中华按蚊幼虫肠道细菌宏基因组的组成研究

Taxonomic Composition of Metagenomic Community in the Larval Gut of Mosquito Anopheles sinensis (Diptera ∶ Culicidae)

目的了解野外采集的中华按蚊幼虫肠道细菌的多样性。方法应用高通量测序技术测定采自上海嘉定稻田(分成两组,即L1和L2)和海南文昌渗出小积水(AS)的中华按蚊幼虫肠道细菌16S rDNA V4变异区序列,应用Qiime和Mothur等软件整理和统计样品序列数目和操作分类单元(OTUs)数量,分析物种的丰度、分布和Alpha多样性,以及物种丰度的差异性。结果本研究获得用于分析的序列和OTU数为253 724/3 930(L1)、225 203/4 312(L2)和73 990/2 380(AS);稀疏曲线表明测序深度充分,OTU的数量接近实际情况。L1、L2和AS等3个样品的丰富度指数分别为5 942.61/6 534.88、6 328.17/7 235.89和4 228.66/5 651.20,多样性指数分别为4.63/0.03、5.10/0.02和0.14/3.94。中华按蚊幼虫肠道优势菌种均隶属变形菌门,分别占87%(AS)和90%(L),除此之外,共有的优势门还有厚壁菌门、拟杆菌门和放线菌门。比较样品L与AS的细菌丰度,发现除变形菌门和异常球菌-栖热菌门外,其余18门均差异显著;样品L1和L2相比,仅9个门差异显著。结论获得了稻田和海边渗出水环境中华按蚊幼虫肠道细菌的均匀度、丰富度和菌群结构。

Objective To investigate the bacteria diversity in larval gut of field-collected Anopheles sinensis. Methods The 16S rDNA V4 region of An. sinensis larvae collected from paddy on Jiading District of Shanghai （L1/L2）and small seeping water on Wenchang City of Hainan（AS） was sequenced by high-throughput pyrosequencing. Using Qiime and Mothur softwares, the number of sequences and operational taxonomic units（OTUs） for each sample was sorted and calculated, the species abundance and distribution, Alpha diversity index and difference times of species abundance among samples were analyzed. Results The number of sequences and OTUs for each sample were 253 724/3 930（L1）, 225 203/4 312 （L2） and 73 990/2 380 （AS）. The rarefaction curves showed that adequate sampling was achieved. The number of OTUs was close to actual situation. The value of richness index was 5 942.61/6 534.88（L1）, 6 328.17/7 235.89（L2） and 4 228.66/5 651.20 （AS）; diversity index was 4.63/0.03 （L1）, 5.10/0.02 （L2） and 0.14/3.94 （AS）. The dominant species of An. sinensis larvae gut microbiota all belonged to the phylum Proteobacteria, with a percentage of 87%（AS） and 90% （L）. In addition, the dominant phyla among them were Firmicutes, Bacteroidetes and Actinobacteria. The comparison of bacterial abundance between L and AS showed that there were 18 phyla with significant difference, except the Proteobacteria and Deinococcus-Thermus; only 9 phyla were different significantly between L1 and L2. Conclusion Evenness and richness of bacteria flora in the An. sinensis larvae gut collected from paddy and small seeping waters were obtained.