Symes S J, Gunesekere I C, Marshall J A, Wright P J
Department of Microbiology, Monash University, Clayton, Victoria, Australia.
Arch Virol. 2007;152(6):1075-86. doi: 10.1007/s00705-007-0938-9. Epub 2007 Feb 28.
We describe an outbreak of gastroenteritis in which the nucleic acid of three distinct noroviruses was amplified from the same fecal sample. To enable the separate amplification of each virus, an inclusion/exclusion RT-PCR primer design strategy was developed. This paired a virus-specific exclusion primer (designed with the exact sequence of one virus in a region displaying low conservation among the three viruses) with a virus-nonspecific inclusion primer (designed in a conserved region). Thus, in each reaction the exclusion primer provided specificity for a single virus, and the inclusion primer increased the sensitivity and allowed hybridization in a region of unknown sequence. Analysis of the partial genomic sequences of the three viruses (3.6-3.8 kb) indicated that each virus belonged to a separate genogroup II cluster, and each displayed evidence of a potential recombination event when the sequences were compared with other published norovirus sequences. Our results, which show a mixed norovirus infection in a single individual, confirm the need to be aware of the possibility of mixed norovirus infections, and of the possibility of genomic recombination causing anomalies in phylogenetic analyses in such instances.
我们描述了一起肠胃炎暴发事件，在该事件中，从同一个粪便样本中扩增出了三种不同诺如病毒的核酸。为了能够分别扩增每种病毒，我们开发了一种包含/排除逆转录聚合酶链反应（RT-PCR）引物设计策略。该策略将病毒特异性排除引物（在三种病毒间保守性较低的区域设计，其序列与一种病毒的精确序列相同）与病毒非特异性包含引物（在保守区域设计）配对。因此，在每个反应中，排除引物为单一病毒提供特异性，而包含引物提高了灵敏度，并允许在未知序列区域进行杂交。对这三种病毒部分基因组序列（3.6 - 3.8 kb）的分析表明，每种病毒都属于不同的基因组II簇，并且当将这些序列与其他已发表的诺如病毒序列进行比较时，每种病毒都显示出潜在重组事件的证据。我们的结果显示在单个个体中存在混合诺如病毒感染，证实有必要意识到混合诺如病毒感染的可能性，以及在这种情况下基因组重组可能导致系统发育分析出现异常的可能性。
