PCR-变性梯度凝胶电泳技术分析肺炎儿童下呼吸道菌群多样性

目的：初步探讨PCR-变性梯度凝胶电泳(PCR-DGGE)检测肺炎儿童下呼吸道菌群多样性的作用。方法收集10份培养阳性的患儿肺泡灌洗液2~5 mL，经细菌DNA提取、16S rDNA-PCR后进行变性梯度凝胶电泳(DGGE)分析，对优势条带进行测序。结果经培养10份标本中7份鉴定为一种菌感染，3份培养出两种病原菌；经PCR-DGGE分析8份标本出现多条明显条带，2份得到一条条带，共筛选出12个优势条带；DGGE法得到的细菌种类多于培养法。结论 PCR-DGGE可较好地鉴定混合细菌感染性肺炎病原菌并能检测到培养法没有检出的细菌；儿童肺炎主要以肺炎链球菌、大肠埃希菌、草绿色链球菌等病原菌为主。

PCR-denaturing gradient gel electrophoresis analysis of lower respiratory tract microbial diversity in children with pneumonia

Objective To explore the value of PCR-denaturing gradient gel electrophoresis (PCR-DGGE) in analysis of lower respiratory tract microbial diversity in children with pneumonia. Methods A total of 10 bron-choalveolar lavage fluid samples which has been identified bacterial infection by culture were included. The bacteri-al DNA was extracted, and 16S rDNA-V3 was amplified, following by DGGE, sequencing the predominant bonds and BLAST. Results In the 10 samples, there was one bacterium in 7 samples, two bacteria in 3 samples by bacterial cul-ture. There were multiple bonds in 8 samples by DGGE, and a total of 12 predominant bonds were identified. DGGE method can detect more bacterial species than bacterial culture method. Conclusion PCR-DGGE can identify mixed bacteria which caused pneumonia and detect the pathogen that cannot be detected by bacterial culture. The pneumonia in children is mainly caused by Streptococcus pneumoniae, Escherichia coli, Streptococcus viridans.