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Sample records for em xylella fastidiosa

  1. Efeito da Xylella fastidiosa em cafeeiros em diferentes regiões edafoclimáticas Effect of Xylella fastidiosa in coffee plants at different edaphoclimatic regions

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    Rachel Benetti Queiroz-Voltan

    2005-01-01

    Full Text Available A bactéria Xylella fastidiosa vem causando problemas para a cafeicultura, uma vez que sua presença, associada a diversos fatores de estresse, provoca um decréscimo na produção devido à diminuição no número e tamanho dos frutos e à morte de alguns ramos. Este trabalho objetiva avaliar o efeito da X. fastidiosa sobre cultivares de Coffea arabica (enxertados ou não através da quantificação da proporção de vasos do xilema obstruídos pela bactéria, nas diferentes partes da planta e entre ramos com e sem sintoma da doença, em experimentos desenvolvidos em diferentes regiões edafoclimáticas. Avaliou-se também a distribuição das classes de infecção nas diferentes partes da planta nos materiais genéticos estudados. Os experimentos foram instalados em 1986 em Mococa e Garça (SP e as amostras para o estudo anatômico, retiradas em abril de 1998 e 2000 (período de estresse hídrico, respectivamente, das plantas de cafeeiros dessas áreas. Na região de Mococa, observou-se que a nervura principal e o pecíolo foram os tecidos com proporção maior de vasos do xilema obstruídos pela X. fastidiosa; na região de Garça, foram o pecíolo e o caule. Não houve diferenças significativas na obstrução de elementos de vaso do xilema do cafeeiro ocasionado pela bactéria entre as duas regiões estudadas. Não houve tolerância à bactéria nos materiais genéticos, havendo no entanto variação dentro de cada um deles. Na região de Garça, nas plantas de café, observou-se alta proporção de vasos obstruídos nas raízes (3%, entretanto, não houve dano maior na parte aérea.The Xylella fastidiosa bacterium causes problems to coffee cultivation because of a relationship with various stress factors, leading to reduction on coffee production by decreasing fruit number and size, as well as senescence of branches. This research aimed to evaluate the effect of Xylella fastidiosa to Coffea arabica cultivars (grafted or not through the

  2. Avaliação da suscetibilidade à Xylella fastidiosa em diferentes espécies de cafeeiro Susceptibity valuation to Xylella fastidiosa in different coffee species

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    Rachel Benetti Queiroz-Voltan

    2005-01-01

    Full Text Available A bactéria Xylella fastidiosa Wells et al. foi detectada pela primeira vez em cafeeiro no Brasil, em 1995, entretanto acredita-se que a cultura foi infectada por essa bactéria há muitos anos, embora os sintomas fossem atribuídos a um estresse nutricional. Até o momento têm sido realizados estudos principalmente com espécies de C. arabica e C. canephora, porém, em outras espécies do gênero, somente foi detectada sua presença. Neste trabalho, objetivou-se avaliar a proporção de elementos de vaso do xilema obstruídos, total e parcialmente, pela X. fastidiosa, naturalmente infectadas, em diferentes espécies de cafeeiro do Banco de Germoplasma do IAC, visando identificar material resistente a essa bactéria para ser utilizado no programa de melhoramento genético. Os acessos estudados foram: C. canephora (progenitora da 'Guarini', C. liberica var. liberica, os quatro acessos de C. liberica var. dewevrei (Ugandae, Dibowskii, Abeokutae, Excelsa e o híbrido interespecífico Piatã (C. arabica X C. liberica var. dewevrei. Todos eles mostraram-se menos suscetíveis à X. fastidiosa. A porcentagem de obstrução dos elementos de vasos na folha não foi maior que 0,6% na maioria dos acessos, com exceção de Excelsa e do híbrido Piatã com até 2% de obstrução, sendo bem menos suscetíveis a essa bactéria do que as cultivares de C. arabica. Trata-se, portanto, de materiais genéticos importantes para serem utilizados no programa de melhoramento do cafeeiro visando à resistência ao agente dessa doença.Xylella fastidiosa Wells et al. bacteria was firstly detected in coffee plants in Brazil in 1995. However it is believed to be attacking this crop this time. Disease symptoms have been attributed mostly to nutritional unbalances. Up to date studies have comprised only the species C. arabica and C. canephora. However X. fastidiosa was also detected in other Coffea species, but without disease symptoms. Aiming to identify in the IAC

  3. Efeito da poda do tipo decote no controle da xylella fastidiosa em cultivares de cafeeiro "Decote" type pruning effect upon xylella fastidiosa control in coffee cultivars

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    Rachel Benetti Queiroz-Voltan

    2007-01-01

    Full Text Available A bactéria Xylella fastidiosa causa prejuízos à cafeicultura e o emprego de produtos químicos, até o presente, não tem possibilitado o controle econômico dessa bactéria. O manejo adequado do cafezal, desde o plantio, com o uso de mudas isentas da bactéria e o controle das cigarrinhas vetoras, são medidas que atenuam a incidência da doença. A utilização de podas, que tem sido recomendada como medida de controle em citros e videiras, não tem ainda eficiência comprovada para o cafeeiro. Neste trabalho, objetivou-se estudar a eficácia do emprego da poda do tipo decote em cafeeiros arábica como controle de X. fastidiosa. Para tanto, após o emprego desse tipo de poda em cafeeiros infectados pela bactéria, quantificou-se a proporção de elementos de vaso do xilema obstruídos pela bactéria, e avaliou-se a severidade dos sintomas externos de infecção provocados pela X. fastidiosa. Oito meses após a aplicação da poda, no mês de junho de 2003 (período seco, observou-se que as plantas estavam com 4% dos elementos de vaso do pecíolo, 2% na nervura principal e 1% no caule obstruído pela X. fastidiosa. No período chuvoso, 14 meses após a poda, a proporção de obstrução dos elementos de vaso diminuiu para 2% no pecíolo, 1% no caule e na nervura principal. A prática da poda diminuiu ligeiramente a proporção de elementos de vaso obstruídos pela bactéria apenas no período seco, uma vez que foi observado antes da poda um máximo de 6% de obstrução no pecíolo. Os novos ramos que brotavam no cafeeiro, na estação chuvosa, pareciam compensar a obstrução dos ramos mais velhos, diminuindo a proporção de obstrução dos elementos de vaso na planta. Em 2003, não houve diferenças na severidade do sintoma externo entre os tratamentos nos dois períodos, seca e chuvoso. Entretanto, no período seco de 2004, as cultivares Catuaí Vermelho IAC 81 e Mundo Novo IAC 515-20, enxertadas sobre o porta-enxerto 'Apoatã IAC 2258

  4. Eficiência da poda em cafeeiros no controle da Xylella fastidiosa Prune efficiency in the control of Xylella fastidiosa in coffee trees

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    Rachel Benetti Queiroz-Voltan

    2006-01-01

    Full Text Available A bactéria Xylella fastidiosa coloniza os vasos do xilema dos seus hospedeiros, bloqueando o movimento da água e nutrientes, afetando a produção. Até o momento, o manejo adequado do cafezal, desde o plantio, com o uso de mudas isentas da bactéria e o controle das cigarrinhas vetoras, são medidas que atenuam a incidência da doença. A poda é uma prática importante para a otimização da produção do cafezal, e o tipo de poda depende da cultivar e do ambiente, usando-se podas tradicionais ou drásticas. Neste trabalho, objetivou-se avaliar a eficiência do emprego de diferentes tipos de poda no controle de X. fastidiosa nas cultivares comerciais de café arábica `Acaiá IAC 474-19' e `Catuaí Vermelho IAC 81'. Oito plantas de cada cultivar foram submetidas aos tipos de podas: decote, esqueletamento e recepa, em outubro de 2003, e oito delas foram mantidas sem poda, como testemunha. Para o estudo anatômico, anteriormente à poda, foram retirados cinco ramos das plantas utilizadas como testemunha e, em outubro de 2004 (período chuvoso e junho de 2005 (período seco, retiraram-se outros cinco ramos de cada planta dos quatro tratamentos. Na cultivar Acaiá IAC 474-19, notou-se baixa proporção de obstrução dos elementos de vaso de xilema, não sendo observadas diferenças entre os tratamentos. Na `Catuaí Vermelho IAC 81', embora as diferenças entre os tratamentos também não tenham sido significativas, constatou-se uma tendência em diminuir a proporção de obstrução de elementos de vasos do xilema pela bactéria nas podas mais drásticas do tipo esqueletamento e recepa, nos dois períodos (chuvoso e seco. Sugere-se que a prática da poda dos tipos esqueletamento e recepa sejam mais vantajosas para o controle da Xylella em situações de alta severidade.Xylella fastidiosa bacteria colonize the host plant xylem vessels and block the water and nutrient flow with consequent yield reduction. Up to now, adequate coffee plantation

  5. Comparações sazonais do efeito da Xylella fastidiosa em cultivares de cafeeiro Seasonal comparactions of Xylella fastidiosa effect in coffee cultivars

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    Rachel Benetti Queiroz-Voltan

    2004-12-01

    Full Text Available Há muitos anos, o cafeeiro vem apresentando problemas de atrofia e seca dos ramos, atribuídos a um esgotamento nutricional devido às altas taxas de produção. Entretanto, hoje sabe-se que esse problema é causado pela presença da bactéria Xylella fastidiosa Wells et al. cujos sintomas estão relacionados a fatores de estresse. Embora essa bactéria tenha sido muito estudada nos Estados Unidos, devido aos danos causados à videira, pouco se conhece sobre a relação patógeno-hospedeiro-vetor nas diferentes culturas. O objetivo deste trabalho foi o de avaliar a obstrução de elementos de vaso do xilema nas diferentes partes da planta, em ramos com e sem sintomas externos de infecção, de cultivares de cafeeiro (porta-enxertados ou pé-franco, em duas épocas do ano, a fim de verificar o efeito da bactéria na estrutura da planta e na sua nutrição. Foi utilizado um experimento instalado em 1986, em Garça (SP, sendo as amostras retiradas em 2000. Para o estudo anatômico foram obtidas amostras em dois períodos: abril/maio (estação seca e novembro/dezembro (estação chuvosa e para as análises foliar e edáfica foram retiradas amostras em abril. A proporção de obstrução de elementos de vaso do xilema devido ao efeito ocasionado pela bactéria foi maior na estação seca, no período de estresse hídrico, do que na estação chuvosa. O órgão que apresentou uma proporção maior de obstrução de elementos de vaso foi o caule, seguido do pecíolo, limbo e raiz. Não houve diferença significativa na proporção de elementos de vaso obstruídos entre os tratamentos nos dois períodos do ano; também, não houve diferença significativa na composição nutricional foliar nas condições do estudo, porém as amostras retiradas dos ramos com sintoma de infecção, de alguns tratamentos, apresentaram menores concentrações de alguns elementos químicos em relação àquelas amostras retiradas de ramos sem sintoma da presença da bact

  6. Cloning and expression of cellulase XF-818 of Xylella fastidiosa in Escherichia Coli Clonagem e expressão da celulase Xf-818 de Xylella Fastidiosa em Escherichia Coli

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    Nelson Arno Wulff

    2003-12-01

    Full Text Available Xylella fastidiosa's genome was the first of a plant pathogen to be completely sequenced. Through comparative sequence analysis many genes were identified and, among them, several potentially involved in plant-pathogen interaction. However, the biological role of each gene should be assigned experimentally. On this regard, heterologous protein expression is a powerful tool to produce proteins from such genes, allowing their characterization. X. fastidiosa lives inside xylem vessels and eventually would degrade pit membranes from xylem cells to move radialy into the host. The identification of several putative plant cell wall degrading enzymes on X. fastidiosa genome prompted the assession of the function of such proteins. The open reading frame (ORF Xf-818 was cloned into expression vector pET20b and E. coli cells harboring such plasmid exhibited cellulase activity. Using IPTG at 0.4 mmol L-1 with a 12 h incubation at 32°C are the best conditions to produce higher amounts of heterologous protein. The enzyme degrades cellulose confirming the endoglucanase activity of Xf-818.Xylella fastidiosa foi a primeira bactéria fitopatogênica que teve seu genoma completamente seqüenciado. A identificação de diversos genes, através de similaridade de seqüências, indicou os possíveis mecanismos de patogenicidade da bactéria. Entretanto, a determinação da função de um gene requer a confirmação experimental e, neste aspecto, a expressão heteróloga é uma poderosa ferramenta. X. fastidiosa coloniza somente o xilema das plantas hospedeiras e a identificação putativa de diversos genes semelhantes a enzimas que degradam a parede celular vegetal, estimularam o presente estudo de catacterização destas enzimas. A clonagem da ORF Xf-818 de X. fastidiosa no vetor de expressão pET20b possibilitou a produção da proteína heterologamente em E. coli. O emprego de IPTG a 0,4 mmol L-1 com 12 h a 32°C, possibilitou as melhores condições para E. coli

  7. Spatial-temporal distribution of sharpshooters (Hemyptera: Cicadellidae insect vectors of Xylella fastidiosa in citrus orchards = Distribuição espaço-temporal de cigarrinhas (Hemiptera: Cicadellidae vetores da Xylella fastidiosa em pomares cítricos

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    Rúbia de Oliveira Molina

    2016-07-01

    Full Text Available Variegated chlorosis (CVC is a citrus disease, reported initially in the northwest of São Paulo state and in the Triângulo Mineiro region of Minas Gerais state in 1987. The CVC is caused by the xylematic bacteria Xylella fastidiosa. The bacteria is spread through contaminated bubbles or by insect vectors belonging to the Hemyptera order and Cicadellidae family. The aimed of this study was to identify the species of Xylella fastidiosa insect vector and to determine its spatial and temporal distribution in commercial orchards of sweet orange [Citrus sinensis (L. Osbeck]. The experiment was conducted in a commercial area of sweet orange, Pêra variety, grafted on Rangpur lime, located in northwest Paraná. For sampling, yellow sticky traps were used, distributed in the peripheral and central area of the orchard with four replicates per street sampled (5, 30, 55 and 80th plant, each plant was considered a sample unit. Were evaluated ten plots per street, totaling 40 traps for sampling. Every thirty days during the evaluation period, the traps were renewed in the orchard. The main species caught were Acrogonia citrine and Dilobopterus costalimai. The highest incidences occurred from winter to spring, and summer to autumn of the next year. According to the geostatistical analysis, the spatial distribution of these species concentrated in the peripheral zone of the portion where a higher incidence of these species was captured. The results show that it is necessary to adopt pest management practices for the Cicadellidae vector of X. fastidiosa differentiated in space and time. = A clorose variegada dos citros (CVC é uma doença de plantas cítricas, constatada, em 1987, inicialmente nos municípios do noroeste paulista e da região do triângulo mineiro. Ela é causada por uma bactéria de xilema, denominada Xylella fastidiosa. Sua disseminação ocorre através de borbulhas contaminadas ou por meio de insetos vetores da ordem Hemiptera e fam

  8. Diversidade genética de Xylella fastidiosa em regiões produtoras de citros na Bahia

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    Vinicius Oliveira Casais

    2014-01-01

    Full Text Available O objetivo deste trabalho foi avaliar, por meio de marcadores SSR, a diversidade genética de Xylella fastidiosa no Estado da Bahia. Foram estudadas duas das principais regiões produtoras de citros no Estado, o Litoral Norte e o Recôncavo Sul. Para fins comparativos, utilizaram-se dez amostras provenientes do Estado de São Paulo. Foram empregados os seguintes iniciadores: ASSR20, OSSR9, OSSR17, CSSR4, CSSR12 e CSSR20, dos quais os quatro últimos permitiram identificar 22 loci polimórficos. As populações de X. fastidiosa presentes em citros no Estado da Bahia apresentam elevada diversidade genética, com base nos marcadores SSR, com pools gênicos distintos e agrupamento geográfico. No Litoral Norte, as populações do isolado apresentam maior diversidade genética do que as da região do Recôncavo Sul da Bahia.

  9. Plasmid transfer by conjugation in Xylella fastidiosa.

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    Recombination and horizontal gene transfer have been implicated in the adaption of Xylella fastidiosa (Xf) to infect a wide variety of different plant species. There is evidence that certain strains of Xf carry native plasmids equipped with transfer and mobilization genes, suggesting conjugation as ...

  10. Monitoramento populacional das cigarrinhas vetoras de Xylella fastidiosa, através de armadilhas adesivas amarelas em pomares comerciais de citros Population monitoring of leafhopper vectors of Xylella fastidiosa, through yellow sticky traps in commercial citrus orchards

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    Rúbia de Oliveira Molina

    2010-12-01

    Full Text Available A Clorose variegada dos citros (CVC é uma doença causada pela bactéria de xilema Xylella fastidiosa Wells. A disseminação ocorre por meio de insetos vetores pertencente à ordem Hemiptera, família Cicadellidae (subfamília Cicadellinae, os quais transmitem a bactéria depois de se alimentarem em plantas contaminadas. Neste trabalho, objetivou-se identificar e monitorar as espécies de cigarrinhas vetoras em um pomar comercial no município de Paranavaí, Paraná. O experimento foi realizado em um talhão comercial de laranja doce (Citrus sinensis (L. Osbeck, variedade Pera, com 1.000 plantas de dez anos de idade. A amostragem foi realizada por meio de armadilhas adesivas amarelas, distribuídas na área periférica e central do pomar, com duas repetições por rua amostrada. As etiquetas foram distribuídas entre a 5ª e 30ª plantas em 10 ruas, totalizando 20 armadilhas que foram renovadas no pomar, a cada trinta dias, durante o período de avaliação que foi entre junho de 2005 e setembro de 2006. As principais espécies de Cicadellinae capturadas foram Acrogonia citrina Marucci & Cavichioli, Dilobopterus costalimai Young e Macugonalia cavifrons Stal. Essas espécies apresentaram ocorrência constante e frequência de 3,97%, 4,2%, 13,0% respectivamente, em relação ao total de cigarrinhas coletadas.Citrus variegated chlorosis (CVC is the disease caused by the bacteria Xylella fastidiosa Wells. Dissemination occurs through insect vectors belonging to the order Hemiptera, family Cicadellidae (subfamily Cicadellinae, which transmit the bacteria after feeding on infected plants. The objective of this study was to identify the species of insect vectors in an orchard in the municipality of Paranavaí, in the State of Paraná. The experiment was conducted in a commercial stand of sweet orange (Citrus sinensis (L. Osbeck, Pêra variety with 1,000 10-year-old plants. Monitoring was performed using yellow sticky traps, distributed in the central

  11. Morphological evidence for phages in Xylella fastidiosa

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    Civerolo Edwin L

    2008-06-01

    Full Text Available Abstract Presumptive phage particles associated with Xylella fastidiosa strain Temecula-1 grown in PW broth were observed by transmission electron microscopy (TEM in ultrathin sections of bacterial cell-containing low speed centrifugation pellets and in partially purified preparations from CsCl equilibrium centrifugation density gradients. Ultrathin-sectioned cell pellets contained icosahedral particles of about 45 nm in diameter. Samples collected from CsCl density gradients revealed mostly non-tailed icosahedral but also tailed particles. The icosahedral particles could be divided into two types: a large type (about 45 nm and a small type (about 30 nm. Filamentous phage-like particles (17 × 120 to 6,300 nm were also observed. The presence of different types of phage-like particles resembling to those in several bacteriophage families provides new physical evidence, in addition to X. fastidiosa genomic information, that X. fastidiosa possesses active phages. This is the first report of phage particles released in X. fastidiosa cultures.

  12. Draft genome sequence of Xylella fastidiosa subsp. fastidiosa strain Stag’s Leap

    Science.gov (United States)

    Xylella fastidiosa subsp. fastidiosa causes Pierce’s disease of grapevine. Presented here is the draft genome sequence of the Stag’s Leap strain, previously used in pathogenicity/virulence assays to evaluate grapevine germplasm bearing Pierce’s disease....

  13. Draft Genome Sequence of Xylella fastidiosa subsp. fastidiosa Strain Stag?s Leap

    OpenAIRE

    Chen, J.; Wu, F.; Zheng, Z.; Deng, X.; Burbank, L. P.; Stenger, D. C.

    2016-01-01

    Xylella fastidiosa subsp. fastidiosa causes Pierce?s disease of grapevine. Presented here is the draft genome sequence of the Stag?s Leap strain, previously used in pathogenicity/virulence assays to evaluate grapevine germplasm bearing Pierce?s disease resistance and a phenotypic assessment of knockout mutants to determine gene function.

  14. Antibacterial Activity of Phenolic Compounds Against the Phytopathogen Xylella fastidiosa

    OpenAIRE

    Maddox, Christina E.; Laur, Lisa M.; Tian, Li

    2010-01-01

    Xylella fastidiosa is a pathogenic bacterium that causes diseases in many crop species, which leads to considerable economic loss. Phenolic compounds (a group of secondary metabolites) are widely distributed in plants and have shown to possess antimicrobial properties. The anti-Xylella activity of 12 phenolic compounds, representing phenolic acid, coumarin, stilbene and flavonoid, was evaluated using an in vitro agar dilution assay. Overall, these phenolic compounds were effective in inhibiti...

  15. Xylella fastidiosa esterase rather than hydroxynitrile lyase.

    Science.gov (United States)

    Torrelo, Guzman; Ribeiro de Souza, Fayene Zeferino; Carrilho, Emanuel; Hanefeld, Ulf

    2015-03-02

    In 2009, we reported that the product of the gene SCJ21.16 (XFa0032) from Xylella fastidiosa, a xylem-restricted plant pathogen that causes a range of diseases in several important crops, encodes a protein (XfHNL) with putative hydroxynitrile lyase activity. Sequence analysis and activity tests indicated that XfHNL exhibits an α/β-hydrolase fold and could be classified as a member of the family of FAD-independent HNLs. Here we provide a more detailed sequence analysis and new experimental data. Using pure heterologously expressed XfHNL we show that this enzyme cannot catalyse the cleavage/synthesis of mandelonitrile and that this protein is in fact a non-enantioselective esterase. Homology modelling and ligand docking simulations were used to study the active site and support these results. This finding could help elucidate the common ancestor of esterases and hydroxynitrile lyases with an α/β -hydrolase fold. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  16. Fractal analysis of Xylella fastidiosa biofilm formation

    Science.gov (United States)

    Moreau, A. L. D.; Lorite, G. S.; Rodrigues, C. M.; Souza, A. A.; Cotta, M. A.

    2009-07-01

    We have investigated the growth process of Xylella fastidiosa biofilms inoculated on a glass. The size and the distance between biofilms were analyzed by optical images; a fractal analysis was carried out using scaling concepts and atomic force microscopy images. We observed that different biofilms show similar fractal characteristics, although morphological variations can be identified for different biofilm stages. Two types of structural patterns are suggested from the observed fractal dimensions Df. In the initial and final stages of biofilm formation, Df is 2.73±0.06 and 2.68±0.06, respectively, while in the maturation stage, Df=2.57±0.08. These values suggest that the biofilm growth can be understood as an Eden model in the former case, while diffusion-limited aggregation (DLA) seems to dominate the maturation stage. Changes in the correlation length parallel to the surface were also observed; these results were correlated with the biofilm matrix formation, which can hinder nutrient diffusion and thus create conditions to drive DLA growth.

  17. Host colonization differences between citrus and coffee isolates of Xylella fastidiosa in reciprocal inoculation Diferenças em colonização do hospedeiro por isolados de Xylella fastidiosa de citros e cafeeiro em inoculações recíprocas

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    Simone de Souza Prado

    2008-01-01

    Full Text Available Citrus variegated chlorosis (CVC and coffee stem atrophy (CSA are important diseases in Brazil associated with closely-related strains of Xylella fastidiosa, but little is know about host aoverlappingnd importance of citrus and coffee as inoculum sources of these strains. In this study, reciprocal-inoculation experiments were performed to determine if CVC and CSA isolates are biologically similar within citrus and coffee plants. These two hosts were mechanically inoculated with a CVC and a CSA isolate of X. fastidiosa at four concentrations ranging between10³ and 10(9 colony forming units CFU mL-1. At two, four and eight months after inoculation, the infection efficiency and bacterial populations of the isolates in each host were determined by culturing. The CVC isolate infected both citrus and coffee plants, but developed lower populations in coffee. The CSA isolate did not colonize citrus. Inoculation of coffee plants with the CVC isolate resulted in low rates of infection and required an inoculum concentration ten-fold higher than that necessary to obtain a similar (25% rate of infection in citrus. The relatively low infection rates and bacterial numbers of the CVC isolate in coffee plants compared with those observed in citrus suggest that coffee is not a suitable host to serve as a source of inoculum of the CVC strain for primary spread to citrus or within coffee plantations.Clorose variegada dos citros (CVC e atrofia dos ramos do cafeeiro (ARC são doenças importantes no Brasil, associadas a estirpes de Xylella fastidiosa que são geneticamente próximas. Entretanto, pouco se sabe a respeito de plantas hospedeiras em comum e da importância de citros e cafeeiro como fontes de inóculo dessas estirpes. Neste estudo, realizaram-se experimentos de inoculação recíproca para determinar se isolados de X. fastidiosa de CVC e de ARC são biologicamente semelhantes em plantas de citros e café. Estes dois hospedeiros foram mecanicamente

  18. Biological and bacteriological characterization of the strain of Xylella fastidiosa Xf-PGAICR and detection of X. fastidiosa from leafhoppers

    International Nuclear Information System (INIS)

    Marin Chaves, Maria Priscilla

    2014-01-01

    Laboratory tools are developed and optimized for the biological characterization of a strain of Xylella fastidiosa (Xf-PGAICR) isolated of guava. The testing of pathogenesis robustly has allowed the characterization of other strains of X. fastidiosa. The detection of X. fastidiosa is carried out in their insect vectors to establish the relationship with the presence of phytopathogen in the vector, the presence and dissemination of X. fastidiosa to nearby areas [es

  19. Characterization of a diffusible signaling factor from Xylella fastidiosa.

    Science.gov (United States)

    Beaulieu, Ellen D; Ionescu, Michael; Chatterjee, Subhadeep; Yokota, Kenji; Trauner, Dirk; Lindow, Steven

    2013-01-08

    Cell-cell signaling in Xylella fastidiosa has been implicated in the coordination of traits enabling colonization in plant hosts as well as insect vectors. This cell density-dependent signaling has been attributed to a diffusible signaling factor (DSF) produced by the DSF synthase RpfF. DSF produced by related bacterial species are unsaturated fatty acids, but that of X. fastidiosa was thought to be different from those of other taxa. We describe here the isolation and characterization of an X. fastidiosa DSF (XfDSF) as 2(Z)-tetradecenoic acid. This compound was isolated both from recombinant Erwinia herbicola expressing X. fastidiosa rpfF and from an X. fastidiosa rpfC deletion mutant that overproduces DSF. Since an rpfF mutant is impaired in biofilm formation and underexpresses the hemagglutinin-like protein-encoding genes hxfA and hxfB, we demonstrate that these traits can be restored by ca. 0.5 µM XfDSF but not by myristic acid, the fully saturated tetradecenoic acid. A phoA-based X. fastidiosa biosensor that assesses DSF-dependent expression of hxfA or hxfB revealed a high level of molecular specificity of DSF signaling. X. fastidiosa causes diseases in many important plants, including grape, where it incites Pierce's disease. Virulence of X. fastidiosa for grape is coordinated by cell-cell signaling molecules, designated DSF (Diffusible Signaling Factor). Mutants blocked in DSF production are hypervirulent for grape, suggesting that virulence is suppressed upon DSF accumulation and that disease could be controlled by artificial elevation of the DSF level in plants. In this work, we describe the isolation of the DSF produced by X. fastidiosa and the verification of its biological activity as an antivirulence factor. We also have developed X. fastidiosa DSF biosensors to evaluate the specificity of cell-cell signaling to be investigated.

  20. Avaliação da resistência à Xylella fastidiosa em germoplasma de tangerina e híbridos introduzidos da Itália e Córsega

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    GONZÁLEZ JAIMES ELENA PAOLA

    2002-01-01

    Full Text Available A Clorose Variegada dos Citros (CVC, causada pela bactéria Xylella fastidiosa, é atualmente uma das doenças que mais afeta a citricultura brasileira, sendo as variedades de laranja-doce as mais afetadas. Ensaio instalado na Estação Experimental de Bebedouro (EECB, em condições de estufa, teve como objetivo avaliar o comportamento em relação à CVC de germoplasma de citros introduzidos pela EECB, Fundecitrus e Cenargen. Os materiais foram multiplicados sobre diversos porta-enxertos e, quando atingiram o tamanho adequado, inoculados por garfagem lateral de ramo doente. Cada variedade constou de quatro plantas, três das quais foram inoculadas, e a outra sem inocular deixada como padrão. As avaliações consistiram na observação de sintomas, teste de ELISA e PCR. Os primeiros sintomas nos materiais contaminados começaram a surgir 7 meses após a inoculação. Encontraram-se 18 variedades positivas no teste de PCR, o que indica sua suscetibilidade à bactéria Xylella fastidiosa. Entretanto, as variedades que foram detectadas pelo teste ELISA e não pelo PCR não foram contadas como suscetíveis e, sim, como falsos positivos.

  1. Xylella fastidiosa Isolates from Both subsp. multiplex and fastidiosa Cause Disease on Southern Highbush Blueberry (Vaccinium sp.) Under Greenhouse Conditions.

    Science.gov (United States)

    Oliver, J E; Cobine, P A; De La Fuente, L

    2015-07-01

    Xylella fastidiosa is a xylem-limited gram-negative plant pathogen that affects numerous crop species, including grape, citrus, peach, pecan, and almond. Recently, X. fastidiosa has also been found to be the cause of bacterial leaf scorch on blueberry in the southeastern United States. Thus far, all X. fastidiosa isolates obtained from infected blueberry have been classified as X. fastidiosa subsp. multiplex; however, X. fastidiosa subsp. fastidiosa isolates are also present in the southeastern United States and commonly cause Pierce's disease of grapevines. In this study, seven southeastern U.S. isolates of X. fastidiosa, including three X. fastidiosa subsp. fastidiosa isolates from grape, one X. fastidiosa subsp. fastidiosa isolate from elderberry, and three X. fastidiosa subsp. multiplex isolates from blueberry, were used to infect the southern highbush blueberry 'Rebel'. Following inoculation, all isolates colonized blueberry, and isolates from both X. fastidiosa subsp. multiplex and X. fastidiosa subsp. fastidiosa caused symptoms, including characteristic stem yellowing and leaf scorch symptoms as well as dieback of the stem tips. Two X. fastidiosa subsp. multiplex isolates from blueberry caused more severe symptoms than the other isolates examined, and infection with these two isolates also had a significant impact on host mineral nutrient content in sap and leaves. These findings have potential implications for understanding X. fastidiosa host adaptation and expansion and the development of emerging diseases caused by this bacterium.

  2. Xylella fastidiosa gene expression analysis by DNA microarrays

    OpenAIRE

    Travensolo,Regiane F.; Carareto-Alves,Lucia M.; Costa,Maria V.C.G.; Lopes,Tiago J.S.; Carrilho,Emanuel; Lemos,Eliana G.M.

    2009-01-01

    Xylella fastidiosa genome sequencing has generated valuable data by identifying genes acting either on metabolic pathways or in associated pathogenicity and virulence. Based on available information on these genes, new strategies for studying their expression patterns, such as microarray technology, were employed. A total of 2,600 primer pairs were synthesized and then used to generate fragments using the PCR technique. The arrays were hybridized against cDNAs labeled during reverse transcrip...

  3. EXPLOITING PATHOGEN CONFUSION STRATEGY TO ACHIEVE XYLELLA FASTIDIOSA BIOCONTROL

    OpenAIRE

    G. D'Attoma, A. Giampetruzzi, V.N. Savino, D. Boscia, P. Saldarelli.

    2016-01-01

    The recent findings of the plant pathogenic bacterium Xylella fastidiosa (Xf), infecting several plant species in Italy and France, raised major concerns for its potential impact on the EU and Mediterranean agriculture. In the current EU outbreaks, olive is the predominantly affected crop, in which the bacterium has been consistently associated with a new severe syndrome, denoted “Olive Quick Decline”. So far, no effective treatments are available to cure infected plants. However, several app...

  4. Chitin utilization by the insect-transmitted bacterium Xylella fastidiosa.

    Science.gov (United States)

    Killiny, Nabil; Prado, Simone S; Almeida, Rodrigo P P

    2010-09-01

    Xylella fastidiosa is an insect-borne bacterium that colonizes xylem vessels of a large number of host plants, including several crops of economic importance. Chitin is a polysaccharide present in the cuticle of leafhopper vectors of X. fastidiosa and may serve as a carbon source for this bacterium. Biological assays showed that X. fastidiosa reached larger populations in the presence of chitin. Additionally, chitin induced phenotypic changes in this bacterium, notably increasing adhesiveness. Quantitative PCR assays indicated transcriptional changes in the presence of chitin, and an enzymatic assay demonstrated chitinolytic activity by X. fastidiosa. An ortholog of the chitinase A gene (chiA) was identified in the X. fastidiosa genome. The in silico analysis revealed that the open reading frame of chiA encodes a protein of 351 amino acids with an estimated molecular mass of 40 kDa. chiA is in a locus that consists of genes implicated in polysaccharide degradation. Moreover, this locus was also found in the genomes of closely related bacteria in the genus Xanthomonas, which are plant but not insect associated. X. fastidiosa degraded chitin when grown on a solid chitin-yeast extract-agar medium and grew in liquid medium with chitin as the sole carbon source; ChiA was also determined to be secreted. The gene encoding ChiA was cloned into Escherichia coli, and endochitinase activity was detected in the transformant, showing that the gene is functional and involved in chitin degradation. The results suggest that X. fastidiosa may use its vectors' foregut surface as a carbon source. In addition, chitin may trigger X. fastidiosa's gene regulation and biofilm formation within vectors. Further work is necessary to characterize the role of chitin and its utilization in X. fastidiosa.

  5. Locais e período de alimentação da cigarrinha vetora de Xylella fastidiosa, Bucephalogonia xanthophis (Berg (Hemiptera: Cicadellidae, em mudas cítricas Feeding sites and food intake of Bucephalogonia xanthophis (Berg (Hemiptera: Cicadellidae, a sharpshooter vector of Xylella fastidiosa, on citrus plants

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    Marcelo Pedreira de Miranda

    2008-12-01

    Full Text Available A cigarrinha Bucephalogonia xanthophis (Berg (Hemiptera: Cicadellidae é um importante vetor da bactéria Xylella fastidiosa, agente causal da clorose variegada dos citros. Este trabalho teve como objetivo identificar o local preferido de alimentação e o período de maior atividade alimentar desta cigarrinha em citros, no sentido de elucidar o comportamento alimentar relacionado à transmissão da bactéria. O local de alimentação foi estudado em ensaio de escolha, no qual 30 insetos adultos foram liberados em gaiolas de observação (n = 10 contendo uma muda de laranja-doce [Citrus sinensis (L. Osbeck]. Após 1; 15; 21; 25; 39; 45 e 49 h da liberação, contaram-se os insetos na parte superior (ramos com brotações e inferior (haste principal, até H"40 cm de altura da muda. Nos ramos da parte superior, avaliou-se a preferência entre a haste, o pecíolo e o limbo foliar. Em um segundo ensaio, 20 machos e 20 fêmeas de B. xanthophis foram confinados individualmente sobre a haste de 'seedlings' de laranja-doce para determinar os períodos de alimentação, quantificando-se a excreção de 'honeydew' (medida indireta da ingestão em períodos sucessivos de dia e noite, durante 48 h. A maioria dos indivíduos de B. xanthophis preferiu a haste dos ramos novos (62%, na parte superior da muda cítrica (91%. Nos 'seedlings', observou-se maior volume de excreção e proporção de indivíduos excretando durante a fotofase, independentemente do sexo. Portanto, em estudos de transmissão de X. fastidiosa, deve-se considerar a preferência de B. xanthophis pela haste de brotações cítricas e sua maior atividade alimentar durante a fotofase.The sharpshooter Bucephalogonia xanthophis (Berg (Hemiptera: Cicadellidae is an important vector of Xylella fastidiosa, a bacterium that causes citrus variegated chlorosis. This study aimed to identify the preferred feeding sites and periods of this vector on citrus, in order to improve our understanding of the

  6. In vitro Determination of Extracellular Proteins from Xylella fastidiosa.

    Science.gov (United States)

    Mendes, Juliano S; Santiago, André S; Toledo, Marcelo A S; Horta, Maria A C; de Souza, Alessandra A; Tasic, Ljubica; de Souza, Anete P

    2016-01-01

    The phytopathogen Xylella fastidiosa causes economic losses in important agricultural crops. Xylem vessel occlusion caused by biofilm formation is the major mechanism underlying the pathogenicity of distinct strains of X. fastidiosa . Here, we provide a detailed in vitro characterization of the extracellular proteins of X. fastidiosa . Based on the results, we performed a comparison with a strain J1a12, which cannot induce citrus variegated chlorosis symptoms when inoculated into citrus plants. We then extend this approach to analyze the extracellular proteins of X. fastidiosa in media supplemented with calcium. We verified increases in extracellular proteins concomitant with the days of growth and, consequently, biofilm development (3-30 days). Outer membrane vesicles carrying toxins were identified beginning at 10 days of growth in the 9a5c strain. In addition, a decrease in extracellular proteins in media supplemented with calcium was observed in both strains. Using mass spectrometry, 71 different proteins were identified during 30 days of X. fastidiosa biofilm development, including proteases, quorum-sensing proteins, biofilm formation proteins, hypothetical proteins, phage-related proteins, chaperones, toxins, antitoxins, and extracellular vesicle membrane components.

  7. Network analysis reveals why Xylella fastidiosa will persist in Europe.

    Science.gov (United States)

    Strona, Giovanni; Carstens, Corrie Jacobien; Beck, Pieter S A

    2017-03-06

    The insect vector borne bacterium Xylella fastidiosa was first detected in olive trees in Southern Italy in 2013, and identified as the main culprit behind the 'olive quick decline syndrome'. Since then, the disease has spread rapidly through Italy's main olive oil producing region. The epidemiology of the outbreak is largely unstudied, with the list of X. fastidiosa hosts and vectors in Europe likely incomplete, and the role humans play in dispersal unknown. These knowledge gaps have led to management strategies based on general assumptions that require, among others, local vector control and, in certain areas, the destruction of infected plants and healthy ones around them in an attempt to eradicate or halt the spreading pest. Here we show that, regardless of epidemiological uncertainties, the mere distribution of olive orchards in Southern Italy makes the chances of eradicating X. fastidiosa from the region extremely slim. Our results imply that Southern Italy is becoming a reservoir for X. fastidiosa. As a consequence, management strategies should keep the prevalence of X. fastidiosa in the region as low as possible, primarily through vector control, lest the pathogen, that has also been detected in southern France and the island of Mallorca (Spain), continues spreading through Italy and Europe.

  8. Transmission of Xylella fastidiosa to Grapevine by the Meadow Spittlebug.

    Science.gov (United States)

    Cornara, D; Sicard, A; Zeilinger, A R; Porcelli, F; Purcell, A H; Almeida, R P P

    2016-11-01

    There is little information available on Xylella fastidiosa transmission by spittlebugs (Hemiptera, Cercopoidea). This group of insect vectors may be of epidemiological relevance in certain diseases, so it is important to better understand the basic parameters of X. fastidiosa transmission by spittlebugs. We used grapevines as a host plant and the aphrophorid Philaenus spumarius as a vector to estimate the effect of plant access time on X. fastidiosa transmission to plants; in addition, bacterial population estimates in the heads of vectors were determined and correlated with plant infection status. Results show that transmission efficiency of X. fastidiosa by P. spumarius increased with plant access time, similarly to insect vectors in another family (Hemiptera, Cicadellidae). Furthermore, a positive correlation between pathogen populations in P. spumarius and transmission to plants was observed. Bacterial populations in insects were one to two orders of magnitude lower than those observed in leafhopper vectors, and population size peaked within 3 days of plant access period. These results suggest that P. spumarius has either a limited number of sites in the foregut that may be colonized, or that fluid dynamics in the mouthparts of these insects is different from that in leafhoppers. Altogether our results indicate that X. fastidiosa transmission by spittlebugs is similar to that by leafhoppers. In addition, the relationship between cell numbers in vectors and plant infection may have under-appreciated consequences to pathogen spread.

  9. Potential complications when developing gene deletion clones in Xylella fastidiosa.

    Science.gov (United States)

    Johnson, Kameka L; Cursino, Luciana; Athinuwat, Dusit; Burr, Thomas J; Mowery, Patricia

    2015-04-16

    The Gram-negative xylem-limited bacterium, Xylella fastidiosa, is an important plant pathogen that infects a number of high value crops. The Temecula 1 strain infects grapevines and induces Pierce's disease, which causes symptoms such as scorching on leaves, cluster collapse, and eventual plant death. In order to understand the pathogenesis of X. fastidiosa, researchers routinely perform gene deletion studies and select mutants via antibiotic markers. Site-directed pilJ mutant of X. fastidiosa were generated and selected on antibiotic media. Mutant cultures were assessed by PCR to determine if they were composed of purely transformant cells or included mixtures of non-transformants cells. Then pure pilJ mutant and wildtype cells were mixed in PD2 medium and following incubation and exposure to kanamycin were assessed by PCR for presence of mutant and wildtype populations. We have discovered that when creating clones of targeted mutants of X. fastidiosa Temecula 1 with selection on antibiotic plates, X. fastidiosa lacking the gene deletion often persist in association with targeted mutant cells. We believe this phenomenon is due to spontaneous antibiotic resistance and/or X. fastidiosa characteristically forming aggregates that can be comprised of transformed and non-transformed cells. A combined population was confirmed by PCR, which showed that targeted mutant clones were mixed with non-transformed cells. After repeated transfer and storage the non-transformed cells became the dominant clone present. We have discovered that special precautions are warranted when developing a targeted gene mutation in X. fastidiosa because colonies that arise following transformation and selection are often comprised of transformed and non-transformed cells. Following transfer and storage the cells can consist primarily of the non-transformed strain. As a result, careful monitoring of targeted mutant strains must be performed to avoid mixed populations and confounding results.

  10. Chromosome-based genetic complementation system for Xylella fastidiosa.

    Science.gov (United States)

    Matsumoto, Ayumi; Young, Glenn M; Igo, Michele M

    2009-03-01

    Xylella fastidiosa is a xylem-limited, gram-negative bacterium that causes Pierce's disease of grapevine. Here, we describe the construction of four vectors that facilitate the insertion of genes into a neutral site (NS1) in the X. fastidiosa chromosome. These vectors carry a colE1-like (pMB1) replicon and DNA sequences from NS1 flanking a multiple-cloning site and a resistance marker for one of the following antibiotics: chloramphenicol, erythromycin, gentamicin, or kanamycin. In X. fastidiosa, vectors with colE1-like (pMB1) replicons have been found to result primarily in the recovery of double recombinants rather than single recombinants. Thus, the ease of obtaining double recombinants and the stability of the resulting insertions at NS1 in the absence of selective pressure are the major advantages of this system. Based on in vitro and in planta characterizations, strains carrying insertions within NS1 are indistinguishable from wild-type X. fastidiosa in terms of growth rate, biofilm formation, and pathogenicity. To illustrate the usefulness of this system for complementation analysis, we constructed a strain carrying a mutation in the X. fastidiosa cpeB gene, which is predicted to encode a catalase/peroxidase, and showed that the sensitivity of this mutant to hydrogen peroxide could be overcome by the introduction of a wild-type copy of cpeB at NS1. Thus, this chromosome-based complementation system provides a valuable genetic tool for investigating the role of specific genes in X. fastidiosa cell physiology and virulence.

  11. Production of DNA microarray and expression analysis of genes from Xylella fastidiosa in different culture media

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    Regiane de Fátima Travensolo

    2009-06-01

    Full Text Available DNA Microarray was developed to monitor the expression of many genes from Xylella fastidiosa, allowing the side by-side comparison of two situations in a single experiment. The experiments were performed using X. fastidiosa cells grown in two culture media: BCYE and XDM2. The primers were synthesized, spotted onto glass slides and the array was hybridized against fluorescently labeled cDNAs. The emitted signals were quantified, normalized and the data were statistically analyzed to verify the differentially expressed genes. According to the data, 104 genes were differentially expressed in XDM2 and 30 genes in BCYE media. The present study showed that DNA microarray technique efficiently differentiate the expressed genes under different conditions.DNA Microarray foi desenvolvida para monitorar a expressão de muitos genes de Xylella fastidiosa, permitindo a comparação de duas situações distintas em um único experimento. Os experimentos foram feitos utilizando células de X. fastidiosa cultivada em dois meios de cultura: BCYE e XDM2. Pares de oligonucleotídeos iniciadores foram sintetizados, depositados em lâminas de vidro e o arranjo foi hibridizado contra cDNAs marcados fluorescentemente. Os sinais emitidos foram quantificados, normalizados e os dados foram estatisticamente analisados para verificar os genes diferencialmente expressos. De acordo com nossos dados, 104 genes foram diferencialmente expressos para o meio de cultura XDM2 e 30 genes para o BCYE. No presente estudo, nós demonstramos que a técnica de DNA microarrays eficientemente diferencia genes expressos sob diferentes condições de cultivo.

  12. Diversity of endophytic bacterial populations and their interaction with Xylella fastidiosa in citrus plants

    NARCIS (Netherlands)

    Araujo, W.L.; Marcon, J.; jr. Maccheroni, W.; Elsas, van J.D.; Vuurde, van J.W.L.; Azevedo, de J.L.

    2002-01-01

    Citrus variegated chlorosis (CVC) is caused by Xylella fastidiosa, a phytopathogenic bacterium that can infect all Citrus sinensis cultivars. The endophytic bacterial communities of healthy, resistant, and CVC-affected citrus plants were studied by using cultivation as well as

  13. The Xylella fastidiosa PD1063 protein is secreted in association with outer membrane vesicles.

    Science.gov (United States)

    Pierce, Brittany K; Voegel, Tanja; Kirkpatrick, Bruce C

    2014-01-01

    Xylella fastidiosa is a gram-negative, xylem-limited plant pathogenic bacterium that causes disease in a variety of economically important agricultural crops including Pierce's disease of grapevines. Xylella fastidiosa biofilms formed in the xylem vessels of plants play a key role in early colonization and pathogenicity by providing a protected niche and enhanced cell survival. Here we investigate the role of Xylella fastidiosa PD1063, the predicted ortholog of Xanthomonas oryzae pv. oryzae PXO_03968, which encodes an outer membrane protein. To assess the function of the Xylella fastidiosa ortholog, we created Xylella fastidiosa mutants deleted for PD1063 and then assessed biofilm formation, cell-cell aggregation and cell growth in vitro. We also assessed disease severity and pathogen titers in grapevines mechanically inoculated with the Xylella fastidiosa PD1063 mutant. We found a significant decrease in cell-cell aggregation among PD1063 mutants but no differences in cell growth, biofilm formation, disease severity or titers in planta. Based on the demonstration that Xanthomonas oryzae pv. oryzae PXO_03968 encodes an outer membrane protein, secreted in association with outer membrane vesicles, we predicted that PD1063 would also be secreted in a similar manner. Using anti-PD1063 antibodies, we found PD1063 in the supernatant and secreted in association with outer membrane vesicles. PD1063 purified from the supernatant, outer membrane fractions and outer membrane vesicles was 19.2 kD, corresponding to the predicted size of the processed protein. Our findings suggest Xylella fastidiosa PD1063 is not essential for development of Pierce's disease in Vitis vinifera grapevines although further research is required to determine the function of the PD1063 outer membrane protein in Xylella fastidiosa.

  14. Temperature dependent RNA metabolism in Xylella fastidiosa during cold stress and grapevine infection

    Science.gov (United States)

    Re-occurrence of Pierce’s disease of grapes, caused by Xylella fastidiosa, is known to be influenced by environmental factors, particularly cold temperatures during overwintering. Grapevines in colder regions are often cured of X. fastidiosa infection over the winter season, depending on cultivar, t...

  15. Identification and characterization of plasmids from Xylella fastidiosa using next generation sequencing analyses

    Science.gov (United States)

    Xylella fastidiosa causes diseases in many economically important crops, e.g. Pierce’s disease (PD) of grapevines and blueberry bacterial leaf scorch (BBLS) disease in the U.S. Biological research on X. fastidiosa has been difficult due to its nutritional fastidiousness. Genomic research provides a ...

  16. Xylella fastidiosa in rabbiteye blueberry: A newly studied host of an old foe

    Science.gov (United States)

    The bacterium Xylella fastidiosa causes a number of plant diseases, including bacterial leaf scorch of southern highbush blueberry. In Louisiana, X. fastidiosa has been detected in rabbiteye blueberry orchards, and we wanted to know if it affected yield in rabbiteye blueberry plants. We detected X...

