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Desde inicios del decenio de 1990 y de manera casi simultánea, se empezaron a observar afecciones desconocidas en numerosos cultivos, sobre todo en regiones tropicales y subtropicales. Dichas enfermedades eran causadas por begomovirus hasta entonces poco estudiados. Su vector es la mosca blanca Bemisia tabaci (Hemiptera: Aleyrodidae), de la que no se habían registrado poblaciones tan desmedidas en áreas agrícolas. Ello originó una seria crisis de producción a nivel mundial, con pérdidas millonarias para los agricultores de numerosos países, incluyendo la región de Mesoamérica. Por fortuna, en Costa Rica, algunos centros de investigación, con la colaboración de especialistas extranjeros, han podido profundizar en las causas de este fenómeno, para buscar soluciones fundamentadas en información de carácter epidemiológico. Además de la especie de B. tabaci New World (NW), nativa y previamente conocida, en años recientes han ingresado sendas especies del Mediterráneo (MED) y el Medio Oriente-Asia Menor 1 (MEAM1). Asimismo, se ha detectado la presencia de begomovirus bipartitas, nativos y exóticos, especialmente en frijol común, cucurbitáceas, tomate y chile; además, se detectó el begomovirus monopartito conocido a nivel mundial Tomato yellow leaf curl virus (TYLCV) en tomate. Con base en el conocimiento biológico y ecológico acumulado hasta ahora, en la presente revisión se aporta una panorámica del dinamismo con que se han expresado en Costa Rica las interacciones entre las diferentes especies vectoras y los distintos begomovirus, con énfasis en pato-sistemas de hortalizas. Se espera que la información aquí presentada permita mejorar el tipo de respuestas prácticas y eficaces de parte de agricultores, extensionistas agrícolas e investigadores involucrados en la producción de hortalizas, frente a problemas actuales o imprevistos. Since the early 1990s, and almost simultaneously, unknown diseases started to be observed in many crops, especially in tropical and subtropical regions. These diseases were predominantly caused by begomoviruses, which were poorly known at that time. Their vector, the whitefly Bemisia tabaci (Hemiptera: Aleyrodidae), often reached unprecedented huge populations in agricultural areas. This elicited a serious production crisis worldwide, that caused losses of millions of dollars for farmers in many countries, including the Mesoamerican region. Fortunately, in Costa Rica, some local research centers, with the collaboration of foreign specialists, have been able to study the causes of this phenomenon, in the search for solutions based on solid epidemiological information. In addition to the previously reported native Bemisia tabaci species, New World (NW), two exotic species, - Mediterranean (MED) and Middle East-Asia Minor 1 (MEAM1) - were found. Moreover, native and exotic bipartite begomoviruses have been detected, especially in common bean, cucurbits, tomato and sweet pepper, as well as the worldwide spread monopartite begomovirus Tomato yellow leaf curl virus (TYLCV). Based upon biological and ecological knowledge accumulated to date, this review offers a comprehensive overview of the very dynamic ways in which the interactions of the different whiteflies and begomovirus species have expressed in Costa Rica, with emphasis on vegetable pathosystems. Hopefully, the information provided in this paper may allow farmers, extension agents, and researchers involved in vegetable production to develop sound practical responses to current and unforeseen problems regarding whiteflies and their associated viruses. Universidad de Costa Rica/[]/UCR/Costa Rica Ministerio de Economía y Competitivi-dad/[ AGL2016-75819-C2-2]//España European Regional Development Fund/[AGL2016-75819-C2-2]/ERDF/Unión Europea European Social Fund/[AGL2016-75819-C2-2]/ESF/Unión Europea National Council for Scientific and Technological Development/[]/CNPq/Brasil UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Agroalimentarias::Centro Nacional de Ciencia y Tecnología de Alimentos (CITA) UCR::Vicerrectoría de Docencia::Ciencias Agroalimentarias::Facultad de Ciencias Agroalimentarias::Escuela de Tecnología de Alimentos UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Básicas::Centro de Investigación en Biología Celular y Molecular (CIBCM)

Begomoviruses (genus Begomovirus, family Geminiviridae) have emerged as important plant pathogens in tropical and subtropical regions worldwide. Although these viruses were reported during the 1970s in Costa Rica, they are still poorly known. Therefore, the objective of this study was to analyse the diversity and distribution of begomoviruses in commercial tomato and sweet pepper fields from different agricultural production systems of the major growing regions of Costa Rica. A total of 651 plants were randomly sampled from greenhouses and open field crops during 2011 and 2012 in three different geographical locations. The bipartite begomoviruses Tomato yellow mottle virus, Tomato leaf curl Sinaloa virus and Pepper golden mosaic virus, and the monopartite begomovirus Tomato yellow leaf curl virus were detected in the collected samples. The complete genome of isolates from each species was cloned and sequenced. The frequency of detection of these four begomoviruses in the analysed samples ranged from 0 to 9%, the presence, and the prevalent virus varied largely according to the geographical location, the host (tomato and pepper), and the production system (greenhouses or open fields). An association between geographical region and begomovirus species was observed suggesting that in Costa Rica the heterogeneity on climate, topography and agricultural system might influence the distribution of begomovirus species in the country. A broader survey needs to be conducted to confirm it, although these preliminary results may contribute to the management of begomoviruses in Costa Rica. Ciencia y Tecnología para el Desarrollo/[11RT0433]/CYTED/Costa Rica Universidad de Costa Rica/[801-B1-650]/UCR/Costa Rica UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Básicas::Centro de Investigación en Biología Celular y Molecular (CIBCM) UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Agroalimentarias::Centro de Investigación en Protección de Cultivos (CIPROC)

