Guan Li, Zhao Lin-Qing, Zhou Hang-Yu, Nie Kai, Li Xin-Na, Zhang Dan, Song Juan, Qian Yuan, Ma Xue-Jun
Key Laboratory for Medical Virology, National Health and Family Planning Commission, Chinese Center for Disease Control and Prevention, National Institute for Viral Disease Control and Prevention, No.155 Changbai Road, Changping, Beijing, 102206, China.
Laboratory of Virology, Beijing Key Laboratory of Etiology of Viral Diseases in Children, Capital Institute of Pediatrics, 2 Yabao Road, Beijing, 100020, China.
Arch Virol. 2016 Jul;161(7):1891-8. doi: 10.1007/s00705-016-2858-z. Epub 2016 Apr 30.
Human rhinoviruses (HRVs) have long been recognized as the cause of more than one-half of acute viral upper respiratory illnesses, and they are associated with more-serious diseases in children, such as asthma, acute otitis media and pneumonia. A rapid and universal test for of HRV infection is in high demand. In this study, a reverse transcription genome exponential amplification reaction (RT-GEAR) assay targeting the HRV 5' untranslated region (UTR) was developed for pan-HRV detection. The reaction was performed in a single tube in one step at 65 °C for 60 min using a real-time fluorometer (Genie(®)II; Optigene). The RT-GEAR assay showed no cross-reactivity with common human enteroviruses, including HEV71, CVA16, CVA6, CVA10, CVA24, CVB5, Echo30, and PV1-3 or with other common respiratory viruses including FluA H3, FluB, PIV1-4, ADV3, RSVA, RSVB and HMPV. With in vitro-transcribed RNA containing the amplified regions of HRV-A60, HRV-B06 and HRV-C07 as templates, the sensitivity of the RT-GEAR assay was 5, 50 and 5 copies/reaction, respectively. Experiments to evaluate the clinical performance of the RT-GEAR assay were also carried out with a panel of 143 previously verified samples, and the results were compared with those obtained using a published semi-nested PCR assay followed by sequencing. The tested panel comprised 91 HRV-negative samples and 52 HRV-positive samples (18 HRV-A-positive samples, 3 HRV-B-positive samples and 31 HRV-C-positive samples). The sensitivity and specificity of the pan-HRVs RT-GEAR assay was 98.08 % and 100 %, respectively. The kappa correlation between the two methods was 0.985. The RT-GEAR assay based on a portable Genie(®)II fluorometer is a sensitive, specific and rapid assay for the universal detection of HRV infection.
人鼻病毒（HRV）长期以来被认为是超过一半的急性病毒性上呼吸道疾病的病因，并且它们与儿童的更严重疾病有关，如哮喘、急性中耳炎和肺炎。因此，对HRV感染进行快速通用检测的需求很高。在本研究中，开发了一种针对HRV 5'非翻译区（UTR）的逆转录基因组指数扩增反应（RT-GEAR）检测方法，用于泛HRV检测。该反应在单管中于65°C一步进行60分钟，使用实时荧光计（Genie(®)II；Optigene）。RT-GEAR检测方法与常见的人类肠道病毒（包括HEV71、CVA16、CVA6、CVA10、CVA24、CVB5、Echo30和PV1-3）或其他常见呼吸道病毒（包括FluA H3、FluB、PIV1-4、ADV3、RSVA、RSVB和HMPV）均无交叉反应。以含有HRV-A60、HRV-B06和HRV-C07扩增区域的体外转录RNA为模板，RT-GEAR检测方法的灵敏度分别为5、50和5拷贝/反应。还使用一组143个先前验证的样本进行了评估RT-GEAR检测方法临床性能的实验，并将结果与使用已发表的半巢式PCR检测方法随后进行测序获得的结果进行比较。测试组包括91个HRV阴性样本和52个HRV阳性样本（18个HRV-A阳性样本、3个HRV-B阳性样本和31个HRV-C阳性样本）。泛HRV RT-GEAR检测方法的灵敏度和特异性分别为98.08%和100%。两种方法之间的kappa相关性为0.985。基于便携式Genie(®)II荧光计的RT-GEAR检测方法是一种用于泛HRV感染通用检测的灵敏性、特异性和快速检测方法。
