滇重楼新病毒ParV1的分子变异分析及致病性测定  

Molecular variation analysis and pathogenicity determination of ParV1,a novel virus from Paris polyphylla var.yunnanensis

作　　者：王喆 陈潞 魏朝霞 余代宏 陈泽历 高丽珂 李尚贇 文国松[2] 赵明富[1] WANG Zhe;CHEN Lu;WEI Zhao-xia;YU Dai-hong;CHEN Ze-li;GAO Li-ke;LI Shang-yun;WEN Guo-song;ZHAO Ming-fu(Key Laboratory of Agricultural Biodiversity and Disease Control,Ministry of Education,Yunnan Agricultural University,Kunming 650201,China;College of Agronomy and Biotechnology,Yunnan Agricultural University,Kunming 650201,China;Yuanjiang County Plant Protection and Plant Inspection Station,Yuxi 653301,Yunnan,China)

机构地区：[1]云南农业大学农业生物多样性与病害控制教育部重点实验室,昆明650201 [2]云南农业大学农学与生物技术学院,昆明650201 [3]元江县植保植检站,云南玉溪653301

出　　处：《湖北农业科学》2022年第19期152-159,163,共9页Hubei Agricultural Sciences

基　　金：国家自然科学基金项目(81860774)。

摘　　要：为明确云南省引起滇重楼(Paris polyphylla var.yunnanensis)病毒病的病原,以昆明市发现的花叶、斑驳症状的滇重楼叶片为样品,进行透射电镜观察、高通量测序和反转录(RT-PCR)检测。结果表明,观察到大小约(700~1000)nm×13 nm的线状病毒粒子;基于高通量测序结果设计特异性引物进行RTPCR扩增并克隆,得到1条全长为10066个核苷酸(nt)的病毒序列,将该病毒暂命名为Paris virus 1(ParV1);BLAST比对分析表明,ParV1与马铃薯Y病毒属(Potyvirus)中的浙贝母花叶病毒(Thunberg fritillary mosaic virus,TFMV)具有最高为65.1%、68.9%的核苷酸及氨基酸(aa)序列同源性;多序列比对分析显示,ParV1多聚蛋白与potyviruses具有49.6%~65.1%的nt序列同源性及47.1%~68.9%的aa序列同源性;基于多聚蛋白氨基酸序列构建的系统发育树表明,该病毒与TFMV亲缘关系较近且聚为一个单独的分支。综上所述,确定ParV1属于Potyvirus的一种新病毒。将含ParV1的病毒汁液以机械摩擦的方法接种到心叶烟、本氏烟上对ParV1进行致病性初步测定。接种后的心叶烟植株叶片表现出明显病毒病症状,经RT-PCR及分子克隆在接种的心叶烟中检测到ParV1的存在,表明心叶烟植株受到ParV1的侵染,而接种的本氏烟上暂未检测到该病毒的存在。In order to clarify the pathogen of Paris polyphylla var.yunnanensis virus disease in Yunnan Province,the leaves of Paris polyphylla var.yunnanensis with mosaic and mottle symptoms found in Kunming City were used as samples for transmission electron microscopy,high-throughput sequencing and RT-PCR detection.The results showed that the linear virus particles with a size of about(700~1000)nm×13 nm were observed.Based on the results of high-throughput sequencing,specific primers were designed for RTPCR amplification and cloning,and a virus sequence with a total length of 10066 nucleotides(nt)was obtained.The virus was temporarily named Paris virus 1(ParV1);BLAST alignment analysis showed that ParV1 had the highest nucleotide and amino acid(aa)sequence homology of 65.1% and 68.9% with Thunberg fritillary mosaic virus(TFMV)in the genus Potyvirus.Multiple sequence alignment analysis showed that ParV1 had 49.6%~65.1% nt sequence homology and 47.1%~68.9% aa sequence homology with potyviruses.The phylogenetic tree based on the polyprotein amino acid sequence showed that the virus was closely related to TFMV and clustered into a separate branch.In summary,ParV1 is a new virus of Potyvirus.The virus juice containing ParV1 was inoculated into Nicotiana tabacum and Nicotiana benthamiana by mechanical friction to determine the pathogenicity of ParV1.The leaves of the inoculated Nicotiana glutinosa plants showed obvious symptoms of virus disease.The presence of ParV1 was detected in the inoculated Nicotiana glutinosa by RT-PCR and molecular cloning,indicating that the Nicotiana glutinosa plants were infected by ParV1,but the presence of the virus was not detected on the inoculated Nicotiana benthamiana.

关 键 词：滇重楼(Paris polyphylla var.yunnanensis) Paris virus 1 致病性测定 

分 类 号：S432.41[农业科学—植物病理学]