  17. Functional characterization of the role of the PilG in Xylella fastidiosa

    Science.gov (United States)

    Type IV pili of Xylella fastidiosa are regulated by pilG, a chemotaxis regulator in the Pil-Chp operon involving signal transduction pathways. To elucidate the role of pilG in twitching motility and pathogenicity of X. fastidiosa, phenotypes of wild type, a pilG-mutant, and a complementary strain we...

  18. Conjugative plasmid transfer in Xylella fastidiosa is dependent on tra and trb operon functions

    Science.gov (United States)

    The insect-transmitted plant pathogen Xylella fastidiosa is capable of efficient horizontal gene transfer and recombination, leading to diversity between strains and the categorization of X. fastidiosa into multiple subspecies. Although natural transformation is shown to occur at high rates in X. fa...

  19. Model Plants for Studying the Interaction between Methylobacterium mesophilicum and Xylella fastidiosa

    NARCIS (Netherlands)

    Andreote, F.D.; Lacava, P.T.; Araújo, W.L.; Maccheroni Jr., W.; Overbeek, van L.S.; Elsas, van J.D.; Azevedo, J.L.

    2006-01-01

    Over the last few years, endophytic bacterial communities associated with citrus have been studied as key components interacting with Xylella fastidiosa. In this study, we investigated the possible interaction between the citrus endophyte Methylobacterium mesophilicum SR1.6/6 and X. fastidiosa in

  20. Model plants for studying the interaction between Methylobacterium mesophilicum and Xylella fastidiosa

    NARCIS (Netherlands)

    Andreote, FD; Lacava, PT; Gai, CS; Araujo, WL; Maccheroni, W; van Overbeek, LS; van Elsas, JD; Azevedo, JL

    Over the last few years, endophytic bacterial communities associated with citrus have been studied as key components interacting with Xylella fastidiosa. In this study, we investigated the possible interaction between the citrus endophyte Methylobacterium mesophilicum SR1.6/6 and X. fastidiosa in

  1. Xylella fastidiosa gene expression analysis by DNA microarrays

    Directory of Open Access Journals (Sweden)

    Regiane F. Travensolo

    2009-01-01

    Full Text Available Xylella fastidiosa genome sequencing has generated valuable data by identifying genes acting either on metabolic pathways or in associated pathogenicity and virulence. Based on available information on these genes, new strategies for studying their expression patterns, such as microarray technology, were employed. A total of 2,600 primer pairs were synthesized and then used to generate fragments using the PCR technique. The arrays were hybridized against cDNAs labeled during reverse transcription reactions and which were obtained from bacteria grown under two different conditions (liquid XDM2 and liquid BCYE. All data were statistically analyzed to verify which genes were differentially expressed. In addition to exploring conditions for X. fastidiosa genome-wide transcriptome analysis, the present work observed the differential expression of several classes of genes (energy, protein, amino acid and nucleotide metabolism, transport, degradation of substances, toxins and hypothetical proteins, among others. The understanding of expressed genes in these two different media will be useful in comprehending the metabolic characteristics of X. fastidiosa, and in evaluating how important certain genes are for the functioning and survival of these bacteria in plants.

  2. Conjugative Plasmid Transfer in Xylella fastidiosa Is Dependent on tra and trb Operon Functions

    OpenAIRE

    Burbank, Lindsey P.; Van Horn, Christopher R.

    2017-01-01

    The insect-transmitted plant pathogen Xylella fastidiosa is capable of efficient horizontal gene transfer (HGT) and recombination. Natural transformation occurs at high rates in X. fastidiosa, but there also is evidence that certain strains of X. fastidiosa carry native plasmids equipped with transfer and mobilization genes, suggesting conjugation as an additional mechanism of HGT in some instances. Two operons, tra and trb, putatively encoding a conjugative type IV secretion system, are foun...

  3. Xylella fastidiosa: Host Range and Advance in Molecular Identification Techniques

    Science.gov (United States)

    Baldi, Paolo; La Porta, Nicola

    2017-01-01

    In the never ending struggle against plant pathogenic bacteria, a major goal is the early identification and classification of infecting microorganisms. Xylella fastidiosa, a Gram-negative bacterium belonging to the family Xanthmonadaceae, is no exception as this pathogen showed a broad range of vectors and host plants, many of which may carry the pathogen for a long time without showing any symptom. Till the last years, most of the diseases caused by X. fastidiosa have been reported from North and South America, but recently a widespread infection of olive quick decline syndrome caused by this fastidious pathogen appeared in Apulia (south-eastern Italy), and several cases of X. fastidiosa infection have been reported in other European Countries. At least five different subspecies of X. fastidiosa have been reported and classified: fastidiosa, multiplex, pauca, sandyi, and tashke. A sixth subspecies (morus) has been recently proposed. Therefore, it is vital to develop fast and reliable methods that allow the pathogen detection during the very early stages of infection, in order to prevent further spreading of this dangerous bacterium. To this purpose, the classical immunological methods such as ELISA and immunofluorescence are not always sensitive enough. However, PCR-based methods exploiting specific primers for the amplification of target regions of genomic DNA have been developed and are becoming a powerful tool for the detection and identification of many species of bacteria. The aim of this review is to illustrate the application of the most commonly used PCR approaches to X. fastidiosa study, ranging from classical PCR, to several PCR-based detection methods: random amplified polymorphic DNA (RAPD), quantitative real-time PCR (qRT-PCR), nested-PCR (N-PCR), immunocapture PCR (IC-PCR), short sequence repeats (SSRs, also called VNTR), single nucleotide polymorphisms (SNPs) and multilocus sequence typing (MLST). Amplification and sequence analysis of specific

  4. Xylella fastidiosa: Host Range and Advance in Molecular Identification Techniques

    Directory of Open Access Journals (Sweden)

    Paolo Baldi

    2017-06-01

    Full Text Available In the never ending struggle against plant pathogenic bacteria, a major goal is the early identification and classification of infecting microorganisms. Xylella fastidiosa, a Gram-negative bacterium belonging to the family Xanthmonadaceae, is no exception as this pathogen showed a broad range of vectors and host plants, many of which may carry the pathogen for a long time without showing any symptom. Till the last years, most of the diseases caused by X. fastidiosa have been reported from North and South America, but recently a widespread infection of olive quick decline syndrome caused by this fastidious pathogen appeared in Apulia (south-eastern Italy, and several cases of X. fastidiosa infection have been reported in other European Countries. At least five different subspecies of X. fastidiosa have been reported and classified: fastidiosa, multiplex, pauca, sandyi, and tashke. A sixth subspecies (morus has been recently proposed. Therefore, it is vital to develop fast and reliable methods that allow the pathogen detection during the very early stages of infection, in order to prevent further spreading of this dangerous bacterium. To this purpose, the classical immunological methods such as ELISA and immunofluorescence are not always sensitive enough. However, PCR-based methods exploiting specific primers for the amplification of target regions of genomic DNA have been developed and are becoming a powerful tool for the detection and identification of many species of bacteria. The aim of this review is to illustrate the application of the most commonly used PCR approaches to X. fastidiosa study, ranging from classical PCR, to several PCR-based detection methods: random amplified polymorphic DNA (RAPD, quantitative real-time PCR (qRT-PCR, nested-PCR (N-PCR, immunocapture PCR (IC-PCR, short sequence repeats (SSRs, also called VNTR, single nucleotide polymorphisms (SNPs and multilocus sequence typing (MLST. Amplification and sequence analysis of

  5. Conjugative Plasmid Transfer in Xylella fastidiosa Is Dependent on tra and trb Operon Functions.

    Science.gov (United States)

    Burbank, Lindsey P; Van Horn, Christopher R

    2017-11-01

    The insect-transmitted plant pathogen Xylella fastidiosa is capable of efficient horizontal gene transfer (HGT) and recombination. Natural transformation occurs at high rates in X. fastidiosa , but there also is evidence that certain strains of X. fastidiosa carry native plasmids equipped with transfer and mobilization genes, suggesting conjugation as an additional mechanism of HGT in some instances. Two operons, tra and trb , putatively encoding a conjugative type IV secretion system, are found in some but not all X. fastidiosa isolates, often on native plasmids. X. fastidiosa strains that carry the conjugative transfer genes can belong to different subspecies and frequently differ in host ranges. Using X. fastidiosa strain M23 ( X. fastidiosa subsp. fastidiosa ) or Dixon ( X. fastidiosa subsp. multiplex ) as the donor strain and Temecula ( X. fastidiosa subsp. fastidiosa ) as the recipient strain, plasmid transfer was characterized using the mobilizable broad-host-range vector pBBR5pemIK. Transfer of plasmid pBBR5pemIK was observed under in vitro conditions with both donor strains and was dependent on both tra and trb operon functions. A conjugative mechanism likely contributes to gene transfer between diverse strains of X. fastidiosa , possibly facilitating adaptation to new environments or different hosts. IMPORTANCE Xylella fastidiosa is an important plant pathogen worldwide, infecting a wide range of different plant species. The emergence of new diseases caused by X. fastidiosa , or host switching of existing strains, is thought to be primarily due to the high frequency of HGT and recombination in this pathogen. Transfer of plasmids by a conjugative mechanism enables movement of larger amounts of genetic material at one time, compared with other routes of gene transfer such as natural transformation. Establishing the prevalence and functionality of this mechanism in X. fastidiosa contributes to a better understanding of HGT, adaptation, and disease emergence

  6. Evaluation of assembling methods on determination of whole genome sequence of Xylella fastidiosa blueberry bacterial leaf scorch strain

    Science.gov (United States)

    Blueberry bacterial leaf scorch (BBLS) disease, a threat to blueberry production in the Southern USA and potentially elsewhere, is caused by Xylella fastidiosa. Efficient control of BBLS requires knowledge of the pathogen. However, this is challenging because Xylella fastidiosa is difficult to cultu...

  7. Genetic analysis of a novel Xylella fastidiosa subspecies found in the southwestern United States.

    Science.gov (United States)

    Randall, Jennifer J; Goldberg, Natalie P; Kemp, John D; Radionenko, Maxim; French, Jason M; Olsen, Mary W; Hanson, Stephen F

    2009-09-01

    Xylella fastidiosa, the causal agent of several scorch diseases, is associated with leaf scorch symptoms in Chitalpa tashkentensis, a common ornamental landscape plant used throughout the southwestern United States. For a number of years, many chitalpa trees in southern New Mexico and Arizona exhibited leaf scorch symptoms, and the results from a regional survey show that chitalpa trees from New Mexico, Arizona, and California are frequently infected with X. fastidiosa. Phylogenetic analysis of multiple loci was used to compare the X. fastidiosa infecting chitalpa strains from New Mexico, Arizona, and trees imported into New Mexico nurseries with previously reported X. fastidiosa strains. Loci analyzed included the 16S ribosome, 16S-23S ribosomal intergenic spacer region, gyrase-B, simple sequence repeat sequences, X. fastidiosa-specific sequences, and the virulence-associated protein (VapD). This analysis indicates that the X. fastidiosa isolates associated with infected chitalpa trees in the Southwest are a highly related group that is distinct from the four previously defined taxons X. fastidiosa subsp. fastidiosa (piercei), X. fastidiosa subsp. multiplex, X. fastidiosa subsp. sandyi, and X. fastidiosa subsp. pauca. Therefore, the classification proposed for this new subspecies is X. fastidiosa subsp. tashke.

  8. The induction of differentially expressed proteins of Xylella fastidiosa with citrus extract Indução de proteínas de Xylella fastidiosa expressas diferencialmente com extrato de citros

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    Cláudia de M. Bellato

    2004-09-01

    Full Text Available An in vitro system was developed to induce and identify Xylella fastidiosa proteins that were differentially expressed in the presence of callus-derived extracts from its host, the citrus cultivar Pêra. To optimize the induction, we first developed a single culture medium for the growth of both, host and bacteria. This medium, CPXPm7, which mimics the citrus xylem sap, showed that X. fastidiosa at 72 h post-incubation had 10(8 colony forming units mL-1, while Pêra cells had the highest fresh weight content (0.79 g. After testing various methods of co-cultivation of the bacteria and host callus grown in this single medium, the best induction procedure was to grow X. fastidiosa in a solid medium amended with an extract of Pêra callus grown in CPXPm7. Analysis, by two-dimensional electrophoresis, of the X. fastidiosa proteins (120 µg of total proteins grown in the presence of Pêra callus extract revealed 414 differentially expressed protein spots when compared to the protein profile obtained in the absence of the extract. The system developed in this study improves the induction and analysis of differentially expressed proteins of X. fastidiosa, which may be involved in pathogenicity.Estudos in vitro foram desenvolvidos para obter proteínas de Xylella fastidiosa expressas diferencialmente na presença de calos do hospedeiro, citros cultivar Pêra. Para otimizar a indução, desenvolveu-se um meio de cultura comum, o qual foi baseado na seiva do xilema de citros, para cultivar a bacteria e os calos de Pêra. Dados mostraram, após 72 h de cultivo neste meio, 10(8 unidades formadoras de colônias de X. fastidiosa por mL, e 0,79 g de peso seco de células de Pêra. Após testar diferentes métodos de co-cultivo da bactéria com calos de Pêra neste meio, observou-se que a melhor taxa de indução ocorreu quando X. fastidiosa foi cultivada em meio sólido enriquecido com um extrato derivado dos calos de Pêra. Análise em gel bidimensional (2DE

  9. Diversity Evaluation of Xylella fastidiosa from Infected Olive Trees in Apulia (Southern Italy

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    Stefania M. Mang

    2016-04-01

    Full Text Available Olive culture is very important in the Mediterranean Basin. A severe outbreak of Olive Quick Decline Syndrome (OQDS caused by Xylella fastidiosa infection was first noticed in 2013 on olive trees in the southern part of Apulia region (Lecce province, southern Italy. Studies were carried out for detection and diversity evaluation of the Apulian strain of Xylella fastidiosa. The presence of the pathogen in olive samples was detected by PCR amplifying the 16S rDNA, gyrase B subunit (gyrB and HL hypothetical protein genes and single nucleotide polymorphisms (SNPs assessment was performed to genotype X. fastidiosa. Twelve SNPs were recorded over gyrB and six SNPs were found for HL gene. Less variations were detected on 16S rDNA gene. Only gyrB and HL provided sufficient information for dividing the Apulian X. fastidiosa olive strains into subspecies. Using HL nucleotide sequences was possible to separate X. fastidiosa into subspecies pauca and fastidiosa. Whereas, nucleotide variation present on gyrB gene allowed separation of X. fastidiosa subsp. pauca from the other subspecies multiplex and fastidiosa. The X. fastidiosa strain from Apulia region was included into the subspecies pauca based on three genes phylogenetic analyses.

  10. Analysis of the biofilm proteome of Xylella fastidiosa

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    Labate Carlos A

    2011-09-01

    Full Text Available Abstract Background Xylella fastidiosa is limited to the xylem of the plant host and the foregut of insect vectors (sharpshooters. The mechanism of pathogenicity of this bacterium differs from other plant pathogens, since it does not present typical genes that confer specific interactions between plant and pathogens (avr and/or hrp. The bacterium is injected directly into the xylem vessels where it adheres and colonizes. The whole process leads to the formation of biofilms, which are considered the main mechanism of pathogenicity. Cells in biofilms are metabolically and phenotypically different from their planktonic condition. The mature biofilm stage (phase of higher cell density presents high virulence and resistance to toxic substances such as antibiotics and detergents. Here we performed proteomic analysis of proteins expressed exclusively in the mature biofilm of X. fastidiosa strain 9a5c, in comparison to planktonic growth condition. Results We found a total of 456 proteins expressed in the biofilm condition, which correspond to approximately 10% of total protein in the genome. The biofilm showed 37% (or 144 proteins different protein than we found in the planktonic growth condition. The large difference in protein pattern in the biofilm condition may be responsible for the physiological changes of the cells in the biofilm of X. fastidiosa. Mass spectrometry was used to identify these proteins, while real-time quantitative polymerase chain reaction monitored expression of genes encoding them. Most of proteins expressed in the mature biofilm growth were associated with metabolism, adhesion, pathogenicity and stress conditions. Even though the biofilm cells in this work were not submitted to any stress condition, some stress related proteins were expressed only in the biofilm condition, suggesting that the biofilm cells would constitutively express proteins in different adverse environments. Conclusions We observed overexpression of proteins

  11. Xylella fastidiosa requires polygalacturonase for colonization and pathogenicity in Vitis vinifera grapevines.

    Science.gov (United States)

    Roper, M Caroline; Greve, L Carl; Warren, Jeremy G; Labavitch, John M; Kirkpatrick, Bruce C

    2007-04-01

    Xylella fastidiosa is the causal agent of Pierce's disease of grape, an economically significant disease for the grape industry. X. fastidiosa systemically colonizes the xylem elements of grapevines and is able to breach the pit pore membranes separating xylem vessels by unknown mechanisms. We hypothesized that X. fastidiosa utilizes cell wall degrading enzymes to break down pit membranes, based on the presence of genes involved in plant cell wall degradation in the X. fastidiosa genome. These genes include several beta-1,4 endoglucanases, several xylanases, several xylosidases, and one polygalacturonase (PG). In this study, we demonstrated that the pglA gene encodes a functional PG. A mutant in pglA lost pathogenicity and was compromised in its ability to systemically colonize Vitis vinifera grapevines. The results indicate that PG is required for X. fastidiosa to successfully infect grapevines and is a critical virulence factor for X. fastidiosa pathogenesis in grapevine.

  12. The endophyte Curtobacterium flaccumfaciens reduces symptoms caused by Xylella fastidiosa in Catharanthus roseus.

    Science.gov (United States)

    Lacava, Paulo Teixeira; Li, Wenbin; Araújo, Welington Luiz; Azevedo, João Lúcio; Hartung, John Stephen

    2007-10-01

    Citrus variegated chlorosis (CVC) is a disease of the sweet orange [Citrus sinensis (L.)], which is caused by Xylella fastidiosa subsp. pauca, a phytopathogenic bacterium that has been shown to infect all sweet orange cultivars. Sweet orange trees have been occasionally observed to be infected by Xylella fastidiosa without evidencing severe disease symptoms, whereas other trees in the same grove may exhibit severe disease symptoms. The principal endophytic bacterial species isolated from such CVC-asymptomatic citrus plants is Curtobacterium flaccumfaciens. The Madagascar periwinkle [Citrus sinensis (L.)] is a model plant which has been used to study X. fastidiosa in greenhouse environments. In order to characterize the interactions of X. fastidiosa and C. flaccumfaciens, periwinkle plants were inoculated separately with C. flaccumfaciens, X. fastidiosa, and both bacteria together. The number of flowers produced by the plants, the heights of the plants, and the exhibited disease symptoms were evaluated. PCR-primers for C. flaccumfaciens were designed in order to verify the presence of this endophytic bacterium in plant tissue, and to complement an existing assay for X. fastidiosa. These primers were capable of detecting C. flaccumfaciens in the periwinkle in the presence of X. fastidiosa. X. fastidiosa induced stunting and reduced the number of flowers produced by the periwinkle. When C. flaccumfaciens was inoculated together with X. fastidiosa, no stunting was observed. The number of flowers produced by our doubly- inoculated plants was an intermediate between the number produced by the plants inoculated with either of the bacteria separately. Our data indicate that C. flaccumfaciens interacted with X. fastidiosa in C. roseus, and reduced the severity of the disease symptoms induced by X. fastidiosa. Periwinkle is considered to be an excellent experimental system by which the interaction of C. flaccumfaciens and other endophytic bacteria with X. fastidiosa can be

  13. Contribution of rpfB to cell-to-cell signal synthesis, virulence, and vector transmission of Xylella fastidiosa.

    Science.gov (United States)

    Almeida, Rodrigo P P; Killiny, Nabil; Newman, Karyn L; Chatterjee, Subhadeep; Ionescu, Michael; Lindow, Steven E

    2012-04-01

    In Xylella fastidiosa the fatty acid signal molecule diffusible signaling factor (DSF) is produced and sensed by components of the regulation of pathogenicity factors (rpf) cluster; lack of DSF production in RpfF mutants results in a non-vector-transmissible phenotype yet cells are hypervirulent to grape. rpfB has not been characterized in Xylella fastidiosa, although its homolog has been suggested to be required for DSF synthesis in Xanthomonas campestris pv. campestris. We show that RpfB is involved in DSF processing in both Xylella fastidiosa and Xanthomonas campestris, affecting the profile of DSF-like fatty acids observed in thin-layer chromatography. Although three fatty acids whose production is dependent on RpfF were detected in Xylella fastidiosa and Xanthomonas campestris wild-type strains, their respective rpfB mutants accumulated primarily one chemical species. Although no quantifiable effect of rpfB on plant colonization by Xylella fastidiosa was found, insect colonization and transmission was reduced. Thus, RpfB apparently is involved in DSF processing, and like Xanthomonas campestris, Xylella fastidiosa also produces multiple DSF molecules. It is possible that Xylella fastidiosa coordinates host vector and plant colonization by varying the proportions of different forms of DSF signals via RpfB.

  14. RpfF-dependent regulon of Xylella fastidiosa.

    Science.gov (United States)

    Wang, Nian; Li, Jian-Liang; Lindow, Steven E

    2012-11-01

    ABSTRACT Xylella fastidiosa regulates traits important to both virulence of grape as well as colonization of sharpshooter vectors via its production of a fatty acid signal molecule known as DSF whose production is dependent on rpfF. Although X. fastidiosa rpfF mutants exhibit increased virulence to plants, they are unable to be spread from plant to plant by insect vectors. To gain more insight into the traits that contribute to these processes, a whole-genome Agilent DNA microarray for this species was developed and used to determine the RpfF-dependent regulon by transcriptional profiling. In total, 446 protein coding genes whose expression was significantly different between the wild type and an rpfF mutant (false discovery rate genes were downregulated in the rpfF mutant compared with the wild-type strain whereas 281 genes were over-expressed. RpfF function was required for regulation of 11 regulatory and σ factors, including rpfE, yybA, PD1177, glnB, rpfG, PD0954, PD0199, PD2050, colR, rpoH, and rpoD. In general, RpfF is required for regulation of genes involved in attachment and biofilm formation, enhancing expression of hemagglutinin genes hxfA and hxfB, and suppressing most type IV pili and gum genes. A large number of other RpfF-dependent genes that might contribute to virulence or insect colonization were also identified such as those encoding hemolysin and colicin V, as well as genes with unknown functions.

  15. Xylella fastidiosa: an examination of a re-emerging plant pathogen.

    Science.gov (United States)

    Rapicavoli, Jeannette; Ingel, Brian; Blanco-Ulate, Barbara; Cantu, Dario; Roper, Caroline

    2018-04-01

    Xylella fastidiosa is a Gram-negative bacterial plant pathogen with an extremely wide host range. This species has recently been resolved into subspecies that correlate with host specificity. This review focuses on the status of X. fastidiosa pathogenic associations in plant hosts in which the bacterium is either endemic or has been recently introduced. Plant diseases associated with X. fastidiosa have been documented for over a century, and much about what is known in the context of host-pathogen interactions is based on these hosts, such as grape and citrus, in which this pathogen has been well described. Recent attention has focused on newly emerging X. fastidiosa diseases, such as in olives. Bacteria; Gammaproteobacteria; family Xanthomonadaceae; genus Xylella; species fastidiosa. Gram-negative rod (0.25-0.35 × 0.9-3.5 μm), non-flagellate, motile via Type IV pili-mediated twitching, fastidious. Xylella fastidiosa has a broad host range that includes ornamental, ecological and agricultural plants belonging to over 300 different species in 63 different families. To date, X. fastidiosa has been found to be pathogenic in over 100 plant species. In addition, it can establish non-symptomatic associations with many plants as a commensal endophyte. Here, we list the four distinct subspecies of X. fastidiosa and some of the agriculturally relevant diseases caused by them: X. fastidiosa ssp. fastidiosa causes Pierce's disease (PD) of grapevine (Vitis vinifera); X. fastidiosa ssp. multiplex causes almond leaf scorch (ALS) and diseases on other nut and shade tree crops; X. fastidiosa ssp. pauca causes citrus variegated chlorosis (CVC) (Citrus spp.), coffee leaf scorch and olive quick decline syndrome (OQDS) (Olea europaea); X. fastidiosa ssp. sandyi causes oleander leaf scorch (OLS) (Nerium oleander). Significant host specificity seemingly exists for some of the subspecies, although this could be a result of technical biases based on the limited number of

  16. Promiscuous Diffusible Signal Factor Production and Responsiveness of the Xylella fastidiosa Rpf System

    OpenAIRE

    Ionescu, Michael; Yokota, Kenji; Antonova, Elena; Garcia, Angelica; Beaulieu, Ellen; Hayes, Terry; Iavarone, Anthony T.; Lindow, Steven E.

    2016-01-01

    © 2016 Ionescu et al. Cell density-dependent regulation of gene expression in Xylella fastidiosa that is crucial to its switching between plant hosts and insect vectors is dependent on RpfF and its production of 2-enoic acids known as diffusible signal factor (DSF). We show that X. fastidiosa produces a particularly large variety of similar, relatively long-chain-length 2-enoic acids that are active in modulating gene expression. Both X. fastidiosa itself and a Pantoea agglomerans surrogate h...

  17. Multilocus sequence typing of Xylella fastidiosa causing Pierce's disease and oleander leaf scorch in the United States.

    Science.gov (United States)

    Yuan, Xiaoli; Morano, Lisa; Bromley, Robin; Spring-Pearson, Senanu; Stouthamer, Richard; Nunney, Leonard

    2010-06-01

    Using a modified multilocus sequence typing (MLST) scheme for the bacterial plant pathogen Xylella fastidiosa based on the same seven housekeeping genes employed in a previously published MLST, we studied the genetic diversity of two subspecies, X. fastidiosa subsp. fastidiosa and X. fastidiosa subsp. sandyi, which cause Pierce's disease and oleander leaf scorch, respectively. Typing of 85 U.S. isolates (plus one from northern Mexico) of X. fastidiosa subsp. fastidiosa from 15 different plant hosts and 21 isolates of X. fastidiosa subsp. sandyi from 4 different hosts in California and Texas supported their subspecific status. Analysis using the MLST genes plus one cell-surface gene showed no significant genetic differentiation based on geography or host plant within either subspecies. Two cases of homologous recombination (with X. fastidiosa subsp. multiplex, the third U.S. subspecies) were detected in X. fastidiosa subsp. fastidiosa. Excluding recombination, MLST site polymorphism in X. fastidiosa subsp. fastidiosa (0.048%) and X. fastidiosa subsp. sandyi (0.000%) was substantially lower than in X. fastidiosa subsp. multiplex (0.240%), consistent with the hypothesis that X. fastidiosa subspp. fastidiosa and sandyi were introduced into the United States (probably just prior to 1880 and 1980, respectively). Using whole-genome analysis, we showed that MLST is more effective at genetic discrimination at the specific and subspecific level than other typing methods applied to X. fastidiosa. Moreover, MLST is the only technique effective in detecting recombination.

  18. Reação de variedades e clones de laranjas a Xylella fastidiosa Reaction of oranges varieties and clones to Xylella fastidiosa

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    Paulo Sergio de Souza

    2006-04-01

    Full Text Available A clorose variegada dos citros (CVC é uma doença grave, causada por Xylella fastidiosa. As medidas usuais de controle mostram-se pouco eficientes ou práticas e com alto custo. Dessa forma, o uso de variedades resistentes e/ou tolerantes desponta como a alternativa mais eficiente, razão pela qual se julgou oportuna a realização deste trabalho. O presente estudo teve como objetivo avaliar o comportamento de variedades e clones de laranjas introduzidas em relação a X. fastidiosa. Foram estudados 59 variedades e clones de laranjas doces e 2 de laranjas azedas introduzidos da França, Itália e Portugal. O delineamento experimental foi o de blocos casualizados (DBC, com 62 tratamentos e 4 repetições, incluindo a variedade 'Pêra', como padrão. Cada parcela continha duas plantas, sendo uma inoculada e a outra sem inoculação. Para a inoculação do patógeno, foi empregado o método de encostia, utilizando-se de mudas infectadas. Para a avaliação da incidência da doença, utilizou-se de dados qualitativos, positivos ou negativos, enquanto para severidade empregou-se escala de notas, que foi estabelecida baseando-se nos sintomas de CVC, confirmados através dos testes de PCR. As variedades de laranjas azedas Beja e Sr. Pinto e as laranjas doces Navelina ISA 315, Navelina SRA 332 e Newhall Navel SRA 343 não apresentaram sintomas em folhas até 27 meses após a inoculação.Citrus variegated chlorosis (CVC is a serious disease, caused by Xylella fastidiosa, being the most important to the Brazilian citriculture. The usual measures of control show less efficient or practice and they are very expensive. The use of resistant varieties is showed as the most efficient, consisting the aimed of this research. The objective of the present work was study the behavior of varieties and clones introduced in relation to X. fastidiosa. Were studied 59 varieties and clones of sweet oranges and 2 sour oranges introduced from France, Italy and Portugal

  19. RNA metabolism in Xylella fastidiosa during cold adaptation and survival responses

    Science.gov (United States)

    Fastidious plant pathogen Xylella fastidiosa has a reduced ability to adapt to cold temperatures, limiting persistence in perennial hosts, such as grapevine, growing in colder regions. RNA metabolism is an essential part of bacterial response to low temperature, including inducible expression of RNA...

  20. Identification of a low copy number plasmid in Xylella fastidiosa Strain Stag’s Leap

    Science.gov (United States)

    Xylella fastidiosa (Xf) causes Pierce’s Disease (PD) in grapevine. The Stag’s Leap strain is known for its high virulence level and is a model for PD research. Research on Xf has been difficult due to its nutritional fastidiousness. One difficult research issue is the low copy number plasmid. Plasmi...

  1. The DinJ/RelE toxin-antitoxin system suppresses virulence in Xylella fastidiosa

    Science.gov (United States)

    Xylella fastidiosa, the causal agent of a number agriculturally important plant diseases, encodes multiple toxin-antitoxin (TA) systems. TA modules consist of a toxin protein co-expressed with a specific antitoxin, and are often acquired through horizontal gene transfer. Antitoxin molecules (RNA or ...

  2. The role of Xylella fastidiosa cold shock proteins in Pierce’s disease of grapes

    Science.gov (United States)

    Pierce’s disease of grapevine, caused by the bacterial pathogen Xylella fastidiosa (Xf) is limited to warmer climates, and plant infection can be eliminated by cold winter conditions. Milder winters can increase the likelihood of pathogen persistence from one growing season to the next. Cold adaptat...

  3. Functional characterization of the role of rpfA in Xylella fastidiosa

    Science.gov (United States)

    Xylella fastidiosa coordinates virulence in grapevines via quorum sensing signal molecules that are regulated and synthesized by the rpf gene cluster (regulation of pathogenicity factors). rpfA encodes aconitate hydratase and could play a regulator role involved in virulence. To elucidate the role o...

  4. Genomic insights into Xylella fastidiosa interactions with plant and insect hosts

    Science.gov (United States)

    Utilization of genomic data and widespread availability of genomics tools are still incipient in plant pathology. The first genome of a plant pathogen, the bacterium Xylella fastidiosa, was only completed in 2000. Since then, a large number of bacterial plant pathogens have been sequenced, but much ...

  5. Draft genome sequence of Xylella fastidiosa pear leaf scorch strain in Taiwan

    Science.gov (United States)

    The draft genome sequence of Xylella fastidiosa pear leaf scorch strain (PLS229) isolated from pear cultivar Hengshan (Pyrus pyrifolia) in Taiwan is reported. The bacterium has a genome size of 2,733,013 bp with a G+C content of 53.1%. The PLS229 strain genome was annotated to have 3,259 open readin...

  6. Distribution of Xylella fastidiosa in Sycamore associated with low temperature and host resistance

    Science.gov (United States)

    T.S.M. Henneberger; K.L. Stevenson; C.J. Chang

    2004-01-01

    Experiments were conducted in the field and laboratory to determine effects of low temperatures 4% on Xylella fastidiosa populations in American sycamore. Roots and shoots from naturally infected trees at two locations were collected monthly. Sap extracted from the samples was tested by enzyme-linked immunosorbent assay for presence of X...

  7. Probe-based real-time PCR method for multilocus melt typing of Xylella fastidiosa strains.

    Science.gov (United States)

    Brady, Jeff A; Faske, Jennifer B; Ator, Rebecca A; Castañeda-Gill, Jessica M; Mitchell, Forrest L

    2012-04-01

    Epidemiological studies of Pierce's disease (PD) can be confounded by a lack of taxonomic detail on the bacterial causative agent, Xylella fastidiosa (Xf). PD in grape is caused by strains of Xylella fastidiosa subsp. fastidiosa, but is not caused by other subspecies of Xf that typically colonize plants other than grape. Detection assays using ELISA and qPCR are effective at detecting and quantifying Xf presence or absence, but offer no information on Xf subspecies or strain identity. Surveying insects or host plants for Xf by current ELISA or qPCR methods provides only presence/absence and quantity information for any and all Xf subspecies, potentially leading to false assessments of disease threat. This study uses a series of adjacent-hybridizing DNA melt analysis probes that are capable of efficiently discriminating Xf subspecies and strain relationships in rapid real-time PCR reactions. Copyright © 2012 Elsevier B.V. All rights reserved.

  8. Evaluation of Xylella fastidiosa genetic diversity by fAFPL markers Diversidade genética de Xylella fastidiosa avaliada por marcadores fAFPL

    Directory of Open Access Journals (Sweden)

    Luciano Takeshi Kishi

    2008-03-01

    Full Text Available The first phytopathogenic bacterium with its DNA entirely sequenced is being detected and isolated from different host plants in several geographic regions. Although it causes diseases in cultures of economic importance, such as citrus, coffee, and grapevine little is known about the genetic relationships among different strains. Actually, all strains are grouped as a single species, Xylella fastidiosa, despite colonizing different hosts, developing symptoms, and different physiological and microbiological observed conditions. The existence of genetic diversity among X. fastidiosa strains was detected by different methodological techniques, since cultural to molecular methods. However, little is know about the phylogenetic relationships developed by Brazilian strains obtained from coffee and citrus plants. In order to evaluate it, fAFLP markers were used to verify genetic diversity and phylogenetic relationships developed by Brazilian and strange strains. fAFLP is an efficient technique, with high reproducibility that is currently used for bacterial typing and classification. The obtained results showed that Brazilian strains present genetic diversity and that the strains from this study were grouped distinctly according host and geographical origin like citrus-coffee, temecula-grapevine-mulberry and plum-elm.A primeira bactéria fitopatogênica a ter seu genoma totalmente seqüenciado foi detectada e isolada em diferentes hospedeiros em diferentes regiões geográficas. Embora seja causadora de doenças em culturas economicamente importantes, como citros, cafeeiro e videira, pouco se conhece acerca das relações genéticas estabelecidas entre isolados da bactéria. Atualmente, todos os isolados são agrupados como uma única espécie, Xylella fastidiosa, apesar de colonizarem diferentes hospedeiros que desenvolvem sintomas diferenciados e possuir diferentes condições fisiológicas e microbiológicas. A existência de diversidade gen

  9. Genetic characterization of Xylella fastidiosa isolated from citrus and coffee plants Caracterização genética de Xylella fastidiosa isolada de plantas de citros e café

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    Vicente Savonitti Miranda

    2007-10-01

    Full Text Available The Citrus Variegated Chlorosis and the Coffee Leaf Scorch are some of the many destructive diseases caused by Xylella fastidiosa, a gram-negative bacterium limited to the xylem of affected plants. As its genetic characterization is still not well established, different isolates of X. fastidiosa from citrus and coffee were evaluated through RAPD (Random Amplified Polymorphic DNA technique to characterize and classify these isolates based on similarity coefficients. Sixteen isolates of X. fastidiosa were used on this trial, obtained from citrus, coffee and almond. The genetic polymorphism evaluation was performed using six arbitrary 10-base primer pairs. It was possible to establish a dendogram in which the isolates were classified into five groups (A, B, C, D and E. A prevalence of citrus isolates in groups A and D was observed. In groups B and C, there was a prevalence of coffee isolates meanwhile the group D consisted of the almond isolate, solely.A Clorose Variegada dos Citros e a Requeima das Folhas do Cafeeiro são algumas das várias doenças destrutivas causadas pela Xylella fastidiosa, que é uma bactéria gram-negativa e limitada ao xilema de plantas afetadas. Como a sua caracterização genética ainda não está determinada, diferentes isolados da X. fastidiosa de citros e café foram avaliados pela técnica RAPD (Polimorfismo do DNA Amplificado ao Acaso para caracterizar e classificar estes isolados com base em coeficientes de similaridade. Foram utilizados 16 isolados de X. fastidiosa provenientes de citros, café e amêndoa. A avaliação do polimorfismo genético foi realizada utilizando seis iniciadores randômicos de 10 pares de base. Foi possível estabelecer um dendograma no qual os isolados foram classificados em cinco grupos (A, B, C, D e E. Nos grupos A e D existe uma forte predominância de isolados de citros. Nos grupos B e C há predominância de isolados de café enquanto no grupo E ficou apenas o isolado de amêndoa.

  10. Diversity of Xylella fastidiosa host suitability among siblings from a non-traditional almond X peach cross

    Science.gov (United States)

    Ten F2 clones from an initial hybridization of Prunus webbii X P. persica cv Harrow Blood were evaluated under greenhouse conditions for their reaction to Xylella fastidiosa subsp. fastidiosa strain M23 during two growing seasons. Clonal accessions used for the study were selected on the basis of ho...

  11. Genome-wide comparison and taxonomic relatedness of multiple Xylella fastidiosa strains reveal the occurrence of three subspecies and a new Xylella species.

    Science.gov (United States)

    Marcelletti, Simone; Scortichini, Marco

    2016-10-01

    A total of 21 Xylella fastidiosa strains were assessed by comparing their genomes to infer their taxonomic relationships. The whole-genome-based average nucleotide identity and tetranucleotide frequency correlation coefficient analyses were performed. In addition, a consensus tree based on comparisons of 956 core gene families, and a genome-wide phylogenetic tree and a Neighbor-net network were constructed with 820,088 nucleotides (i.e., approximately 30-33 % of the entire X. fastidiosa genome). All approaches revealed the occurrence of three well-demarcated genetic clusters that represent X. fastidiosa subspecies fastidiosa, multiplex and pauca, with the latter appeared to diverge. We suggest that the proposed but never formally described subspecies 'sandyi' and 'morus' are instead members of the subspecies fastidiosa. These analyses support the view that the Xylella strain isolated from Pyrus pyrifolia in Taiwan is likely to be a new species. A widely used multilocus sequence typing analysis yielded conflicting results.

  12. Transcriptome analysis of the phytobacterium Xylella fastidiosa growing under xylem-based chemical conditions.

    Science.gov (United States)

    Ciraulo, Maristela Boaceff; Santos, Daiene Souza; Rodrigues, Ana Claudia de Freitas Oliveira; de Oliveira, Marcus Vinícius; Rodrigues, Tiago; de Oliveira, Regina Costa; Nunes, Luiz R

    2010-01-01

    Xylella fastidiosa is a xylem-limited bacterium responsible for important plant diseases, like citrus-variegated chlorosis (CVC) and grapevine Pierce's disease (PD). Interestingly, in vitro growth of X. fastidiosa in chemically defined media that resemble xylem fluid has been achieved, allowing studies of metabolic processes used by xylem-dwelling bacteria to thrive in such nutrient-poor conditions. Thus, we performed microarray hybridizations to compare transcriptomes of X. fastidiosa cells grown in 3G10-R, a medium that resembles grape sap, and in Periwinkle Wilt (PW), the complex medium traditionally used to cultivate X. fastidiosa. We identified 299 transcripts modulated in response to growth in these media. Some 3G10R-overexpressed genes have been shown to be upregulated in cells directly isolated from infected plants and may be involved in plant colonization, virulence and environmental competition. In contrast, cells cultivated in PW show a metabolic switch associated with increased aerobic respiration and enhanced bacterial growth rates.

  13. Effect of oxygen on the growth and biofilm formation of Xylella fastidiosa in liquid media.

    Science.gov (United States)

    Shriner, Anthony D; Andersen, Peter C

    2014-12-01

    Xylella fastidiosa is a xylem-limited bacterial pathogen, and is the causative agent of Pierce's disease of grapevines and scorch diseases of many other plant species. The disease symptoms are putatively due to blocking of the transpiration stream by bacterial-induced biofilm formation and/or by the formation of plant-generated tylosis. Xylella fastidiosa has been classified as an obligate aerobe, which appears unusual given that dissolved O2 levels in the xylem during the growing season are often hypoxic (20-60 μmol L(-1)). We examined the growth and biofilm formation of three strains of X. fastidiosa under variable O2 conditions (21, 2.1, 0.21 and 0 % O2), in comparison to that of Pseudomonas syringae (obligate aerobe) and Erwinia carotovora (facultative anaerobe) under similar conditions. The growth of X. fastidiosa more closely resembled that of the facultative anaerobe, and not the obligate aerobe. Xanthomonas campestris, the closest genetic relative of X. fastidiosa, exhibited no growth in an N2 environment, whereas X. fastidiosa was capable of growing in an N2 environment in PW(+), CHARDS, and XDM2-PR media. The magnitude of growth and biofilm formation in the N2 (0 % O2) treatment was dependent on the specific medium. Additional studies involving the metabolism of X. fastidiosa in response to low O2 are warranted. Whether X. fastidiosa is classified as an obligate aerobe or a facultative anaerobe should be confirmed by gene activation and/or the quantification of the metabolic profiles under hypoxic conditions.

  14. Differential expression of genes of Xylella fastidiosa in xylem fluid of citrus and grapevine.

    Science.gov (United States)

    Shi, Xiangyang; Bi, Jianlong; Morse, Joseph G; Toscano, Nick C; Cooksey, Donald A

    2010-03-01

    Xylella fastidiosa causes a serious Pierce's disease (PD) in grapevine. Xylella fastidiosa cells from a PD strain were grown in a pure xylem fluid of a susceptible grapevine cultivar vs. xylem fluid from citrus, which is not a host for this strain of X. fastidiosa. When grown in grapevine xylem fluid, cells of the PD strain formed clumps and biofilm formed to a greater extent than in citrus xylem fluid, although the PD strain did grow in xylem fluid of three citrus varieties. The differential expression of selected genes of a PD X. fastidiosa strain cultured in the two xylem fluids was analyzed using a DNA macroarray. Compared with citrus xylem fluid, grapevine xylem fluid stimulated the expression of X. fastidiosa genes involved in virulence regulation, such as gacA, algU, xrvA, and hsq, and also genes involved in the biogenesis of pili and twitching motility, such as fimT, pilI, pilU, and pilY1. Increased gene expression likely contributes to PD expression in grapevine, whereas citrus xylem fluid did not support or possibly suppressed the expression of these virulence genes.

  15. Quantification of Xylella fastidiosa from Citrus Trees by Real-Time Polymerase Chain Reaction Assay.

    Science.gov (United States)

    Oliveira, Antonio C; Vallim, Marcelo A; Semighini, Camile P; Araújo, Welington L; Goldman, Gustavo H; Machado, Marcos A

    2002-10-01

    ABSTRACT Xylella fastidiosa is the causal agent of citrus variegated chlorosis (CVC), a destructive disease of sweet orange cultivars in Brazil. Polymerase chain reaction (PCR)-based assays constitute the principal diagnostic method for detection of these bacteria. In this work, we established a real-time quantitative PCR (QPCR) assay to quantify X. fastidiosa in naturally and artificially infected citrus. The X. fastidiosa cell number detected in the leaves increased according to the age of the leaf, and bacteria were not detected in the upper midrib section in young leaves, indicating temporal and spatial distribution patterns of bacteria, respectively. In addition, the X. fastidiosa cell number quantified in leaves of 'Pera' orange and 'Murcott' tangor reflected the susceptible and resistant status of these citrus cultivars. None of the 12 endophytic citrus bacteria or the four strains of X. fastidiosa nonpathogenic to citrus that were tested showed an increase in the fluorescence signal during QPCR. In contrast, all 10 CVC-causing strains exhibited an increase in fluorescence signal, thus indicating the specificity of this QPCR assay. Our QPCR provides a powerful tool for studies of different aspects of the Xylella-citrus interactions, and can be incorporated into breeding programs in order to select CVC-resistant plants more quickly.

  16. Evaluation of Arabidopsis thaliana as a model host for Xylella fastidiosa.

    Science.gov (United States)

    Rogers, Elizabeth E

    2012-06-01

    The bacterium Xylella fastidiosa causes a number of plant diseases of significant economic impact. To date, progress determining mechanisms of host-plant susceptibility, tolerance, or resistance has been slow, due in large part to the long generation time and limited available genetic resources for grape, almond, and other known hosts of X. fastidiosa. To overcome many of these limitations, Arabidopsis thaliana has been evaluated as a host for X. fastidiosa. A pin-prick inoculation method has been developed to infect Arabidopsis with X. fastidiosa. Following infection, X. fastidiosa multiplies and can be detected by microscopy, polymerase chain reaction, and isolation. The ecotypes Van-0, LL-0, and Tsu-1 all allow more growth of strain X. fastidiosa Temecula than the reference ecotype Col-0. Affymetrix ATH1 microarray analysis of inoculated vs. noninoculated Tsu-1 reveals gene expression changes that differ greatly from changes seen after infection with apoplast-colonizing bacteria such as Psuedomonas syringae pvs. tomato or syringae. Many genes responsive to oxidative stress are differentially regulated, while classic pathogenesis-related genes are not induced by X. fastidiosa infection.

  17. Isolation and pathogenicity of Xylella fastidiosa from grapevine and almond in Iran

    Directory of Open Access Journals (Sweden)

    Naser AMANIFAR

    2014-09-01

    Full Text Available Symptoms similar to those of Pierce’s disease (PD of grapevine and leaf scorch of almond were observed in vineyards and almond orchards in several provinces of Iran. Grafting of scions from symptomatic almond trees onto seedlings of a local almond (cv. Mamaee under greenhouse conditions resulted in the transmission of the leaf scorch agent. A number of symptomatic samples from orchard and greenhouse plants were positive for presence of Xylella fastidiosa when tested by DAS-ELISA and PCR with X. fastidiosa specific antibodies and primers. A Gram-negative bacterium similar to X. fastidiosa was isolated on ‘periwinkle wilt’ (PW medium. Selected isolates induced symptoms similar to those caused by X. fastidiosa when inoculated on Nicotiana tabacum, seedlings of almond and grapevine under greenhouse conditions. DAS-ELISA and PCR confirmed the identity of the isolated bacteria. On the basis of disease symptoms, graft transmission, isolation on specific X. fastidiosa culture medium, pathogenicity tests and positive reactions in DAS-ELISA and PCR, X. fastidiosa is associated with almond leaf scorch and Pierce’s disease in grapevine in Iran. This is the first report on the presence of X. fastidiosa in the Middle East and western Asia.

  18. Twitching motility and biofilm formation are associated with tonB1 in Xylella fastidiosa.

    Science.gov (United States)

    Cursino, Luciana; Li, Yaxin; Zaini, Paulo A; De La Fuente, Leonardo; Hoch, Harvey C; Burr, Thomas J

    2009-10-01

    A mutation in the Xylella fastidiosa tonB1 gene resulted in loss of twitching motility and in significantly less biofilm formation as compared with a wild type. The altered motility and biofilm phenotypes were restored by complementation with a functional copy of the gene. The mutation affected virulence as measured by Pierce's disease symptoms on grapevines. The role of TonB1 in twitching and biofilm formation appears to be independent of the characteristic iron-uptake function of this protein. This is the first report demonstrating a functional role for a tonB homolog in X. fastidiosa.

  19. Comparative genomic characterization of citrus-associated Xylella fastidiosa strains

    Directory of Open Access Journals (Sweden)

    Nunes Luiz R

    2007-12-01

    Full Text Available Abstract Background The xylem-inhabiting bacterium Xylella fastidiosa (Xf is the causal agent of Pierce's disease (PD in vineyards and citrus variegated chlorosis (CVC in orange trees. Both of these economically-devastating diseases are caused by distinct strains of this complex group of microorganisms, which has motivated researchers to conduct extensive genomic sequencing projects with Xf strains. This sequence information, along with other molecular tools, have been used to estimate the evolutionary history of the group and provide clues to understand the capacity of Xf to infect different hosts, causing a variety of symptoms. Nonetheless, although significant amounts of information have been generated from Xf strains, a large proportion of these efforts has concentrated on the study of North American strains, limiting our understanding about the genomic composition of South American strains – which is particularly important for CVC-associated strains. Results This paper describes the first genome-wide comparison among South American Xf strains, involving 6 distinct citrus-associated bacteria. Comparative analyses performed through a microarray-based approach allowed identification and characterization of large mobile genetic elements that seem to be exclusive to South American strains. Moreover, a large-scale sequencing effort, based on Suppressive Subtraction Hybridization (SSH, identified 290 new ORFs, distributed in 135 Groups of Orthologous Elements, throughout the genomes of these bacteria. Conclusion Results from microarray-based comparisons provide further evidence concerning activity of horizontally transferred elements, reinforcing their importance as major mediators in the evolution of Xf. Moreover, the microarray-based genomic profiles showed similarity between Xf strains 9a5c and Fb7, which is unexpected, given the geographical and chronological differences associated with the isolation of these microorganisms. The newly

  20. The complex biogeography of the plant pathogen Xylella fastidiosa: genetic evidence of introductions and Subspecific introgression in Central America.

