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1

OLIVEIRA, Janaína Cortêz de. "Caracterização de isolados de Acidovorax avenae subsp. citrulli." Universidade Federal Rural de Pernambuco, 2006. http://www.tede2.ufrpe.br:8080/tede2/handle/tede2/6568.

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Submitted by (lucia.rodrigues@ufrpe.br) on 2017-03-13T15:35:27Z No. of bitstreams: 1 Janaina Cortez de Oliveira.pdf: 511674 bytes, checksum: 0e608fbee62310affc8c094a69725337 (MD5)<br>Made available in DSpace on 2017-03-13T15:35:27Z (GMT). No. of bitstreams: 1 Janaina Cortez de Oliveira.pdf: 511674 bytes, checksum: 0e608fbee62310affc8c094a69725337 (MD5) Previous issue date: 2006-02-03<br>Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq<br>The fruit blotch caused by Acidovorax avenae subsp. citrulli is the main bacterial disease of melon (Cucumis melo) in the Northeast of Brazil. In this work 49 isolates of A. avenae subsp. citrulli were studied aiming: to investigate the production of enzymes (pectinolytic, amylolytic, cellulolytic, lipolytic and proteolytic), phytohormone (indoleacetic acid), polysaccharide (levan) and toxin (syringomycin) by isolates of A. avenae subsp. citrulli and; to characterize this population based upon disease severity on seedlings, plants and fruits of melon and watermelon (Citrullus lanatus), hypersensitivity in tobacco (Nicotiana tabacum), utilization of amino acids as carbon and energy sources, and in vitro sensitivity to copper and antibiotics. All isolates of A. avenae subsp. citrulli produced lipase and indoleacetic acid, and none showed pectinolytic, amylolytic, cellulolytic and proteolytic activity nor produced levan and syringomycin. All isolatesinduced typical symptoms of fruit blotch on seedlings, plants and fruits of melon and watermelon. The Scott-Knott test (P = 0.05) separated the isolates for disease index in 5 and 7 groups respectively for melon and watermelon seedlings and 2 groups for plants of these two hosts. In fruits all isolates were separated in 3 and 10 groups for the variable diameter of extern lesion and 2 and 9 groups for lesion depth, respectively for melon and watermelon. All isolates also induced hypersensitivity in tobacco; utilized the amino acids asparagine, L-leucine and DL-acid lactic; showed sensitivity to copper oxychloride (120 μg ml-1), cuprous oxide (120 μg ml-1), copper hydroxide (138.2 μg ml-1), streptomycin sulfate (25 μg ml-1) and Agrimaicin 500 (428 μg ml-1); and resistance to kasugamycin (87 μg ml-1), agrimicin (200 μg ml-1), erythromycin (15 μg), gentamicin (10 μg), amoxicilin (10 μg), neomycin (30 μg) streptomycin (10 μg), norfloxacin (10 μg)and rifampicin (5 μg). Variability was found among the 41 isolates in relation toand rifampicin (5 μg). Variability was found among the 41 isolates in relation to<br>A mancha-aquosa causada por Acidovorax avenae subsp. citrulli é a principal doença bacteriana do meloeiro (Cucumis melo) no Nordeste do Brasil. Neste trabalho, foram estudados 41 isolados de A. avenae subsp. citrulli com os objetivos de: investigar a produção de enzimas (pectinolíticas, amilolíticas, celulolíticas, lipolíticas e proteolíticas), fitohormônio (ácido indol acético), polissacarídeo (levana) e toxina (siringomicina) por isolados de A. avenae subsp. citrulli e; caracterizar essa população com base na severidade da doença em plântulas, plantas e frutos de meloeiro e melancieira (Citrullus lanatus), reação de hipersensibilidade em fumo (Nicotiana tabacum), utilização de aminoácidos como fonte de carbono e energia e sensibilidade in vitro a cúpricos e antibióticos. Todos os isolados de A. avenae subsp. citrulli produziram enzimas lipolíticas e ácido indol acético, e nenhum apresentou atividade pectinolítica, amilolítica, celulolíticas e proteolítica ou produção do polissacarídeo levana e da toxina siringomicina. Em plântulas, plantas efrutos, todos os isolados induziram sintomas típicos da mancha-aquosa em meloeiro e melancieira. Pelo teste de agrupamento de Scott-Knott (P = 0,05) os isolados foram separados quanto ao índice de doença em 5 e 7 grupos, respectivamente para plântulas meloeiro e melancieira, e em 2 grupos para plantas das duas hospedeiras. Em frutos, os isolados foram separados em 3 e 10 grupos para a variável diâmetro da lesão externa e 2 e 9 grupos para profundidade da lesão, respectivamente para meloeiro e melancieira. Todos os isolados também induziram reação de hipersensibilidade em fumo; utilizaram os aminoácidos asparagina, L-leucina e DL-ácido lático; foram sensíveis a oxicloreto de cobre (120 μg ml-1), óxido cuproso (120 μg ml-1), hidróxido de cobre (138,2 μg ml-1), sulfato de estreptomicina (25 μg ml-1) e Agrimaicin® 500 (428 μg ml-1); e resistentes a kasugamicina(87 μg ml-1), agrimicina (200 μg ml-1), eritromicina (15 μg), gentamicina (10 μg),amoxicilina (10 μg), neomicina (30 μg), estreptomicina (10 μg), norfloxacina (10 μg) e rifampicina (5 μg). Foi constatada variabilidade entre os 41 isolados de A. avenae subsp. citrulli quanto a sensibilidade à tetraciclina (30 μg), sendo 41,5% resistentes, 46,32% moderadamente sensíveis e 12,2% altamente sensíveis
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2

SOUZA, Elineide Barbosa de. "Variabilidade de Acidovorax avenae subsp. citrulli e epidemiologia da mancha-aquosa do melão." Universidade Federal Rural de Pernambuco, 2002. http://www.tede2.ufrpe.br:8080/tede2/handle/tede2/6587.

