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番茄斑驳双生病毒侵染墨西哥尤卡坦半岛番茄的首次报道

First Report of Tomato Mottle Geminivirus Infecting Tomatoes in Yucatan, Mexico.

作者信息

Garrido-Ramirez E R, Gilbertson R L

机构信息

Department of Plant Pathology, University of California, Davis 95616.

出版信息

Plant Dis. 1998 May;82(5):592. doi: 10.1094/PDIS.1998.82.5.592B.

Abstract

Whitefly-transmitted geminiviruses are a major constraint on tomato production in Mexico (3). In the Yucatan State, these viruses can cause serious losses in late season plantings. As part of an effort to characterize these viruses, leaf samples from four tomato plants showing symptoms of geminivirus infection, such as stunted growth and leaf mottling and deformation, were collected from a single field in the Yucatan State in February, 1996. Geminivirus nucleic acids were detected in leaf samples from all four plants by squash blot hybridization analysis with a general DNA probe for Western Hemisphere whitefly-transmitted geminiviruses (2). Nicotiana benthamiana plants inoculated with sap prepared with leaf tissue from one plant developed stunted growth and leaf mottling and deformation. When graft-transmitted from N. benthamiana to tomato, the geminivirus(es) induced leaf mottling and deformation, which were similar to symptoms in the field-collected tomato plants. The presence of geminivirus DNA in the sap- and graft-inoculated plants was confirmed with the polymerase chain reaction (PCR) and degenerate primers for the DNA-A (PAL1v1978 and PAR1c496) or DNA-B (PBL1v2040 and PCRc1) components of whitefly-transmitted geminiviruses (4). Using PCR and these degenerate primers, approximately 1.1-kb DNA-A and approximately 0.6-kb DNA-B fragments were amplified from DNA extracts prepared from leaves of each of the four Yucatan tomato plants. No DNA fragments were amplified from these extracts with primers for pepper huasteco geminivirus (pAL1c2329 and pAL1v1471, or pBR1c840 and pBL1v1830). To determine the identity of the geminivirus(es) infecting these tomato plants, the PCR-amplified DNA-A and DNA-B fragments from one of the samples were cloned and sequenced. Comparisons made with these sequences revealed two distinct types of DNA-A and DNA-B clones, indicating a mixed infection of at least two bipartite geminiviruses. DNA-A and DNA-B sequences of one set of clones were >97% identical to sequences of tomato mottle geminivirus (ToMoV) from Florida (1). The presence of ToMoV in all four tomato leaf samples was demonstrated by the PCR-mediated amplification of a 0.9-kb DNA-A fragment with ToMoV-specific primers (pAL1v2295 and pAR1c580). The identity of this 0.9-kb DNA fragment was further confirmed based upon its hybridization with a full-length clone of ToMoV DNA-A under high stringency conditions (2). A data base search made with the sequence of the other type of DNA-A clone revealed sequence identities of <70% with various bipartite geminiviruses (e.g., identities of 70% with tomato mottle, 69% with Sida golden mosaic, 67% with bean dwarf mosaic, and 66% with taino tomato mottle and with potato yellow mosaic), which confirmed that a second geminivirus was present in a mixed infection with ToMoV in this tomato leaf sample. To confirm the bipartite nature of this geminivirus, a DNA-B fragment that contained the common region (CR) sequence was amplified from the same sample with PCR and primers PBL1v2040 and PBR1c970 (a degenerate primer that anneals within the BV1 open reading frame; F. M. Zerbini and R. L. Gil-bertson, unpublished data), cloned, and sequenced. The CR sequence of this DNA-B fragment was 96% identical to that of the DNA-A fragment, which establishes the presence of another bipartite geminivirus in this sample. This is the first report of ToMoV in Mexico. These results also suggest that at least two bipartite geminiviruses may infect tomatoes in the Yucatan Peninsula. References: (1) A. M. Abouzid et al. J. Gen. Virol. 73:3225, 1992. (2) R. L. Gilbertson et al. Plant Dis. 75:336, 1991. (3) J. E. Polston and P. K. Anderson. Plant Dis. 81:1358, 1997. (4) M. R. Rojas et al. Plant Dis. 77:340, 1993.

