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目的 研究同时携带两种CTXΦ基因组的O139群霍乱弧菌中CTXΦ基因组的多样性。方法 Southernblot检测分离自福建的部分O139霍乱弧菌菌株中ctxAB和zot基因的多态性 ,然后选取ctxAB和zot基因拷贝数差异较大的 3株菌 ,扩增其调控抑制基因rstR基因片段 ,克隆并进行序列测定、分析。结果 此 3株菌染色体zot杂交条带多于ctxAB 2～ 5条 ,提示 1个菌株染色体上有编码ctxAB的CTXΦ以及不编码ctxAB的CTXΦ两种基因组。以保守区设计的引物扩增此 3株菌的rstR区 ,均得到 2条大小不同的条带 ,分别克隆后进行测序 ,经GenBank同源性检索 ,发现在此 3个菌株染色体中分别有来自ElTor型的CTXETΦ与不编码ctxAB的nct CTXO13 9Φ、来自加尔各答型的CTXcalcΦ与nct CTXO13 9Φ、以及CTXETΦ与新报道的一种具有独特rstR序列的CTXΦ类基因组两两共存于 1个菌株染色体上。这 3个菌株还具有不同的 16S核糖体基因杂交图谱。结论 不同rstR序列的CTXΦ基因组可以共存于 1个菌株染色体中 ,提示CTXΦ家族进化过程中的复杂分化 ,CTXΦ家族在不同霍乱菌株间的出现及传递、在霍乱弧菌菌株的变异分化中具有相关性
Objective To study the diversity of CTXΦ genome in V. cholerae O139 harboring two CTXΦ genomes. Methods Southern blot was used to detect the polymorphisms of ctxAB and zot genes in some strains of Vibrio cholerae O139 isolated from Fujian Province. Three strains with different ctxAB and zot gene copy numbers were selected to amplify the rstR gene fragment, And sequence determination, analysis. Results The zot bands of these three strains were more than 2 to 5 of ctxAB, suggesting that one strain had CTXΦ encoding ctxAB on chromosome and CTXΦ not encoding ctxAB. Two primers were designed to amplify the rstR region of these three strains. Two bands of different sizes were obtained and cloned separately for sequencing. GenBank was used to search for the homology of the three strains. ElTor CTXETΦ and nct CTXO13 9Φ not encoding ctxAB, CTXcalcΦ and nct CTXO13 9Φ from Calcutta, and CTXETΦ coexist with one newly reported CTXΦ genome with unique rstR sequences on one strain chromosome. These three strains also have different 16S ribosomal gene hybridization patterns. Conclusion CTXΦ genomes with different rstR sequences can coexist in chromosomes of one strain, suggesting complex differentiation of CTXΦ family members, appearance and transmission of CTXΦ family among different strains of cholera, and their correlation with the variation and differentiation of V. cholera strains