艰难梭菌(Clostridium difficile)通过分泌两种毒素蛋白（TcdA/B）致病，是全球范围导致抗生素相关腹泻和假膜结肠炎的主因。作为严重威胁公共生命安全的感染性疾病，艰难梭菌通过毒素侵染的机制还不十分清楚，这严重阻碍了有效治疗手段的研发。TcdB被证明是艰难梭菌致病的主要原因，而且它是通过受体介导进入到宿主细胞内部致病的，然后人们对该毒素受体几十年的寻找至今一无所获。我们计划利用shRNAmir基因沉默文库以及我们刚研发成功的基于CRISPR/Cas9系统的sgRNA基因敲除文库（Nature 2014），筛选TcdB的宿主细胞受体蛋白。我们初步结果已经获得候选的毒素受体，我们将使用多种手段证明寻找到的受体，以及继续筛选鉴定存在的第二个受体。这项研究将深化人们对艰难梭菌引起的感染性腹泻的发病机理的认识，并对发展新型治疗手段提供新的药物作用靶点。

As a gram-positive, spore-forming anaerobic bacillus, Clostridium difficile is responsible for severe and fatal pseudomembranous colitis, and poses the most urgent antibiotic resistance threat worldwide. Epidemic C. difficile is the leading cause of antibiotic-associated diarrhoea globally, especially diarrhoea due to the emergence of hypervirulent strains associated with high mortality and morbidity. C. difficile causes disease is mainly through its secretion of two toxic proteins: TcdA and TcdB. TcdB is proved to be the major virulence factor, which enters host cells through a poorly understood mechanism to elicit its pathogenic effect. We propose to identify the cellular receptor for TcdB through both shRNAmir library gene silencing screening and sgRNA library gene knockout screening, a newly developed technique by us using CRISPR/Cas9 system (Nature 2014). We have already obtained a candidate TcdB receptor, and we will finish the validation and characterization of this important target. Meanwhile, we propose to screen for the second receptor that also functions to mediate TcdB endocytosis. The success of this study will help deepen our understanding on the infectious mechanism of C. difficile, and provide insights for the development of host-oriented countermeasures against this deadly disease.