Whole genome sequence analysis of the arctic-lineage strain responsible for distemper in Italian wolves and dogs through a fast and robust next generation sequencing protocol
Whole genome sequence analysis of the arctic-lineage strain responsible for distemper in Italian wolves and dogs through a fast and robust next generation sequencing protocol
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DOI：
10.1016/j.jviromet.2014.02.027
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发表时间：
2014-06-15
影响因子：
3.1
通讯作者：
Lorusso, Alessio
中科院分区：
文献类型：
--
作者：
Marcacci, Maurilia;Ancora, Massimo;Lorusso, Alessio
犬瘟热病毒（CDV）流行株的动态监测和特征是预防可能的疫苗突破事件的必要条件。本研究描述了用于获得CDV分离株（CDV 2784/2013）完整基因组序列的快速和稳健的下一代测序（NGS）离子PGM（TM）方案的设置。CDV 2784/2013是2013年期间在意大利引起犬和狼严重临床犬瘟热的CDV毒株的原型。在细胞培养物上分离CDV 2784/2013，并将总RNA用于NGS样品制备。使用MIRA版本4.0rc4从头组装总共112.3 Mb的读段，其产生总数为403个重叠群，覆盖率为12.1%。成功地回收了全基因组（15，690 bp），并与现有的CDV全基因组进行了比较。显示CDV 2784/2013与Onderstepoort疫苗株具有92%的nt同一性。这项研究首次描述了一种快速和稳健的基于Ion PGM（TM）平台的全基因组扩增方案，用于从总细胞纯化RNA开始的非分段负链RNA病毒。此外，这是第一项报告北极血统菌株全基因组分析的研究，该菌株已知在欧洲，亚洲和美国广泛传播。(C)2014爱思唯尔有限公司版权所有。
Dynamic surveillance and characterization of canine distemper virus (CDV) circulating strains are essential against possible vaccine breakthroughs events. This study describes the setup of a fast and robust next-generation sequencing (NGS) Ion PGM (TM) protocol that was used to obtain the complete genome sequence of a CDV isolate (CDV2784/2013). CDV2784/2013 is the prototype of CDV strains responsible for severe clinical distemper in dogs and wolves in Italy during 2013. CDV2784/2013 was isolated on cell culture and total RNA was used for NGS sample preparation. A total of 112.3 Mb of reads were assembled de nova using MIRA version 4.0rc4, which yielded a total number of 403 contigs with 12.1% coverage. The whole genome (15,690 bp) was recovered successfully and compared to those of existing CDV whole genomes. CDV2784/2013 was shown to have 92% nt identity with the Onderstepoort vaccine strain. This study describes for the first time a fast and robust Ion PGM (TM) platform-based whole genome amplification protocol for non-segmented negative stranded RNA viruses starting from total cell-purified RNA. Additionally, this is the first study reporting the whole genome analysis of an Arctic lineage strain that is known to circulate widely in Europe, Asia and USA. (C) 2014 Elsevier B.V. All rights reserved.