Several species of Xanthomonas cause bacterial leaf spot, a disease that affects solanaceous crops worldwide. The diversity of 64 Australian isolates of Xanthomonas spp. associated with bacterial leaf spot in tomato, capsicum and chilli crops in eastern Australia was determined using multi-locus sequence analysis of atpD, dnaK, efp and gyrB genes, species-specific PCR assays and biochemical analyses. At least five species of Xanthomonas associated with bacterial leaf spot were identified in Australian tomato, capsicum and chilli crops and their pathogenicity assessed. Phylogenetic and biochemical analyses identified X. euvesicatoria, X. perforans and X. vesicatoria as the most frequently recovered pathogenic species. Non-pathogenic and weakly pathogenic species were also identified. The suitability of the identification methods used and the implications of the detection of these species will be discussed.

Tomato bacterial spot is caused by Xanthomonas euvesicatoria, X. vesicatoria, X. perforans and X. gardneri. In order to determine the distribution, frequency of occurrence, and diversity of these species in the Brazilian commercial tomato fields, a survey was conducted between 2009 and 2012. In this period, 204 strains were obtained from 33 counties (22 with processing tomatoes and 11 with fresh-market tomatoes). Pathogenicity tests, BOX-PCR, PCR with species-specific primers, and sequence analysis of the avirulence gene avrXv3 were performed in order to identify the strains at species and race level. Xanthomonas perforans predominated among the strains (92%) and was present in most counties. In addition, this species was prevalent in most areas of both fresh-market tomatoes (63.6% of counties surveyed) and processing tomatoes (95.4% of counties surveyed). Fifteen strains (7.5%) were identified as X. gardneri, which was found mostly in fresh-market fields located at regions with altitude higher than 900 m, and only one strain of X. euvesicatoria (0.5%) was found in a processing tomato field. High genetic diversity was observed within X. perforans, with 137 BOX-PCR haplotypes. Race T3 prevailed (97.5%), but reported here for the first time is the occurrence of five strains identified as race T4 in fresh-market fields in the state of São Paulo. The race T4 phenotype of these strains resulted from the presence of an 859 bp insertion in the avirulence gene avrXv3. This insertion is related to amino acid sequences of a transposase found in X. gardneri, and to amino acid sequences of X. campestris.

Aims: To establish protocols for the simultaneous detection and identification of Xanthomonas species causing tomato bacterial spot. Methods and Results: We verified the specificity and sensitivity of the previously reported sets of primers designed for strains of the four species of Brazilian tomato bacterial spot xanthomonads, consisting of 30 of Xanthomonas euvesicatoria, 30 of X. vesicatoria, 50 of X. perforans and 50 of X. gardneri. Furthermore, we tested a multiplex PCR protocol for the purpose of concurrent species identification. The possibility of direct detection of the pathogens in diseased leaf samples was also verified. The primers were highly specific, amplifying only target DNA. The sensitivity of the primers in conventional PCR was 50 pg ll À1 for purified DNA and ranged from 5 9 10 2 to 5 9 10 4 CFU ml À1 when bacterial suspensions were analysed. The multiplex PCR was suitable for the detection of all four species and showed similar sensitivity to conventional PCR when tested on purified DNA. When using bacterial suspensions, its sensitivity was similar to conventional PCR only when a biological amplification step (Bio-PCR) was included. Both methods were able to detect the pathogens in symptomatic tomato leaves. Conclusions: Brazilian Xanthomonas strains causing tomato bacterial spot can be differentiated and identified at species level by a PCR-based method and by a multiplex PCR. Significance and Impact of the Study: This protocol may be a feasible alternative tool for the identification and detection of these pathogens in plant material and may be used for routine diagnostic purposes in plant pathology laboratories.