    Science.gov (United States)

    Nunney, Leonard; Ortiz, Beatriz; Russell, Stephanie A; Ruiz Sánchez, Rebeca; Stouthamer, Richard

    2014-01-01

    The bacterium Xylella fastidiosa is a plant pathogen with a history of economically damaging introductions of subspecies to regions where its other subspecies are native. Genetic evidence is presented demonstrating the introduction of two new taxa into Central America and their introgression into the native subspecies, X. fastidiosa subsp. fastidiosa. The data are from 10 genetic outliers detected by multilocus sequence typing (MLST) of isolates from Costa Rica. Six (five from oleander, one from coffee) defined a new sequence type (ST53) that carried alleles at six of the eight loci sequenced (five of the seven MLST loci) diagnostic of the South American subspecies Xylella fastidiosa subsp. pauca which causes two economically damaging plant diseases, citrus variegated chlorosis and coffee leaf scorch. The two remaining loci of ST53 carried alleles from what appears to be a new South American form of X. fastidiosa. Four isolates, classified as X. fastidiosa subsp. fastidiosa, showed a low level of introgression of non-native DNA. One grapevine isolate showed introgression of an allele from X. fastidiosa subsp. pauca while the other three (from citrus and coffee) showed introgression of an allele with similar ancestry to the alleles of unknown origin in ST53. The presence of X. fastidiosa subsp. pauca in Central America is troubling given its disease potential, and establishes another route for the introduction of this economically damaging subspecies into the US or elsewhere, a threat potentially compounded by the presence of a previously unknown form of X. fastidiosa.

  1. The complex biogeography of the plant pathogen Xylella fastidiosa: genetic evidence of introductions and Subspecific introgression in Central America.

    Directory of Open Access Journals (Sweden)

    Leonard Nunney

    Full Text Available The bacterium Xylella fastidiosa is a plant pathogen with a history of economically damaging introductions of subspecies to regions where its other subspecies are native. Genetic evidence is presented demonstrating the introduction of two new taxa into Central America and their introgression into the native subspecies, X. fastidiosa subsp. fastidiosa. The data are from 10 genetic outliers detected by multilocus sequence typing (MLST of isolates from Costa Rica. Six (five from oleander, one from coffee defined a new sequence type (ST53 that carried alleles at six of the eight loci sequenced (five of the seven MLST loci diagnostic of the South American subspecies Xylella fastidiosa subsp. pauca which causes two economically damaging plant diseases, citrus variegated chlorosis and coffee leaf scorch. The two remaining loci of ST53 carried alleles from what appears to be a new South American form of X. fastidiosa. Four isolates, classified as X. fastidiosa subsp. fastidiosa, showed a low level of introgression of non-native DNA. One grapevine isolate showed introgression of an allele from X. fastidiosa subsp. pauca while the other three (from citrus and coffee showed introgression of an allele with similar ancestry to the alleles of unknown origin in ST53. The presence of X. fastidiosa subsp. pauca in Central America is troubling given its disease potential, and establishes another route for the introduction of this economically damaging subspecies into the US or elsewhere, a threat potentially compounded by the presence of a previously unknown form of X. fastidiosa.

  2. Differentiation of Xylella fastidiosa Strains via Multilocus Sequence Analysis of Environmentally Mediated Genes (MLSA-E)

    OpenAIRE

    Parker, Jennifer K.; Havird, Justin C.; De La Fuente, Leonardo

    2012-01-01

    Isolates of the plant pathogen Xylella fastidiosa are genetically very similar, but studies on their biological traits have indicated differences in virulence and infection symptomatology. Taxonomic analyses have identified several subspecies, and phylogenetic analyses of housekeeping genes have shown broad host-based genetic differences; however, results are still inconclusive for genetic differentiation of isolates within subspecies. This study employs multilocus sequence analysis of enviro...

  3. The exopolysaccharide of Xylella fastidiosa is essential for biofilm formation, plant virulence, and vector transmission

    OpenAIRE

    Killiny, N; Hernandez Martinez, R; Korsi Dumenyo, C; Cooksey, DA; Almeida, RPP

    2013-01-01

    Exopolysaccharides (EPS) synthesized by plant-pathogenic bacteria are generally essential for virulence. The role of EPS produced by the vector-transmitted bacterium Xylella fastidiosa was investigated by knocking out two genes implicated in the EPS biosynthesis, gumD and gumH. Mutant strains were affected in growth characteristics in vitro, including adhesion to surfaces and biofilm formation. In addition, different assays were used to demonstrate that the mutant strains produced significant...

  4. Promiscuous Diffusible Signal Factor Production and Responsiveness of the Xylella fastidiosa Rpf System

    Directory of Open Access Journals (Sweden)

    Michael Ionescu

    2016-07-01

    Full Text Available Cell density-dependent regulation of gene expression in Xylella fastidiosa that is crucial to its switching between plant hosts and insect vectors is dependent on RpfF and its production of 2-enoic acids known as diffusible signal factor (DSF. We show that X. fastidiosa produces a particularly large variety of similar, relatively long-chain-length 2-enoic acids that are active in modulating gene expression. Both X. fastidiosa itself and a Pantoea agglomerans surrogate host harboring X. fastidiosa RpfF (XfRpfF is capable of producing a variety of both saturated and unsaturated free fatty acids. However, only 2-cis unsaturated acids were found to be biologically active in X. fastidiosa. X. fastidiosa produces, and is particularly responsive to, a novel DSF species, 2-cis-hexadecanoic acid that we term XfDSF2. It is also responsive to other, even longer 2-enoic acids to which other taxa such as Xanthomonas campestris are unresponsive. The 2-enoic acids that are produced by X. fastidiosa are strongly affected by the cellular growth environment, with XfDSF2 not detected in culture media in which 2-tetradecenoic acid (XfDSF1 had previously been found. X. fastidiosa is responsive to much lower concentrations of XfDSF2 than XfDSF1. Apparently competitive interactions can occur between various saturated and unsaturated fatty acids that block the function of those agonistic 2-enoic fatty acids. By altering the particular 2-enoic acids produced and the relative balance of free enoic and saturated fatty acids, X. fastidiosa might modulate the extent of DSF-mediated quorum sensing.

  5. Direct Evidence of Egestion and Salivation of Xylella fastidiosa Suggests Sharpshooters Can Be "Flying Syringes".

    Science.gov (United States)

    Backus, Elaine A; Shugart, Holly J; Rogers, Elizabeth E; Morgan, J Kent; Shatters, Robert

    2015-05-01

    Xylella fastidiosa is unique among insect-transmitted plant pathogens because it is propagative but noncirculative, adhering to and multiplying on the cuticular lining of the anterior foregut. Any inoculation mechanism for X. fastidiosa must explain how bacterial cells exit the vector's stylets via the food canal and directly enter the plant. A combined egestion-salivation mechanism has been proposed to explain these unique features. Egestion is the putative outward flow of fluid from the foregut via hypothesized bidirectional pumping of the cibarium. The present study traced green fluorescent protein-expressing X. fastidiosa or fluorescent nanoparticles acquired from artificial diets by glassy-winged sharpshooters, Homalodisca vitripennis, as they were egested into simultaneously secreted saliva. X. fastidiosa or nanoparticles were shown to mix with gelling saliva to form fluorescent deposits and salivary sheaths on artificial diets, providing the first direct, conclusive evidence of egestion by any hemipteran insect. Therefore, the present results strongly support an egestion-salivation mechanism of X. fastidiosa inoculation. Results also support that a column of fluid is transiently held in the foregut without being swallowed. Evidence also supports (but does not definitively prove) that bacteria were suspended in the column of fluid during the vector's transit from diet to diet, and were egested with the held fluid. Thus, we hypothesize that sharpshooters could be true "flying syringes," especially when inoculation occurs very soon after uptake of bacteria, suggesting the new paradigm of a nonpersistent X. fastidiosa transmission mechanism.

  6. The Secreted Protease PrtA Controls Cell Growth, Biofilm Formation and Pathogenicity in Xylella fastidiosa.

    Science.gov (United States)

    Gouran, Hossein; Gillespie, Hyrum; Nascimento, Rafael; Chakraborty, Sandeep; Zaini, Paulo A; Jacobson, Aaron; Phinney, Brett S; Dolan, David; Durbin-Johnson, Blythe P; Antonova, Elena S; Lindow, Steven E; Mellema, Matthew S; Goulart, Luiz R; Dandekar, Abhaya M

    2016-08-05

    Pierce's disease (PD) is a deadly disease of grapevines caused by the Gram-negative bacterium Xylella fastidiosa. Though disease symptoms were formerly attributed to bacteria blocking the plant xylem, this hypothesis is at best overly simplistic. Recently, we used a proteomic approach to characterize the secretome of X. fastidiosa, both in vitro and in planta, and identified LesA as one of the pathogenicity factors of X. fastidiosa in grapevines that leads to leaf scorching and chlorosis. Herein, we characterize another such factor encoded by PD0956, designated as an antivirulence secreted protease "PrtA" that displays a central role in controlling in vitro cell proliferation, length, motility, biofilm formation, and in planta virulence. The mutant in X. fastidiosa exhibited reduced cell length, hypermotility (and subsequent lack of biofilm formation) and hypervirulence in grapevines. These findings are supported by transcriptomic and proteomic analyses with corresponding plant infection data. Of particular interest, is the hypervirulent response in grapevines observed when X. fastidiosa is disrupted for production of PrtA, and that PD-model tobacco plants transformed to express PrtA exhibited decreased symptoms after infection by X. fastidiosa.

  7. Initial crystallographic studies of a small heat-shock protein from Xylella fastidiosa

    International Nuclear Information System (INIS)

    Tada, Susely F. S.; Saraiva, Antonio Marcos; Lorite, Gabriela S.; Rosselli-Murai, Luciana K.; Pelloso, Alexandre César; Santos, Marcelo Leite dos; Trivella, Daniela B. B.; Cotta, Mônica A.; Souza, Anete Pereira de; Aparicio, Ricardo

    2012-01-01

    Initial crystallographic studies of the X. fastidiosa small heat-shock protein HSP17.9 are reported. The ORF XF2234 in the Xylella fastidiosa genome was identified as encoding a small heat-shock protein of 17.9 kDa (HSP17.9). HSP17.9 was found as one of the proteins that are induced during X. fastidiosa proliferation and infection in citrus culture. Recombinant HSP17.9 was crystallized and surface atomic force microscopy experiments were conducted with the aim of better characterizing the HSP17.9 crystals. X-ray diffraction data were collected at 2.7 Å resolution. The crystal belonged to space group P4 3 22, with unit-cell parameters a = 68.90, b = 68.90, c = 72.51 Å, and is the first small heat-shock protein to crystallize in this space group

  8. New Coffee Plant-Infecting Xylella fastidiosa Variants Derived via Homologous Recombination.

    Science.gov (United States)

    Jacques, Marie-Agnès; Denancé, Nicolas; Legendre, Bruno; Morel, Emmanuelle; Briand, Martial; Mississipi, Stelly; Durand, Karine; Olivier, Valérie; Portier, Perrine; Poliakoff, Françoise; Crouzillat, Dominique

    2015-12-28

    Xylella fastidiosa is a xylem-limited phytopathogenic bacterium endemic to the Americas that has recently emerged in Asia and Europe. Although this bacterium is classified as a quarantine organism in the European Union, importation of plant material from contaminated areas and latent infection in asymptomatic plants have engendered its inevitable introduction. In 2012, four coffee plants (Coffea arabica and Coffea canephora) with leaf scorch symptoms growing in a confined greenhouse were detected and intercepted in France. After identification of the causal agent, this outbreak was eradicated. Three X. fastidiosa strains were isolated from these plants, confirming a preliminary identification based on immunology. The strains were characterized by multiplex PCR and by multilocus sequence analysis/typing (MLSA-MLST) based on seven housekeeping genes. One strain, CFBP 8073, isolated from C. canephora imported from Mexico, was assigned to X. fastidiosa subsp. fastidiosa/X. fastidiosa subsp. sandyi. This strain harbors a novel sequence type (ST) with novel alleles at two loci. The two other strains, CFBP 8072 and CFBP 8074, isolated from Coffea arabica imported from Ecuador, were allocated to X. fastidiosa subsp. pauca. These two strains shared a novel ST with novel alleles at two loci. These MLST profiles showed evidence of recombination events. We provide genome sequences for CFBP 8072 and CFBP 8073 strains. Comparative genomic analyses of these two genome sequences with publicly available X. fastidiosa genomes, including the Italian strain CoDiRO, confirmed these phylogenetic positions and provided candidate alleles for coffee plant adaptation. This study demonstrates the global diversity of X. fastidiosa and highlights the diversity of strains isolated from coffee plants. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  9. Identification of three potential insect vectors of Xylella fastidiosa in southern Italy

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    Toufic ELBEAINO

    2014-09-01

    Full Text Available In order to identify potential vectors of Xylella fastidiosa in olive orchards in Puglia (southern Italy, Hemiptera insects were collected from October to December, 2013, in olive orchards with high incidences of X. fastidiosa associated with “rapid decline” symptoms. The study focused on species in the Auchenorrhyncha (sharpshooter leafhoppers and froghoppers or spittlebugs, a group that includes known vectors of X. fastidiosa.  Adults of three species, i.e. Philaenus spumarius L. (Aphrophoridae, Neophilaenus campestris Fallén (Aphrophoridae and Euscelis lineolatus Brullé (Cicadellidae were captured, from which total DNA was extracted and assayed by PCR using three sets of specific primers designed for X. fastidiosa detection. Results of PCR showed that 38 out of a total of 84 tested insects were positive for X. fastidiosa, i.e. eight (of 20 P. spumarius, 14 (of 18 N. campestris and 16 (of 46 E. lineolatus. PCR amplicons of the RNA polymerase sigma-70 factor gene from six specimens (two of each insect species were sequenced. The sequences obtained were 99.3‒99.4% identical. BlastN analyses demonstrated these sequences to be similar to those of X. fastidiosa isolates from olive OL-X and OL-G reported from Puglia, whereas they displayed distant molecular identity (always less than 98% with X. fastidiosa subspecies from other countries. The detection of X. fastidiosa in P. spumarius and, for the first time, in N. campestris and E. lineolatus (which, unlike the others, is a phloem feeder, indicates potential vectoring roles of these insects for the spread of the bacterium in Puglia. Further investigations and specific infectivity trials are required to definitively determine the roles of these insects as effective vectors of this pathogen.

  10. Comparative genomic analysis of coffee-infecting Xylella fastidiosa strains isolated from Brazil.

    Science.gov (United States)

    Barbosa, Deibs; Alencar, Valquíria Campos; Santos, Daiene Souza; de Freitas Oliveira, Ana Cláudia; de Souza, Alessandra A; Coletta-Filho, Helvecio D; de Oliveira, Regina Souza; Nunes, Luiz R

    2015-05-01

    Strains of Xylella fastidiosa constitute a complex group of bacteria that develop within the xylem of many plant hosts, causing diseases of significant economic importance, such as Pierce's disease in North American grapevines and citrus variegated chlorosis in Brazil. X. fastidiosa has also been obtained from other host plants, in direct correlation with the development of diseases, as in the case of coffee leaf scorch (CLS)--a disease with potential to cause severe economic losses to the Brazilian coffee industry. This paper describes a thorough genomic characterization of coffee-infecting X. fastidiosa strains, initially performed through a microarray-based approach, which demonstrated that CLS strains could be subdivided in two phylogenetically distinct subgroups. Whole-genomic sequencing of two of these bacteria (one from each subgroup) allowed identification of ORFs and horizontally transferred elements (HTEs) that were specific to CLS-related X. fastidiosa strains. Such analyses confirmed the size and importance of HTEs as major mediators of chromosomal evolution amongst these bacteria, and allowed identification of differences in gene content, after comparisons were made with previously sequenced X. fastidiosa strains, isolated from alternative hosts. Although direct experimentation still needs to be performed to elucidate the biological consequences associated with such differences, it was interesting to verify that CLS-related bacteria display variations in genes that produce toxins, as well as surface-related factors (such as fimbrial adhesins and LPS) that have been shown to be involved with recognition of specific host factors in different pathogenic bacteria. © 2015 The Authors.

  11. The diversity of citrus endophytic bacteria and their interactions with Xylella fastidiosa and host plants

    Science.gov (United States)

    Azevedo, João Lúcio; Araújo, Welington Luiz; Lacava, Paulo Teixeira

    2016-01-01

    Abstract The bacterium Xylella fastidiosa is the causal agent of citrus variegated chlorosis (CVC) and has been associated with important losses in commercial orchards of all sweet orange [Citrus sinensis (L.)] cultivars. The development of this disease depends on the environmental conditions, including the endophytic microbial community associated with the host plant. Previous studies have shown that X. fastidiosa interacts with the endophytic community in xylem vessels as well as in the insect vector, resulting in a lower bacterial population and reduced CVC symptoms. The citrus endophytic bacterium Methylobacterium mesophilicum can trigger X. fastidiosa response in vitro, which results in reduced growth and induction of genes associated with energy production, stress, transport, and motility, indicating that X. fastidiosa has an adaptive response to M. mesophilicum. Although this response may result in reduced CVC symptoms, the colonization rate of the endophytic bacteria should be considered in studies that intend to use this endophyte to suppress CVC disease. Symbiotic control is a new strategy that uses symbiotic endophytes as biological control agents to antagonize or displace pathogens. Candidate endophytes for symbiotic control of CVC must occupy the xylem of host plants and attach to the precibarium of sharpshooter insects to access the pathogen. In the present review, we focus on interactions between endophytic bacteria from sweet orange plants and X. fastidiosa, especially those that may be candidates for control of CVC. PMID:27727362

  12. The diversity of citrus endophytic bacteria and their interactions with Xylella fastidiosa and host plants

    Directory of Open Access Journals (Sweden)

    João Lúcio Azevedo

    Full Text Available Abstract The bacterium Xylella fastidiosa is the causal agent of citrus variegated chlorosis (CVC and has been associated with important losses in commercial orchards of all sweet orange [Citrus sinensis (L.] cultivars. The development of this disease depends on the environmental conditions, including the endophytic microbial community associated with the host plant. Previous studies have shown that X. fastidiosa interacts with the endophytic community in xylem vessels as well as in the insect vector, resulting in a lower bacterial population and reduced CVC symptoms. The citrus endophytic bacterium Methylobacterium mesophilicum can trigger X. fastidiosa response in vitro, which results in reduced growth and induction of genes associated with energy production, stress, transport, and motility, indicating that X. fastidiosa has an adaptive response to M. mesophilicum. Although this response may result in reduced CVC symptoms, the colonization rate of the endophytic bacteria should be considered in studies that intend to use this endophyte to suppress CVC disease. Symbiotic control is a new strategy that uses symbiotic endophytes as biological control agents to antagonize or displace pathogens. Candidate endophytes for symbiotic control of CVC must occupy the xylem of host plants and attach to the precibarium of sharpshooter insects to access the pathogen. In the present review, we focus on interactions between endophytic bacteria from sweet orange plants and X. fastidiosa, especially those that may be candidates for control of CVC.

  13. Identification of an operon, Pil-Chp, that controls twitching motility and virulence in Xylella fastidiosa.

    Science.gov (United States)

    Cursino, Luciana; Galvani, Cheryl D; Athinuwat, Dusit; Zaini, Paulo A; Li, Yaxin; De La Fuente, Leonardo; Hoch, Harvey C; Burr, Thomas J; Mowery, Patricia

    2011-10-01

    Xylella fastidiosa is an important phytopathogenic bacterium that causes many serious plant diseases, including Pierce's disease of grapevines. Disease manifestation by X. fastidiosa is associated with the expression of several factors, including the type IV pili that are required for twitching motility. We provide evidence that an operon, named Pil-Chp, with genes homologous to those found in chemotaxis systems, regulates twitching motility. Transposon insertion into the pilL gene of the operon resulted in loss of twitching motility (pilL is homologous to cheA genes encoding kinases). The X. fastidiosa mutant maintained the type IV pili, indicating that the disrupted pilL or downstream operon genes are involved in pili function, and not biogenesis. The mutated X. fastidiosa produced less biofilm than wild-type cells, indicating that the operon contributes to biofilm formation. Finally, in planta the mutant produced delayed and less severe disease, indicating that the Pil-Chp operon contributes to the virulence of X. fastidiosa, presumably through its role in twitching motility.

  14. Radicinin from Cochliobolus sp. inhibits Xylella fastidiosa, the causal agent of Pierce's Disease of grapevine.

    Science.gov (United States)

    Aldrich, Thomas J; Rolshausen, Philippe E; Roper, M Caroline; Reader, Jordan M; Steinhaus, Matthew J; Rapicavoli, Jeannette; Vosburg, David A; Maloney, Katherine N

    2015-08-01

    The fastidious phytopathogenic bacterium, Xylella fastidiosa, poses a substantial threat to many economically important crops, causing devastating diseases including Pierce's Disease of grapevine. Grapevines (Vitis vinifera L.) planted in an area under Pierce's Disease pressure often display differences in disease severity and symptom expression, with apparently healthy vines growing alongside the dying ones, despite the fact that all the vines are genetic clones of one another. Under the hypothesis that endophytic microbes might be responsible for this non-genetic resistance to X. fastidiosa, endophytic fungi were isolated from vineyard cvs. 'Chardonnay' and 'Cabernet Sauvignon' grown under high Pierce's Disease pressure. A Cochliobolus sp. isolated from a Cabernet Sauvignon grapevine inhibited the growth of X. fastidiosa in vitro. Bioassay-guided isolation of an organic extract of Cochliobolus sp. yielded the natural product radicinin as the major active compound. Radicinin also inhibited proteases isolated from the culture supernatant of X. fastidiosa. In order to assess structure-activity relationships, three semi-synthetic derivatives of radicinin were prepared and tested for activity against X. fastidiosa in vitro. Assay results of these derivatives are consistent with enzyme inactivation by conjugate addition to carbon-10 of radicinin, as proposed previously. Copyright © 2015 Elsevier Ltd. All rights reserved.

  15. Multiplexed lateral flow microarray assay for detection of citrus pathogens Xylella fastidiosa and Xanthomonas axonopodis pv citri

    Energy Technology Data Exchange (ETDEWEB)

    Cary,; Bruce, R [Santa Fe, NM; Stubben, Christopher J [Los Alamos, NM

    2011-03-22

    The invention provides highly sensitive and specific assays for the major citrus pathogens Xylella fastidiosa and Xanthomonas axonopodis, including a field deployable multiplexed assay capable of rapidly assaying for both pathogens simultaneously. The assays are directed at particular gene targets derived from pathogenic strains that specifically cause the major citrus diseases of citrus variegated chlorosis (Xylella fastidiosa 9a5c) and citrus canker (Xanthomonas axonopodis pv citri). The citrus pathogen assays of the invention offer femtomole sensitivity, excellent linear dynamic range, and rapid and specific detection.

  16. Visualization of Twitching Motility and Characterization of the Role of the PilG in Xylella fastidiosa.

    Science.gov (United States)

    Shi, Xiangyang; Lin, Hong

    2016-04-08

    Xylella fastidiosa is a Gram-negative non-flagellated bacterium that causes a number of economically important diseases of plants. The twitching motility provides X. fastidiosa a means for long-distance intra-plant movement and colonization, contributing toward pathogenicity in X. fastidiosa. The twitching motility of X. fastidiosa is operated by type IV pili. Type IV pili of Xylella fastidiosa are regulated by pilG, a chemotaxis regulator in Pil-Chp operon encoding proteins that are involved with signal transduction pathways. To elucidate the roles of pilG in the twitching motility of X. fastidiosa, a pilG-deficient mutant XfΔpilG and its complementary strain XfΔpilG-C containing native pilG were developed. A microfluidic chambers integrated with a time-lapse image recording system was used to observe twitching motility in XfΔpilG, XfΔpilG-C and its wild type strain. Using this recording system, it permits long-term spatial and temporal observations of aggregation, migration of individual cells and populations of bacteria via twitching motility. X. fastidiosa wild type and complementary XfΔpilG-C strain showed typical twitching motility characteristics directly observed in the microfluidic flow chambers, whereas mutant XfΔpliG exhibited the twitching deficient phenotype. This study demonstrates that pilG contributes to the twitching motility of X. fastidiosa. The microfluidic flow chamber is used as a means for observing twitching motility.

  17. Differentiation of Xylella fastidiosa strains via multilocus sequence analysis of environmentally mediated genes (MLSA-E).

    Science.gov (United States)

    Parker, Jennifer K; Havird, Justin C; De La Fuente, Leonardo

    2012-03-01

    Isolates of the plant pathogen Xylella fastidiosa are genetically very similar, but studies on their biological traits have indicated differences in virulence and infection symptomatology. Taxonomic analyses have identified several subspecies, and phylogenetic analyses of housekeeping genes have shown broad host-based genetic differences; however, results are still inconclusive for genetic differentiation of isolates within subspecies. This study employs multilocus sequence analysis of environmentally mediated genes (MLSA-E; genes influenced by environmental factors) to investigate X. fastidiosa relationships and differentiate isolates with low genetic variability. Potential environmentally mediated genes, including host colonization and survival genes related to infection establishment, were identified a priori. The ratio of the rate of nonsynonymous substitutions to the rate of synonymous substitutions (dN/dS) was calculated to select genes that may be under increased positive selection compared to previously studied housekeeping genes. Nine genes were sequenced from 54 X. fastidiosa isolates infecting different host plants across the United States. Results of maximum likelihood (ML) and Bayesian phylogenetic (BP) analyses are in agreement with known X. fastidiosa subspecies clades but show novel within-subspecies differentiation, including geographic differentiation, and provide additional information regarding host-based isolate variation and specificity. dN/dS ratios of environmentally mediated genes, though gene dN/dS ratios and correlate with increased sequence variability. MLSA-E can more precisely resolve relationships between closely related bacterial strains with low genetic variability, such as X. fastidiosa isolates. Discovering the genetic relationships between X. fastidiosa isolates will provide new insights into the epidemiology of populations of X. fastidiosa, allowing improved disease management in economically important crops.

  18. Promiscuous Diffusible Signal Factor Production and Responsiveness of the Xylella fastidiosa Rpf System.

    Science.gov (United States)

    Ionescu, Michael; Yokota, Kenji; Antonova, Elena; Garcia, Angelica; Beaulieu, Ellen; Hayes, Terry; Iavarone, Anthony T; Lindow, Steven E

    2016-07-19

    Cell density-dependent regulation of gene expression in Xylella fastidiosa that is crucial to its switching between plant hosts and insect vectors is dependent on RpfF and its production of 2-enoic acids known as diffusible signal factor (DSF). We show that X. fastidiosa produces a particularly large variety of similar, relatively long-chain-length 2-enoic acids that are active in modulating gene expression. Both X. fastidiosa itself and a Pantoea agglomerans surrogate host harboring X. fastidiosa RpfF (XfRpfF) is capable of producing a variety of both saturated and unsaturated free fatty acids. However, only 2-cis unsaturated acids were found to be biologically active in X. fastidiosa X. fastidiosa produces, and is particularly responsive to, a novel DSF species, 2-cis-hexadecanoic acid that we term XfDSF2. It is also responsive to other, even longer 2-enoic acids to which other taxa such as Xanthomonas campestris are unresponsive. The 2-enoic acids that are produced by X. fastidiosa are strongly affected by the cellular growth environment, with XfDSF2 not detected in culture media in which 2-tetradecenoic acid (XfDSF1) had previously been found. X. fastidiosa is responsive to much lower concentrations of XfDSF2 than XfDSF1. Apparently competitive interactions can occur between various saturated and unsaturated fatty acids that block the function of those agonistic 2-enoic fatty acids. By altering the particular 2-enoic acids produced and the relative balance of free enoic and saturated fatty acids, X. fastidiosa might modulate the extent of DSF-mediated quorum sensing. X. fastidiosa, having a complicated lifestyle in which it moves and multiplies within plants but also must be vectored by insects, utilizes DSF-based quorum sensing to partition the expression of traits needed for these two processes within different cells in this population based on local cellular density. The finding that it can produce a variety of DSF species in a strongly

  19. Xylella fastidiosa comparative genomic database is an information resource to explore the annotation, genomic features, and biology of different strains

    Directory of Open Access Journals (Sweden)

    Alessandro M. Varani

    2012-01-01

    Full Text Available The Xylella fastidiosa comparative genomic database is a scientific resource with the aim to provide a user-friendly interface for accessing high-quality manually curated genomic annotation and comparative sequence analysis, as well as for identifying and mapping prophage-like elements, a marked feature of Xylella genomes. Here we describe a database and tools for exploring the biology of this important plant pathogen. The hallmarks of this database are the high quality genomic annotation, the functional and comparative genomic analysis and the identification and mapping of prophage-like elements. It is available from web site http://www.xylella.lncc.br.

  20. ASPECTOS ESTRUTURAIS DE CAFEEIRO INFECTADO COM XYLELLA FASTIDIOSA

    Directory of Open Access Journals (Sweden)

    RACHEL BENETTI QUEIROZ-VOLTAN

    1998-01-01

    Full Text Available Estudaram-se as alterações anatômicas em folhas e caules de ramos de cafeeiros infectados com X. fastidiosa visando compreender as causas dos seus distúrbios fisiológicos. Analisaram-se amostras dos cultivares comerciais de Coffea arabica L. - Catuaí Vermelho IAC H 2077-25-81 e Mundo Novo 515-20 enxertados sobre a progênie IAC 2258 de C. canephora - e de Catuaí Vermelho IAC H 2077-25-81 sem enxertia (pé franco. As amostras foram coletadas no campo, no Núcleo Experimental de Campinas, IAC, em cafezais onde foi detectada a presença da bactéria. Analisou-se também a estrutura da folha e do caule de ramos com e sem sintomas visuais da bactéria, sendo estimada a porcentagem de obstrução dos vasos do xilema do caule. Os ramos infectados apresentaram um encurtamento dos entrenós e, com o agravamento da doença, senescência foliar precoce na sua base, resultando em pequeno número de folhas no ápice. O número e a posição dos ramos por indivíduo com tais sintomas variaram entre as plantas, sendo mais freqüentes na região basal. Esses ramos também mostraram pecíolos e área foliar reduzidos e frutos menores e agrupados, em vista de um encurtamento dos pedicelos e dos entrenós, além de uma deposição de "goma" nos vasos do xilema do caule, pecíolo e folha, assim como divisões celulares anormais no xilema, floema e córtex daquelas partes. As células do mesofilo das folhas afetadas apresentaram número reduzido de cloroplastos, associado a maior concentração de cristais de oxalato de cálcio. Sugere-se que a presença da bactéria esteja induzindo a senescência foliar que se relaciona a um ou mais fatores de estresse.Leaf and stem anatomical changes were studied in coffee plants infected with X. fastidiosa looking for the causes of the observed physiological disturbances. Samples of commercial cultivars of Coffea arabica L. were analysed - Catuaí Vermelho IAC H 2077-25-81 and Mundo Novo 515-20 grafted on the progenie IAC

  1. Twitching motility and biofilm formation are associated with tonB1 in Xylella fastidiosa

    OpenAIRE

    Cursino, Luciana; Li, Yaxin; Zaini, Paulo A.; De La Fuente, Leonardo; Hoch, Harvey C.; Burr, Thomas J.

    2017-01-01

    A mutation in the Xylella fastidiosa tonB1 gene resulted in loss of twitching motility and in significantly less biofilm formation as compared with a wild type. The altered motility and biofilm phenotypes were restored by complementation with a functional copy of the gene. The mutation affected virulence as measured by Pierce's disease symptoms on grapevines. The role of TonB1 in twitching and biofilm formation appears to be independent of the characteristic iron-uptake function of this prote...

  2. Hidden Markov models applied to a subsequence of the Xylella fastidiosa genome

    Directory of Open Access Journals (Sweden)

    Silva Cibele Q. da

    2003-01-01

    Full Text Available Dependencies in DNA sequences are frequently modeled using Markov models. However, Markov chains cannot account for heterogeneity that may be present in different regions of the same DNA sequence. Hidden Markov models are more realistic than Markov models since they allow for the identification of heterogeneous regions of a DNA sequence. In this study we present an application of hidden Markov models to a subsequence of the Xylella fastidiosa DNA data. We found that a three-state model provides a good description for the data considered.

  3. Xylella fastidiosa outer membrane vesicles modulate plant colonization by blocking attachment to surfaces.

    Science.gov (United States)

    Ionescu, Michael; Zaini, Paulo A; Baccari, Clelia; Tran, Sophia; da Silva, Aline M; Lindow, Steven E

    2014-09-16

    Outer membrane vesicles (OMVs) of Gram-negative bacteria have been studied intensively in recent years, primarily in their role in delivering virulence factors and antigens during pathogenesis. However, the near ubiquity of their production suggests that they may play other roles, such as responding to envelope stress or trafficking various cargoes to prevent dilution or degradation by other bacterial species. Here we show that OMVs produced by Xylella fastidiosa, a xylem-colonizing plant pathogenic bacterium, block its interaction with various surfaces such as the walls of xylem vessels in host plants. The release of OMVs was suppressed by the diffusible signal factor-dependent quorum-sensing system, and a X. fastidiosa ΔrpfF mutant in which quorum signaling was disrupted was both much more virulent to plants and less adhesive to glass and plant surfaces than the WT strain. The higher virulence of the ΔrpfF mutant was associated with fivefold higher numbers of OMVs recovered from xylem sap of infected plants. The frequency of attachment of X. fastidiosa to xylem vessels was 20-fold lower in the presence of OMVs than in their absence. OMV production thus is a strategy used by X. fastidiosa cells to adjust attachment to surfaces in its transition from adhesive cells capable of insect transmission to an "exploratory" lifestyle for systemic spread within the plant host which would be hindered by attachment. OMV production may contribute to the movement of other bacteria in porous environments by similarly reducing their contact with environmental constituents.

  4. Transcriptome Analysis of the Phytobacterium Xylella fastidiosa Growing under Xylem-Based Chemical Conditions

    Directory of Open Access Journals (Sweden)

    Maristela Boaceff Ciraulo

    2010-01-01

    Full Text Available Xylella fastidiosa is a xylem-limited bacterium responsible for important plant diseases, like citrus-variegated chlorosis (CVC and grapevine Pierce's disease (PD. Interestingly, in vitro growth of X. fastidiosa in chemically defined media that resemble xylem fluid has been achieved, allowing studies of metabolic processes used by xylem-dwelling bacteria to thrive in such nutrient-poor conditions. Thus, we performed microarray hybridizations to compare transcriptomes of X. fastidiosa cells grown in 3G10-R, a medium that resembles grape sap, and in Periwinkle Wilt (PW, the complex medium traditionally used to cultivate X. fastidiosa. We identified 299 transcripts modulated in response to growth in these media. Some 3G10R-overexpressed genes have been shown to be upregulated in cells directly isolated from infected plants and may be involved in plant colonization, virulence and environmental competition. In contrast, cells cultivated in PW show a metabolic switch associated with increased aerobic respiration and enhanced bacterial growth rates.

  5. Comparative analysis of differentially expressed sequence tags of sweet orange and mandarin infected with Xylella fastidiosa

    Directory of Open Access Journals (Sweden)

    Alessandra A. de Souza

    2007-01-01

    Full Text Available The Citrus ESTs Sequencing Project (CitEST conducted at Centro APTA Citros Sylvio Moreira/IAC has identified and catalogued ESTs representing a set of citrus genes expressed under relevant stress responses, including diseases such as citrus variegated chlorosis (CVC, caused by Xylella fastidiosa. All sweet orange (Citrus sinensis L. Osb. varieties are susceptible to X. fastidiosa. On the other hand, mandarins (C. reticulata Blanco are considered tolerant or resistant to the disease, although the bacterium can be sporadically detected within the trees, but no disease symptoms or economic losses are observed. To study their genetic responses to the presence of X. fastidiosa, we have compared EST libraries of leaf tissue of sweet orange Pêra IAC (highly susceptible cultivar to X. fastidiosa and mandarin ‘Ponkan’ (tolerant artificially infected with the bacterium. Using an in silico differential display, 172 genes were found to be significantly differentially expressed in such conditions. Sweet orange presented an increase in expression of photosynthesis related genes that could reveal a strategy to counterbalance a possible lower photosynthetic activity resulting from early effects of the bacterial colonization in affected plants. On the other hand, mandarin showed an active multi-component defense response against the bacterium similar to the non-host resistance pattern.

  6. Origins of the Xylella fastidiosa prophage-like regions and their impact in genome differentiation.

    Directory of Open Access Journals (Sweden)

    Alessandro de Mello Varani

    Full Text Available Xylella fastidiosa is a Gram negative plant pathogen causing many economically important diseases, and analyses of completely sequenced X. fastidiosa genome strains allowed the identification of many prophage-like elements and possibly phage remnants, accounting for up to 15% of the genome composition. To better evaluate the recent evolution of the X. fastidiosa chromosome backbone among distinct pathovars, the number and location of prophage-like regions on two finished genomes (9a5c and Temecula1, and in two candidate molecules (Ann1 and Dixon were assessed. Based on comparative best bidirectional hit analyses, the majority (51% of the predicted genes in the X. fastidiosa prophage-like regions are related to structural phage genes belonging to the Siphoviridae family. Electron micrograph reveals the existence of putative viral particles with similar morphology to lambda phages in the bacterial cell in planta. Moreover, analysis of microarray data indicates that 9a5c strain cultivated under stress conditions presents enhanced expression of phage anti-repressor genes, suggesting switches from lysogenic to lytic cycle of phages under stress-induced situations. Furthermore, virulence-associated proteins and toxins are found within these prophage-like elements, thus suggesting an important role in host adaptation. Finally, clustering analyses of phage integrase genes based on multiple alignment patterns reveal they group in five lineages, all possessing a tyrosine recombinase catalytic domain, and phylogenetically close to other integrases found in phages that are genetic mosaics and able to perform generalized and specialized transduction. Integration sites and tRNA association is also evidenced. In summary, we present comparative and experimental evidence supporting the association and contribution of phage activity on the differentiation of Xylella genomes.

  7. Detection and typing of Xylella fastidiosa from glassy-winged sharpshooter for Pierce’s disease epidemiology

    Science.gov (United States)

    Epidemiology of Pierce’s disease of grape, caused by the bacterial pathogen Xylella fastidiosa (Xf), is largely dependent on populations of insect vectors such as the invasive glassy-winged sharpshooter (GWSS) (Homalodisca vitripennis). In the grape-growing regions of the southern San Joaquin Valley...

  8. Plasmid vectors for Xylella fastidiosa utilizing a toxin-antitoxin system for plasmid stability in the absence of antibiotic selection

    Science.gov (United States)

    The phytopathogen Xylella fastidiosa causes disease in a variety of important crop and landscape plants. Functional genetic studies have led to a broader understanding of virulence mechanisms used by this pathogen in the grapevine host. Plasmid shuttle vectors are important tools in studies of bacte...

  9. Xylella fastidiosa in rabbiteye blueberry in Louisiana is genetically similar to a strain found in Southern highbush blueberry in Georgia

    Science.gov (United States)

    During the past ten years, Xylella fastidiosa has been confirmed as a pathogen of Southern highbush blueberry (Vaccinium corymbosum interspecific hybrids) in Georgia and Florida. Recent work in Louisiana has shown that it is also associated with reduced yield and altered fruit quality in ‘Tifblue’ ...

  10. The distribution and biology of potential vectors of Xylella fastidiosa on coffee and citrus in Puerto Rico

    Science.gov (United States)

    Plant diseases caused by Xylella fastidiosa (Wells et al.) (Xf) surround the Caribbean Basin. Two major commodities of Puerto Rico, coffee and citrus, are highly susceptible to Xf. We surveyed potential vectors of Xf in coffee and citrus farms in western Puerto Rico over an 18 month period. Cicadel...

  11. Plasmid transfer by conjugation as a possible route of horizontal gene transfer and recombination in Xylella fastidiosa

    Science.gov (United States)

    Horizontal gene transfer is an important component of evolution and adaptation of bacterial species. Xylella fastidiosa has the ability to incorporate exogenous DNA into its genome by homologous recombination at relatively high rates. This genetic recombination is believed to play a role in adaptati...

  12. De Novo whole genome sequence of Xylella fastidiosa subsp. multiplex strain BB01 from blueberry in Georgia, USA

    Science.gov (United States)

    This study reports a de novo assembled draft genome sequence of Xylella fastidiosa subsp. multiplex strain BB01 causing blueberry bacterial leaf scorch in Georgia, USA. The BB01 genome is 2,517,579 bp with a G+C content of 51.8% and 2,943 open reading frames (ORFs) and 48 RNA genes....

  13. Association of Xylella fastidiosa with Yield Loss and Altered Fruit Quality in a Naturally Infected Rabbiteye Blueberry Orchard

    Science.gov (United States)

    Xylella fastidiosa causes disease in a number of plants in the southeastern United States, including southern highbush blueberry, but little was known concerning its potential impact in rabbiteye blueberry (Vaccinium virgatum). In a naturally infected orchard in Louisiana, mean yields of X. fastidi...

  14. Population structure of Xylella fastidiosa associated with almond leaf scorch disease in the San Joaquin Valley of California

    Science.gov (United States)

    Xylella fastidiosa (Xf) causes disease in many commercial crops including almond leaf scorch (ALS) disease in susceptible almond (Prunus dulcis). In this study, genetic diversity and population structure of Xf associated with ALS disease were evaluated. Strains isolated from two almond production si...

  15. Identification of a non-host plant of Xylella fastidiosa to rear healthy sharpshooter vectors Identificação de uma planta não-hospedeira de Xylella fastidiosa para criação de insetos vetores sadios

    Directory of Open Access Journals (Sweden)

    Rosangela Cristina Marucci

    2003-12-01

    Full Text Available Rearing leafhopper (Hemiptera: Cicadellidae vectors free of Xylella fastidiosa is a requirement for studies of various aspects of vector-pathogen interactions. The selection of a plant that allows vector development but not bacterial multiplication is desirable to produce healthy vectors. In this study, two leafhopper hosts, Vernonia condensata ('boldo' and Aloysia virgata ('lixeira' were needle inoculated with citrus and coffee strains of X. fastidiosa to evaluate if these plants support pathogen colonization. The inoculated plants did not present symptoms and the pathogen was not detected by culture and PCR tests, neither soon after inoculation (7-14 days nor later, at 1, 4, 6 and 12 months after inoculation. To obtain healthy adults of the leafhopper vectors Acrogonia citrina, Bucephalogonia xanthophis, Dilobopterus costalimai, Homalodisca ignorata and Oncometopia facialis, early-instar nymphs were reared on V. condensata. X. fastidiosa was not detected in any of 175 adults obtained. V. condensata and A. virgata are nonpropagative hosts of X. fastidiosa and enable the production of healthy leafhoppers for vector studies.A obtenção de cigarrinhas (Hemiptera: Cicadellidae livres de Xylella fastidiosa é importante para estudos de interação entre essa bactéria e seus vetores, sendo desejável a seleção de uma planta que permita a criação desses insetos, mas não a multiplicação da bactéria. Neste estudo, duas plantas hospedeiras de cigarrinhas, Vernonia condensata (boldo e Aloysia virgata (lixeira, foram inoculadas por agulha com as estirpes de citros e de cafeeiro de X. fastidiosa, para avaliar a possibilidade deste patógeno colonizá-las. Não foram observados sintomas, nem se detectou a bactéria por isolamento em meio de cultura e/ou PCR em períodos curtos (7 e 14 dias ou longos (1, 4, 6 e 12 meses após a inoculação. Para obtenção de adultos sadios das cigarrinhas vetoras, Acrogonia citrina, Bucephalogonia xanthophis

  16. Characterization of novel virulent broad-host-range phages of Xylella fastidiosa and Xanthomonas.

    Science.gov (United States)

    Ahern, Stephen J; Das, Mayukh; Bhowmick, Tushar Suvra; Young, Ry; Gonzalez, Carlos F

    2014-01-01

    The xylem-limited bacterium Xylella fastidiosa is the causal agent of several plant diseases, most notably Pierce's disease of grape and citrus variegated chlorosis. We report the isolation and characterization of the first virulent phages for X. fastidiosa, siphophages Sano and Salvo and podophages Prado and Paz, with a host range that includes Xanthomonas spp. Phages propagated on homologous hosts had observed adsorption rate constants of ~4 × 10(-12) ml cell(-1) min(-1) for X. fastidiosa strain Temecula 1 and ~5 × 10(-10) to 7 × 10(-10) ml cell(-1) min(-1) for Xanthomonas strain EC-12. Sano and Salvo exhibit >80% nucleotide identity to each other in aligned regions and are syntenic to phage BcepNazgul. We propose that phage BcepNazgul is the founding member of a novel phage type, to which Sano and Salvo belong. The lysis genes of the Nazgul-like phage type include a gene that encodes an outer membrane lipoprotein endolysin and also spanin gene families that provide insight into the evolution of the lysis pathway for phages of Gram-negative hosts. Prado and Paz, although exhibiting no significant DNA homology to each other, are new members of the phiKMV-like phage type, based on the position of the single-subunit RNA polymerase gene. The four phages are type IV pilus dependent for infection of both X. fastidiosa and Xanthomonas. The phages may be useful as agents for an effective and environmentally responsible strategy for the control of diseases caused by X. fastidiosa.

  17. Three New Pierce's Disease Pathogenicity Effectors Identified Using Xylella fastidiosa Biocontrol Strain EB92-1.

    Science.gov (United States)

    Zhang, Shujian; Chakrabarty, Pranjib K; Fleites, Laura A; Rayside, Patricia A; Hopkins, Donald L; Gabriel, Dean W

    2015-01-01

    Xylella fastidiosa (X. fastidiosa) infects a wide range of plant hosts and causes economically serious diseases, including Pierce's Disease (PD) of grapevines. X. fastidiosa biocontrol strain EB92-1 was isolated from elderberry and is infectious and persistent in grapevines but causes only very slight symptoms under ideal conditions. The draft genome of EB92-1 revealed that it appeared to be missing genes encoding 10 potential PD pathogenicity effectors found in Temecula1. Subsequent PCR and sequencing analyses confirmed that EB92-1 was missing the following predicted effectors found in Temecula1: two type II secreted enzymes, including a lipase (LipA; PD1703) and a serine protease (PD0956); two identical genes encoding proteins similar to Zonula occludens toxins (Zot; PD0915 and PD0928), and at least one relatively short, hemagglutinin-like protein (PD0986). Leaves of tobacco and citrus inoculated with cell-free, crude protein extracts of E. coli BL21(DE3) overexpressing PD1703 exhibited a hypersensitive response (HR) in less than 24 hours. When cloned into shuttle vector pBBR1MCS-5, PD1703 conferred strong secreted lipase activity to Xanthomonas citri, E. coli and X. fastidiosa EB92-1 in plate assays. EB92-1/PD1703 transformants also showed significantly increased disease symptoms on grapevines, characteristic of PD. Genes predicted to encode PD0928 (Zot) and a PD0986 (hemagglutinin) were also cloned into pBBR1MCS-5 and moved into EB92-1; both transformants also showed significantly increased symptoms on V. vinifera vines, characteristic of PD. Together, these results reveal that PD effectors include at least a lipase, two Zot-like toxins and a possibly redundant hemagglutinin, none of which are necessary for parasitic survival of X. fastidiosa populations in grapevines or elderberry.

  18. Detecting genetic introgression: high levels of intersubspecific recombination found in Xylella fastidiosa in Brazil.

    Science.gov (United States)

    Nunney, Leonard; Yuan, Xiaoli; Bromley, Robin E; Stouthamer, Richard

    2012-07-01

    Documenting the role of novel mutation versus homologous recombination in bacterial evolution, and especially in the invasion of new hosts, is central to understanding the long-term dynamics of pathogenic bacteria. We used multilocus sequence typing (MLST) to study this issue in Xylella fastidiosa subsp. pauca from Brazil, a bacterium causing citrus variegated chlorosis (CVC) and coffee leaf scorch (CLS). All 55 citrus isolates typed (plus one coffee isolate) defined three similar sequence types (STs) dominated by ST11 (85%), while the remaining 22 coffee isolates defined two STs, mainly ST16 (74%). This low level of variation masked unusually large allelic differences (>1% divergence with no intermediates) at five loci (leuA, petC, malF, cysG, and holC). We developed an introgression test to detect whether these large differences were due to introgression via homologous recombination from another X. fastidiosa subspecies. Using additional sequencing around these loci, we established that the seven randomly chosen MLST targets contained seven regions of introgression totaling 2,172 bp of 4,161 bp (52%), only 409 bp (10%) of which were detected by other recombination tests. This high level of introgression suggests the hypothesis that X. fastidiosa subsp. pauca became pathogenic on citrus and coffee (crops cultivated in Brazil for several hundred years) only recently after it gained genetic variation via intersubspecific recombination, facilitating a switch from native hosts. A candidate donor is the subspecies infecting plum in the region since 1935 (possibly X. fastidiosa subsp. multiplex). This hypothesis predicts that nonrecombinant native X. fastidiosa subsp. pauca (not yet isolated) does not cause disease in citrus or coffee.

