关键词: 致倦库蚊, 细菌多样性, 16S rDNA, 血餐, 乙脑病毒, 抗菌肽

Abstract: 【Aim】 To study the midgut bacterial diversity and difference in Culex quinquefasciatus adults before and after feeding with blood meal and infection with the Japanese encephalitis virus (JEV), then further reveal the relationship between C. quinquefasciatus and its midgut bacteria, as well as the reasons for the changes of the midgut bacteria during blood digestion and viral infection.【Methods】 The bacterial genome DNA of the midguts from C. quinquefasciatus adults fed with sugar water (10% sucrose solution), blood meal without JEV and blood meal with JEV was extracted. The 16S rDNA highly variable region sequence of the midgut bacteria was amplified by PCR, sequenced using the Illumina NovaSeq6000 sequencing platform and analyzed by bioinformatics. The total RNA was extracted from the midguts of C. quinquefasciatus adults fed with blood meal without JEV and blood meal with JEV, and the expression levels of the antimicrobial peptide (AMP) genes CqAttacin, CqCecropin, CqCecropin A, CqDefensin A and CqDefensin C in the midgut were detected by RT-qPCR.【Results】 There were 204 amplicon sequence variants (ASVs) in the midgut bacteria of C. quinquefasciatus adults before and after blood meal, belonging to 4 phyla, 9 classes, 17 orders, 27 families and 36 genera, and 120 ASVs in the midgut bacteria before and after infection with JEV, belonging to 6 phyla, 13 classes, 23 orders, 38 families and 47 genera. At the genus level, Elizabethkingia was widely distributed in the midgut bacteria as a dominant genus. The abundance of Enterobacter in the midgut of C. quinquefasciatus adults was the highest on the day 2 after blood meal. Morganella, Rhizobium and Bordetella mainly existed in the midgut of C. quinquefasciatus adults infected with JEV. The difference in the metabolic pathways of the midgut bacteria between C. quinquefasciatus adults on the day 2 after being fed with sugar water and blood meal was the largest. The main differential metabolic pathways were the phosphotransferase system, biosynthesis of siderophore group nonribosomal peptides, N-glycan biosynthesis, fatty acid biosynthesis and the metabolic pathways of amino acids. There were differences in the midgut microbiota between C. quinquefasciatus adults infected with and without JEV, and the midgut bacterial abundance of C. quinquefasciatus adults was higher after JEV infection. Blood meal upregulated the expression levels of the AMP genes CqAttacin, CqCecropin, CqCecropin A, CqDefensinA and CqDefensin C in the midgut compared with the control group fed with sugar water, while infection with JEV significantly downregulated the expression levels of the AMP genes CqCecropin A, CqDefensin A and CqDefensin C in the midgut of C. quinquefasciatus adults compared with blood meal without JEV.【Conclusion】 Both the blood meal and JEV infection lead to the changes of midgut bacteria of C. quinquefasciatus adults. The former shows that the blood digestion process is selective for the midgut bacteria, while the latter may be related to the downregulation of the expression levels of AMP genes in the midgut of C. quinquefasciatus adults caused by JEV infection.

Key words: Culex quinquefasciatus, bacterial diversity, 16S rDNA, blood meal; Japanese encephalitis virus, antimicrobial peptide