Carbohydrates and the oxidative branch of the pentose phosphate pathway modify Bacteroides thetaiotaomicron phage resistance by phase variable S-layers

The human gut microbiota consists of hundreds of bacterial species, some of which persist in the presence of lytic phage that infect them. Bacteroides employ numerous phase-variable strategies to survive in the presence of phage, including capsular polysaccharides (CPS) and S-layer lipoproteins. We previously reported that a Bacteroides thetaiotaomicron strain lacking CPS exhibits almost complete resistance to multiple phages when forced to express the S-layer protein BT1927. However, this strain was only resistant after certain growth conditions, suggesting nutritional variables alter infection and resistance. We grew this strain on various simple sugars and polysaccharides finding that some substrates (fructose, glucose) promote strong resistance to a single phage (ARB25) while others like N-acetylgalactosamine (GalNAc) and mucin O-glycans increase susceptibility. Mixing fructose and GalNAc indicates the effects of GalNAc are dominant. Despite increasing ARB25 susceptibility, GalNAc did not reduce BT1927 transcript or protein levels. Instead, GalNAc reduced the amount of BT1927 displayed on the cell surface and increased outer membrane vesiculation. Mutants in any of the 3 steps of the oxidative branch of the pentose phosphate pathway—grown in fructose—behaved similarly to wild-type cells grown in GalNAc, illuminating this pathway in regulation of sugar-mediated phage-resistance. Despite promoting strong resistance, cells grown in glucose/fructose sometimes displayed sub-populations that appeared to completely lack surface BT1927, suggesting another checkpoint exists to control whether this phage defense is deployed. Finally, we show the mucin sugar GalNAc increases susceptibility to several other phage, which has implications for B. thetaiotaiomicron persistence in niches like the mucus layer.

人类肠道微生物群（human gut microbiota）由数百种细菌物种构成，其中部分类群可在感染自身的裂解性噬菌体（lytic phage）存在下存活。拟杆菌属（Bacteroides）演化出多种相变异策略以在噬菌体存在的环境中存活，相关策略包括荚膜多糖（CPS）与S层脂蛋白。本团队此前报道，缺失荚膜多糖的多形拟杆菌（Bacteroides thetaiotaomicron）菌株经诱导表达S层蛋白BT1927后，对多种噬菌体几乎完全耐药。但该菌株仅在特定生长条件下才表现出耐药性，提示营养变量会改变噬菌体感染过程与宿主的耐药状态。本研究将该菌株接种于多种单糖与多糖培养基中培养，发现部分底物如果糖、葡萄糖可显著增强其对单一噬菌体ARB25的耐药性，而另一些底物如N-乙酰半乳糖胺（GalNAc）与黏蛋白O-聚糖则会提升宿主的噬菌体易感性。将果糖与GalNAc混合培养的实验结果表明，GalNAc的调控效应占主导地位。尽管GalNAc会提升宿主对ARB25的易感性，但它并未降低BT1927的转录本水平或蛋白表达量。相反，GalNAc减少了BT1927在细胞表面的展示量，并增强了外膜囊泡形成。在果糖培养基中培养的、磷酸戊糖途径（pentose phosphate pathway）氧化分支3个步骤中任意一个发生突变的菌株，其表型与在GalNAc中培养的野生型细胞一致，这阐明了该途径在糖介导的噬菌体耐药调控中的关键作用。尽管葡萄糖/果糖培养可显著提升宿主的噬菌体耐药性，但部分细胞亚群似乎完全缺失表面BT1927，提示存在另一调控检查点以管控该噬菌体防御系统的激活。最后，本团队证实黏蛋白来源的糖GalNAc会提升宿主对多种其他噬菌体的易感性，这对多形拟杆菌在黏液层等微生境中的定植与存活具有重要意义。