Koitabashi Tsutomu, Vuddhakul Varaporn, Radu Son, Morigaki Tadaaki, Asai Norio, Nakaguchi Yoshitsugu, Nishibuchi Mitsuaki
Graduate School of Medicine, Kyoto University, Japan.
Microbiol Immunol. 2006;50(2):135-48. doi: 10.1111/j.1348-0421.2006.tb03779.x.
Nine Escherichia coli O157: H7/- strains isolated primarily from non-clinical sources in Thailand and Japan carried the stx(2) gene but did not produce Stx2 toxin in a reversed passive latex agglutination (RPLA) assay. A strain (EDL933) bearing a stx(2) phage (933W) was compared to a strain (Thai-12) that was Stx2-negative but contained the stx(2) gene. To study the lack of Stx2 production, the Thai-12 stx(2) gene and its upstream nucleotide sequence were analyzed. The Thai-12 stx(2) coding region was intact and Stx2 was expressed from a cloned stx(2) gene using a plasmid vector and detected using RPLA. A lacZ fusion analysis found the Thai-12 stx(2) promoter non-functional. Because the stx(2) gene is downstream of the late promoter in the stx(2) phage genome, the antitermination activity of Q protein is essential for strong stx(2) transcription. Thai-12 had the q gene highly homologous to that of Phi21 phage but not to the 933W phage. High-level expression of exogenous q genes demonstrated Q antitermination activity was weak in Thai-12. Replication of stx(2) phage was not observed in Stx2-negative strains. The q-stx(2) gene sequence of Thai-12 was well conserved in all Stx2-negative strains. A PCR assay to detect the Thai-12 q-stx(2) sequence demonstrated that 30% of O157 strains from marketed Malaysian beef carried this sequence and they produced little or no Stx2. These results suggest that stx(2)-positive O157 strains that produce little or no Stx2 may be widely distributed in the Asian environment.
主要从泰国和日本的非临床来源分离出的9株大肠杆菌O157:H7/-菌株携带stx(2)基因，但在反向被动乳胶凝集试验（RPLA）中不产生Stx2毒素。将携带stx(2)噬菌体（933W）的菌株（EDL933）与Stx2阴性但含有stx(2)基因的菌株（泰国-12）进行比较。为了研究Stx2不产生的原因，对泰国-12的stx(2)基因及其上游核苷酸序列进行了分析。泰国-12的stx(2)编码区完整，使用质粒载体从克隆的stx(2)基因中表达出Stx2，并通过RPLA检测到。lacZ融合分析发现泰国-12的stx(2)启动子无功能。由于stx(2)基因位于stx(2)噬菌体基因组晚期启动子的下游，Q蛋白的抗终止活性对于stx(2)的强转录至关重要。泰国-12的q基因与Phi21噬菌体的q基因高度同源，但与933W噬菌体的q基因不同源。外源q基因的高水平表达表明泰国-12中的Q抗终止活性较弱。在Stx2阴性菌株中未观察到stx(2)噬菌体的复制。泰国-12的q-stx(2)基因序列在所有Stx2阴性菌株中都高度保守。检测泰国-12的q-stx(2)序列的PCR分析表明，市售马来西亚牛肉中的O157菌株中有30%携带该序列，且它们产生的Stx2很少或不产生。这些结果表明，产生很少或不产生Stx2的stx(2)阳性O157菌株可能在亚洲环境中广泛分布。
