CAJ | 학술논문

目的建立并应用检测儿童下呼吸道感染WU多瘤病毒(WU polyomavirus)的实时荧光定量PCR( real-time fluorescent quantitative PCR,FQ-PCR)方法.方法选择WU多瘤病毒的VP2基因作为检测的目标基因,设计FQ-PCR引物和检测探针,以重组质粒为标准品建立标准曲线,并对该方法的特异性、灵敏度、重复性进行评价；应用该方法对温州医学院附属温岭医院收集的临床下呼吸道感染患儿的痰液、咽拭子846份及血清标本846份进行定量检测.结果本研究建立FQ-PCR检测方法,质粒标准曲线的方差系数为0.998,灵敏度可达到50拷贝；应用该方法检测700份痰液标本,检测到7例阳性标本,146份咽试子标本中未检测到阳性标本,总阳性率为1.00％ (7/700),846份血清标本未检测到阳性标本.结论本研究建立的FQ-PCR方法可以特异、快速、灵敏地对儿童下呼吸道感染的WU多瘤病毒进行定量检测；痰液标本较咽拭子或血清标本更适用于WU多瘤病毒感染的核酸检测.
목적 건립병응용검측인동하호흡도감염WU다류병독(WU polyomavirus)적실시형광정량PCR( real-time fluorescent quantitative PCR,FQ-PCR)방법.방법 선택WU다류병독적VP2기인작위검측적목표기인,설계FQ-PCR인물화검측탐침,이중조질립위표준품건립표준곡선,병대해방법적특이성、령민도、중복성진행평개；응용해방법대온주의학원부속온령의원수집적림상하호흡도감염환인적담액、인식자846빈급혈청표본846빈진행정량검측.결과 본연구건립FQ-PCR검측방법,질립표준곡선적방차계수위0.998,령민도가체도50고패；응용해방법검측700빈담액표본,검측도7례양성표본,146빈인시자표본중미검측도양성표본,총양성솔위1.00％ (7/700),846빈혈청표본미검측도양성표본.결론 본연구건립적FQ-PCR방법가이특이、쾌속、령민지대인동하호흡도감염적WU다류병독진행정량검측；담액표본교인식자혹혈청표본경괄용우WU다류병독감염적핵산검측.
Objective Development and application of a real time fluorescent quantitative PCR (FQ-PCR) assay for detecting WU polyomavirus in children with low respiratory tract infections.Methods The VP2 gene of WU polyomavirus was selected as the detection target,from which the real time primers and probes were designed.The standard curve was established by using recombinant plasmid as template.And the FQ-PCR assay for specific detection of WU polyomavirus was established.The speciflcity,sensitivity and reproducibility of the method were evaluated. Furthermore,the clinical specimens from children with respiratory tract infections collected in Wenling First People's Hospital were quantitatively detected using this method.Results In this study,the FQ-PCR method was established to detect a specific fragment in VP2gene of WU polyomavirus.The standard curve coefficient R2 was 0.998.And this method can detect as low as 50 copies recombinant plasmid.The clinical specimens of sputum and throat swab from children with respiratory tract infections were quantitatively detected using this method.7 sputum specimens were detected as WU polyomavirus positive in 700 sputum specimens,the positive ratio was 1.00％.No positive specimens were detected in 146 specimens of throat swabs and 846 blood samples from same patient population.Conclusion .The results indicated that the FQ-PCR assay method established in this study was specific,rapid and sensitive for detecting WU polyomavirus in children with lower respiratory tract infections.The sputum specimen is more suitable to be used for gene detection of WU polyomavirus than throat swab or blood.