四川番茄黄化曲叶病病原分子鉴定及变异分析

摘要

[目的]明确引起四川番茄黄化曲叶病(Tomato yellow leaf Curl disease,TYLCD)的病原.[方法]对采自四川攀枝花市田间表现矮化、黄化和曲叶症状的番茄植株SC64-67,通过PCR、克隆及测序等技术获得病毒及卫星DNA的全基因组序列,并对序列进行变异分析.[结果]利用双生病毒简并引物PA/PB从4个样品中均扩增得到约500 bp的片段,随机选择SC65进行DNA-A全基因组扩增和序列测定,该DNA分子全长为2 732 nts,系统进化分析表明,其与已报道的中国番茄黄化曲叶病毒(Tomato yellow leaf curl China vlrus,TYLCCNV)来自云南元谋的菜豆分离物(TYLCCNV-Bean-YM)的核苷酸序列相似性最高,为96.0%.检测发现,所有分高物均伴随有卫星DNAβ分子.全序列测定表明SC65 DNAp全长1 338 nts,与分离自云南楚雄番茄上的TYLCCNV-Y25伴随的卫星DNAβ亲缘关系最近,核苷酸序列相似性为77.5%.[结论]研究表明四川番茄黄化曲叶病样品均受到TYLCCNV/DNAβ病害复合体的侵染,但其病毒DNA-A和卫星DNAβ分子来源于TYLCCNV的不同分离物.%[ Objective] In this study, pathogenic agents associated with tomato yellow leaf curl disease in Panzhihua, Sichuan Province in China were identified. [Method] Diseased tomato plants exhabiting typical stunting, yellowing and leaf-curling symptoms were collected in Panzhihua, Sichuan Province. Using PCR, cloning, and sequencing techniques a begomovirus disease complex associated with tomato was identified for phylogenetic analysis. [Result] 500 bp fragments were amplified from four samples (SC64-67) using the begomovirus degenerate primer pair PA/PB. The complete nucleotide sequence of isolate SC65 was determined to be 2 732 nts and shared the highest sequence identity (96.0％) with an isolate of Tomato yellow leaf curl China virus (TYLCCNV-Bean-YM) isolated from kidney bean in Yuanmou, Yunnan Province. All tomato samples were found to be associated with the satellite DNAβ molecules. The satellite DNAβ molecule associated with SC65 (SC65 DNAβ) consisted of 1 338 nts and shared 77.5％ nucleotide sequence identity with DNAβ associated with TYLCCNV-Y25 isolated from tomato plants in Chuxiong,Yunnan Province. [ Conclusion ] These results suggested that tomato yellow leaf curl disease in Sichuan Province is associated with the TYLCCNV/DNAβ disease complex which may originate from DNA-A and DNAβ molecules of different TYLCCNV isolates.