中文摘要：目前对噬菌体-细菌相互作用机制的了解还很有限。在大肠杆菌中，RapZ调节葡萄糖胺-6-磷酸（GlcN6P）代谢，其形成启动了包括脂多糖（LPS）在内的细菌细胞膜的合成。然而，RapZ对噬菌体侵染能力的作用还有待研究。本研究采用噬菌体喷雾实验，分离到产肠毒素大肠杆菌（ETEC）菌株，该菌株对其特异性裂解噬菌体vB_EcoM_JS09（JS09）具有耐药性。对该菌株进行全基因组分析，发现rapZ基因在227氨基酸处提前停止突变。本文报道了通过rapZ基因突变，抑制了93.5%的噬菌体吸附从而产生抗性。此外，该突变改变了噬菌斑的形态，降低了电镀效率和噬菌体繁殖效率，还降低了噬菌体JS09对ETEC的侵染性。利用扫描电子显微镜分析，发现噬菌体无法吸附是由于RapZ缺陷菌株丧失了受体。LPS谱分析表明，RapZ缺陷菌株通过改变大肠杆菌的LPS谱来抑制噬菌体侵染。将噬菌体JS09与从野生型(WT)菌株提取的LPS预孵育后，侵染被阻断，表明LPS是噬菌体JS09吸附宿主的受体。可见，ETEC通过rapZ基因突变，增加glmS表达并改变了噬菌体受体-LPS谱，从而获得抗噬菌体JS09感染。这些发现为rapZ基因对J609噬菌体高效感染的作用提供了思路。
重要性：噬菌体耐药菌的发展是噬菌体治疗一个具有挑战性的问题，而目前对噬菌体耐药性机制的认识仍然有限。RapZ是rapZ基因编码的RNase衔接蛋白，在革兰氏阳性菌和革兰氏阴性菌中起重要作用。本文报道了一株抗噬菌体的产肠毒素大肠杆菌(ETEC)的全基因组分析，发现rapZ基因发生了一个提前停止突变(E227Stop)。研究显示，rapZ的提前停止突变破坏了噬菌体JS09在ETEC中的感染性。此外，ETEC通过rapZ基因突变，增加glmS的表达，改变噬菌体受体-LPS谱，从而对JS09噬菌体的吸附和感染产生抗性。本文首次报道RapZ对JS09噬菌体的有效感染是至关重要的。
外文摘要：Our understanding of the mechanisms underlying phage-bacterium interactions remains limited. In Escherichia coli, RapZ regulates glucosamine-6-phosphate (GlcN6P) metabolism, the formation of which initiates synthesis of the bacterial cell envelope, including lipopolysaccharides (LPS). However, the role of RapZ, if any, on phage infectivity remains to be investigated. Here, we isolated strains of enterotoxigenic E. coli (ETEC) resistant to its specific lytic bacteriophage vB_EcoM _JS09 (JS09) in a phage aerosol spray experiment. Whole-genome analysis of phage-resistant bacteria revealed the rapZ gene acquired a premature stop mutation at amino acid 227. Here, we report that the mutation in the rapZ gene confers resistance by inhibiting 93.5% phage adsorption. Furthermore, this mutation changes the morphology of phage plaques, reduces efficiency of plating and phage propagation efficiency, and impairs the infectivity of phage JS09 against ETEC. Using scanning electron microscopy assays, we attribute the inability of the phage to adsorb to the loss of receptors in strains with defective RapZ. Analysis of the LPS profile shows that strains with defective RapZ inhibit phage infection by changing the LPS profile in E. coli. Preincubation of phage JS09 with LPS extracted from a wild-type (WT) strain blocked infection, suggesting LPS is the host receptor for phage J509 adsorption. Our data uncover the mechanism by which ETEC resists infection of phage JS09 by mutating the ropZ gene and then increasing the expression of glmS and changing the phage receptor-LPS profile. These findings provide insight into the function of the rapZ gene for efficient infection of phage J609.
   IMPORTANCE The development of phage-resistant bacteria is a challenging problem for phage therapy. However, our knowledge of phage resistance mechanisms is still limited. RapZ is an RNase adaptor protein encoded by the rapZ gene and plays an important function in Gram-positive and Gram-negative bacteria. Here, we report the whole-genome analysis of a phage-resistant enterotoxigenic Escherichia coli (ETEC) strain, which revealed that the rapZ gene acquired a premature stop mutation (E227Stop). We show that the premature stop mutation of rapZ impairs the infectivity of phage JS09 in ETEC. Furthermore, our findings indicate that ETEC becomes resistant against the adsorption and infection of phage JS09 by mutating the rapZ gene, increasing the expression of glmS, and changing the phage receptor-LPS profile. It is also first reported here that RapZ is essential for efficient infection of phage JS09.
外文关键词：rapZ；lytic bacteriophage；phage resistance；infectivity；enterotoxigenic E. coli；LPS；glmS
作者：Zhou, Y；Bao, HD；Zhang, H；Pang, MD；Zhu, SJ；Wang, R
作者单位：Jiangsu Acad Agr Sci
期刊名称：MSPHERE
期刊影响因子：4.282
出版年份：2021
出版刊次：2
点击下载：一个自发的rapZ突变体降低了裂解性噬菌体vB EcoM JS09对产肠毒素大肠杆菌的感染性