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Submitted by (lucia.rodrigues@ufrpe.br) on 2017-03-16T14:38:17Z No. of bitstreams: 1 Elineide Barbosa de Souza.pdf: 390725 bytes, checksum: 343f1ce1b17f2c507a8bc4587f0077ac (MD5)<br>Made available in DSpace on 2017-03-16T14:38:17Z (GMT). No. of bitstreams: 1 Elineide Barbosa de Souza.pdf: 390725 bytes, checksum: 343f1ce1b17f2c507a8bc4587f0077ac (MD5) Previous issue date: 2002-06-28<br>The variability of 20 Acidovorax avenae subsp. citrulli isolates in relation to melon fruit blotch components, hypersensitive reaction, and the bacterial transmission by seeds from inoculated fruits were studied. The influence of duration (0, 6, 24 and 48 h), the onset of leaf wetness period (0, 6, 24 and 48 h after inoculation), and inoculum concentration of A. avenae subsp. citrulli (3.4 x 101 to 3.4 x 107 CFU mL-1) on severity of fruit blotch in melon plants were also evaluated. The effects of temperature (15, 20, 25, 30, 35 and 40° C), humidity (0 and 6 h of moist chamber), inoculum concentration (3.4 x 101 to 3.4 x 107 CFU mL-1) and fruit age (40, 50, 60 and 70 days) on the development of melon fruit blotch were also verified. Seeds, plants and fruits were inoculated through vacuum infiltration, atomization and sub-epidermal injection, respectively. Seedlings and plants were assessed in relation to incubation period, diseaseindex, area under disease progress curve and disease progress rate; incubation period, diameter of external lesion and lesion depth were assessed on the fruits. The data were submitted to mean comparison tests, clustering tests or regression analysis. The Euclidian distance–single linkage confirmed the variability among the A. avenae subsp. citrulli strains allowing their separation in four similarity groups. Seed transmission ranged from 30 to 64 % and all strains induced hypersensitive reaction on tobacco and tomato leaves. The regression equations for the analyzed variables in melon plants were better adjusted by the quadratic or logarithmic models. The incubation period ranged from 1.3 to 2.7 days and was higher in plants without leaf wetness, although the disease index and area under disease progress curve increased as the duration of leaf wetness increased. The beginning of the leaf wetness period at 48 h after inoculation elevatedthe incubation period and disease progress rate in relation to the other periods. The disease progress rate, disease index and area under disease progress curve increased as the inoculum concentration increases, reaching maximum values of 4.4 infection units/day, 73.7 % and 18.9 at 3.4 x 107 UFC mL-1, respectively, at 3.4 x 101 CFU mL-1. The temperature and humidity influenced significantly (P=0,05) the severity of melon fruit blotch, however, the incubation period was not affected. The larger external lesions were observed in the fruits incubated at 35 and 30° C without moist chamber, and at 30° C in moist chamber for six h. In relation to lesion depth, those lesions in fruits incubated without moist chamber were deeper at 25 and 30° C. However, with moist chamber the lesions at 30° C were deeper than the others. No disease symptoms were observed onfruits incubated at 15 and 20° C. The humidity significantly (P=0,05) influenced the development of external lesions and lesions depth at 35 and 25° C, respectively. The diameter and depth of lesions increased as the inoculum concentration was higher and were reduced as the fruit age increased. No external or internal lesions were detected on fruits with 60 and 70 days inoculated with the pathogen at 3.4 x 101 CFU mL-1.<br>Foi estudada a variabilidade de 20 isolados de Acidovorax avenae subsp. citrulli quanto aos componentes da mancha-aquosa do melão e reação de hipersensibilidade, e analisada a transmissão da bactéria pelas sementes dos frutos inoculados. Foi também avaliada a influência da duração (0, 6, 24 e 48 horas) e início do período de molhamento foliar (0, 6, 24 e 48 horas após a inoculação), bem como da concentração de inóculo de A. avenae subsp. citrulli (3,4 x 101 a 3,4 x 107 UFC mL-1) na severidade da mancha-aquosa em meloeiro. A influência da temperatura (15, 20, 25, 30, 35 e 40° C), da umidade (0 e 6 horas de câmara úmida), da concentração de inóculo ( 3,4 x 101 a 3,4 x 107 UFC mL-1) e da idade do fruto (40, 50, 60 e 70 dias) no desenvolvimento da mancha-aquosa em melão, foram ainda verificadas. Sementes, plantas e frutos foram inoculados pelos métodos de infiltração a vácuo, pulverização e injeção subepidérmica, respectivamente. Plântulas e plantas foram avaliadas quanto ao período de incubação,índice de doença, área abaixo da curva do progresso da doença e taxa de progresso da doença, e os frutos, quanto ao período de incubação, diâmetro da lesão externa e profundidade da lesão. Os dados obtidos foram submetidos a testes de comparação de médias, testes de agrupamento ou análises de regressão. A análise da distância Euclidiana por ligações simples, confirmou a variabilidade entre os isolados de A. avenae subsp. citrulli, permitindo a separação destes em quatro grupos de similaridade. A transmissão da bactéria por sementes variou de 30 a 64 % e todos os isolados induziram reação de hipersensibilidade em folhas de fumo e tomate. As equações de regressão para as variáveis analisadas em meloeiros foram melhor ajustadas pelos modelos quadrático ou logarítmico. O período de incubação variou de 1,3 a 2,7 dias, emaior nas plantas sem molhamento foliar, contudo o índice de doença e a área abaixo da curva de progresso da doença aumentaram com o incremento da duração do molhamento foliar. O início do período de molhamento foliar às 48 horas após a inoculação elevou o período de incubação e a taxa de progresso da doença em relação aos demais períodos. O incremento da concentração de inóculo elevou a taxa de progresso da doença, índice de doença e área abaixo da curva de progresso da doença, os quais atingiram valores máximos de 4,4 unidades de infecção/dia, 73,7 % e 18,9, respectivamente, na concentração 3,4 x 107 UFC mL-1. A temperatura e umidade influenciaram significativamente a severidade da mancha-aquosa nos frutos, embora o período de incubação não tenha sido afetado. As maiores lesões externas foram observadas em frutos incubados a 35 e 30 ° C sem câmara úmida, e a 30° C em câmaraúmida por seis horas. Com relação à profundidade, as lesões nos frutos incubados sem câmara úmida foram maiores às temperaturas de 25 e 30° C. Em câmara úmida, as lesões a 30° C foram maiores que as demais. Não foi observado desenvolvimento da mancha-aquosa em frutos incubados a 15 e 20° C. A umidade influenciou significativamente (P=0,05) o diâmetro e profundidade da lesão, exceto às temperaturas de 35 e 25° C, respectivamente. O diâmetro e profundidade das lesões aumentaram com a elevação da concentração de inóculo e foram reduzidos com o aumento da idade do fruto. Na concentração 3,4 x 101 UFC mL-1 não foi observada a presença de sintomas da doença na casca e na polp
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Roza, dos Santos Elizama. "Controle biológico da mancha-aquosa do melão causada por Acidovorax avenae subsp. citrulli." Universidade Federal de Pernambuco, 2004. https://repositorio.ufpe.br/handle/123456789/1420.

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Made available in DSpace on 2014-06-12T15:49:51Z (GMT). No. of bitstreams: 2 arquivo4438_1.pdf: 555928 bytes, checksum: f7ae357b001b006fb69c9507a2f06dd5 (MD5) license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5) Previous issue date: 2004<br>A mancha-aquosa, causada pela bactéria Acidovorax avenae subsp. citrulli, é hoje um problema para a cultura do melão nas áreas produtoras do Nordeste, principalmente nos Estados do Ceará e Rio Grande do Norte. A doença, sobretudo durante o período chuvoso, chega a dizimar boa parte da produção em algumas lavouras. Microrganismos têm sido usados no controle de doenças de plantas e o gênero Bacillus é citado como um dos mais utilizados, devido, em grande parte, à sua capacidade de produzir substâncias antimicrobianas, entre as quais lipopeptídeos. O objetivo deste trabalho foi investigar o controle da mancha aquosa do melão por espécies de Bacillus. Foram realizados testes in vivo, aplicando-se caldos fermentados de B. subtilis R14, B. megaterium pv. cerealis RAB7, B. megaterium C116 e Bacillus sp. MEN2, com e sem de células, em sementes de melão anteriormente inoculadas com A. avenae subsp. citrulli. As fermentações foram realizadas em mesa agitadora e o plantio das sementes em bandeja. Testes de antagonismo in vitro foram também realizados pelo método de difusão em ágar. A avaliação dos testes in vivo foi realizada através das seguintes variáveis: incidência (INC = porcentagem de plantas com sintomas) e período de incubação (PI = número de dias transcorridos do plantio até o aparecimento dos sintomas). A severidade foi avaliada, de acordo com escala de notas, diariamente, durante cinco dias, calculando-se o índice de doença (IDO) e área abaixo da curva de progresso da doença (AACPD). Os resultados foram submetidos à análise de variância e as médias comparadas pelo teste de Tukey. B. subtilis R14, B. megaterium. pv. cerealis RAB7, B. megaterium C116 e Bacillus sp. MEN2 apresentaram atividade in vitro contra A. avenae subsp. citrulli e controlaram a doença in vivo, sem diferença significativa entre as quatro linhagens de Bacillus. Os tratamentos, com e sem células, também não apresentaram diferença significativa, indicando que a inibição do crescimento do fitopatógeno ocorreu devido à presença de compostos bioativos produzidos durante as fermentações. Estes compostos foram parcialmente caracterizados como lipopeptídeos, através de testes de hemólise e de atividade surfactante
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Frare, Vanessa Cristina. "Tratamento de sementes de melão (Cucumis melo L.) para o controle de Acidovorax avenae subsp. citrulli." Universidade de São Paulo, 2010. http://www.teses.usp.br/teses/disponiveis/11/11135/tde-25052010-092252/.

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O melão é uma fruta de grande importância para o comércio de exportação do Brasil e, embora restrito a um pequeno número de estados produtores, o cultivo dessa fruta ampliou-se de forma significativa nos últimos anos. Um dos maiores problemas para essa cultura é a presença de patógenos, como a bactéria Acidovorax avenae subsp. citrulli (Aac), responsável por perdas de até 100%. O uso de sementes garantidamente sadias é a principal e mais efetiva medida de controle preconizada para essa bacteriose. Este trabalho teve como objetivo selecionar, in vitro e posteriormente in vivo, produtos capazes de erradicar a bactéria Aac de sementes de melão. Sessenta produtos fitossanitários comerciais foram selecionados para a avaliação do controle in vitro de quatro isolados, sendo nove bactericidas e 51 fungicidas, correspondendo a 6 e 45 ingredientes ativos distintos, respectivamente. Além desses produtos, avaliou-se também o efeito do óleo de melaleuca e do ácido peroxiacético sobre o crescimento dos isolados. Nos ensaios in vivo, sementes inoculadas e naturalmente infectadas foram tratadas, de maneira convencional e a vácuo, com os produtos e doses selecionados no ensaio in vitro. Todas as plântulas foram avaliadas quanto à presença de sintomas característicos de mancha-aquosa aos 21 dias após a emergência. Os antibióticos casugamicina (100 e 200 ug/L), oxitetraciclina (10, 100 e 200 ug/L), oxitetraciclina+sulfato de cobre (10, 100 e 200 ug/L), os fungicidas captana, carboxina+tiram, cloreto de benzalcônio, mancozebe+oxicloreto de cobre, metiram, metiram+piraclostrobina e tebuconazol, nas doses de 100 e 200 ug/L e o ácido peroxiacético a partir da dose de 300 ug/L apresentaram resultados satisfatórios na avaliação in vitro. No ensaio in vivo nenhum tratamento foi eficiente na erradicação da bactéria das sementes.<br>Melon is a fruit of great importance for the export trade of Brazil. Although cultivated in a small number of states, the cropping of this fruit has expanded significantly in recent years. Pathogens represent the most limiting factors, among which stands out the bacterium Acidovorax avenae subsp. citrulli (Aac), responsible for up to 100% losses. The use of healthy seed is the main and the most effective control measure of this bacterium. This study aimed to select first in vitro and subsequently in vivo, erradicant chemicals of Aac in melon seeds. Sixty commercial products were were tested in vitro in the control of four isolates, being nine bactericides and 51 fungicides, corresponding to six and 45 distinct active ingredients, respectively. Besides these products the effects of oil of the melaleuca tea tree and peroxyacetic acid were also evaluated. In the in vivo tests, inoculated and naturally infected seeds were treated by conventional manner and under vacuum, with the products and doses selected in the in vitro test. All seedlings were evaluated for the presence of characteristic bacterial fruit blotch symptoms at 21 days after emergence. The antibiotics kasugamycin (100 and 200 ug/L), oxytetracycline (10, 100 and 200 ug/L), oxytetracycline+copper sulphate (10, 100 and 200 ug/L), the fungicides captan, carboxin+thiram, benzalkonium chloride, mancozeb+copper oxychloride, metiran, metiran+pyraclostrobin and tebuconazole at doses of 100 and 200 ug/L and peroxyacetic acid at the dose of 300 ug/L showed satisfactory results in the in vitro control of the bacterium. However, no chemical efficiently eradicated the bacterium from the seeds in the \"in vivo\" tests.
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OLIVEIRA, Aldenir de. "Colonização de Acidovorax avenae subsp. citrulli em meloeiro e sobrevivência em restos de cultura e no solo." Universidade Federal Rural de Pernambuco, 2008. http://www.tede2.ufrpe.br:8080/tede2/handle/tede2/6397.

