Nicholls L, Grant T H, Robins-Browne R M
Microbiological Research Unit, Department of Microbiology and Infectious Diseases, Royal Children's Hospital, University of Melbourne, Parkville, Victoria 3052, Australia.
Mol Microbiol. 2000 Jan;35(2):275-88. doi: 10.1046/j.1365-2958.2000.01690.x.
Enterohaemorrhagic Escherichia coli (EHEC) are food-borne intestinal pathogens with a low infectious dose. Adhesion of some EHEC strains to epithelial cells is attributed, in part, to intimin, but other factors may be required for the intestinal colonizing ability of these bacteria. In order to identify additional adherence factors of EHEC, we generated transposon mutants of a clinical EHEC isolate of serotype O111:H-, which displayed high levels of adherence to cultured Chinese hamster ovary (CHO) cells. One mutant was markedly deficient in CHO cell adherence, human red blood cell agglutination and autoaggregation. Sequence analysis of the gene disrupted in this mutant revealed a 9669 bp novel chromosomal open reading frame (ORF), which was designated efa1, for EHEC factor for adherence. efa1 displayed 28% amino acid identity with the predicted product of a recently described ORF from the haemolysin-encoding plasmid of EHEC O157:H7. The amino termini of the putative products of these two genes exhibit up to 38% amino acid similarity to Clostridium difficile toxins A and B. efa1 occurred within a novel genetic locus, at least 15 kb in length, which featured a low G+C content, several insertion sequence homologues and a homologue of the Shigella flexneri enterotoxin ShET2. DNA probes prepared from different regions of efa1 hybridized with all of 116 strains of attaching-effacing E. coli (AEEC) of a variety of serotypes, including enteropathogenic E. coli (EPEC) and EHEC, but with none of 91 non-AEEC strains. Nevertheless, efa1 was not required for the attachment-effacement phenotype, and the efa1 locus was not physically linked to the locus for enterocyte effacement (LEE) pathogenicity island, which is responsible for this phenotype in EPEC. These findings suggest that efa1 encodes a novel virulence-associated determinant of AEEC, which contributes to the adhesive capacity of these bacteria.
肠出血性大肠杆菌（EHEC）是食源性肠道病原体，感染剂量低。一些EHEC菌株对上皮细胞的黏附部分归因于紧密黏附素，但这些细菌的肠道定植能力可能还需要其他因素。为了鉴定EHEC的其他黏附因子，我们构建了一株血清型为O111:H-的临床EHEC分离株的转座子突变体，该菌株对培养的中国仓鼠卵巢（CHO）细胞表现出高水平的黏附。一个突变体在CHO细胞黏附、人红细胞凝集和自凝方面明显缺陷。对该突变体中被破坏的基因进行序列分析，发现一个9669 bp的新型染色体开放阅读框（ORF），命名为efa1，即EHEC黏附因子。efa1与最近描述的来自EHEC O157:H7溶血素编码质粒的一个ORF的预测产物具有28%的氨基酸同一性。这两个基因推定产物的氨基末端与艰难梭菌毒素A和B表现出高达38%的氨基酸相似性。efa1位于一个至少15 kb长的新型基因座内，其特点是G+C含量低、有几个插入序列同源物以及痢疾志贺氏菌肠毒素ShET2的一个同源物。从efa1不同区域制备的DNA探针与116株各种血清型的紧密黏附-抹平性大肠杆菌（AEEC）杂交，包括肠致病性大肠杆菌（EPEC）和EHEC，但不与91株非AEEC菌株杂交。然而，efa1对于紧密黏附-抹平表型不是必需的，并且efa1基因座与肠上皮细胞抹平（LEE）致病岛没有物理连接，LEE致病岛在EPEC中负责这种表型。这些发现表明，efa1编码一种新型的与AEEC毒力相关的决定因素，它有助于这些细菌的黏附能力。
