A complete genome of a novel ectromelia virus isolated in Guangdong province, China.

Samples used for virus isolation were obtained from Rattus rattoides during a field investigation of scrub typhus in Guangzhou, Guangdong Province, in 2009. We inoculated the original samples onto Vero and HeLa cells. An obvious cytopathic Effect (CPE) was observed on day three from a spleen tissue samples collected from Rattus rattoides. Total genomic DNA was extracted from a 200 µL virus stock using the QIAamp DNA Mini Kit.Long reads were generated via the GridION sequencing platform, and genome assembly was conducted using Canu v2.2 and polished with NGS datasets. Genomic annotation was performed using Softberry FGENESV. Through Nanopore sequencing, a contiguous linear genomic sequence spanning 208,281 bp was obtained, leading to the identification of ECTV-C-Tan-GD01. This strain shows an A/T typical OPV composition of 67%. Evolutionary analysis with other ectromelia virus strains and representatives of other orthopoxviruses revealed that ECTV-C-Tan-GD01 clusters with other ECTV strains. The annotated ECTV-C-Tan-GD01 genome comprised 199 putative genes, including 1 gene truncated at the C-terminal, with 91 and 108 genes on the positive and negative strands, respectively.

2009年，在广东省广州市开展恙虫病（scrub typhus）野外调查期间，我们从黄毛鼠（Rattus rattoides）体内获取了用于病毒分离的样本。我们将原始样本接种至Vero细胞与HeLa细胞中。从该黄毛鼠采集的脾脏组织样本中，接种后第3天即可观察到明显的细胞病变效应（CPE）。我们使用QIAamp DNA迷你提取试剂盒（QIAamp DNA Mini Kit）从200 μL病毒原液中提取了总基因组DNA。利用GridION测序平台生成长读长测序数据，并使用Canu v2.2进行基因组组装，再通过下一代测序（Next-Generation Sequencing, NGS）数据集完成序列抛光。使用Softberry公司的FGENESV软件完成基因组注释。通过纳米孔（Nanopore）测序，我们获得了一条长度为208281 bp的连续线性基因组序列，由此鉴定出病毒毒株ECTV-C-Tan-GD01。该毒株的A/T碱基占比为67%，符合正痘病毒（Orthopoxvirus, OPV）的典型碱基组成特征。针对其他鼠痘病毒（ectromelia virus, ECTV）毒株以及其他正痘病毒属代表毒株开展进化分析后发现，ECTV-C-Tan-GD01与其他鼠痘病毒毒株聚类为同一分支。经注释的ECTV-C-Tan-GD01基因组共包含199个推定基因，其中1个基因在C端发生截短，正链与负链上分别分布有91个和108个基因。