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Submitted by (lucia.rodrigues@ufrpe.br) on 2017-02-16T14:20:21Z No. of bitstreams: 1 Aldenir de Oliveira.pdf: 717930 bytes, checksum: eb22d1cac453b579b74b66da5fd0bc67 (MD5)<br>Made available in DSpace on 2017-02-16T14:20:21Z (GMT). No. of bitstreams: 1 Aldenir de Oliveira.pdf: 717930 bytes, checksum: eb22d1cac453b579b74b66da5fd0bc67 (MD5) Previous issue date: 2008-02-29<br>This dissertation aimed to study: colonization of Acidovorax avenae subsp. citrulli in melons after inoculation of the first pair of true leaves, seeds and hermaphrodite flowers; bacterial survival in fruit and leaf residues incorporated to the soil at 0, 5, 10 and 15 cm depth and in soils without the host plant, under the influence of different soil type, temperature (10, 15, 20, 25, 30 and 35ºC) and humidity (50 and 100% of field capacity). In all studies a mutant resistant to 100 ppm of rifampicin (Aac1Rif) was utilized. Bacterial colonization was detected until 30 days after inoculation in the 10th pair of true leaves, with populations of 3.1 log UFC g-1 of leaf. In the same period, the shoot segment between the 10th and 11th leaf pair showed population of 3.52 log UFC g-1 of shoot. After seed inoculation the pathogen colonized the hypocotyl, roots, cotyledonary leaves, true leaves and shoots, until reach undetectable levels at 27 days after inoculation. Flower colonization by the bacteria was not verified. Aac1Rif was found in fruit and leaf residues at 0, 5 and 10 cm during 21 days and at 15 cm during 14 days. Highest population relative extinction rates (TERP)were presented by fruits on soil surface [0.1464 log (UFC) day -1] and on leaves at 10cm [0.084 log (UFC) day -1]. Aac1Rif survived on seven soil types only during three days and the soil C showed the highest TERP [0.9062 log (UFC) day-1]. Higher concentrations of Na+ and silt as well as higher populations of actinomycetes and Trichoderma correlated to faster extinction of Aac1Rif populations in soil. Generally for all soils the lowers TERP were found at 10 or 15ºC and the higher, at 30 or 35ºC. There was no significant (P=0.05) interaction between soil and humidity, however the T test showed significant difference (P=0.05) between the TERP at 100% [0.6685 log (UFC) day -1] and 50% [0.504591 log (UFC) day -1] of field capacity. Independent of temperature and humidity, Aac1Rif also survived in soil only during three days.<br>Esta dissertação teve como objetivos estudar: colonização de A. avenae subsp. citrulli em meloeiro a partir da inoculação no primeiro par de folhas verdadeiras, sementes e flores hermafroditas; sobrevivência da bactéria em restos de folhas e frutos incorporados ao solo a diferentes profundidades (0, 5, 10 e 15 cm) e em solos na ausência da planta hospedeira, sob a influência de diferentes tipos de solo (sete solos), temperaturas (10, 15, 20, 25, 30 e 35ºC) e umidades (50 e 100% da capacidade de campo). Para os estudos foi utilizado um mutante resistente a 100 ppm de rifampicina (Aac1Rif). Quando a bactéria foi inoculada nas folhas, a colonização foi detectada até os 30 dias, no 10º par de folhas verdadeiras, com população de 3,1 log UFC g-1 de folha. Nesse mesmo período, observou-se a colonização no segmento de ramo compreendido entre o 10º e 11º par de folhas com população de 3,52 log UFC g-1 de ramo. A partir de sementes, a bactéria colonizou o hipocótilo, raízes, folhas cotiledonares, folhas verdadeiras e ramos, até atingir níveis não detectáveis aos 27 dias após a inoculação. Não foi verificada colonização das flores pela bactéria. Aac1Rif foi encontrada em restosde frutos e folhas de meloeiro a 0, 5 e 10 cm durante 21 dias e a 15 cm por 14 dias. As maiores taxas de extinção relativa da população (TERP) ocorreram nos frutos na superfície do solo [0,1464 log (UFC) dia-1] e nas folhas a 10 cm [0,084 log (UFC) dia-1]. Aac1Rif sobreviveu nos sete tipos de solo apenas durante três dias e o solo C apresentou a maior TERP [0,9062 log (UFC) dia-1]. Maiores concentrações de Na+ e silte bem como maiores populações de actinomicetos e Trichoderma estiveram correlacionadas a mais rápida extinção da população de Aac1Rif no solo. Para a maioria dos solos, as menores TERP foram atingidas a 10 ou 15ºC e as maiores, a 30 ou 35ºC. Não houve interação significativa (P=0,05) entre solos e umidade, contudo o teste de T evidenciou diferença significativa (P=0,1) entre as TERP a 100% [0,6685 log (UFC) dia-1] e 50% [0,504591 log (UFC) dia-1] da capacidade de campo. Independente da temperatura e umidade, Aac1Rif também sobreviveu nos solos apenas por três dias.
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SILVA, Valter Alexandre Vieira da. "Levantamento da intensidade da mancha-aquosa em Juazeiro(Bahia) e sobrevivência de Acidovorax avenae subsp. citrulli em meloeiro." Universidade Federal Rural de Pernambuco, 2005. http://www.tede2.ufrpe.br:8080/tede2/handle/tede2/6659.

