RESUMO
Eggplant (Solanum melongena L.) is an economically important vegetable crop in Brazil, especially in family-based farming. Eggplant hybrids 'Ciça' and 'Napoli' (≈ 400 plants) were detected exhibiting virus-like symptoms (5-20% incidence) in field surveys (2015-2018) in Brasília-DF (Figure 1). Symptoms included chlorosis, mosaic and apical leaf deformation. Six symptomatic leaf samples were collected from fruit-bearing plants (around 100 days after planting) aiming at verifying the potential orthotospovirus infection. Double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) was carried out with polyclonal antibodies (produced at Centro Nacional de Pesquisa de Hortaliças - CNPH) against the N gene coat protein of the three major orthotospoviruses: tomato spotted wilt orthotospovirus (TSWV), groundnut ringspot orthotospovirus (GRSV) and, tomato chlorotic spot orthotospovirus (TCSV). Strong serological reactions were observed only against GRSV antibodies in the extracts from symptomatic samples, but not in the controls. To confirm the causal agent of those symptoms, total RNA was extracted from infected leaf samples via the standard Trizol® (Sigma) protocol and subsequently used in a two-step reverse transcriptase polymerase chain reaction (RT-PCR) approach. Synthesis of the cDNA was carried out with the J13 primer (5'-CCC GGA TCC AGA GCA AT-3') (Cortez et al., 2001) followed by PCR assays with the primer pair BR60 (5'-AGA GCA ATC GTG TCA-3`) and BR65 (5`-ATC AAG CCT TCT GAA AGT CAT-3') (Eiras et al., 2002). This primer set amplifies a fragment of 453 bp including the 3' untranslated region at the 3' terminus of the S RNA and the protein N-coding gene of at least five species: TSWV, GRSV, TCSV, chrysanthemum stem necrosis orthotospovirus (CSNV) and zucchini lethal chlorosis orthotospovirus (ZLCV). In addition, GRSV-specific primers (LNA Reis, unpublished) were used for amplification of all three segments: L segment: LF/LR (5'-AAC AGG ATT CAG CAA TAT GG-3'/ 5'-AAT TCC TTG AAG ACA ATT GTG T -3'); M segment: MF/MR (5'-TTT GTC CAA CCA TAC CAG ACC C- 3' / 5'-GGC TTC AAT AAA GGC TTG GG-3') and, S segment: SF/SR (5'-TTC AAA CTC AGT TGT ACT CTG A-3'/5'-TTA CTT TCG ATC TGG TTG AA- 3'). Amplicons with 509 bp (MT043204), 289 bp (MT043205), and 901 bp (MT043203) were obtained for L, M and S segments of the eggplant isolate DF-687. PCR amplicons corresponding to a segment of the N-coding gene (396 bp) of a second eggplant isolate (BJL02; MK176337) were obtained with the primer pair BR60/BR65 and subjected to Sanger dideoxy sequencing at CNPH. Alignments of nucleotide sequences of both isolates revealed identity levels varying around 99% to the corresponding genomic regions of a large set of GRSV isolates from GenBank database. PCR assays using total RNA as template yielded 494 bp amplicons solely with GRSV-specific primers (Webster et al., 2011), but no products were obtained with TSWV-specific primers (Adkins and Rosskopf, 2002), confirming the former as the sole causal agent of the field symptoms. Leaves of eggplant cv. 'Ciça' and indicator hosts, including Nicotiana rustica, Capsicum chinense 'PI 159236' (with the Tsw gene), and S. lycopersicum cv. Santa Clara were rub inoculated with extracts prepared from eggplant samples naturally infected with GRSV. Mosaic, necrotic ringspots and systemic leaf deformation symptoms were observed around ten days after inoculation on newly emerged leaves of all inoculated plants. GRSV infection was confirmed by DAS-ELISA and RT-PCR ten days after inoculation. Eggplant was erroneously listed as a host of GRSV in Brazil (Kitajima, 2020). Hence, this is the first report of eggplant infection by this virus in South America. No significant yield losses have been observed in eggplant due to GRSV infection since the overall symptoms are often mild. However, this natural host of GRSV might impact disease management strategies since eggplant is quite often cultivated under family-based farming conditions as a companion crop of highly susceptible tomato, sweet-pepper, and lettuce cultivars. References: Adkins, S., and Rosskopf, E. N. 2002. Plant Dis. 86: 1310. Cortez, I., et al. 2001. Arch. Virol. 146:265. Eiras, M. et al., 2002. Fitopatol. Bras. 27:285. Kitajima, E.W. 2020. Biota Neotrop. 20: e2019932. Webster, C. G., et al. 2011. Virology 413: 216.
