Comparative Genomics of the Herbivore Gut Symbiont Lactobacillus reuteri Reveals Genetic Diversity and Lifestyle Adaptation. Limosilactobacillus reuteri

Lactobacillus reuteri is a catalase-negative, Gram-positive, non-motile, obligately heterofermentative bacterial species that has been used as a model to describe the ecology and evolution of vertebrate gut symbionts. However, the genetic features and evolutionary strategies of L. reuteri from the gastrointestinal tract of herbivores remain unknown. Therefore, 16 L. reuteri strains isolated from goat, sheep, cow, and horse in Inner Mongolia, China were sequenced in this study. A comparative genomic approach was used to assess genetic diversity and gain insight into the distinguishing features related to the different hosts based on 21 published genomic sequences. Genome size, G + C content, and average nucleotide identity values of the L. reuteri strains from different hosts indicated that the strains have broad genetic diversity. The pan-genome of 37 L. reuteri strains contained 8,680 gene families, and the core genome contained 726 gene families. A total of 92,270 nucleotide mutation sites were discovered among 37 L. reuteri strains, and all core genes displayed a Ka/Ks ratio much lower than 1, suggesting strong purifying selective pressure (negative selection). A highly robust maximum likelihood tree based on the core genes shown in the herbivore isolates were divided into three clades; clades A and B contained most of the herbivore isolates and were more closely related to human isolates and vastly distinct from clade C. Some functional genes may be attributable to host-specific of the herbivore, omnivore, and sourdough groups. Moreover, the numbers of genes encoding cell surface proteins and active carbohydrate enzymes were host-specific. This study provides new insight into the adaptation of L. reuteri to the intestinal habitat of herbivores, suggesting that the genomic diversity of L. reuteri from different ecological origins is closely associated with their living environment.

罗伊氏乳杆菌（Lactobacillus reuteri）是一种过氧化氢酶阴性、革兰氏阳性、无运动性的专性异型发酵细菌，曾作为模式物种用于研究脊椎动物肠道共生菌的生态学与演化过程。然而，目前学界对分离自草食动物胃肠道的罗伊氏乳杆菌的遗传特征与演化策略仍缺乏系统认知。为此，本研究针对采集自中国内蒙古地区山羊、绵羊、奶牛与马匹的16株罗伊氏乳杆菌开展全基因组测序，并结合已公开的21株罗伊氏乳杆菌基因组序列，采用比较基因组学方法解析其遗传多样性，揭示不同宿主来源菌株的特异性特征。不同宿主来源的罗伊氏乳杆菌菌株在基因组大小、G+C含量及平均核苷酸一致性（average nucleotide identity, ANI）等指标上均存在显著差异，提示该物种具有丰富的遗传多样性。本研究纳入的37株罗伊氏乳杆菌的泛基因组（pan-genome）共包含8680个基因家族，核心基因组（core genome）则包含726个基因家族。37株菌株中共鉴定到92270个核苷酸突变位点，且所有核心基因的Ka/Ks比值均远小于1，表明其经历了强烈的纯化选择（负选择）压力。基于核心基因构建的高置信度最大似然系统发育树显示，草食动物来源的罗伊氏乳杆菌分离株可分为三个进化枝：进化枝A与进化枝B包含绝大多数草食动物分离株，且与人类来源分离株亲缘关系更近，与进化枝C则差异显著。部分功能基因可能与草食动物、杂食动物及酸面团（sourdough）来源菌群的宿主特异性特征相关。此外，编码细胞表面蛋白与活性碳水化合物酶的基因数量同样呈现宿主特异性差异。本研究为解析罗伊氏乳杆菌对草食动物肠道生境的适应性机制提供了新视角，同时表明不同生态来源的罗伊氏乳杆菌基因组多样性与其生存环境紧密相关。