基于rpsD功能基因测序分析家族成员肠道拟杆菌群落组成及其跨代传递

Amplicon sequencing of rpsD reveals the community composition and transgenerational transmission of Bacteroides in family members

摘要:

人体肠道专性共栖细菌在维持微生态平衡方面发挥着关键作用，一般认为通过母乳喂养实现跨代垂直传递。然而，与双歧杆菌(Bifidobacterium)相比，拟杆菌(Bacteroides)作为人体专性共栖菌的代表，其在人群中的传递、扩散机制鲜有报道，且在家族成员中菌群的发生与跨代传递研究较少。目的 探究家族成员间拟杆菌的垂直传递及共发生模式，揭示人体肠道微生物组的构建机制，为肠道微生物健康干预和个性化调控提供理论基础。方法 采集中国新疆3-4代家族共50位家族成员的粪便样本，基于拟杆菌rpsD基因的高通量测序技术构建数据库，通过比对注释，在种水平和特征序列(amplicon sequence variant, ASV)水平上解析家族拟杆菌群落组成和多样性。结果 4个家族共注释拟杆菌16个种、3 704个ASVs，其中4个家族共有的ASVs数量为1 293。ASVs数量最多的5个种为脆弱拟杆菌(B. fragilis，653)、卵形拟杆菌(B. ovatus，619)、单形拟杆菌(B. uniformis，507)、粪拟杆菌(B. caccae，463)和芬氏拟杆菌(B. finegoldii，314)，这5个种也是相对丰度和发生率最高的。家族间拟杆菌菌群存在差异，以B. fragilis、B. uniformis和毛丝鼠粪拟杆菌(B. faecichinchillae)的差异最为显著。不同性别和年龄段家族成员的α多样性无显著差异(P>0.05)。基于Bray-Curtis距离的β多样性分析显示，性别与年龄组间差异不明显，但家族间差异显著(P=0.001)。基于Bray-Curtis距离和ASVs共有率分析表明，不同社会关系家族成员之间的拟杆菌群落组成相似度为母-子、兄弟姐妹对明显高于父-子、夫-妻和无亲缘关系成员。结论 拟杆菌的群落结构和多样性具有家族人群特征，家族间差异显著；依据社会关系分析，母子和兄弟姐妹间的菌群相似性最高，支持在菌株水平上的跨代垂直传递，未来需要采用菌株分离与宏基因组深度测序相结合的大数据方法进一步验证。

Abstract:

Obligate symbiotic bacteria in the human gut play a key role in maintaining microecological balance and are generally thought to be transmitted vertically across generations through breastfeeding. However, compared with that on Bifidobacterium, there is limited literature on the transmission and diffusion mechanisms of Bacteroides, a representative of obligate symbiotic microbiota in the human body, within populations. Additionally, the occurrence and cross-generational transmission of Bacteroides within families remain poorly understood.Objective We explored the vertical transmission and co-occurrence patterns of Bacteroides among family members, aiming to reveal the assembly mechanism of gut microbiota in the human body and provide a theoretical basis for the recommendation of microbiome-based interventions and the realization of personalized gut regulation.Methods A high-throughput sequencing dataset for Bacteroides-specific rpsD in fecal samples from four families with 50 members in 3-4 generations in Xinjiang, China was established. Then, the composition and diversity of Bacteroides communities among different families and member groups were measured at the species level and amplicon sequence variant (ASV) level by comparison and annotation.Results A total of 16 Bacteroides species and 3 704 ASVs were identified, of which 1 293 ASVs were common among the four families. The five species with the highest number of ASVs were B. fragilis (653), B. ovatus (619), B. uniformis (507), B. caccae (463), and B. finegoldii (314), which were also the five species with the highest relative abundance and prevalence. There were significant differences in community composition and abundance of Bacteroides among families, with B. fragilis, B. uniformis, and B. faecichinchillae being the most significant representatives. There was no significant difference in alpha or beta diversity among family members grouped according to gender and age (P>0.05). By contrast, the beta diversity analysis based on Bray-Curtis distance showed differences between families (P=0.001). According to the sharing rate of ASVs and Bray-Curtis distance, the Bacteroides strain similarity of mother-child and siblings was significantly higher than that of father-child, couple, and unrelated members.Conclusion The community structure and diversity of Bacteroides were characterized by family convergence, with significant differences among families. The similarity of Bacteroides strains was the highest within the mother-child and sibling groups among different social relationship groups, supporting transgenerational vertical transmission at the strain level. The conclusion remains to be validated by the combination of the strain isolation method and metagenomic sequencing.