培养与聚合酶链反应联合进行住院患儿肺炎细菌病原学检测

Bacterial etiology of pneumonia in hospitalized children： combined detection with culture and polymerase chain reaction

目的探讨细菌培养与病原特异性DNA联合检测住院患儿肺炎细菌病原的应用价值。方法对187例肺炎患儿的深部呼吸道吸引物进行肺炎链球菌、流感嗜血杆菌选择性培养和普通培养，并且对同一标本采用靶序列富集多重PCR（Tem-PCR）扩增结合Luminex液态芯片检测平台进行定量测定，检测肺炎链球菌、流感嗜血杆菌、金黄色葡萄球菌、肺炎克雷伯杆菌、大肠杆菌、嗜肺军团菌、绿脓杆菌、鲍曼不动杆菌等14种病原菌的特异性DNA。结果细菌培养的总检出率为40．1％（75／187，含3例检出2种病原菌），病原菌依次为流感嗜血杆菌17．1％、大肠杆菌8．6％、肺炎克雷伯杆菌6．4％、金黄色葡萄球菌4．8％、肺炎链球菌3．7％、绿脓杆菌1．6％、鲍曼不动杆菌1．1％和阴沟肠杆菌1．1％。以细菌培养或Tem-PCR任一阳性为标准，联合检测的总检出率为78．6％（147／187），病原菌依次为流感嗜血杆菌28．9％、肺炎链球菌19．3％、大肠杆菌8．6％、肺炎克雷伯杆菌6．4％、金黄色葡萄球菌5．9％、鲍曼不动杆菌5．9％、绿脓杆菌2．7％和阴沟肠杆菌1．1％。结论Tem-PCR能提高流感嗜血杆菌、肺炎链球菌、金黄色葡萄球菌、绿脓杆菌和鲍曼不动杆菌的检出例数。细菌培养与病原特异性DNA联合检测应用能显著提高肺炎病原的检出率，可能更真实地反映肺炎的细菌病原学情况。

Objective Bacterial cuhures from respiratory aspirate or sputum have been the conventional diagnostic method for pneumonia, but the results of culture was often affected by early extensive use of antibiotics, sample collection and delivery. The objective of this study was to explore application of the combined detection of culture and polymerase chain reaction （PCR） assay in hospitalized children with pneumonia. Methods Totally 187 hospitalized children with pneumonia were enrolled. The age of the patients ranged from 1 month to 10 years, 124 were male, 63 female; 175 of the patients received antibiotics treatment before admission. Deep respiratory aspirate sample from patients was cultured by Streptococcus pneumoniae selective plate, Hemophilus influenzae selective plate and conventional plate. The aspirate samples were also amplified for DNA of 14 bacteria with target enriched multiplex polymerase chain reaction （Tem-PCR） and detected with Luminex xMAP technology platform. Results The total positive rate by bacterial culture was 40. 1% （75/187）, of which 17. 1% （24/187） were Hemophilus influenzae b, 8. 6% （16/187） were Escherichia coli, 6.4% （ 12/187 ） were Klebsiella pneumoniae, 4. 8% （ 9/187 ） were Staphylococcus aureus, 3.7% （7/187） were Streptococcus paeumoniae, 1.6% （ 3/187 ） were Pseudomonas aeruginosa, 1.1% （2/187） were Acinetobacter baumannii, and 1.1% （2/187） were Enterobacter cloacae. The total positive rate by combined detection of culture and Tem-PCR assay were 78. 6% （ 147/187 ） , of which 28.9% （54/187） were Hemophilus inflaenzae b, 19.3% （36/187） were Streptococcus pneumoniae, 8.6% （ 16/187 ） were Escherichia coli, 6.4% （ 12/187 ） were Klebsiella pneumoniae, 5.9% （ 11/187 ） were Staphylococcus aureus, 5.9% （11/187） were Acinetobacter baumannii, 2.7% （5/187） were Pseudomonas aeruginosa, and 1.1% （2/187） were Enterobacter cloacae. Conclusion The Tem-PCR assay may increase the detection rate of Hemophilus inflaenzae b, Streptococcus pneumoniae , Staphylococcus aureus , Pseudomonas aeruginosa and Acinetobacter baumannii. The Combined detection may increase the positive rate of bacterial pathogens in hospitalized children with pneumonia, and the results might reflect the real patterns of bacterial etiology. The Tem-PCR needs further improvement for diagnosis of Escherichia coli and Klebsiella pneumoniae.