摘要

粉虱传播的双生病毒是墨西哥番茄生产的主要限制因素(3)。在尤卡坦州,这些病毒会导致晚季种植严重减产。作为对这些病毒进行特性描述工作的一部分,1996年2月从尤卡坦州的一块田地中采集了4株表现出双生病毒感染症状(如生长受阻、叶片斑驳和变形)的番茄植株的叶片样本。通过使用针对西半球粉虱传播双生病毒的通用DNA探针进行压片杂交分析,在所有4株植株的叶片样本中均检测到双生病毒核酸(2)。用其中一株植株的叶片组织制备的汁液接种本氏烟植株后,植株出现生长受阻、叶片斑驳和变形。当从本氏烟嫁接到番茄上时,双生病毒诱导叶片出现斑驳和变形,这与田间采集的番茄植株症状相似。通过聚合酶链反应(PCR)以及用于粉虱传播双生病毒DNA-A(PAL1v1978和PAR1c496)或DNA-B(PBL1v2040和PCRc1)组分的简并引物,证实了汁液接种和嫁接接种植株中存在双生病毒DNA(4)。使用PCR和这些简并引物,从4株尤卡坦番茄植株的叶片制备的DNA提取物中扩增出了约1.1 kb的DNA-A片段和约0.6 kb的DNA-B片段。用针对辣椒瓦斯泰科双生病毒的引物(pAL1c2329和pAL1v1471,或pBR1c840和pBL1v1830)从这些提取物中未扩增出DNA片段。为确定感染这些番茄植株的双生病毒的身份,将其中一个样本经PCR扩增的DNA-A和DNA-B片段进行克隆并测序。与这些序列的比较揭示了两种不同类型的DNA-A和DNA-B克隆,表明至少两种双分体双生病毒混合感染。一组克隆的DNA-A和DNA-B序列与来自佛罗里达的番茄斑驳双生病毒(ToMoV)的序列一致性>97%(1)。通过用ToMoV特异性引物(pAL1v2295和pAR1c580)进行PCR介导的0.9 kb DNA-A片段扩增,证实了所有4个番茄叶片样本中均存在ToMoV。基于该0.9 kb DNA片段在高严格条件下与ToMoV DNA-A全长克隆的杂交,进一步确认了其身份(2)。用另一种类型的DNA-A克隆序列进行数据库搜索,发现与各种双分体双生病毒的序列一致性<70%(例如,与番茄斑驳病毒一致性为70%,与 sida 金色花叶病毒一致性为69%) ,与菜豆矮化花叶病毒一致性为67%,与泰诺番茄斑驳病毒和马铃薯黄花叶病毒一致性为66%),这证实了该番茄叶片样本中存在第二种双生病毒与ToMoV混合感染。为确认这种双生病毒的双分体性质,用PCR和引物PBL1v2040和PBR1c970(一种在BV1开放阅读框内退火的简并引物;F.M.泽尔比尼和R.L.吉尔伯森,未发表数据)从同一样本中扩增出包含共同区域(CR)序列的DNA-B片段,进行克隆并测序。该DNA-B片段的CR序列与DNA-A片段的CR序列一致性为96%,这证实了该样本中存在另一种双分体双生病毒。这是墨西哥首次报道ToMoV。这些结果还表明,至少两种双分体双生病毒可能感染尤卡坦半岛的番茄。参考文献:(1)A.M.阿布齐德等人,《普通病毒学杂志》73:3225,1992年。(2)R.L.吉尔伯森等人,《植物病害》75:336,1991年。(3)J.E.波尔斯顿和P.K.安德森,《植物病害》81:1358,1997年。(4)M.R.罗哈斯等人,《植物病害》77:340,1993年。

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