  19. Three New Pierce's Disease Pathogenicity Effectors Identified Using Xylella fastidiosa Biocontrol Strain EB92-1.

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    Shujian Zhang

    Full Text Available Xylella fastidiosa (X. fastidiosa infects a wide range of plant hosts and causes economically serious diseases, including Pierce's Disease (PD of grapevines. X. fastidiosa biocontrol strain EB92-1 was isolated from elderberry and is infectious and persistent in grapevines but causes only very slight symptoms under ideal conditions. The draft genome of EB92-1 revealed that it appeared to be missing genes encoding 10 potential PD pathogenicity effectors found in Temecula1. Subsequent PCR and sequencing analyses confirmed that EB92-1 was missing the following predicted effectors found in Temecula1: two type II secreted enzymes, including a lipase (LipA; PD1703 and a serine protease (PD0956; two identical genes encoding proteins similar to Zonula occludens toxins (Zot; PD0915 and PD0928, and at least one relatively short, hemagglutinin-like protein (PD0986. Leaves of tobacco and citrus inoculated with cell-free, crude protein extracts of E. coli BL21(DE3 overexpressing PD1703 exhibited a hypersensitive response (HR in less than 24 hours. When cloned into shuttle vector pBBR1MCS-5, PD1703 conferred strong secreted lipase activity to Xanthomonas citri, E. coli and X. fastidiosa EB92-1 in plate assays. EB92-1/PD1703 transformants also showed significantly increased disease symptoms on grapevines, characteristic of PD. Genes predicted to encode PD0928 (Zot and a PD0986 (hemagglutinin were also cloned into pBBR1MCS-5 and moved into EB92-1; both transformants also showed significantly increased symptoms on V. vinifera vines, characteristic of PD. Together, these results reveal that PD effectors include at least a lipase, two Zot-like toxins and a possibly redundant hemagglutinin, none of which are necessary for parasitic survival of X. fastidiosa populations in grapevines or elderberry.

  20. Direct tissue blot immunoassay for detection of Xylella fastidiosa in olive trees

    Directory of Open Access Journals (Sweden)

    Khaled DJELOUAH

    2015-01-01

    Full Text Available A direct tissue blot immunoassay (DTBIA technique has been compared with ELISA and PCR for detection of Xylella fastidiosa in olive trees from Apulia (southern Italy. Fresh cross-sections of young twigs and leaf petioles were printed onto nitrocellulose membranes and analyzed in the laboratory. Analyses of a first group of 61 samples gave similar efficiency for the three diagnostic techniques for detection the bacterium (24 positive and 36 negative samples, except for a single sample which was positive only with DTBIA and PCR. Similar results were obtained by separately analyzing suckers and twigs collected from different sectors of tree canopies of a second group of 20 olive trees (ten symptomatic and ten symptomless. In this second test the three diagnostic techniques confirmed the irregular distribution of the bacterium in the tree canopies and erratic detectability of the pathogen in the young suckers. It is therefore necessary to analyse composite samples per tree which should be prepared with twigs collected from different sides of the canopy. The efficiency comparable to ELISA and PCR, combined with the advantages of easier handling, speed and cost, make DTBIA a valid alternative to ELISA in large-scale surveys for occurrence of X. fastidiosa. Moreover, the printing of membranes directly in the field prevents infections spreading to Xylella-free areas, through movement of plant material with pathogen vectors for laboratory testing.

  1. A new paradigm for vector inoculation of Xylella fastidiosa: Direct evidence of egestion and salivation supports that sharpshooters can be “flying syringes”

    Science.gov (United States)

    Despite nearly 70 years of research, the inoculation mechanism of Xylella fastidiosa by its sharpshooter vectors remains unproven. X. fastidiosa is unique among insect-transmitted plant pathogens because it is propagative but non-circulative, adhering to and multiplying on the cuticular lining of th...

  2. A conjugative 38 kB plasmid is present in multiple subspecies of Xylella fastidiosa.

    Science.gov (United States)

    Rogers, Elizabeth E; Stenger, Drake C

    2012-01-01

    A ≈ 38kB plasmid (pXF-RIV5) was present in the Riv5 strain of Xylella fastidiosa subsp. multiplex isolated from ornamental plum in southern California. The complete nucleotide sequence of pXF-RIV5 is almost identical to that of pXFAS01 from X. fastidiosa subsp. fastidiosa strain M23; the two plasmids vary at only 6 nucleotide positions. BLAST searches and phylogenetic analyses indicate pXF-RIV5 and pXFAS01 share some similarity to chromosomal and plasmid (pXF51) sequences of X. fastidiosa subsp. pauca strain 9a5c and more distant similarity to plasmids from a wide variety of bacteria. Both pXF-RIV5 and pXFAS01 encode homologues of a complete Type IV secretion system involved in conjugation and DNA transfer among bacteria. Mating pair formation proteins (Trb) from Yersinia pseudotuberculosis IP31758 are the mostly closely related non-X. fastidiosa proteins to most of the Trb proteins encoded by pXF-RIV5 and pXFAS01. Unlike many bacterial conjugative plasmids, pXF-RIV5 and pXFAS01 do not carry homologues of known accessory modules that confer selective advantage on host bacteria. However, both plasmids encode seven hypothetical proteins of unknown function and possess a small transposon-associated region encoding a putative transposase and associated factor. Vegetative replication of pXF-RIV5 and pXFAS01 appears to be under control of RepA protein and both plasmids have an origin of DNA replication (oriV) similar to that of pRP4 and pR751 from Escherichia coli. In contrast, conjugative plasmids commonly encode TrfA and have an oriV similar to those found in IncP-1 incompatibility group plasmids. The presence of nearly identical plasmids in single strains from two distinct subspecies of X. fastidiosa is indicative of recent horizontal transfer, probably subsequent to the introduction of subspecies fastidiosa to the United States in the late 19(th) century.

  3. A conjugative 38 kB plasmid is present in multiple subspecies of Xylella fastidiosa.

    Directory of Open Access Journals (Sweden)

    Elizabeth E Rogers

    Full Text Available A ≈ 38kB plasmid (pXF-RIV5 was present in the Riv5 strain of Xylella fastidiosa subsp. multiplex isolated from ornamental plum in southern California. The complete nucleotide sequence of pXF-RIV5 is almost identical to that of pXFAS01 from X. fastidiosa subsp. fastidiosa strain M23; the two plasmids vary at only 6 nucleotide positions. BLAST searches and phylogenetic analyses indicate pXF-RIV5 and pXFAS01 share some similarity to chromosomal and plasmid (pXF51 sequences of X. fastidiosa subsp. pauca strain 9a5c and more distant similarity to plasmids from a wide variety of bacteria. Both pXF-RIV5 and pXFAS01 encode homologues of a complete Type IV secretion system involved in conjugation and DNA transfer among bacteria. Mating pair formation proteins (Trb from Yersinia pseudotuberculosis IP31758 are the mostly closely related non-X. fastidiosa proteins to most of the Trb proteins encoded by pXF-RIV5 and pXFAS01. Unlike many bacterial conjugative plasmids, pXF-RIV5 and pXFAS01 do not carry homologues of known accessory modules that confer selective advantage on host bacteria. However, both plasmids encode seven hypothetical proteins of unknown function and possess a small transposon-associated region encoding a putative transposase and associated factor. Vegetative replication of pXF-RIV5 and pXFAS01 appears to be under control of RepA protein and both plasmids have an origin of DNA replication (oriV similar to that of pRP4 and pR751 from Escherichia coli. In contrast, conjugative plasmids commonly encode TrfA and have an oriV similar to those found in IncP-1 incompatibility group plasmids. The presence of nearly identical plasmids in single strains from two distinct subspecies of X. fastidiosa is indicative of recent horizontal transfer, probably subsequent to the introduction of subspecies fastidiosa to the United States in the late 19(th century.

  4. VapD in Xylella fastidiosa Is a Thermostable Protein with Ribonuclease Activity.

    Science.gov (United States)

    Mendes, Juliano S; Santiago, André da S; Toledo, Marcelo A S; Rosselli-Murai, Luciana K; Favaro, Marianna T P; Santos, Clelton A; Horta, Maria Augusta C; Crucello, Aline; Beloti, Lilian L; Romero, Fabian; Tasic, Ljubica; de Souza, Alessandra A; de Souza, Anete P

    2015-01-01

    Xylella fastidiosa strain 9a5c is a gram-negative phytopathogen that is the causal agent of citrus variegated chlorosis (CVC), a disease that is responsible for economic losses in Brazilian agriculture. The most well-known mechanism of pathogenicity for this bacterial pathogen is xylem vessel occlusion, which results from bacterial movement and the formation of biofilms. The molecular mechanisms underlying the virulence caused by biofilm formation are unknown. Here, we provide evidence showing that virulence-associated protein D in X. fastidiosa (Xf-VapD) is a thermostable protein with ribonuclease activity. Moreover, protein expression analyses in two X. fastidiosa strains, including virulent (Xf9a5c) and nonpathogenic (XfJ1a12) strains, showed that Xf-VapD was expressed during all phases of development in both strains and that increased expression was observed in Xf9a5c during biofilm growth. This study is an important step toward characterizing and improving our understanding of the biological significance of Xf-VapD and its potential functions in the CVC pathosystem.

  5. A chitinase is required for Xylella fastidiosa colonization of its insect and plant hosts.

    Science.gov (United States)

    Labroussaa, Fabien; Ionescu, Michael; Zeilinger, Adam R; Lindow, Steven E; Almeida, Rodrigo P P

    2017-04-01

    Xylella fastidiosa colonizes the xylem network of host plant species as well as the foregut of its required insect vectors to ensure efficient propagation. Disease management strategies remain inefficient due to a limited comprehension of the mechanisms governing both insect and plant colonization. It was previously shown that X. fastidiosa has a functional chitinase (ChiA), and that chitin likely serves as a carbon source for this bacterium. We expand on that research, showing that a chiA mutant strain is unable to grow on chitin as the sole carbon source. Quantitative PCR assays allowed us to detect bacterial cells in the foregut of vectors after pathogen acquisition; populations of the wild-type and complemented mutant strain were both significantly larger than the chiA mutant strain 10 days, but not 3 days, post acquisition. These results indicate that adhesion of the chiA mutant strain to vectors may not be impaired, but that cell multiplication is limited. The mutant was also affected in its transmission by vectors to plants. In addition, the chiA mutant strain was unable to colonize host plants, suggesting that the enzyme has other substrates associated with plant colonization. Lastly, ChiA requires other X. fastidiosa protein(s) for its in vitro chitinolytic activity. The observation that the chiA mutant strain is not able to colonize plants warrants future attention to be paid to the substrates for this enzyme.

  6. The Bacterial Pathogen Xylella fastidiosa Affects the Leaf Ionome of Plant Hosts during Infection

    Science.gov (United States)

    De La Fuente, Leonardo; Parker, Jennifer K.; Oliver, Jonathan E.; Granger, Shea; Brannen, Phillip M.; van Santen, Edzard; Cobine, Paul A.

    2013-01-01

    Xylella fastidiosa is a plant pathogenic bacterium that lives inside the host xylem vessels, where it forms biofilm believed to be responsible for disrupting the passage of water and nutrients. Here, Nicotiana tabacum was infected with X. fastidiosa, and the spatial and temporal changes in the whole-leaf ionome (i.e. the mineral and trace element composition) were measured as the host plant transitioned from healthy to diseased physiological status. The elemental composition of leaves was used as an indicator of the physiological changes in the host at a specific time and relative position during plant development. Bacterial infection was found to cause significant increases in concentrations of calcium prior to the appearance of symptoms and decreases in concentrations of phosphorous after symptoms appeared. Field-collected leaves from multiple varieties of grape, blueberry, and pecan plants grown in different locations over a four-year period in the Southeastern US showed the same alterations in Ca and P. This descriptive ionomics approach characterizes the existence of a mineral element-based response to X. fastidiosa using a model system suitable for further manipulation to uncover additional details of the role of mineral elements during plant-pathogen interactions. This is the first report on the dynamics of changes in the ionome of the host plant throughout the process of infection by a pathogen. PMID:23667547

  7. Population structure of the bacterial pathogen Xylella fastidiosa among street trees in Washington D.C.

    Science.gov (United States)

    Harris, Jordan Lee; Balci, Yilmaz

    2015-01-01

    Bacterial leaf scorch, associated with the bacterial pathogen Xylella fastidiosa, is a widely established and problematic disease of landscape ornamentals in Washington D.C. A multi-locus sequence typing analysis was performed using 10 housekeeping loci for X. fastidiosa strains in order to better understand the epidemiology of leaf scorch disease in this municipal environment. Samples were collected from 7 different tree species located throughout the District of Columbia, consisting of 101 samples of symptomatic and asymptomatic foliage from 84 different trees. Five strains of the bacteria were identified. Consistent with prior data, these strains were host specific, with only one strain associated with members of the red oak family, one strain associated with American elm, one strain associated with American sycamore, and two strains associated with mulberry. Strains found for asymptomatic foliage were the same as strains from the symptomatic foliage on individual trees. Cross transmission of the strains was not observed at sites with multiple species of infected trees within an approx. 25 m radius of one another. X. fastidiosa strain specificity observed for each genus of tree suggests a highly specialized host-pathogen relationship.

  8. The bacterial pathogen Xylella fastidiosa affects the leaf ionome of plant hosts during infection.

    Directory of Open Access Journals (Sweden)

    Leonardo De La Fuente

    Full Text Available Xylella fastidiosa is a plant pathogenic bacterium that lives inside the host xylem vessels, where it forms biofilm believed to be responsible for disrupting the passage of water and nutrients. Here, Nicotiana tabacum was infected with X. fastidiosa, and the spatial and temporal changes in the whole-leaf ionome (i.e. the mineral and trace element composition were measured as the host plant transitioned from healthy to diseased physiological status. The elemental composition of leaves was used as an indicator of the physiological changes in the host at a specific time and relative position during plant development. Bacterial infection was found to cause significant increases in concentrations of calcium prior to the appearance of symptoms and decreases in concentrations of phosphorous after symptoms appeared. Field-collected leaves from multiple varieties of grape, blueberry, and pecan plants grown in different locations over a four-year period in the Southeastern US showed the same alterations in Ca and P. This descriptive ionomics approach characterizes the existence of a mineral element-based response to X. fastidiosa using a model system suitable for further manipulation to uncover additional details of the role of mineral elements during plant-pathogen interactions. This is the first report on the dynamics of changes in the ionome of the host plant throughout the process of infection by a pathogen.

  9. VapD in Xylella fastidiosa Is a Thermostable Protein with Ribonuclease Activity.

    Directory of Open Access Journals (Sweden)

    Juliano S Mendes

    Full Text Available Xylella fastidiosa strain 9a5c is a gram-negative phytopathogen that is the causal agent of citrus variegated chlorosis (CVC, a disease that is responsible for economic losses in Brazilian agriculture. The most well-known mechanism of pathogenicity for this bacterial pathogen is xylem vessel occlusion, which results from bacterial movement and the formation of biofilms. The molecular mechanisms underlying the virulence caused by biofilm formation are unknown. Here, we provide evidence showing that virulence-associated protein D in X. fastidiosa (Xf-VapD is a thermostable protein with ribonuclease activity. Moreover, protein expression analyses in two X. fastidiosa strains, including virulent (Xf9a5c and nonpathogenic (XfJ1a12 strains, showed that Xf-VapD was expressed during all phases of development in both strains and that increased expression was observed in Xf9a5c during biofilm growth. This study is an important step toward characterizing and improving our understanding of the biological significance of Xf-VapD and its potential functions in the CVC pathosystem.

  10. Population structure of the bacterial pathogen Xylella fastidiosa among street trees in Washington D.C.

    Directory of Open Access Journals (Sweden)

    Jordan Lee Harris

    Full Text Available Bacterial leaf scorch, associated with the bacterial pathogen Xylella fastidiosa, is a widely established and problematic disease of landscape ornamentals in Washington D.C. A multi-locus sequence typing analysis was performed using 10 housekeeping loci for X. fastidiosa strains in order to better understand the epidemiology of leaf scorch disease in this municipal environment. Samples were collected from 7 different tree species located throughout the District of Columbia, consisting of 101 samples of symptomatic and asymptomatic foliage from 84 different trees. Five strains of the bacteria were identified. Consistent with prior data, these strains were host specific, with only one strain associated with members of the red oak family, one strain associated with American elm, one strain associated with American sycamore, and two strains associated with mulberry. Strains found for asymptomatic foliage were the same as strains from the symptomatic foliage on individual trees. Cross transmission of the strains was not observed at sites with multiple species of infected trees within an approx. 25 m radius of one another. X. fastidiosa strain specificity observed for each genus of tree suggests a highly specialized host-pathogen relationship.

  11. Homologous Recombination and Xylella fastidiosa Host-Pathogen Associations in South America.

    Science.gov (United States)

    Coletta-Filho, Helvécio D; Francisco, Carolina S; Lopes, João R S; Muller, Christiane; Almeida, Rodrigo P P

    2017-03-01

    Homologous recombination affects the evolution of bacteria such as Xylella fastidiosa, a naturally competent plant pathogen that requires insect vectors for dispersal. This bacterial species is taxonomically divided into subspecies, with phylogenetic clusters within subspecies that are host specific. One subspecies, pauca, is primarily limited to South America, with the exception of recently reported strains in Europe and Costa Rica. Despite the economic importance of X. fastidiosa subsp. pauca in South America, little is known about its genetic diversity. Multilocus sequence typing (MLST) has previously identified six sequence types (ST) among plant samples collected in Brazil (both subsp. pauca and multiplex). Here, we report on a survey of X. fastidiosa genetic diversity (MLST based) performed in six regions in Brazil and two in Argentina, by sampling five different plant species. In addition to the six previously reported ST, seven new subsp. pauca and two new subsp. multiplex ST were identified. The presence of subsp. multiplex in South America is considered to be the consequence of a single introduction from its native range in North America more than 80 years ago. Different phylogenetic approaches clustered the South American ST into four groups, with strains infecting citrus (subsp. pauca); coffee and olive (subsp. pauca); coffee, hibiscus, and plum (subsp. pauca); and plum (subsp. multiplex). In areas where these different genetic clusters occurred sympatrically, we found evidence of homologous recombination in the form of bidirectional allelic exchange between subspp. pauca and multiplex. In fact, the only strain of subsp. pauca isolated from a plum host had an allele that originated from subsp. multiplex. These signatures of bidirectional homologous recombination between endemic and introduced ST indicate that gene flow occurs in short evolutionary time frames in X. fastidiosa, despite the ecological isolation (i.e., host plant species) of genotypes.

  12. A cell-cell signaling sensor is required for virulence and insect transmission of Xylella fastidiosa.

    Science.gov (United States)

    Chatterjee, Subhadeep; Wistrom, Christina; Lindow, Steven E

    2008-02-19

    Cell-cell signaling in Xylella fastidiosa, a xylem-colonizing plant pathogenic bacterium, mediated by a fatty acid Diffusible Signaling Factor (DSF), is required to colonize insect vectors and to suppress virulence to grape. Here, we show that a hybrid two-component regulatory protein RpfC is involved in negative regulation of DSF synthesis by RpfF in X. fastidiosa. X. fastidiosa rpfC mutants hyperexpress rpfF and overproduce DSF and are deficient in virulence and movement in the xylem vessels of grape. The expression of the genes encoding the adhesins FimA, HxfA, and HxfB is much higher in rpfC mutants, which also exhibit a hyperattachment phenotype in culture that is associated with their inability to migrate in xylem vessels and cause disease. rpfF mutants deficient in DSF production have the opposite phenotypes for all of these traits. RpfC is also involved in the regulation of other signaling components including rpfG, rpfB, a GGDEF domain protein that may be involved in intracellular signaling by modulating the levels of cyclic-di-GMP, and the virulence factors tolC and pglA required for disease. rpfC mutants are able to colonize the mouthparts of insect vectors and wild-type strains but are not transmitted as efficiently to new host plants, apparently because of their high levels of adhesiveness. Because of the conflicting contributions of adhesiveness and other traits to movement within plants and vectoring to new host plants, X. fastidiosa apparently coordinates these traits in a population-size-dependent fashion involving accumulation of DSF.

  13. Global gene expression under nitrogen starvation in Xylella fastidiosa: contribution of the σ54 regulon

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    da Silva Neto José F

    2010-08-01

    Full Text Available Abstract Background Xylella fastidiosa, a Gram-negative fastidious bacterium, grows in the xylem of several plants causing diseases such as citrus variegated chlorosis. As the xylem sap contains low concentrations of amino acids and other compounds, X. fastidiosa needs to cope with nitrogen limitation in its natural habitat. Results In this work, we performed a whole-genome microarray analysis of the X. fastidiosa nitrogen starvation response. A time course experiment (2, 8 and 12 hours of cultures grown in defined medium under nitrogen starvation revealed many differentially expressed genes, such as those related to transport, nitrogen assimilation, amino acid biosynthesis, transcriptional regulation, and many genes encoding hypothetical proteins. In addition, a decrease in the expression levels of many genes involved in carbon metabolism and energy generation pathways was also observed. Comparison of gene expression profiles between the wild type strain and the rpoN null mutant allowed the identification of genes directly or indirectly induced by nitrogen starvation in a σ54-dependent manner. A more complete picture of the σ54 regulon was achieved by combining the transcriptome data with an in silico search for potential σ54-dependent promoters, using a position weight matrix approach. One of these σ54-predicted binding sites, located upstream of the glnA gene (encoding glutamine synthetase, was validated by primer extension assays, confirming that this gene has a σ54-dependent promoter. Conclusions Together, these results show that nitrogen starvation causes intense changes in the X. fastidiosa transcriptome and some of these differentially expressed genes belong to the σ54 regulon.

  14. Intersubspecific recombination in Xylella fastidiosa Strains native to the United States: infection of novel hosts associated with an unsuccessful invasion.

    Science.gov (United States)

    Nunney, Leonard; Hopkins, Donald L; Morano, Lisa D; Russell, Stephanie E; Stouthamer, Richard

    2014-02-01

    The bacterial pathogen Xylella fastidiosa infects xylem and causes disease in many plant species in the Americas. Different subspecies of this bacterium and different genotypes within subspecies infect different plant hosts, but the genetics of host adaptation are unknown. Here we examined the hypothesis that the introduction of novel genetic variation via intersubspecific homologous recombination (IHR) facilitates host shifts. We investigated IHR in 33 X. fastidiosa subsp. multiplex isolates previously identified as recombinant based on 8 loci (7 multilocus sequence typing [MLST] loci plus 1 locus). We found significant evidence of introgression from X. fastidiosa subsp. fastidiosa in 4 of the loci and, using published data, evidence of IHR in 6 of 9 additional loci. Our data showed that IHR regions in 2 of the 4 loci were inconsistent (12 mismatches) with X. fastidiosa subsp. fastidiosa alleles found in the United States but consistent with alleles from Central America. The other two loci were consistent with alleles from both regions. We propose that the recombinant forms all originated via genomewide recombination of one X. fastidiosa subsp. multiplex ancestor with one X. fastidiosa subsp. fastidiosa donor from Central America that was introduced into the United States but subsequently disappeared. Using all of the available data, 5 plant hosts of the recombinant types were identified, 3 of which also supported non-IHR X. fastidiosa subsp. multiplex, but 2 were unique to recombinant types from blueberry (7 isolates from Georgia, 3 from Florida); and blackberry (1 each from Florida and North Carolina), strongly supporting the hypothesis that IHR facilitated a host shift to blueberry and possibly blackberry.

  15. Crystal structure of a small heat-shock protein from Xylella fastidiosa reveals a distinct high-order structure.

    Science.gov (United States)

    Fonseca, Emanuella Maria Barreto; Scorsato, Valéria; Dos Santos, Marcelo Leite; Júnior, Atilio Tomazini; Tada, Susely Ferraz Siqueira; Dos Santos, Clelton Aparecido; de Toledo, Marcelo Augusto Szymanski; de Souza, Anete Pereira; Polikarpov, Igor; Aparicio, Ricardo

    2017-04-01

    Citrus variegated chlorosis is a disease that attacks economically important citrus plantations and is caused by the plant-pathogenic bacterium Xylella fastidiosa. In this work, the structure of a small heat-shock protein from X. fastidiosa (XfsHSP17.9) is reported. The high-order structures of small heat-shock proteins from other organisms are arranged in the forms of double-disc, hollow-sphere or spherical assemblies. Unexpectedly, the structure reported here reveals a high-order architecture forming a nearly square cavity.

  16. Search of Xylella fastidiosa in plants with symptoms of chlorosis and leaf scorch present in ornamental areas in the university district and nearness

    International Nuclear Information System (INIS)

    Pacheco Acevedo, Maria Jose

    2010-01-01

    The presence of Xylella fastidiosa is detected in plants considered as hosts of urban ornament with characteristic symptoms, present in gardens in the area of the university district and around in the area. Urban ornamental plants are identified with suggestive symptomatology of infection by X. fastidiosa in gardens of urban areas. Plants are classified according to the presence or absence of symptoms. In the study were gathered 97 samples, belonging to 29 vegetable species with symptoms of chlorosis on leaves, leaf scorch, delay in the development and loss of foliage. The identified plants are screened by techniques of ELISA, and immunofluorescence for the detection of X. fastidiosa. Xylella fastidiosa is isolated from urban ornamental plants. The isolates of Xylella obtained, are characterized phenotypic and molecularly. The performance of two standardized immunological techniques are compared for the serological detection of Xylella fastidiosa. The presence of X. fastidiosa is detected, using the DAS-ELISA technique on 48 of the 97 processed samples, corresponding to 46,1% of the samples. Parallely, the samples processed using the IFA technique, have detected the presence of X. fastidiosa in the same proportions [es

  17. Nanofilms of hyaluronan/chitosan assembled layer-by-layer: An antibacterial surface for Xylella fastidiosa.

    Science.gov (United States)

    Hernández-Montelongo, Jacobo; Nascimento, Vicente F; Murillo, Duber; Taketa, Thiago B; Sahoo, Prasana; de Souza, Alessandra A; Beppu, Marisa M; Cotta, Monica A

    2016-01-20

    In this work, nanofilms of hyaluronan/chitosan (HA/CHI) assembled layer by layer were synthesized; their application as a potential antimicrobial material was demonstrated for the phytopathogen Xylella fastidiosa, a gram-negative bacterium, here used as a model. For the synthesis, the influence of pH and ionic strength of these natural polymer stem-solutions on final characteristics of the HA/CHI nanofilms was studied in detail. The antibacterial effect was evaluated using widefield fluorescence microscopy. These results were correlated with the chemical properties of the nanofilms, studied by FTIR and Raman spectroscopy, as well as with their morphology and surface properties characterized using SEM and AFM. The present findings can be extended to design and optimize HA/CHI nanofilms with enhanced antimicrobial behavior for other type of phytopathogenic gram-negative bacteria species, such as Xanthomonas citri, Xanthomas campestri and Ralstonia solanacearum. Copyright © 2015 Elsevier Ltd. All rights reserved.

  18. Lipopolysaccharide O-antigen delays plant innate immune recognition of Xylella fastidiosa.

    Science.gov (United States)

    Rapicavoli, Jeannette N; Blanco-Ulate, Barbara; Muszyński, Artur; Figueroa-Balderas, Rosa; Morales-Cruz, Abraham; Azadi, Parastoo; Dobruchowska, Justyna M; Castro, Claudia; Cantu, Dario; Roper, M Caroline

    2018-01-26

    Lipopolysaccharides (LPS) are among the known pathogen-associated molecular patterns (PAMPs). LPSs are potent elicitors of PAMP-triggered immunity (PTI), and bacteria have evolved intricate mechanisms to dampen PTI. Here we demonstrate that Xylella fastidiosa (Xf), a hemibiotrophic plant pathogenic bacterium, possesses a long chain O-antigen that enables it to delay initial plant recognition, thereby allowing it to effectively skirt initial elicitation of innate immunity and establish itself in the host. Lack of the O-antigen modifies plant perception of Xf and enables elicitation of hallmarks of PTI, such as ROS production specifically in the plant xylem tissue compartment, a tissue not traditionally considered a spatial location of PTI. To explore translational applications of our findings, we demonstrate that pre-treatment of plants with Xf LPS primes grapevine defenses to confer tolerance to Xf challenge.

  19. Expression of Xylella fastidiosa RpfF in citrus disrupts signaling in Xanthomonas citri subsp. citri and thereby its virulence.

    Science.gov (United States)

    Caserta, R; Picchi, S C; Takita, M A; Tomaz, J P; Pereira, W E L; Machado, M A; Ionescu, M; Lindow, S; De Souza, A A

    2014-11-01

    Xylella fastidiosa and Xanthomonas citri subsp. citri, that cause citrus variegated chlorosis (CVC) and citrus canker diseases, respectively, utilize diffusible signal factor (DSF) for quorum sensing. DSF, produced by RpfF, are similar fatty acids in both organisms, although a different set of genes is regulated by DSF in each species. Because of this similarity, Xylella fastidiosa DSF might be recognized and affect the biology of Xanthomonas citri. Therefore, transgenic Citrus sinensis and Carrizo citrange plants overexpressing the Xylella fastidiosa rpfF were inoculated with Xanthomonas citri and changes in symptoms of citrus canker were observed. X. citri biofilms formed only at wound sites on transgenic leaves and were thicker; however, bacteria were unable to break through the tissue and form pustules elsewhere. Although abundant growth of X. citri occurred at wound sites on inoculated transgenic leaves, little growth was observed on unwounded tissue. Genes in the DFS-responsive core in X. citri were downregulated in bacteria isolated from transgenic leaves. DSF-dependent expression of engA was suppressed in cells exposed to xylem sap from transgenic plants. Thus, altered symptom development appears to be due to reduced expression of virulence genes because of the presence of antagonists of DSF signaling in X. citri in rpfF-expressing plants.

  20. Spatiotemporal distribution of different extracellular polymeric substances and filamentation mediate Xylella fastidiosa adhesion and biofilm formation.

    Science.gov (United States)

    Janissen, Richard; Murillo, Duber M; Niza, Barbara; Sahoo, Prasana K; Nobrega, Marcelo M; Cesar, Carlos L; Temperini, Marcia L A; Carvalho, Hernandes F; de Souza, Alessandra A; Cotta, Monica A

    2015-04-20

    Microorganism pathogenicity strongly relies on the generation of multicellular assemblies, called biofilms. Understanding their organization can unveil vulnerabilities leading to potential treatments; spatially and temporally-resolved comprehensive experimental characterization can provide new details of biofilm formation, and possibly new targets for disease control. Here, biofilm formation of economically important phytopathogen Xylella fastidiosa was analyzed at single-cell resolution using nanometer-resolution spectro-microscopy techniques, addressing the role of different types of extracellular polymeric substances (EPS) at each stage of the entire bacterial life cycle. Single cell adhesion is caused by unspecific electrostatic interactions through proteins at the cell polar region, where EPS accumulation is required for more firmly-attached, irreversibly adhered cells. Subsequently, bacteria form clusters, which are embedded in secreted loosely-bound EPS, and bridged by up to ten-fold elongated cells that form the biofilm framework. During biofilm maturation, soluble EPS forms a filamentous matrix that facilitates cell adhesion and provides mechanical support, while the biofilm keeps anchored by few cells. This floating architecture maximizes nutrient distribution while allowing detachment upon larger shear stresses; it thus complies with biological requirements of the bacteria life cycle. Using new approaches, our findings provide insights regarding different aspects of the adhesion process of X. fastidiosa and biofilm formation.

  1. The exopolysaccharide of Xylella fastidiosa is essential for biofilm formation, plant virulence, and vector transmission.

    Science.gov (United States)

    Killiny, N; Martinez, R Hernandez; Dumenyo, C Korsi; Cooksey, D A; Almeida, R P P

    2013-09-01

    Exopolysaccharides (EPS) synthesized by plant-pathogenic bacteria are generally essential for virulence. The role of EPS produced by the vector-transmitted bacterium Xylella fastidiosa was investigated by knocking out two genes implicated in the EPS biosynthesis, gumD and gumH. Mutant strains were affected in growth characteristics in vitro, including adhesion to surfaces and biofilm formation. In addition, different assays were used to demonstrate that the mutant strains produced significantly less EPS compared with the wild type. Furthermore, gas chromatography-mass spectrometry showed that both mutant strains did not produce oligosaccharides. Biologically, the mutants were deficient in movement within plants, resulting in an avirulent phenotype. Additionally, mutant strains were affected in transmission by insects: they were very poorly transmitted by and retained within vectors. The gene expression profile indicated upregulation of genes implicated in cell-to-cell signaling and adhesins while downregulation in genes was required for within-plant movement in EPS-deficient strains. These results suggest an essential role for EPS in X. fastidiosa interactions with both plants and insects.

  2. Production of Xylella fastidiosa diffusible signal factor in transgenic grape causes pathogen confusion and reduction in severity of Pierce's disease.

    Science.gov (United States)

    Lindow, Steven; Newman, Karyn; Chatterjee, Subhadeep; Baccari, Clelia; Lavarone, Anthony T; Ionescu, Michael

    2014-03-01

    The rpfF gene from Xylella fastidiosa, encoding the synthase for diffusible signal factor (DSF), was expressed in 'Freedom' grape to reduce the pathogen's growth and mobility within the plant. Symptoms in such plants were restricted to near the point of inoculation and incidence of disease was two- to fivefold lower than in the parental line. Both the longitudinal and lateral movement of X. fastidiosa in the xylem was also much lower. DSF was detected in both leaves and xylem sap of RpfF-expressing plants using biological sensors, and both 2-Z-tetradecenoic acid, previously identified as a component of X. fastidiosa DSF, and cis-11-methyl-2-dodecenoic acid were detected in xylem sap using electrospray ionization mass spectrometry. A higher proportion of X. fastidiosa cells adhered to xylem vessels of the RpfF-expressing line than parental 'Freedom' plants, reflecting a higher adhesiveness of the pathogen in the presence of DSF. Disease incidence in RpfF-expressing plants in field trials in which plants were either mechanically inoculated with X. fastidiosa or subjected to natural inoculation by sharpshooter vectors was two- to fourfold lower in than that of the parental line. The number of symptomatic leaves on infected shoots was reduced proportionally more than the incidence of infection, reflecting a decreased ability of X. fastidiosa to move within DSF-producing plants.

  3. Xylella fastidiosa differentially accumulates mineral elements in biofilm and planktonic cells.

    Science.gov (United States)

    Cobine, Paul A; Cruz, Luisa F; Navarrete, Fernando; Duncan, Daniel; Tygart, Melissa; De La Fuente, Leonardo

    2013-01-01

    Xylella fastidiosa is a bacterial plant pathogen that infects numerous plant hosts. Disease develops when the bacterium colonizes the xylem vessels and forms a biofilm. Inductively coupled plasma optical emission spectroscopy was used to examine the mineral element content of this pathogen in biofilm and planktonic states. Significant accumulations of copper (30-fold), manganese (6-fold), zinc (5-fold), calcium (2-fold) and potassium (2-fold) in the biofilm compared to planktonic cells were observed. Other mineral elements such as sodium, magnesium and iron did not significantly differ between biofilm and planktonic cells. The distribution of mineral elements in the planktonic cells loosely mirrors the media composition; however the unique mineral element distribution in biofilm suggests specific mechanisms of accumulation from the media. A cell-to-surface attachment assay shows that addition of 50 to 100 µM Cu to standard X. fastidiosa media increases biofilm, while higher concentrations (>200 µM) slow cell growth and prevent biofilm formation. Moreover cell-to-surface attachment was blocked by specific chelation of copper. Growth of X. fastidiosa in microfluidic chambers under flow conditions showed that addition of 50 µM Cu to the media accelerated attachment and aggregation, while 400 µM prevented this process. Supplementation of standard media with Mn showed increased biofilm formation and cell-to-cell attachment. In contrast, while the biofilm accumulated Zn, supplementation to the media with this element caused inhibited growth of planktonic cells and impaired biofilm formation. Collectively these data suggest roles for these minerals in attachment and biofilm formation and therefore the virulence of this pathogen.

  4. O antigen modulates insect vector acquisition of the bacterial plant pathogen Xylella fastidiosa.

    Science.gov (United States)

    Rapicavoli, Jeannette N; Kinsinger, Nichola; Perring, Thomas M; Backus, Elaine A; Shugart, Holly J; Walker, Sharon; Roper, M Caroline

    2015-12-01

    Hemipteran insect vectors transmit the majority of plant pathogens. Acquisition of pathogenic bacteria by these piercing/sucking insects requires intimate associations between the bacterial cells and insect surfaces. Lipopolysaccharide (LPS) is the predominant macromolecule displayed on the cell surface of Gram-negative bacteria and thus mediates bacterial interactions with the environment and potential hosts. We hypothesized that bacterial cell surface properties mediated by LPS would be important in modulating vector-pathogen interactions required for acquisition of the bacterial plant pathogen Xylella fastidiosa, the causative agent of Pierce's disease of grapevines. Utilizing a mutant that produces truncated O antigen (the terminal portion of the LPS molecule), we present results that link this LPS structural alteration to a significant decrease in the attachment of X. fastidiosa to blue-green sharpshooter foreguts. Scanning electron microscopy confirmed that this defect in initial attachment compromised subsequent biofilm formation within vector foreguts, thus impairing pathogen acquisition. We also establish a relationship between O antigen truncation and significant changes in the physiochemical properties of the cell, which in turn affect the dynamics of X. fastidiosa adhesion to the vector foregut. Lastly, we couple measurements of the physiochemical properties of the cell with hydrodynamic fluid shear rates to produce a Comsol model that predicts primary areas of bacterial colonization within blue-green sharpshooter foreguts, and we present experimental data that support the model. These results demonstrate that, in addition to reported protein adhesin-ligand interactions, O antigen is crucial for vector-pathogen interactions, specifically in the acquisition of this destructive agricultural pathogen. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  5. Xylella fastidiosa differentially accumulates mineral elements in biofilm and planktonic cells.

    Directory of Open Access Journals (Sweden)

    Paul A Cobine

    Full Text Available Xylella fastidiosa is a bacterial plant pathogen that infects numerous plant hosts. Disease develops when the bacterium colonizes the xylem vessels and forms a biofilm. Inductively coupled plasma optical emission spectroscopy was used to examine the mineral element content of this pathogen in biofilm and planktonic states. Significant accumulations of copper (30-fold, manganese (6-fold, zinc (5-fold, calcium (2-fold and potassium (2-fold in the biofilm compared to planktonic cells were observed. Other mineral elements such as sodium, magnesium and iron did not significantly differ between biofilm and planktonic cells. The distribution of mineral elements in the planktonic cells loosely mirrors the media composition; however the unique mineral element distribution in biofilm suggests specific mechanisms of accumulation from the media. A cell-to-surface attachment assay shows that addition of 50 to 100 µM Cu to standard X. fastidiosa media increases biofilm, while higher concentrations (>200 µM slow cell growth and prevent biofilm formation. Moreover cell-to-surface attachment was blocked by specific chelation of copper. Growth of X. fastidiosa in microfluidic chambers under flow conditions showed that addition of 50 µM Cu to the media accelerated attachment and aggregation, while 400 µM prevented this process. Supplementation of standard media with Mn showed increased biofilm formation and cell-to-cell attachment. In contrast, while the biofilm accumulated Zn, supplementation to the media with this element caused inhibited growth of planktonic cells and impaired biofilm formation. Collectively these data suggest roles for these minerals in attachment and biofilm formation and therefore the virulence of this pathogen.

  6. Ionome changes in Xylella fastidiosa-infected Nicotiana tabacum correlate with virulence and discriminate between subspecies of bacterial isolates.

    Science.gov (United States)

    Oliver, J E; Sefick, S A; Parker, J K; Arnold, T; Cobine, P A; De La Fuente, L

    2014-10-01

    Characterization of ionomes has been used to uncover the basis of nutrient utilization and environmental adaptation of plants. Here, ionomic profiles were used to understand the phenotypic response of a plant to infection by genetically diverse isolates of Xylella fastidiosa, a gram-negative, xylem-limited bacterial plant pathogen. In this study, X. fastidiosa isolates were used to infect a common model host (Nicotiana tabacum 'SR1'), and leaf and sap concentrations of eleven elements together with plant colonization and symptoms were assessed. Multivariate statistical analysis revealed that changes in the ionome were significantly correlated with symptom severity and bacterial populations in host petioles. Moreover, plant ionome modification by infection could be used to differentiate the X. fastidiosa subspecies with which the plant was infected. This report establishes host ionome modification as a phenotypic response to infection.

  7. Population Structure of Xylella fastidiosa Associated with Almond Leaf Scorch Disease in the San Joaquin Valley of California.

    Science.gov (United States)

    Lin, Hong; Islam, Md Sajedul; Cabrera-La Rosa, Juan C; Civerolo, Edwin L; Groves, Russell L

    2015-06-01

    Xylella fastidiosa causes disease in many commercial crops, including almond leaf scorch (ALS) disease in susceptible almond (Prunus dulcis). In this study, genetic diversity and population structure of X. fastidiosa associated with ALS disease were evaluated. Isolates obtained from two almond orchards in Fresno and Kern County in the San Joaquin Valley of California were analyzed for two successive years. Multilocus simple-sequence repeat (SSR) analysis revealed two major genetic clusters that were associated with two host cultivars, 'Sonora' and 'Nonpareil', respectively, regardless of the year of study or location of the orchard. These relationships suggest that host cultivar selection and adaptation are major driving forces shaping ALS X. fastidiosa population structure in the San Joaquin Valley. This finding will provide insight into understanding pathogen adaptation and host selection in the context of ALS disease dynamics.

  8. An Evaluation of the Genetic Diversity of Xylella fastidiosa Isolated from Diseased Citrus and Coffee in São Paulo, Brazil.

    Science.gov (United States)

    Qin, X; Miranda, V S; Machado, M A; Lemos, E G; Hartung, J S

    2001-06-01

    ABSTRACT Strains of Xylella fastidiosa, isolated from sweet orange trees (Citrus sinensis) and coffee trees (Coffea arabica) with symptoms of citrus variegated chlorosis and Requeima do Café, respectively, were indistinguishable based on repetitive extragenic palindromic polymerase chain reaction (PCR) and enterobacterial repetitive intergenic consensus PCR assays. These strains were also indistinguishable with a previously described PCR assay that distinguished the citrus strains from all other strains of Xylella fastidiosa. Because we were not able to document any genomic diversity in our collection of Xylella fastidiosa strains isolated from diseased citrus, the observed gradient of increasing disease severity from southern to northern regions of São Paulo State is unlikely due to the presence of significantly different strains of the pathogen in the different regions. When comparisons were made to reference strains of Xylella fastidiosa isolated from other hosts using these methods, four groups were consistently identified consistent with the hosts and regions from which the strains originated: citrus and coffee, grapevine and almond, mulberry, and elm, plum, and oak. Independent results from random amplified polymorphic DNA (RAPD) PCR assays were also consistent with these results; however, two of the primers tested in RAPD-PCR were able to distinguish the coffee and citrus strains. Sequence comparisons of a PCR product amplified from all strains of Xylella fastidiosa confirmed the presence of a CfoI polymorphism that can be used to distinguish the citrus strains from all others. The ability to distinguish Xylella fastidiosa strains from citrus and coffee with a PCR-based assay will be useful in epidemiological and etiological studies of this pathogen.

  9. Calcium-Enhanced Twitching Motility in Xylella fastidiosa Is Linked to a Single PilY1 Homolog.

    Science.gov (United States)

    Cruz, Luisa F; Parker, Jennifer K; Cobine, Paul A; De La Fuente, Leonardo

    2014-12-01

    The plant-pathogenic bacterium Xylella fastidiosa is restricted to the xylem vessel environment, where mineral nutrients are transported through the plant host; therefore, changes in the concentrations of these elements likely impact the growth and virulence of this bacterium. Twitching motility, dependent on type IV pili (TFP), is required for movement against the transpiration stream that results in basipetal colonization. We previously demonstrated that calcium (Ca) increases the motility of X. fastidiosa, although the mechanism was unknown. PilY1 is a TFP structural protein recently shown to bind Ca and to regulate twitching and adhesion in bacterial pathogens of humans. Sequence analysis identified three pilY1 homologs in X. fastidiosa (PD0023, PD0502, and PD1611), one of which (PD1611) contains a Ca-binding motif. Separate deletions of PD0023 and PD1611 resulted in mutants that still showed twitching motility and were not impaired in attachment or biofilm formation. However, the response of increased twitching at higher Ca concentrations was lost in the pilY1-1611 mutant. Ca does not modulate the expression of any of the X. fastidiosa PilY1 homologs, although it increases the expression of the retraction ATPase pilT during active movement. The evidence presented here suggests functional differences between the PilY1 homologs, which may provide X. fastidiosa with an adaptive advantage in environments with high Ca concentrations, such as xylem sap. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  10. Initial biochemical and functional characterization of a 5'-nucleotidase from Xylella fastidiosa related to the human cytosolic 5'-nucleotidase I.

    Science.gov (United States)

    Santos, Clelton A; Saraiva, Antonio M; Toledo, Marcelo A S; Beloti, Lilian L; Crucello, Aline; Favaro, Marianna T P; Horta, Maria A C; Santiago, André S; Mendes, Juliano S; Souza, Alessandra A; Souza, Anete P

    2013-01-01

    The 5'-nucleotidases constitute a ubiquitous family of enzymes that catalyze either the hydrolysis or the transfer of esterified phosphate at the 5' position of nucleoside monophosphates. These enzymes are responsible for the regulation of nucleotide and nucleoside levels in the cell and can interfere with the phosphorylation-dependent activation of nucleoside analogs used in therapies targeting solid tumors and viral infections. In the present study, we report the initial biochemical and functional characterization of a 5'-nucleotidase from Xylella fastidiosa that is related to the human cytosolic 5'-nucleotidase I. X. fastidiosa is a plant pathogenic bacterium that is responsible for numerous economically important crop diseases. Biochemical assays confirmed the phosphatase activity of the recombinant purified enzyme and revealed metal ion dependence for full enzyme activity. In addition, we investigated the involvement of Xf5'-Nt in the formation of X. fastidiosa biofilms, which are structures that occlude the xylem vessels of susceptible plants and are strictly associated with bacterial pathogenesis. Using polyclonal antibodies against Xf5'-Nt, we observed an overexpression of Xf5'-Nt during the initial phases of X. fastidiosa biofilm formation that was not observed during X. fastidiosa planktonic growth. Our results demonstrate that the de/phosphorylation network catalyzed by 5'-nucleotidases may play an important role in bacterial biofilm formation, thereby contributing novel insights into bacterial nucleotide metabolism and pathogenicity. Copyright © 2013 Elsevier Ltd. All rights reserved.

  11. A Temperature-Independent Cold-Shock Protein Homolog Acts as a Virulence Factor in Xylella fastidiosa.

    Science.gov (United States)

    Burbank, Lindsey P; Stenger, Drake C

    2016-05-01

    Xylella fastidiosa, causal agent of Pierce's disease (PD) of grapevine, is a fastidious organism that requires very specific conditions for replication and plant colonization. Cold temperatures reduce growth and survival of X. fastidiosa both in vitro and in planta. However, little is known regarding physiological responses of X. fastidiosa to temperature changes. Cold-shock proteins (CSP), a family of nucleic acid-binding proteins, act as chaperones facilitating translation at low temperatures. Bacterial genomes often encode multiple CSP, some of which are strongly induced following exposure to cold. Additionally, CSP contribute to the general stress response through mRNA stabilization and posttranscriptional regulation. A putative CSP homolog (Csp1) with RNA-binding activity was identified in X. fastidiosa Stag's Leap. The csp1 gene lacked the long 5' untranslated region characteristic of cold-inducible genes and was expressed in a temperature-independent manner. As compared with the wild type, a deletion mutant of csp1 (∆csp1) had decreased survival rates following cold exposure and salt stress in vitro. The deletion mutant also was significantly less virulent in grapevine, as compared with the wild type, in the absence of cold stress. These results suggest an important function of X. fastidiosa Csp1 in response to cellular stress and during plant colonization.

