Alabi O J, Crosslin J M, Saidov N, Naidu R A
Department of Plant Pathology, Washington State University, Irrigated Agriculture Research and Extension Center, Prosser 99350.
USDA-ARS Vegetable and Forage Crops Research Unit, Prosser, WA 99350.
Plant Dis. 2012 Jul;96(7):1074. doi: 10.1094/PDIS-03-12-0249-PDN.
Potato (Solanum tuberosum L.) is widely grown as a staple food and cash crop in Tajikistan and is an important food security crop in the country. In June 2011, we conducted a survey of potatoes in farmers' fields in the Buston and Dushanbe regions (about 200 miles apart) of Tajikistan. Potato plants with stunted growth and leaves showing chlorotic spots, curling, and necrotic spots and rings were observed with the disease incidence monitored in 10 fields each in Buston and Dushanbe areas varying between 10 and 60%. Representative samples from symptomatic plants tested positive for Potato virus Y (PVY) using virus-specific immunostrips (Agdia Inc., Elkhart, IN). Leaf samples from symptomatic plants were collected from Buston and Dushanbe areas, imprinted on FTA Classic Cards (Whatman International Ltd., Maidstone, UK), air dried, and shipped to the lab at Washington State University for confirmatory diagnostic tests. Total nucleic acids were eluted from FTA cards (1) and subjected to reverse transcription (RT)-PCR with primers (PVY/Y4A and PVY/Y3S) specific to the coat protein of PVY (3). Samples infected with PVY ordinary strain (PVY), tuber necrosis strain (PVY), tobacco veinal necrosis strains (PVY and PVY), and a recombinant strain (PVY) were included as references to validate RT-PCR results. A single DNA product of approximately 480 bp was amplified from potato samples that tested positive with PVY-specific immunostrips. The amplified fragments from two samples from Dushanbe and six from Buston areas were cloned separately into pCR2.1 (Invitrogen Corp., Carlsbad, CA) and two independent clones per amplicon were sequenced from both orientations. Pairwise comparison of these sequences showed 90 to 100% identity among the cloned amplicons (GenBank Accession Nos. JQ743609 to JQ743616) and 90 to 100% with corresponding nucleotide sequence of reference PVY strains (GenBank Accession Nos. JQ743617 to JQ743621). A global phylogenetic analysis of sequences revealed the presence of PVY in both samples from Dushanbe and one sample from Buston regions and presence of PVY in the remaining five samples from the Buston region. Because of the possible occurrence of mixed infections of PVY strains (2), further studies are needed to determine the presence of mixed infections of two or more strains of PVY and their specificity to potato cultivars. To our knowledge, this study represents the first confirmed report of two distinct strains of PVY in potato in Tajikistan. The occurrence of PVY, a quarantine pathogen in many countries (2), warrants additional investigations to improve sanitary status of potato fields and to facilitate the availability of virus-free seed in clean plant programs for significant yield increases in Tajikistan. References: (1) O. J. Alabi et al. J. Virol. Methods 154:111, 2008. (2) S. Gray et al. Plant Dis. 94:1384, 2010. (3) R. P. Singh et al. J. Virol. Methods 59:189, 1996.
土豆(Solanum tuberosum L.)在塔吉克斯坦作为主食和经济作物广泛种植,是该国重要的粮食安全作物。2011年6月,我们对塔吉克斯坦布斯顿和杜尚别地区(相距约200英里)农民田地里的土豆进行了调查。观察到生长发育不良的土豆植株,其叶片出现褪绿斑点、卷曲以及坏死斑点和环纹,在布斯顿和杜尚别地区各10块田地中监测到的发病率在10%至60%之间。使用病毒特异性免疫试纸条(Agdia公司,美国印第安纳州埃尔克哈特)对有症状植株的代表性样本进行检测,结果显示感染了马铃薯Y病毒(PVY)。从布斯顿和杜尚别地区有症状的植株上采集叶片样本,印在FTA Classic卡片(Whatman国际有限公司,英国梅德斯通)上,风干后运往华盛顿州立大学实验室进行确诊诊断检测。从FTA卡片上洗脱总核酸(1),并用针对PVY外壳蛋白的引物(PVY/Y4A和PVY/Y3S)进行逆转录(RT)-PCR(3)。感染PVY普通株系(PVY)、块茎坏死株系(PVY)、烟草脉坏死株系(PVY和PVY)以及重组株系(PVY)的样本作为对照,用于验证RT-PCR结果。从用PVY特异性免疫试纸条检测呈阳性的土豆样本中扩增出一条约480 bp的单一DNA产物。从杜尚别地区的两个样本和布斯顿地区的六个样本中扩增得到的片段分别克隆到pCR2.1(Invitrogen公司,美国加利福尼亚州卡尔斯巴德)中,每个扩增子的两个独立克隆从两个方向进行测序。这些序列的两两比较显示,克隆的扩增子之间的同一性为90%至100%(GenBank登录号JQ743609至JQ743616),与参考PVY株系的相应核苷酸序列的同一性为90%至100%(GenBank登录号JQ743617至JQ743621)。对序列进行的全球系统发育分析表明,在杜尚别地区的两个样本和布斯顿地区的一个样本中存在PVY普通株系,在布斯顿地区的其余五个样本中存在PVY坏死株系。由于可能存在PVY株系的混合感染(2),需要进一步研究以确定是否存在两种或更多PVY株系的混合感染及其对土豆品种的特异性。据我们所知,本研究是塔吉克斯坦首次关于土豆中两种不同PVY株系的确诊报告。PVY在许多国家属于检疫性病原菌(2),其出现需要进行更多调查,以改善土豆田的卫生状况,并在清洁植物计划中促进无病毒种子的供应,从而在塔吉克斯坦大幅提高产量。参考文献:(1) O. J. Alabi等人,《病毒学方法杂志》154:111,2008年。(2) S. Gray等人,《植物病害》94:1384,2010年。(3) R. P. Singh等人,《病毒学方法杂志》59:189,1996年。