【摘要】 为明确苹果坏死花叶病毒（Apple necrotic mosaic virus, ApNMV）在河北的发生及其基因组特性，本试验利用RT-PCR技术对河北保定两地ApNMV发病情况进行检测，然后通过高通量测序技术对带毒样本进行序列测定和分析。结果表明，河北农大果园‘红珍珠’‘锦锈红’‘信侬红’3个品种及保定唐县苹果实验基地富士品种ApNMV检出率分别为9.09%、18.18%、100%和65%。ApNMV阳性果树高通量测序结果显示，果树为ApNMV、苹果褪绿叶斑病毒（Apple chlorotic leaf spot virus, ACLSV）、苹果茎沟病毒（Apple stem grooving virus, ASGV）和苹果茎痘病毒（Apple stem pitting virus, ASPV）复合侵染，ApNMV河北分离物（ApNMV-HB）的近全基因组序列包含RNA1～RNA3序列，GenBank登录号分别为OP019626、OP019627和OP019628。RNA1编码120 kD的甲基转移酶/NTP-binding解旋酶（MET/HEL）,RNA2编码97 kD的RNA聚合酶（POL）,RNA3编码运动蛋白（Movement protein, MP）和外壳蛋白（Coatprotein,CP）,4个基因核苷酸和氨基酸序列与代表性分离物相似性分别为89.44%～97.02%、92.14%～98.67%。系统发育关系分析表明，以ApNMV RNA1、RNA2、RNA3(CP和MP)编码的氨基酸为基础构建系统发育树，ApNMV-HB分别与AM95、AM125、XJ、ZZ亲缘关系最近。本研究首次揭示了ApNMV河北分离物的近全基因组序列，丰富了该病毒的基因组序列信息，明确了该分离物与其它分离物的系统发育关系，为进一步了解该病毒的系统发育及遗传进化提供参考。更多还原

【Abstract】 In order to clarify the occurrence and genomic characteristics of apple necrotic mosaic virus(ApNMV) in Hebei, the incidence of ApNMV in two regions of Baoding was detected by RT-PCR, and then the infected sample was sequenced by high-throughput sequencing technology. The results showed that the positive rates of ApNMV were 9.09%, 18.18%, 100% and 65% in the cultivars of ‘Hongzhenzhu’, ‘Jinxiuhong’ and ‘Xinnonghong’ in the Apple Experimental Garden of Agricultural University of Hebei, and ‘Fuji’ in the the Apple Experimental Site of Tangxian County, Baoding. High-throughput sequencing revealed that the apple trees infected by ApNMV were also infected by apple chlorotic leaf spot virus(ACLSV), apple stem grooving virus(ASGV) and apple stem pitting virus(ASPV). The near-complete genome sequence of ApNMV was obtained from the sequencing results. The RNA1～RNA3 GenBank accession numbers were OP019626, OP019627 and OP019628, respectively. The isolate was named ApNMV Hebei isolate(ApNMV-HB). RNA1 encoded a 120 kD methyltransferase/NTP-binding helicase(MET/HEL). RNA2 encoded a 97 kD RNA polymerase(POL). And RNA3 encoded movement protein(MP) and coat protein(CP). The four genes shared sequence similarity with the representative isolates in 89.44%—97.02%of the nucleotide and 92.14%—98.67% of the amino acid. The phylogenetic analysis showed that all representative isolates were clustered in one branch based on the amino acids of ApNMV RNA1, and ApNMV-HB was closest to AM95. All representative isolates were clustered in one branch, except for AM72, based on the amino acids encoded by ApNMV RNA2. ApNMV-HB was closest related to AM125. In the phylogenetic tree based on ApNMV CP, the representative isolates clustered in one branch rooted with AM72, and ApNMV-HB and XJ were relatively close. In the phylogenetic tree based on ApNMV MP, all isolates were divided into 4 branches. ApNMV-HB was clustered with AM75, ApNMV-Qu, AM125 and ZZ in one branch, and was closest to ZZ. We revealed the near-complete genome sequence of the ApNMV Hebei isolate for the first time, which enriches the genome sequence information of the virus. Clarification of the phylogenetic relationship between the isolate and other isolates provides reference for further understanding of the phylogeny and genetic evolution of the virus.更多还原