  12. A preliminary survey on the presence of Xylella fastidiosa in olive, citrus and grapevine groves in Morocco

    Directory of Open Access Journals (Sweden)

    Ahmed AARABE

    2018-03-01

    Full Text Available The bacterium Xylella fastidiosa is gram negative, xylem-inhabiting, devastating pathogen which causes various diseases on more than 300 plant hosts. Given the recent confirmed findings of X. fastidiosa in the European Union, this bacterium is becoming a serious threat to the Moroccan agricultural sector. A survey was conducted during May-September 2015 on the presence of X. fastidiosa in several commercial groves, covering olive, citrus and grapevine growing areas. In a few trees, severe symptoms which could be associated to the bacterium were observed. A total of 900 samples of different crops from different regions were randomly collected: 220 olive trees (cv. Picholine Marocaine from two regions, 410 citrus trees belonging to 7 different cultivars collected in 4 regions and 270 grapevine plants belonging to 6 different cultivars from 3 regions; all these samples were tested for the presence of X. fastidiosa by using an ELISA commercial kit. The obtained results did not show any positive sample. These preliminary results are taken as an encouraging indication, considering that X. fastidiosa was not found in Morocco, at least in the surveyed crops. However, frequent extensive surveys in different regions are needed to prevent its entrance into the country.

  13. A triply cloned strain of xylella fastidiosa multiplies and induces symptoms of citrus variegated chlorosis in sweet orange

    Science.gov (United States)

    Li; Zreik; Fernandes; Miranda; Teixeira; Ayres; Garnier; Bov

    1999-08-01

    Xylella fastidiosa isolate 8.1.b obtained from a sweet orange tree affected by citrus variegated chlorosis in the state of Sao Paulo, Brazil, and shown in 1993 to be the causal agent of the disease, was cloned by repeated culture in liquid and on solid PW medium, yielding triply cloned strain 9a5c. The eighth and the 16th passages of strain 9a5c were mechanically inoculated into sweet orange plants. Presence of X. fastidiosa in sweet orange leaves of shoots having grown after inoculation (first-flush shoots) was detected by DAS-ELISA and PCR. Thirty-eight days after inoculation, 70% of the 20 inoculated plants tested positive, and all plants gave strong positive reactions 90 days after inoculation. Symptoms first appeared after 3 months and were conspicuous after 5 months. X. fastidiosa was reisolated from sweet orange leaves, 44 days after inoculation. These results indicate that X. fastidiosa strain 9a5c, derived from pathogenic isolate 8.1.b by triply cloning, is also pathogenic. Strain 9a5c is now used for the X. fastidiosa genome sequencing project undertaken on a large scale in Brazil.http://link. springer-ny.com/link/service/journals/00284/bibs/39n2p106.html

  14. Functional characterization of replication and stability factors of an incompatibility group P-1 plasmid from Xylella fastidiosa.

    Science.gov (United States)

    Lee, Min Woo; Rogers, Elizabeth E; Stenger, Drake C

    2010-12-01

    Xylella fastidiosa strain riv11 harbors a 25-kbp plasmid (pXF-RIV11) belonging to the IncP-1 incompatibility group. Replication and stability factors of pXF-RIV11 were identified and used to construct plasmids able to replicate in X. fastidiosa and Escherichia coli. Replication in X. fastidiosa required a 1.4-kbp region from pXF-RIV11 containing a replication initiation gene (trfA) and the adjacent origin of DNA replication (oriV). Constructs containing trfA and oriV from pVEIS01, a related IncP-1 plasmid of the earthworm symbiont Verminephrobacter eiseniae, also were competent for replication in X. fastidiosa. Constructs derived from pXF-RIV11 but not pVEIS01 replicated in Agrobacterium tumefaciens, Xanthomonas campestris, and Pseudomonas syringae. Although plasmids bearing replication elements from pXF-RIV11 or pVEIS01 could be maintained in X. fastidiosa under antibiotic selection, removal of selection resulted in plasmid extinction after 3 weekly passages. Addition of a toxin-antitoxin addiction system (pemI/pemK) from pXF-RIV11 improved plasmid stability such that >80 to 90% of X. fastidiosa cells retained plasmid after 5 weekly passages in the absence of antibiotic selection. Expression of PemK in E. coli was toxic for cell growth, but toxicity was nullified by coexpression of PemI antitoxin. Deletion of N-terminal sequences of PemK containing the conserved motif RGD abolished toxicity. In vitro assays revealed a direct interaction of PemI with PemK, suggesting that antitoxin activity of PemI is mediated by toxin sequestration. IncP-1 plasmid replication and stability factors were added to an E. coli cloning vector to constitute a stable 6.0-kbp shuttle vector (pXF20-PEMIK) suitable for use in X. fastidiosa.

  15. Seasonal Fluctuations of Sap-Feeding Insect Species Infected by Xylella fastidiosa in Apulian Olive Groves of Southern Italy.

    Science.gov (United States)

    Ben Moussa, Issam Eddine; Mazzoni, Valerio; Valentini, Franco; Yaseen, Thaer; Lorusso, Donato; Speranza, Stefano; Digiaro, Michele; Varvaro, Leonardo; Krugner, Rodrigo; D'Onghia, Anna Maria

    2016-08-01

    A study on seasonal abundance of Auchenorrhyncha species and their infectivity by Xylella fastidiosa in the Apulia region of Italy was conducted to identify ideal periods for monitoring and adoption of potential control measures against insect vectors. Adult populations of Auchenorrhyncha species were monitored monthly over a 2-yr period from five olive groves. A total of 15 species were captured, identified, and tested for presence of X. fastidiosa by polymerase chain reaction (PCR). For three species, Philaenus spumarius L., Neophilaenus campestris (Fallèn), and Euscelis lineolatus Brullé, positive reactions to X. fastidiosa were obtained, on average, in 16.3, 15.9 and 18.4% of adult insects, respectively. Philaneous spumarius was the dominant species (39.8% of total Auchenorrhyncha captured) with the highest adult abundance in summer months. Adult P. spumarius and N. campestris were first detected between March and May in both years, and all insects tested during these periods (year 1: n = 42, year 2: n = 132) gave negative reactions to X. fastidiosa by PCR. Similarly, first adults of E. lineolatus that appeared from October to November (year 1: n = 20, year 2: n = 15) tested negative for presence of X. fastidiosa Given the lack of transstadial and transovarial transmission of X. fastidiosa and considering that P. spumarius is univoltine, control measures against nymphal stages of P. spumarius should be investigated as means of population suppression to reduce spread of X. fastidiosa in olive groves. © The Authors 2016. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  16. Natural Competence of Xylella fastidiosa Occurs at a High Frequency Inside Microfluidic Chambers Mimicking the Bacterium's Natural Habitats.

    Science.gov (United States)

    Kandel, Prem P; Lopez, Samantha M; Almeida, Rodrigo P P; De La Fuente, Leonardo

    2016-09-01

    Xylella fastidiosa is a xylem-limited bacterium that is the causal agent of emerging diseases in a number of economically important crops. Genetic diversity studies have demonstrated homologous recombination occurring among X. fastidiosa strains, which has been proposed to contribute to host plant shifts. Moreover, experimental evidence confirmed that X. fastidiosa is naturally competent for recombination in vitro Here, as an approximation of natural habitats (plant xylem vessels and insect mouthparts), recombination was studied in microfluidic chambers (MCs) filled with media amended with grapevine xylem sap. First, different media were screened for recombination in solid agar plates using a pair of X. fastidiosa strains that were previously reported to recombine in coculture. The highest frequency of recombination was obtained with PD3 medium, compared to those with the other two media (X. fastidiosa medium [XFM] and periwinkle wilt [PW] medium) used in previous studies. Dissection of the media components led to the identification of bovine serum albumin as an inhibitor of recombination that was correlated to its previously known effect on inhibition of twitching motility. When recombination was performed in liquid culture, the frequencies were significantly higher under flow conditions (MCs) than under batch conditions (test tubes). The recombination frequencies in MCs and agar plates were not significantly different from each other. Grapevine xylem sap from both susceptible and tolerant varieties allowed high recombination frequency in MCs when mixed with PD3. These results suggest that X. fastidiosa has the ability to be naturally competent in the natural growth environment of liquid flow, and this phenomenon could have implications in X. fastidiosa environmental adaptation. Xylella fastidiosa is a plant pathogen that lives inside xylem vessels (where water and nutrients are transported inside the plant) and the mouthparts of insect vectors. This bacterium

  17. Calcium Increases Xylella fastidiosa Surface Attachment, Biofilm Formation, and Twitching Motility

    Science.gov (United States)

    Cruz, Luisa F.; Cobine, Paul A.

    2012-01-01

    Xylella fastidiosa is a plant-pathogenic bacterium that forms biofilms inside xylem vessels, a process thought to be influenced by the chemical composition of xylem sap. In this work, the effect of calcium on the production of X. fastidiosa biofilm and movement was analyzed under in vitro conditions. After a dose-response study with 96-well plates using eight metals, the strongest increase of biofilm formation was observed when medium was supplemented with at least 1.0 mM CaCl2. The removal of Ca by extracellular (EGTA, 1.5 mM) and intracellular [1,2-bis(o-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid acetoxymethyl ester (BAPTA/AM), 75 μM] chelators reduced biofilm formation without compromising planktonic growth. The concentration of Ca influenced the force of adhesion to the substrate, biofilm thickness, cell-to-cell aggregation, and twitching motility, as shown by assays with microfluidic chambers and other assays. The effect of Ca on attachment was lost when cells were treated with tetracycline, suggesting that Ca has a metabolic or regulatory role in cell adhesion. A double mutant (fimA pilO) lacking type I and type IV pili did not improve biofilm formation or attachment when Ca was added to the medium, while single mutants of type I (fimA) or type IV (pilB) pili formed more biofilm under conditions of higher Ca concentrations. The concentration of Ca in the medium did not significantly influence the levels of exopolysaccharide produced. Our findings indicate that the role of Ca in biofilm formation may be related to the initial surface and cell-to-cell attachment and colonization stages of biofilm establishment, which rely on critical functions by fimbrial structures. PMID:22194297

  18. Intrinsic bent DNA sites in the chromosomal replication origin of Xylella fastidiosa 9a5c

    Directory of Open Access Journals (Sweden)

    F. Gimenes

    2008-04-01

    Full Text Available The features of the nucleotide sequences in both replication and promoter regions have been investigated in many organisms. Intrinsically bent DNA sites associated with transcription have been described in several prokaryotic organisms. The aim of the present study was to investigate intrinsic bent DNA sites in the segment that holds the chromosomal replication origin, oriC, of Xylella fastidiosa 9a5c. Electrophoretic behavior analyses, as well as in silico analyses of both the 2-D projection and helical parameters, were performed. The chromosomal segment analyzed contains the initial sequence of the rpmH gene, an intergenic region, the dnaA gene, the oriC sequence, and the 5' partial sequence of the dnaN gene. The analysis revealed fragments with reduced electrophoretic mobility, which indicates the presence of curved DNA segments. The analysis of the helical parameter ENDS ratio revealed three bent DNA sites (b1, b2, and b3 located in the rpmH-dnaA intergenic region, the dnaA gene, and the oriC 5' end, respectively. The chromosomal segment of X. fastidiosa analyzed here is rich in phased AT tracts and in CAnT motifs. The 2-D projection indicated a segment whose structure was determined by the cumulative effect of all bent DNA sites. Further, the in silico analysis of the three different bacterial oriC sequences indicated similar negative roll and twist >34.00° values. The DnaA box sequences, and other motifs in them, may be associated with the intrinsic DNA curvature.

  19. Diffusible Signal Factor (DSF) Synthase RpfF of Xylella fastidiosa Is a Multifunction Protein Also Required for Response to DSF

    OpenAIRE

    Ionescu, Michael; Baccari, Clelia; Da Silva, Aline Maria; Garcia, Angelica; Yokota, Kenji; Lindow, Steven E.

    2013-01-01

    Xylella fastidiosa, like related Xanthomonas species, employs an Rpf cell-cell communication system consisting of a diffusible signal factor (DSF) synthase, RpfF, and a DSF sensor, RpfC, to coordinate expression of virulence genes. While phenotypes of a ΔrpfF strain in Xanthomonas campestris could be complemented by its own DSF, the DSF produced by X. fastidiosa (XfDSF) did not restore expression of the XfDSF-dependent genes hxfA and hxfB to a ΔrpfF strain of X. fastidiosa, suggesting that Rp...

  20. Molecular characteristics of a strain (Salento-1 of Xylella fastidiosa isolated in Apulia (Italy from an olive plant with the quick decline syndrome

    Directory of Open Access Journals (Sweden)

    Gianluca BLEVE

    2016-05-01

    Full Text Available DNA-based approaches were used to characterize a strain (Salento-1 of Xylella fastidiosa obtained from an olive plant suffering from the syndrome of quick decline in Apulia (South Italy. Salento-1 was indistinguishable from strain CoDiRO previously isolated from olive in Apulia and assigned to  X. fastidiosa subsp. pauca. Based on our results and comparative analysis with reported data, the subspecies pauca, multiplex, and fastidiosa may invade olive throughout the world (California, Italy, Argentina and Brazil. The strain Salento-1 has been deposited in the National Collection of Plant Pathogenic Bacteria (NCPPB, England, and in the Belgian Coordinated Collections of Microorganisms (BCCM, Belgium.

  1. Characterization of regulatory pathways in Xylella fastidiosa: genes and phenotypes controlled by algU.

    Science.gov (United States)

    Shi, Xiang Yang; Dumenyo, C Korsi; Hernandez-Martinez, Rufina; Azad, Hamid; Cooksey, Donald A

    2007-11-01

    Many virulence genes in plant bacterial pathogens are coordinately regulated by "global" regulatory genes. Conducting DNA microarray analysis of bacterial mutants of such genes, compared with the wild type, can help to refine the list of genes that may contribute to virulence in bacterial pathogens. The regulatory gene algU, with roles in stress response and regulation of the biosynthesis of the exopolysaccharide alginate in Pseudomonas aeruginosa and many other bacteria, has been extensively studied. The role of algU in Xylella fastidiosa, the cause of Pierce's disease of grapevines, was analyzed by mutation and whole-genome microarray analysis to define its involvement in aggregation, biofilm formation, and virulence. In this study, an algU::nptII mutant had reduced cell-cell aggregation, attachment, and biofilm formation and lower virulence in grapevines. Microarray analysis showed that 42 genes had significantly lower expression in the algU::nptII mutant than in the wild type. Among these are several genes that could contribute to cell aggregation and biofilm formation, as well as other physiological processes such as virulence, competition, and survival.

  2. Analysis of expressed sequence tags from Citrus sinensis L. Osbeck infected with Xylella fastidiosa

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    Alessandra A. de Souza

    2007-01-01

    Full Text Available In order to understand the genetic responses resulting from physiological changes that occur in plants displaying citrus variegated chlorosis (CVC symptoms, we adopted a strategy of comparing two EST libraries from sweet orange [Citrus sinensis (L. Osbeck]. One of them was prepared with plants showing typical CVC symptoms caused by Xylella fastidiosa and the other with non-inoculated plants. We obtained 15,944 ESTs by sequencing the two cDNA libraries. Using an in silico hybridization strategy, 37 genes were found to have significant variation at the transcriptional level. Within this subset, 21 were up-regulated and 16 were down-regulated in plants with CVC. The main functional categories of the down-regulated transcripts in plants with CVC were associated with metabolism, protein modification, energy and transport facilitation. The majority of the up-regulated transcripts were associated with metabolism and defense response. Some transcripts associated with adaptation to stress conditions were up-regulated in plants with CVC and could explain why plants remain alive even under severe water and nutritional stress. Others of the up-regulated transcripts are related to defense response suggesting that sweet orange plants activate their defense machinery. The genes associated with stress response might be expressed as part of a secondary response related to physiological alterations caused by the infection.

  3. Fatty acid synthesis in Xylella fastidiosa: correlations between genome studies, 13C NMR data, and molecular models

    International Nuclear Information System (INIS)

    Osiro, Denise; Muniz, Joao Renato C.; Coleta Filho, Helvecio Della; Alves de Sousa, Alessandra; Machado, Marcos Antonio; Garratt, Richard C.; Colnago, Luiz Alberto

    2004-01-01

    Xylella fastidiosa was the first plant pathogen to have its complete genome sequence elucidated. Routine database analyses suggested that two enzymes essential for fatty acid synthesis were missing, one of these is the holo-acyl-carrier-protein synthase. However, here we demonstrate, using 13 C NMR spectroscopy, that X. fastidiosa is indeed able to synthesize fatty acids from acetate via an apparently conventional metabolic pathway. We further identify a gene product HetI, an alternative phosphopantetheinyl transferase, which we propose to fill the missing link. Homology modeling of HetI shows conservation of the Coenzyme A binding site suggesting it to be an active enzyme and reveals several interesting structural features when compared with the surfactin synthase-activating enzyme, on which the model was built. These include a simplified topology due to N- and C-terminal deletions and the observation of a novel serine ladder

  4. Short communication. A survey of potential insect vectors of the plant pathogenic bacterium Xylella fastidiosa in three regions of Spain

    Directory of Open Access Journals (Sweden)

    Joao R. S. Lopes

    2014-07-01

    Full Text Available The emergence of a rapid-spreading olive disease associated with Xylella fastidiosa in southern Italy represents a high risk to susceptible crops in other countries of the Mediterranean basin, if insect vectors occur in the region. The goal of this study was to identify xylem-feeding Auchenorrhyncha that could potentially act as vectors of X. fastidiosa in three regions of Spain (Andalucía, Murcia and Madrid. Samplings with sweep net and stem tap were carried out in October/2004 on grapevines and adjacent crops (olives, nectarine, citrus, Prunus spp., ornamental trees and herbaceous weeds. Yellow sticky cards were placed in ten vineyards located across 100 km in Andalucía and in three vineyards distant 10-15 km apart in Murcia. Specimens of frequently-trapped species were tested by nested- or multiplex-PCR for the presence of X. fastidiosa. The Typhlocybinae leafhopper, Austroasca (Jacobiasca lybica (Hemiptera: Cicadellidae was the most abundant species in vineyards and citrus orchards. Planthoppers (Hemiptera: Fulgoroidea and psyllids (Hemiptera: Psylloidea were prevalent on olives. Cicadellinae leafhoppers (known as sharpshooters, which are major vectors of X. fastidiosa in the Americas, were not found in the samples. The only potential vectors were spittlebugs (Hemiptera: Cercopoidea collected on Populus sp., herbaceous and on conifer trees (Pinus halepense; the spittlebug Neophileanus sp. was common on conifer trees adjacent to a vineyard in Jumilla. None of the insect samples tested positive for X. fastidiosa by PCR assays. However, spittlebugs already associated with susceptible crops in Spain may allow fast spread of X. fastidiosa in case this pathogen is introduced.

  5. Structural and Biochemical Characterization of Xylella fastidiosa DsbA Family Members: New insightsinto the Enzyme-Substrate Interaction

    Energy Technology Data Exchange (ETDEWEB)

    Rinaldi, F.; Meza, A; Gulmarges, B

    2009-01-01

    Disulfide oxidoreductase DsbA catalyzes disulfide bond formation in proteins secreted to the periplasm and has been related to the folding process of virulence factors in many organisms. It is among the most oxidizing of the thioredoxin-like proteins, and DsbA redox power is understood in terms of the electrostatic interactions involving the active site motif CPHC. The plant pathogen Xylella fastidiosa has two chromosomal genes encoding two oxidoreductases belonging to the DsbA family, and in one of them, the canonical motif CPHC is replaced by CPAC. Biochemical assays showed that both X. fastidiosa homologues have similar redox properties and the determination of the crystal structure of XfDsbA revealed substitutions in the active site of X. fastidiosa enzymes, which are proposed to compensate for the lack of the conserved histidine in XfDsbA2. In addition, electron density maps showed a ligand bound to the XfDsbA active site, allowing the characterization of the enzyme interaction with an 8-mer peptide. Finally, surface analysis of XfDsbA and XfDsbA2 suggests that X. fastidiosa enzymes may have different substrate specificities.

  6. Tolerance to oxidative stress is required for maximal xylem colonization by the xylem-limited bacterial phytopathogen, Xylella fastidiosa.

    Science.gov (United States)

    Wang, Peng; Lee, Yunho; Igo, Michele M; Roper, M Caroline

    2017-09-01

    Bacterial plant pathogens often encounter reactive oxygen species (ROS) during host invasion. In foliar bacterial pathogens, multiple regulatory proteins are involved in the sensing of oxidative stress and the activation of the expression of antioxidant genes. However, it is unclear whether xylem-limited bacteria, such as Xylella fastidiosa, experience oxidative stress during the colonization of plants. Examination of the X. fastidiosa genome uncovered only one homologue of oxidative stress regulatory proteins, OxyR. Here, a knockout mutation in the X. fastidiosa oxyR gene was constructed; the resulting strain was significantly more sensitive to hydrogen peroxide (H 2 O 2 ) relative to the wild-type. In addition, during early stages of grapevine infection, the survival rate was 1000-fold lower for the oxyR mutant than for the wild-type. This supports the hypothesis that grapevine xylem represents an oxidative environment and that X. fastidiosa must overcome this challenge to achieve maximal xylem colonization. Finally, the oxyR mutant exhibited reduced surface attachment and cell-cell aggregation and was defective in biofilm maturation, suggesting that ROS could be a potential environmental cue stimulating biofilm development during the early stages of host colonization. © 2016 BSPP AND JOHN WILEY & SONS LTD.

  7. Response of Xylella fastidiosa to zinc: decreased culturability, increased exopolysaccharide production, and formation of resilient biofilms under flow conditions.

    Science.gov (United States)

    Navarrete, Fernando; De La Fuente, Leonardo

    2014-02-01

    The bacterial plant pathogen Xylella fastidiosa produces biofilm that accumulates in the host xylem vessels, affecting disease development in various crops and bacterial acquisition by insect vectors. Biofilms are sensitive to the chemical composition of the environment, and mineral elements being transported in the xylem are of special interest for this pathosystem. Here, X. fastidiosa liquid cultures were supplemented with zinc and compared with nonamended cultures to determine the effects of Zn on growth, biofilm, and exopolysaccharide (EPS) production under batch and flow culture conditions. The results show that Zn reduces growth and biofilm production under both conditions. However, in microfluidic chambers under liquid flow and with constant bacterial supplementation (closer to conditions inside the host), a dramatic increase in biofilm aggregates was seen in the Zn-amended medium. Biofilms formed under these conditions were strongly attached to surfaces and were not removed by medium flow. This phenomenon was correlated with increased EPS production in stationary-phase cells grown under high Zn concentrations. Zn did not cause greater adhesion to surfaces by individual cells. Additionally, viability analyses suggest that X. fastidiosa may be able to enter the viable but nonculturable state in vitro, and Zn can hasten the onset of this state. Together, these findings suggest that Zn can act as a stress factor with pleiotropic effects on X. fastidiosa and indicate that, although Zn could be used as a bactericide treatment, it could trigger the undesired effect of stronger biofilm formation upon reinoculation events.

  8. Grape Cultivar and Sap Culture Conditions Affect the Development of Xylella fastidiosa Phenotypes Associated with Pierce's Disease.

    Science.gov (United States)

    Hao, Lingyun; Zaini, Paulo A; Hoch, Harvey C; Burr, Thomas J; Mowery, Patricia

    2016-01-01

    Xylella fastidiosa is a xylem-limited bacterium in plant hosts and causes Pierce's disease (PD) of grapevines, which differ in susceptibility according to the Vitis species (spp.). In this work we compared X. fastidiosa biofilm formation and population dynamics when cultured in xylem saps from PD-susceptible and -resistant Vitis spp. under different conditions. Behaviors in a closed-culture system were compared to those in different sap-renewal cultures that would more closely mimic the physicochemical environment encountered in planta. Significant differences in biofilm formation and growth in saps from PD-susceptible and -resistant spp. were only observed using sap renewal culture. Compared to saps from susceptible V. vinifera, those from PD-resistant V. aestivalis supported lower titers of X. fastidiosa and less biofilm and V. champinii suppressed both growth and biofilm formation, behaviors which are correlated with disease susceptibility. Furthermore, in microfluidic chambers X. fastidiosa formed thick mature biofilm with three-dimensional (3-D) structures, such as pillars and mounds, in saps from all susceptible spp. In contrast, only small aggregates of various shapes were formed in saps from four out of five of the resistant spp.; sap from the resistant spp. V. mustangensis was an exception in that it also supported thick lawns of biofilm but not the above described 3-D structures typically seen in a mature biofilm from the susceptible saps. Our findings provide not only critical technical information for future bioassays, but also suggest further understanding of PD susceptibility.

  9. Effects of the antimicrobial peptide gomesin on the global gene expression profile, virulence and biofilm formation of Xylella fastidiosa.

    Science.gov (United States)

    Fogaça, Andréa C; Zaini, Paulo A; Wulff, Nelson A; da Silva, Patrícia I P; Fázio, Marcos A; Miranda, Antônio; Daffre, Sirlei; da Silva, Aline M

    2010-05-01

    In the xylem vessels of susceptible hosts, such as citrus trees, Xylella fastidiosa forms biofilm-like colonies that can block water transport, which appears to correlate to disease symptoms. Besides aiding host colonization, bacterial biofilms play an important role in resistance against antimicrobial agents, for instance antimicrobial peptides (AMPs). Here, we show that gomesin, a potent AMP from a tarantula spider, modulates X. fastidiosa gene expression profile upon 60 min of treatment with a sublethal concentration. DNA microarray hybridizations revealed that among the upregulated coding sequences, some are related to biofilm production. In addition, we show that the biofilm formed by gomesin-treated bacteria is thicker than that formed by nontreated cells or cells exposed to streptomycin. We have also observed that the treatment of X. fastidiosa with a sublethal concentration of gomesin before inoculation in tobacco plants correlates with a reduction in foliar symptoms, an effect possibly due to the trapping of bacterial cells to fewer xylem vessels, given the enhancement in biofilm production. These results warrant further investigation of how X. fastidiosa would respond to the AMPs produced by citrus endophytes and by the insect vector, leading to a better understanding of the mechanism of action of these molecules on bacterial virulence.

  10. Plasmid Vectors for Xylella fastidiosa Utilizing a Toxin-Antitoxin System for Stability in the Absence of Antibiotic Selection.

    Science.gov (United States)

    Burbank, Lindsey P; Stenger, Drake C

    2016-08-01

    The phytopathogen Xylella fastidiosa causes disease in a variety of important crop and landscape plants. Functional genetic studies have led to a broader understanding of virulence mechanisms used by this pathogen in the grapevine host. Plasmid shuttle vectors are important tools in studies of bacterial genetics but there are only a limited number of plasmid vectors available that replicate in X. fastidiosa, and even fewer that are retained without antibiotic selection. Two plasmids are described here that show stable replication in X. fastidiosa and are effective for gene complementation both in vitro and in planta. Plasmid maintenance is facilitated by incorporation of the PemI/PemK plasmid addiction system, consisting of PemK, an endoribonuclease toxin, and its cognate antitoxin, PemI. Vector pXf20pemIK utilizes a native X. fastidiosa replication origin as well as a high-copy-number pUC origin for propagation in Escherichia coli cloning strains. Broad-host-range vector pBBR5pemIK is a medium- to low-copy-number plasmid based on the pBBR1 backbone. Both plasmids are maintained for extended periods of time in the absence of antibiotic selection, as well as up to 14 weeks in grapevine, without affecting bacterial fitness. These plasmids present an alternative to traditional complementation and expression vectors which rely on antibiotic selection for plasmid retention.

  11. Grape Cultivar and Sap Culture Conditions Affect the Development of Xylella fastidiosa Phenotypes Associated with Pierce's Disease

    Science.gov (United States)

    Hoch, Harvey C.; Burr, Thomas J.; Mowery, Patricia

    2016-01-01

    Xylella fastidiosa is a xylem-limited bacterium in plant hosts and causes Pierce’s disease (PD) of grapevines, which differ in susceptibility according to the Vitis species (spp.). In this work we compared X. fastidiosa biofilm formation and population dynamics when cultured in xylem saps from PD-susceptible and -resistant Vitis spp. under different conditions. Behaviors in a closed-culture system were compared to those in different sap-renewal cultures that would more closely mimic the physicochemical environment encountered in planta. Significant differences in biofilm formation and growth in saps from PD-susceptible and -resistant spp. were only observed using sap renewal culture. Compared to saps from susceptible V. vinifera, those from PD-resistant V. aestivalis supported lower titers of X. fastidiosa and less biofilm and V. champinii suppressed both growth and biofilm formation, behaviors which are correlated with disease susceptibility. Furthermore, in microfluidic chambers X. fastidiosa formed thick mature biofilm with three-dimensional (3-D) structures, such as pillars and mounds, in saps from all susceptible spp. In contrast, only small aggregates of various shapes were formed in saps from four out of five of the resistant spp.; sap from the resistant spp. V. mustangensis was an exception in that it also supported thick lawns of biofilm but not the above described 3-D structures typically seen in a mature biofilm from the susceptible saps. Our findings provide not only critical technical information for future bioassays, but also suggest further understanding of PD susceptibility. PMID:27508296

  12. Grape Cultivar and Sap Culture Conditions Affect the Development of Xylella fastidiosa Phenotypes Associated with Pierce's Disease.

    Directory of Open Access Journals (Sweden)

    Lingyun Hao

    Full Text Available Xylella fastidiosa is a xylem-limited bacterium in plant hosts and causes Pierce's disease (PD of grapevines, which differ in susceptibility according to the Vitis species (spp.. In this work we compared X. fastidiosa biofilm formation and population dynamics when cultured in xylem saps from PD-susceptible and -resistant Vitis spp. under different conditions. Behaviors in a closed-culture system were compared to those in different sap-renewal cultures that would more closely mimic the physicochemical environment encountered in planta. Significant differences in biofilm formation and growth in saps from PD-susceptible and -resistant spp. were only observed using sap renewal culture. Compared to saps from susceptible V. vinifera, those from PD-resistant V. aestivalis supported lower titers of X. fastidiosa and less biofilm and V. champinii suppressed both growth and biofilm formation, behaviors which are correlated with disease susceptibility. Furthermore, in microfluidic chambers X. fastidiosa formed thick mature biofilm with three-dimensional (3-D structures, such as pillars and mounds, in saps from all susceptible spp. In contrast, only small aggregates of various shapes were formed in saps from four out of five of the resistant spp.; sap from the resistant spp. V. mustangensis was an exception in that it also supported thick lawns of biofilm but not the above described 3-D structures typically seen in a mature biofilm from the susceptible saps. Our findings provide not only critical technical information for future bioassays, but also suggest further understanding of PD susceptibility.

  13. Detection and characterization of pXFSL21, a novel single-copy plasmid from Xylella fastidiosa Strain Stag’s Leap

    Science.gov (United States)

    Xylella fastidiosa Strain Stag’s Leap, originally isolated from Napa Valley, California, is highly virulent in causing Pierce’s Disease (PD) of grapevine. Plasmids are extrachromosomal genetic elements associated with bacterial environmental adaptation such as virulence development. In this study, t...

  14. Preliminary findings suggest that vector feeding behaviors controlling inoculation of Xylella fastidiosa are performed less on Vitis arizonica than on V. vinifera ‘Chardonnay’

    Science.gov (United States)

    The most successful example of classical grapevine breeding for resistance to Xylella fastidiosa (Xf) is the PdR1 gene, which mediates resistance to Xf multiplication and spread in the host, once Xf has been inoculated. No effort has been made to determine whether resistance of PdR1 or its parent wi...

  15. Plant water stress effects on stylet probing behaviors of Homalodisca vitripennis (Hemiptera: Cicadellidae) associated with acquisition and inoculation of the bacterium Xylella fastidiosa

    Science.gov (United States)

    The glassy-winged sharpshooter, Homalodisca vitripennis, is a xylem fluid-ingesting leafhopper that transmits Xylella fastidiosa, the causal agent of several plant diseases in the Americas. While the role of plant water stress on the population density and dispersal of H. vitripennis has been studie...

  16. Wintercuring of Prunus dulcis cv ‘Butte,’ P. webbii and their interspecific hybrid in response to Xylella fastidiosa infections

    Science.gov (United States)

    Clonal replicates of Prunus dulcis cv ‘Butte,’ P. webbii and their interspecific hybrid P 63-61 were inoculated with Xylella fastidiosa strain M23 and evaluated for Almond Leaf Scorch Disease and subsequent wintercuring of infections during three growing seasons. Initial inoculations established gr...

  17. Host selection and adaptation are major driving forces shaping ALS Xylella fastidiosa population structure in the San Joaquin Valley of California

    Science.gov (United States)

    Xylella fastidiosa (Xf) causes disease in many commercial crops including almond leaf scorch (ALS) disease in susceptible almond (Prunus dulcis). In this study, genetic diversity and population structure of Xf associated with ALS disease were evaluated. Strains from two almond production sites in th...

  18. A zinc, copper and citric acid biocomplex shows promise for control of Xylella fastidiosa subsp. pauca in olive trees in Apulia region (southern Italy)

    Science.gov (United States)

    The bacterium Xylella fastidiosa subsp. pauca is associated with the “olive quick decline syndrome” in the Apulia region of southern Italy. To investigate control of this phytopathogen, a compound containing zinc and copper complexed with citric-acid hydracids (Dentamet®) was evaluated for in vitro ...

  19. Seasonal Abundance and Natural Inoculativity of Insect Vectors of Xylella fastidiosa in Oklahoma Tree Nurseries and Vineyards.

    Science.gov (United States)

    Overall, Lisa M; Rebek, Eric J

    2015-12-01

    Xylella fastidiosa is the causative agent of diseases of perennial plants including peach, plum, elm, oak, pecan, and grape. This bacterial pathogen is transmitted by xylem-feeding insects. In recent years, Pierce's disease of grape has been detected in 10 counties in central and northeastern Oklahoma, prompting further investigation of the disease epidemiology in this state. We surveyed vineyards and tree nurseries in Oklahoma for potential insect vectors to determine species composition, infectivity, and natural inoculativity of commonly captured insect vectors. Yellow sticky cards were used to sample insect fauna at each location. Insects were removed from sticky cards and screened for X. fastidiosa using immunocapture-PCR to determine their infectivity. A second objective was to test the natural inoculativity of insect vectors that are found in vineyards. Graphocephala versuta (Say), Graphocephala coccinea (Forster), Paraulacizes irrorata (F.), Oncometopia orbona (F.), Cuerna costalis (F.), and Entylia carinata Germar were collected from vineyards and taken back to the lab to determine their natural inoculativity. Immunocapture-PCR was used to test plant and insect samples for presence of X. fastidiosa. The three most frequently captured species from vineyards and tree nurseries were G. versuta, Clastoptera xanthocephala Germar, and O. orbona. Of those insects screened for X. fastidiosa, 2.4% tested positive for the bacterium. Field-collected G. versuta were inoculative to both ragweed and alfalfa. Following a 7-d inoculation access period, a higher percentage of alfalfa became infected than ragweed. Results from this study provide insight into the epidemiology of X. fastidiosa in Oklahoma. © The Authors 2015. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  20. Factors affecting the initial adhesion and retention of the plant pathogen Xylella fastidiosa in the foregut of an insect vector.

    Science.gov (United States)

    Killiny, Nabil; Almeida, Rodrigo P P

    2014-01-01

    Vector transmission of bacterial plant pathogens involves three steps: pathogen acquisition from an infected host, retention within the vector, and inoculation of cells into susceptible tissue of an uninfected plant. In this study, a combination of plant and artificial diet systems were used to determine the importance of several genes on the initial adhesion and retention of the bacterium Xylella fastidiosa to an efficient insect vector. Mutant strains included fimbrial (fimA and pilB) and afimbrial (hxfA and hxfB) adhesins and three loci involved in regulatory systems (rpfF, rpfC, and cgsA). Transmission assays with variable retention time indicated that HxfA and HxfB were primarily important for early adhesion to vectors, while FimA was necessary for both adhesion and retention. The long pilus protein PilB was not deficient in initial adhesion but may be important for retention. Genes upregulated under the control of rpfF are important for both initial adhesion and retention, as transmission rates of this mutant strain were initially low and decreased over time, while disruption of rpfC and cgsA yielded trends similar to that shown by the wild-type control. Because induction of an X. fastidiosa transmissible state requires pectin, a series of experiments were used to test the roles of a polygalacturonase (pglA) and the pectin and galacturonic acid carbohydrates on the transmission of X. fastidiosa. Results show that galacturonic acid, or PglA activity breaking pectin into its major subunit (galacturonic acid), is required for X. fastidiosa vector transmission using an artificial diet system. This study shows that early adhesion and retention of X. fastidiosa are mediated by different factors. It also illustrates that the interpretation of results of vector transmission experiments, in the context of vector-pathogen interaction studies, is highly dependent on experimental design.

  1. Development of single chain variable fragment (scFv) antibodies against Xylella fastidiosa subsp. pauca by phage display.

    Science.gov (United States)

    Yuan, Qing; Jordan, Ramon; Brlansky, Ronald H; Istomina, Olga; Hartung, John

    2015-10-01

    Xylella fastidiosa is a member of the gamma proteobacteria. It is fastidious, insect-vectored and xylem-limited and causes a variety of diseases, some severe, on a wide range of economically important perennial crops, including grape and citrus. Antibody based detection assays are commercially available for X. fastidiosa, and are effective at the species, but not at the subspecies level. We have made a library of scFv antibody fragments directed against X. fastidiosa subsp. pauca strain 9a5c (citrus) by using phage display technology. Antibody gene repertoires were PCR-amplified using 23 primers for the heavy chain variable region (V(H)) and 21 primers for the light chain variable region (V(L)). The V(H) and V(L) were joined by overlap extension PCR, and then the genes of the scFv library were ligated into the phage vector pKM19. The library contained 1.2×10(7) independent clones with full-length scFv inserts. In each of 3cycles of affinity-selection with 9a5c, about 1.0×10(12) phage were used for panning with 4.1×10(6), 7.1×10(6), 2.1×10(7) phage recovered after the first, second and third cycles, respectively. Sixty-six percent of clones from the final library bound X. fastidiosa 9a5c in an ELISA. Some of these scFv antibodies recognized strain 9a5c and did not recognize X. fastidiosa strains that cause Pierce's disease of grapevine. Published by Elsevier B.V.

  2. Detection of siderophores in endophytic bacteria Methylobacterium spp. associated with Xylella fastidiosa subsp. pauca Detecção de sideróforos nas bactérias endofíticas Methylobacterium spp. associadas com Xylella fastidiosa subsp. pauca

    Directory of Open Access Journals (Sweden)

    Paulo Teixeira Lacava

    2008-04-01

    Full Text Available The objective of this work was to study the production of siderophores by endophytic bacteria Methylobacterium spp., which occupy the same ecological niche as Xylella fastidiosa subsp. pauca (Xfp in citrus plants. The siderophore production of Methylobacterium strains was tested according to chromeazurol agar assay test (CAS, Csáky test (hydroxamate-type and Arnow test (catechol-type. In addition, the ability of Xfp to use siderophores, in vitro, produced by endophytic bacteria as source of iron, was evaluated. All 37 strains of Methylobacterium spp. tested were CAS-positive for siderophore production. Methylobacterium spp. produced hydroxamate-type, but not catechol-type siderophores. In vitro growth of Xfp was stimulated by the presence of supernatant siderophores of endophytic Methylobacterium mesophilicum.O objetivo deste trabalho foi estudar a produção de sideróforos pelas bactérias endofíticas Methylobacterium spp., que ocupam o mesmo nicho ecológico que Xylella fastidiosa subsp. pauca (Xfp, em plantas cítricas. A produção de sideróforos, pelas linhagens de Methylobacterium, foi testada por meio do ensaio de cromoazarol-ágar (chromeazurol agar assay-CAS, teste de Csáky (tipo hidroxamato e do teste de Arnow (tipo catecol. Além disso, a habilidade de Xfp em utilizar sideróforos produzidos por bactérias endofíticas, como fonte de ferro, in vitro, foi avaliada. Todas as 37 linhagens de Methylobacterium spp. testadas foram positivas para a produção de sideróforos, pelo teste CAS-ágar. Methylobacterium spp. foram capazes de produzir sideróforos do tipo hidroxamato, mas não do tipo catecol. O crescimento in vitro de Xfp foi estimulado pela presença de sideróforos no sobrenadante de Methylobacterium mesophilicum endofítica.

  3. Role of cyclic di-GMP in Xylella fastidiosa biofilm formation, plant virulence, and insect transmission.

    Science.gov (United States)

    Chatterjee, Subhadeep; Killiny, Nabil; Almeida, Rodrigo P P; Lindow, Steven E

    2010-10-01

    Xylella fastidiosa must coordinately regulate a variety of traits contributing to biofilm formation, host plant and vector colonization, and transmission between plants. Traits such as production of extracellular polysaccharides (EPS), adhesins, extracellular enzymes, and pili are expressed in a cell-density-dependent fashion mediated by a cell-to-cell signaling system involving a fatty acid diffusible signaling factor (DSF). The expression of gene PD0279 (which has a GGDEF domain) is downregulated in the presence of DSF and may be involved in intracellular signaling by modulating the levels of cyclic di-GMP. PD0279, designated cyclic di-GMP synthase A (cgsA), is required for biofilm formation, plant virulence, and vector transmission. cgsA mutants exhibited a hyperadhesive phenotype in vitro and overexpressed gumJ, hxfA, hxfB, xadA, and fimA, which promote attachment of cells to surfaces and, hence, biofilm formation. The mutants were greatly reduced in virulence to grape albeit still transmissible by insect vectors, although at a reduced level compared with transmission rates of the wild-type strain, despite the fact that similar numbers of cells of the cgsA mutant were acquired by the insects from infected plants. High levels of EPS were measured in cgsA mutants compared with wild-type strains, and scanning electron microscopy analysis also revealed a thicker amorphous layer surrounding the mutants. Overexpression of cgsA in a cgsA-complemented mutant conferred the opposite phenotypes in vitro. These results suggest that decreases of cyclic di-GMP result from the accumulation of DSF as cell density increases, leading to a phenotypic transition from a planktonic state capable of colonizing host plants to an adhesive state that is insect transmissible.

  4. Characterization of the Xylella fastidiosa PD1311 gene mutant and its suppression of Pierce's disease on grapevines.

    Science.gov (United States)

    Hao, Lingyun; Johnson, Kameka; Cursino, Luciana; Mowery, Patricia; Burr, Thomas J

    2017-06-01

    Xylella fastidiosa causes Pierce's disease (PD) on grapevines, leading to significant economic losses in grape and wine production. To further our understanding of X. fastidiosa virulence on grapevines, we examined the PD1311 gene, which encodes a putative acyl-coenzyme A (acyl-CoA) synthetase, and is highly conserved across Xylella species. It was determined that PD1311 is required for virulence, as the deletion mutant, ΔPD1311, was unable to cause disease on grapevines. The ΔPD1311 strain was impaired in behaviours known to be associated with PD development, including motility, aggregation and biofilm formation. ΔPD1311 also expressed enhanced sensitivity to H 2 O 2 and polymyxin B, and showed reduced survival in grapevine sap, when compared with wild-type X. fastidiosa Temecula 1 (TM1). Following inoculation, ΔPD1311 could not be detected in grape shoots, which may be related to its altered growth and sensitivity phenotypes. Inoculation with ΔPD1311 2 weeks prior to TM1 prevented the development of PD in a significant fraction of vines and eliminated detectable levels of TM1. In contrast, vines inoculated simultaneously with TM1 and ΔPD1311 developed disease at the same level as TM1 alone. In these vines, TM1 populations were distributed similarly to populations in TM1-only inoculated plants. These findings suggest that, through an indirect mechanism, pretreatment of vines with ΔPD1311 suppresses pathogen population and disease. © 2016 BSPP AND JOHN WILEY & SONS LTD.

  5. Specific Detection and Identification of American Mulberry-Infecting and Italian Olive-Associated Strains of Xylella fastidiosa by Polymerase Chain Reaction.

    Directory of Open Access Journals (Sweden)

    Wei Guan

    Full Text Available Xylella fastidiosa causes bacterial leaf scorch in many landscape trees including elm, oak, sycamore and mulberry, but methods for specific identification of a particular tree host species-limited strain or differentiation of tree-specific strains are lacking. It is also unknown whether a particular landscape tree-infecting X. fastidiosa strain is capable of infecting multiple landscape tree species in an urban environment. We developed two PCR primers specific for mulberry-infecting strains of X. fastidiosa based on the nucleotide sequence of a unique open reading frame identified only in mulberry-infecting strains among all the North and South American strains of X. fastidiosa sequenced to date. PCR using the primers allowed for detection and identification of mulberry-infecting X. fastidiosa strains in cultures and in samples collected from naturally infected mulberry trees. In addition, no mixed infections with or non-specific detections of the mulberry-infecting strains of X. fastidiosa were found in naturally X. fastidiosa-infected oak, elm and sycamore trees growing in the same region where naturally infected mulberry trees were grown. This genotype-specific PCR assay will be valuable for disease diagnosis, studies of strain-specific infections in insects and plant hosts, and management of diseases caused by X. fastidiosa. Unexpectedly but interestingly, the unique open reading frame conserved in the mulberry-infecting strains in the U. S. was also identified in the recently sequenced olive-associated strain CoDiRO isolated in Italy. When the primer set was tested against naturally infected olive plant samples collected in Italy, it allowed for detection of olive-associated strains of X. fastidiosa in Italy. This PCR assay, therefore, will also be useful for detection and identification of the Italian group of X. fastidiosa strains to aid understanding of the occurrence, evolution and biology of this new group of X. fastidiosa strains.

  6. Specific Detection and Identification of American Mulberry-Infecting and Italian Olive-Associated Strains of Xylella fastidiosa by Polymerase Chain Reaction.

    Science.gov (United States)

    Guan, Wei; Shao, Jonathan; Elbeaino, Toufic; Davis, Robert E; Zhao, Tingchang; Huang, Qi

    2015-01-01

    Xylella fastidiosa causes bacterial leaf scorch in many landscape trees including elm, oak, sycamore and mulberry, but methods for specific identification of a particular tree host species-limited strain or differentiation of tree-specific strains are lacking. It is also unknown whether a particular landscape tree-infecting X. fastidiosa strain is capable of infecting multiple landscape tree species in an urban environment. We developed two PCR primers specific for mulberry-infecting strains of X. fastidiosa based on the nucleotide sequence of a unique open reading frame identified only in mulberry-infecting strains among all the North and South American strains of X. fastidiosa sequenced to date. PCR using the primers allowed for detection and identification of mulberry-infecting X. fastidiosa strains in cultures and in samples collected from naturally infected mulberry trees. In addition, no mixed infections with or non-specific detections of the mulberry-infecting strains of X. fastidiosa were found in naturally X. fastidiosa-infected oak, elm and sycamore trees growing in the same region where naturally infected mulberry trees were grown. This genotype-specific PCR assay will be valuable for disease diagnosis, studies of strain-specific infections in insects and plant hosts, and management of diseases caused by X. fastidiosa. Unexpectedly but interestingly, the unique open reading frame conserved in the mulberry-infecting strains in the U. S. was also identified in the recently sequenced olive-associated strain CoDiRO isolated in Italy. When the primer set was tested against naturally infected olive plant samples collected in Italy, it allowed for detection of olive-associated strains of X. fastidiosa in Italy. This PCR assay, therefore, will also be useful for detection and identification of the Italian group of X. fastidiosa strains to aid understanding of the occurrence, evolution and biology of this new group of X. fastidiosa strains.

  7. Insights into the Activity and Substrate Binding of Xylella fastidiosa Polygalacturonase by Modification of a Unique QMK Amino Acid Motif Using Protein Chimeras.

    Science.gov (United States)

    Warren, Jeremy G; Lincoln, James E; Kirkpatrick, Bruce C

    2015-01-01

    Polygalacturonases (EC 3.2.1.15) catalyze the random hydrolysis of 1, 4-alpha-D-galactosiduronic linkages in pectate and other galacturonans. Xylella fastidiosa possesses a single polygalacturonase gene, pglA (PD1485), and X. fastidiosa mutants deficient in the production of polygalacturonase are non-pathogenic and show a compromised ability to systemically infect grapevines. These results suggested that grapevines expressing sufficient amounts of an inhibitor of X. fastidiosa polygalacturonase might be protected from disease. Previous work in our laboratory and others have tried without success to produce soluble active X. fastidiosa polygalacturonase for use in inhibition assays. In this study, we created two enzymatically active X. fastidiosa / A. vitis polygalacturonase chimeras, AX1A and AX2A to explore the functionality of X. fastidiosa polygalacturonase in vitro. The AX1A chimera was constructed to specifically test if recombinant chimeric protein, produced in Escherichia coli, is soluble and if the X. fastidiosa polygalacturonase catalytic amino acids are able to hydrolyze polygalacturonic acid. The AX2A chimera was constructed to evaluate the ability of a unique QMK motif of X. fastidiosa polygalacturonase, most polygalacturonases have a R(I/L)K motif, to bind to and allow the hydrolysis of polygalacturonic acid. Furthermore, the AX2A chimera was also used to explore what effect modification of the QMK motif of X. fastidiosa polygalacturonase to a conserved RIK motif has on enzymatic activity. These experiments showed that both the AX1A and AX2A polygalacturonase chimeras were soluble and able to hydrolyze the polygalacturonic acid substrate. Additionally, the modification of the QMK motif to the conserved RIK motif eliminated hydrolytic activity, suggesting that the QMK motif is important for the activity of X. fastidiosa polygalacturonase. This result suggests X. fastidiosa polygalacturonase may preferentially hydrolyze a different pectic substrate or

  8. Surface physicochemical properties at the micro and nano length scales: role on bacterial adhesion and Xylella fastidiosa biofilm development.

