福氏志贺菌历史菌株血清型转换的分子特征研究

Study on the mechanism of serotype conversion of historical isolates of Shigella flexneri

目的研究1949—1965年间出现血清型转换的福氏志贺菌的分子特征。方法255株福氏志贺菌均来源于中国医学细菌保藏管理中心（CMCC）。其中，进行血清型转换研究的20株福氏志贺菌收集自1949—1965年，分别来源于中国12株、苏联3株、捷克斯洛伐克1株、美国2株、匈牙利1株、法国1株。分别使用玻片凝集和多重PCR法进行血清分型，并进行gtrⅡ基因检测。对出现血清型转换的菌株进行gtrⅡ基因序列比对以及PFGE分型。结果255株福氏志贺菌中，79株携带gtrⅡ基因；其中19株玻片凝集为Y血清型菌株，多重PCR法检测分别为福氏2a血清型；1株玻片凝集为X血清型，多重PCR法检测为2b血清型；20株菌株发现了血清型转换，并出现了多处位点的核酸突变，除3株菌出现了3个氨基酸突变外，其余17株菌氨基酸序列均提前出现了终止密码子，导致gtrⅡ功能性失活。PFGE分析发现，携带gtrⅡ基因的Y血清型菌株与2a血清型菌株相似性为75.8%-100%，携带gtrⅡ基因的X血清型菌株与2b血清型菌株相似性为81.6%-100%。结论发生血清型转换的福氏Y或X血清型菌株，gtrⅡ基因出现的突变更加复杂；发生血清型转换的福氏Y或X血清型菌株可能来源于福氏2a或2b血清型，并将血清型转换出现的时间向前推进至1949年。

ObjectiveThe serotype screening of Shigella flexneri from 1934 to 1965 preserved by the National Center for Medical Culture Collections was carried out, and the molecular characteristics of the serotype conversion strains were studied.MethodsSerotyping of Shigella flexneri in this study was conducted by slide agglutination and multiplex PCR, respectively. The gtrⅡ gene sequence alignment and pulsed field gel electrophoresis typing were performed on the serotype conversion strains.ResultsAmong the 255 strains of Shigella flexneri preserved in CMCC （B） from 1934 to 1965, 79 were carrying gtrⅡ gene, of which 19 strains and 1 strain were agglutinated with the Y serotype and X serotype, respectively, and furthermore, the multiplex PCR assays results showed serotypes 2a and 2b, respectively, and the strains were considered to have serotype conversion. The 20 strains carrying the gtrⅡ gene showed multiple nucleotide mutations. Besides 3 strains of 3 amino acid mutations, the amino acid sequences of the other 17 strains showed a stop codon in advance, resulting in functional inactivation of gtrⅡ. PFGE analysis revealed that the similarity between the serotype Y strain carrying the gtrⅡ gene and the serotype 2a strain was 75.8%-100%, and the similarity between the serotype X strain carrying the gtrⅡ gene and the serotype 2b strain was 81.6%-100%.ConclusionMutations in the gtrⅡ gene are more complicated in serotype-transforming Shigella flexneri serotype Y or X strains. Molecular typing suggests that the serotype-transforming Shigella flexneri serotype Y or X strains may be derived from the Shigella flexneri serotype 2a or 2b, and advance the serotype conversion to 1949.