The complete nucleotide sequence of the genomic RNA 1 (8745 nt) and RNA 2 (4986 nt) of Citrus leprosis virus cytoplasmic type (CiLV-C) was determined using cloned cDNA. RNA 1 contains two open reading frames (ORFs), which correspond to 286 and 29 kDa proteins. The 286 kDa protein is a polyprotein putatively involved in virus replication, which contains four conserved domains: methyltransferase, protease, helicase and polymerase. RNA 2 contains four ORFs corresponding to 15, 61, 32 and 24 kDa proteins, respectively. The 32 kDa protein is apparently involved in cell-to-cell movement of the virus, but none of the other putative proteins exhibit any conserved domain. The 59 regions of the two genomic RNAs contain a 'cap' structure and poly(A) tails were identified in the 39-terminals. Sequence analyses and searches for structural and non-structural protein similarities revealed conserved domains with members of the genera Furovirus, Bromovirus, Tobravirus and Tobamovirus, although phylogenetic analyses strongly suggest that CiLV-C is a member of a distinct, novel virus genus and family, and definitely demonstrate that it does not belong to the family Rhabdoviridae, as previously proposed. Based on these results it was proposed that Citrus leprosis virus be considered as the type member of a new genus of viruses, Cilevirus.
Solanum violaefolium is an ornamental plant, with prostrate, trailing growth habit and is cultivated in shaded areas. A virus that causes ringspot symptoms on its leaves, tentatively named as Solanum violaefolium ringspot virus (SvRSV) and transmitted by Brevipalpus phoenicis (Acari: Tenuipalpidae) was found in Piracicaba city, São Paulo State. It is a bacilliform virus that resembles other cytoplasmic viruses transmitted by Brevipalpus sp. The objective of this work is to describe the biological properties and establish partial molecular characterization of the SvRSV. The SvRSV can be transmitted mechanically to several plant species causing local lesions. Among the tested species, Datura stramonium was proved to be the best experimental host. It was observed that S. violaefolium plants were infested by B. obovatus that also transmitted SvRSV in preliminary Ferreira, P.T.O., Locali-Fabris, E.C., Freitas-Astúa, J., Antonioli-Luizon, R., Gomes, R.T., Machado, M.A., Kitajima, E.W. Characterization of a bacilliform virus isolated from Solanum violaefolium transmitted by the tenuipalpid mites Brevipalpus phoenicis and Brevipalpus obovatus. Summa Phytopathologica, v.33, n.3, p.264-269, 2007 RESUMO Solano-violeta (Solanum violaefolium) é uma planta ornamental rasteira usada para cobrir solos de áreas sombreadas. Um vírus que induz manchas anelares nas folhas desta planta, tentativamente designado Solanum violaefolium ringspot virus -SvRSV, transmitido pelo ácaro Brevipalpus phoenicis (Acari: Tenuipalpidae) foi encontrado em Piracicaba, SP. Trata-se de um vírus baciliforme que se assemelha a outros vírus do tipo citoplasmático transmitidos por Brevipalpus sp. Este trabalho teve como objetivo relatar propriedades biológicas e estabelecer uma caracterização molecular parcial do SvRSV. O vírus pode ser transmitido mecanicamente a várias outras espécies botânicas, causando lesões localizadas. Entre as espécies avaliadas, Datura stramonium mostrou-se a melhor hospedeira experimental. Observou-se também a manifestação de sintomas nestas plantas após infestação das mesmas por B. obovatus previamente alimentado em Ferreira, P.T.O., Locali-Fabris, E.C., Freitas-Astúa, J., Antonioli-Luizon, R., Gomes, R.T., Machado, M.A., Kitajima, E.W. Caracterização de um vírus baciliforme isolado de Solanum violaefolium transmitido pelos ácaros Brevipalpus phoenicis e Brevipalpus obovatus (Acari: Tenuipalpidae). Summa Phytopathologica, v.33, n.3, p. [264][265][266][267][268][269] 2007 Palavras-chave adicionais: SvRSV, CiLV-C, dsRNA, sequenciamento, efeito citopático lesões de SvRSV, confirmando esta outra espécie de ácaro como vetor do vírus. Suas propriedades físicas in vitro foram: temperatura de inativação 40-45 ºC; ponto final de diluição 10 -3 -10 -4 ; longevidade in vitro 12 dias. Em secções ultrafinas, as partículas do SvRSV mostraramse levemente mais delgadas e mais longas que as de outros vírus do mesmo grupo. A partir do dsRNA do SvRSV foi construída uma biblioteca de cDNA e foram identificadas duas possíveis regiões c...
