リバースジェネティクス法を用いた新遺伝子型ＲＳウイルス（ＯＮ１）の増殖能の解析

我在2013年、2014年和2015年的三个季节中分析了菲律宾Biliran岛HRSV的分子特征。鼻咽拭子采集自Biliran省立医院的重症肺炎住院儿童。通过使用实时聚合酶链反应（PCR）筛选样品。采用常规PCR和G基因第二高变区（HVR）测序确定基因型。在检测的1，240份样本中，356份（28.7%）为HRSV阳性，包括183份（51.4%）HRSV-A和173份（48.6%）HRSV-B。HRSV-A又分为两种基因型：NA 1和ON 1。NA 1和ON 1在2013年共同循环，只有ON 1在2014年仍在循环。2014年检测到一种新的ON 1变体，其在遗传学上与其他ON 1菌株不同。这些新的ON 1病毒在G基因的第二个HVR中具有三个独特的氨基酸替换（K212 R、K213 G和K223 G），这在其他国家还没有发现。除两份样本被鉴定为GB 2外，所有HRSV-B病毒均被鉴定为基因型BA 9。2013年和2014年两个季节HRSV-A和-B均为共流，但2015年季节检出的HRSV以HRSV-B为主。G基因第2个HVR的大部分氨基酸位点处于纯化/负选择状态。这项研究可以建立在分子特征的基础上，为菲律宾HRSV的分子遗传学研究做出贡献。此外，我去年在菲律宾完成了HRSV-A的完整基因组序列的手稿。

I analyzed molecular characterization of HRSV in Biliran Island, Philippines over three seasons in 2013, 2014, and 2015. Nasopharyngeal swabs were collected from hospitalized children with severe pneumonia at the Biliran Provincial Hospital. Samples were screened by using real-time polymerase chain reaction (PCR). Genotypes were determined using conventional PCR and sequencing of the 2nd hypervariable region (HVR) of the G gene. Of 1,240 sample tested, 356 (28.7%) were positive for HRSV including 183 (51.4%) HRSV-A and 173 (48.6%) HRSV-B. HRSV-A were further divided into two genotypes; NA1 and ON1. Both NA1 and ON1 were co-circulating in 2013, only ON1 remained to be circulating in 2014. A novel ON1 variant, which was phylogenetically distinct from other ON1 strains, was detected in 2014. These novel ON1 viruses possess three unique amino acid substitutions (K212R, K213G, and K223G) in the 2nd HVR of the G gene, which had not been found in other countries. All HRSV-B viruses were identified as genotype BA9 except two samples identified as GB2. Both HRSV-A and -B were co-circulating in 2013 and 2014 seasons, but most of HRSV detected in 2015 season were HRSV-B. Most of amino acid sites of the 2nd HVR of the G gene were under purifying/negative selection. This study can build on the molecular characterization to contribute the advance of molecular genetics on HRSV in the Philippines. Moreover, I made the manuscript of complete genome sequences of HRSV-A in the Philippines that I did last year.