Beyond Serotypes and Virulence-Associated Factors: Detection of Genetic Diversity among O153:H45 CFA/I Heat-Stable Enterotoxigenic Escherichia coli Strains

Characterization of enterotoxigenic Escherichia coli (ETEC) has been based almost exclusively on the detection of phenotypic traits such as serotypes and virulence-associated factors: heat-labile (LT) and heat-stable (ST) toxins and colonization factors (CFs). In the present work we show that the analysis of band patterns generated by randomly amplified polymorphic DNA (RAPD) analysis and pulsed-field gel electrophoresis (PFGE) of digested chromosomal DNA can be used to detect genetic diversity among ETEC strains expressing identical phenotypic traits. The study included 29 ETEC isolates from Latin America and Spain expressing the phenotype O153:H45 CFA/I ST plus 1 rough derivative, 2 nonmotile derivatives, and 1 O78:H12 CFA/I ST isolate, and a representative of a genetically distinct ETEC group. The results showed that the O153:H45 CFA/I ST ETEC isolates belong to a single clonal cluster whose isolates share on average, 84% of the RAPD bands and 77% of the PFGE restriction fragments, while the O78:H12 isolate shared only 44 and 4% of the RAPD bands and PFGE fragments, respectively, with the isolates of the O153:H45 group. More relevantly, RAPD and PFGE fingerprints disclosed the presence of different clonal lineages among the isolates of the O153:H45 cluster. Some of the genetic variants were isolated from defined geographic areas, while places like São Paulo City in Brazil and the middle-eastern part of Argentina were populated by several genetic variants of related, but not identical, ETEC strains. These results show that molecular biology-based typing methods can disclose strain diversity, which is usually missed in studies restricted to phenotypic typing of ETEC.

产肠毒素大肠杆菌（enterotoxigenic Escherichia coli, ETEC）的特性表征此前几乎完全基于表型性状检测，涵盖血清型、毒力相关因子：不耐热毒素（heat-labile, LT）、耐热毒素（heat-stable, ST）以及定植因子（colonization factors, CFs）。本研究证实，对酶切染色体DNA开展随机扩增多态性DNA（randomly amplified polymorphic DNA, RAPD）分析与脉冲场凝胶电泳（pulsed-field gel electrophoresis, PFGE）得到的条带图谱进行解析，可用于检测表型性状一致的ETEC菌株间的遗传多样性。本研究共纳入29株来自拉丁美洲与西班牙的ETEC分离株，其表型均为O153:H45 CFA/I ST，另包含1株粗糙型衍生株、2株无动力衍生株、1株O78:H12 CFA/I ST分离株，以及1株遗传背景迥异的ETEC类群代表株。结果显示，O153:H45 CFA/I ST型ETEC分离株隶属于单一克隆簇，该簇内菌株平均共享84%的RAPD条带与77%的PFGE限制性片段；而O78:H12分离株与O153:H45组菌株仅分别共享44%的RAPD条带与4%的PFGE片段。尤为重要的是，RAPD与PFGE指纹图谱揭示了O153:H45克隆簇内分离株存在不同的克隆谱系。部分遗传变异株分离自特定地理区域，而巴西圣保罗市与阿根廷中部地区则存在多株亲缘相关但并非完全一致的ETEC遗传变异株。本研究结果表明，基于分子生物学的分型方法可揭示菌株多样性，而这类多样性在仅局限于ETEC表型分型的研究中通常会被遗漏。