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Submitted by (lucia.rodrigues@ufrpe.br) on 2017-03-23T15:39:15Z No. of bitstreams: 1 Valter Alexandre Vieira da Silva.pdf: 281801 bytes, checksum: 0bb776ffcea4e506c269e4726b3eafa8 (MD5)<br>Made available in DSpace on 2017-03-23T15:39:15Z (GMT). No. of bitstreams: 1 Valter Alexandre Vieira da Silva.pdf: 281801 bytes, checksum: 0bb776ffcea4e506c269e4726b3eafa8 (MD5) Previous issue date: 2005-02-23<br>Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq<br>One of the main diseases occurring in melon fields in Brazilian Northeastern is the bacterial fruit blotch caused by the bacterium Acidovorax avenae subsp. citrulli. This study aimed to survey the bacterial fruit blotch incidence in 12 melon planting areas in Juazeiro (BA), in two consecutive plantings, and determine the ability of A. avenae subsp. citrulli to survive epiphytically and endophytically on the leaves and roots, and also in the rhizosphere of melon by using a mutant resistant to rifampicin (Aac1Rif). The surveys were conducted to determine the disease incidence and prevalence in 12 commercial melon areas chosen randomly with plants at harvesting time. In each area 25 sub-plots with 100 m2 were marked and 20 fruits evaluated in a diagonal line. In the first survey performed in January 2004, the prevalence of bacterial blotch was 33.33% and there was low incidence in the four areas where the diseaseoccurred, ranging from 0.2 to 4.4%. In the second survey performed in the same areas in April 2004 the disease was not detected. For the survival study on leaves, melon plants with 18-day, grown in greenhouse and field, were sprayed with mutant suspensions at concentrations 3.4 x 102, 3.4 x 103 and 3.4 x 104 CFU mL-1. To determine survival on roots and rhizosphere, seeds of melon type Yellow were sown in soil infested with suspensions of Aac1Rif at 3.4 x 105, 3.4 x 106 and 3.4 x 107 CFU mL-1. At 6-day intervals samples of leaves, roots and rhizosphere soil were collected and processed for isolation on NYDA medium amended with refampicin. Bacterial populations were then determined as UFC g-1 of sample and the data were transformed to log10 for regression analysis. On melon leaves, in greenhouse and field Aac1Rifsurvived epiphytically for 54 days. These epiphytic bacterial populations increasedinitially and decreased after certain time. The final populations were similar in the two conditions and ranged from 103 to 104 CFU g-1 leaf. On the roots and rhizosphere in greenhouse bacterial populations decreased according to the inoculum concentration and at 60 days after planting ranged from 102 to 103 CFU g-1 roots and 101 to 102 CFU g-1 soil. AacRif was not detected as endophytic in leaves or roots of melon.<br>Uma das principais doenças que vem ocorrendo em campos de meloeiro do Nordeste brasileiro é a mancha-aquosa, causada pela bactéria Acidovorax avenae subsp. citrulli. Este trabalho teve por objetivos realizar o levantamento epidemiológico da incidência da mancha-aquosa em 12 áreas comerciais de meloeiro em Juazeiro (BA), em dois plantios sucessivos, e determinar a capacidade de sobrevivência de A. avenae subsp. citrulli como epifítica e endofítica nas folhas e raízes, e na rizosfera de meloeiro, através da utilização de um mutante resistente a rifampicina (Aac1Rif). Levantamentos foram conduzidos para determinar a prevalência e a incidência da mancha-aquosa em 12 áreas comerciais de meloeiro, escolhidas ao acaso, com plantas em estádio de colheita. Em cada área foram demarcadas 25 subparcelas de 100 m2 e avaliados 20 frutos/subparcela ao longo da diagonal. No primeiro levantamento, realizado em janeirode 2004, a prevalência da mancha-aquosa foi de 33,33%, com baixa incidência nas quatro áreas em que foi registrada, variando de 0,2 a 4,4%. No segundo levantamento, realizado nas mesmas áreas em abril de 2004, a doença não foi constatada. Para o estudo de sobrevivência em folhas, meloeiros com 18 dias, cultivados em casa de vegetação e no campo, foram pulverizados com suspensões do mutante nas concentrações (3,4 x 102, 3,4 x 103 e 3,4 x 104 UFC mL-1). Para determinar a sobrevivência em raízes e na rizosfera, sementes de melão Amarelo foram semeadas em solo infestado com suspensões de Aac1Rif a 3,4 x 105, 3,4 x 106 e 3,4 x 107 UFC mL-1. A intervalos de seis dias, amostras de folhas, raízes e solo rizosférico foram coletadas e processadas para isolamento em meio ágar nutritivo-extrato de levedura-dextrosecontendo rifampicina. As populações bacterianas foram determinadas em UFC g-1 deamostra e os dados transformados em log10 para análise de regressão. Nas folhas de meloeiro, em casa de vegetação e campo, o mutante Aac1Rif sobreviveu epifiticamente durante 54 dias, observando-se inicialmente aumento da população bacteriana epifítica, com posterior declínio, sendo as populações finais semelhantes nas duas condições estudadas, com valores variando de 103 a 104 UFC g-1 de folha. Nas raízes e rizosfera, em casa de vegetação, a população bacteriana decresceu variando em função da concentração utilizada até atingir aos 60 dias após o plantio, níveis de 102 a 103 UFC g-1 de raiz e 101 a 102 UFC g-1 de solo. AacRif não foi detectado sobrevivendo endofiticamente em folhas ou raízes de meloeiro.
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Frare, Vanessa Cristina. "Desenvolvimento de um meio semi-seletivo para detecção de Acidovorax avenae subsp. citrulli em sementes de melão (Cucumis melo L.)." Universidade de São Paulo, 2006. http://www.teses.usp.br/teses/disponiveis/11/11135/tde-22022006-145257/.

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Um dos principais fatores limitantes da produção do melão é a ocorrência de doenças. Entre os patógenos mais importantes estão as bactérias, que causam perdas significativas na produção. Causada pela bactéria Acidovorax avenae subsp. citrulli (Aac), a mancha-aquosa-do-melão, também conhecida como mancha-bacteriana-dofruto, é uma doença grave, que tem preocupado produtores do nordeste, sendo que todos os tipos de melão apresentam suscetibilidade ao patógeno. A principal fonte de inóculo para esta bactéria é a semente infectada. Este trabalho teve como objetivo o desenvolvimento de um meio semi-seletivo para detecção e identificação de Acidovorax avenae subsp. citrulli em sementes de melão, para testes de rotina em laboratórios de patologia de sementes. Por meio de testes de fungitoxicidade, antibiogramas qualitativos e quantitativos e testes bioquímicos foi desenvolvido o seguinte meio semiseletivo para a detecção de Aac em sementes de melão: dextrose (5 g/L), NaCl (5 g/L), peptona (5 g/L), KH2PO4 (2 g/L), MgSO4.7H2O (0,2 g/L), vermelho de fenol (0,012 g/L), uréia (25 g/L), agar (17 g/L), benomil (100 mg/L), nistatina (200 mg/L) e amoxicilina (15 mg/L).<br>One of the main limiting factors of the melon production is the occurrence of diseases. The bacteria are among the most important pathogens causing significant losses in the production. Caused by the bacterium Acidovorax avenae subsp. citrulli (Aac), the bacterial-fruit-blotch is a serious disease that affects all types of melon and has worried northeast producers. The main source of inoculum for this bacterium is the infected seed. This work had as objective the development of a semi-selective medium to detect and identify Acidovorax avenae subsp. citrulli in melon seeds, for routine tests in seed pathology laboratories. By means of fungitoxicity tests, qualitative and quantitative antibiograms and biochemical tests the following semi-selective medium to detect Aac in melon seeds was developed: dextrose (5 g/L), NaCl (5 g/L), peptone (5 g/L), KH2PO4 (2 g/L), MgSO4.7H2O (0,2 g/L), phenol red (0,012 g/L), urea (25 g/L), agar (17 g/L), benomil (100 mg/L), nistatina (200 mg/L) and amoxicilin (15 mg/L).
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SILVA, Elias Inácio da. "Levantamento da incidência da mancha-aquosa do melão nos municípios de Mossoró e Baraúna (Rio Grande do Norte, Brasil) e determinação do tamanho da amostra para quantificação da doença." Universidade Federal Rural de Pernambuco, 2002. http://www.tede2.ufrpe.br:8080/tede2/handle/tede2/6558.

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Submitted by (lucia.rodrigues@ufrpe.br) on 2017-03-10T14:34:25Z No. of bitstreams: 1 Elias Inacio da Silva.pdf: 496927 bytes, checksum: d6971c4e240310675bba44bb481b6833 (MD5)<br>Made available in DSpace on 2017-03-10T14:34:25Z (GMT). No. of bitstreams: 1 Elias Inacio da Silva.pdf: 496927 bytes, checksum: d6971c4e240310675bba44bb481b6833 (MD5) Previous issue date: 2002-09-30<br>Melon is one of the most popular cucurbits in the world. The bacterial blotch of melon, caused by Acidovorax avenae subsp. citrulli has caused 40 to 50% losses in the production and made the fruits inappropriate for commercialization in the State of Rio Grande do Norte. This study aimed to survey the bacterial blotch incidence in 18 melon planting areas of the counties Mossoró and Baraúna in Rio Grande do Norte, and to determine the ideal sample size for disease assessment. Prevalence of 100% was reported in the studied fields. The incidence of the bacterial fruit blotch ranged from 4.30 to 47.29%. Incidence levels were under 20% in 50% of the areas and higher than 40% in 17% of the areas. There was significant difference for the variables melon type (P=0.01) and years of melon culture in the area (P=0.05). The t test showed the melon Piel de sapo was more susceptible than the yellow type and the disease incidence in areas with less than 10 years of melon cropping was higher than those with more than20 years. However, there were no significant differences (P=0.05) for disease incidence averages among areas planted with corn (18%) and corn plus other grasses (20%) between melon seasons. Using the incidence data from 18 areas as pilot-samples, the sample size for disease assessment was determined according to the mean variability coefficient. The Pierson test showed no significant correlations (P=0.05) between the levels of disease incidence and sample sizes. Based on the data obtained andconsidering 10% of acceptable error, it is recommended that in future surveys of melon bacterial blotch incidence in fields in Rio Grande do Norte, a sample composed by 12 subsamples comprising 100 m2/ha and 20 fruit evaluated per subsample must be analyzed.<br>O meloeiro é uma das cucurbitáceas mais populares do mundo. A mancha-aquosa do melão, causada pela bactéria Acidovorax avenae subsp. citrulli, tem causado perdas de 40 a 50% na produção e depreciação no valor comercial do fruto no estado do Rio Grande do Norte. Os objetivos deste estudo foram realizar o levantamento da incidência da mancha-aquosa em 18 áreas de plantio de meloeiro dos municípios Mossoró e Baraúna, no Rio Grande do Norte e determinar o tamanho ideal das amostras para quantificação da incidência da doença no campo. Foi registrada a prevalência da doença em 100% dos campos. A incidência da mancha-aquosa variou entre 4,30% e 47,29%. Em 50% das áreas foram constatados níveis de incidência da doença inferiores a 20%, enquanto em 17% foram registrados valores superiores a 40%. Houve diferença significativa na incidência da doença em relação aos tipos de melão (P=0,01) e aos anosde plantio na mesma área (P=0,05). Pelo teste t o melão tipo Pele de sapo foi mais suscetível que o Amarelo e a incidência da doença foi maior em áreas com menos de 10 anos de plantio em relação àquelas com mais de 20 anos de plantio dessa cucurbitácea. Não foram constatadas diferenças significativas (P=0,05) pelo teste t, para médias de incidência da mancha-aquosa entre áreas com plantio de milho (18%) e milho com outras gramíneas (20%) na entressafra. Utilizando os dados de incidência da doença nas 18 áreas como amostragens-piloto, o tamanho das amostras para quantificação da doença foi determinado com base no coeficiente de variação da média. O teste de Pierson não constatou correlações significativas (P=0,05) entre os níveis de incidência da doença e o tamanho das amostras. Considerando os resultados obtidos e um erroaceitável de 10%, em futuros levantamentos da incidência da mancha-aquosa em plantios de meloeiro no Rio Grande do Norte, recomenda-se a utilização de uma amostra de 12 subparcelas com 100 m2/ha e 20 frutos avaliados por subparcela
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Cabral, Clêidio da Paz. "Eficiência de indutores de resistência no controle da mancha-aquosa em meloeiro." Universidade Federal Rural de Pernambuco, 2009. http://www.tede2.ufrpe.br:8080/tede2/handle/tede2/6451.

