Watson Debbie, Sleator Roy D, Casey Pat G, Hill Colin, Gahan Cormac G M
Alimentary Pharmabiotic Centre, Department of Microbiology, University College Cork, Western Rd., Cork, Ireland.
Infect Immun. 2009 Nov;77(11):4895-904. doi: 10.1128/IAI.00153-09. Epub 2009 Sep 8.
The food-borne pathogenic bacterium Listeria monocytogenes has the potential to adapt to an array of suboptimal growth environments encountered within the host. The pathogen is relatively bile tolerant and has the capacity to survive and grow within both the small intestine and the gallbladder in murine models of oral infection. We have previously demonstrated a role for the principal carnitine transport system of L. monocytogenes (OpuC) in gastrointestinal survival of the pathogen (R. Sleator, J. Wouters, C. G. M. Gahan, T. Abee, and C. Hill, Appl. Environ. Microbiol. 67:2692-2698, 2001). However, the mechanisms by which OpuC, or indeed carnitine, protects the pathogen in this environment are unclear. In the current study, systematic analysis of strains with mutations in osmolyte transporters revealed a role for OpuC in resisting the acute toxicity of bile, with a minor role also played by BetL, a secondary betaine uptake system which also exhibits a low affinity for carnitine. In addition, the toxic effects of bile on wild-type L. monocytogenes cells were ameliorated when carnitine (but not betaine) was added to the medium. lux-promoter fusions to the promoters of the genes encoding the principal osmolyte uptake systems Gbu, BetL, and OpuC and the known bile tolerance system BilE were constructed. Promoter activity for all systems was significantly induced in the presence of bile, with the opuC and bilE promoters exhibiting the highest levels of bile-dependent expression in vitro and the betL and bilE promoters showing the highest expression levels in the intestines of orally inoculated mice. A direct comparison of all osmolyte transporter mutants in a murine oral infection model confirmed a major role for OpuC in intestinal persistence and systemic invasion and a minor role for the BetL transporter in fecal carriage. This study therefore demonstrates a previously unrecognized function for osmolyte uptake systems in bile tolerance in L. monocytogenes.
食源性病原体单核细胞增生李斯特菌有能力适应宿主内遇到的一系列非最佳生长环境。该病原体对胆汁有相对耐受性，并且在口服感染的小鼠模型中，有能力在小肠和胆囊内存活并生长。我们之前已经证明了单核细胞增生李斯特菌的主要肉碱转运系统（OpuC）在该病原体胃肠道存活中的作用（R. 斯利特、J. 沃特斯、C. G. M. 加汉、T. 阿贝、C. 希尔，《应用与环境微生物学》67:2692 - 2698，2001年）。然而，OpuC或者说肉碱在这种环境中保护病原体的机制尚不清楚。在当前研究中，对渗透溶质转运蛋白发生突变的菌株进行系统分析后发现，OpuC在抵抗胆汁的急性毒性方面发挥作用，次要作用也由BetL发挥，BetL是一种次要的甜菜碱摄取系统，对肉碱也表现出低亲和力。此外，当向培养基中添加肉碱（而非甜菜碱）时，胆汁对野生型单核细胞增生李斯特菌细胞的毒性作用得到缓解。构建了与编码主要渗透溶质摄取系统Gbu、BetL和OpuC以及已知胆汁耐受系统BilE的基因启动子的lux - 启动子融合体。在有胆汁存在的情况下，所有系统的启动子活性均显著诱导，其中opuC和bilE启动子在体外表现出最高水平的胆汁依赖性表达，betL和bilE启动子在口服接种小鼠的肠道中表现出最高表达水平。在小鼠口服感染模型中对所有渗透溶质转运蛋白突变体进行直接比较，证实了OpuC在肠道持续性和全身侵袭中起主要作用，而BetL转运蛋白在粪便携带中起次要作用。因此，本研究证明了渗透溶质摄取系统在单核细胞增生李斯特菌胆汁耐受性方面有此前未被认识到的功能。
