Chrysanthemum stunt viroid in Brazil: survey, identification, biological and molecular characterization and detection methods

[EN] In Brazil, the ornamental flowers and plants market moves in the wholesale and retail more than two billion US dollars annually, and chrysanthemum stands out as one of the most valuable commercial species. The stunting disease induced by Chrysanthemum stunt viroid (CSVd) has become a serious problem in chrysanthemum production systems worldwide. CSVd incites also colour breaking and retards flowering, but in many situations it does not induce visible symptoms, facilitating its spread in the field and passing international borders unnoticed. In Brazil there are few studies on this pathogen, with a single report of its possible occurrence in chrysanthemum in the State of Sao Paulo. In this work we aimed at surveying, identifying and characterizyng viroids in chrysanthemum crops in the State of Sao Paulo; challenging chrysanthemum varieties with a Brazilian CSVd isolate; and establishing diagnostic strategies to strengthen quarantine and indexing programs. Our survey showed that CSVd is widely disseminated in chrysanthemum crops in the State of Sao Paulo. All evaluated chrysanthemum varieties were susceptible of infection, although infection was sometimes symptomless. The complete genome sequencing of seven CSVd isolates showed high nucleotide identity when compared with other isolates from databases. Specific oligonucleotides were designed for conventional RT-PCR and RT-qPCR CSVd analysis. Results show that CSVd can be identified with a sensitivity about 100,000 times higher than PAGE. Dot-blot proved also to be highly sensitive and, consequently, most useful for large scale diagnosis. This is the first biological and molecular identification and characterization of CSVd isolates in Brazil.The authors gratefully acknowledge financial support given by Conselho Nacional de Desenvolvimento Cientifico e Tecnologico (CNPq - proc. 471796/2011-5). D. G. was recipient of student fellowship of FAPESP (proc. 2011/02721-1 and proc. 2012/20570-3); M. E. and R. H. are supported by a CNPq research fellowship. Work in J.A.D. laboratory was supported by grant BIO2011-26741 from the Ministerio de Economia y Competitividad (Spain). The authors are indebted to Mrs. Veronica Aragones (IBMCP, Valencia, Spain) by the aid provided in the sequencing of the CSVd isolates.Gobatto, D.; Chaves, A.; Harakava, R.; Marque, JM.; Daros Arnau, JA.; Eiras, M. (2014). Chrysanthemum stunt viroid in Brazil: survey, identification, biological and molecular characterization and detection methods. Journal of Plant Pathology. 96(1):111-119. https://doi.org/10.4454/JPP.V96I1.037S11111996

Ocorrência generalizada do Lettuce mottle virus em três regiões produtoras de alface comercial do Estado de São Paulo Occurrence of Lettuce mottle virus on three lettuce producing areas from São Paulo State

Os sequivírus são vírus isométricos transmitidos por afídeos. Lettuce mottle virus (LeMoV), um provável sequivirus foi descrito no Brasil em 1982 e causa sintomas de mosaico semelhantes aos observados pelo Lettuce mosaic virus (LMV). Um levantamento para ocorrência do LeMoV nos campos de produção de alface de três diferentes regiões do Estado de São Paulo (Mogi das Cruzes, Campinas e Bauru) foi realizado durante 2002 a 2005. RNA total foi extraído e utilizado na detecção, em RT-PCR, com oligonucleotídeos específicos para o LeMoV. Do total de 1362 amostras, 137 (10,05%) foram positivas para o LeMoV. Infecção mista com o LMV foi verificada em 43 amostras (31,4%). Foi verificada a ocorrência do LeMoV nas três diferentes regiões analisadas, porém sua ocorrência foi baixa nas diferentes épocas do ano.<br>Sequiviruses are isometric aphid-borne plant viruses. Lettuce mottle virus (LeMoV), a putative sequivirus was first described in Brazil on 1982 causing similar mosaic symptoms as Lettuce mosaic virus (LMV). A survey for the occurrence of LeMoV on open field conditions was carried out during 2002 to 2005 on Mogi das Cruzes, Campinas and Bauru in São Paulo state. Total RNA was extracted and used on RT-PCR with specific LeMoV primers. On 1362 samples tested, 137 (10,05%) were positive for LeMoV. Mixed infections with LMV was observed on 43 samples (31,4%). The presence of LeMoV was observed in the three different regions, but with low incidence during the year

Infecção mista pelo Sugarcane mosaic virus e Maize rayado fino virus provoca danos na cultura do milho no estado de São Paulo Mixed infection by Sugarcane mosaic virus and Maize rayado fino virus causing breaking yields in maize in São Paulo state

Os dois principais vírus que infectam o milho no Brasil são o Sugarcane mosaic virus (SCMV) e o Maize rayado fino virus (MRFV), cujos principais vetores são o afídeo Rhopalosiphum maidis e a cigarrinha Dalbulus maidis, respectivamente. O MRFV é freqüentemente encontrado em infecções mistas com fitoplasmas e espiroplasmas, causando as doenças denominadas enfezamentos do milho. Em uma lavoura de milho próxima a Santo Antonio da Posse, SP, cercada por campos de cana-de-açúcar, foi encontrada alta incidência de plantas apresentando mosaico, riscas, nanismo e espigas com falhas no enchimento de grãos. Análises serológicas com anti-soros específicos detectaram a presença do SCMV e MRFV nessas plantas. A infecção pelo SCMV também foi confirmada por RT-PCR com primers específicos e análise de seqüências. Em observações de preparações contrastadas negativamente em TEM, partículas flexuosas (ca.770 nm) e isométricas (ca.30 nm) foram detectadas. Em cortes ultrafinos, inclusões citoplasmáticas, típicas de Potyviridae, foram observadas; não foi encontrada a presença de espiroplasmas nem de fitoplasmas. Esses resultados mostram que a infecção conjunta por SCMV e MRFV pode ser responsável pelos danos encontrados nessa lavoura.<br>The two main viruses infecting maize in Brazil are Sugarcane mosaic virus (SCMV) and Maize rayado fino virus (MRFV), whose main vectors are the aphid Rhopalosiphum maidis and the leafhopper Dalbulus maidis, respectively. MRFV is frequently found in mixed infections with phytoplasms and spiroplasms causing the so-called corn stunting diseases. In a maize commercial field close to Santo Antonio da Posse, São Paulo state, surrounded by sugarcane fields, it was found a high incidence of plants showing mosaic, striping, stunting and cobs with decreased number of grains. Serological analysis with specific antisera detected the presence of SCMV and MRFV in these plants. SCMV infection was also confirmed by RT-PCR with specific primers and sequence analysis. In TEM observations of negatively stained preparations, flexuous (ca.770 nm) and isometric particles (ca.30 nm) were detected. In ultrathin sections, cytoplasmic inclusions, typical of Potyviridae, were observed; neither presence of spiroplasms nor phytoplasms could be noted. These results show that mixed infection by SCMV and MRFV might be responsible for the crop losses