按蚊复合体的亲缘关系较远。关于未订名种NBC（Anophelessp．NBG）的分类地位，依据分子特征鉴定其为浅色按蚊B，可能是圣代克按蚊复合体新的成员种。结论鉴定形态在个体间变异大的种类时．客观的分子特征更具重要性，迷走按蚊与浅色按蚊MC／D具较近的亲缘关系。

Objective To clarify the taxonomic status of anopheline mosquitoes in Hainan Island and reconstruct the molecular phylogenetic relationship among Anopheles vagus and its cryptic species. Methods Anopheline mosquitoes from 8 sites in Hainan Island were distinguished based on morphology and molecular markers, with ribosomal DNA second internal transcribed spacer （rDNA-ITS2）and the third domain of 28S （28S-D3） sequences. The phylogenetic tree, using maximum likelihood method, was reconstructed by rDNA- ITS2 and 28S-D3 dataset, after the sequences of mosquito species within Pyretophorus series were analyzed. Results Total 407 individuals were identified in this study. The agreement between results of molecular iden- tification and morphology in adults was 84.9%. In 228 ITS2 sequences of A n. vagus, some individuals at two positions exhibitd double-peaks （G/A）, the frequencies were 57.9%, 64.8%, respectively, both for 57%. Other positions were intraspecific conservation. There were no differences of D3 sequences within An. vagus individuals. The ML tree based on ITS2 sequences showed that An. vagus and An. subpictus cytotype A/C/D were close on the same clade, while far away with the other clades with An. subpictus cytotype B and An. sundia-cus complex. Anopheles sp. NBG was An. subpictus cytotype B by ITS2 marker. However, it was also likely a new member of Art. sundiacus complex. Conclusion Molecular marker is more important in distinguishing anopheline mosquitoes species while there are many intraspeeific morphology variations. The relationship between An. vagus and An. subpictus eytotype A/C/D is closer than that between subpictus cytotype B and An. subpictus cytotype A/C/D.