    Science.gov (United States)

    Lorite, Gabriela S; Janissen, Richard; Clerici, João H; Rodrigues, Carolina M; Tomaz, Juarez P; Mizaikoff, Boris; Kranz, Christine; de Souza, Alessandra A; Cotta, Mônica A

    2013-01-01

    The phytopathogen Xylella fastidiosa grows as a biofilm causing vascular occlusion and consequently nutrient and water stress in different plant hosts by adhesion on xylem vessel surfaces composed of cellulose, hemicellulose, pectin and proteins. Understanding the factors which influence bacterial adhesion and biofilm development is a key issue in identifying mechanisms for preventing biofilm formation in infected plants. In this study, we show that X. fastidiosa biofilm development and architecture correlate well with physicochemical surface properties after interaction with the culture medium. Different biotic and abiotic substrates such as silicon (Si) and derivatized cellulose films were studied. Both biofilms and substrates were characterized at the micro- and nanoscale, which corresponds to the actual bacterial cell and membrane/ protein length scales, respectively. Our experimental results clearly indicate that the presence of surfaces with different chemical composition affect X. fastidiosa behavior from the point of view of gene expression and adhesion functionality. Bacterial adhesion is facilitated on more hydrophilic surfaces with higher surface potentials; XadA1 adhesin reveals different strengths of interaction on these surfaces. Nonetheless, despite different architectural biofilm geometries and rates of development, the colonization process occurs on all investigated surfaces. Our results univocally support the hypothesis that different adhesion mechanisms are active along the biofilm life cycle representing an adaptation mechanism for variations on the specific xylem vessel composition, which the bacterium encounters within the infected plant.

  9. Calcium transcriptionally regulates the biofilm machinery of Xylella fastidiosa to promote continued biofilm development in batch cultures.

    Science.gov (United States)

    Parker, Jennifer K; Chen, Hongyu; McCarty, Sara E; Liu, Lawrence Y; De La Fuente, Leonardo

    2016-05-01

    The functions of calcium (Ca) in bacteria are less characterized than in eukaryotes, where its role has been studied extensively. The plant-pathogenic bacterium Xylella fastidiosa has several virulence features that are enhanced by increased Ca concentrations, including biofilm formation. However, the specific mechanisms driving modulation of this feature are unclear. Characterization of biofilm formation over time showed that 4 mM Ca supplementation produced denser biofilms that were still developing at 96 h, while biofilm in non-supplemented media had reached the dispersal stage by 72 h. To identify changes in global gene expression in X. fastidiosa grown in supplemental Ca, RNA-Seq of batch culture biofilm cells was conducted at three 24-h time intervals. Results indicate that a variety of genes are differentially expressed in response to Ca, including genes related to attachment, motility, exopolysaccharide synthesis, biofilm formation, peptidoglycan synthesis, regulatory functions, iron homeostasis, and phages. Collectively, results demonstrate that Ca supplementation induces a transcriptional response that promotes continued biofilm development, while biofilm cells in nonsupplemented media are driven towards dispersion of cells from the biofilm structure. These results have important implications for disease progression in planta, where xylem sap is the source of Ca and other nutrients for X. fastidiosa. © 2016 Society for Applied Microbiology and John Wiley & Sons Ltd.

  10. Zinc Detoxification Is Required for Full Virulence and Modification of the Host Leaf Ionome by Xylella fastidiosa.

    Science.gov (United States)

    Navarrete, Fernando; De La Fuente, Leonardo

    2015-04-01

    Zinc (Zn) is an essential element for all forms of life because it is a structural or catalytic cofactor of many proteins, but it can have toxic effects at high concentrations; thus, microorganisms must tightly regulate its levels. Here, we evaluated the role of Zn homeostasis proteins in the virulence of the xylem-limited bacterium Xylella fastidiosa, causal agent of Pierce's disease of grapevine, among other diseases. Two mutants of X. fastidiosa 'Temecula' affected in genes which regulate Zn homeostasis (zur) and Zn detoxification (czcD) were constructed. Both knockouts showed increased sensitivity to Zn at physiologically relevant concentrations and increased intracellular accumulation of this metal compared with the wild type. Increased Zn sensitivity was correlated with decreased growth in grapevine xylem sap, reduced twitching motility, and downregulation of exopolysaccharide biosynthetic genes. Tobacco plants inoculated with either knockout mutant showed reduced foliar symptoms and a much reduced (czcD) or absent (zur) modification of the leaf ionome (i.e., the mineral nutrient and trace element composition), as well as reduced bacterial populations. The results show that detoxification of Zn is crucial for the virulence of X. fastidiosa and verifies our previous findings that modification of the host leaf ionome correlates with bacterial virulence.

  11. Genomics and X-ray microanalysis indicate that Ca2+ and thiols mediate the aggregation and adhesion of Xylella fastidiosa

    Directory of Open Access Journals (Sweden)

    Leite B.

    2002-01-01

    Full Text Available The availability of the genome sequence of the bacterial plant pathogen Xylella fastidiosa, the causal agent of citrus variegated chlorosis, is accelerating important investigations concerning its pathogenicity. Plant vessel occlusion is critical for symptom development. The objective of the present study was to search for information that would help to explain the adhesion of X. fastidiosa cells to the xylem. Scanning electron microscopy revealed that adhesion may occur without the fastidium gum, an exopolysaccharide produced by X. fastidiosa, and X-ray microanalysis demonstrated the presence of elemental sulfur both in cells grown in vitro and in cells found inside plant vessels, indicating that the sulfur signal is generated by the pathogen surface. Calcium and magnesium peaks were detected in association with sulfur in occluded vessels. We propose an explanation for the adhesion and aggregation process. Thiol groups, maintained by the enzyme peptide methionine sulfoxide reductase, could be active on the surface of the bacteria and appear to promote cell-cell aggregation by forming disulfide bonds with thiol groups on the surface of adjacent cells. The enzyme methionine sulfoxide reductase has been shown to be an auxiliary component in the adhesiveness of some human pathogens. The negative charge conferred by the ionized thiol group could of itself constitute a mechanism of adhesion by allowing the formation of divalent cation bridges between the negatively charged bacteria and predominantly negatively charged xylem walls.

  12. Citrus Functional Genomics and Molecular Modeling in Relation to Citrus sinensis (Sweet Orange) Infection with Xylella fastidiosa (Citrus Variegated Chlorosis).

    Science.gov (United States)

    Dwivedi, Upendra N; Tiwari, Sameeksha; Prasanna, Pragya; Awasthi, Manika; Singh, Swati; Pandey, Veda P

    2016-08-01

    Citrus are among the economically most important fruit tree crops in the world. Citrus variegated chlorosis (CVC), caused by Xylella fastidiosa infection, is a serious disease limiting citrus production at a global scale. With availability of citrus genomic resources, it is now possible to compare citrus expressed sequence tag (EST) data sets and identify single-nucleotide polymorphisms (SNPs) within and among different citrus cultivars that can be exploited for citrus resistance to infections, citrus breeding, among others. We report here, for the first time, SNPs in the EST data sets of X. fastidiosa-infected Citrus sinensis (sweet orange) and their functional annotation that revealed the involvement of eight C. sinensis candidate genes in CVC pathogenesis. Among these genes were xyloglucan endotransglycosylase, myo-inositol-1-phosphate synthase, and peroxidase were found to be involved in plant cell wall metabolism. These have been further investigated by molecular modeling for their role in CVC infection and defense. Molecular docking analyses of the wild and the mutant (SNP containing) types of the selected three enzymes with their respective substrates revealed a significant decrease in the binding affinity of substrates for the mutant enzymes, thus suggesting a decrease in the catalytic efficiency of these enzymes during infection, thereby facilitating a favorable condition for infection by the pathogen. These findings offer novel agrigenomics insights in developing future molecular targets and strategies for citrus fruit cultivation in ways that are resistant to X. fastidiosa infection, and by extension, with greater harvesting efficiency and economic value.

  13. Insights into enzyme-substrate interaction and characterization of catalytic al intermediates of a Xylella Fastidiosa antioxidant protein

    International Nuclear Information System (INIS)

    Oliveira, Marcos Antonio de; Cussiol, Jose Renato Rosa; Soares Netto, Luis Eduardo; Guimaraes, Beatriz Gomes; Medrano, Francisco Javier; Gozzo, Fabio Cesar

    2005-01-01

    Plants and animals have developed various strategies to defend themselves from pathogens. One of them is the generation of oxidants such as organic Hydroperoxides (OHP). OHP can be generated through free radicals as well as enzymatic oxidation of unsaturated fatty acids. To counteract this oxidative stress, bacteria have evolved several antioxidant mechanisms. Organic hydroperoxide resistance protein (Ohr) was initially identified as a factor involved in the resistance of bacteria, most of them pathogenic, to OHP, but not H 2 O 2 . We have cloned Ohr gene from Xylella fastidiosa and expressed in in Escherichia coli. The biochemical role of Ohr remained unknown for a long time until this work and the work of Nikolov's group (Cornell University, New York) independently showed that these proteins are thiol dependent peroxidases, whose activity is generated by a reactive cysteine. (author)

  14. Insights into enzyme-substrate interaction and characterization of catalytic al intermediates of a Xylella Fastidiosa antioxidant protein

    Energy Technology Data Exchange (ETDEWEB)

    Oliveira, Marcos Antonio de; Cussiol, Jose Renato Rosa; Soares Netto, Luis Eduardo [Sao Paulo Univ. (USP), SP (Brazil). Inst. de Biociencias. Dept. de Genetica e Biologia Evolutiva; Guimaraes, Beatriz Gomes; Medrano, Francisco Javier; Gozzo, Fabio Cesar [Laboratorio Nacional de Luz Sincrotron (LNLS), Campinas, SP (Brazil). Centro de Biologia Molecular Estrutural

    2005-07-01

    Plants and animals have developed various strategies to defend themselves from pathogens. One of them is the generation of oxidants such as organic Hydroperoxides (OHP). OHP can be generated through free radicals as well as enzymatic oxidation of unsaturated fatty acids. To counteract this oxidative stress, bacteria have evolved several antioxidant mechanisms. Organic hydroperoxide resistance protein (Ohr) was initially identified as a factor involved in the resistance of bacteria, most of them pathogenic, to OHP, but not H{sub 2}O{sub 2}. We have cloned Ohr gene from Xylella fastidiosa and expressed in in Escherichia coli. The biochemical role of Ohr remained unknown for a long time until this work and the work of Nikolov's group (Cornell University, New York) independently showed that these proteins are thiol dependent peroxidases, whose activity is generated by a reactive cysteine. (author)

  15. Diffusible signal factor (DSF) synthase RpfF of Xylella fastidiosa is a multifunction protein also required for response to DSF.

    Science.gov (United States)

    Ionescu, Michael; Baccari, Clelia; Da Silva, Aline Maria; Garcia, Angelica; Yokota, Kenji; Lindow, Steven E

    2013-12-01

    Xylella fastidiosa, like related Xanthomonas species, employs an Rpf cell-cell communication system consisting of a diffusible signal factor (DSF) synthase, RpfF, and a DSF sensor, RpfC, to coordinate expression of virulence genes. While phenotypes of a ΔrpfF strain in Xanthomonas campestris could be complemented by its own DSF, the DSF produced by X. fastidiosa (XfDSF) did not restore expression of the XfDSF-dependent genes hxfA and hxfB to a ΔrpfF strain of X. fastidiosa, suggesting that RpfF is involved in XfDSF sensing or XfDSF-dependent signaling. To test this conjecture, rpfC and rpfF of X. campestris were replaced by those of X. fastidiosa, and the contribution of each gene to the induction of a X. campestris DSF-dependent gene was assessed. As in X. fastidiosa, XfDSF-dependent signaling required both X. fastidiosa proteins RpfF and RpfC. RpfF repressed RpfC signaling activity, which in turn was derepressed by XfDSF. A mutated X. fastidiosa RpfF protein with two substitutions of glutamate to alanine in its active site was incapable of XfDSF production yet enabled a response to XfDSF, indicating that XfDSF production and the response to XfDSF are two separate functions in which RpfF is involved. This mutant was also hypervirulent to grape, demonstrating the antivirulence effects of XfDSF itself in X. fastidiosa. The Rpf system of X. fastidiosa is thus a novel example of a quorum-sensing signal synthase that is also involved in the response to the signal molecule that it synthesizes.

  16. Crystallization and preliminary X-ray diffraction analysis of a glutathione S-transferase from Xylella fastidiosa

    International Nuclear Information System (INIS)

    Garcia, Wanius; Travensolo, Regiane F.; Rodrigues, Nathalia C.; Muniz, João R. C.; Caruso, Célia S.; Lemos, Eliana G. M.; Araujo, Ana Paula U.; Carrilho, Emanuel

    2008-01-01

    Glutathione S-transferase from X. fastidiosa (xfGST) has been overexpressed in E. coli, purified and crystallized. Diffraction data were collected to 2.23 Å. Glutathione S-transferases (GSTs) form a group of multifunctional isoenzymes that catalyze the glutathione-dependent conjugation and reduction reactions involved in the cellular detoxification of xenobiotic and endobiotic compounds. GST from Xylella fastidiosa (xfGST) was overexpressed in Escherichia coli and purified by conventional affinity chromatography. In this study, the crystallization and preliminary X-ray analysis of xfGST is described. The purified protein was crystallized by the vapour-diffusion method, producing crystals that belonged to the triclinic space group P1. The unit-cell parameters were a = 47.73, b = 87.73, c = 90.74 Å, α = 63.45, β = 80.66, γ = 94.55°. xfGST crystals diffracted to 2.23 Å resolution on a rotating-anode X-ray source

  17. Cigarrinhas dos Citros, Vetoras da Bactéria Xylella fastidiosa Wells et al.: Pragas Potenciais para a Citricultura Sergipana

    Directory of Open Access Journals (Sweden)

    Ruberval Azevedo

    2015-04-01

    Abstract. The citrus industry in Brazil plays a role of great economic, social, generating jobs, income and development. Brazil is the largest producer of citrus, the State of Sergipe stands out in 5th place in national production. Among the many pest problems faced by Brazilian citrus is Citrus Variegated Chlorosis (CVC, known as the yellowing caused by the bacterium Xylella fastidiosa Wells et al. The CVC was officially identified in Brazil in 1987, in orchards of “Triângulo Mineiro” and North and northwest of the state of São Paulo. In the Northeast Region of Brazil, was found in 1996 in the municipality of Boquim Sergipe, and Bahia in 1997, the municipalities of Rio Real and Itapicuru. The aim was to review the literature on the species of leafhoppers vectors of CVC, and verify that occur in the state of Sergipe. The first symptoms are seen in the leaves, then go for the fruits and end up affecting the entire plant, and to be perceived can take between five months and two years. The main vectors of X. fastidiosa in citrus are the sharpshooters of the family Cicadellidae. In Brazil 12 sharpshooters species have already been confirmed. For the state of Sergipe, is scarce information about the Cicadellidae vectors, the data are limited to the northern coast of Bahia, except for vague quote about four genus (Oncometropia, Acrogonia, Dilobopterus and Homolodisca and three species (Homolodisca ignorata Melichar, Acrogonia sp. and Homolodisca spottii Takiya, Cavichioli & McKamey.

  18. Crystallization and preliminary X-ray diffraction analysis of a glutathione S-transferase from Xylella fastidiosa

    Energy Technology Data Exchange (ETDEWEB)

    Garcia, Wanius, E-mail: wanius@if.sc.usp.br [Laboratório de Biofísica Molecular ‘Sérgio Mascarenhas’, Instituto de Física de São Carlos, Universidade de São Paulo (USP), São Carlos (Brazil); Travensolo, Regiane F. [Grupo de Bioanalítica, Microfabricação e Separações, Instituto de Química de São Carlos, Universidade de São Paulo (USP), São Carlos (Brazil); Rodrigues, Nathalia C.; Muniz, João R. C. [Laboratório de Biofísica Molecular ‘Sérgio Mascarenhas’, Instituto de Física de São Carlos, Universidade de São Paulo (USP), São Carlos (Brazil); Caruso, Célia S. [Grupo de Bioanalítica, Microfabricação e Separações, Instituto de Química de São Carlos, Universidade de São Paulo (USP), São Carlos (Brazil); Lemos, Eliana G. M. [Laboratório de Bioquímica de Microrganismos e de Plantas, Departamento de Tecnologia, UNESP, Jaboticabal (Brazil); Araujo, Ana Paula U. [Laboratório de Biofísica Molecular ‘Sérgio Mascarenhas’, Instituto de Física de São Carlos, Universidade de São Paulo (USP), São Carlos (Brazil); Carrilho, Emanuel, E-mail: wanius@if.sc.usp.br [Grupo de Bioanalítica, Microfabricação e Separações, Instituto de Química de São Carlos, Universidade de São Paulo (USP), São Carlos (Brazil); Laboratório de Biofísica Molecular ‘Sérgio Mascarenhas’, Instituto de Física de São Carlos, Universidade de São Paulo (USP), São Carlos (Brazil)

    2008-02-01

    Glutathione S-transferase from X. fastidiosa (xfGST) has been overexpressed in E. coli, purified and crystallized. Diffraction data were collected to 2.23 Å. Glutathione S-transferases (GSTs) form a group of multifunctional isoenzymes that catalyze the glutathione-dependent conjugation and reduction reactions involved in the cellular detoxification of xenobiotic and endobiotic compounds. GST from Xylella fastidiosa (xfGST) was overexpressed in Escherichia coli and purified by conventional affinity chromatography. In this study, the crystallization and preliminary X-ray analysis of xfGST is described. The purified protein was crystallized by the vapour-diffusion method, producing crystals that belonged to the triclinic space group P1. The unit-cell parameters were a = 47.73, b = 87.73, c = 90.74 Å, α = 63.45, β = 80.66, γ = 94.55°. xfGST crystals diffracted to 2.23 Å resolution on a rotating-anode X-ray source.

  19. Spatial Genetic Structure of Coffee-Associated Xylella fastidiosa Populations Indicates that Cross Infection Does Not Occur with Sympatric Citrus Orchards.

    Science.gov (United States)

    Francisco, Carolina S; Ceresini, Paulo C; Almeida, Rodrigo P P; Coletta-Filho, Helvécio D

    2017-04-01

    Xylella fastidiosa, an economically important plant-pathogenic bacterium, infects both coffee and citrus trees in Brazil. Although X. fastidiosa in citrus is well studied, knowledge about the population structure of this bacterium infecting coffee remains unknown. Here, we studied the population structure of X. fastidiosa infecting coffee trees in São Paulo State, Brazil, in four regions where citrus is also widely cultivated. Genotyping of over 500 isolates from coffee plants using 14 genomic microsatellite markers indicated that populations were largely geographically isolated, as previously found with populations of X. fastidiosa infecting citrus. These results were supported by a clustering analysis, which indicated three major genetic groups among the four sampled regions. Overall, approximately 38% of isolates showed significant membership coefficients not related to their original geographical populations (i.e., migrants), characterizing a significant degree of genotype flow among populations. To determine whether admixture occurred between isolates infecting citrus and coffee plants, one site with citrus and coffee orchards adjacent to each other was selected; over 100 isolates were typed from each host plant. No signal of natural admixture between citrus- and coffee-infecting isolates was found; artificial cross-infection assays with representative isolates also yielded no successful cross infection. A comparison determined that X. fastidiosa populations from coffee have higher genetic diversity and allelic richness compared with citrus. The results showed that coffee and citrus X. fastidiosa populations are effectively isolated from each other and, although coffee populations are spatially structured, migration has an important role in shaping diversity.

  20. Characterization of the LysR-type transcriptional regulator YcjZ-like from Xylella fastidiosa overexpressed in Escherichia coli.

    Science.gov (United States)

    Santiago, André S; Santos, Clelton A; Mendes, Juliano S; Toledo, Marcelo A S; Beloti, Lilian L; Souza, Alessandra A; Souza, Anete P

    2015-09-01

    The Xylella fastidiosa 9a5c strain is a xylem-limited phytopathogen that is the causal agent of citrus variegated chlorosis (CVC). This bacterium is able to form a biofilm and occlude the xylem vessels of susceptible plants, which leads to significant agricultural and economic losses. Biofilms are associated with bacterial pathogenicity because they are very resistant to antibiotics and other metal-based chemicals that are used in agriculture. The X. fastidiosa YcjZ-like (XfYcjZ-like) protein belongs to the LysR-type transcriptional regulator (LTTR) family and is involved in various cellular functions that range from quorum sensing to bacterial survival. In the present study, we report the cloning, expression and purification of XfYcjZ-like, which was overexpressed in Escherichia coli. The secondary folding of the recombinant and purified protein was assessed by circular dichroism, which revealed that XfYcjZ-like contains a typical α/β fold. An initial hydrodynamic characterization showed that XfYcjZ-like is a globular tetramer in solution. In addition, using a polyclonal antibody against XfYcjZ-like, we assessed the expression profile of this protein during the different developmental phases of X. fastidiosa in in vitro cultivated biofilm cells and demonstrated that XfYcjZ-like is upregulated in planktonic cells in response to a copper shock treatment. Finally, the ability of XfYcjZ-like to interact with its own predicted promoter was confirmed in vitro, which is a typical feature of LysR. Taken together, our findings indicated that the XfYcjZ-like protein is involved in both the organization of the architecture and the maturation of the bacterial biofilm and that it is responsive to oxidative stress. Copyright © 2015 Elsevier Inc. All rights reserved.

  1. Isolation of Xylella fastidiosa from Citrus sinensis (L) Osb. And Vitis vinifera and study of genetic diversity in Costa Rica

    International Nuclear Information System (INIS)

    Aguilar Alvarez, Estela Yamileth

    2007-01-01

    Xylella fastidiosa is a plant pathogenic bacterium that causes diseases in different crops. Symptoms similar to those caused by citrus variegated chlorosis (CVC) were observed in sweet orange trees which served as shade and fences in coffee plantations in Costa Rica, in 2002. A total of 35 citrus trees and 24 vines from eight different districts and 3 respectively were evaluated by 'double antibody sandwich enzyme-linked immunosorbent assay' (DASELISA), resulting in 21 citrus and 19 positive vid. From four citrus trees and six of vines, were obtained six isolates and seven isolates respectively in solid medium, whose morphological and biochemical characteristics coincided with those reported in the literature as characteristic of X. fastidiosa. The identity of the isolates is confirmed by the chain polymerase reaction (PCR) using primers 272-1/272-2int and RST31/RST33. Three isolates from Grecia (Alajuela Province) amplified a band of 500pb using specific primers 272-2int/CVC-1 for strains of X. fastidiosa that cause CVC. The genetic variability of isolates from each other in comparison with isolates of coffee in Costa Rica, U.S. grapes and citrus in Brazil have been studied using techniques of random amplification polymorphism DNA (RAPD) and length polymorphisms of restriction fragments (RFLPs) of the products obtained with primers int/272-2int JB-1/JB-2 and 272-1. The results showed a clear separation between citrus isolates of Costa Rica; and, an association of three of them with the strains of citrus in Brasil. Also, an association between strains of coffee of Costa Rica with grape vines in the U.S. An association of molecular analysis confirmed the data variance. (author) [es

  2. Salivary enzymes are injected into xylem by the glassy-winged sharpshooter, a vector of Xylella fastidiosa.

    Science.gov (United States)

    Backus, Elaine A; Andrews, Kim B; Shugart, Holly J; Carl Greve, L; Labavitch, John M; Alhaddad, Hasan

    2012-07-01

    A few phytophagous hemipteran species such as the glassy-winged sharpshooter, Homalodisca vitripennis, (Germar), subsist entirely on xylem fluid. Although poorly understood, aspects of the insect's salivary physiology may facilitate both xylem-feeding and transmission of plant pathogens. Xylella fastidiosa is a xylem-limited bacterium that causes Pierce's disease of grape and other scorch diseases in many important crops. X. fastidiosa colonizes the anterior foregut (precibarium and cibarium) of H. vitripennis and other xylem-feeding vectors. Bacteria form a dense biofilm anchored in part by an exopolysaccharide (EPS) matrix that is reported to have a β-1,4-glucan backbone. Recently published evidence supports the following, salivation-egestion hypothesis for the inoculation of X. fastidiosa during vector feeding. The insect secretes saliva into the plant and then rapidly takes up a mixture of saliva and plant constituents. During turbulent fluid movements in the precibarium, the bacteria may become mechanically and enzymatically dislodged; the mixture is then egested back out through the stylets into plant cells, possibly including xylem vessels. The present study found that proteins extracted from dissected H. vitripennis salivary glands contain several enzyme activities capable of hydrolyzing glycosidic linkages in polysaccharides such as those found in EPS and plant cell walls, based on current information about the structures of those polysaccharides. One of these enzymes, a β-1,4-endoglucanase (EGase) was enriched in the salivary gland protein extract by subjecting the extract to a few, simple purification steps. The EGase-enriched extract was then used to generate a polyclonal antiserum that was used for immunohistochemical imaging of enzymes in sharpshooter salivary sheaths in grape. Results showed that enzyme-containing gelling saliva is injected into xylem vessels during sharpshooter feeding, in one case being carried by the transpiration stream away

  3. On-site detection of Xylella fastidiosa in host plants and in “spy insects” using the real-time loop-mediated isothermal amplification method

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    Thaer YASEEN

    2015-12-01

    Full Text Available A recent severe outbreak of Xylella fastidiosa associated with ‘olive quick decline syndrome’ (OQDS was reported in Apulia (Southern Italy. In this study an on-site real-time loop-mediated isothermal amplification (real-time LAMP was developed for detecting X. fastidiosa in host plants and insects. A marked simplification of the DNA extraction procedure was obtained by heating the samples in a portable Smart-Dart device and using an optimized enhancer reaction buffer. The connection to a tablet or Smartphone allowed to visualize the results of the reaction in real time. Compared to PCR and ELISA, with which it showed comparable results in terms of sensitivity and reliability in the X. fastidiosa detection, this simplified real-time LAMP procedure proved to be “user friendly”, displaying the advantages to be an on-site detection method of easy handling, rapid execution and low cost.

  4. Crystallization and preliminary X-ray analysis of stationary phase survival protein E (SurE) from Xylella fastidiosa in two crystal forms

    International Nuclear Information System (INIS)

    Reis, Marcelo Augusto dos; Saraiva, Antonio Marcos; Santos, Marcelo Leite dos; Souza, Anete Pereira de; Aparicio, Ricardo

    2012-01-01

    The crystallization and preliminary analysis of two crystal forms of survival protein E from X. fastidiosa are reported. The bacterium Xylella fastidiosa is a phytopathogenic organism that causes citrus variegated chlorosis, a disease which attacks economically important crops, mainly oranges. In this communication, the crystallization and preliminary X-ray crystallographic analysis of XfSurE, a survival protein E from X. fastidiosa, are reported. Data were collected for two crystal forms, I and II, to 1.93 and 2.9 Å resolution, respectively. Crystal form I belonged to space group C2, with unit-cell parameters a = 172.36, b = 84.18, c = 87.24 Å, α = γ = 90, β = 96.59°, whereas crystal form II belonged to space group C2, with unit-cell parameters a = 88.05, b = 81.26, c = 72.84 Å, α = γ = 90, β = 94.76°

  5. Large-scale intersubspecific recombination in the plant-pathogenic bacterium Xylella fastidiosa is associated with the host shift to mulberry.

    Science.gov (United States)

    Nunney, Leonard; Schuenzel, Erin L; Scally, Mark; Bromley, Robin E; Stouthamer, Richard

    2014-05-01

    Homologous recombination plays an important role in the structuring of genetic variation of many bacteria; however, its importance in adaptive evolution is not well established. We investigated the association of intersubspecific homologous recombination (IHR) with the shift to a novel host (mulberry) by the plant-pathogenic bacterium Xylella fastidiosa. Mulberry leaf scorch was identified about 25 years ago in native red mulberry in the eastern United States and has spread to introduced white mulberry in California. Comparing a sequence of 8 genes (4,706 bp) from 21 mulberry-type isolates to published data (352 isolates representing all subspecies), we confirmed previous indications that the mulberry isolates define a group distinct from the 4 subspecies, and we propose naming the taxon X. fastidiosa subsp. morus. The ancestry of its gene sequences was mixed, with 4 derived from X. fastidiosa subsp. fastidiosa (introduced from Central America), 3 from X. fastidiosa subsp. multiplex (considered native to the United States), and 1 chimeric, demonstrating that this group originated by large-scale IHR. The very low within-type genetic variation (0.08% site polymorphism), plus the apparent inability of native X. fastidiosa subsp. multiplex to infect mulberry, suggests that this host shift was achieved after strong selection acted on genetic variants created by IHR. Sequence data indicate that a single ancestral IHR event gave rise not only to X. fastidiosa subsp. morus but also to the X. fastidiosa subsp. multiplex recombinant group which infects several hosts but is the only type naturally infecting blueberry, thus implicating this IHR in the invasion of at least two novel native hosts, mulberry and blueberry.

  6. Attempts to develop sustainable biocontrol strategies of Xylella fastidiosa infections in olive

    OpenAIRE

    D'Attoma G., Morelli M., Cicco S., Saponari M. and Saldarelli P.

    2017-01-01

    X. fastidiosa is a plant pathogenic bacterium, which is causing the ‘Olive Quick Decline Syndrome’ (OQDS), on olive trees in the southern part of Apulia region. The knowledge of mechanisms regulating olive- X. fastidiosa interactions is fundamental to develop biocontrol strategies. In Pierce’s Disease (PD) the pathogen virulence relies on a fine balance between motile cells, which move and proliferate in xylem vessels, and sticky cells forming a biofilm and responsible for vessels...

  7. Survey of potential sharpshooter and spittlebug vectors of Xylella fastidiosa to grapevines at the São Francisco River Valley, Brazil

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    Rudiney Ringenberg

    2014-06-01

    Full Text Available Survey of potential sharpshooter and spittlebug vectors of Xylella fastidiosa to grapevines at the São Francisco River Valley, Brazil. Pierce's disease of grapevines, caused by Xylella fastidiosa, is a serious problem in some regions of North America, not yet reported in Brazil. In this study, a survey of potential sharpshooter (Hemiptera, Cicadellidae, Cicadellinae and spittlebug (Hemiptera, Cercopidae vectors of X. fastidiosa was conducted in vineyards at the São Francisco River Valley, a major grape growing region in Brazil. Four vineyards of Vitis vinifera L. were sampled fortnightly from June/2005 to June/2007, using yellow sticky cards, each placed at two different heights (45 cm aboveground and 45 cm above the crop canopy in 10 sampling localities. A total of 4,095 specimens of sharpshooters were collected, nearly all from 3 Proconiini species, Homalodisca spottii Takiya, Cavichioli & McKamey, 2006 (96.8% of the specimens, Tapajosa fulvopunctata (Signoret, 1854 (3.1%, and Tretogonia cribrata Melichar, 1926 (1 specimen. Hortensia similis (Walker, 1851 (2 specimens was the only Cicadellini species. Only 1 cercopid specimen, belonging to Aeneolamia colon (Germar, 1821, was trapped. Even though they are not considered potential Xylella vectors, 2 Gyponini leafhoppers were collected: Curtara samera DeLong & Freytag, 1972 (11 specimens and Curtara inflata DeLong & Freytag, 1976 (1 specimen. Homalodisca spottii was observed feeding and mating on green branches of grapevines, in addition to egg masses. Because of its prevalence on the crop canopy, occurrence throughout the year (with peaks from February to August, and ability to colonize grapevines, H. spottii could be an important vector if a X. fastidiosa strain pathogenic to grapevines becomes introduced at the São Francisco River Valley.

  8. Differential gene expression in Xylella fastidiosa 9a5c during co-cultivation with the endophytic bacterium Methylobacterium mesophilicum SR1.6/6.

    Science.gov (United States)

    Dourado, Manuella Nóbrega; Santos, Daiene Souza; Nunes, Luiz Roberto; Costa de Oliveira, Regina Lúcia Batista da; de Oliveira, Marcus Vinicius; Araújo, Welington Luiz

    2015-12-01

    Xylella fastidiosa, the causal agent of citrus variegated chlorosis (CVC), colonizes plant xylem, reducing sap flow, and inducing internerval chlorosis, leaf size reduction, necrosis, and harder and smaller fruits. This bacterium may be transmitted from plant to plant by sharpshooter insects, including Bucephalogonia xanthopis. The citrus endophytic bacterium Methylobacterium mesophilicum SR1.6/6 colonizes citrus xylem and previous studies showed that this strain is also transferred from plant to plant by B. xanthopis (Insecta), suggesting that this endophytic bacterium may interact with X. fastidiosa in planta and inside the insect vector during co-transmission by the same insect vector. To better understand the X. fastidiosa behavior in the presence of M. mesophilicum, we evaluated the X. fastidiosa transcriptional profile during in vitro interaction with M. mesophilicum SR1.6/6. The results showed that during co-cultivation, X. fastidiosa down-regulated genes related to growth and up-regulated genes related to energy production, stress, transport, and motility, suggesting the existence of a specific adaptive response to the presence of M. mesophilicum in the culture medium. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  9. Functional and structural studies of the disulfide isomerase DsbC from the plant pathogen Xylella fastidiosa reveals a redox-dependent oligomeric modulation in vitro.

    Science.gov (United States)

    Santos, Clelton A; Toledo, Marcelo A S; Trivella, Daniela B B; Beloti, Lilian L; Schneider, Dilaine R S; Saraiva, Antonio M; Crucello, Aline; Azzoni, Adriano R; Souza, Alessandra A; Aparicio, Ricardo; Souza, Anete P

    2012-10-01

    Xylella fastidiosa is a Gram-negative bacterium that grows as a biofilm inside the xylem vessels of susceptible plants and causes several economically relevant crop diseases. In the present study, we report the functional and low-resolution structural characterization of the X. fastidiosa disulfide isomerase DsbC (XfDsbC). DsbC is part of the disulfide bond reduction/isomerization pathway in the bacterial periplasm and plays an important role in oxidative protein folding. In the present study, we demonstrate the presence of XfDsbC during different stages of X. fastidiosa biofilm development. XfDsbC was not detected during X. fastidiosa planktonic growth; however, after administering a sublethal copper shock, we observed an overexpression of XfDsbC that also occurred during planktonic growth. These results suggest that X. fastidiosa can use XfDsbC in vivo under oxidative stress conditions similar to those induced by copper. In addition, using dynamic light scattering and small-angle X-ray scattering, we observed that the oligomeric state of XfDsbC in vitro may be dependent on the redox environment. Under reducing conditions, XfDsbC is present as a dimer, whereas a putative tetrameric form was observed under nonreducing conditions. Taken together, our findings demonstrate the overexpression of XfDsbC during biofilm formation and provide the first structural model of a bacterial disulfide isomerase in solution. © 2012 The Authors Journal compilation © 2012 FEBS.

  10. Functional and Evolutionary Characterization of a UDP-Xylose Synthase Gene from the Plant Pathogen Xylella fastidiosa, Involved in the Synthesis of Bacterial Lipopolysaccharide.

    Science.gov (United States)

    Alencar, Valquíria Campos; Jabes, Daniela Leite; Menegidio, Fabiano Bezerra; Sassaki, Guilherme Lanzi; de Souza, Lucas Rodrigo; Puzer, Luciano; Meneghetti, Maria Cecília Zorél; Lima, Marcelo Andrade; Tersariol, Ivarne Luis Dos Santos; de Oliveira, Regina Costa; Nunes, Luiz R

    2017-02-07

    Xylella fastidiosa is a plant-infecting bacillus, responsible for many important crop diseases, such as Pierce's disease of vineyards, citrus variegated chlorosis, and coffee leaf scorch (CLS), among others. Recent genomic comparisons involving two CLS-related strains, belonging to X. fastidiosa subsp. pauca, revealed that one of them carries a frameshift mutation that inactivates a gene encoding an oxidoreductase of the short-chain dehydrogenase/reductase (SDR) superfamily, which may play important roles in determining structural variations in bacterial glycans and glycoconjugates. However, the exact nature of this SDR has been a matter of controversy, as different annotations of X. fastidiosa genomes have implicated it in distinct reactions. To confirm the nature of this mutated SDR, a comparative analysis was initially performed, suggesting that it belongs to a subgroup of SDR decarboxylases, representing a UDP-xylose synthase (Uxs). Functional assays, using a recombinant derivative of this enzyme, confirmed its nature as XfUxs, and carbohydrate composition analyses, performed with lipopolysaccharide (LPS) molecules obtained from different strains, indicate that inactivation of the X. fastidiosa uxs gene affects the LPS structure among CLS-related X. fastidiosa strains. Finally, a comparative sequence analysis suggests that this mutation is likely to result in a morphological and evolutionary hallmark that differentiates two subgroups of CLS-related strains, which may influence interactions between these bacteria and their plant and/or insect hosts.

  11. Characterization of the Xylella fastidiosa PD1671 gene encoding degenerate c-di-GMP GGDEF/EAL domains, and its role in the development of Pierce's disease.

    Science.gov (United States)

    Cursino, Luciana; Athinuwat, Dusit; Patel, Kelly R; Galvani, Cheryl D; Zaini, Paulo A; Li, Yaxin; De La Fuente, Leonardo; Hoch, Harvey C; Burr, Thomas J; Mowery, Patricia

    2015-01-01

    Xylella fastidiosa is an important phytopathogenic bacterium that causes many serious plant diseases including Pierce's disease of grapevines. X. fastidiosa is thought to induce disease by colonizing and clogging xylem vessels through the formation of cell aggregates and bacterial biofilms. Here we examine the role in X. fastidiosa virulence of an uncharacterized gene, PD1671, annotated as a two-component response regulator with potential GGDEF and EAL domains. GGDEF domains are found in c-di-GMP diguanylate cyclases while EAL domains are found in phosphodiesterases, and these domains are for c-di-GMP production and turnover, respectively. Functional analysis of the PD1671 gene revealed that it affected multiple X. fastidiosa virulence-related phenotypes. A Tn5 PD1671 mutant had a hypervirulent phenotype in grapevines presumably due to enhanced expression of gum genes leading to increased exopolysaccharide levels that resulted in elevated biofilm formation. Interestingly, the PD1671 mutant also had decreased motility in vitro but did not show a reduced distribution in grapevines following inoculation. Given these responses, the putative PD1671 protein may be a negative regulator of X. fastidiosa virulence.

  12. Characterization of the Xylella fastidiosa PD1671 gene encoding degenerate c-di-GMP GGDEF/EAL domains, and its role in the development of Pierce's disease.

    Directory of Open Access Journals (Sweden)

    Luciana Cursino

    Full Text Available Xylella fastidiosa is an important phytopathogenic bacterium that causes many serious plant diseases including Pierce's disease of grapevines. X. fastidiosa is thought to induce disease by colonizing and clogging xylem vessels through the formation of cell aggregates and bacterial biofilms. Here we examine the role in X. fastidiosa virulence of an uncharacterized gene, PD1671, annotated as a two-component response regulator with potential GGDEF and EAL domains. GGDEF domains are found in c-di-GMP diguanylate cyclases while EAL domains are found in phosphodiesterases, and these domains are for c-di-GMP production and turnover, respectively. Functional analysis of the PD1671 gene revealed that it affected multiple X. fastidiosa virulence-related phenotypes. A Tn5 PD1671 mutant had a hypervirulent phenotype in grapevines presumably due to enhanced expression of gum genes leading to increased exopolysaccharide levels that resulted in elevated biofilm formation. Interestingly, the PD1671 mutant also had decreased motility in vitro but did not show a reduced distribution in grapevines following inoculation. Given these responses, the putative PD1671 protein may be a negative regulator of X. fastidiosa virulence.

  13. The Major Outer Membrane Protein MopB Is Required for Twitching Movement and Affects Biofilm Formation and Virulence in Two Xylella fastidiosa strains.

    Science.gov (United States)

    Chen, Hongyu; Kandel, Prem P; Cruz, Luisa F; Cobine, Paul A; De La Fuente, Leonardo

    2017-11-01

    MopB is a major outer membrane protein (OMP) in Xylella fastidiosa, a bacterial plant pathogen that causes losses on many economically important crops. Based on in silico analysis, the uncharacterized MopB protein of X. fastidiosa contains a β-barrel structure with an OmpA-like domain and a predicted calcium-binding motif. Here, MopB function was studied by mutational analysis taking advantage of the natural competence of X. fastidiosa. Mutants of mopB were constructed in two different X. fastidiosa strains, the type strain Temecula and the more virulent WM1-1. Deletion of the mopB gene impaired cell-to-cell aggregation, surface attachment, and biofilm formation in both strains. Interestingly, mopB deletion completely abolished twitching motility. Electron microscopy of the bacterial cell surface revealed that mopB deletion eliminated type IV and type I pili formation, potentially caused by destabilization of the outer membrane. Both mopB mutants showed reduced virulence using tobacco (Nicotiana tabacum) as a host under greenhouse conditions. These results suggest that MopB has pleiotropic functions in biofilm formation and twitching motility and is important for virulence of X. fastidiosa.

  14. Carbohydrate metabolism of Xylella fastidiosa: Detection of glycolytic and pentose phosphate pathway enzymes and cloning and expression of the enolase gene

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    Facincani Agda Paula

    2003-01-01

    Full Text Available The objective of this work was to assess the functionality of the glycolytic pathways in the bacterium Xylella fastidiosa. To this effect, the enzymes phosphoglucose isomerase, aldolase, glyceraldehyde-3-phosphate dehydrogenase and pyruvate kinase of the glycolytic pathway, and glucose 6-phosphate dehydrogenase of the Entner-Doudoroff pathway were studied, followed by cloning and expression studies of the enolase gene and determination of its activity. These studies showed that X. fastidiosa does not use the glycolytic pathway to metabolize carbohydrates, which explains the increased duplication time of this phytopatogen. Recombinant enolase was expressed as inclusion bodies and solubilized with urea (most efficient extractor, Triton X-100, and TCA. Enolase extracted from X. fastidiosa and from chicken muscle and liver is irreversibly inactivated by urea. The purification of enolase was partial and resulted in a low yield. No enzymatic activity was detected for either recombinant and native enolases, aldolase, and glyceraldehyde-3-phosphate dehydrogenase, suggesting that X. fastidiosa uses the Entner-Doudoroff pathway to produce pyruvate. Evidence is presented supporting the idea that the regulation of genes and the presence of isoforms with regulation patterns might make it difficult to understand the metabolism of carbohydrates in X. fastidiosa.

  15. Ingestão de seiva do xilema de laranjeiras 'Pêra' e 'Valência' (Citrus sinensis (L. Osbeck sadias e infectadas por Xylella fastidiosa, pelas cigarrinhas vetoras Oncometopia facialis e Dilobopterus costalimai (Hemiptera: Cicadellidae Xylem sap ingestion form healthy "Pera" and "Valencia" sweet orange (Citrus sinensis (L. Osbeck and infected ones by Xylella fastidiosa, Oncometopia facialis and Dilobopterus costalimai (Hemiptera: Cicadellidae

    Directory of Open Access Journals (Sweden)

    Luiz Henrique Montesino

    2006-08-01

    Full Text Available Estudou-se o efeito da infecção pela bactéria Xylella fastidiosa, agente causal da Clorose Variegada dos Citros (CVC, sobre a taxa de ingestão de seiva do xilema de plantas cítricas por duas espécies de cigarrinhas vetoras (Hemiptera: Cicadellidae. Foram utilizados pés-francos de laranjeira-doce (Citrus sinensis das variedades 'Pêra' e 'Valência', infectadas por X. fastidiosa da linhagem 9a5c, por meio de inoculação mecânica. Os insetos utilizados nos experimentos foram coletados em campo, sendo um representante da Tribo Cicadellini (Dilobopterus costalimai e um da Proconiini (Oncometopia facialis. A taxa de ingestão de seiva do xilema por O. facialis foi quantificada nos ramos das plantas e a de D. costalimai nas folhas e ramos, por meio da avaliação do volume do líquido (honeydew excretado por unidade de tempo. O consumo pela cigarrinha O. facialis nas plantas doentes foi menor do que nas plantas sadias. Na variedade 'Pêra' doente, o consumo foi baixo, não permitindo a quantificação da seiva eliminada. Na 'Pêra' sadia e na 'Valência' doente e sadia, O. facialis apresentou valores expressivos de excreção, com maior alimentação no período diurno. Nas plantas sadias das duas variedades, o consumo pela cigarrinha D. costalimai foi maior do que nas plantas com CVC. Comparando-se as variedades, o consumo foi superior na variedade 'Valência', e, em relação às partes da planta, folha e ramo, a taxa de ingestão foi maior no ramo das duas variedades, apresentando consumo maior no período diurno.It was studied the effect of Xylella fastidiosa infection, causal agent of Citrus Variegated Chlorosis (CVC, on the xylem sap ingestion rate of citrus plants by two sharpshooters species (Hemiptera: Cicadellidae. Seedlings of sweet orange Pera and Valencia (Citrus sinensis were used and infected by X. fastidiosa, strain 9a5c, obtained by mechanical inoculation. The insects used in the experiments were collected in the field, one

  16. Recent evolutionary radiation and host plant specialization in the Xylella fastidiosa subspecies native to the United States.

    Science.gov (United States)

    Nunney, Leonard; Vickerman, Danel B; Bromley, Robin E; Russell, Stephanie A; Hartman, John R; Morano, Lisa D; Stouthamer, Richard

    2013-04-01

    The bacterial pathogen, Xylella fastidiosa, infects many plant species in the Americas, making it a good model for investigating the genetics of host adaptation. We used multilocus sequence typing (MLST) to identify isolates of the native U.S. subsp. multiplex that were largely unaffected by intersubspecific homologous recombination (IHR) and to investigate how their evolutionary history influences plant host specialization. We identified 110 "non-IHR" isolates, 2 minimally recombinant "intermediate" ones (including the subspecific type), and 31 with extensive IHR. The non-IHR and intermediate isolates defined 23 sequence types (STs) which we used to identify 22 plant hosts (73% trees) characteristic of the subspecies. Except for almond, subsp. multiplex showed no host overlap with the introduced subspecies (subspecies fastidiosa and sandyi). MLST sequences revealed that subsp. multiplex underwent recent radiation (<25% of subspecies age) which included only limited intrasubspecific recombination (ρ/θ = 0.02); only one isolated lineage (ST50 from ash) was older. A total of 20 of the STs grouped into three loose phylogenetic clusters distinguished by nonoverlapping hosts (excepting purple leaf plum): "almond," "peach," and "oak" types. These host differences were not geographical, since all three types also occurred in California. ST designation was a good indicator of host specialization. ST09, widespread in the southeastern United States, only infected oak species, and all peach isolates were ST10 (from California, Florida, and Georgia). Only ST23 had a broad host range. Hosts of related genotypes were sometimes related, but often host groupings crossed plant family or even order, suggesting that phylogenetically plastic features of hosts affect bacterial pathogenicity.

  17. Chemical characterization of Citrus sinensis grafted on C. limonia and the effect of some isolated compounds on the growth of Xylella fastidiosa.

    Science.gov (United States)

    Ribeiro, Alan Bezerra; Abdelnur, Patrícia Verardi; Garcia, Cleverson Fernando; Belini, Adriana; Severino, Vanessa G Pasqualotto; da Silva, M Fátima das G F; Fernandes, João B; Vieira, Paulo C; de Carvalho, Sérgio A; de Souza, Alessandra A; Machado, Marcos A

    2008-09-10

    Citrus sinensis grafted on C. limonia produces a considerable number of compounds that are not common in both plants developed from germination of seeds. The chemical profile of scion and rootstock differ notably for absence in the form of flavonoids and coumarins containing C5 prenyl groups attached to the carbon atoms of aromatic and heterocyclic systems or to oxygen. Only linear pyranocoumarins xanthyletin and xanthoxyletin were found in scion. This observation indicates that the prenylated compounds once biosynthesized in the roots could have been translocated to other organs. Xylella fastidiosa colonizes the xylem of plants causing diseases on several economically important crops such as citrus variegated chlorosis (CVC). A number of flavonoids, coumarins, alkaloids, dihydrocinnamic acid derivative, anacardic acid, triterpenes, and limonoids were tested for in vitro activity on the growth of Xylella fastidiosa. Azadirachtin A was the most active. Hesperidin, which occurs in great amounts in cells of the mesophyll of the affected leaves with CVC, showed a moderate activity suggesting that it can act as a good barrier for small-size colonies from X. fastidiosa.

