霍乱弧菌VPI毒力岛的tcpI-P和tcpH-A间隔区序列在不同菌株中的差异分析

目的：TcpA是霍乱弧菌主要定居因子毒素共调菌毛的主要亚单位，基因位于VPI毒力岛上。古典生物型和EI Tor生物型霍乱弧菌菌株的tcpA序列有较大差异，同时毒力岛上其他一些区域，主要包括部分基因的间隔区序列也存在差异。我们已在不同EI Tor流行株与非流行株以及非01／非O139菌株中又发现4种新的tcpA序列。笔拟探明这4种不同类型tcpA序列的菌株中，VPI毒力岛上其他几个差异片段(tcpI—P间隔区和tcpH—A间隔区)是否与相应tcpA序列有平行的变异关系、进而分析毒力岛和菌株分化的多样性。方法：在tcpI—P和tcpH—A间隔区外侧保守区中设计引物，扩增包含这两个间隔区的片段并进行测序，计算比较序列差异。结果：基因tcpH与tcpA之间的间隔区序列两两间比较的一致性在63．3％至95．3％之间；基因tcpI与tcpP之间的间隔区序列两两间比较的一致性在26．7％至100％之间。这些间隔区序列并没有像tcpA序列在这些菌株中的差异那样显示出类似的变异水平，甚至一些菌株的tcpA序列有较大差异时、某些区域反而高度一致。结论：在不同tcpA序列的菌株中tcpI—P间隔区和tcpH—A间隔区及tcpA这三个片段分化变异的速率不同，并不具有平行性。这些序列的多样性反映了霍乱弧菌VPI毒力岛以及菌株分化的多样性。这种分化也提示可能与这些区域具有结合一些调控因子、从而对环境信号产生不同方式或程度的应答的功能有关。

Diversity Analysis of Intergenic Sequence of TcpI- P and TcpH- A within VPI in Different Vibrio Cholerae Strains

Objective TcpA is the major su bunit of toxin-coregulated pili,the critical colonization factor of Vibrio chole rae tcpA gene,which locates in Vibrio Pathogenicity Island(VPI),is obviously d i fferent in nucleotide sequence between the strains of classical and El Tor bioty pes It has been showed that some other loci in VPI,including some intergentic s equence(spacer),are also diverse in their sequences We have found 4 novel tcpA sequences from different El Tor strains and non-O1/non-O139 strains In this st u dy we try to determine if the diverse of the spacer sequences of tcpI-P and tcpH -A were parallel with that of the tcpA sequences, and to evaluate the differenti ation of VPI and strains Methods The spac ers of tcpI-P and tcpH-A were amplified,the PCR products were purified and direc tly sequenced The sequences comparison was made by phylogenesis analysis Results The identity of tcpH-A and tcpI-P spacers in different strains are 63 3%-95 3% and 26 7%-100%,respectively,a nd doesn't show similar variation of tcpA Furthermore,in some strains though the tcpA is far more diverse,the spacers are identical instead Conclusion The rates of the variation of t cpA,tcpI-P and tcpH-A spacers were not in parallel each other,and the polymorphi sm of these sequences showed the high diversity of VPI These differentiations m ay suggest that these regions are possibly involved in binding with some signal regulators,and responding to environmental signals in different ways and at diff erent levels