多重PCR技术在正畸治疗患者口腔细菌检测中的应用  

Multiplex PCR detection of four species of bacteria in oral specimens from orthodontic patients

作　　者：黄香娥[1] 韩谅 肖水清[2] 

机构地区：[1]济南市第四人民医院口腔科,济南250031 [2]济南市口腔医院口腔正畸科,济南250001

出　　处：《山东大学学报（医学版）》2010年第6期76-79,83,共5页Journal of Shandong University:Health Sciences

基　　金：济南市科委(2007-16006);济南市卫生局资助项目(2004-37)

摘　　要：目的建立多重PCR方法检测正畸治疗患者口腔中伴放线放线杆菌(Aa)、福塞氏类杆菌(Bf)、具核梭杆菌(Fn)、牙龈卟啉单胞菌(Pg)4种细菌的存在,并与牙龈指数进行相关性分析。方法选择55例正畸治疗至少2个月的青少年患者为矫治组,34例未带矫治器的牙周健康者为对照组,记录牙龈指数,分别采龈沟液标本进行细菌DNA提取及聚合酶链式反应;同时,以细菌的厌氧培养和生化反应鉴定为标准进行对比验证,并做了PCR的敏感性和特异性实验。结果建立的多重PCR最低可检测出细菌DNA1pg,约相当于20个Aa、Fn和Pg,80个Bf;能扩增出Aa、Bf、Fn、Pg4种参考菌株的目的条带,而对大肠杆菌的扩增没有目的条带;多重PCR法与常规细菌培养法对细菌的检测阳性率较为一致(P>0.05);Aa、Fn、Pg在两组间有明显差异(P<0.05),对Bf的检测未见明显差异(P>0.05);Aa、Fn、Pg的阳性率与牙龈指数之间呈正相关(P<0.01);而Bf与牙龈指数之间无相关性(P>0.05)。结论建立的多重PCR有较高的敏感性和特异性,可同时检测龈沟液中4种常见牙周致病菌,观察固定矫治器使用过程中牙周细菌的变化。Objective To establish a multiplex PCR for the detection of four species of bacteria （ Aa,Bf,Fn and Pg） in oral specimens from orthodontic patients,and to analyze the correlation between the presence of bacteria and the gingival index. Methods Periodomal pocket specimens from fifty-five patients who had worn fixed orthodontic appliances for at least 2 months and thirty-four healthy individuals without orthodontic appliances were collected. DNAs of the bacteria were extracted,and then PCR was used to amplify the target genes. At the same time,anaerobic culture and biochemical events were used to verify PCR results. Sensitivity and specificity of PCR were also evaluated with the reference bacteria. In addition,the gingival index of each individual was recorded. Results The multiplex PCR was able to detect a minimum of 1pg bacteria DNA corresponding to 20 cells of Aa,Fn and Pg as well as 80 cells of Bf. With good specificity,target fragments of four species of bacteria were detected but the E.coli was not. The positive rate of PCR assay was coincident with the bacteria culture （P0.05）. There were statistically significant differences in detectionresults of Aa,Fn and Pg between patients and healthy individuals（P0.05）,while these was no difference in the detection result of Bf（P0.05）. The analysis of Spearman rank correlation indicated that the gingival index had a positive correlation with the presence of Aa,Fn and Pg （P0.01） but no correlation with Bf （P0.05）. Conclusion With high sensitivity and specificity,the multiplex PCR can be used to simultaneously detect Aa,Bf,Fn and Pg and to observe changes of bacteria induced by wearing fixed orthodontic appliances.

关 键 词：固定矫治器 多重PCR 细菌检测 

分 类 号：R78[医药卫生—口腔医学]