  18. Detection of siderophores in endophytic bacteria Methylobacterium spp. associated with Xylella fastidiosa subsp. pauca Detecção de sideróforos nas bactérias endofíticas Methylobacterium spp. associadas com Xylella fastidiosa subsp. pauca

    OpenAIRE

    Paulo Teixeira Lacava; Maria Estela Silva-Stenico; Welington Luiz Araújo; Ana Valéria Colnaghi Simionato; Emanuel Carrilho; Siu Mui Tsai; João Lúcio Azevedo

    2008-01-01

    The objective of this work was to study the production of siderophores by endophytic bacteria Methylobacterium spp., which occupy the same ecological niche as Xylella fastidiosa subsp. pauca (Xfp) in citrus plants. The siderophore production of Methylobacterium strains was tested according to chromeazurol agar assay test (CAS), Csáky test (hydroxamate-type) and Arnow test (catechol-type). In addition, the ability of Xfp to use siderophores, in vitro, produced by endophytic bacteria as source ...

  19. Populational fluctuation of vectors of Xylella fastidiosa, wells in sweet orange [Citrus sinensis (L. Osbeck] varieties of northwest Paraná State, Brazil

    Directory of Open Access Journals (Sweden)

    Rúbia de Oliveira Molina

    2010-06-01

    Full Text Available The aim of the present study was to assess the population flutuation of the sharpshooters species subfamily Cicadellinae belonging to the tribes Cicadellini and Proconiini, in sweet orange [Citrus sinensis( L. Osbeck] commercial orchards of the northwest region of Paraná State , Brazil. Samplings were carried out the employing every time 24 yellow sticky cards. Identification of the species showed that the most representative were Dilobopterus costalimai of the Cicadellini tribe and Acrogonia citrina of the Proconiini tribe.A Clorose variegada dos citros (CVC é uma importante doença que ocorre nos citros, cujo agente causal é a bactéria Xylella fastidiosa, Wells. A bactéria depende, obrigatoriamente, de insetos vetores para sua disseminação, que são as cigarrinhas sugadoras do xilema (Hemiptera: Cicadellidae, Cicadellinae. No presente estudo objetivou-se avaliar a flutuação populacional de espécies de cigarrinhas nas diferentes variedades de laranja doce [Citrus sinensis (L. Osbeck], Natal, Pêra, Valência e Folha Murcha, em um pomar comercial localizado na região Noroeste do Paraná, no período de janeiro de 2000 a dezembro de 2002. Amostragens quinzenais foram realizadas com o uso de armadilhas adesivas amarelas, num total de 24 armadilhas em cada avaliação. Após a identificação das espécies observou-se, que as mais representativas foram Dilobopterus costalimai da tribo Cicadellini e Acrogonia citrina da tribo Proconiini, sendo que a variedade de laranja Pêra apresentou o maior número de espécies vetoras durante os anos avaliados.

  20. Analysis of the genome-wide variations among multiple strains of the plant pathogenic bacterium Xylella fastidiosa

    Directory of Open Access Journals (Sweden)

    Walker M Andrew

    2006-09-01

    Full Text Available Abstract Background The Gram-negative, xylem-limited phytopathogenic bacterium Xylella fastidiosa is responsible for causing economically important diseases in grapevine, citrus and many other plant species. Despite its economic impact, relatively little is known about the genomic variations among strains isolated from different hosts and their influence on the population genetics of this pathogen. With the availability of genome sequence information for four strains, it is now possible to perform genome-wide analyses to identify and categorize such DNA variations and to understand their influence on strain functional divergence. Results There are 1,579 genes and 194 non-coding homologous sequences present in the genomes of all four strains, representing a 76. 2% conservation of the sequenced genome. About 60% of the X. fastidiosa unique sequences exist as tandem gene clusters of 6 or more genes. Multiple alignments identified 12,754 SNPs and 14,449 INDELs in the 1528 common genes and 20,779 SNPs and 10,075 INDELs in the 194 non-coding sequences. The average SNP frequency was 1.08 × 10-2 per base pair of DNA and the average INDEL frequency was 2.06 × 10-2 per base pair of DNA. On an average, 60.33% of the SNPs were synonymous type while 39.67% were non-synonymous type. The mutation frequency, primarily in the form of external INDELs was the main type of sequence variation. The relative similarity between the strains was discussed according to the INDEL and SNP differences. The number of genes unique to each strain were 60 (9a5c, 54 (Dixon, 83 (Ann1 and 9 (Temecula-1. A sub-set of the strain specific genes showed significant differences in terms of their codon usage and GC composition from the native genes suggesting their xenologous origin. Tandem repeat analysis of the genomic sequences of the four strains identified associations of repeat sequences with hypothetical and phage related functions. Conclusion INDELs and strain specific genes

  1. The DinJ/RelE Toxin-Antitoxin System Suppresses Bacterial Proliferation and Virulence of Xylella fastidiosa in Grapevine.

    Science.gov (United States)

    Burbank, Lindsey P; Stenger, Drake C

    2017-04-01

    Xylella fastidiosa, the causal agent of Pierce's disease of grapes, is a slow-growing, xylem-limited, bacterial pathogen. Disease progression is characterized by systemic spread of the bacterium through xylem vessel networks, causing leaf-scorching symptoms, senescence, and vine decline. It appears to be advantageous to this pathogen to avoid excessive blockage of xylem vessels, because living bacterial cells are generally found in plant tissue with low bacterial cell density and minimal scorching symptoms. The DinJ/RelE toxin-antitoxin system is characterized here for a role in controlling bacterial proliferation and population size during plant colonization. The DinJ/RelE locus is transcribed from two separate promoters, allowing for coexpression of antitoxin DinJ with endoribonuclease toxin RelE, in addition to independent expression of RelE. The ratio of antitoxin/toxin expressed is dependent on bacterial growth conditions, with lower amounts of antitoxin present under conditions designed to mimic grapevine xylem sap. A knockout mutant of DinJ/RelE exhibits a hypervirulent phenotype, with higher bacterial populations and increased symptom development and plant decline. It is likely that DinJ/RelE acts to prevent excessive population growth, contributing to the ability of the pathogen to spread systemically without completely blocking the xylem vessels and increasing probability of acquisition by the insect vector.

  2. Genome-Wide Analysis Provides Evidence on the Genetic Relatedness of the Emergent Xylella fastidiosa Genotype in Italy to Isolates from Central America.

    Science.gov (United States)

    Giampetruzzi, Annalisa; Saponari, Maria; Loconsole, Giuliana; Boscia, Donato; Savino, Vito Nicola; Almeida, Rodrigo P P; Zicca, Stefania; Landa, Blanca B; Chacón-Diaz, Carlos; Saldarelli, Pasquale

    2017-07-01

    Xylella fastidiosa is a plant-pathogenic bacterium recently introduced in Europe that is causing decline in olive trees in the South of Italy. Genetic studies have consistently shown that the bacterial genotype recovered from infected olive trees belongs to the sequence type ST53 within subspecies pauca. This genotype, ST53, has also been reported to occur in Costa Rica. The ancestry of ST53 was recently clarified, showing it contains alleles that are monophyletic with those of subsp. pauca in South America. To more robustly determine the phylogenetic placement of ST53 within X. fastidiosa, we performed a comparative analysis based on single nucleotide polymorphisms (SNPs) and the study of the pan-genome of the 27 currently public available whole genome sequences of X. fastidiosa. The resulting maximum-parsimony and maximum likelihood trees constructed using the SNPs and the pan-genome analysis are consistent with previously described X. fastidiosa taxonomy, distinguishing the subsp. fastidiosa, multiplex, pauca, sandyi, and morus. Within the subsp. pauca, the Italian and three Costa Rican isolates, all belonging to ST53, formed a compact phylotype in a clade divergent from the South American pauca isolates, also distinct from the recently described coffee isolate CFBP8072 imported into Europe from Ecuador. These findings were also supported by the gene characterization of a conjugative plasmid shared by all the four ST53 isolates. Furthermore, isolates of the ST53 clade possess an exclusive locus encoding a putative ATP-binding protein belonging to the family of histidine kinase-like ATPase gene, which is not present in isolates from the subspecies multiplex, sandyi, and pauca, but was detected in ST21 isolates of the subspecies fastidiosa from Costa Rica. The clustering and distinctiveness of the ST53 isolates supports the hypothesis of their common origin, and the limited genetic diversity among these isolates suggests this is an emerging clade within subsp

  3. Plant water stress effects on stylet probing behaviors of Homalodisca vitripennis (Hemiptera: Cicadellidae) associated with acquisition and inoculation of the bacterium Xylella fastidiosa.

    Science.gov (United States)

    Krugner, Rodrigo; Backus, Elaine A

    2014-02-01

    ABSTRACT The glassy-winged sharpshooter, Homalodisca vitripennis (Germar) (Hemiptera: Cicadellidae), is a xylem fluid-ingesting leafhopper that transmits Xylella fastidiosa Wells et al., a plant-infecting bacterium that causes several plant diseases in the Americas. Although the role of plant water stress on the population density and dispersal ofH. vitripennis has been studied, nothing is known about the effects of plant water stress on the transmission of X. fastidiosa by H. vitripennis. A laboratory study was conducted to determine the influence of plant water stress on the sharpshooter stylet probing behaviors associated with the acquisition and inoculation of X. fastidiosa. Electrical penetration graph was used to monitor H. vitripennis feeding behaviors for 20-h periods on citrus [Citrus sinensis (L.) Osbeck] and almond [Prunus dulcis (Miller) D.A. Webb] plants subjected to levels of water stress. Adult H. vitripennis successfully located xylem vessels, then performed behaviors related to the evaluation of the xylem cell and fluid, and finally ingested xylem fluid from citrus and almond plants under the tested fluid tensions ranging from -5.5 to -33.0 bars and -6.0 to -24.5 bars, respectively. In general, long and frequent feeding events associated with the acquisition and inoculation of X. fastidiosa were observed only in fully irrigated plants (i.e., >-10 bars), which suggests that even low levels of plant water stress may reduce the spread of X. fastidiosa. Results provided insights to disease epidemiology and support the hypothesis that application of regulated deficit irrigation has the potential to reduce the incidence of diseases caused by X.fastidiosa by reducing the number of vectors and by decreasing pathogen transmission efficiency.

  4. Specific Fluorescence in Situ Hybridization (FISH) Test to Highlight Colonization of Xylem Vessels by Xylella fastidiosa in Naturally Infected Olive Trees (Olea europaea L.)

    Science.gov (United States)

    Cardinale, Massimiliano; Luvisi, Andrea; Meyer, Joana B.; Sabella, Erika; De Bellis, Luigi; Cruz, Albert C.; Ampatzidis, Yiannis; Cherubini, Paolo

    2018-01-01

    The colonization behavior of the Xylella fastidiosa strain CoDiRO, the causal agent of olive quick decline syndrome (OQDS), within the xylem of Olea europaea L. is still quite controversial. As previous literature suggests, even if xylem vessel occlusions in naturally infected olive plants were observed, cell aggregation in the formation of occlusions had a minimal role. This observation left some open questions about the whole behavior of the CoDiRO strain and its actual role in OQDS pathogenesis. In order to evaluate the extent of bacterial infection in olive trees and the role of bacterial aggregates in vessel occlusions, we tested a specific fluorescence in situ hybridization (FISH) probe (KO 210) for X. fastidiosa and quantified the level of infection and vessel occlusion in both petioles and branches of naturally infected and non-infected olive trees. All symptomatic petioles showed colonization by X. fastidiosa, especially in the larger innermost vessels. In several cases, the vessels appeared completely occluded by a biofilm containing bacterial cells and extracellular matrix and the frequent colonization of adjacent vessels suggested a horizontal movement of the bacteria. Infected symptomatic trees had 21.6 ± 10.7% of petiole vessels colonized by the pathogen, indicating an irregular distribution in olive tree xylem. Thus, our observations point out the primary role of the pathogen in olive vessel occlusions. Furthermore, our findings indicate that the KO 210 FISH probe is suitable for the specific detection of X. fastidiosa. PMID:29681910

  5. RNA-Seq analysis of Citrus reticulata in the early stages of Xylella fastidiosa infection reveals auxin-related genes as a defense response.

    Science.gov (United States)

    Rodrigues, Carolina M; de Souza, Alessandra A; Takita, Marco A; Kishi, Luciano T; Machado, Marcos A

    2013-10-03

    Citrus variegated chlorosis (CVC), caused by Xylella fastidiosa, is one the most important citrus diseases, and affects all varieties of sweet orange (Citrus sinensis L. Osb). On the other hand, among the Citrus genus there are different sources of resistance against X. fastidiosa. For these species identifying these defense genes could be an important step towards obtaining sweet orange resistant varieties through breeding or genetic engineering. To assess these genes we made use of mandarin (C. reticulata Blanco) that is known to be resistant to CVC and shares agronomical characteristics with sweet orange. Thus, we investigated the gene expression in Ponkan mandarin at one day after infection with X. fastidiosa, using RNA-seq. A set of genes considered key elements in the resistance was used to confirm its regulation in mandarin compared with the susceptible sweet orange. Gene expression analysis of mock inoculated and infected tissues of Ponkan mandarin identified 667 transcripts repressed and 724 significantly induced in the later. Among the induced transcripts, we identified genes encoding proteins similar to Pattern Recognition Receptors. Furthermore, many genes involved in secondary metabolism, biosynthesis and cell wall modification were upregulated as well as in synthesis of abscisic acid, jasmonic acid and auxin. This work demonstrated that the defense response to the perception of bacteria involves cell wall modification and activation of hormone pathways, which probably lead to the induction of other defense-related genes. We also hypothesized the induction of auxin-related genes indicates that resistant plants initially recognize X. fastidiosa as a necrotrophic pathogen.

  6. Development and systematic validation of qPCR assays for rapid and reliable differentiation of Xylella fastidiosa strains causing citrus variegated chlorosis.

    Science.gov (United States)

    Li, Wenbin; Teixeira, Diva C; Hartung, John S; Huang, Qi; Duan, Yongping; Zhou, Lijuan; Chen, Jianchi; Lin, Hong; Lopes, Silvio; Ayres, A Juliano; Levy, Laurene

    2013-01-01

    The xylem-limited, Gram-negative, fastidious plant bacterium Xylella fastidiosa is the causal agent of citrus variegated chlorosis (CVC), a destructive disease affecting approximately half of the citrus plantations in the State of São Paulo, Brazil. The disease was recently found in Central America and is threatening the multi-billion U.S. citrus industry. Many strains of X. fastidiosa are pathogens or endophytes in various plants growing in the U.S., and some strains cross infect several host plants. In this study, a TaqMan-based assay targeting the 16S rDNA signature region was developed for the identification of X. fastidiosa at the species level. Another TaqMan-based assay was developed for the specific identification of the CVC strains. Both new assays have been systematically validated in comparison with the primer/probe sets from four previously published assays on one platform and under similar PCR conditions, and shown to be superior. The species specific assay detected all X. fastidiosa strains and did not amplify any other citrus pathogen or endophyte tested. The CVC-specific assay detected all CVC strains but did not amplify any non-CVC X. fastidiosa nor any other citrus pathogen or endophyte evaluated. Both sets were multiplexed with a reliable internal control assay targeting host plant DNA, and their diagnostic specificity and sensitivity remained unchanged. This internal control provides quality assurance for DNA extraction, performance of PCR reagents, platforms and operators. The limit of detection for both assays was equivalent to 2 to 10 cells of X. fastidiosa per reaction for field citrus samples. Petioles and midribs of symptomatic leaves of sweet orange harbored the highest populations of X. fastidiosa, providing the best materials for detection of the pathogen. These new species specific assay will be invaluable for molecular identification of X. fastidiosa at the species level, and the CVC specific assay will be very powerful for the

  7. Multilocus sequence typing of Xylella fastidiosa isolated from olive affected by “olive quick decline syndrome” in Italy

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    Toufic ELBEAINO

    2015-01-01

    Full Text Available The recent finding of Xylella fastidiosa (Xf in olive trees in southern Italy, the scanty molecular information on this bacterium and its association with the olive quick decline syndrome (OQDS prompted the necessity to isolate and acquire more genetic data on the type of strain present in that region. For the first time, the bacterium was isolated from infected olive on culture media. Genetic information were obtained through genomic comparison with other subspecies or strains. The sequences of thirteen genes from its genome, comprising seven housekeeping genes (leuA, petC, lacF, cysG, holC, nuoL and gltT usually used in multilocus sequence typing (MLST systems, and six genes involved in different biochemical functions (RNA Pol sigma-70 factor, hypothetical protein HL, 16S rRNA, rfbD, nuoN, and pilU, were analyzed. The sequences of the biochemical function genes were explored  individually to study the genetic structure of this bacterium, while the MLST genes were linked together into one concatameric sequence (4161 bp long to increase the resolution of the phylogenetic analysis when compared with Xf strains previously reported. Sequence analyses of single genes showed that the Xf olive strain is distinct from the four previously defined taxons (Xf subsp. fastidiosa, Xf subsp. multiplex, Xf subsp. sandyi and Xf subsp. pauca with a dissimilarity rate that reached 4%. In particular, Xf from olive shared the greatest identity with the strain “9a5c” (subsp. pauca, but was nevertheless distinct from it. Similarly, the MLST based on concatameric sequences confirmed the genetic variance of Xf from olive by generating a novel sequence type profile (ST53. Phylogenetic tree analyses showed that Xf from olive clustered in one clade close to subspecies pauca (strains “9a5c” and “CVC0018”, but was nevertheless distinct from them. These results indicate molecular divergence of this olive bacterium with all other strains yet reported.

  8. Directed evolution induces tributyrin hydrolysis in a virulence factor of Xylella fastidiosa using a duplicated gene as a template.

    Science.gov (United States)

    Gouran, Hossein; Chakraborty, Sandeep; Rao, Basuthkar J; Asgeirsson, Bjarni; Dandekar, Abhaya

    2014-01-01

    Duplication of genes is one of the preferred ways for natural selection to add advantageous functionality to the genome without having to reinvent the wheel with respect to catalytic efficiency and protein stability. The duplicated secretory virulence factors of Xylella fastidiosa (LesA, LesB and LesC), implicated in Pierce's disease of grape and citrus variegated chlorosis of citrus species, epitomizes the positive selection pressures exerted on advantageous genes in such pathogens. A deeper insight into the evolution of these lipases/esterases is essential to develop resistance mechanisms in transgenic plants. Directed evolution, an attempt to accelerate the evolutionary steps in the laboratory, is inherently simple when targeted for loss of function. A bigger challenge is to specify mutations that endow a new function, such as a lost functionality in a duplicated gene. Previously, we have proposed a method for enumerating candidates for mutations intended to transfer the functionality of one protein into another related protein based on the spatial and electrostatic properties of the active site residues (DECAAF). In the current work, we present in vivo validation of DECAAF by inducing tributyrin hydrolysis in LesB based on the active site similarity to LesA. The structures of these proteins have been modeled using RaptorX based on the closely related LipA protein from Xanthomonas oryzae. These mutations replicate the spatial and electrostatic conformation of LesA in the modeled structure of the mutant LesB as well, providing in silico validation before proceeding to the laborious in vivo work. Such focused mutations allows one to dissect the relevance of the duplicated genes in finer detail as compared to gene knockouts, since they do not interfere with other moonlighting functions, protein expression levels or protein-protein interaction.

  9. Transcriptional regulation of the grape cytochrome P450 monooxygenase gene CYP736B expression in response to Xylella fastidiosa infection

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    Walker M Andrew

    2010-07-01

    Full Text Available Abstract Background Plant cytochrome P450 monooxygenases (CYP mediate synthesis and metabolism of many physiologically important primary and secondary compounds that are related to plant defense against a range of pathogenic microbes and insects. To determine if cytochrome P450 monooxygenases are involved in defense response to Xylella fastidiosa (Xf infection, we investigated expression and regulatory mechanisms of the cytochrome P450 monooxygenase CYP736B gene in both disease resistant and susceptible grapevines. Results Cloning of genomic DNA and cDNA revealed that the CYP736B gene was composed of two exons and one intron with GT as a donor site and AG as an acceptor site. CYP736B transcript was up-regulated in PD-resistant plants and down-regulated in PD-susceptible plants 6 weeks after Xf inoculation. However, CYP736B expression was very low in stem tissues at all evaluated time points. 5'RACE and 3'RACE sequence analyses revealed that there were three candidate transcription start sites (TSS in the upstream region and three candidate polyadenylation (PolyA sites in the downstream region of CYP736B. Usage frequencies of each transcription initiation site and each polyadenylation site varied depending on plant genotype, developmental stage, tissue, and treatment. These results demonstrate that expression of CYP736B is regulated developmentally and in response to Xf infection at both transcriptional and post-transcriptional levels. Multiple transcription start and polyadenylation sites contribute to regulation of CYP736B expression. Conclusions This report provides evidence that the cytochrome P450 monooxygenase CYP736B gene is involved in defense response at a specific stage of Xf infection in grapevines; multiple transcription initiation and polyadenylation sites exist for CYP736B in grapevine; and coordinative and selective use of transcription initiation and polyadenylation sites play an important role in regulation of CYP736B expression

  10. Grapevine rootstock effects on scion sap phenolic levels, resistance to Xylella fastidiosa infection, and progression of Pierce’s disease

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    Christopher Michael Wallis

    2013-12-01

    Full Text Available The xylem-limited bacterium Xylella fastidiosa (Xf causes Pierce’s disease (PD, an important disease of grapevine, Vitis vinifera L.. Grapevine rootstocks were developed to provide increased resistance to root disease, but rootstock effects on cane and vine diseases remain unclear. Grapevines that consisted of Cabernet Sauvignon or Chardonnay grafted to 13 different rootstocks were inoculated with Xf and evaluated for PD severity and Xf titer after six months. A subset of six rootstock/scion combinations had xylem sap phenolic levels assessed in non-infected and Xf-infected grapevines. Vigor also was analyzed by measuring root lengths and masses. Cabernet Sauvignon grafted to 101-14MG, 1103P, 420A, or Schwarzmann had reduced PD severity compared to Cabernet Sauvignon grafted to 110R, 5BB, or SO4. Chardonnay grafted to Salt Creek or Freedom had reduced PD severity compared to Chardonnay grafted to RS3 or Schwarzmann. Chardonnay grafted to RS3 had greater Xf titer than Chardonnay grafted to 101-14MG, Freedom, or Salt Creek. No other differences in Xf titer among rootstocks were observed. Of the six scion/rootstock combinations which had xylem sap phenolics analyzed, Chardonnay/ RS3 had the highest levels of most phenolics whereas Cabernet Sauvignon/101-14MG had the lowest phenolic levels. However, Chardonnay/101-14MG, which had mild PD symptoms, had greater sap levels of caftaric acid than other scion/rootstock combinations. Sap levels of caftaric acid, methyl salicylate, a procyanidin trimer, and quinic acid were greater in Xf-infected versus non-infected grapevines. Chardonnay on 101-14MG or Salt Creek had greater root mass than Chardonnay on RS3. Cabernet Sauvignon on 101-14MG had greater root mass than Cabernet Sauvignon on 110R. These results identified rootstocks with the capacity for reducing PD symptom progression. Rootstocks also were shown to affect Xf titer, xylem sap phenolic levels, and plant vigor.

  11. DNA microarray-based genome comparison of a pathogenic and a nonpathogenic strain of Xylella fastidiosa delineates genes important for bacterial virulence.

    Science.gov (United States)

    Koide, Tie; Zaini, Paulo A; Moreira, Leandro M; Vêncio, Ricardo Z N; Matsukuma, Adriana Y; Durham, Alan M; Teixeira, Diva C; El-Dorry, Hamza; Monteiro, Patrícia B; da Silva, Ana Claudia R; Verjovski-Almeida, Sergio; da Silva, Aline M; Gomes, Suely L

    2004-08-01

    Xylella fastidiosa is a phytopathogenic bacterium that causes serious diseases in a wide range of economically important crops. Despite extensive comparative analyses of genome sequences of Xylella pathogenic strains from different plant hosts, nonpathogenic strains have not been studied. In this report, we show that X. fastidiosa strain J1a12, associated with citrus variegated chlorosis (CVC), is nonpathogenic when injected into citrus and tobacco plants. Furthermore, a DNA microarray-based comparison of J1a12 with 9a5c, a CVC strain that is highly pathogenic and had its genome completely sequenced, revealed that 14 coding sequences of strain 9a5c are absent or highly divergent in strain J1a12. Among them, we found an arginase and a fimbrial adhesin precursor of type III pilus, which were confirmed to be absent in the nonpathogenic strain by PCR and DNA sequencing. The absence of arginase can be correlated to the inability of J1a12 to multiply in host plants. This enzyme has been recently shown to act as a bacterial survival mechanism by down-regulating host nitric oxide production. The lack of the adhesin precursor gene is in accordance with the less aggregated phenotype observed for J1a12 cells growing in vitro. Thus, the absence of both genes can be associated with the failure of the J1a12 strain to establish and spread in citrus and tobacco plants. These results provide the first detailed comparison between a nonpathogenic strain and a pathogenic strain of X. fastidiosa, constituting an important step towards understanding the molecular basis of the disease.

  12. The MqsRA Toxin-Antitoxin System from Xylella fastidiosa Plays a Key Role in Bacterial Fitness, Pathogenicity, and Persister Cell Formation

    Science.gov (United States)

    Merfa, Marcus V.; Niza, Bárbara; Takita, Marco A.; De Souza, Alessandra A.

    2016-01-01

    Through the formation of persister cells, bacteria exhibit tolerance to multidrug and other environmental stresses without undergoing genetic changes. The toxin-antitoxin (TA) systems are involved in the formation of persister cells because they are able to induce cell dormancy. Among the TA systems, the MqsRA system has been observed to be highly induced in persister cells of Xylella fastidiosa (causal agent of citrus variegated chlorosis—CVC) activated by copper stress, and has been described in Escherichia coli as related to the formation of persister cells and biofilms. Thus, we evaluated the role of this TA system in X. fastidiosa by overexpressing the MqsR toxin, and verified that the toxin positively regulated biofilm formation and negatively cell movement, resulting in reduced pathogenicity in citrus plants. The overexpression of MqsR also increased the formation of persister cells under copper stress. Analysis of the gene and protein expression showed that this system likely has an autoregulation mechanism to express the toxin and antitoxin in the most beneficial ratio for the cell to oppose stress. Our results suggest that this TA system plays a key role in the adaptation and survival of X. fastidiosa and reveal new insights into the physiology of phytopathogen-host interactions. PMID:27375608

  13. Global expression profile of biofilm resistance to antimicrobial compounds in the plant-pathogenic bacterium Xylella fastidiosa reveals evidence of persister cells.

    Science.gov (United States)

    Muranaka, Lígia S; Takita, Marco A; Olivato, Jacqueline C; Kishi, Luciano T; de Souza, Alessandra A

    2012-09-01

    Investigations of biofilm resistance response rarely focus on plant-pathogenic bacteria. Since Xylella fastidiosa is a multihost plant-pathogenic bacterium that forms biofilm in the xylem, the behavior of its biofilm in response to antimicrobial compounds needs to be better investigated. We analyzed here the transcriptional profile of X. fastidiosa subsp. pauca in response to inhibitory and subinhibitory concentrations of copper and tetracycline. Copper-based products are routinely used to control citrus diseases in the field, while antibiotics are more widely used for bacterial control in mammals. The use of antimicrobial compounds triggers specific responses to each compound, such as biofilm formation and phage activity for copper. Common changes in expression responses comprise the repression of genes associated with metabolic functions and movement and the induction of toxin-antitoxin systems, which have been associated with the formation of persister cells. Our results also show that these cells were found in the population at a ca. 0.05% density under inhibitory conditions for both antimicrobial compounds and that pretreatment with subinhibitory concentration of copper increases this number. No previous report has detected the presence of these cells in X. fastidiosa population, suggesting that this could lead to a multidrug tolerance response in the biofilm under a stressed environment. This is a mechanism that has recently become the focus of studies on resistance of human-pathogenic bacteria to antibiotics and, based on our data, it seems to be more broadly applicable.

  14. Characterization of the TolB-Pal trans-envelope complex from Xylella fastidiosa reveals a dynamic and coordinated protein expression profile during the biofilm development process.

    Science.gov (United States)

    Santos, Clelton A; Janissen, Richard; Toledo, Marcelo A S; Beloti, Lilian L; Azzoni, Adriano R; Cotta, Monica A; Souza, Anete P

    2015-10-01

    The intriguing roles of the bacterial Tol-Pal trans-envelope protein complex range from maintenance of cell envelope integrity to potential participation in the process of cell division. In this study, we report the characterization of the XfTolB and XfPal proteins of the Tol-Pal complex of Xylella fastidiosa. X. fastidiosa is a major plant pathogen that forms biofilms inside xylem vessels, triggering the development of diseases in important cultivable plants around the word. Based on functional complementation experiments in Escherichia coli tolB and pal mutant strains, we confirmed the role of xftolB and xfpal in outer membrane integrity. In addition, we observed a dynamic and coordinated protein expression profile during the X. fastidiosa biofilm development process. Using small-angle X-ray scattering (SAXS), the low-resolution structure of the isolated XfTolB-XfPal complex in solution was solved for the first time. Finally, the localization of the XfTolB and XfPal polar ends was visualized via immunofluorescence labeling in vivo during bacterial cell growth. Our results highlight the major role of the components of the cell envelope, particularly the TolB-Pal complex, during the different phases of bacterial biofilm development. Copyright © 2015 Elsevier B.V. All rights reserved.

  15. Identification of a response regulator involved in surface attachment, cell-cell aggregation, exopolysaccharide production and virulence in the plant pathogen Xylella fastidiosa.

    Science.gov (United States)

    Voegel, Tanja M; Doddapaneni, Harshavardhan; Cheng, Davis W; Lin, Hong; Stenger, Drake C; Kirkpatrick, Bruce C; Roper, M Caroline

    2013-04-01

    Xylella fastidiosa, the causal agent of Pierce's disease of grapevine, possesses several two-component signal transduction systems that allow the bacterium to sense and respond to changes in its environment. Signals are perceived by sensor kinases that autophosphorylate and transfer the phosphate to response regulators (RRs), which direct an output response, usually by acting as transcriptional regulators. In the X. fastidiosa genome, 19 RRs were found. A site-directed knockout mutant in one unusual RR, designated XhpT, composed of a receiver domain and a histidine phosphotransferase output domain, was constructed. The resulting mutant strain was analysed for changes in phenotypic traits related to biofilm formation and gene expression using microarray analysis. We found that the xhpT mutant was altered in surface attachment, cell-cell aggregation, exopolysaccharide (EPS) production and virulence in grapevine. In addition, this mutant had an altered transcriptional profile when compared with wild-type X. fastidiosa in genes for several biofilm-related traits, such as EPS production and haemagglutinin adhesins. © 2012 BSPP AND BLACKWELL PUBLISHING LTD.

  16. Ectopic Expression of Xylella fastidiosa rpfF Conferring Production of Diffusible Signal Factor in Transgenic Tobacco and Citrus Alters Pathogen Behavior and Reduces Disease Severity.

    Science.gov (United States)

    Caserta, R; Souza-Neto, R R; Takita, M A; Lindow, S E; De Souza, A A

    2017-11-01

    The pathogenicity of Xylella fastidiosa is associated with its ability to colonize the xylem of host plants. Expression of genes contributing to xylem colonization are suppressed, while those necessary for insect vector acquisition are increased with increasing concentrations of diffusible signal factor (DSF), whose production is dependent on RpfF. We previously demonstrated that transgenic citrus plants ectopically expressing rpfF from a citrus strain of X. fastidiosa subsp. pauca exhibited less susceptibility to Xanthomonas citri subsp. citri, another pathogen whose virulence is modulated by DSF accumulation. Here, we demonstrate that ectopic expression of rpfF in both transgenic tobacco and sweet orange also confers a reduction in disease severity incited by X. fastidiosa and reduces its colonization of those plants. Decreased disease severity in the transgenic plants was generally associated with increased expression of genes conferring adhesiveness to the pathogen and decreased expression of genes necessary for active motility, accounting for the reduced population sizes achieved in the plants, apparently by limiting pathogen dispersal through the plant. Plant-derived DSF signal molecules in a host plant can, therefore, be exploited to interfere with more than one pathogen whose virulence is controlled by DSF signaling.

  17. Análise espaço-temporal da clorose variegada dos citros no Noroeste do Paraná, com uso de PCR para detecção de Xylella fastidiosa = Spatio-temporal analysis of the citrus variegated chlorosis (CVC in the Northwest of Paraná, using PCR for detection of Xylella fastidiosa

    Directory of Open Access Journals (Sweden)

    William Mário de Carvalho Nunes

    2006-07-01

    Full Text Available A citricultura é afetada por inúmeras doenças, como a clorose variegada do citros (CVC, causada pela bactéria Xylella fastidiosa. O objetivo deste trabalho foi determinar a distribuição espacial da doença dentro de pomares comerciais do Noroeste do Paraná com o uso de métodos moleculares. Foram selecionados pomares sintomáticos para CVC com as variedades ‘Pêra’, ‘Valência’ e ‘Folha Murcha’ (Citrus sinensis Osbeck. Foram marcadas para cada variedade, 4 plantas-referência positivas para CVC (por sintomas e análise molecular e 8 plantas ao redor de cada uma das plantas-referência foram amostradas, num total de 36 plantas por variedade. Realizou-se o teste da Reação da Polimerase em Cadeia (PCR para detecção da bactéria e na mesma época foram conduzidas avaliações visuais de sintomas de CVC. Os resultados da análise temporal, utilizando-se os modelos Monomolecular, Logístico e Gompertz, apontaram o modelo Logístico como o que melhor se ajustou para descrever o comportamento da doença no tempo, para todas as variedades estudadas. Observou-se que o comportamento espacial da doença diferiu quando a mesma área foiavaliada pelos métodos visual e molecular, resultando em uma diferença no padrão espacial das áreas avaliadas. Portanto, ambos os métodos empregados, sintomas e PCR, foram capazes de constatar asmudanças no padrão espacial apresentado, sendo que a análise molecular (PCR foi mais sensível para detectar as mudanças ocorridas.Countless diseases affect the citriculture, as the citrus variegated chlorosis (CVC which is caused by the bacteria Xylella fastidiosa.The aim of this work was to determine the space distribution of the disease inside commercial orchards in the Northwest of Paraná, using molecular methods. Symptomatic orchards were selected for CVC with the varieties 'Pêra', 'Valência' and 'Folha Murcha' (Citrus sinensis Osbeck. For each variety, 4 positive reference-plant for CVC

  18. The chemotaxis regulator pilG of Xylella fastidiosa is required for virulence in Vitis vinifera grapevines

    Science.gov (United States)

    Type IV pili of X. fastidiosa are regulated by pilG, a response regulator protein putatively involved in chemotaxis-like operon sensing stimuli through signal transduction pathways. To elucidate roles of pilG in pathogenicity of X. fastidiosa, the pilG-deletion mutant and complementary strain contai...

  19. A rhamnose-rich O-antigen mediates adhesion, virulence, and host colonization for the xylem-limited phytopathogen Xylella fastidiosa.

    Science.gov (United States)

    Clifford, Jennifer C; Rapicavoli, Jeannette N; Roper, M Caroline

    2013-06-01

    Xylella fastidiosa is a gram-negative, xylem-limited bacterium that causes a lethal disease of grapevine called Pierce's disease. Lipopolysaccharide (LPS) composes approximately 75% of the outer membrane of gram-negative bacteria and, because it is largely displayed on the cell surface, it mediates interactions between the bacterial cell and its surrounding environment. LPS is composed of a conserved lipid A-core oligosaccharide component and a variable O-antigen portion. By targeting a key O-antigen biosynthetic gene, we demonstrate the contribution of the rhamnose-rich O-antigen to surface attachment, cell-cell aggregation, and biofilm maturation: critical steps for successful infection of the host xylem tissue. Moreover, we have demonstrated that a fully formed O-antigen moiety is an important virulence factor for Pierce's disease development in grape and that depletion of the O-antigen compromises its ability to colonize the host. It has long been speculated that cell-surface polysaccharides play a role in X. fastidiosa virulence and this study confirms that LPS is a major virulence factor for this important agricultural pathogen.

  20. N-Acetylcysteine in Agriculture, a Novel Use for an Old Molecule: Focus on Controlling the Plant–Pathogen Xylella fastidiosa

    Science.gov (United States)

    Muranaka, Lígia S.; Giorgiano, Thais E.; Takita, Marco A.; Forim, Moacir R.; Silva, Luis F. C.; Coletta-Filho, Helvécio D.; Machado, Marcos A.; de Souza, Alessandra A.

    2013-01-01

    Xylella fastidiosa is a plant pathogen bacterium that causes diseases in many different crops. In citrus, it causes Citrus Variegated Chlorosis (CVC). The mechanism of pathogenicity of this bacterium is associated with its capacity to colonize and form a biofilm in the xylem vessels of host plants, and there is not yet any method to directly reduce populations of this pathogen in the field. In this study, we investigated the inhibitory effect of N-Acetylcysteine (NAC), a cysteine analogue used mainly to treat human diseases, on X. fastidiosa in different experimental conditions. Concentrations of NAC over 1 mg/mL reduced bacterial adhesion to glass surfaces, biofilm formation and the amount of exopolysaccharides (EPS). The minimal inhibitory concentration of NAC was 6 mg/mL. NAC was supplied to X. fastidiosa-infected plants in hydroponics, fertigation, and adsorbed to organic fertilizer (NAC-Fertilizer). HPLC analysis indicated that plants absorbed NAC at concentrations of 0.48 and 2.4 mg/mL but not at 6 mg/mL. Sweet orange plants with CVC symptoms treated with NAC (0.48 and 2.4 mg/mL) in hydroponics showed clear symptom remission and reduction in bacterial population, as analyzed by quantitative PCR and bacterial isolation. Experiments using fertigation and NAC-Fertilizer were done to simulate a condition closer to that normally is used in the field. For both, significant symptom remission and a reduced bacterial growth rate were observed. Using NAC-Fertilizer the lag for resurgence of symptoms on leaves after interruption of the treatment increased to around eight months. This is the first report of the anti-bacterial effect of NAC against a phytopathogenic bacterium. The results obtained in this work together with the characteristics of this molecule indicate that the use of NAC in agriculture might be a new and sustainable strategy for controlling plant pathogenic bacteria. PMID:24009716

  1. Crystallization and preliminary X-ray analysis of BigR, a transcription repressor from Xylella fastidiosa involved in biofilm formation

    International Nuclear Information System (INIS)

    Barbosa, Rosicler Lázaro; Rinaldi, Fábio Cupri; Guimarães, Beatriz Gomes; Benedetti, Celso Eduardo

    2007-01-01

    In order to gain new insights into the protein structure and its possible interaction with a metal ion or effector ligand, BigR from X. fastidiosa was crystallized in native and selenomethionine (SeMet) labelled forms using the hanging-drop vapour-diffusion method. BigR (biofilm growth-associated repressor) is a novel repressor protein that regulates the transcription of an operon implicated in biofilm growth in both Xylella fastidiosa and Agrobacterium tumefaciens. This protein binds to a palindromic TA-rich element located in the promoter of the BigR operon and strongly represses transcription of the operon. BigR contains a helix–turn–helix (HTH) domain that is found in some members of the ArsR/SmtB family of metal sensors, which control metal resistance in bacteria. Although functional studies have suggested that BigR does not act as a metal sensor, the presence of two cysteines and a methionine in its primary structure raised the possibility of BigR being a metal-ligand protein. In order to gain new insights into the protein structure and its possible interaction with a metal ion or effector ligand, BigR from X. fastidiosa was crystallized in native and selenomethionine (SeMet) labelled forms using the hanging-drop vapour-diffusion method. X-ray diffraction data were collected from native and SeMet crystals to resolutions of 1.95 and 2.2 Å, respectively. Both crystals belong to space group P321 and contain one molecule per asymmetric unit

  2. N-acetylcysteine in agriculture, a novel use for an old molecule: focus on controlling the plant-pathogen Xylella fastidiosa.

    Science.gov (United States)

    Muranaka, Lígia S; Giorgiano, Thais E; Takita, Marco A; Forim, Moacir R; Silva, Luis F C; Coletta-Filho, Helvécio D; Machado, Marcos A; de Souza, Alessandra A

    2013-01-01

    Xylella fastidiosa is a plant pathogen bacterium that causes diseases in many different crops. In citrus, it causes Citrus Variegated Chlorosis (CVC). The mechanism of pathogenicity of this bacterium is associated with its capacity to colonize and form a biofilm in the xylem vessels of host plants, and there is not yet any method to directly reduce populations of this pathogen in the field. In this study, we investigated the inhibitory effect of N-Acetylcysteine (NAC), a cysteine analogue used mainly to treat human diseases, on X. fastidiosa in different experimental conditions. Concentrations of NAC over 1 mg/mL reduced bacterial adhesion to glass surfaces, biofilm formation and the amount of exopolysaccharides (EPS). The minimal inhibitory concentration of NAC was 6 mg/mL. NAC was supplied to X. fastidiosa-infected plants in hydroponics, fertigation, and adsorbed to organic fertilizer (NAC-Fertilizer). HPLC analysis indicated that plants absorbed NAC at concentrations of 0.48 and 2.4 mg/mL but not at 6 mg/mL. Sweet orange plants with CVC symptoms treated with NAC (0.48 and 2.4 mg/mL) in hydroponics showed clear symptom remission and reduction in bacterial population, as analyzed by quantitative PCR and bacterial isolation. Experiments using fertigation and NAC-Fertilizer were done to simulate a condition closer to that normally is used in the field. For both, significant symptom remission and a reduced bacterial growth rate were observed. Using NAC-Fertilizer the lag for resurgence of symptoms on leaves after interruption of the treatment increased to around eight months. This is the first report of the anti-bacterial effect of NAC against a phytopathogenic bacterium. The results obtained in this work together with the characteristics of this molecule indicate that the use of NAC in agriculture might be a new and sustainable strategy for controlling plant pathogenic bacteria.

  3. Sequence/structural analysis of xylem proteome emphasizes pathogenesis-related proteins, chitinases and β-1, 3-glucanases as key players in grapevine defense against Xylella fastidiosa

    Directory of Open Access Journals (Sweden)

    Sandeep Chakraborty

    2016-05-01

    Full Text Available Background. Xylella fastidiosa, the causative agent of various plant diseases including Pierce’s disease in the US, and Citrus Variegated Chlorosis in Brazil, remains a continual source of concern and economic losses, especially since almost all commercial varieties are sensitive to this Gammaproteobacteria. Differential expression of proteins in infected tissue is an established methodology to identify key elements involved in plant defense pathways. Methods. In the current work, we developed a methodology named CHURNER that emphasizes relevant protein functions from proteomic data, based on identification of proteins with similar structures that do not necessarily have sequence homology. Such clustering emphasizes protein functions which have multiple copies that are up/down-regulated, and highlights similar proteins which are differentially regulated. As a working example we present proteomic data enumerating differentially expressed proteins in xylem sap from grapevines that were infected with X. fastidiosa. Results. Analysis of this data by CHURNER highlighted pathogenesis related PR-1 proteins, reinforcing this as the foremost protein function in xylem sap involved in the grapevine defense response to X. fastidiosa. β-1, 3-glucanase, which has both anti-microbial and anti-fungal activities, is also up-regulated. Simultaneously, chitinases are found to be both up and down-regulated by CHURNER, and thus the net gain of this protein function loses its significance in the defense response. Discussion. We demonstrate how structural data can be incorporated in the pipeline of proteomic data analysis prior to making inferences on the importance of individual proteins to plant defense mechanisms. We expect CHURNER to be applicable to any proteomic data set.

  4. N-acetylcysteine in agriculture, a novel use for an old molecule: focus on controlling the plant-pathogen Xylella fastidiosa.

    Directory of Open Access Journals (Sweden)

    Lígia S Muranaka

    Full Text Available Xylella fastidiosa is a plant pathogen bacterium that causes diseases in many different crops. In citrus, it causes Citrus Variegated Chlorosis (CVC. The mechanism of pathogenicity of this bacterium is associated with its capacity to colonize and form a biofilm in the xylem vessels of host plants, and there is not yet any method to directly reduce populations of this pathogen in the field. In this study, we investigated the inhibitory effect of N-Acetylcysteine (NAC, a cysteine analogue used mainly to treat human diseases, on X. fastidiosa in different experimental conditions. Concentrations of NAC over 1 mg/mL reduced bacterial adhesion to glass surfaces, biofilm formation and the amount of exopolysaccharides (EPS. The minimal inhibitory concentration of NAC was 6 mg/mL. NAC was supplied to X. fastidiosa-infected plants in hydroponics, fertigation, and adsorbed to organic fertilizer (NAC-Fertilizer. HPLC analysis indicated that plants absorbed NAC at concentrations of 0.48 and 2.4 mg/mL but not at 6 mg/mL. Sweet orange plants with CVC symptoms treated with NAC (0.48 and 2.4 mg/mL in hydroponics showed clear symptom remission and reduction in bacterial population, as analyzed by quantitative PCR and bacterial isolation. Experiments using fertigation and NAC-Fertilizer were done to simulate a condition closer to that normally is used in the field. For both, significant symptom remission and a reduced bacterial growth rate were observed. Using NAC-Fertilizer the lag for resurgence of symptoms on leaves after interruption of the treatment increased to around eight months. This is the first report of the anti-bacterial effect of NAC against a phytopathogenic bacterium. The results obtained in this work together with the characteristics of this molecule indicate that the use of NAC in agriculture might be a new and sustainable strategy for controlling plant pathogenic bacteria.

  5. Crystallization and preliminary X-ray analysis of BigR, a transcription repressor from Xylella fastidiosa involved in biofilm formation

    Energy Technology Data Exchange (ETDEWEB)

    Barbosa, Rosicler Lázaro; Rinaldi, Fábio Cupri; Guimarães, Beatriz Gomes, E-mail: beatriz@lnls.br; Benedetti, Celso Eduardo, E-mail: beatriz@lnls.br [Center for Molecular and Structural Biology, Brazilian Synchrotron Light Laboratory, Campinas, SP, CP 6192, CEP 13083-970 (Brazil)

    2007-07-01

    In order to gain new insights into the protein structure and its possible interaction with a metal ion or effector ligand, BigR from X. fastidiosa was crystallized in native and selenomethionine (SeMet) labelled forms using the hanging-drop vapour-diffusion method. BigR (biofilm growth-associated repressor) is a novel repressor protein that regulates the transcription of an operon implicated in biofilm growth in both Xylella fastidiosa and Agrobacterium tumefaciens. This protein binds to a palindromic TA-rich element located in the promoter of the BigR operon and strongly represses transcription of the operon. BigR contains a helix–turn–helix (HTH) domain that is found in some members of the ArsR/SmtB family of metal sensors, which control metal resistance in bacteria. Although functional studies have suggested that BigR does not act as a metal sensor, the presence of two cysteines and a methionine in its primary structure raised the possibility of BigR being a metal-ligand protein. In order to gain new insights into the protein structure and its possible interaction with a metal ion or effector ligand, BigR from X. fastidiosa was crystallized in native and selenomethionine (SeMet) labelled forms using the hanging-drop vapour-diffusion method. X-ray diffraction data were collected from native and SeMet crystals to resolutions of 1.95 and 2.2 Å, respectively. Both crystals belong to space group P321 and contain one molecule per asymmetric unit.

  6. Xylella fastidiosa CoDiRO strain associated with the olive quick decline syndrome in southern Italy belongs to a clonal complex of the subspecies pauca that evolved in Central America.

    Science.gov (United States)

    Marcelletti, Simone; Scortichini, Marco

    2016-12-01

    Xylella fastidiosa, a xylem-limited bacterium transmitted by xylem-fluid-feeding Hemiptera insects, causes economic losses of both woody and herbaceous plant species. A Xyl. fastidiosa subsp. pauca strain, namely CoDiRO, was recently found to be associated with the 'olive quick decline syndrome' in southern Italy (i.e. Apulia region). Recently, some Xyl. fastidiosa strains intercepted in France from Coffea spp. plant cuttings imported from Central and South America were characterized. The introduction of infected plant material from Central America in Apulia was also postulated even though an ad hoc study to confirm this hypothesis is lacking. In the present study, we assessed the complete and draft genome of 27 Xyl. fastidiosa strains. Through a genome-wide approach, we confirmed the occurrence of three subspecies within Xyl. fastidiosa, namely fastidiosa, multiplex and pauca, and demonstrated the occurrence of a genetic clonal complex of four Xyl. fastidiosa strains belonging to subspecies pauca which evolved in Central America. The CoDiRO strain displayed 13 SNPs when compared with a strain isolated in Costa Rica from Coffea sp. and 32 SNPs when compared with two strains obtained from Nerium oleander in Costa Rica. These results support the close relationships of the two strains. The four strains in the clonal complex contain prophage-like genes in their genomes. This study strongly supports the possibility of the introduction of Xyl. fastidiosa in southern Italy via coffee plants grown in Central America. The data also stress how the current global circulation of agricultural commodities potentially threatens the agrosystems worldwide.

