昆虫病原线虫共生菌对二化螟体内微生物群落结构的影响

doi:10.13560/j.cnki.biotech.bull.1985.2025-1150

摘要/Abstract

摘要：

目的明确昆虫病原线虫共生菌—嗜线虫致病杆菌（Xenorhabdus nematophila）对二化螟幼虫的致病力，解析该菌对幼虫体内微生物群落结构的影响，以揭示其致病机制，为开发新型微生物杀虫剂提供理论依据。方法设置小卷蛾斯氏线虫自主寄生（SC组）和嗜线虫致病杆菌HNS01微针注射（XN组）2种处理，生理盐水为对照（CA、CB组）。以二化螟3龄幼虫为对象，测定杀虫活性。收集各组幼虫组织进行宏基因组测序，分析幼虫体内微生物群落结构及基因功能。结果 SC组和XN组对二化螟3龄幼虫致死率都达100%，致死时间分别为48 h和96 h。Alpha多样性显示，SC组Shannon指数显著（P<0.05）低于对照组CA，4组Simpson指数无显著变化，SC组Ace和Chao1指数极显著（P<0.01）高于CA组。Beta多样性显示，4组样本在坐标空间中既有重叠又有分离部分。在物种组成上，SC组和XN组的嗜线虫致病杆菌丰度最高。LEfSe分析发现CA组和SC组差异19个种，CB组和XN组差异8个种。KEGG注释表明，SC组和XN组在ABC转运蛋白和双组分系统等通路富集。CAZy注释表明，SC组和XN组令CBM50、GT4、GT2等酶基因的相对丰度升高，而AA3、GH16、GH31等酶基因的相对丰度降低；GH22和GH13酶基因的相对丰度，SC组使其上调，而XN组使其下调。结论嗜线虫致病杆菌HNS01具有杀二化螟3龄幼虫活性，其成功定殖于幼虫体内，并主导宿主微生物群落，通过增强营养利用与环境适应、抑制寄主免疫应答等发挥致病作用。

关键词: 二化螟, 昆虫病原线虫共生菌, 嗜线虫致病杆菌, 昆虫病原线虫, 小卷蛾斯氏线虫, 宏基因组

Abstract:

Objective This study clarified pathogenicity of Xenorhabdus nematophila, a mutualistic bacterium from entomopathogenic nematodes, to the larvae of Chilo suppressalis. We analyzed the changes in microbial community structure in larvae caused by this bacterium, in order to reveal their pathogenic mechanisms and provide theoretical basis for the development of new microbial insecticides. Method Two treatments were set: Natural infection by Steinernema carpocapsae (SC group) and microneedle injection of X. nematophila HNS01 (XN group), with physiological saline as the control (CA and CB groups). Using 3rd-instar larvae of C. suppressalis as the object, insecticidal activity was assessed. Larvae tissues from each group were collected for metagenomic sequencing to analyze the microbial community structure and gene function. Result The mortality rates of SC group and XN group against 3rd-instar larvae of C. suppressalis were both 100%, with lethal times of 48 h and 96 h, respectively. Alpha diversity showed that Shannon index of SC group was significantly (P<0.05) lower than that of control group CA. There was no significant change in the Simpson index of four groups. Ace and Chao1 indices of SC group were extremely significantly (P<0.01) higher than CA group. Beta diversity showed that the four groups of samples had both overlapping and separated parts in the coordinate space. In terms of species composition, SC group and XN group had the highest abundance of X. nematophila. LEfSe analysis revealed 19 differences between CA and SC groups, and 8 differences between CB and XN groups. KEGG annotation indicated that SC group and XN group are enriched in pathways such as ABC transporter and two-component system. CAZy annotation indicated that SC and XN groups increased the relative abundances of enzyme genes such as CBM50, GT4, and GT2, while the relative abundances of enzyme genes such as AA3, GH16, and GH31 decreased. The relative abundances of GH22 and GH13 enzyme genes were upregulated in SC group, but downregulated in XN group. Conclusion X. nematophila HNS01 has activity of killing 3rd-instar larvae of C. suppressalis. It successfully colonizes the larvae and dominates the host microbial community, exerting pathogenic effects by enhancing nutrient utilization and environmental adaptation, as well as inhibiting host immune response.