The diagnosis of plant diseases caused by Brevipalpus-transmitted viruses (BrTVs) has been done through the analyses of symptoms, transmission electron microscopy, and RT-PCR of infected plant tissues. Here, we report the detection of Citrus leprosis virus C, Orchid fleck virus, Clerodendrum chlorotic spot virus and Solanum violaefolium ringspot virus in their viruliferous vectors Brevipalpus spp. using specific primer pairs for each of the viruses. The efficiency of virus transmission by Brevipalpus mites is low, so the detection of these pathogens in their vectors could constitute an important tool for studies involving virus-vector relationships, transmission, and monitoring the pathogen prior to the appearance of symptoms in the field.
The variability of a fragment of the nucleocapsid gene of orchid fleck virus (OFV) was investigated by single-strand conformational polymorphism (SSCP) analysis and nucleotide sequencing. Forty-eight samples of 18 genera of orchids were collected from Brazil, Costa Rica and Australia. The SSCP analysis yielded six different band patterns, and phylogenetic analysis based on the nucleotide fragment sequence obtained in this work and six available in GenBank showed two different groups, one with isolates 023Germany and So-Japan, and other with the rest of the isolates. None of the analyses showed geographic correlation among the Brazilian strains. The data obtained in this study showed a low genetic variation in this region of the genome; the d(N)/d(S) ratio of 0.251-0.405 demonstrated a negative selective pressure that maintains the stability of the analyzed fragments.
In Colombia, citrus is cultivated in mostly small plantings that total 55,000 ha by approximately 25,000 farmers. Production includes 1,200 tons of fresh fruits and 60 tons of juice for domestic consumption, resulting in a net worth of US$650,000 per year. Most of the production comes from areas located between the Cordillera Occidental and Cordillera Central mountain ranges (departments of Antioquia, Caldas, Quindio, and Risaralda) near coffee plantations. The departments of Meta and Casanare, located at the east plains (Llanos Orientales), include a zone parallel (4 to 5°N, 72 to 74°W) to the east mountain range and generate approximately 10% of the total Colombian citrus production. Suspected citrus leprosis symptoms on leaves and fruits of sweet oranges (Citrus sinensis (L.) Osb.) were first observed by plant pathologists for CORPOICA (Colombian National Agricultural Research Organization) in citrus orchards in Casanare in 2003, and later in 2004, in Meta. To confirm the visual identification, leaves and fruits from Valencia sweet orange exhibiting typical lesions of leprosis were collected from several locations in the departments of Casanare (Yopal, Aguazul) and Meta (Guamal, Villavicencio, and Cumaral). Samples were fixed in cacodylate-buffered paraformaldehyde/glutaraldehyde solution and subsequently processed for examination in thin sections using electron microscopy. Samples were processed and examined at the Citrus Research and Educational Center (CREC) of the University of Florida, Lake Alfred, and the Agricultural College (ESALQ) of the Universidade de São Paulo at Piracicaba, SP, Brazil. Some leaf samples collected in Meta were also dried and used for detection of Citrus leprosis virus, cytoplasmic type (CiLV-C) by reverse transcription-polymerase chain reaction (RT-PCR) at the Centro APTA Citros Sylvio Moreira at Cordeirópolis (CAPTACSM). The RT-PCR was performed with primers that specifically amplify a fragment of the viral genome that codes for the putative cell-to-cell movement protein (1). Locations at CREC and ESALQ each observed, using electron microscopy, cell changes characteristic of CiLV-C that include short bacilliform particles in the endoplasmic reticulum and dense, vacuolated, and irregularly shaped viroplasm in the cytoplasm (2) in samples from Casanare and Meta. RT-PCR amplified cDNA fragments of the expected size for samples collected in Meta and one of the amplicons was sequenced (GenBank Accession No. DQ272491). The sequence obtained was found to have 98% nucleotide sequence identity to the Brazilian CiLV-C isolate (GenBank Accession No. AY289190.1). Mites collected from affected plants from the department of Meta were identified at ESALQ as Brevipalpus phoenicis (Geijskes), a known principal vector of CiLV-C (2). These several lines of evidence confirmed that the symptoms observed in sweet oranges at Meta and Casanare are due to the infection by CiLV-C. To our knowledge, this is the first report of this virus in Colombia. References:(1) E. C. Locali et al. Plant Dis. 87:1317, 2003, (2) J. C. V. Rodrigues et al. Exp. Appl. Acarol. 30:161, 2003.
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