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RESUMOO objetivo deste trabalho foi identificar isolados de Xanthomonas causadores da mancha bacteriana do tomateiro para consumo in natura, provenientes da região do Alto Vale do Rio do Peixe, SC, Brasil, bem como caracterizá-los quanto à sensibilidade in vitro ao cobre. As espécies foram determinadas por similaridades de perfis genômicos gerados por BOX-PCR e a sensibilidade ao cobre foi estabelecida utilizando-se o meio CYE suplementado com sulfato de cobre nas concentrações de 50, 100 e 200 μg/mL. Do total de 44 isolados, 80% foram identificados como X. gardneri, 11% como X. perforans e 9% como X. vesicatoria. De acordo com a reação ao cobre, os isolados foram agrupados em quatro classes (S, sensíveis; MS, moderadamente sensíveis; MI, moderadamente insensíveis; I, insensíveis). Quanto à sensibilidade ao cobre, 98% de todos os isolados foram sensíveis a 200 μg/mL, sugerindo que aparentemente, a dosagem recomendada de produtos à base de cobre em campo, 10 vezes maior que a dosagem limítrofe utilizada nos testes in vitro, ainda é eficiente para as diferentes espécies da bactéria. Palavras-chave: Xanthomonas euvesicatoria, X. gardneri, X. perforans, X. vesicatoria, BOX-PCR, cobre, controle químico. ABSTRACT Occurrence and characterization of the species complex causing tomato bacterial spot in "Alto Vale do Rio do Peixe", SC, BrazilThe aim of this study was to identify at the species level Xanthomonas strains causing tomato bacterial spot in the region of "Alto Vale do Rio do Peixe", state of Santa Catarina, Brazil, as well as to determine their in vitro sensitivity to copper. Species were determined by similarity analysis of genomic profiles generated by BOX-PCR and sensitivity to copper was established using the CYE medium supplemented with copper sulfate at concentrations of 50, 100 and 200 μg/mL. Of the 44 isolates, 80% were identified as X. gardneri, 11% as X. perforans and 9% as X. vesicatoria. According to the response to copper, the isolates were divided into four classes (S, sensitive; MS, moderately sensitive; MI, moderately insensitive; I, insensitive). Regarding the sensitivity to copper, 98% of all isolates were sensitive at 200 μg/mL, suggesting that the recommended dosage of copper-based products registered for the crop may still provide effective control of the different bacterial species.

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This work evaluated the field performance and resistance to Peronospora destructor of 46 onion cultivars in two different experiments. Two open-pollinated experimental cultivars: BBola Precoce-Agroecológica, and BSuperprecoce-Agroecológicaŵ ere more productive and resistant. The onion breeding by screenings in agroecological production systems may be an alternative to obtain germplasm less sensitive to biotic and abiotic stresses. Keywords Adaptability. Allium cepa. Downy mildew. Local cultivars. Plant breeding Onion (Allium cepa) is an important vegetable crop grown all over the world. It is the third most economically important vegetable in Brazil, with an estimated production value of BRL 1.81 billion (approximately US$ 600 million) in 2015 (IBGE 2017). The southern region of Brazil concentrates about 33% of this production value. Despite the intensive cultivation of onion in this region, two factors can significantly limit yield: (i) the high occurrence and severity of downy mildew; (ii) and use of cultivars not adapted. Onion downy mildew, caused by Peronospora destructor, has global occurrence and becomes more relevant in temperate regions (Domingues and Tofoli 2009). According to Wordell Filho and Boff (2006), downy mildew is one of the main onion diseases in southern Brazil. van der Meer and de Vries (1990) reported complete resistance to downy mildew found in Allium roylei. Kofoet et al. (1990) determined that the inheritance of resistance was due to a single dominant gene, Pd1. However, to date, there is no resistant commercial cultivar in Brazil. The use of onion cultivars not adapted to a particular region may hinder the formation of bulbs, reducing the yield. There are 437 onion cultivars registered and four protected cultivars in Brazil (MAPA 2017). This paper aimed to evaluate the field performance of 46 onion cultivars in the southern region of Brazil, as well as evaluate the severity and yield loss caused by downy mildew. Thirty-one commercial open-pollinated (OP) onion cultivars, eight experimental hybrids, two commercial hybrids, and five experimental OP cultivars originated by mass selection in agroecological systems (both during the bulb and the seed production phases) were evaluated in two different experiments. The identification of the cultivars is presented in Table 1. The experimental OP cultivar denominated BPoranga-Agroecológica^was originated from BSCS366 Poranga^for two complete cycles in agroecological systems. The following cultivars were selected in five complete cycles in the agroecological system: cv. BSuperprecoce-Agroecológico^originated from Epagri 363 Superprecoce^; BBola Precoce-Agroecológica^originated from BEmpasc 352 Bola Precoce^; BJuporanga-Agroecológica^from BEmpasc 355 Juporangaâ nd BCrioula Alto Vale-Agroecógica^from BEpagri 362 Crioula Alto Vale^. Onion seedlings were grown on soil bed and approximately 60 days after sowing, they were transplanted to the two experiments that remained from July to December in 2015. The light hours required by cultivars of short or i...