Jossey S, Babadoost M
Department of Crop Sciences, University of Illinois, Urbana 61801.
Plant Dis. 2006 Oct;90(10):1361. doi: 10.1094/PD-90-1361B.
During a survey of commercial pumpkin and squash fields for viruses, conducted in Illinois in 2005, Tobacco ringspot virus (TRSV) was identified for the first time in symptomatic pumpkin samples collected during August and September from Douglas, Kankakee, Piatt, and Tazewell counties in one of three, one of three, one of one, and one of seven samples tested, respectively. In an earlier study from southern Illinois, the only viruses detected in pumpkins were Cucumber mosaic virus, Papaya ringspot virus, Squash mosaic virus, Watermelon mosaic virus, and Zucchini yellow mosaic virus (2). TRSV has been reported in cucurbits from some states in the United States (1). We detected TRSV in symptomatic leaves exhibiting mild mosaic with leaf yellowing using a double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) kit (Agdia, Inc., Elkhart, IN). Samples were considered positive if the absorbance readings at 405 nm exceeded 3× the absorbance of the negative control. The presence of TRSV was confirmed by reverse transcription-polymerase chain reactions (RT-PCR). Total RNA was extracted from the symptomatic plants using TRIzol Reagent and reverse transcribed by M-MLV Reverse Transcriptase (Invitrogen, Carlsbad, CA). PCR was conducted using forward primer 5'-CTTGCGGCCCAAATCT ATAA-3' and reverse primer 5'-ACTTGTGCCCAGGAGAGCTA-3', which anneal to the conserved region in the coat protein gene. The reaction produced an amplification product of the expected size (348 bp). Hence, utilizing ELISA and RT-PCR tests, the presence of TRSV in pumpkin was determined, to our knowledge, for the first time in Illinois. References: (1) R. Provvidenti. Tobacco ringspot. Page 42 in: Compendium of Cucurbit Diseases. T. A. Zitter et al., eds. The American Phytopathological Society, St. Paul, MN. 1996. (2) S. A. Walters et al. HortScience 38:65, 2003.
2005年在伊利诺伊州对商业南瓜和西葫芦种植田进行病毒调查期间，首次在8月和9月从道格拉斯县、坎卡基县、皮亚特县和塔泽韦尔县采集的有症状南瓜样本中鉴定出烟草环斑病毒（TRSV），在分别检测的三个样本中的一个、三个样本中的一个、一个样本以及七个样本中的一个中检测到该病毒。在伊利诺伊州南部的一项早期研究中，在南瓜中检测到的唯一病毒是黄瓜花叶病毒、木瓜环斑病毒、西葫芦花叶病毒、西瓜花叶病毒和小西葫芦黄花叶病毒（2）。在美国的一些州，已在葫芦科作物中报道过烟草环斑病毒（1）。我们使用双抗体夹心酶联免疫吸附测定（DAS - ELISA）试剂盒（Agdia公司，印第安纳州埃尔克哈特），在表现出轻度花叶并伴有叶片发黄的有症状叶片中检测到了烟草环斑病毒。如果405nm处的吸光度读数超过阴性对照吸光度的3倍，则样本被视为阳性。通过逆转录 - 聚合酶链反应（RT - PCR）确认了烟草环斑病毒的存在。使用TRIzol试剂从有症状的植物中提取总RNA，并通过M - MLV逆转录酶（Invitrogen公司，加利福尼亚州卡尔斯巴德）进行逆转录。使用正向引物5'-CTTGCGGCCCAAATCTATAA-3'和反向引物5'-ACTTGTGCCCAGGAGAGCTA-3'进行PCR，这两个引物与外壳蛋白基因中的保守区域退火。反应产生了预期大小（348bp）的扩增产物。因此，据我们所知，利用ELISA和RT - PCR检测，首次在伊利诺伊州确定了南瓜中存在烟草环斑病毒。参考文献：（1）R. Provvidenti。烟草环斑病毒。载于：《葫芦科病害简编》。T. A. Zitter等人编。美国植物病理学会，明尼苏达州圣保罗。1996年。（2）S. A. Walters等人。《园艺科学》38：65，2003年。
