摘要
目的运用基于Taqman探针实时荧光定量PCR技术检测胆囊结石中的华支睾吸虫DNA。方法选取2012年3～4月份40例实施内镜取石保胆手术患者的胆囊结石,通过傅里叶红外光谱法分析结石成分,对结石研磨涂片镜检观察,并提取其DNA,运用实时荧光定量PCR技术检测华支睾吸虫基因。结果 40例患者的胆囊结石按主要成分可分为胆色素性结石(25例),胆固醇性结石(6例),混合性结石(9例)。结石涂片镜检有23例发现华支睾吸虫虫卵(阳性率57.5%),其中胆色素性结石19例,混合性结石4例。实时荧光定量PCR检测华支睾吸虫DNA阳性率为65.0%(26/40),其中包括23例镜检虫卵阳性标本及3例镜检阴性标本。结论运用实时荧光定量PCR在胆囊结石中检测出华支睾吸虫DNA,敏感性高于镜检法查虫卵。这不仅为华支睾吸虫感染诱发胆囊结石形成提供了分子生物学证据,同时也提供了一种诊断华支睾吸虫感染的有效手段,对胆囊结石病人的病因诊断、临床治疗以及术后预防复发等具有重要的指导意义。
Objective Detection of Clonorchis sinensis DNA in gallbladder stones of patients with cholecystolithiasis by a TaqMan based real-time PCR.Methods Forty patients with cholecystolithiasis were chosen randomly who accepted endoscopic gallbladder-preserving cholelithotomy from March to April 2012.The stones obtained from clinical operation were first analyzed through Fourier infrared spectrometry,and then grinded for microscopic examination.Specific primers and TaqMan probe targeting the cytochrome c oxidase gene of C.sinensis were designed and synthesized,and then the DNA of gallbladder stones was extracted and detected by a real-time PCR for C.sinensis gene.Results 40 gallbladder stones were divided into three types based on the Fourier infrared spectrometry analysis： bile pigment stone（25 cases）,cholesterol stone（6 cases） and mixed stone（9 cases）.Parasite eggs were found in 23 stones including 19 pigment and 4 mixed stones（positive rate 57.5%）.No eggs were found in the cholesterol stones.The positive rate of real-time PCR was 65.0%（26/40）,C.sinensis DNA was detected in all the 23 egg-positive stones and 3 other egg-negative stones.Conclusion In this study,C.sinensis DNA was detected in gallbladder stones through real-time PCR,which was superior to microscopic examination and considered as the most effective means for assessing C.sinensis infection-status.The research not only presenting the molecular biological evidence for the formation of gallbladder stone induced by C.sinensis infection,but also providing a new efficient approach for the detection of C.sinensis infection.It has a great guiding significance for those patients with cholecystolithiasis in the diagnosis,treatment,prevention and so on.
出处
《中国病原生物学杂志》
CSCD
北大核心
2012年第10期767-769,766,共4页
Journal of Pathogen Biology
