多重 RT-PCR 与液相悬浮芯片检测临床腹泻相关病毒的比较  

Comparing the multiplex RT-PCR method and liquichip technology in the detection of diarrhea-related virus

作　　者：罗欣[1] 余楠[1] 郭勇晖[1] 邓间开[1] 丁细霞[1] 王瑞莲[1] 富宁[1] 车小燕[1] 

机构地区：[1]南方医科大学珠江医院检验医学部,广州510282

出　　处：《中华检验医学杂志》2015年第6期387-391,共5页Chinese Journal of Laboratory Medicine

摘　　要：目的：比较实验室建立的多重RT-PCR与液相悬浮芯片对于临床腹泻相关病毒（腺病毒40／41型、诺如病毒GⅠ型和GⅡ型，星状病毒和札如病毒）的检测效能。方法多重RT-PCR方法学建立。收集2013年9月至2014年2月因腹泻至南方医科大学珠江医院就医的患者的粪便标本共107份。利用液相悬浮芯片技术和多重RT-PCR分别检测标本的腺病毒40／41型、诺如病毒GⅠ型和GⅡ型，对液相悬浮芯片未涵盖的星状病毒和札如病毒，以RT-PCR为参考，评估多重RT-PCR方法的特异度、敏感度。采用配对计数资料的Kappa系数检验，比较方法间的吻合度。多重RT-PCR和RT-PCR平行扩增10倍连续稀释的5种病毒质粒，测定多重RT-PCR的检测限。结果建立的多重RT-PCR方法与液相悬浮芯片、RT-PCR的检测一致性高，Kappa值分别为0.885和1.000，P均＝0.000。以液相悬浮芯片为标准，多重RT-PCR检测敏感度为80.8％（21／26），特异度100.0％（295／295）。多重RT-PCR对5种病毒的检测限为104～106拷贝／μl。结论多重RT-PCR与液相悬浮芯片对于临床标本检测显示一致性佳，特异度和敏感度高，快速、高通量，适用于临床微生物实验室常见腹泻相关病毒的多重检测。（中华检验医学杂志，2015，38：387-391）Objective To compare the detection efficiency between multiplex RT-PCR method and liquichip technology for screening the viral etiological agents of diarrhea.Methods The development of the multiplex RT-PCR method.A total of 107 feces samples from patients who suffered from diarrhea and attended to Zhujiang Hospital of Southern University from September 2013 to February 2014 were collected and tested in parallel by both multiplex RT-PCR and xTAG Gastrointestinal Pathogen Panel （ xTAG GPP） for Adenovirus, Norovirus genogroupⅠandⅡ, as well as by both multiplex RT-PCR and monoplex RT-PCR for Astrovirus and Sapovirus.To evaluate the sensitivity and specificity of multiplex RT-PCR, xTAG GPP and monoplex RT-PCR were used as reference.Kappa coefficient test was used to evaluate the consistency among the methods.The detection limit and accuracy of multiplex RT-PCR were evaluated by detection of serial dilution of positive plasmids and products sequencing for the five viral agents.Results The multiplex RT-PCR showed high consistency with xTAG GPP and monoplex RT-PCR, in which Kappa value was 0.885 and 1.000 respectively（ P=0.000 ）.Compared to xTAG GPP, the sensitivity and specificity of the multiplex RT-PCR were at average of 80.8%（ 21/26 ） and 100%（ 295/295 ） respectively.The detection limit and accuracy of multiplex RT-PCR were 104 copies /μl-106 copies/μl.Conclusion The high consistency indicated that both the multiplex RT-PCR and xTAG GPP are useful as a special,sensitive, high throughput and rapid diagnostic tools for the detection of the major viral pathogens related to diarrhea in clinical laboratory.

关 键 词：腹泻 腺病毒科 诺罗病毒 星状病毒属 多重聚合酶链式反应 逆转录聚合酶链反应 

分 类 号：R440[医药卫生—诊断学]