广东省575株肺炎链球菌的毒力因子psaA、ply基因检测分析

发布时间：2018-02-13 18:57

  本文关键词： 肺炎链球菌 psaA基因 ply基因 PCR　出处：《中国病原生物学杂志》2015年08期 　论文类型：期刊论文

【摘要】：目的采用PCR检测肺炎链球菌(S.pn)毒力基因psaA和ply,研究分析两个毒力基因的特异性,为S.pn临床诊断提供依据。方法对广东省12家医疗机构诊断为S.pn感染患者的654份标本,经过细菌培养和乳胶凝集试验筛选出575株,以psaA、ply基因核心区域序列设计合成扩增引物,以抽提的S.pn分离株DNA为模板,采用常规聚合酶链反应(PCR)扩增目标基因psaA、ply片段,长度分别为838bp和209bp,调查分析不同来源标本的分离株psaA和ply基因检出率;采用乳胶凝集试验与多重PCR分型,比较两种结果中S.pn菌株psaA和ply检出率差异,以及广东S.pn主要血清型与非主要血清型菌株的psaA和ply检出率情况。结果经乳胶凝集试验鉴定为S.pn的560株菌株中,psaA和ply基因检出率分别为78.21%和83.57%,ply基因检出率高于psaA基因,psaA基因阳性的S.pn菌株,其ply基因均阳性;来源于血液和脑脊液标本的S.pn分离株的psaA和ply基因检出率分别为93.18%和95.45%,脓液标本分离的S.pn菌株psaA和ply基因检出率为100%。在荚膜肿胀反应和多重PCR检测为S.pn可分型血清型菌株中,前者psaA和ply基因阳性率分别为93.90%和98.57%,后者为96.34%和98.31%;而在多重PCR检定为不可分型血清型菌株中,psaA和ply基因检出率分别为47.06%和57.84%,显著低于荚膜肿胀试验的68.59%和74.64%,但ply基因检出率也均比psaA高;可分型血清型中主要血清型菌株的psaA和ply基因检出率高于非主要血清型。结论广东地区S.pn临床分离株中ply基因检出率比psaA高,保守性高,但其他链球菌亦能扩增出ply基因从而出现假阳性,因此,对于S.pn菌株的鉴定,采用PCR同时扩增psaA和ply基因,并结合临床常规实验,鉴定更为可靠。此外,血液和脑脊液psaA和ply基因阳性率高可能与psaA和ply是S.pn入侵血和脑的关键因子有关。
[Abstract]:Objective to detect the virulence genes psaA and plyl of S. pneumoniae by PCR, and to analyze the specificity of the two virulence genes in order to provide the basis for the clinical diagnosis of S.pn. Methods 654 samples of patients with S.pn infection were collected from 12 medical institutions in Guangdong Province. 575 strains were screened by bacterial culture and latex agglutination test. Primers were designed and synthesized from the core region of psaAply gene, and the target gene psaAply fragment was amplified by routine polymerase chain reaction (PCR) with the extracted S.pn isolate DNA as template. The length was 838bp and 209bp, respectively. The detection rate of psaA and ply genes in different samples were investigated and analyzed, and the detection rates of S.pn strain psaA and ply were compared by latex agglutination test and multiple PCR typing. The detection rate of psaA and ply in the main serotype and non-serotype strains of S.pn in Guangdong Province were 78.21% and 83.57ply, respectively, which were higher than that of psaA in the 560strains identified as S.pn by latex agglutination test. S.pn, which is positive for psaA gene, All of them were positive for ply gene. The detection rates of psaA and ply genes of S.pn isolates from blood and cerebrospinal fluid samples were 93.18% and 95.45, respectively. The detection rates of psaA and ply genes of S.pn strains isolated from pus samples were 100. The positive rates of psaA and ply genes were 93.90% and 98.57 in the former, 96.34% and 98.31 in the latter, and 47.06% and 57.84, respectively, in the multiplex PCR untyped serotype strains, which were significantly lower than 68.59% and 74.64 in the capsule swelling test, but ply. The detection rate of gene was higher than that of psaA. The detection rate of psaA and ply genes in the main serotype strains was higher than that in non-major serotypes. Conclusion the detection rate of ply gene in clinical isolates of S.pn in Guangdong is higher than that in psaA, and the detection rate of ply gene is higher than that of psaA. However, other streptococcus could amplify the ply gene, so it is more reliable to use PCR to amplify both psaA and ply genes of S.pn strain, and to combine with clinical routine experiments to identify the S.pn strain. In addition, the identification of S.pn is more reliable. The high positive rate of psaA and ply gene in blood and cerebrospinal fluid may be related to psaA and ply which are the key factors of S.pn invading blood and brain.
【作者单位】： 华南农业大学制药工程系;广东省疾病预防控制中心;广东省应急病原学重点实验室;广东省生物制品和药物研究所;
【基金】：中美新发和再发传染病合作项目 广州市科技计划重点项目(No.11C32100704)
【分类号】：R378.12
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