关键词: 松墨天牛, 肠道细菌, 16S rDNA, 群落结构, 功能预测

Abstract:

【Objective】 The aim of this study was to understand the bacterial community structure in the foregut, midgut and hindgut of Monochamus alternatus larvae and to compare the differences in the bacterial diversity and dominant flora among the different intestinal segments to provide a reference for revealing the mechanism of intestinal bacteria when M. alternatus obtains nutrients and overcomes the chemical defense of host plants. 【Method】The gut DNA of five foreguts, midguts and hindguts, repeated three times respectively, of fourth instar larvae of M. alternatus were extracted. The library construction and high-throughput sequencing of the 16S rDNA V3-V4 region of the intestinal bacteria in M. alternatus were performed using Illumina HiSeq techniques. The original sequences were quality controlled and spliced using Trimmomatic software and FLASH software, respectively. The operational taxonomic unit (OTU) clustering of sequences was performed using USEARCH software, the number of OTUs was counted, and Venn diagrams were drawn. The community composition and species richness of each sample were determined at the phylum and genus levels. The alpha and beta diversity were used to reflect the diversity and similarity of the flora in different samples. The functions of the intestinal bacteria in M. alternatus larvae mapped to the KEGG database were predicted using PICRUSt software, and the potential functions of different intestinal bacterial communities were explored. 【Result】A total of 643 404 high-quality sequences were obtained and clustered to 1 614 OTUs with 97% similarity, which were annotated into 35 phyla, 63 classes, 137 orders, 250 families, 554 genera and 844 species. The OTUs were the least abundant in the foregut and were the most abundant in the hindgut. There were similarities and differences in the OTU composition of each segment. Proteobacteria was the most dominant phylum in all segments of the intestine, Gluconobacter was the most dominant genus in the foregut, Serratia was the most dominant genus in the midgut, and Gluconobacter and Serratia were the most dominant genera in the hindgut. The alpha diversity showed that the bacterial communities of the midgut and hindgut were more abundant, whereas the beta diversity showed that the bacterial composition in the three gut segments was different, but was similar in the midgut and hindgut. The bacterial function prediction analysis showed that metabolism was the most abundant function in all the intestinal bacteria, in which carbohydrate metabolism and amino acid metabolism were the main functions, which were concentrated in the midgut and hindgut. 【Conclusion】 There are differences in the bacterial community structures and potential functions of bacteria in different intestinal segments of indoor populations of M. alternatus larvae, which are caused by the differences in the physicochemical properties in different intestinal segments and their digestion functions. Intestinal bacteria and M. alternatus larvae form a symbiotic functional body, which plays an important role in assisting the larvae metabolism, obtaining nutrients and overcoming the chemical defense of host plants. The microbial function in this experiment was based on the prediction results of the PICRUSt software.

Key words: Monochamus alternatus, intestinal bacteria, 16S rDNA, community structure, function prediction