Beyond the Consensus: Dissecting Within-Host Viral Population Diversity of Foot-and-Mouth Disease Virus by Using Next-Generation Genome Sequencing
Beyond the Consensus: Dissecting Within-Host Viral Population Diversity of Foot-and-Mouth Disease Virus by Using Next-Generation Genome Sequencing
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DOI：
10.1128/jvi.01396-10
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发表时间：
2011-03-01
影响因子：
5.4
通讯作者：
King, Donald P.
中科院分区：
文献类型：
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作者：
Wright, Caroline F.;Morelli, Marco J.;King, Donald P.
个体宿主内病毒群体的不同序列是RNA病毒（如口蹄疫病毒（FMDV））的选择和随后进化的起始材料。使用在基因组分析仪平台（Illumina）上进行的下一代测序（NGS），本研究比较了来自单个动物的两个牛上皮样本（足部病变）中的病毒群与用于启动实验感染的接种物。在重复测序运行中测定基因组序列，并且通过NGS测定的接种物的共有序列与先前使用桑格方法测定的一致。然而，NGS揭示了病毒群体的精细多态性亚结构，从以略低于50%的频率存在的核苷酸变异到以1%的分数存在的核苷酸变异。一些频率较高的多态性确定了与硫酸乙酰肝素结合相关的密码子内的编码变化，并存在于两个足部病变中，揭示了在哺乳动物宿主内复制的组织培养适应病毒进化的中间阶段。我们在接种物和两个足部病变中分别确定了2，622、1，434和1，703个多态性：大多数替换仅发生在一小部分人群中，代表了在任何选择压力发生之前最近细胞复制的后代。我们估计细胞内病毒全基因组突变率的上限为7.8 × 10 - 4个核苷酸。通过NGS实现的更大的检测深度表明，该方法是用于解剖宿主内的FMDV群体的强大且有价值的工具。
The diverse sequences of viral populations within individual hosts are the starting material for selection and subsequent evolution of RNA viruses such as foot-and-mouth disease virus (FMDV). Using next-generation sequencing (NGS) performed on a Genome Analyzer platform (Illumina), this study compared the viral populations within two bovine epithelial samples (foot lesions) from a single animal with the inoculum used to initiate experimental infection. Genomic sequences were determined in duplicate sequencing runs, and the consensus sequence of the inoculum determined by NGS was identical to that previously determined using the Sanger method. However, NGS revealed the fine polymorphic substructure of the viral population, from nucleotide variants present at just below 50% frequency to those present at fractions of 1%. Some of the higher-frequency polymorphisms identified encoded changes within codons associated with heparan sulfate binding and were present in both foot lesions, revealing intermediate stages in the evolution of a tissue culture-adapted virus replicating within a mammalian host. We identified 2,622, 1,434, and 1,703 polymorphisms in the inoculum and in the two foot lesions, respectively: most of the substitutions occurred in only a small fraction of the population and represented the progeny from recent cellular replication prior to onset of any selective pressures. We estimated the upper limit for the genome-wide mutation rate of the virus within a cell to be 7.8 x 10(-4) per nucleotide. The greater depth of detection achieved by NGS demonstrates that this method is a powerful and valuable tool for the dissection of FMDV populations within hosts.