Mucin microcosm culture of complex and defined synthetic gut community

The behavior of microbial communities depends on both taxonomic composition and physical structure. Metagenomic sequencing of fecal samples has revealed the composition of human gut microbiomes, but we remain less familiar with the spatial organization of microbes between regions such as lumen and mucosa, as well as the microbial genes that regulate this organization. To discover the determinants of spatial organization in the gut, we simulate mucosal colonization over time using an in vitro culture approach incorporating mucin hydrogel microcosms with a complex yet defined community of 123 human strains for which we generated high-quality genome assemblies. Tracking strain abundance longitudinally using shotgun metagenomic measurements, we observe distinct and strain-specific spatial organization in our cultures with strains enriched on mucin microcosms versus in supernatant, reminiscent of mucosa versus lumen enrichment in vivo. Our high taxonomic resolution data enables a comprehensive search for microbial genes that underlie this spatial organization. We identify gene families positively associated with microcosm-enrichment, including several known for biofilm and adhesion functions such as efflux pumps, gene expression regulation, and membrane proteases, as well as a novel link between a coenzyme F420 hydrogenase gene family and lipo/exopolysaccharide biosynthesis. Our strain-resolved abundance measurements also demonstrate that incorporation of microcosms yields a more diverse community than liquid-only culture by allowing co-existence of closely related strains. Altogether these findings demonstrate that microcosm culture with synthetic communities can effectively simulate lumen versus mucosal regions in the gut, providing measurements of microbial organization with high taxonomic resolution to enable identification of specific bacterial genes and functions associated with spatial structure.

微生物群落的行为同时由其分类组成与物理结构共同决定。对粪便样本开展宏基因组测序（metagenomic sequencing）已揭示了人类肠道微生物组的组成，但我们对于肠腔（lumen）与黏膜（mucosa）等区域间微生物的空间组织模式，以及调控该组织的微生物基因仍缺乏深入认知。为探明肠道空间组织的决定因素，本研究采用体外培养策略，将黏蛋白水凝胶微培养体系（mucin hydrogel microcosms）与由123株人类来源菌株构成的复杂且定义明确的群落相结合，对黏膜定植过程进行时序模拟，并为这些菌株生成了高质量基因组组装。通过鸟枪法宏基因组测序（shotgun metagenomic sequencing）对菌株丰度进行纵向追踪，我们在培养体系中观察到了显著且具菌株特异性的空间组织模式：相较于上清液（supernatant），菌株在黏蛋白微培养体系中呈现富集状态，这与体内黏膜相较于肠腔的富集特征高度相似。我们的高分类学分辨率数据支持对介导该空间组织的微生物基因开展全面筛选。本研究鉴定出了与微培养体系富集呈正相关的基因家族，包括若干已知与生物膜形成、黏附功能相关的家族，例如外排泵（efflux pumps）、基因表达调控因子以及膜蛋白酶（membrane proteases）；同时还发现了辅酶F420氢化酶（coenzyme F420 hydrogenase）基因家族与脂/外多糖生物合成之间的全新关联。基于菌株分辨率的丰度测量结果还显示，相较于仅使用液体培养基的培养体系，黏蛋白微培养体系的引入可使紧密相关菌株实现共存，进而构建出多样性更高的群落。综上，本研究结果证实，采用合成群落的微培养体系可有效模拟肠道内肠腔与黏膜区域的差异，提供了具备高分类学分辨率的微生物组织特征测量数据，从而能够精准鉴定出与空间结构相关的特定细菌基因及功能。