Although the Sw‐5 gene cluster has been cloned, and Sw‐5b has been identified as the functional gene copy that confers resistance to Tomato spotted wilt virus (TSWV), its avirulence (Avr) determinant has not been identified to date. Nicotiana tabacum ‘SR1‘ plants transformed with a copy of the Sw‐5b gene are immune without producing a clear visual response on challenge with TSWV, whereas it is shown here that N. benthamiana transformed with Sw‐5b gives a rapid and conspicuous hypersensitive response (HR). Using these plants, from all structural and non‐structural TSWV proteins tested, the TSWV cell‐to‐cell movement protein (NS(M)) was confirmed as the Avr determinant using a Potato virus X (PVX) replicon or a non‐replicative pEAQ‐HT expression vector system. HR was induced in Sw‐5b‐transgenic N. benthamiana as well as in resistant near‐isogenic tomato lines after agroinfiltration with a functional cell‐to‐cell movement protein (NS(M)) from a resistance‐inducing (RI) TSWV strain (BR‐01), but not with NS(M) from a Sw‐5 resistance‐breaking (RB) strain (GRAU). This is the first biological demonstration that Sw‐5‐mediated resistance is triggered by the TSWV NS(M) cell‐to‐cell movement protein.

The begomoviruses (fam. Geminiviridae) have an ssDNA genome, are transmitted by whiteflies, and cause significant losses in tomato fields in Brazil. Nowadays, begomovirus species identification is carried out through the analysis of their complete genome sequence. Due to the high costs and difficulty in applying this technique to large-scale analysis, we aimed to develop a species-specific detection method based on PCR. Three species were targeted for amplification: Tomato severe rugose virus (ToSRV), Tomato rugose mosaic virus (ToRMV) and Tomato yellow vein streak virus (ToYVSV). Thirteen primer combinations were designed, and four were finally selected and tested against a group of 82 samples, including cloned DNA and viral DNA from infected plants previously identified by direct sequencing of PCR products. Three primer combinations were selected that could distinguish the three species, and confirmed by sequencing of amplified products. These combinations were validated by evaluation of field-collected tomato samples infected by begomovirus and this demonstrated that PCR can be a useful tool to distinguish the viruses and detect mixed infections caused by three begomovirus species. Os begomovírus (fam. Geminiviridae) possuem um genoma de ssDNA, são transmitidos por mosca-branca, e causam danos importantes em campos de produção de tomate no Brasil. Atualmente, a identificação de espécies de begomovírus é realizada pela análise da seqüência completa do genoma. Devido ao alto custo e dificuldade em se aplicar essa técnica para análises em larga escala, o objetivo desse trabalho foi desenvolver um método de detecção espécie-específico baseado em PCR. Três espécies foram avaliadas para amplificação: Tomato severe rugose virus (ToSRV), Tomato rugose mosaic virus (ToRMV) e Tomato yellow vein streak virus (ToYVSV). Foram testadas treze combinações de primers, e quatro foram selecionadas e avaliadas em um grupo de 82 amostras, incluindo DNA clonado e DNA viral total extraído de plantas infectadas com begomovírus previamente identificados por seqüenciamento direto de produto de PCR. Três combinações de primers capazes de distinguir as três espécies foram selecionadas, e confirmadas por seqüenciamento dos produtos amplificados. Estas combinações foram validadas com amostras de tomate infectadas por begomovírus em campo, demonstrando que a PCR pode ser uma ferramenta útil para separar as espécies virais e detectar infecções mistas causadas pelas três espécies de begomovírus.

O aumento na ocorrência de begomoviroses (geminiviroses) em tomateiros, Lycopersicon esculentum Mill., em várias regiões brasileiras, vem causando grandes prejuízos para o agronegócio de tomate, devido à ocorrência do inseto vetor, Bemisia argentifolii (Bemisia tabaci biotipo B). A diagnose é realizada em geral por "polymerase chain reaction" (PCR) ou por hibridização com sondas radioativas. A PCR é um método de alta sensibilidade, porém apresenta a desvantagem da possibilidade de obtenção de resultados falso-positivos, devido a contaminações, ou falso-negativos causados por inibidores contaminantes da reação, ou pela extrema especificidade dos iniciadores. A hibridização de ácidos nucléicos com sondas radioativas tem o seu uso limitado devido à necessidade de infra-estrutura especial, treinamento de pessoal, riscos para a saúde do manipulador e demanda constante de radioquímicos. O presente trabalho tem por finalidade demonstrar a viabilidade do uso do método de hibridização com sondas não-radioativas para a detecção de um begomovírus de tomateiro do Brasil. A sensibilidade do teste foi alta, obtendo-se detecção de até 0,1fg do DNA homólogo e em extrato bruto foliar diluído até 100 vezes. Major outbreaks of tomato begomoviruses (geminiviruses) in several tomato Lycopersicon esculentum Mill growing in many parts of Brazil have been imposing significant losses upon the tomato agribusiness, due to the introduction of the Bemisia argentifolii (Bemisia tabaci biotype B). Polymerase chain reaction (PCR) and hybridization are generally used for diagnosis. The PCR is a detection method with high sensitivity, however it has the disadvantage of producing false-positives, due to contamination, or false-negatives caused by inhibitors or because of the high primer specifity. The use of nucleic acid hybridization with radio-labelled probes is restricted due to the requirement of special infrastructure and handling experience, the risk for the user’s health and a regular radiochemical element supply. This study is aimed at demonstrating the usefulness of the hybridization method with non-radioactive probes for detection of a tomato begomoviruses in Brazil. The sensitivity of this method was high enabling the detection of 0.1fg of homologous DNA and in crude sap extract diluted up to 100-fold.