ABSTRACT: Seventy-five pectinolytic strains collected from vegetables grown in the counties of Santarém, Belterra, and Mojuí dos Campos, located in the western region of the Pará State, Brazil, were studied according to their pathological and genetic variability. The strains were grouped in 5 clusters according to pathogenicity in potato, pepper, carrot, and onion, and 38 strains were selected for genetic analysis using rep-PCR. These strains were divided into 35 genetic groups according to rep-PCR at 70% similarity. These results indicated high pathological and genetic variability of the strains causing soft rot in vegetables in the western region of the Pará State, which will be used in etiological research and for the development and assessment of management techniques for the soft rot in vegetables in this region. ABSTRACT Seventyfive Seventy five Santarém Belterra Campos State Brazil potato pepper carrot onion 3 repPCR. repPCR rep PCR. PCR 70 similarity 7

RESUMO: Setenta e cinco isolados de bactérias pectinolíticas coletados de hortaliças cultivadas nos municípios de Santarém, Belterra e Mojuí dos Campos, região oeste do Estado do Pará, Brasil, foram estudadas de acordo com sua variabilidade patológica e genética. Os isolados foram agrupados em cinco grupos de acordo com a patogenicidade em batata, pimenta, cenoura e cebola, e 38 isolados foram selecionados para análise genética por rep-PCR. Esses isolados foram divididos em 35 grupos genéticos de acordo com rep-PCR a 70% de similaridade. Esses resultados indicaram alta variabilidade patológica e genética dos isolados causadores da podridão mole em hortaliças da região oeste do Pará, os quais serão utilizados em pesquisas etiológicas e para o desenvolvimento e avaliação de técnicas de manejo da podridão mole em hortaliças nesta região. RESUMO Santarém Campos Pará Brasil batata pimenta cebola 3 repPCR. repPCR rep PCR. PCR 70 similaridade 7

Abstract Banana tree bacterial wilt is caused by the Ralstonia solanacearum Moko ecotype. These strains vary in their symptom progression in banana, and are classified as typical Moko variants (phylotype IIA and IIB strains from across Central and South America), Bugtok variant (Philippines), and Sergipe facies (the states of Sergipe and Alagoas, Brazil). This study used comparative genomic and phylogenomic approaches to identify a correlation between the symptom progression of the Moko ecotypes based on the analysis of 23 available genomes. Average nucleotide identity and in silico DNA-DNA hybridization revealed a high correlation (>96% and >78%, respectively) between the genomes of Moko variants. Pan-genome analysis identified 21.3% of inheritable regions between representatives of the typical Moko and Sergipe facies variants, which could be traced to an abundance of exclusive homolog clusters. Moko ecotype genomes shared 1,951 orthologous genes, but representatives with typical symptoms did not display unique orthologues. Moreover, Bugtok disease and Sergipe facies genomes did not share any unique genes, suggesting convergent evolution to a shared symptom progression. Overall, genomic and phylogenomic analyses were insufficient to differentiate the Moko variants based on symptom progression.

Abstract Banana vascular wilt or Moko is a disease caused by Ralstonia solanacearum. This study aimed to sequence, assemble, annotate, and compare the genomes of R. solanacearum Moko ecotypes of different sequevar strains from Brazil. Average nucleotide identity analyses demonstrated a high correlation (> 96%) between the genome sequences of strains CCRMRs277 (sequevar IIA-24), CCRMRs287 (IIB-4), CCRMRs304 (IIA-24), and CCRMRsB7 (IIB-25), which were grouped into phylotypes IIA and IIB. The number of coding sequences present in chromosomes and megaplasmids varied from 3,070 to 3,521 and 1,669 to 1,750, respectively. Pangenome analysis identified 3,378 clusters in the chromosomes, of which 2,604 were shared by all four analyzed genomes and 2,580 were single copies. In megaplasmids, 1,834 clusters were identified, of which 1,005 were shared by all four genomes and 992 were identified as single copies. Strains CCRMRsB7 and CCRMRs287 differed from the others by having unique clusters in both their chromosomes and megaplasmids, and CCRMRsB7 possessed the largest genome among all Moko ecotype strains sequenced to date. Therefore, the genomic information obtained in this study provides a theoretical basis for the identification, characterization, and phylogenetic analysis of R. solanacearum Moko ecotypes.

