细菌培养及实时定量PCR方法研究儿童脑膜炎病原菌分布

Etiology of meningitis in children investigated by culture and real-time PCR

目的了解2005—2006年度我院中枢神经系统感染患儿脑脊液病原菌谱、血清型及耐药特点。方法脑脊液细菌鉴定采用细菌培养及实时定量PCR方法。肺炎链球菌血清分型采用荚膜肿胀试验,流感嗜血杆菌及脑膜炎奈瑟菌血清分型采用乳胶凝集法。抗菌药物敏感性试验采用K-B法及E试验。结果入组的新生儿患者28例,脑脊液细菌检测总阳性率39.3%,大肠埃希菌6例,占21.4%,居首位,其中4例ESBL阳性。其次是肺炎链球菌,占7.1%(2例)。未检测到脑膜炎奈瑟菌。1个月至11岁年龄组90例,细菌检测总阳性率33.3%,第1位是肺炎链球菌,占16.7%(15例),其中5例培养阳性,菌株血清型为19F型,2例对青霉素中介。其次是脑膜炎奈瑟菌,占5.6%(5例),3例培养阳性,2例为A群,1例为C群。结论2005—2006年,我院收治的不同年龄组脑膜炎患儿病原菌分布有所不同,新生儿患儿病原菌主要是大肠埃希菌,其次是肺炎链球菌。其他年龄组患儿则以肺炎链球菌、脑膜炎奈瑟菌为主要病原菌。部分菌株已高度耐药。采用实时定量PCR方法可提高肺炎链球菌及脑膜炎奈瑟菌检测的阳性率。

Objective To investigate the etiology, serotypes, and antimicrobial resistance of the pathogens of meningitis in children at Children＇s Hospital of Fudan University from 2005 - 2006. Methods The cerebrospinal fluid （CSF） was tested by real- time PCR and culture to identify potential pathogens. The antibiotic susceptibility testing was carried out by KB and E-test methods. The serotypes of Streptococcus pneurnoniae （SPN） were analyzed by capsule swelling test. The latex agglutination was adopted to determine the serotypes of Haemophilus influenzae （HIN） and Neisseria rneningitidis （NME）. Results During the 24-month surveillance period, a total of 118 cases meeting the criteria of meningitis were enrolled. The age of the patients ranged from newborn to 11 years old. Of the 118 children, 28 were neonates, 90 were 1 month to 11 years old. CSF was positive for microorganisms in 39.3% of the neonatal patients, mainly Escherichia coli （6/28, 21.4%）, 4 of the 6 isolates were ESBL positive. SPN was identified from 2 （7.1 %） patients. Pathogen was positive in 33.3% of the children at age of 1 month to 11 years. SPN infection was identified in 15/90 （16.7%） of the patients. Among the SPN positive patients, 5 were confirmed by culture. The serotype was 19F. Two of the five SPN isolates were intermediate to penicillin （PISP）. NME infection was found in 5.6% （5/90） of the patients. Three NME were isolated. The serotype of NME was A （2 isolates） and C （1 isolate）. Conclusions The pathogen of bacterial meningitis is different between neonates and young children in our hospital. Escherichia coli is the main pathogen of neonatal meningitis. SPN and NME are the primary pathogens in young children with bacterial meningitis. Some of the isolates were highly resistant to antibiotics. Real-time PCR is a sensitive and specific assay to identify SPN and NME from CSF.