| Anopheles lesteri Baisas & Hu 1936, belonging to Anopheles hyrcanus group of the Genus Anopheles, is distributed in Philippines, China, Japan, South Korea, Guam and so on. In terms of medical importance, its higher malaria vector capacity is only in central China. There are different ecological habits and transmission capacity of An. lesteri among geographical distribution in China, through years of investigations. Thus, its population genetic variation needs further research. In this study, anopheline samples were identified by molecular markers, as rDNA-ITS2 and D3. Basing on the research of the microsatellite DNA characteristics of An. lesteri, we used the microsatellite DNA loci to determine the population genetic structure. The accuracy of species identification was improved by molecular markers; the local mosquito information was supplemented and updated certainly. The elaboration of An. lesteri's population genetic variation and divergence should provide theoretical evidence on malaria control strategy. The results were as follows:1. The 126 specimen was collected from Yunnan, Hubei, Guizhou, Henan, Guangdong, Shangdong, Liaoning, Shaanxi and Hainan provinces. The sequence analysis results showed that the species included An.sinensis (n=44), An. vagus (n=22), An. pullus (n=18), An. minimus (n=13), An. lindesayi (n=10), An. kleini (n=4), An. junlianensis (n=4), An. kochi (n=4), An. belenrae (n=3), An. kweiyangensis (n=3) and An. aconitus (n=1).2. Four types of sequences were found in this study, with LL1, LL2, LL3 and LL4, and cannot name. They were most similar with the Hyrcanus group except LL4, and they should be the new members of this group. In addition, there was a double peak in many Anopheles individual sequences, suggesting that these species were during active divergence and speciation recently.3. The characteristics of 11 microsatellite loci were studied by An. lesteri field samples. These results provided new molecular markers for further population genetic structure.4. Two hundred and sixteen An. lesteri samples from China and Korea were detected by 9 microsatellite loci. The closely collecting sites were combined with 6 populations for further analysis.5. The results of genetic variation within population of An. lesteri were as following. The range of expected heterozygosity and observed heterozygosity was from 0.4848 (JS) to 0.7134 (LN) and 0.3309 (JS) to 0.4704 (SK). The number of private allele was highest in of LN (9), lowst in YN (0). The inbreeding coefficient value was from 0.2101 (HN) to 0.5073 (HB), with an average of 0.3508. There were 30 loci deviating from Hardy–Weinberg Equilibrium C. There were 7 loci deviating from HWE in LN populations, but no locus deviating from HWE in SK, YN, HB populations. Linkage disequilibrium test result showed that 38 pairs of loci were associated (P < 0.01), ANL11 and ANL10 were associated in all populations, the others were disordered.6. We removed the population of JS (laboratory colony) and ANL11 when studying genetic variation among population of An. Lesteri. The results were as following. FST value of pair population ranged from -0.0127 (LN-SK) to 0.2539 (YN-HN). The Mantel test showed there was positive correlation between geographic distances with genetic differences among populations. The population genetic structure accord with Isolation-by-distance Model. The AMOVA analysis result showed the percentage of variation within population (86.55%) was greater than among population (13.45%). Bayesian analysis revealed two main clusters, cluster I was included GD and LN populations, and II included SK, HEN, HB, GD, HN and SC populations.7. The analysis results of effective population size and demographic stability showed the HB, LN, HEH and HN populations were recent demographic expansion. Under linkage disequilibrium models, the effective population size ranged from 1.2 (HB) to 33.7 (GD), the overall population size was 44.6, with the 95% confidence interval from 40.6 to 49.0.8. Comprehensive analysis of the An. lesteri's population genetic structure, we can speculate the spread pathway. The ancestors of An. lesteri were in Yunnan, other populations were all spread from there. The pathway was to north and east, the population colonized on the way, and some variations emerged in order to adapt to the local ecological environment. An. lesteri is widely distribution, geographical barriers blocked gene flow in some degree, but the level of genetic divergence among populations was low, the population genetic structure accord with Isolation-by-distance Model. The large quantity usage of insectcide in the distribution region by its medical importance resulted in lower population quantity of An. lesteri. So there were more inbreeding and deviation from HWE.
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