6群肺炎链球菌荚膜多糖合成相关基因的分子生物学研究

Biomolecular analysis of capsular polysaccharide synthesis genes from Streptococcus pneumoniae serogroup 6

目的利用分子生物学方法,对25株6群肺炎链球菌荚膜多糖合成相关基因(cps loci)进行研究。方法采用多位点序列分型(multilocus sequence typing,MLST)技术,对25株6群肺炎链球菌进行分型;依据Gen Bank中收录的6群肺炎链球菌cps loci设计合成引物,以25株肺炎链球菌基因组DNA作为模板进行PCR扩增,并对扩增产物进行基因测定及分析;采用相邻合并分析(neighbour-joining analysis),根据MLST的7个管家基因和wciP、wzy、wzx这三个cps loci的代表基因分别绘制出系统发育树。结果根据MLST技术分析发现7株新的ST型菌株。25株6群肺炎链球菌cps loci的G+C含量为35.4%~35.5%,均属于I类;所有6C型wzy基因均有6个碱基的缺失。根据MLST的7个管家基因绘制的系统发育树,并根据cps loci的3个代表基因wciP、wzy和wzx绘制的系统发育树,差异很大。3个代表基因绘制的系统发育树,6C型为进化距离比较远的分枝,6A型和6B型的分枝进化距离相对较近;6D型与6B型在同一分枝内。结论获得了25株6群肺炎链球菌ST型和全长cps loci,完善了6群肺炎链球菌的菌种档案。

Objective To investigate the capsular polysaccharide synthesis genes（cps loci） in 25 strains of streptococcus pneumoniae serogroup 6 by using biomolecular methods. Methods Multilocus Sequence Typing（MLST） technique was introduced to determine sequence type of 25 strains from streptococcus pneumoniae serogroup 6. Six pairs of primers were designed according to cps loci from Gen Bank,which were used in amplification of target gene products from 25 strains of streptococcus pneumoniae serogroup 6,and the PCR products were sequenced and analyzed. Using Neighbour-joining analysis,the phylogenetic trees were mapped based on the 7 house-keeping genes of MLST and cps sequence profiles,respectively. R esults A totalof 7 new STs were found by MLST. The G＋C contents of the cps loci were 35.4%-35.5% from obtained 25 cps loci of streptococcus pneumoniae serogroup 6,and they all belong to class I. All 6C strains had a unique wzy allele with a 6 bp deletion. The two phylogenetic trees were different,and the phylogenetic tree of cps sequence profiles showed a distinct clade with 6C sequence and moderately distinct clades with 6A and 6B sequences. The serotype 6D cps profile was found within the6 B clade. Conclusion The STs and the capsular polysaccharide synthesis genes of 25 strains of streptococcus pneumoniae serogroup 6 were obtained and analyzed,on which the files of streptococcus pneumoniae serogroup 6 was enriched.