猕猴桃病毒A和B的RT-PCR检测及分子变异初步研究  

The Detection of Actinidia virus A and Actinidia virus B by RT-PCR and Their Molecular Variation Analysis

作　　者：郑亚洲 王国平[1,2] 周菊芳[1] 朱晨熹 王利平[1] 徐文兴[1] 洪霓[1,2] 

机构地区：[1]华中农业大学植物科技学院湖北省作物病害监测与安全控制重点实验室,武汉430070 [2]农业微生物学国家重点实验室,武汉430070

出　　处：《园艺学报》2015年第4期665-671,共7页Acta Horticulturae Sinica

基　　金：国家公益性行业(农业)科研专项项目(nyhyzx200903017-08)

摘　　要：为明确猕猴桃病毒A(Actinidia virus A,Ac VA)和猕猴桃病毒B(Actinidia virus B,Ac VB)在中国猕猴桃(Actinidia sp.)上的侵染状况和分子特性,采用RT-PCR技术对151份猕猴桃样品的Ac VA和Ac VB进行检测,检出率分别为15.2%和17.9%,所收集的6种猕猴桃样品中均检测到一种或两种病毒。发现Ac VA的ORF1、ORF4和ORF5扩增片段序列存在较大分子变异,不同分离物的269 bp片段(ORF1)核苷酸序列相似性为77.3%~99.3%。在基于各扩增片段核苷酸序列的系统进化树中,Ac VA分离物聚为2~3组。采用引物Ac VB5F/5R扩增获得20个Ac VB分离物的342 bp或338 bp的ORF5片段,核苷酸序列相似性为81%~100%,与新西兰Ac VB分离物TP7-93B相应片段的核苷酸序列相似性为83.9%~99.7%,在系统进化树中聚为3组。To understand the infectious status and molecular characteristics of Actinidia virus A(Ac VA)and Actinidia virus B(Ac VB)in kiwifruit(Actinidia sp.)grown in China,151 plants of kiwifruit were tested for the presence of the two viruses by Reverse transcription PCR(RT-PCR). Results revealed that the average infection frequencies of Ac VA and Ac VB in kiwifruit samples were 15.2% and 17.9%,respectively. All six Actinidia species tested were positive to one or two viruses. Sequence analyzes of amplified fragments from ORF,ORF4 and ORF5 of Ac VA genome showed that the virus isolates were molecularly divergent and the 269 bp fragments of ORF1 from different isolates shared nucleotide similarities of 77.3%–99.3%. In phylogenetic trees based on nucleotide sequences of three fragments,all Ac VA isolates separated into two or three clades. The ORF5 fragment of 20 Ac VB isolates amplified using primer set Ac VB5F/5R had a size of 342 bp or 338 bp. All those isolates shared nucleotide similarities of 81%–100% with each other and 83.9%–99.7% with the corresponding sequence of the New Zealand isolate TP7-93 B,and clustered into three clades in the phylogenetic tree constructed basing on nucleotide sequences of the fragment.

关 键 词：猕猴桃 猕猴桃病毒A 猕猴桃病毒B RT-PCR 序列分析 

分 类 号：S436.634.1[农业科学—农业昆虫与害虫防治]