Specific primer amplification of the VP1 region directed by 5′ UTR sequence analysis: Enterovirus testing and identification in clinical samples from hand-foot-and-mouth disease patients
Specific primer amplification of the VP1 region directed by 5′ UTR sequence analysis: Enterovirus testing and identification in clinical samples from hand-foot-and-mouth disease patients
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DOI：
10.1016/j.jviromet.2013.06.009
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发表时间：
2013-11-01
影响因子：
3.1
通讯作者：
Xia, Ningshao
中科院分区：
文献类型：
--
作者：
Ge, Shengxiang;Yan, Qiang;Xia, Ningshao
肠道病毒(EV)病原体的多种基因型别可引起临床手足口病(HFMD)。因此，手足口病病原体的快速鉴定和监测可能很困难，特别是从原始临床标本中。本研究设计了两套通用的泛肠道病毒和EV71/CA16VP1特异性引物，并用于检测手足口病患者的临床标本。根据5‘非翻译区和VP1扩增产物的初步序列分析，重新设计了VP1区的额外引物，用于对剩余的小部分非EV71/非CA16标本进行进一步的基因分型。在已知的专家小组中，使用5‘-UTR序列分型和VP1分型可以识别16个成员中的15个，这表明该方法具有良好的可检测性和基因分型。1株未经5‘-UTR分型的毒株为重组病毒。应用该方法对44例手足口病疑似患者的临床标本进行检测，检出41例EV阳性。只有一例未能鉴定出VP1序列。检出CA16(26/41，63.4%)、EV71-C4(6/41，14.6%)、CA6(5/41，12.2%)和CA10(3/41，7.3%)4种EV。总之，5‘端非编码区扩增测序和随后的VP1特异性引物扩增确保了在临床样本检测中的高检出率和良好的基因分型准确性。这一检测策略可用于手足口病的常规评估和监测，以跟踪当地EV感染的趋势。(C)Elsevier B.V.出版的2013年
Many genotypes of the enterovirus (EV) pathogens can cause clinical hand-foot-and-mouth disease (HFMD). Therefore, rapid identification and monitoring of HFMD pathogens can be difficult, especially from the original clinical specimens. In this study, both universal pan-enterovirus and EV71/CA16 VP1-specific primer sets were designed and used to examine clinical specimens from HFMD patients. Based on the initial sequence analysis of the 5'-untanslated region (5'-UTR) and VP1 amplification products, additional primers for the VP1 region were redesigned for further genotyping of the remaining small portion non-EV71/non-CA16 specimens. With a known panel, it was possible to identify 15 out of 16 members using 5'-UTR sequence typing and VP1 typing, suggesting good detectability and genotyping of this method. One strain that was not typed by 5'-UTR was shown to be a recombinant virus. When this method was applied to examine clinical specimens from 44 suspected HFMD patients, 41 were detected as EV positive. In only one case, the VP1 sequence could not be identified. Four types of EVs, including CA16 (26/41, 63.4%), EV71-C4 (6/41, 14.6%), CA6 (5/41, 12.2%) and CA10 (3/41, 7.3%), were detected. In conclusion, 5' UTR amplification sequencing and subsequent VP1 specific primer amplification ensures a high detection rate and good genotyping accuracy in the examination of clinical samples. This detection strategy can be used for routine evaluation and monitoring of HFMD to follow local trends of EV infection. (C) 2013 Published by Elsevier B.V.