Table_1_CRISPR-Cas systems feature and targeting phages diversity in Lacticaseibacillus rhamnosus strains.DOCX

One of the most important adaptive immune systems in bacteria against phages is clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated (CAS) genes. In this investigation, an approach based on genome mining was employed to characterize the CRISPR-Cas systems of Lacticaseibacillus rhamnosus strains. The analysis involved retrieving complete genome sequences of L. rhamnosus strains, and assessing the diversity, prevalence, and evolution of their CRISPR-Cas systems. Following this, an analysis of homology in spacer sequences from identified CRISPR arrays was carried out to investigate and characterize the range of target phages. The findings revealed that 106 strains possessed valid CRISPR-Cas structures (comprising CRISPR loci and Cas genes), constituting 45% of the examined L. rhamnosus strains. The diversity observed in the CRISPR-Cas systems indicated that all identified systems belonged to subtype II-A. Analyzing the homology of spacer sequences with phage and prophage genomes discovered that strains possessing only CRISPR-Cas subtype II targeted a broader spectrum of foreign phages. In summary, this study suggests that while there is not significant diversity among the CRISPR-Cas systems identified in L. rhamnosus strains, there exists notable variation in subtype II-A systems between L. rhamnosus and other lactobacilli. The diverse nature of these CRISPR-Cas systems underscores their natural activity and importance in adaptive immunity.

细菌抵御噬菌体的关键适应性免疫系统之一，为成簇规律间隔短回文重复序列（clustered regularly interspaced short palindromic repeats，CRISPR）及其相关（CRISPR-associated，CAS）基因。本研究采用基于基因组挖掘的方法，对鼠李糖乳杆菌（Lacticaseibacillus rhamnosus）菌株的CRISPR-Cas系统进行鉴定与表征。本次分析首先获取了所有受试鼠李糖乳杆菌菌株的完整基因组序列，并评估其CRISPR-Cas系统的多样性、流行率与演化历程；随后，针对已鉴定CRISPR基因座中的间隔序列开展同源性分析，以探究并明确其靶向噬菌体的范围。研究结果显示，共有106株菌株携带完整的CRISPR-Cas结构（包含CRISPR基因座与Cas基因），占受试鼠李糖乳杆菌菌株总数的45%。对CRISPR-Cas系统多样性的分析表明，所有鉴定得到的系统均属于II-A亚型。通过将间隔序列与噬菌体及前噬菌体基因组进行同源性比对，发现仅携带CRISPR-Cas II亚型的菌株可靶向更广谱的外源噬菌体。综上，本研究表明：尽管在鼠李糖乳杆菌菌株中鉴定得到的CRISPR-Cas系统整体多样性并不显著，但鼠李糖乳杆菌的II-A亚型系统与其他乳杆菌的同类系统之间存在显著差异。这些CRISPR-Cas系统的多样性特征，凸显了其在适应性免疫中的天然活性与重要意义。