应用分子生物学方法鉴定6群肺炎链球菌血清型

Identification of streptococcus pneumonia serogroup 6by molecular biological technology

目的利用分子生物学方法,鉴定26株6群肺炎链球菌的血清型。方法利用6群肺炎链球菌荚膜多糖相关基因设计合成6对特异性引物,PCR扩增26株肺炎链球菌,并对PCR产物进行基因序列的测定及分析。结果26株肺炎链球菌cpsD蛋白229个氨基酸中有7个突变点,第220~222位均没有缺失;其中,19株肺炎链球菌具有与wci Nα蛋白氨基酸序列完全一致的氨基酸组成,第150位均为Ala,第38位均为Asp,其余7株与wciN_α蛋白氨基酸序列没有相似性,但与wciN_β蛋白氨基酸序列相似性超过99%;15株wciP蛋白第195位为Ser,11株wci P蛋白第195位为Asn。综合分析比对后,26株6群肺炎链球菌中,11株属于6A型肺炎链球菌,8株属于6B型肺炎链球菌,4株属于6C型肺炎链球菌,3株属于6D型肺炎链球菌。结论分子生物学方法可用于6群肺炎链球菌血清型的鉴定,为完善6群肺炎链球菌的菌种档案提供了实验依据。

Objective To Identify 26 strains of streptococcus pneumonia serogroup 6 by molecular biological methods.Methods Six pairs of primers based on the cps loci sequences were designed to be used in PCR amplification of 26 strains of streptococcus pneumonia serogroup 6,and the PCR products were sequenced and analysed. The predicted translation products of cps D、wci N and wci P genes were aligned with BLAST tool. Results Among 229 amino acid residues of the cps D protein in 26 strains of streptococcus pneumonia serogroup 6,there were 7 mutation points,and no deletion was found in the 220-222 sequence. The produced sequences showed a compete identity for amino acids component in 19 strains from streptococcus pneumonia serogroup 6 with wciN_α protein,and Ala-150 and Asp-38 were found in these strains. The other 7 strains had no identity in protein sequences with wciN_α protein,but more than 99% identity was found with wciN_β protein sequence. It showed Ser-195 of wci P protein in 15 strains and Asn-195 in the other 11 strains,among the 26 strains of streptococcus pneumonia serogroup 6. By a comparitive investigation,11 strains belong to serotype 6A,8 strains to serotype 6B,4strains to serotype 6C,3 strains to serotype 6D among total strains. Conclusion Molucular biological technology is feasible in identification of the serotypes in streptococcus pneumonia serogroup 6,on which an experimental basis was set up in completion of the bacterial profile for this serogroup.