基于性别二分类的斑须蝽成虫肠道微生物群落异性

Gut microbiota differences between female and male adults of Dolycoris baccarum

摘要:

昆虫肠道微生物与宿主之间存在相互依赖的共生关系，这对宿主昆虫的代谢、免疫、发育及农药抗性等功能发挥着重要作用。肠道微生物受到食物、性别和饲养环境等因素的影响，目前尚未见针对植食性昆虫斑须蝽(Dolycoris baccarum)雌雄之间群落结构的相关报道。目的 探究性别对斑须蝽微生物组成及丰度的影响，以挖掘性别相关的微生物，为利用不同性别的微生物防治斑须蝽提供依据。方法 基于PacBio平台，采用第三代16S rRNA基因扩增子测序技术和生物信息学分析，研究斑须蝽雌性和雄性肠道内细菌的多样性及组成。使用传统细菌分离培养法获取斑须蝽的肠道菌，并结合形态学及16S rRNA基因序列分析进行鉴定。结果 斑须蝽共生菌种类丰富，共注释到14门29纲60目109科165属。从门(phylum)、纲(class)、目(order)、科(family)、属(genus)、种(species) 6个微生物分类水平分析可知，只有奇异球菌门(Deinococcota)和奇异球菌纲(Deinococci)是雄性特有的；而绿屈挠菌门(Chloroflexi)、脱硫杆菌门(Desulfobacterota)、蓝菌门(Cyanobacteria)、色球藻纲(Cyanobacteriia)、乳杆菌属(Lacticaseibacillus)、肌氨酸酐水解谷氨酸杆菌(Glutamicibacter creatinolyticus)、不动杆菌(Acinetobacter)和类干酪乳杆菌(Lacticaseibacillus paracasei)则是雌性特有的。研究发现，在不同分类阶元中，雄性在黏质沙雷氏菌(Serratia marcescens)中的数量显著高于雌性。在雌雄肠道中，γ-变形菌纲(Gammaproteobacteria)的含量分别为51.223%和95.512%；肠杆菌目(Enterobacterales)的占比分别为49.784%和95.492%；耶尔辛氏菌科(Yersiniaceae)的占比分别为2.385%和10.377%；沙雷氏菌属在雌性和雄性中的占比分别为2.383%和10.372%；黏质沙雷氏菌在雌性和雄性中的占比分别为2.310%和10.375%。经过形态学和16S rRNA基因鉴定的28株体外分离的菌均属于沙雷氏菌属(Serratia)。结论 斑须蝽雌雄成虫的肠道微生物组成存在显著差异。雄性成虫的肠道微生物多样性显著高于雌性成虫，并且表现出性别二态性。通过体外分离的菌株均属于沙雷氏菌属。这为研究斑须蝽雌雄成虫的肠道微生物潜在功能及斑须蝽的生物防治提供了理论支持。

Abstract:

Insects have a mutually dependent symbiotic relationship with their gut microbiota, which plays an important role in the insect metabolism, immunity, development, and pesticide resistance. Gut microbiota is influenced by factors such as diet, sex, and rearing environment. Currently, little is known about the gut microbiota differences between males and females of the phytophagous insect Dolycoris baccarum.Objective To study the effects of sex on the composition and abundance of gut microbiota in D. baccarum, explore sex-related microorganisms, and provide a basis for utilizing different sex-associated strains in the biocontrol of D. baccarum.Methods The PacBio platform was used for third-generation 16S rRNA gene amplicon sequencing and the bioinformatics analysis was performed to reveal the diversity and composition of gut microbiota in male and female D. baccarum. The bacterial isolation and culture method was employed to obtain gut microbiota strains from D. baccarum, which were then identified by morphological analysis and 16S rRNA gene sequencing.Results The gut microbiota of D. baccarum was diverse, including a total of 165 genera belonging to 109 families, 60 orders, 29 classes of 14 phyla. The gut microbiota was analyzed at six taxonomic levels (phylum, class, order, family, genus, and species). Only the phylum Deinococcota and the class Deinococci were specific to males, while the phyla Chloroflexi, Desulfobacterota, and Cyanobacteria, the class Cyanobacteriia, the genus Lacticaseibacillus, and the species Glutamicibacter creatinolyticus, Acinetobacter, and Lacticaseibacillus paracasei were specific to females. The relative abundance of Serratia marcescens was significantly higher in males than in females. In the guts of females and males, the relative abundance of Gammaproteobacteria, Enterobacterales, Yersiniaceae, Serratia, and S. marcescens was 51.223% and 95.512%, 49.784% and 95.492%, 2.385% and 10.377%, 2.383% and 10.372%, and 2.310% and 10.375%, respectively. All 28 strains isolated in vitro and identified based on morphological characteristics and 16S rRNA sequences belonged to Serratia.Conclusion There are significant differences in the gut microbiota composition between male and female D. baccarum. The diversity of gut microbiota in male adults of D. baccarum is significantly higher than that in female adults. All strains isolated belong to Serratia. This study provides theoretical support for investigating the potential functions of gut microbiota in male and female D. baccarum as well as for the biocontrol of D. baccarum.