Identification of a New Enamovirus Associated with Citrus Vein Enation Disease by Deep Sequencing of Small RNAs
Identification of a New Enamovirus Associated with Citrus Vein Enation Disease by Deep Sequencing of Small RNAs
复制标题
DOI：
10.1094/phyto-03-13-0068-r
复制
发表时间：
2013-10-01
期刊：
影响因子：
3.2
通讯作者：
Navarro, Luis
中科院分区：
文献类型：
--
作者：
Carmen Vives, Mari;Velazquez, Karelia;Navarro, Luis
为了确定柑橘叶脉病的致病因子，我们通过深度测序 (Solexa-Illumina) 检查了受感染和健康 Etrog 香橼植物的小 RNA (sRNA) 部分。我们的结果表明，病毒衍生的 sRNA (vsRNA)：(i) 约占总 sRNA 群体的 14.21%，(ii) 主要由 21 和 24 个核苷酸组成，其 5' 核苷酸分布有偏差，并且 (+) 极性的分布明显普遍，(iii) 源自所有病毒基因组，尽管突出的热点存在于 5' 近端区域。由 vsRNA 组装而成的重叠群显示出与黄病毒序列的相似性，特别是与豌豆花叶病毒（Enamovirus 属的类型成员）的相似性。一种新病毒（暂时命名为柑橘脉病毒（CVEV））的基因组 RNA (gRNA) 序列已完成并进行了表征。 CVEV gRNA 是单链、正义的，大小为 5,983 个核苷酸，有 5 个开放阅读框。基于 RNA 聚合酶和外壳蛋白的氨基酸特征的系统发育比较清楚地将 CVEV 归入 Enamovirus 属。开发了斑点印迹杂交和逆转录聚合酶链反应测试来检测受静脉生成病影响的植物中的 CVEV。这些方法的 CVEV 检测已被用于西班牙柑橘检疫、卫生和认证项目。
To identify the causal agent of citrus vein enation disease, we examined by deep sequencing (Solexa-Illumina) the small RNA (sRNA) fraction from infected and healthy Etrog citron plants. Our results showed that virus-derived sRNAs (vsRNAs): (i) represent about 14.21% of the total sRNA population, (ii) are predominantly of 21 and 24 nucleotides with a biased distribution of their 5' nucleotide and with a clear prevalence of those of (+) polarity, and (iii) derive from all the viral genome, although a prominent hotspot is present at a 5'-proximal region. Contigs assembled from vsRNAs showed similarity with luteovirus sequences, particularly with Pea enation mosaic virus, the type member of the genus Enamovirus. The genomic RNA (gRNA) sequence of a new virus, provisionally named Citrus vein enation virus (CVEV), was completed and characterized. The CVEV gRNA was found to be single-stranded, positive-sense, with a size of 5,983 nucleotides and five open reading frames. Phylogenetic comparisons based on amino acid signatures of the RNA polymerase and the coat protein clearly classifies CVEV within the genus Enamovirus. Dot-blot hybridization and reverse transcription-polymerase chain reaction tests were developed to detect CVEV in plants affected by vein enation disease. CVEV detection by these methods has already been adopted for use in the Spanish citrus quarantine, sanitation, and certification programs.