Abstract Ralstonia solanacearum is the causal agent of Moko disease in bananas, which in the state of Sergipe in northeastern Brazil causes “Sergipe facies”. This disease induces atypical symptoms similar to those of Bugtok disease in the Philippines. This study was conducted to sequence, assemble, and annotate the genomes of the Sergipe facies-causing isolates SFC and IBSBF2570 (sequevar IIA-53) and compare their genomes with two representative isolates causing Bugtok disease. The genomes were sequenced and assembled, resulting in lengths of 5.58 Mb (SFC) and 5.46 Mb (IBSBF2570) in 185 and 174 contigs, respectively. The isolates of Sergipe facies and Bugtok disease showed similarities in their gene contents. We identified 5,668 information clusters, 3,752 of which were shared by all genomes (core genes). Moreover, 3,585 single-copy genes were identified. Isolates causing Bugtok disease exclusively shared 266 more information clusters than the isolates causing Sergipe facies. These results suggest that Sergipe facies and Bugtok disease isolates show high genomic similarity. However, the similarity is even greater between the Bugtok disease isolates. This may be because of their longer period of interaction compared to Sergipe facies isolates.

Resumo A sobrevivência de Acidovorax citrulli Aac1Rif foi estudada em tecidos infectados de frutos e folhas de melão incorporadas ao solo a 0; 5; 10 e 15 cm de profundidade, em sete diferentes tipos de solo, às temperaturas de 10; 15; 20; 25; 30 e 35 ºC e umidade na capacidade de campo de 50 e 100%, na ausência da planta hospedeira. Aac1Rif foi detectado em frutos de melão e tecidos foliares a 0; 5 e 10 cm até 21 dias, e a 15 cm até 14 dias. A taxa de extinção relativa mais alta e mais baixa da população (TERP) para Aac1Rif ocorreu, respectivamente, nos tecidos de frutos e nos tecidos foliares, às profundidades de 0 e 5 cm. Para a maioria dos solos, a TERP mais baixa ocorreu a 10 ou 15 ºC, e a mais alta, a 30 ou 35 ºC. Maiores concentrações de Na+, silte e maiores populações de actinomicetos e Trichoderma foram correlacionadas com maior TERP de Aac1Rif no solo. Houve diferença significativa entre a TERP em 100% e 50% da capacidade do campo. O solo não foi considerado fonte primária potencial de inóculo de A. citrulli. Os frutos e folhas de melão infectados no solo foram considerados como fontes de inóculo, pelo menos por 21 dias.

Abstract The survival of Acidovorax citrulli Aac1Rif was accessed in infected melon tissues (fruits and leaves) incorporated to the soil at 0, 5, 10 and 15 cm depth, in seven different types of soil, at temperatures 10, 15, 20, 25, 30 and 35 ºC and moisture field capacity of 50 and 100% in the absence of the host plant. Aac1Rif was detected in melon tissues at 0, 5 and 10 cm until 21 days and at 15 cm until 14 days. The highest and lowest relative extinction rate of the population (RERP) for Aac1Rif occurred respectively in fruit tissues and leaf tissues at depths of 0 and 5 cm. Aac1Rif survived in seven types of soil only for three days. The lowest RERP occurred at 10 or 15 ºC and the highest at 30 or 35 ºC. Greater concentrations of Na+, silt, and greater populations of actinomycetes and Trichoderma were correlated with highest RERP of the Aac1Rif in the soil. There was significant difference between RERP at 100% and 50% of field capacity. The soil was not considered potential primary source of A. citrulli inoculum. Infected melon fruits and leaves in soil were considered as such sources, at least for 21 days.