Key words: Chilo suppressalis, mutualistic bacteria from entomopathogenic nematodes, Xenorhabdus nematophila, entomopathogenic nematodes, Steinernema carpocapsae, metagenomics

朱华珺, 段德勇, 吴胜莲, 周小玲, 方勇, 黄思娣, 刘明星, 刘絮宁, 许隽, 刘洋. 昆虫病原线虫共生菌对二化螟体内微生物群落结构的影响[J]. 生物技术通报, 2026, 42(5): 213-221.

ZHU Hua-jun, DUAN De-yong, WU Sheng-lian, ZHOU Xiao-ling, FANG yong, HUANG Si-di, LIU Ming-xing, LIU Xu-ning, XU Jun, LIU Yang. Effects of Mutualistic Bacteria from Entomopathogenic Nematodes on the Microbial Community Structure in Chilo suppressalis[J]. Biotechnology Bulletin, 2026, 42(5): 213-221.

图/表 8

表1 样品测序数据处理统计结果

Table 1 Statistical results of sample sequencing data processing

样品 Sample	原始测序数据量 Raw base （G）	质控后的数据量 Clean base （G）	Q20 （%）	Q30 （%）	GC （%）
SC1	15.42	14.46	97.88	94.16	41.88
SC2	13.18	12.86	97.19	92.79	41.66
SC3	12.60	12.33	97.60	93.61	39.63
CA1	13.62	13.40	97.75	94.18	38.71
CA2	11.35	11.12	97.75	94.12	38.60
CA3	12.31	12.12	98.55	95.90	37.53
XN1	14.60	14.39	97.78	94.11	37.45
XN2	12.55	12.33	97.91	94.45	41.44
XN3	15.33	14.91	98.03	94.73	39.92
CB1	12.81	12.59	97.68	93.87	38.38
CB2	13.89	13.66	97.54	93.54	38.17
CB3	12.75	12.51	97.71	93.95	38.95

表2 二化螟杀虫活性的检测

Table 2 Detection of insecticidal activity of C. suppressalis

处理 Treatment	校正死亡率 Corrected mortality rate （%）
处理 Treatment	24 h	48 h	72 h	96 h
SC	40.65±2.03d	100.00±0.00a	100.00±0.00a	100.00±0.00a
XN	33.33±3.33d	54.81±7.76c	73.06±8.56b	100.00±0.00a

图1 二化螟体内微生物群落Alpha多样性分析A： Shannon指数； B： Simpson指数； C： Ace指数； D： Chao1指数； *表示差异显著（P<0.05）； **表示差异极显著（P<0.01）

Fig. 1 Alpha diversity analysis of microbial community in C. suppressalisA: Shannon index. B: Simpson index. C: Ace index. D: Chao1 index. * indicated significant difference (P<0.05); ** indicated extremely significant difference (P<0.01)

图2 二化螟体内微生物群落Beta多样性分析A： PCA指数； B： PCoA指数

Fig. 2 Beta diversity analysis of microbial community in C. suppressalisA: PCA index; B: PCoA index

图3 二化螟体内微生物的相对丰度A：微生物群落门水平组成；B：微生物群落种水平组成

Fig. 3 Relative abundance of microorganisms in C. suppressalisA: Taxonomic composition of microbial communities at the phylum level. B: Taxonomic composition of microbial communities at the species level

图4 二化螟体内微生物群落的LEfSe分析A：对照组CA和处理组SC的比较；B：对照组CB和处理组XN的比较

Fig. 4 LEfSe analysis of microbial communities in C. suppressalisA: Comparison between control CA and treatment SC. B: Comparison between control CB and treatment XN

图5 二化螟体内微生物群落信号通路的分析A：2级水平上的KEGG注释；B：3级水平上的KEGG注释

Fig. 5 Signaling pathway analysis of microbial communities in C. suppressalisA: KEGG annotations at level 2. B: KEGG annotations at level 3

图6 二化螟体内微生物群落碳水化合物活性酶的分析A：1级水平上的CAZy基因组成分布图；B：2级水平上的CAZy差异功能基因丰度热图

Fig. 6 Carbohydrate active enzyme analysis of microbial communities in C. suppressalisA: Distribution of CAZy gene at level 1. B: Heatmap of differential functional gene abundance in CAZy at level 2

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