Complete Nucleotide Sequence and Analysis of the Locus of Enterocyte Effacement from Rabbit Diarrheagenic Escherichia coli RDEC-1

The pathogenicity island termed the locus of enterocyte effacement (LEE) is found in diverse attaching and effacing pathogens associated with diarrhea in humans and other animal species. To explore the relation of variation in LEE sequences to host specificity and genetic lineage, we determined the nucleotide sequence of the LEE region from a rabbit diarrheagenic Escherichia coli strain RDEC-1 (O15:H−) and compared it with those from human enteropathogenic E. coli (EPEC, O127:H6) and enterohemorrhagic E. coli (EHEC, O157:H7) strains. Differing from EPEC and EHEC LEEs, the RDEC-1 LEE is not inserted at selC and is flanked by an IS2 element and the lifA toxin gene. The RDEC-1 LEE contains a core region of 40 open reading frames, all of which are shared with the LEE of EPEC and EHEC. orf3 and the ERIC (enteric repetitive intergenic consensus) sequence present in the LEEs of EHEC and EPEC are absent from the RDEC-1 LEE. The predicted promoters of LEE1, LEE2, LEE3, tir, and LEE4 operons are highly conserved among the LEEs, although the upstream regions varied considerably for tir and the crucial LEE1 promoter, suggesting differences in regulation. Among the shared genes, high homology (>95% identity) between the RDEC-1 and the EPEC and EHEC LEEs at the predicted amino acid level was observed for the components of the type III secretion apparatus, the Ces chaperones, and the Ler regulator. In contrast, more divergence (66 to 88% identity) was observed in genes encoding proteins involved in host interaction, such as intimin (Eae) and the secreted proteins (Tir and Esps). A comparison of the highly variable genes from RDEC-1 with those from a number of attaching and effacing pathogens infecting different species and of different evolutionary lineages was performed. Although RDEC-1 diverges from some human-infecting EPEC and EHEC, most of the variation observed appeared to be due to evolutionary lineage rather than host specificity. Therefore, much of the observed hypervariability in genes involved in pathogenesis may not represent specific adaptation to different host species.

被命名为肠细胞脱落位点（locus of enterocyte effacement，LEE）的致病岛，广泛存在于多种与人类及其他动物腹泻相关的黏附脱落型致病菌（attaching and effacing pathogens）中。为探究LEE序列变异与宿主特异性及遗传谱系的关联，我们测定了兔致腹泻大肠杆菌RDEC-1（O15:H−）菌株的LEE区域核苷酸序列，并将其与人类肠致病性大肠杆菌（enteropathogenic E. coli，EPEC，O127:H6）及肠出血性大肠杆菌（enterohemorrhagic E. coli，EHEC，O157:H7）菌株的LEE序列进行比对分析。与EPEC和EHEC的LEE不同，RDEC-1的LEE并非插入于selC位点，其上下游侧翼分别为IS2元件与lifA毒素基因。RDEC-1的LEE包含一段由40个开放阅读框（open reading frames）构成的核心区域，所有开放阅读框均与EPEC和EHEC的LEE保守共享；EHEC与EPEC的LEE中所携带的orf3及肠道重复基因间序列共识（enteric repetitive intergenic consensus，ERIC）序列，在RDEC-1的LEE中并未出现。尽管tir与关键的LEE1启动子的上游区域存在显著序列差异，但LEE1、LEE2、LEE3、tir及LEE4操纵子（operons）的预测启动子在各LEE中均呈现高度保守性，这提示其转录调控机制存在差异。在保守共享的基因中，RDEC-1的LEE与EPEC、EHEC的LEE在预测氨基酸水平上的同源性高达95%以上，涉及Ⅲ型分泌系统（type III secretion apparatus）组分、Ces分子伴侣（Ces chaperones）及Ler调控因子（Ler regulator）。与之相反，编码宿主互作相关蛋白的基因则呈现出更高的序列分化度（同源性为66%至88%），例如黏附素（intimin，Eae）以及分泌蛋白Tir与Esps。我们将RDEC-1的高变异基因，与多种感染不同宿主、隶属于不同进化谱系的黏附脱落型致病菌的对应基因进行了比对分析。尽管RDEC-1与部分人类感染性EPEC及EHEC存在序列分化，但所观察到的绝大多数变异似乎源于进化谱系差异，而非宿主特异性。因此，在致病相关基因中所观察到的高度变异，或许并不代表对不同宿主物种的特异性适应性演化。