A disseminação de Xanthomonas campestris pv. viticola (Xcv), agente do cancro bacteriano da videira, ocorre, dentre outras formas, por meio de mudas e bacelos infectados. Foi estudada a obtenção de material propagativo livre do patógeno, testando a eficiência do tratamento de bacelos com termoterapia, bactericidas e sanitizantes. Os isolados de Xcv foram testados quanto à patogenicidade e realizado o teste de sensibilidade in vitro aos produtos, em diferentes concentrações. A erradicação de Xcv em bacelos de videira foi testada em experimentos com termoterapia (50ºC por 30 e 40 min; 53ºC por 5 e 10 min); bactericidas [oxitetraciclina+sulfato de cobre (150+2.000; 165+2.200; 180+2.400 e 195+2.600 mg L-1 de H2O) e oxitetraciclina (600; 700; 800 e 900 mg L-1)]; e sanitizantes [cloreto de dodecildimetil amônio (600; 1.200; 1.800; 2.400 e 3.000 µL L-1); hipoclorito de sódio (5.000; 10.000; 20.000; 30.000 e 40.000 µL L-1) e cloreto de benzalcônio (125; 167;250; 334 e 500 µL L-1)]. Foram avaliados período de incubação, incidência e severidade da doença. O bactericida oxitetraciclina e os sanitizantes cloreto de dodecildimetil amônio e hipoclorito de sódio proporcionaram os maiores halos de inibição de Xcv in vitro. No entanto, apesar dos diversos tratamentos testados, não foi possível recomendar tratamento termoterápico ou produto que erradicasse Xcv de bacelos infectados. Porém, ficou confirmada a grande importância destes na disseminação do agente do cancro bacteriano da videira.

The spread of Xanthomonas campestris pv. viticola (Xcv), causal agent of grapevine bacterial canker, occurs among other ways, through infected plants and grapevine cuttings. It was studied how to obtain propagation material free of Xcv testing the effectiveness of treatments of cuttings using thermotherapy, bactericides and sanitizers. The Xcv isolates were tested for pathogenicity and in vitro sensitivity to bactericides and sanitizers at different concentrations. The eradication of Xcv on cuttings was tested in experiments with thermotherapy (50ºC for 30 and 40 min; 53ºC for 5 and 10 min); bactericides [oxytetracycline+copper sulphate (150+2000, 165+2200, 180+2400 and 195+2600 mg L-1 of H2O) and oxytetracycline (600, 700, 800 and 900 mg L-1)] and sanitizers [dodecyldimethyl ammonium chloride (600, 1200, 1800, 2400 and 3000 µl L-1) sodium hypochlorite (5000, 10000, 20000, 30000 and 40000 µl L-1) and benzalkonium chloride (125, 167, 250, 334 and 500 µl L-1)]. We evaluated the incubation period, the incidence and severity of disease. The bactericidal oxytetracycline and sanitizers dodecyldimethyl ammonium chloride and sodium hypochlorite provided the largest zones of inhibition, in vitro. However, despite the several treatments tested we cannot recommend an efficient condition of temperature/time, and concentrations of bactericides and sanitizers capable of eradicating Xcv from infected grapevine cuttings, thus confirming their great importance for the spread of grapevine bacterial canker.

A ação de óleos essenciais na redução da podridão mole em couve-chinesa e sua influência na colorimetria e características físico-químicas da hortaliça foram avaliadas. Em casa de vegetação, plantas da cultivar Natsume foram pulverizadas com 11 óleos selecionados em testes preliminares de fitotoxidez: bergamota, capim-limão, copaíba, Eucalyptus citriodora, E. globulus, funcho de erva-doce, gengibre, hortelã, laranja doce, limão siciliano e sálvia esclaréia a 0,5%, e com o antibiótico Mycoshield® (3 g L-1). Após 72 h, as plantas foram inoculadas com Pectobacterium carotovorum subsp. carotovorum (Pcc-c). Os óleos e o Mycoshield® reduziram significativamente (P<0,05) a severidade (SEV) e a área abaixo da curva de progresso da doença (AACPD). Os óleos de bergamota, copaíba, E. citriodora, hortelã e laranja doce foram então testados quanto à estabilidade da eficácia de controle em relação a três isolados de P. carotovorum subsp. carotovorum. Estes óleos reduziram a SEV (30,5 a 38,6%) e a AACPD (23,1 a 26,6%) sem diferirem entre si ou do Mycoshield® (SEV 45,2 e AACPD 32,8%) exceto o de copaíba (20,3%), menos eficiente que o antibiótico na redução da AACPD. Apenas Mycoshield® inibiu o patógeno in vitro. Nenhum tratamento alterou a colorimetria, teor de ácido ascórbico e pH das folhas da couve-chinesa. O óleo de hortelã elevou a acidez total titulável, assim como os óleos de laranja doce, E. citriodora e bergamota elevaram os sólidos solúveis totais. Portanto, a pulverização com os óleos de bergamota, copaíba, E. citriodora, hortelã e laranja doce têm potencial como medida de controle da doença.