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Submitted by (lucia.rodrigues@ufrpe.br) on 2017-02-20T12:00:31Z No. of bitstreams: 1 Cleidio da Paz Cabral.pdf: 307888 bytes, checksum: 839bdb2400d1038e8ac0b20e10e22398 (MD5)<br>Made available in DSpace on 2017-02-20T12:00:31Z (GMT). No. of bitstreams: 1 Cleidio da Paz Cabral.pdf: 307888 bytes, checksum: 839bdb2400d1038e8ac0b20e10e22398 (MD5) Previous issue date: 2009-02-27<br>Bacterial fruit blotch caused by Acidovorax avenae subsp. citrulli (Schaad et al.) Willems et al. is an important melon (Cucumis melo L.) disease which causes fruit value reduction and high production losses reaching 100% in rainy season. At the present there are not effective methods of controlling this disease in Brazil, and therefore research for finding alternative control methods is essential. It was studied the effect of resistance inducers for controlling bacterial fruit blotch in ‘yellow’ melon (AF4945) and ‘Pele de Sapo’ (Nilo), evaluating different application periods (10 and 15 days after plant emergency), inducer dosages (acibenzolar-S-metil- ASM - 25; 50 and 75 mg a.i. L-1; Agro-Mos® - 0.5; 1.0 and 1.5 c.p. L-1; Ecolifeâ - 2; 3 and 4 mL c.p. L-1) and plant physiology in soil supplemented or not with NPK observing plant height, fresh and dry shoot biomass and enzymatic activity. The best period for inducers application was 10 days after plant emergency. For both genotypes, ASM (50 mg a.i. L-1) and Ecolifeâ (3 mL c.p. L-1) increased incubation period until 12 and 8 days; and reduced disease incidence by 87.5 and 60%; disease index by 95.7 and 88%; and area under disease progress curve by 93.7 and 74.5%. The two inducers independent of NPK level reduced plant height, fresh and dry shoot biomass until 24.5; 41.4 and 34.2%, respectively. Onlythe peroxidase activity was detected at 5, 10 and 45 days after inducer application. Although ASM (50 mg i.a. L-1) and Ecolifeâ (3 mL p.c. L-1) had affected plant development they significantly reduced bacterial fruit blotch in both melon types, ‘yellow’ and ‘Pele de Sapo’, and should be used for the integrated management of this disease.<br>A mancha-aquosa causada pela bactéria Acidovorax avenae subsp. citrulli (Schaad et al.) Willems et al. é uma importante doença para a cultura do meloeiro ocasionando depreciação do valor comercial do fruto e grandes perdas na produção, que podem atingir até 100% em períodos chuvosos. Não se dispõe até o momento de medidas efetivas de controle dessa doença no Brasil o que torna imprescindível à realização de pesquisas que visem à obtenção de métodos alternativos de controle. Foi estudado o efeito de indutores de resistência no controle da mancha-aquosa em meloeiros Amarelo (AF4945) e Pele de Sapo (Nilo), em diferentes épocas de aplicação (10 e 15 dias após a emergência das plântulas) e dosagens (acibenzolar-S-metil – ASM - 25; 50 e 75 mg i.a. L-1; Agro-Mos® - 0,5; 1,0 e 1,5 p.c. L-1; Ecolifeâ - 2; 3 e 4 mL p.c. L-1); e na fisiologia da planta em solo com ou sem NPK, avaliando-se altura (AL), biomassa fresca e seca da parte aérea (BFPA, BSPA), e atividade enzimática. A melhor época para aplicação dos indutores foi dez dias após a emergência das plântulas. Considerando os dois genótipos, ASM (50 mg i.a. L-1) e Ecolifeâ (3 mL p.c. L-1) elevaram o período de incubação em até 12,6 e 8 dias e reduziram a incidência da doença em 87,5 e 60%; índice de doença em 95,7 e 88%; e área abaixo da curva de progresso da doença em 93,7 e 74,5%, respectivamente. Esses dois indutores, independente do nível de NPK, reduziram a AL,BFPA e/ou BSPA em até 24,5; 41,4 e 34,2%, respectivamente. Apenas a atividade de peroxidase foi detectada aos 5, 10 e 45 dias após a aplicação dos indutores. ASM (50 mg i.a. L-1) e Ecolifeâ (3 mL p.c. L-1) mesmo tento afetado o desenvolvimento de plantas de meloeiro, apresentaram redução significativa da intensidade da mancha aquosa tanto em meloeiro tipo Amarelo quanto Pele de Sapo, podendo ser inseridos no manejo integrado da mancha-aquosa.
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Guo, Yu-Wun, and 郭昱彣. "Characterization of HrpY and HrpW proteins in Acidovorax avenae subsp. avenae CH12." Thesis, 2013. http://ndltd.ncl.edu.tw/handle/30213248148798845526.

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碩士<br>國立中興大學<br>生物科技學研究所<br>101<br>Acidovorax avenae subsp. avenae (Aaa) CH12 causes bacterial leaf stripe disease on corn. The bacterial proteins secreted via type III secretion system (T3SS) which is encoded by a hrp/hrc cluster are the major virulence factors to cause diseases in host plants or elicit the hypersensitive response (HR) on nonhost plants. The genes residing the region between hrcT and GALA genes in hrp/hrc cluster of A. avenae isolated from different host are variable. To elucidate whether the diversity is involved in virulence or not, the two ORFs annotated in this region from AaaCH12 were characterized in this study. These two ORFs were named as hrpY and hrpW based on their amino acid sequence shared 99% and 98% identities with those from AaaN1141, respectively. Moreover, HrpY and HrpW from AaaN1141 strain were predicted to be harpin proteins, which are glycine rich and thermal stable, could be an HR elicitor, have no N-terminal signal peptide and secreted via T3SS. In this study, HrpY and HrpW proteins from AaaCH12 are glycine rich and have no N-terminal signal peptide. The crude extracted or purified HrpY-His6 and HrpW-His6 proteins which were overexpressed by T7 RNA polymerase system could elicit the HR on Nicotiana tabacum with or without heat treatment, suggesting that HrpY-His6 and HrpW-His6 proteins are thermal stable. A pectate lyase domain residing in C-terminus of HrpW from AaaCH12 has pectate lyase activity based on the pectate lyase assay using polyglacturonic acid as a substrate. Gel filtration assay showed both purified HrpY-His6 and HrpW-His6 could be detected as about 2000 kDa in size. HrpY-His6 protein was applied to raise a polyclonal antibody in rabbit for Western blot analysis. Under T3SS inducing condition by cultured in modified XVM2 medium, both HrpY and HrpW could be secreted in wild type but not in T3SS-deficient hrcV mutant of AaaCH12, suggesting that HrpY and HrpW are secreted via T3SS. The results strongly suggest that HrpY and HrpW are harpin-like proteins. In addition, hrpY, hrpW and hrpYW deletion mutants were generated by using unmarked gene deletion mutagenesis. The hrpY, hrpW and hrpYW mutants elicited the delay HR on N. tabacum. The hrpY and hrpYW deletion mutants also reduced the virulence of Aaa CH12 by decreasing disease lesion length and bacterial growth in corn leaves, but the hrpW deletion mutant did not. Additionally, the hrpYW double mutant is significantly less virulent than hrpY, suggesting that HrpYW proteins have an additive effect in virulence on corns.
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Chen, Man-Shu, and 陳蔓抒. "Genetic diversity of Acidovorax avenae subsp. citrulli in Taiwan." Thesis, 2003. http://ndltd.ncl.edu.tw/handle/81456813293047487072.

