Infection mechanism of E.coli 0157 to native human colon epithelial cells

本课题旨在阐明肠出血性大肠杆菌O157对人肠上皮细胞的感染机制。1)建立了人结肠上皮细胞的原代培养。2)大肠杆菌O157侵入天然结肠上皮细胞的程度与肠沙门氏菌相当，这一点已被庆大霉素杀死细胞外细菌所证明。侵袭依赖于细胞骨架和细胞表面胆固醇含量的作用。3)共焦荧光显微镜和电镜形态学观察证实了细胞内细菌的存在，并进行了庆大霉素保护试验。4)大肠杆菌O157的靶细胞表达上皮标记物，但不表达单核细胞标记物。5)当对另一种上皮细胞Hela进行大肠杆菌0157和沙门氏菌感染检测时，后者侵入结肠细胞的程度相似，而前者则没有。6)利用细菌消隐位点的等基因缺失突变体，证明sepL对细菌的侵袭活性是必需的，而eaeA、espA和Tir不是必需的。

The purpose of this project is to elucidate the infection mechanism of enterohemorrhagic E. coli O157 to human intestinal epithelial cells.1) The primary culture of native epithelial cells from human colon has been established.2) E. coli O157 has been shown to invade the native colon epithelial cells to a comparable extent with Salmonella enterica as proved by killing the extracellular bacteria by gentamicin. The invasion was dependent on the action of cytoskeletons and cell surface cholesterol content.3) The morphological insights using con focal fluorescent microscopy and electromicroscopy confirmed the intracellular bacteria, in addition to gentamicin protection assay.4) The target cells of E. coli O157 were shown to express epithelial markers but not monocyte markers.5) When another type of epithelial cell, Hela was tested for the infection of E. coli 0157 and salmonella, the latter invaded to a similar degree of colon cells but not the former.6) Employing the isogenic deletion mutants of bacterial effacement locus, the sepL was proved to be essential for the invasion activity, whereas not was eaeA, espA or Tir.