The action of essential oils in reducing soft rot in Chinese cabbage, and their influence on the colorimetry and physicochemical characteristics of the vegetable were evaluated. In the greenhouse, plants of the cultivar Natsume were sprayed with 11 oils selected in preliminary tests for phytotoxicity: bergamot, lemongrass, copaiba, Eucalyptus citriodora, blue gum, fennel, ginger, spearmint, sweet orange, lemon and clary sage (0.5%) and also the antibiotic Mycoshield® (3 g L-1). After 72 hours the plants were inoculated with Pectobacterium carotovorum subsp. carotovorum (Pcc-c). The oils and the Mycoshield® significantly reduced (P<0.05) the severity (SEV) and the area under the disease progress curve (AUDPC). The oils of bergamot, copaiba, E. citriodora, spearmint and sweet orange were then tested for the stability of their effectiveness in the control of three isolates of P. carotovora subsp. carotovorum. These oils reduced the SEV (30.5 to 38.6%) and the AUDPC (23.1 to 26.6%) with no differences between them or the Mycoshield® (SEV 45.2 and AUDPC 32.8%), except for the copaiba (20.3%) which was less effective than the antibiotic in the reduction of the AUDPC. In vitro, only Mycoshield® inhibited the pathogen. None of the treatments altered the colorimetry, levels of ascorbic acid or pH of the leaves of the Chinese cabbage. The spearmint oil increased the total titratable acidity in the same way as the oils of sweet orange, E. citriodora and bergamot increased the total soluble solids. Therefore, spraying with the oils of bergamot, copaiba, E. citriodora, spearmint and sweet orange has potential in the control of this disease.

Soft rot, caused by Pectobacterium carotovorum subsp. carotovorum (Pcc), is the main bacterial disease affecting lettuce (Lactuca sativa L.) crops in Brazil and leads to significant yield losses. This study aimed to assess the reaction of lettuce genotypes to soft rot induced by a virulent isolate and the stability of the resistance to three isolates varying in virulence. Using a descriptive ordinal scale ranging from 1 to 9 a classification system was defined: class 1 = resistant (R): severity (Sev) < 1.5; class 2 = moderately resistant (MR): 1.5 < Sev < 2.5; class 3 = susceptible (S): 2.5 < Sev < 3.5; class 4 = highly susceptible (HS): Sev > 3.5). Of the 41 tested genotypes, 14 were classified as MR and 27 as S when inoculated with a Pcc isolate of intermediate virulence. Eleven of these genotypes (four S and seven MR) were selected to test their resistance stability against three other isolates with an increasing degree of virulence (Pcc36 < Pcc-A1.1 < Pcc-23). Out of the 11 genotypes eight retained the original classification and three moved from S to MR resistant class when challenged with the least virulent isolate. Vitória de Santo Antão was the only genotype classified as MR for all tested isolates and is a promising candidate for durable soft rot resistance breeding.

O cancro bacteriano da videira é causado por Xanthomonas campestris pv. viticola (Xcv). Visando à limpeza clonal de mudas de 'Red Globe', foram estudados: tamanho ideal de ápices e gemas axilares para cultivo em meio de Galzy modificado (MGM); efeito da termoterapia (38ºC/30 dias); e ação de antibióticos na eliminação de Xcv em videiras infectadas. Os percentuais de contaminação por Xcv e de regeneração foram analisados, e as plantas obtidas foram indexadas em meio ágar nutritivo-dextrose-extrato de levedura-ampicilina (NYDAM), seguindo-se teste de patogenicidade. O cultivo de explantes com 3 mm possibilitou a obtenção de plantas livres da bactéria, com regeneração 14,3 vezes maior que explantes com 1 mm. A termoterapia de mudas infectadas, associada ao cultivo in vitro, não eliminou o patógeno. O cultivo de explantes com 10 mm, durante 40 dias em MGM + cefotaxima (300 mg L-1), proporcionou limpeza clonal das mudas. A indexação de plantas de videira regeneradas in vitro, quanto à infecção por Xcv utilizando NYDAM, seguida de teste de patogenicidade, é uma alternativa econômica e eficiente para produção de mudas de alta qualidade fitossanitária.