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碩士<br>國立中興大學<br>植物病理學系<br>91<br>Bacterial fruit blotch caused by Acidovorax avenae subsp. citrulli (Acc) has been a destructive and widespread disease on watermelon, muskmelon and bitter gourd in Taiwan since first observed in 1994. The genetic diversity of Aac strains from different hosts was studied by using the genomic fingerprinting techniques and Biolog system to understand the population structure of Aac in Taiwan and its relationship with strains from other countries. The cluster analysis based on the amplified DNA profiles by multiplex PCR using selected four random primers revealed two major clusters, R1 and R2, among 64 strains of Aac from Taiwan. The majority (88%) of strains from watermelon and 18.8% of strains from muskmelon were grouped in cluster R1, and 12% of strains from watermelon, majority (81.2%) of strains from muskmelon, and all strains from bitter gourd were in cluster R2. The analysis of combined genomic fingerprints of BOX- and ERIC-PCR also grouped strains from Taiwan into two clusters, BE1 and BE2. The majority (78.9%) of strains from watermelon and 14.8% of strains from muskmelon were in cluster BE1, and 21.1% of strains from watermelon, majority (85.2%) of strains from muskmelon and all strains from bitter gourd were in cluster BE2. When DNAs of nine strains from other countries were compared, three watermelon strains and one pumpkin strain from U.S.A. and two cantaloupe strains from Australia were grouped in cluster BE2, however, one watermelon strain and two citron strains from U.S.A. were distinct and could be grouped into third cluster, BE3. The analysis of the Biolog system revealed three cluster, B1, B2 and B3. The majority (93%) of watermelon strains and 8.3% of muskmelon strains were in cluster B1, and 7% of watermelon strains, majority (91.7%) of muskmelon strains were in cluster B2. Among the four biter gourd strains tested, three were in cluster B2 and one was in cluster B3. The above results showed that Aac strains from Taiwan could be differentiated into two major clusters. Cluster 1 consisted of strains mainly from watermelon and cluster 2 is composed of strains mainly from muskmelon and bitter gourd. Among strains form U.S.A. and Australia tested, most were in cluster 2. The inoculation tests did not show the relationship between the genetic groups and their pathogenicity to the hosts.
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Yeh, Guan-Lin, and 葉冠麟. "Characterization of the Acidovorax avenae subsp. citrulli HrpW protein." Thesis, 2008. http://ndltd.ncl.edu.tw/handle/70372971416229872700.

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碩士<br>國立中興大學<br>生物科技學研究所<br>96<br>Bacterial fruit blotch (including watermelon, muskmelon and bitter gourd) is a bacterial plant disease caused by Acidovorax avenae subsp. citrulli (Aac) . HrpW protein in Pseudomonas syringae pv. tomato is secreted by type III secretion system (TTSS). In previous study, a hrpW gene was cloned from Aac156 isolated from bitter gourd. The feature of this gene product similar to the HrpW in Pseudomonas syringae pv. tomato is a harpin-like protein with the amino sequence consisting of the harpin domain in N-termini and the pectate lyase domain in C-termini, and can induce hypersensitive response on nonhost leaves. However, hrpW gene doesn’t exist in every strain of A. avenae subsp. citrulli. HrcV is a component of TTSS. In plant pathogenic bacteria ,the TTSS and its secreting proteins are the major virulence factors. So the expression of the TTSS in the medium will benefit for the studies on these proteins and their pathogenic mechanism. The gene composition encoding TTSS in Aac is similar to those in Xanthomonas campestris and Ralstonia solanacearum, but the XVM2 medium which was commonly used for inducing the TTSS in Xanthomonas campestris was not suitable for inducing the TTSS in Aac. In previous study, it was found that the XVM2 medium containing 1mM Tween80 could activate the TTSS promoter of Aac by using a plasmid carrying a TTSS promoter from Aac148 fused to luxA-luxB reporter gene. In this study, the distribution and homology of hrpW in Aac was conferred by Southern blotting with probes prepared from the two domains of hrpW. HrpW protein was overexpressed and was applied to raise polyclonal antibody in rabbit. Based on western blot analysis with HrpW antibody, expression of HrpW in Aac 156 was observed in XVM2 medium and XVM2 medium containing 1mM Tween80 and the protein was detected in supernatant fraction. By construction of hrcV mutant, the secretion of proteins can be investigated whether it is TTSS dependent.
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Chou, Ying-Chen, and 周盈甄. "The development of detection techniques of Acidovorax avenae subsp. citrulli." Thesis, 2009. http://ndltd.ncl.edu.tw/handle/40544139646501851286.

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Yang, Wen-Jen, and 楊文仁. "Detection of Acidovorax avenae subsp. citrulli in watermelon and melon seeds." Thesis, 2001. http://ndltd.ncl.edu.tw/handle/27632034126776115113.

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碩士<br>國立中興大學<br>植物病理學系<br>89<br>The objective of this study was to investigate the detection of Acidovorax avenae subsp. citrulli (Aac) in watermelon and melon seeds. For increasing the efficiency of detection, the infested seeds were incubated in an improved semiselective medium WFB68 containing 25 ppm cefoperazone, 10 ppm ceftriaxane and 25 ppm piperacillin. After enrichment with this medium, enzyme-linked immunosorbent assay(ELISA), direct polymerase chain rection (direct PCR) and immunomagnetic separation and polymerase chain rection(IMS-PCR) were compared for their efficiencies of detection. Detection by direct PCR for artificially infested seed samples(1 and 5% infestation) using 100 seeds as an assay sample were all positive, however, only 50% samples were positive using 200 seeds(0.5 and 2.5% infestation)as an assay sample. With 1000 seeds as an assay sample, Aac on watermelon seed samples (0.1 and 0.5% infestation) were not detected by ELISA and direct PCR, but 80% detection rate (positive tests per total tests) were obtained by IMS-PCR; Aac on melon seed samples(0.1 and 0.5% infestation) was also not detected by ELISA, but 20 and 40% detection rates were obtained by direct PCR, and 80 and 100% detection rates were obtained by IMS-PCR for seed samples with 0.1 and 0.5% infestation, respectively. When three naturally infested seed lots ( two of watermelon and one of melon seeds) were assayed, the detection rates by IMS-PCR(80 ―100% ) were higher than those by direct PCR(22.5 — 62.5%) and ELISA (0 ― 40%). Detection by IMS-PCR was not affected by seed treament with agrochemicals. The primer pair SL1/SR1 used in PCR was aslo specific for all nine strains of A. avenae subsp. citrulli from foreign countries, but the other primer pair L2/R2 only amplified a distinct DNA band from strains belonging to group II. The results indicate that the IMS-PCR could be potentially used as a seed-detection assay for A. avenae subsp. citrulli on watermelon and melon seeds.
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Tang, Chin Jen, and 唐致仁. "Studies on bacterial fruit blotch of watermelon caused by Acidovorax avenae subsp. citrulli." Thesis, 1997. http://ndltd.ncl.edu.tw/handle/39147374711088076522.

