Torrico A K, Cafrune E E, Conci V C
National Council of Scientific and Technical Researches (CONICET), Cordoba, Argentina.
Institute of Phytopathology and Vegetal Physiology of National Institute of Agricultural Technology (IFFIVE-INTA), Cordoba, Argentina.
Plant Dis. 2010 Jul;94(7):915. doi: 10.1094/PDIS-94-7-0915B.
Because of exclusively agamic propagation, garlic is commonly infected with a virus complex mainly composed of species within the genera Potyvirus, Allexivirus, and Carlavirus. This virus complex causes leaf striping that ranges from various shades of green to yellow and results in yield losses (2,4). Onion yellow dwarf virus, Leek yellow stripe virus (potyviruses), Garlic virus A, Garlic virus C (allexiviruses), and Garlic common latent virus (carlavirus) have been detected in Argentina previously (1,2). Recently, Shallot latent virus (SLV; another carlavirus) was detected in 25 of 30 garlic plants (cv. Morado) growing in four different fields near Córdoba, Argentina by double-antibody sandwich (DAS)-ELISA using BIOREBA (Reinach, Switzerland) antibodies. To confirm the presence of the virus, DAS-ELISA-positive plants were also analyzed by one-step reverse transcription (RT)-PCR using the Access RT-PCR system (Promega, Madison, WI) with specific primers reported by Tsuneyoshi et al. (3). RNA extractions were performed from 100 mg of leaves with the Qiagen RNeasy Plant Mini Kit (Qiagen, Valencia, CA). Primers used were Car-V1 (5'-AAACCTTTTGGTTCACTTTAGG-3'); Car-V2 (5'-AGGTGCATTGTTATCATTACTGG-3'); and Car-Cp3 (5'-GCGTGCTATATTTAAGTTGCATAC-3'). Primer pairs Car-V1/Car-Cp3 and Car-V2/Car-Cp3 were used for the amplification of the coat protein (CP) gene of SLV and an isolate of SLV formerly known as Garlic latent virus, respectively. Fragments of 992 bp and 1,079 bp were amplified with these primer pairs, respectively. The RT-PCR products were cloned with the TOPO TA Cloning Kit in the 3.9-kb pCR-TOPO vector (Qiagen). The nucleotide sequences of both fragments were determined and were found to be identical (GenBank No. GU355922) showing 94.2% nt sequence identity with the CP gene of an isolate of SLV from Indonesian garlic (GenBank No. AB004686) formerly referred to as Garlic latent virus (3). Consequently, the Argentinean virus is now considered a garlic isolate of SLV. References: (1) E. Cafrune et al. Plant Dis. 90:898, 2006. (2) V. C. Conci. Virus y Fitoplasmas de Ajo. Page 267 in: 50 Temas Sobre Producción de Ajo. Vol. 3. J. L. Burba, ed. Ediciones INTA, Mendoza, Argentina. 1997. (3) T. Tsuneyoshi et al. Arch. Virol. 143:1093, 1998. (4) D. G. A. Walkey and D. N. Antill. J. Hortic. Sci. 64:53, 1989.
由于大蒜仅通过无性繁殖,它通常感染一种病毒复合体,该复合体主要由马铃薯Y病毒属、葱X病毒属和香石竹潜隐病毒属的病毒种类组成。这种病毒复合体导致叶片出现条斑,颜色从各种深浅不一的绿色到黄色,从而造成产量损失(2,4)。此前在阿根廷已检测到洋葱黄矮病毒、韭葱黄条病毒(马铃薯Y病毒属)、大蒜病毒A、大蒜病毒C(葱X病毒属)和大蒜普通潜隐病毒(香石竹潜隐病毒属)(1,2)。最近,通过使用瑞士雷纳赫市BIOREBA公司的抗体进行双抗体夹心(DAS)-ELISA检测,在阿根廷科尔多瓦附近四个不同田块种植的30株大蒜植株(品种为Morado)中,有25株检测到了青葱潜隐病毒(SLV;另一种香石竹潜隐病毒属病毒)。为了确认该病毒的存在,还使用普洛麦格公司(美国威斯康星州麦迪逊)的Access RT-PCR系统和常吉等(3)报道的特异性引物,通过一步法逆转录(RT)-PCR对DAS-ELISA检测呈阳性的植株进行了分析。使用Qiagen公司(美国加利福尼亚州瓦伦西亚)的RNeasy植物小提试剂盒从100毫克叶片中提取RNA。所用引物为Car-V1(5'-AAACCTTTTGGTTCACTTTAGG-3');Car-V2(5'-AGGTGCATTGTTATCATTACTGG-3');以及Car-Cp3(5'-GCGTGCTATATTTAAGTTGCATAC-3')。引物对Car-V1/Car-Cp3和Car-V2/Car-Cp3分别用于扩增SLV的外壳蛋白(CP)基因以及一种曾被称为大蒜潜隐病毒的SLV分离株。用这两对引物分别扩增出了992 bp和1,079 bp的片段。RT-PCR产物用TOPO TA克隆试剂盒克隆到3.9 kb的pCR-TOPO载体(Qiagen公司)中。测定了这两个片段的核苷酸序列,发现它们是相同的(GenBank登录号:GU355922),与来自印度尼西亚大蒜的一种曾被称为大蒜潜隐病毒的SLV分离株的CP基因(GenBank登录号:AB004686)的核苷酸序列同一性为94.2%(3)。因此,现在认为这种阿根廷病毒是SLV的一个大蒜分离株。参考文献:(1)E. Cafrune等人,《植物病害》90:898,2006年。(2)V. C. Conci,《大蒜病毒与植原体》。载于《大蒜生产的50个主题》第3卷。J. L. Burba主编。阿根廷门多萨INTA出版社,1997年,第267页。(3)T.常吉等人,《病毒学档案》143:1093,1998年。(4)D. G. A. Walkey和D. N. Antill,《园艺学杂志》64:53,1989年。