Bacterial canker is caused by Xanthomonas campestris pv. viticola (Xcv). In order to eliminate Xcv from 'Red Globe' plants it was studied: optimal size of meristem tips and axillary buds for cultivation in modified Galzy's medium (MGM); effects of thermotherapy (38ºC/30 days); and action of antibiotics in the elimination of Xcv in infected grapevines. The percentages of contamination by Xcv and regeneration were analyzed and plants obtained were indexed using the semi-selective culture medium nutrient agar-dextrose-yeast extract-ampicilin (NYDAM) followed by a pathogenicity test. The cultivation of 3 mm explants permitted to obtain plants free of bacteria with regeneration 14.3 times higher than 1 mm explants. The thermotherapy of infected plants associated to the in vitro culture did not eliminate the pathogen. The cultivation of 10 mm explants during 40 days in MGM + cefotaxime (300 mg L-1) eliminated Xcv from grapevine plants. The indexation of micropropagated grapevine plants for Xcv infection by using NYDAM medium followed by a pathogenicity test is an economical and efficient alternative to produce plants of high sanity quality.

Tecidos de plantas infectados constituem uma importante fonte de inóculo primário para fitobacterioses. O objetivo deste trabalho foi investigar a sobrevivência de Xanthomonas campestris pv. viticola (Xcv), agente do cancro bacteriano da videira, em tecidos infectados na superfície do solo e durante a compostagem de restos de poda. Mudas de videira 'Festival' foram inoculadas com mutante resistente à rifampicina Xcv2Rif e mantidas em casa de vegetação até apresentarem alta severidade da doença. No primeiro experimento, ramos fragmentados e folhas inteiras destas plantas foram acondicionados em bolsas de malha plástica e colocados na superfície de microparcelas. No segundo experimento, ramos e folhas fragmentados foram acondicionados em bolsas plásticas e depositados no interior de pilhas de compostagem de restos de poda de videira. A sobrevivência de Xcv2Rif em tecidos infectados foi monitorada a intervalos de 8 e 10 dias a partir do início do primeiro e segundo experimentos, respectivamente. No primeiro experimento, foi também avaliada a decomposição de tecidos, e no segundo, as curvas de temperatura das pilhas, conteúdo de fenóis e microbiota antagonista a Xcv2Rif. O patógeno sobreviveu em altas populações até 80 dias em tecidos infectados na superfície do solo. A compostagem eliminou Xcv2Rif de restos culturais em 10 dias, devido às altas temperaturas, liberação de compostos fenólicos e antagonismo microbiano. Concluindo, em tecidos infectados de videira Xcv sobrevive na superfície do solo por, pelo menos, 80 dias mas é eliminada pela compostagem em 10 dias.

Infected plant tissues are an important primary inoculum sources for plant bacterial diseases. This study investigated the survival of Xanthomonas campestris pv. viticola (Xcv), the agent of bacterial canker of grapevine in infected tissues on soil surface and during composting process of pruning residues. Grapevine plants cv. Festival were inoculated with a mutant resistant to rifampicin Xcv2Rif and maintained under greenhouse conditions until presenting high disease severity. In the first experiment, fragmented shoots and entire leaves from these plants, were placed into mesh bags and located on the surface of microplots. In the second experiment fragmented shoots and leaves were also placed into mesh bags and located inside composting piles of grapevine pruning residues. The survival of Xcv2Rif in infected tissue was monitored at 8 and 10 days intervals from 1 and 2 experiment setting, respectively. In the experiment 1 tissue decomposition was also evaluated and in the experiment 2, pile temperature curves, phenolyc content, and antagonism against Xcv2Rif were analyzed. The pathogen survived in high populations for at least 80 days in grapevine-infected tissues on soil surface. The composting process eliminated Xcv2Rif from crop residues in 10 days due to high temperatures, liberation of phenolyc compounds and microbial antagonism. In conclusion, Xcv survives in grapevine infected tissues on soil surface at least until 80 days but is eradicated by composting in 10 days.