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碩士<br>國立中興大學<br>植物病理學系<br>85<br>The bacterial fruit blotch of watermelon, a newly found disease, occurred in some fields of southern and central parts of Taiwan. In the field, the disease appeared after overhead irrigation or a rainy period. The most pronounced symptom of the disease was the appearence of water-soaked blotches on the upper surface of fruit. The blotch may expand to cover most area of the upper surface, however, the lesions rarely extended into the flesh of the fruit. Water-soaked spots which became dark brown and then necrotic also formed on cotyledons and true leaves of the seedlings. Lesions on leaves of mature plants were generally inconspicuous, light brown to reddish brown in color and often spread along the midrib of the leaves. Based on physiological and biochemical characteristics and inoculation tests, the causal organism was identified as Acidovorax avenae subsp. citrulli. In the field, the bacterium affected watermelon and muskmelon. Artificial inoculation showed that the bacterium also was pathogenic to orie ntal pickling melon, cucumber, bitter gourd, sponge gourd, bottle gourd, pumpkin and wax gourd. Generally,strains of A. avenae subsp. citrulli from watermelon were similarly virulent to watermelon and muskmelon, however, the strains from muskmelon were more virulent to muskmelon than to watermelon. The disease was more severe on younger fruits ( age at 2-3 days after fruit setting ) than on older fruits ( age at 5-7 days or 7-14 days after fruit setting ) when spray inoculated with the pathogen; the diseas e severity was also greater on inoculated fruits covered with plastic bags than those without covering. Seeds collected from the diseased fruits were found to carry the pathogen with the infestation rate of 54.3 - 72.6%. Seed assays by the grow-out test revealed that 15 of 16 cultivars of watermelon tested ( one seed lot for each cultivar ) were infested with Acidovorax avenae subsp. citrulli, and with some cultivars, more than 20% of seeds were infested. The 15 test cultivars of watermelon differed in thei r susceptibility to the bacterium when evaluated on seedlings by spray inoculation and seed-soaking inoculation methods. Among six seed treatments , treatment with 1% HCl was most effective in reducing the disease incidence, followed by treatment with 0.1M acidified ZnSO4 ( 38℃, pH 2.8 ), whereas hot water treatment ( 50℃, 20 min ) was not effective.
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Chang-Chien, Ju, and 張簡如. "Analysis on biological functions of Acidovorax avenae subsp. citrulli harpin-like protein (Aave_0457)." Thesis, 2009. http://ndltd.ncl.edu.tw/handle/69933066215600103742.

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碩士<br>國立中興大學<br>生物科技學研究所<br>97<br>Acidovorax avenae subsp. citrulli (Aac), which causes bacterial fruit blotch disease (BFB) in Cucurbitaceae, is a gram-negative, rod-shaped plant pathogen. The ability of Aac to grow and cause diseases in plants is dependent on the injection of multiple effector proteins into plant cells via the type III secretion system (T3SS), encoded by hrp/hrc genes cluster. Effectors contribute to pathogenesis by suppressing plant defenses and promoting disease symptom in host plants. In nonhost plants, it can elicit the hypersensitive response (HR). However, the DNA sequence between GALA and hrcT in hrp/hrc genes cluster of Aac strains from different hosts are variable. For example, Aac156 isolated from bitter gourd contains hrpW and transposase in this region, but Aac148 and Aac31 isolated from watermelon contain two putative PE_PGRS genes, named Aave_0456 and Aave_0457. The recombinant Aave_0457 protein was overexpressed in E. coli BL21 (λDE3) and applied to raise polyclonal antibody in rabbit. According to western blotting results, Aave_0457 was induced in XVM2 minimal medium and secreted into medium. The amino acid sequence of Aave_0457, which has no N-terminal signal peptide, is homologous to the harpin domain of HrpW in Aac156. The recombinant Aave_0457 protein overexpressed in E. coli BL21 (λDE3) is heat-stable and can elicit the HR in nonhost plant after 24 hours post inoculation, suggesting that Aave_0457 is a harpin-like protein. Aave_0457 mutant elicited the delayed HR on nonhost tobacco leaves, and reduced the disease incidence and disease severity in host plant, but the growth of Aave0457 mutant and wild type was no significant difference in Cucurbitaceae. Furthermore, the non-coding region between Aave_0457 and Aave_0456 might be promoter-less, suggesting that hrcT, Aave_0456 and Aave_0457 belong to an operon.
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Sung, Ping-Feng, and 宋秉峰. "The Polymerase Chain Reaction Technique for Identification and Detection of Acidovorax avenae subsp. citrulli." Thesis, 1999. http://ndltd.ncl.edu.tw/handle/48053765049508984991.

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碩士<br>國立中興大學<br>植物病理學系<br>87<br>Sixty different random primers were used to find specific DNA fragments of Acidovorax avenae subsp. citrulli by using random amplified polymorphic DNA (RAPD). A specific DNA fragment (about 300bp in size) of A. avenae subsp. citrulli amplified by the primer OPD-11 was cloned into the pCRII-TOPO vector, and sequenced to design primer pairs SL1/SR1. The primer pairs could amplify a distinct band of 194 bp that was specific to A. avenae subsp. citrulli isolated from watermelon, muskmelon and bitter gourd by polymerase chain reaction (PCR). And there was no any DNA fragment amplified with primer pairs SL1/SR1 from any other tested bacteria belonging to 16 species and 7 genus. The minimum amount of DNA from A. avenae subsp. citrulli that could be amplified by PCR was 100pg. Sensitivity of PCR for detection of cells of A. avenae subsp. citrulli with primer pairs SL1/SR1 was 1.2’102 cfu, and could be raised to 1.2 cfu by re-amplification of diluted standard PCR products (Second PCR). Nontarget bacteria WS, MS1 and MS2 did not affect the efficiency of specific amplification of A. avenae subsp. citrulli in PCR assay with primer pairs SL1/SR1. PCR technique with primer pairs SL1/SR1 identifies cultures of A. avenae subsp. citrulli within 4hours and detected the bacterium in diseased tissues and infested seeds of watermelon and muskmelon. The results indicated that the primer pairs SL1/SR1 could be a useful tool for rapid identification and detection of A. avenae subsp. citrulli in diseased plant tissues infected with A. avenae subsp. citrulli, and could also be potentially used for detection of contaminated seeds.
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Chen, Yi-Jyun, and 陳羿君. "Characterization of HrpWAaca protein of Acidovorax avenae subsp. cattleyae OAC1 and improvement of hrp gene inducing medium for Acidovorax spp." Thesis, 2010. http://ndltd.ncl.edu.tw/handle/70339702386197467840.

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碩士<br>國立中興大學<br>生物科技學研究所<br>98<br>Acidovorax avenae subsp. cattleyae (Aaca) OAC1 causes a brown spot disease on Cattleyae, while A. a. subsp. citrulli (Aac) causes Bacterial fruit blotch disease (BFB) on cucurbitaceae. These bacteria causing diseases in host plants or eliciting hypersensitive response (HR) on nonhost plants depend on type III secretion system (T3SS), encoded by hrp/hrc gene cluster. In this study, we analyzed the biological function of hrpWAaca of Aaca by loss-of-function strategy. The hrpWAaca mutant elicited a delayed HR on Nicotiana tabacum, but still maintained the same virulence as its wild type on its host Oncidium and Phalaenopsis. Hence, HrpWAaca might be not play an essential role in pathogenicity. The purified or crude extracted HrpWAaca-His6 proteins could elicit HR with or without heat treatment. Moreover, HrpWAaca-His6 protein contains 17.58% glycine but no cysteine, and HrpWAaca could be secreted into bacterial milieu under T3SS inducing condition. Therefore, HrpWAaca is a harpin-like protein. The Pel domain in C-terminus of HrpWAaca has the pectate lyase activity based on the pel assay with polyglacturonic acid as a substrate. Besides, Gel filtration assay showed the purified HrpWAaca-His6 could be detected as dimer (120 kDa)、hexamer (400 kDa) and oligomer (>2000 kDa) in size. To control the tomato bacterial spot caused by Xanthomonas vesicatoria, HrpWAaca-His6 solution was sprayed onto tomato leaves 1 or 4 days before inoculated with X. vesicatoria. The result showed that HrpWAaca-His6 could reduce bacterial spot severity after 4 days harpin treatment. To improve the T3SS induction medium for Aac, extracts of watermelon suspension cell and loofah were total using gfp as a reporter gene under the control of hrcC promoter. The results indicated that both extracts significantly improved induction efficiency of the XVM2 medium.
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Fang, Chun-wei, and 方俊偉. "Studies on detection of Acidovorax avenae subsp. citrulli by using PCR and RT-PCR techniques." Thesis, 2001. http://ndltd.ncl.edu.tw/handle/91261847054354621602.

