丙型副伤寒沙门氏菌噬菌体SPC-P1在不同血清型中的分布及整合位点分析

Distribution in different Salmonella serovars and integration sites of Salmonella paratyphi C phage SPC-P1

目的：确定丙型副伤寒沙门氏菌（Salmonella paratyphi C,SPC）噬菌体SPC-P1在其他血清型中是否存在,并确定其在宿主菌基因组中的整合位点。方法：以丙型副伤寒沙门氏菌RKS4594基因组中SPC-P1为模板,设计6对引物,分别在11株伤寒沙门氏菌、11株甲型副伤寒沙门氏菌、12株乙型副伤寒沙门氏菌和23株丙型副伤寒沙门氏菌中进行SPC-P1片段的扩增。同时下载美国国立生物技术信息中心（National Center for Biotechnology Information,NCBI）基因组数据库中全基因组序列完成的100株20个血清型的沙门氏菌全基因组序列,通过Mauve 2.3.1软件进行比对,以确定SPC-P1在各个血清型中的分布。根据SPC-P1在RKS4594基因组中的整合位点,设计引物,对10株SPC-P1阳性的丙型副伤寒沙门氏菌进行扩增,并对产物测序,根据测序结果分析SPC-P1整合情况。结果：SPC-P1广泛存在于丙型副伤寒沙门氏菌基因组中,14株能够扩增到所有的6个片段,2株扩增到3~5个片段,所有扩增阳性的片段与预期大小吻合。在伤寒沙门氏菌、甲型副伤寒沙门氏菌和乙型副伤寒沙门氏菌中,6个片段均阴性或仅存在1~2个片段,并且片段均比预期小。Mauve比对结果显示,仅在与丙型副伤寒沙门氏菌进化关系较近的猪霍乱沙门氏菌中存在接近完整SPC-P1的片段,但序列相似性存在差异。SPC-P1在丙型副伤寒沙门氏菌基因组中的整合位点是固定的,位于两个相邻基因pgt E和yfd C之间。结论：SPC-P1是丙型副伤寒沙门氏菌独特的致病因子,其存在可能使得丙型副伤寒沙门氏菌的宿主范围、致病性大小等功能方面有别于其他沙门氏菌。

Objective： To determine the prevalence of Salmonella paratyphi C phage（ SPC-P1） in different Salmonella serovars and to identify the integration sites in host genome. Methods： Based on the complete genome of SPC-P1 in S. paratyphi C RKS4594,6 pairs of primers were designed and used to amplify the fragments of SPC-P1 in 11 S. typhi,11 S. paratyphi A,12 S. paratyphi B and 23 S. paratyphi C strains. At the same time,100 complete genomes of Salmonella including 20 serovars available in National Center for Biotechnology Information（ NCBI） database were downloaded and aligned by Mauve2. 3. 1 to determine the prevalence of SPC-P1 in these serovars. Primers were designed according to the integration sites of SPC-P1 in the genome of RKS4594,and used to amplify ten strains having SPC-P1 in the genome. The PCR products were sequenced to investigate the integration sites of SPC-P1. Results：SPC-P1 was widely distributed in S. paratyphi C genome. In the study,14 strains had all 6 fragments and2 strains had 3-5 fragments. All the amplified fragments showed expected sizes. In contrast,in the genomes of S. typhi,S. paratyphi A and S. paratyphi B,no or only 1-2 fragments could be amplified,and the sizes were smaller than expected. The results from Mauve showed that only in the genome of S.choleraesuis,which was a close relative of S. paratyphi C,there existed an almost complete genome of SPC-P1. The insertion site of SPC-P1 in all the ten S. paratyphi C strains tested was between pgt E and yfd C genes. Conclusion： SPC-P1 is a unique virulence factor of S. paratyphi C. It may play roles in the host range and pathogenicity of S. paratyphi C.