RESUMO
Endive (Cichorium endivia L.) is a very important cash crop for small farmers in Brazil. During inspections conducted in the summer season of 2019-2020, leaf samples of C. endivia 'La Spezia' seedlings exhibiting typical symptoms of orthotospoviruses infection (viz. concentric chlorotic spots and apical leaf deformation; ≈ 10%) were collected in commercial greenhouses in Brasília-DF, Central Brazil. Leaves of one healthy and three symptomatic plants were initially evaluated via double antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) with polyclonal antibodies (produced at CNPH) raised against the nucleoprotein of the three major orthotospoviruses: tomato spotted wilt orthotospovirus (TSWV), groundnut ringspot orthotospovirus (GRSV) and tomato chlorotic spot orthotospovirus (TCSV). Strong serological reactions were observed only against GRSV antibodies exclusively in extracts from symptomatic samples. In order to confirm the causal agent of those symptoms, total RNA was extracted (Trizol®; Sigma) from infected leaf samples and used in a two-step reverse transcriptase polymerase chain reaction (RT-PCR) approach. Synthesis of the cDNA was carried out with the J13 primer (5'-CCC GGA TCC AGA GCA AT-3') (Cortez et al., 2001) followed by PCR assays with the primer pair BR60 (5'-AGA GCA ATC GTG TCA-3`) and BR65 (5'-ATC AAG CCT TCT GAA AGT CAT-3') (Eiras et al., 2001). This primer set amplifies a fragment of 453 bp including the untranslated region at the 3' terminus of the small RNA and the protein N-coding gene of at least five orthotospoviruses: TSWV, GRSV, TCSV, chrysanthemum stem necrosis orthotospovirus (CSNV) and zucchini lethal chlorosis orthotospovirus (ZLCV) (Eiras et al., 2001). The obtained amplicons (≈ 432 bp) were subsequently subjected to Sanger dideoxy nucleotide sequencing at CNPH. BLASTn analysis showed >99% identity with a wide array of GRSV isolates available in the GenBank. The nucleotide sequence of Tospo #1 (MT215222) and Tospo #3 (MT215224) isolates displayed 100% identity between them, whereas the Tospo #2 (MT215223) isolate displayed one non-synonymous point mutation in the 3' untranslated region in comparison with the former two isolates. Three plants of C. endivia, Capsicum annuum L. cv. Ikeda, tomato (Solanum lycopersicum L.) cv. Santa Clara and its isoline 'LAM-147' (with the Sw-5 resistance gene), Nicotiana rustica L., Lactuca sativa L. ('Vanda' and 'PI-342444') and Gomphrena globosa L. were mechanically inoculated individually with each GRSV isolate in order to confirm their pathogenicity. Chlorotic lesions and mosaic were observed seven days after inoculation of all plant materials, except the tomato inbred line 'LAM-147', which has the Sw-5 gene that confers broad-spectrum resistance to all Brazilian orthotospoviruses (Boiteux and Giordano, 1993). The GRSV infection was confirmed via DAS-ELISA and RT-PCR 15 days after inoculation, using the same set of antibodies and the primer pair BR60 / BR65. Transmission electron microscopy of ultrathin sections from symptomatic leaf tissues, both from field-infected and experimentally inoculated endive revealed the presence of typical orthotospovirus particles, within endoplasmic reticulum cisternae. Natural infection of endive by TSWV has been reported in Greece (Chatzivassiliou et al., 2000) and by TCSV in São Paulo State, Brazil and in Florida, USA (Subramanya Sastry et al., 2019). To our knowledge, it is the first report of GRSV naturally infecting this Asteraceae species in Brazil. Confirmation of GRSV infection of C. endivia plants is a relevant piece of information aiming to design effective disease management strategies. References: Boiteux, L.S. and Giordano, L. B. 1993. Euphytica 71: 151. Eiras, M. et al. 2001. Fitopatol. Bras. 26: 170. Chatzivassiliou, E.K. et al. 2000 Ann. Appl. Biol. 137: 127. Cortez, I., et al. 2001. Arch. Virol. 146: 265. Subramanya Sastry, K., et al. 2019. Encyclopedia of plant viruses and viroids. Springer, New Delhi. https://doi.org/10.1007/978-81-322-3912-3.
RESUMO
Watermelon (Citrullus lanatus) cultivated in almost all tropical and subtropical regions of the world, has its largest output in China, and then, according to FAO data, Turkey, Iran and Brazil, being one of the main crops cultivated in State of Tocantins, Brazil. In this work was investigated the occurrence and distribution of the watermelon viruses, totaling 752 samples taken in a stratified experimental design in four representative regions of production: Gurupi (150), Lagoa da Confusao (232), Formoso do Araguaia (265) and Porto Nacional (105). The sampling and collecting the leaves of plants with the presence of symptoms were performed once a week during the entire cultivation cycle. As a result, were observed by Dot-ELISA method, different types of viruses, such as Papaya ringspot W (PRSV-W), Zucchini yellow mosaic virus (ZYMV), Watermelon mosaic virus (WMV) (potyvirus), Cucumber mosaic virus ( CMV) (Cucumovirus) and Zucchini lethal chlorosis virus (ZLCV) (Tospovirus). Of these, PRSV-W was predominant (22%), followed by WMV (15%), ZLCV (11%), CMV (5%) and ZYMV (4%). Mixed infections with PRSV-W + WMV and PRSV-W + ZLCV were also observed around 20% frequency (expressed with symptoms differently from a single infection). The results provide important support for the program management viruses.
RESUMO
Efetuou-se revisäo de literatura sobre o descarte de embalagens e de resíduos de agrotóxicos. Foram discutidos os aspectos legais, o impacto ambiental causado pelo uso intensivo de agrotóxicos, os tipos de embalagens e os métodos (biológicos, físicos e químicos) utilizados para o seu descarte, além daqueles para recuperaçäo/reciclagem. Concluiu-se que é necessário o estabelecimento de açäo conjunta entre instituiçöes governamentais, empresas produtoras e usuários destes produtos, visando a conscientizaçäo quanto ao uso adequado e seguro de agrotóxicos, além da destinaçäo final correta de embalagens vazias e de resíduos