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碩士<br>國立高雄師範大學<br>生物科學研究所<br>90<br>Three different primers [ one upstream primer ( Aap5 ) and two downstream primers ( Aap6 and Aap61 ) ] selected from Acidovorax avenae-associated nucleotide sequences in Genbank database of National Center for Biotechnology Information ( NCBI ) on internet were used to clone specific cDNA fragments of A. avenae subsp. citrulli Aa31 by using RT-PCR. cDNA products of ~1600 bp with Aap5/Aap6 primers and ~900 bp with Aap5/Aap61 primers were obtained by using RT-PCR. For further nucleic acid sequence analysis, the two cDNA fragments were amplified with PCR using elongated primers. The ~900 bp cDNA fragment was used for sequencing of nucleic acid sequence. This 887 bp nucleotides sequence of cDNA fragment were analyzed. Six open reading frames were found, designed as Ama01, Ama02, Ama03, Ama04, Ama05, and Ama06, respectively. Among them, Ama04 had 40% identities and 61% positives with transcriptional regulators, transcriptional repressors, and cell division control proteins of some bacteria and phages by searching with NCBI Genbank database. The others had no significant identities. Therefore, Ama04 might play an important physiological role in Aa31. A primer pair L1/R1 was also designed from the nucleic acid sequence of 887 bp cDNA fragment for detection of Aa31. 14 strains of A. avenae subsp. citrulli and 8 kinds of other plant pathogenic bacteria were tested by PCR amplification with primer pair L1/R1. The result indicated that a distinct band of 360 bp was amplified with primer pair L1/R1 from Aa31, a less obvious band of 360 bp was amplified with primer pair L1/R1 from Aa3, Aa27, Aa29, Aa30, Aa32, Aa33 of A. avenae subsp. citrulli, and no amplification from watermelon strains Aa9, Aa24, Aa60 of A. avenae subsp. citrulli and other plant pathogenic bacteria. The minimum amount of DNA from A. avenae subsp. citrulli Aa31 that could be amplified by PCR was 100 pg. Sensitivity of PCR and RT-PCR for detection of cells of A. avenae subsp. citrulli Aa31 with primer pair L1/R1 were 103 cfu and 102 cfu, respectively. From the result, PCR and RT-PCR were able to detect A. avenae subsp. citrull effectively. Genes of the 887 bp cDNA fragment could be potentially used for the further research of A. avenae subsp. citrulli.
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Guo, Song-Yi, and 郭松翊. "Cloning and Overexpression of Specific Antigens of Acidovorax avenae subsp. citrulli and the Development of Serological Detection Techniques." Thesis, 2011. http://ndltd.ncl.edu.tw/handle/80435105426122663570.

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碩士<br>國立中興大學<br>生物科技學研究所<br>99<br>Bacterial fruit blotch (BFB) of watermelon, caused by Acidovorax avenae subsp. citrulli (Aac), can lead to severe fruit losses reaching 100% and cause severe agricultural damages. Aac-contaminated seeds are the primary source of initial inoculum of BFB epidemics that can be controlled by preliminary seed health inspections. However, current methods for the detection of Aac in cucurbit seeds need to be improved in specificity, sensitivity, and convenience in operation. The objective of this study is to establish internationally-accepted standard operation protocol based on serological techniques for the detection of Aac in seeds to prevent the outbreak of BFB and reduce the economical losses. In this study, four approaches were adopted to prepare the serological tools for the specific detection of Aac. Firstly, the polyclonal antiserum was raised against whole particles of Aac and used in two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) to analyze the proteomes of various subspecies of A. avenae to identify Aac-specific antigens. Secondly, PilA protein, the major component of pili required for virulence of Aac, was selected for overexpression in Escherichia coli and antibody production. Thirdly, phage display technique was used to identify the peptides that have specific affinity to Aac. The fourth, approach was to employ monoclonal antibody for the detection of specific antigens of Aac. The preparation of antiserum against whole Aac particle has been accomplished and can be used to differentiate different bacterial strains by western blot with the sensitivity of 105 cfu/ml. Significant differences among the proteomes of different A avenae from different hosts were revealed also by western blot analysis. A 20kDa protein specifically expressed in Aac31 was identified for further analysis. Two phage isolates were identified to exhibit high affinity to A. avenae subspecies. Antisera from two different mice that showed specificity to Aac have been obtained and could provide useful materials for further production of monoclonal antibodies. It is expected that standard operation protocols for the detection of Aac in cucurbit seeds could be established based on the serological tools developed in this study. In the future, the potential pathogenicity-related factors will be analyzed using these serological tools to investigate mechanisms of Aac and to develop efficient management strategies of BFB in cucurbits.
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Hsu, Tsui-Fang, and 許粹芳. "Regulation of the type Ⅲ secretion system mediated by hrpG and hrpX genes in Acidovorax avenae subsp. citrulli." Thesis, 2006. http://ndltd.ncl.edu.tw/handle/73247555777453016157.

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碩士<br>國立中興大學<br>生物科技學研究所<br>94<br>Bacterial fruit blotch of watermelon is caused by Acidovorax avenae subsp. citrulli (Aac). Type III secretion system (TTSS) gene clusters of Aac are very similar to Xanthomonas spp. and Ralstonia solanacearum. The regulatory proteins, HrpG and HrpX, also share high homology with those of Xanthomonas spp.. In Xanthomonas spp, HrpG belongs to an OmpR family in a two-component regulation system, and it functions as an activator. HrpX is a highly conserved member of the AraC family-like transcriptional regulatory protein and can be activated by HrpG. When HrpX is activated, it regulates the expression of downstream genes, whose upstream sequences reveal a conserved sequence motif, designated as the PIP box, and then the TTSS is developed. The synthetic medium XVM2 was used to induce expression of TTSS of Xanthomonas spp. According to previous studies, XVM2 medium can not induce the expression of aac1 gene of Aac, indicating that XVM2 is not a suitable medium for inducing TTSS expression in Aac. In this study, the luciferase genes, luxA and luxB were used as reporter genes to monitor the activites of hrpX, hrpG and hrcC promoters and to detect TTSS expression in Aac. Preliminary results show that the luxA-luxB was expressed in Escherichia coli, indicating that HrpG activates hrpX promoter and HrpX activates hrcC promoter. In addition, the hrpG mutant and hrpX mutant could not cause HR on tobacco leaf. To search for the recipes of Aac TTSS induction medium, I replaced the carbon source of XVM2 with varied chemicals. I found that the medium containing Tween 80 can induce hrcC promoter in Aac.
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Cheng, Ming-Lung, and 鄭明龍. "Cloning of a type III secretion system of Acidovorax avenae subsp. citrulli and involvement of GALA protein in pathogenicity." Thesis, 2005. http://ndltd.ncl.edu.tw/handle/96731103528896529735.

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碩士<br>國立中興大學<br>生物科技學研究所<br>93<br>Bacterial fruit blotch of watermelon caused by Acidovorax avenae subsp. citrulli (Aac) has been widely spread on many watermelon-cultivating areas since it was first reported in Florida, USA in 1989. Since bacterial fruit blotch of watermelon was reported in Taiwan in 1994, the melon and bitter melon were also infected by Aac. Aac is Gram-negative, rod sharp with a single polar flagellium and seed-born. Aac infects seedling and fruit of watermelon and produces the water-soaking symptom at peel. Type III secretion system (TTSS) of plant pathogenic bacteria is necessary for delivering effector proteins to their host cells. TTSS is encoded by hrp (hypersensitive response and pathogenicity) genes and nine high conserved hrc (hrp conserved gene) genes, and its machinery is a needle-like structure. Because the crops infected by Aac are very important and little is known of molecular mechanism of Aac on pathogenicity, the aim of this study is to clone the genes coding TTSS and the effectors. First, the multiple alignment of amino acid sequences of HrcV proteins from five different bacterial strains were performed. According to highly conserved amino acid sequence, two degenerated primers were designed and used in PCR to clone the partial hrcV gene sequence from Aac as a probe. Subsequently, colony hybridization was performed to clone TTSS of Aac from genomic DNA library constructed using Copy ControlTM BAC Cloning Kit. This TTSS gene cluster contains 29 predicted ORFs, including nine hrc genes, seven hrp genes, three hpa genes (hrp association gene) and ten ORFs. Among four ORFs, the genes coding for an Xanthomonas campestris pv. vesicatoria ORF1, Aac1 (GALA) protein and Aac2 are located between hrcN and hrcT gene. Two genes located at the left side of gene cluster are predicted to be involved in bacterial virulence and two genes located at right side of the cluster are involved in biosynthesis of Trp, Phe and Tyr. The arrangement and amino acid sequences of TTSS gene cluster are more similar to xanthomonads and Ralstonia solanacearum. The regulators protein, HrpG and HrpX, are similar to xanthomonads and the putative PIP box-like motif is found at promoter region of among 11 genes. S嶵astien group (2004) reported that the expression of GALA protein of R. solanacearum is regulated by HrpB and the protein is translocated into its host plant but does not interfere the pathogenicity. In this study the aac1 mutant still caused HR on tobacco leaf and GALA-cyaA fusion protein is not secreted under XVM2 hrp inducing medium. Aac148 carrying GALA-cyaA fusion protein also doesn't cause accumulation of cAMP in tobacco and watermelon leaf, indicating that AAC1 protein could not be secreted to cytoplasmic of plant cells and it may be not an effector.
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"Desenvolvimento de um meio semi-seletivo para detecção de Acidovorax avenae subsp. citrulli em sementes de melão (Cucumis melo L.)." Tese, Biblioteca Digital de Teses e Dissertações da USP, 2006. http://www.teses.usp.br/teses/disponiveis/11/11135/tde-22022006-145257/.

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Lessl, Jason Thomas. "The role of bacterial motility in watermelon blossom colonization and seed infestation by Acidovorax avenae subsp. citrulli, causal agent of bacterial fruit blotch." 2004. http://purl.galileo.usg.edu/uga%5Fetd/lessl%5Fjason%5Ft%5F200412%5Fms.

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