  7. Characterization of the Xylella fastidiosa PD1671 Gene Encoding Degenerate c-di-GMP GGDEF/EAL Domains, and Its Role in the Development of Pierce’s Disease

    Science.gov (United States)

    Cursino, Luciana; Athinuwat, Dusit; Patel, Kelly R.; Galvani, Cheryl D.; Zaini, Paulo A.; Li, Yaxin; De La Fuente, Leonardo; Hoch, Harvey C.; Burr, Thomas J.; Mowery, Patricia

    2015-01-01

    Xylella fastidiosa is an important phytopathogenic bacterium that causes many serious plant diseases including Pierce’s disease of grapevines. X. fastidiosa is thought to induce disease by colonizing and clogging xylem vessels through the formation of cell aggregates and bacterial biofilms. Here we examine the role in X. fastidiosa virulence of an uncharacterized gene, PD1671, annotated as a two-component response regulator with potential GGDEF and EAL domains. GGDEF domains are found in c-di-GMP diguanylate cyclases while EAL domains are found in phosphodiesterases, and these domains are for c-di-GMP production and turnover, respectively. Functional analysis of the PD1671 gene revealed that it affected multiple X. fastidiosa virulence-related phenotypes. A Tn5 PD1671 mutant had a hypervirulent phenotype in grapevines presumably due to enhanced expression of gum genes leading to increased exopolysaccharide levels that resulted in elevated biofilm formation. Interestingly, the PD1671 mutant also had decreased motility in vitro but did not show a reduced distribution in grapevines following inoculation. Given these responses, the putative PD1671 protein may be a negative regulator of X. fastidiosa virulence. PMID:25811864

  8. Combined use of a new SNP-based assay and multilocus SSR markers to assess genetic diversity of Xylella fastidiosa subsp. pauca infecting citrus and coffee plants.

    Science.gov (United States)

    Montes-Borrego, Miguel; Lopes, Joao R S; Jiménez-Díaz, Rafael M; Landa, Blanca B

    2015-03-01

    Two haplotypes of Xylella fastidiosa subsp. pauca (Xfp) that correlated with their host of origin were identified in a collection of 90 isolates infecting citrus and coffee plants in Brazil, based on a single-nucleotide polymorphism in the gyrB sequence. A new single-nucleotide primer extension (SNuPE) protocol was designed for rapid identification of Xfp according to the host source. The protocol proved to be robust for the prediction of the Xfp host source in blind tests using DNA from cultures of the bacterium, infected plants, and insect vectors allowed to feed on Xfp-infected citrus plants. AMOVA and STRUCTURE analyses of microsatellite data separated most Xfp populations on the basis of their host source, indicating that they were genetically distinct. The combined use of the SNaPshot protocol and three previously developed multilocus SSR markers showed that two haplotypes and distinct isolates of Xfp infect citrus and coffee in Brazil and that multiple, genetically different isolates can be present in a single orchard or infect a single tree. This combined approach will be very useful in studies of the epidemiology of Xfp-induced diseases, host specificity of bacterial genotypes, the occurrence of Xfp host jumping, vector feeding habits, etc., in economically important cultivated plants or weed host reservoirs of Xfp in Brazil and elsewhere. Copyright© by the Spanish Society for Microbiology and Institute for Catalan Studies.

  9. The Type II Secreted Lipase/Esterase LesA is a Key Virulence Factor Required for Xylella fastidiosa Pathogenesis in Grapevines.

    Science.gov (United States)

    Nascimento, Rafael; Gouran, Hossein; Chakraborty, Sandeep; Gillespie, Hyrum W; Almeida-Souza, Hebréia O; Tu, Aye; Rao, Basuthkar J; Feldstein, Paul A; Bruening, George; Goulart, Luiz R; Dandekar, Abhaya M

    2016-01-12

    Pierce's disease (PD) of grapevines is caused by Xylella fastidiosa (Xf), a xylem-limited gamma-proteobacterium that is responsible for several economically important crop diseases. The occlusion of xylem elements and interference with water transport by Xf and its associated biofilm have been posited as the main cause of PD symptom development; however, Xf virulence mechanisms have not been described. Analysis of the Xf secretome revealed a putative lipase/esterase (LesA) that was abundantly secreted in bacterial culture supernatant and was characterized as a protein ortholog of the cell wall-degrading enzyme LipA of Xanthomonas strains. LesA was secreted by Xf and associated with a biofilm filamentous network. Additional proteomic analysis revealed its abundant presence in outer membrane vesicles (OMVs). Accumulation of LesA in leaf regions associated positively with PD symptoms and inversely with bacterial titer. The lipase/esterase also elicited a hypersensitive response in grapevine. Xf lesA mutants were significantly deficient for virulence when mechanically inoculated into grapevines. We propose that Xf pathogenesis is caused by LesA secretion mediated by OMV cargos and that its release and accumulation in leaf margins leads to early stages of observed PD symptoms.

  10. Transcriptome profiling of two olive cultivars in response to infection by the CoDiRO strain of Xylella fastidiosa subsp. pauca.

    Science.gov (United States)

    Giampetruzzi, Annalisa; Morelli, Massimiliano; Saponari, Maria; Loconsole, Giuliana; Chiumenti, Michela; Boscia, Donato; Savino, Vito N; Martelli, Giovanni P; Saldarelli, Pasquale

    2016-06-27

    The recent Xylella fastidiosa subsp. pauca (Xfp) outbreak in olive (Olea europaea) groves in southern Italy is causing a destructive disease denoted Olive Quick Decline Syndrome (OQDS). Field observations disclosed that Xfp-infected plants of cv. Leccino show much milder symptoms, than the more widely grown and highly susceptible cv. Ogliarola salentina. To determine whether these field observations underlie a tolerant condition of cv. Leccino, which could be exploited for lessening the economic impact of the disease on the local olive industry, transcriptional changes occurring in plants of the two cultivars affected by Xfp were investigated. A global quantitative transcriptome profiling comparing susceptible (Ogliarola salentina) and tolerant (Leccino) olive cultivars, infected or not by Xfp, was done on messenger RNA (mRNAs) extracted from xylem tissues. The study revealed that 659 and 447 genes were differentially regulated in cvs Leccino and Ogliarola upon Xfp infection, respectively, whereas 512 genes were altered when the transcriptome of both infected cultivars was compared. Analysis of these differentially expressed genes (DEGs) shows that the presence of Xfp is perceived by the plants of both cultivars, in which it triggers a differential response strongly involving the cell wall. Up-regulation of genes encoding receptor-like kinases (RLK) and receptor-like proteins (RLP) is the predominant response of cv. Leccino, which is missing in cv. Ogliarola salentina. Moreover, both cultivars react with a strong re-modelling of cell wall proteins. These data suggest that Xfp elicits a different transcriptome response in the two cultivars, which determines a lower pathogen concentration in cv. Leccino and indicates that this cultivar may harbor genetic constituents and/or regulatory elements which counteract Xfp infection. Collectively these findings suggest that cv. Leccino is endowed with an intrinsic tolerance to Xfp, which makes it eligible for further studies

  11. Phenotype overlap in Xylella fastidiosa is controlled by the cyclic di-GMP phosphodiesterase Eal in response to antibiotic exposure and diffusible signal factor-mediated cell-cell signaling.

    Science.gov (United States)

    de Souza, Alessandra A; Ionescu, Michael; Baccari, Clelia; da Silva, Aline M; Lindow, Steven E

    2013-06-01

    Eal is an EAL domain protein in Xylella fastidiosa homologous to one involved in resistance to tobramycin in Pseudomonas aeruginosa. EAL and HD-GYP domain proteins are implicated in the hydrolysis of the secondary messenger bis-(3'-5')-cyclic dimeric GMP (cyclic di-GMP). Cell density-dependent communication mediated by a Diffusible Signal Factor (DSF) also modulates cyclic di-GMP levels in X. fastidiosa, thereby controlling the expression of virulence genes and genes involved in insect transmission. The possible linkage of Eal to both extrinsic factors such as antibiotics and intrinsic factors such as quorum sensing, and whether both affect virulence, was thus addressed. Expression of eal was induced by subinhibitory concentrations of tobramycin, and an eal deletion mutant was more susceptible to this antibiotic than the wild-type strain and exhibited phenotypes similar to those of an rpfF deletion mutant blocked in DSF production, such as hypermotility, reduced biofilm formation, and hypervirulence to grape. Consistent with that, the rpfF mutant was more susceptible than the wild-type strain to tobramycin. Therefore, we propose that cell-cell communication and antibiotic stress can apparently lead to similar modulations of cyclic di-GMP in X. fastidiosa, resulting in similar phenotypes. However, the effect of cell density is dominant compared to that of antibiotic stress, since eal is suppressed by RpfF, which may prevent inappropriate behavioral changes in response to antibiotic stress when DSF accumulates.

  12. Identification of Xylella fastidiosa antivirulence genes: hemagglutinin adhesins contribute a biofilm maturation to X. fastidios and colonization and attenuate virulence.

    Science.gov (United States)

    Guilhabert, Magalie R; Kirkpatrick, Bruce C

    2005-08-01

    Xylella fastidosa, a gram-negative, xylem-limited bacterium, is the causal agent of several economically important plant diseases, including Pierce's disease (PD) and citrus variegated chlorosis (CVC). Until recently, the inability to transform or produce transposon mutants of X. fastidosa had been a major impediment to identifying X. fastidosa genes that mediate pathogen and plant interactions. A random transposon (Tn5) library of X. fastidosa was constructed and screened for mutants showing more severe symptoms and earlier grapevine death (hypervirulence) than did vines infected with the wild type. Seven hypervirulent mutants identified in this screen moved faster and reached higher populations than the wild type in grapevines. These results suggest that X. fastidosa attenuates its virulence in planta and that movement is important in X. fastidosa virulence. The mutated genes were sequenced and none had been described previously as antivirulence genes, although six of them showed similarity with genes of known functions in other organisms. One transposon insertion inactivated a hemagglutinin adhesin gene (PD2118), which we named HxfA. Another mutant in a second putative X. fastidosa hemagglutinin gene, PD1792 (HxfB), was constructed, and further characterization of these hxf mutants suggests that X. fastidosa hemagglutinins mediate contact between X. fastidosa cells, which results in colony formation and biofilm maturation within the xylem vessels.

  13. Quantification and localization of hesperidin and rutin in Citrus sinensis grafted on C. limonia after Xylella fastidiosa infection by HPLC-UV and MALDI imaging mass spectrometry.

    Science.gov (United States)

    Soares, Márcio Santos; da Silva, Danielle Fernandes; Forim, Moacir Rossi; da Silva, Maria Fátima das Graças Fernandes; Fernandes, João Batista; Vieira, Paulo Cezar; Silva, Denise Brentan; Lopes, Norberto Peporine; de Carvalho, Sérgio Alves; de Souza, Alessandra Alves; Machado, Marcos Antônio

    2015-07-01

    A high performance liquid chromatography-ultraviolet (HPLC-UV) method was developed for quantifying hesperidin and rutin levels in leaves and stems of Citrus limonia, with a good linearity over a range of 1.0-80.0 and 1.0-50.0 μg mL(-1) respectively, with r(2)>0.999 for all curves. The limits of detection (LOD) for both flavonoids were 0.6 and 0.5 μg mL(-1), respectively, with quantification (LOQ) being 2.0 and 1.0 μg mL(-1), respectively. The quantification method was applied to Citrus sinensis grafted onto C. limonia with and without CVC (citrus variegated chlorosis) symptoms after Xylella fastidiosa infection. The total content of rutin was low and practically constant in all analyses in comparison with hesperidin, which showed a significant increase in its amount in symptomatic leaves. Scanning electron microscopy studies on leaves with CVC symptoms showed vessel occlusion by biofilm, and a crystallized material was noted. Considering the difficulty in isolating these crystals for analysis, tissue sections were analyzed by matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI MSI) to confirm the presence of hesperidin at the site of infection. The images constructed from MS/MS data with a specific diagnostic fragment ion (m/z 483) also showed higher ion intensities for it in infected plants than in healthy ones, mainly in the vessel regions. These data suggest that hesperidin plays a role in the plant-pathogen interaction, probably as a phytoanticipin. This method was also applied to C. sinensis and C. limonia seedlings, and comparison with the graft results showed that the rootstock had an increased hesperidin content ∼3.6 fold greater in the graft stem than in the stem of C. sinensis seedlings. Increase in hesperidin content by rootstock can be related to induced internal defense mechanisms. Copyright © 2015 Elsevier Ltd. All rights reserved.

  14. Enhanced reliability and accuracy for field deployable bioforensic detection and discrimination of Xylella fastidiosa subsp. pauca, causal agent of citrus variegated chlorosis using razor ex technology and TaqMan quantitative PCR.

    Science.gov (United States)

    Ouyang, Ping; Arif, Mohammad; Fletcher, Jacqueline; Melcher, Ulrich; Ochoa Corona, Francisco Manuel

    2013-01-01

    A reliable, accurate and rapid multigene-based assay combining real time quantitative PCR (qPCR) and a Razor Ex BioDetection System (Razor Ex) was validated for detection of Xylella fastidiosa subsp. pauca (Xfp, a xylem-limited bacterium that causes citrus variegated chlorosis [CVC]). CVC, which is exotic to the United States, has spread through South and Central America and could significantly impact U.S. citrus if it arrives. A method for early, accurate and sensitive detection of Xfp in plant tissues is needed by plant health officials for inspection of products from quarantined locations, and by extension specialists for detection, identification and management of disease outbreaks and reservoir hosts. Two sets of specific PCR primers and probes, targeting Xfp genes for fimbrillin and the periplasmic iron-binding protein were designed. A third pair of primers targeting the conserved cobalamin synthesis protein gene was designed to detect all possible X. fastidiosa (Xf) strains. All three primer sets detected as little as 1 fg of plasmid DNA carrying X. fastidiosa target sequences and genomic DNA of Xfp at as little as 1 - 10 fg. The use of Razor Ex facilitates a rapid (about 30 min) in-field assay capability for detection of all Xf strains, and for specific detection of Xfp. Combined use of three primer sets targeting different genes increased the assay accuracy and broadened the range of detection. To our knowledge, this is the first report of a field-deployable rapid and reliable bioforensic detection and discrimination method for a bacterial phytopathogen based on multigene targets.

  15. A TaqMan-based real time PCR assay for specific detection and quantification of Xylella fastidiosa strains causing bacterial leaf scorch in oleander.

    Science.gov (United States)

    Guan, Wei; Shao, Jonathan; Singh, Raghuwinder; Davis, Robert E; Zhao, Tingchang; Huang, Qi

    2013-02-15

    A TaqMan-based real-time PCR assay was developed for specific detection of strains of X. fastidiosa causing oleander leaf scorch. The assay uses primers WG-OLS-F1 and WG-OLS-R1 and the fluorescent probe WG-OLS-P1, designed based on unique sequences found only in the genome of oleander strain Ann1. The assay is specific, allowing detection of only oleander-infecting strains, not other strains of X. fastidiosa nor other plant-associated bacteria tested. The assay is also sensitive, with a detection limit of 10.4fg DNA of X. fastidiosa per reaction in vitro and in planta. The assay can also be applied to detect low numbers of X. fastidiosa in insect samples, or further developed into a multiplex real-time PCR assay to simultaneously detect and distinguish diverse strains of X. fastidiosa that may occupy the same hosts or insect vectors. Specific and sensitive detection and quantification of oleander strains of X. fastidiosa should be useful for disease diagnosis, epidemiological studies, management of oleander leaf scorch disease, and resistance screening for oleander shrubs. Published by Elsevier B.V.

  16. Conformational variability of the stationary phase survival protein E from Xylella fastidiosa revealed by X-ray crystallography, small-angle X-ray scattering studies, and normal mode analysis.

    Science.gov (United States)

    Machado, Agnes Thiane Pereira; Fonseca, Emanuella Maria Barreto; Reis, Marcelo Augusto Dos; Saraiva, Antonio Marcos; Santos, Clelton Aparecido Dos; de Toledo, Marcelo Augusto Szymanski; Polikarpov, Igor; de Souza, Anete Pereira; Aparicio, Ricardo; Iulek, Jorge

    2017-10-01

    Xylella fastidiosa is a xylem-limited bacterium that infects a wide variety of plants. Stationary phase survival protein E is classified as a nucleotidase, which is expressed when bacterial cells are in the stationary growth phase and subjected to environmental stresses. Here, we report four refined X-ray structures of this protein from X. fastidiosa in four different crystal forms in the presence and/or absence of the substrate 3'-AMP. In all chains, the conserved loop verified in family members assumes a closed conformation in either condition. Therefore, the enzymatic mechanism for the target protein might be different of its homologs. Two crystal forms exhibit two monomers whereas the other two show four monomers in the asymmetric unit. While the biological unit has been characterized as a tetramer, differences of their sizes and symmetry are remarkable. Four conformers identified by Small-Angle X-ray Scattering (SAXS) in a ligand-free solution are related to the low frequency normal modes of the crystallographic structures associated with rigid body-like protomer arrangements responsible for the longitudinal and symmetric adjustments between tetramers. When the substrate is present in solution, only two conformers are selected. The most prominent conformer for each case is associated to a normal mode able to elongate the protein by moving apart two dimers. To our knowledge, this work was the first investigation based on the normal modes that analyzed the quaternary structure variability for an enzyme of the SurE family followed by crystallography and SAXS validation. The combined results raise new directions to study allosteric features of XfSurE protein. © 2017 Wiley Periodicals, Inc.

  17. Structures of the N-acetyltransferase domain of Xylella fastidiosa N-acetyl-L-glutamate synthase/kinase with and without a His tag bound to N-acetyl-L-glutamate.

    Science.gov (United States)

    Zhao, Gengxiang; Jin, Zhongmin; Allewell, Norma M; Tuchman, Mendel; Shi, Dashuang

    2015-01-01

    Structures of the catalytic N-acetyltransferase (NAT) domain of the bifunctional N-acetyl-L-glutamate synthase/kinase (NAGS/K) from Xylella fastidiosa bound to N-acetyl-L-glutamate (NAG) with and without an N-terminal His tag have been solved and refined at 1.7 and 1.4 Å resolution, respectively. The NAT domain with an N-terminal His tag crystallized in space group P4(1)2(1)2, with unit-cell parameters a=b=51.72, c=242.31 Å. Two subunits form a molecular dimer in the asymmetric unit, which contains ∼41% solvent. The NAT domain without an N-terminal His tag crystallized in space group P21, with unit-cell parameters a=63.48, b=122.34, c=75.88 Å, β=107.6°. Eight subunits, which form four molecular dimers, were identified in the asymmetric unit, which contains ∼38% solvent. The structures with and without the N-terminal His tag provide an opportunity to evaluate how the His tag affects structure and function. Furthermore, multiple subunits in different packing environments allow an assessment of the plasticity of the NAG binding site, which might be relevant to substrate binding and product release. The dimeric structure of the X. fastidiosa N-acetytransferase (xfNAT) domain is very similar to that of human N-acetyltransferase (hNAT), reinforcing the notion that mammalian NAGS is evolutionally derived from bifunctional bacterial NAGS/K.

  18. Association of the sharpshooter X wave with xylem inoculation of Xylella fastidiosa leading to systemic, symptomatic Pierce’s diesease infection in grape

    Science.gov (United States)

    Despite several decades of study, the mechanism of inoculation of X. fastidiosa (Xf) to grapevines by its sharpshooter vectors still is not fully understood. Recent research showed that Xf is inoculated into or onto artificial diets by a combination of egestion and salivation. However, the salivatio...

  19. Kennisleemtes over Xylella vragen meer onderzoek

    NARCIS (Netherlands)

    Wolf, van der J.M.

    2016-01-01

    tijdens workshops over Xylella fastidiosa, in augustus in Milaan en onlangs in Brussel, bleek dat er nog veel vragen onbeantwoord zijn over de bacterie, verspreiding en waardplanten. In Noord- en Zuid-Amerika is al veel expertise aanwezig. Dat wordt gebruikt voor verder onderzoek binnen de EU. Doel

  20. Directed evolution induces tributyrin hydrolysis in a virulence factor of Xylella fastidiosa using a duplicated gene as a template [v1; ref status: indexed, http://f1000r.es/48i

    Directory of Open Access Journals (Sweden)

    Hossein Gouran

    2014-09-01

    Full Text Available Duplication of genes is one of the preferred ways for natural selection to add advantageous functionality to the genome without having to reinvent the wheel with respect to catalytic efficiency and protein stability. The duplicated secretory virulence factors of Xylella fastidiosa (LesA, LesB and LesC, implicated in Pierce's disease of grape and citrus variegated chlorosis of citrus species, epitomizes the positive selection pressures exerted on advantageous genes in such pathogens. A deeper insight into the evolution of these lipases/esterases is essential to develop resistance mechanisms in transgenic plants. Directed evolution, an attempt to accelerate the evolutionary steps in the laboratory, is inherently simple when targeted for loss of function. A bigger challenge is to specify mutations that endow a new function, such as a lost functionality in a duplicated gene. Previously, we have proposed a method for enumerating candidates for mutations intended to transfer the functionality of one protein into another related protein based on the spatial and electrostatic properties of the active site residues (DECAAF. In the current work, we present in vivo validation of DECAAF by inducing tributyrin hydrolysis in LesB based on the active site similarity to LesA. The structures of these proteins have been modeled using RaptorX based on the closely related LipA protein from Xanthomonas oryzae. These mutations replicate the spatial and electrostatic conformation of LesA in the modeled structure of the mutant LesB as well, providing in silico validation before proceeding to the laborious in vivo work. Such focused mutations allows one to dissect the relevance of the duplicated genes in finer detail as compared to gene knockouts, since they do not interfere with other moonlighting functions, protein expression levels or protein-protein interaction.

  1. Xylella taiwanensis sp. nov. cause of pear leaf scorch disease in Taiwan

    Science.gov (United States)

    Xylella fastidiosa is a group of xylem-limited and nutritionally fastidious plant pathogenic bacteria. While mostly found in the Americas, new X. fastidiosa strains have been reported from other continents such as Asia, including a pear leaf scorch (PLS) strain from Taiwan. Current taxonomy of X. fa...

  2. A Simple Defined Medium for the Production of True Diketopiperazines in Xylella fastidiosa and Their Identification by Ultra-Fast Liquid Chromatography-Electrospray Ionization Ion Trap Mass Spectrometry.

    Science.gov (United States)

    Silva, Michelli Massaroli da; Andrade, Moacir Dos Santos; Bauermeister, Anelize; Merfa, Marcus Vinícius; Forim, Moacir Rossi; Fernandes, João Batista; Vieira, Paulo Cezar; Silva, Maria Fátima das Graças Fernandes da; Lopes, Norberto Peporine; Machado, Marcos Antônio; Souza, Alessandra Alves de

    2017-06-13

    Diketopiperazines can be generated by non-enzymatic cyclization of linear dipeptides at extreme temperature or pH, and the complex medium used to culture bacteria and fungi including phytone peptone and trypticase peptone, can also produce cyclic peptides by heat sterilization. As a result, it is not always clear if many diketopiperazines reported in the literature are artifacts formed by the different complex media used in microorganism growth. An ideal method for analysis of these compounds should identify whether they are either synthesized de novo from the products of primary metabolism and deliver true diketopiperazines. A simple defined medium ( X. fastidiosa medium or XFM) containing a single carbon source and no preformed amino acids has emerged as a method with a particularly high potential for the grown of X. fastidiosa and to produce genuine natural products. In this work, we identified a range of diketopiperazines from X. fastidiosa 9a5c growth in XFM, using Ultra-Fast Liquid Chromatography coupled with mass spectrometry. Diketopiperazines are reported for the first time from X. fastidiosa , which is responsible for citrus variegated chlorosis. We also report here fatty acids from X. fastidiosa , which were not biologically active as diffusible signals, and the role of diketopiperazines in signal transduction still remains unknown.

  3. A Simple Defined Medium for the Production of True Diketopiperazines in Xylella fastidiosa and Their Identification by Ultra-Fast Liquid Chromatography-Electrospray Ionization Ion Trap Mass Spectrometry

    Directory of Open Access Journals (Sweden)

    Michelli Massaroli da Silva

    2017-06-01

    Full Text Available Diketopiperazines can be generated by non-enzymatic cyclization of linear dipeptides at extreme temperature or pH, and the complex medium used to culture bacteria and fungi including phytone peptone and trypticase peptone, can also produce cyclic peptides by heat sterilization. As a result, it is not always clear if many diketopiperazines reported in the literature are artifacts formed by the different complex media used in microorganism growth. An ideal method for analysis of these compounds should identify whether they are either synthesized de novo from the products of primary metabolism and deliver true diketopiperazines. A simple defined medium (X. fastidiosa medium or XFM containing a single carbon source and no preformed amino acids has emerged as a method with a particularly high potential for the grown of X. fastidiosa and to produce genuine natural products. In this work, we identified a range of diketopiperazines from X. fastidiosa 9a5c growth in XFM, using Ultra-Fast Liquid Chromatography coupled with mass spectrometry. Diketopiperazines are reported for the first time from X. fastidiosa, which is responsible for citrus variegated chlorosis. We also report here fatty acids from X. fastidiosa, which were not biologically active as diffusible signals, and the role of diketopiperazines in signal transduction still remains unknown.

  4. Chromosome-Based Genetic Complementation System for Xylella fastidiosa▿

    OpenAIRE

    Matsumoto, Ayumi; Young, Glenn M.; Igo, Michele M.

    2009-01-01

    Xylella fastidiosa is a xylem-limited, gram-negative bacterium that causes Pierce's disease of grapevine. Here, we describe the construction of four vectors that facilitate the insertion of genes into a neutral site (NS1) in the X. fastidiosa chromosome. These vectors carry a colE1-like (pMB1) replicon and DNA sequences from NS1 flanking a multiple-cloning site and a resistance marker for one of the following antibiotics: chloramphenicol, erythromycin, gentamicin, or kanamycin. In X. fastidio...

  5. Acquisition of Xyllela fastidiosa causes changes to the inoculation behavior (EPG X wave) of an efficient sharpshooter vector

    Science.gov (United States)

    Xylella fastidiosa (Xf) is a foregut-borne bacterium that is inoculated into xylem cells of a healthy plant during feeding by sharpshooter vectors. Inoculation occurs during salivation and egestion behaviors that are likely represented by the sharpshooter X wave. The objective of this study was to t...

  6. Avaliação de tangerinas, tangores e tangelos em relação à clorose variegada dos citros Evaluation of mandarines, tangors and tangelos in relation to the citrus variegated chlorosis (CVC

    Directory of Open Access Journals (Sweden)

    Simone Rodrigues da Silva

    2004-04-01

    Full Text Available A Clorose Variegada dos Citros (CVC, causada pela bactéria Xylella fastidiosa, é uma séria ameaça à citricultura brasileira, constituindo-se, atualmente, numa das principais doenças dos citros no Brasil. O objetivo desse trabalho foi avaliar cultivares de citros introduzidas quanto à suscetibilidade ou resistência à CVC, em condições de campo. O trabalho foi conduzido em Bebedouro-SP. Os materiais genéticos estudados foram cultivares de tangerinas e híbridos (tangores e tangelos introduzidas de bancos de germoplasma da Itália, Portugal, Espanha e Córsega. O trabalho foi constituído por 56 parcelas, com três plantas das quais uma foi inoculada, empregando-se o método de encostia, utilizando-se de mudas previamente infectadas como fontes da bactéria. Foram avaliados os sintomas da doença mediante observação visual através de notas e teste de PCR, específico para Xylella fastidiosa. Os materiais genéticos que se apresentaram positivos ao PCR, mas não apresentaram sintomas, e os que foram negativos ao PCR, possuem um potencial de utilização em programas de melhoramento genético visando à resistência e/ou à tolerância à doença.The Citrus Variegated Chlorosis (CVC, caused by the bacteria Xylella fastidiosa, is a serious threat to the Brazilian citriculture, being, actually, one of the mains diseases to the citrus in Brazil. The objective of this work was evaluate citrus cultivars, introduced according to the susceptibility or resistance to CVC, in field conditions. The research was carried in Bebedouro city - São Paulo state - Brazil. The genotypes studied were cultivars of mandarines and hybrids (tangors and tangelos, introduced from the germplasm collections of Italy, Portugal, Spain and Corsica. The work was composed by 56 portions, with 3 plants, being one of them inoculated, using the approach graft method, with infected plants. Were evaluated the symptoms of the disease by visual observation and PCR test

  7. Universal detection of phytoplasmas and Xylella spp. by TaqMan singleplex and multiplex real-time PCR with dual priming oligonucleotides.

    Directory of Open Access Journals (Sweden)

    Takao Ito

    Full Text Available Phytoplasmas and Xylella spp. are bacteria that cause many economically important plant diseases worldwide. TaqMan probe-based quantitative real-time polymerase chain reaction (qPCR assays have been utilized to universally detect phytoplasmas or Xylella fastidiosa. To develop a superior universal qPCR method, we used a dual priming oligonucleotide (DPO with two annealing sites as a reverse primer to target the well-conserved bacterial 16S rDNA. The new qPCR assays universally detected various species of phytoplasmas and subspecies of X. fastidiosa as well as Xylella taiwanensis, and generally showed superior threshold cycle values when amplifying specific or non-specific products compared to current universal qPCR assays. The proposed qPCR assays were integrated to develop a multiplex qPCR assay that simultaneously detected phytoplasmas, Xylella spp., and an internal plant DNA positive control within 1 hour. This assay could detect a minimum of ten bacterial cells and was compatible with crude extractions used in the rapid screening of various plants. The amplicons were of sufficient lengths to be directly sequenced for preliminary identification, and the primers could be used in universal conventional PCR assays. Additionally, reverse DPO primers can be utilized to improve other probe-based qPCR assays.

  8. Bacterial leaf scorch distribution and isothermal lines (PROJECT NC-EM-08-02)

    Science.gov (United States)

    Gerard C. Adams; Mursel Catall; James Walla; Ann B. Gould

    2013-01-01

    Bacterial leaf scorch (BLS) of shade trees is the common name for a disease caused by Xylella fastidiosa, a xylem-inhabiting bacterium that has fastidious nutritional requirements and is difficult to culture or verify by culturing. Forest trees including oak, sycamore, elm, planetree, sweetgum, mulberry and maple are species susceptible to ...

  9. Espécies de cigarrinhas em cultivo de café no Município de Santa Teresa, Espírito Santo, Brasil (Hemiptera, Cicadellidae, Cicadellinae

    Directory of Open Access Journals (Sweden)

    Rachel A. Carvalho

    2015-01-01

    Full Text Available As cigarrinhas da subfamília Cicadellinae são importantes vetores de patógenos de plantas cultivadas, dentre eles, a bactéria Xylella fastidiosa, que, no Brasil, ataca cultivos de citros, café e também de ameixa. Pouca informação é conhecida sobre a ocorrência, distribuição e biologia de cicadelíneos em plantações de café. Este trabalho apresenta uma lista de 141 espécimes, coletados em meses alternados, no período entre junho de 2009 e abril de 2010, em plantação de café no Município de Santa Teresa no Estado do Espírito Santo, Brasil. Esses espécimes estão distribuídos em 16 gêneros e 21 espécies pertencentes às duas tribos de Cicadellinae: Cicadellini e Proconiini. Dentre os Cicadellini listados, o gênero Graphocephala é, pela primeira vez, registrado para o Brasil.

  10. Influence of Xylella fastidiosa cold shock proteins on pathogenesis in grapevine.

    Science.gov (United States)

    Cold shock proteins (CSPs), a family of nucleic acid binding proteins are an essential part of microbial adaptation to temperature changes. Bacterial CSPs are often expressed in a temperature-dependent manner, and act as chaperones, facilitating translation at low temperature by stabilizing mRNA. In...

  11. Egg maturation by the glassy-winged sharpshooter (Hemiptera: Cicadellidae); a vector of Xylella fastidiosa

    Science.gov (United States)

    Rates of spread of insect-transmitted plant pathogens are a function of vector abundance. Despite this, factors affecting population growth rates of insects that transmit plant pathogens have received limited attention. The glassy-winged sharpshooter (Homalodisca vitripennis) feeds on xylem-sap and ...

  12. Deficiência hídrica agrava os sintomas fisiológicos da clorose variegada dos citros em laranjeira 'Natal' Water deficiency intensifies physiological symptoms of citrus variegated clorosis in 'Natal' sweet orange plants

    Directory of Open Access Journals (Sweden)

    Eduardo Caruso Machado

    2007-01-01

    Full Text Available A clorose variegada dos citros (CVC é uma doença que tem promovido sérios prejuízos aos laranjais das regiões Norte e Nordeste do Estado de São Paulo, onde a deficiência hídrica e as altas temperaturas são mais frequentes. Assim, este trabalho objetivou a avaliação do efeito da deficiência hídrica no desenvolvimento de sintomas fisiológicos em laranjeira 'Natal' com CVC. Foram realizadas medidas do potencial da água na folha, transpiração, condutância estomática e assimilação de CO2, em laranjeiras em condições naturais e submetidas à irrigação. O delineamento experimental foi em blocos ao acaso com cinco repetições. A condutância estomática, a transpiração diária e o potencial da água na folha foram menores nas plantas com CVC. A assimilação diária de CO2 foi menor nas laranjeiras com CVC mesmo quando irrigadas. De fato, a irrigação diminuiu o efeito da CVC, porém não impediu o estabelecimento da doença em laranjeiras inoculadas com Xylella fastidiosa. Em relação aos demais tratamentos, as plantas infectadas e mantidas sob condições naturais (sem irrigação apresentaram maior comprometimento das trocas gasosas, mesmo quando as avaliações fisiológicas foram feitas em período úmido (verão.Citrus variegated chlorosis (CVC is a disease that has caused serious economical losses in citrus grove located in the North and Northeastern regions of São Paulo State, where water deficiency and high temperature occur frequently. Therefore, the aim of this work was to evaluate the influence of water deficiency on the development of physiological symptoms in 'Natal' sweet orange plants with CVC. Measurements of leaf water potential, transpiration, stomatal conductance e CO2 assimilation were taken in plants under natural conditions and submitted to irrigation. The experimental design was in randomized blocks, with five replications. Stomatal conductance, daily transpiration and leaf water potential were

  13. Fluxo de seiva e fotossíntese em laranjeira 'Natal' com clorose variegada dos citros Sap flow and photosynthesis of 'Natal' sweet orange plants with citrus variegated chlorosis

    Directory of Open Access Journals (Sweden)

    Eduardo Caruso Machado

    2006-06-01

    Full Text Available O objetivo deste trabalho foi avaliar os efeitos da clorose variegada dos citros (CVC, no fluxo de seiva, trocas gasosas e atividade fotoquímica em laranjeira 'Natal', com e sem CVC, em condição de campo. O curso diário do fluxo de seiva, potencial da água na folha, assimilação de CO2, transpiração, condutância estomática e eficiência quântica máxima e efetiva do fotossistema II foram avaliados. O delineamento experimental foi em blocos ao acaso com cinco repetições. O fluxo de seiva foi 1,9 vez superior nas plantas sadias em relação às doentes. Em plantas doentes ocorreu queda de 43, 28 e 33% na assimilação de CO2, condutância estomática e transpiração, respectivamente. As plantas com CVC apresentaram fotoinibição dinâmica. Uma vez que a eficiência quântica efetiva apresentou um padrão de resposta semelhante, durante o dia, em ambos os tratamentos, o efeito protetor da fotorrespiração no aparato fotoquímico em plantas com CVC é discutido. As quedas de assimilação de CO2, transpiração e de fluxo de seiva, nas plantas com CVC, foram decorrentes do menor valor da condutância estomática, possivelmente causado pela colonização dos vasos do xilema pela Xylella fastidiosa.The objective of this work was to evaluate the effects of citrus variegated chlorosis (CVC on sap flow, gas exchanges and photochemical activity in 'Natal' sweet orange plants with and without CVC under field condition. Diurnal courses of sap flow, leaf water potential, CO2 assimilation rate and transpiration, stomatal conductance, potential and effective quantum efficiency of photosystem II were evaluated. The experiment was arranged in a random block design with five repetitions. Healthy plants showed sap flow values around 1.9 times higher than injured ones. Injured plants exhibited reductions of 43, 28 and 33% in CO2 assimilation rate, stomatal conductance and leaf transpiration, respectively. CVC-affected plants showed dynamic

  14. Filamentation and spatiotemporal distribution of extracellular polymeric substances: role on X.fastidiosa single cell adhesion and biofilm formation (Conference Presentation)

    Science.gov (United States)

    Janissen, Richard; Murillo, Duber M.; Niza, Barbara; Sahoo, Prasana K.; Monteiro, Moniellen P.; César, Carlos L.; Carvalho, Hernandes F.; de Souza, Alessandra A.; Cotta, Monica A.

    2016-04-01

    Biofilms can be defined as a community of microorganisms attached to a surface, living embedded in a self- produced matrix of hydrated extracellular polymeric substances (EPS) which comprises most of the biofilm mass. We have recently used an extensive pool of microscopy techniques (confocal fluorescence, electron and scanning probe microscopies) at the micro and nanoscales in order to create a detailed temporal observation of Xylella fastidiosa biofilm formation, using both wild type strain and Green Fluorescent Protein (GFP)-modified cells of this citrus phytopathogen. We have identified three different EPS compositions, as well as their spatial and temporal distribution from single cell to mature biofilm formation stages. In the initial adhesion stage, soluble-EPS (S-EPS) accumulates at cell polar regions and forms a surface layer which facilitates irreversible cell attachment and cell cluster formation. These small clusters are subsequently connected by filamentous cells; further S-EPS surface coverage facilitates cell attachment and form filaments, leading to a floating framework of mature biofilms. The important role of EPS in X.fastidiosa biology was further investigated by imunolabelling experiments to detect the distribution of XadA1 adhesin, which is expressed in early stages of biofilm formation and released in outer membrane vesicles. This protein is located mainly in S-EPS covered areas, as well as on the filaments, indicating a molecular pathway to the enhanced cell attachment previously observed. These results suggest that S-EPS may thus represent an important target for disease control, slow plant colonization by the bacteria, keeping the plant more productive in the field.

  15. An EPG waveform library for sharpshooters and preliminary effects of applied voltage on behaviors controlling Xylella fastidiosa inoculation

    Science.gov (United States)

    Electropenetrography (EPG) waveforms represent electrical conductivity of fluids flowing through an insect’s mouthparts. Over the 50 years since its invention, EPG has undergone three major electronic transformations. The newest, third generation of electropenetrograph, the AC-DC EPG monitor, offers...

  16. Nonlinear models for describing the Citrus Variegated Chlorosis in groves of two counties at northwestern Paraná state, Brazil = Modelo não-linear para a curva de progresso de incidência da Clorose Variegada dos Citros (CVC em dois municípios da região noroeste do Estado do Paraná

    Directory of Open Access Journals (Sweden)

    Clara Matiko Ueda

    2010-10-01

    Full Text Available In Brazil, the production of sweet oranges has been threatened by the Citrus Variegated Chlorosis (CVC incited by the gram-negative bacterium Xylella fastidiosa (Wells. Commercial citrus groves in two counties at the Northwestern Paraná state were evaluated to estimate the disease progression by using parameterizations of nonlinear models. Groves of Citrus sinensis Osbeck, variety “Pêra”, “Valência”, “Natal” and “Folha Murcha” had all the plants evaluated for the presence of disease symptoms. Thereafter, different parameterizations of the Logistic and Gompertz models were fitted to these data. The goodness of fit was evaluated by the intrinsic (IN and parameter-effects (PE curvatures of Bates and Watts, the bias of Box and the Hougaard measures of skewness. In Loanda, the best model was the Fermi-Dirac, and in Nova Esperança the data were best fitted to the parameterization L5, which is also a parameterization from the Logistic model.A citricultura é afetada por diversas doenças, dentre as quais a Clorose Variegada dos Citros (CVC causada pela bactéria Xylella fastidiosa (Wells. Para a região noroeste do Estado do Paraná, onde foi avaliada a CVC, propõe-se encontrar modelos não-lineares de curvas de progresso de incidência da CVC que representem o percentual de plantas acometidas pela doença. Para avaliar o comprometimento dos pomares com relação à doença, foram escolhidos pomares comerciais em dois municípios, onde foi determinada a proporção de plantas doentes. Foram selecionados talhões de laranja doce (Citrus sinensis Osbeck nas variedades “Pêra”, “Valência”, “Natal” e “Folha Murcha” e a avaliação de todas as plantas do talhão foi realizada visualmente em relação à presença ou à ausência de sintomas de CVC. Para estimar o modelo que melhor se ajustou aos dados de progresso da proporção da doença em cada talhão, foram considerados modelos não-lineares decrescimento sigmoidal

  17. Estudo fitoquímico de Citrus: resistência a Xylella fastidiosa e interação com Oncometopia facialis.

    OpenAIRE

    Patrícia Verardi Abdelnur

    2006-01-01

    Neste trabalho estão descritas dezesseis substâncias isoladas de C. limonia: xantiletina, seselina, xantoxiletina, trans-kelactona, trans-decursidinol, junosmarina escopoletina, isoescopoletina, demetilsuberosina, xantoarnol, clausarina, limonianina, 5,4-diidróxi-6-(3-metil-2-butenil)-2,2-dimetilpirano(5,6-7,8) flavanona, β-sitosterol, estigmasterol e campesterol; sendo uma destacada, a 5,4- diidróxi-6-(3-metil-2-butenil)-2,2-dimetilpirano(5,6-7,8) flavanona, por ser inédita na lit...

  18. First report of pecan bacterial leaf scorch caused by Xylella fastidiosa in pecan (Carya illinoinensis) in Arizona, New Mexico, California, and Texas

    Science.gov (United States)

    Pecan bacterial leaf scorch (PBLS) is a chronic disease that can cause major yield losses in pecan orchards. In the 2015-16 growing seasons, symptoms consistent with PBLS were observed in commercial pecan cultivars in AZ, NM, CA and TX. Symptoms included tan to light brown necrotic lesions, which ...

  19. Effects of kaolin particle film and imidacloprid on glassy-winged sharpshooter (Homalodisca vitripennis) (Hemiptera: Cicadellidae)populations and the prevention of spread of Xylella fastidiosa in grape

    Science.gov (United States)

    The glassy-winged sharpshooter (GWSS), Homalodisca coagulata (Say), was introduced into California and soon became a major pest of important agronomic, horticultural, landscape, ornamental crops and native trees in California. This pest feeds readily on grape and, in doing so, transmits X. fastidio...

  20. ORF Alignment: NC_004556 [GENIUS II[Archive

    Lifescience Database Archive (English)

    Full Text Available rase ... [Xylella fastidiosa Temecula1] ... Length = 230 ... Query: 10 ... RLHQGDALRLLCGIDSASVDAVITDPP...YCSGAMRMADRFKPTKRKYINNGTKHVAPDFDC 69 ... RLHQGDALRLLCGIDSASVDAVITDPPYCSG...AMRMADRFKPTKRKYINNGTKHVAPDFDC Sbjct: 1 ... RLHQGDALRLLCGIDSASVDAVITDPPYCSGAMRMADRFKPTKRKYINNGTKHVAPDFDC 60 ... Qu

  1. Fatty Acid Methyl Ester (FAME) analyses for characterization and detection of grapevine pathogens

    Science.gov (United States)

    Grapevines can become infected by a variety of devastating pathogens, including the bacterium Xylella fastidiosa and canker fungi. Multiple strains of Xylella fastidiosa exist, each causing different diseases on various hosts. Although sequence-based genotyping can assist in distinguishing these str...

  2. Scapoidus titanus performs a dual-meaning, sharpshooter-style EPG X wave in xylem and phloem: Could S. titanus be a potential vector of Xylella fastidiosa in European vineyards?

    Science.gov (United States)

    In phytophagous hemipterans, salivary sheath-feeding species are described as xylem or phloem sap-ingesting specialists. Because these two food sources have different characteristics, two different feeding tactics are associated with this supposed specialization. Study of feeding behavior is crucial...

  3. Caracterização da comunidade bacteriana endofítica de citros por isolamento, PCR específico e DGGE Characterization of the endophytic bacterial community from citrus by isolation, specific PCR and DGGE

    Directory of Open Access Journals (Sweden)

    Paulo Teixeira Lacava

    2006-04-01

    Full Text Available O objetivo deste trabalho foi caracterizar a comunidade bacteriana endofítica de plantas assintomáticas (escapes e afetadas pela clorose variegada dos citros (CVC por meio de isolamento em meio de cultura, técnica de gradiente desnaturante em gel de eletroforese (DGGE e detecção de Methylobacterium mesophilicum e Xyllela fastidiosa por meio de PCR específico, para estudar esta comunidade e sua relação com a ocorrência da CVC. A análise da comunidade bacteriana via DGGE permitiu a detecção de X. fastidiosa, bem como Klebsiella sp. e Acinetobacter sp. como endófitos de citros. Foram observados também Curtobacterium sp., Pseudomonas sp., Enterobacter sp. e Bacillus spp. Utilizando primers específicos, Methylobacterium mesophilicum e X. fastidiosa também foram observadas, reforçando hipóteses de que estas bactérias podem estar interagindo no interior da planta hospedeira.The aim of this work was to characterize endophytic bacterial community of assintomatic (escape and Citrus Variegated Chlorosis (CVC-affected citrus plants using isolation in culture medium, denaturing gradient gel electrophoresis (DGGE technique and Methylobacterium mesophilicum as well as Xylella fastidiosa specific PCR, allowing to assess this community and its interactions with CVC. The study of bacterial community by DGGE analysis allowed the detection of X. fastidiosa, as well as Klebsiella sp. e Acinetobacter sp., which were not detected previously. Curtobacterium sp., Pseudomonas sp., Enterobacter sp. and Bacillus spp. were also observed as endophyte in citrus plants. Using specific primers Methylobacterium mesophilicum and X. fastidiosa were observed, reinforcing that these bacteria could interact inside the host plant.

  4. Fulltext PDF

    Indian Academy of Sciences (India)

    PRAKASH KUMAR

    Since DNA encrypts the genetic code it cannot be a random ... strains of bacteria such as the plant pathogen Xylella fastidiosa, marine Cyanobacteria Prochlorococcus marinus or ..... This holds even for the highly virulent strain Y. pestis which.

  5. Espécies de cigarrinhas (Hemiptera, Cicadellidae, Cicadellinae em pomares de citros da região de Bebedouro, SP, com descrição de uma espécie nova de Acrogonia Stål

    Directory of Open Access Journals (Sweden)

    Rosangela Cristina Marucci

    2002-01-01

    Full Text Available Sharpshooter leafhoppers (Hemiptera, Cicadellidae, Cicadellinae in citrus groves of the area of Bebedouro, SP, with description of a new species of Acrogonia Stål. Sixteen species of Cicadellidae, subfamily Cicadellinae, were collected and identified: 11 of the tribe Cicadellini and five of the tribe Proconiini. The ten most common species, together with a new species, are characterized based on external morphology and male genitalia: Acrogonia citrina Marucci & Cavichioli sp. nov.; Bucephalogonia xanthophis (Berg, 1879; Dilobopterus costalimai Young, 1977; Ferrariana trivittata (Signoret, 1854; Hortensia similis (Walker, 1851; Macugonalia cavifrons (Stål, 1862; M. leucomelas (Walker, 1851; Oncometopia facialis (Signoret, 1854; Plesiommata corniculata Young, 1977 and P. mollicella (Fowler, 1900. The leafhopper samples were collected in citrus groves and in adjacent habitats (woods, swamp, cultivable plain, spontaneous vegetation of the grove, coffee crop and sugarcane at the counties of Bebedouro and Monte Azul Paulista and in the district of Turvínia. The collecting methods were: insect net, yellow sticky cards, portable suction trap and Malaise trap. Identification keys were elaborated for the 16 species. There is a great diversity of Cicadellinae species in the citric groves. In the three areas sampled, it was collected a larger number of species of the tribe Cicadellini than of the tribe Proconiini. In the Bebedouro area, the vector species of Xylella fastidiosa, A. citrina sp. nov., B. xanthophis, D. costalimai and O. facialis, were collected in the citrus groves and in all adjacent habitats.

  6. Rootstock effects on almond leaf scorch disease incidence and severity

    Science.gov (United States)

    A five-year field study was conducted to evaluate effects of duration and exclusion of Xylella fastidiosa infections on young almond tree performance and their links to tree vigor. ‘Nemaguard’, ‘Okinawa’, ‘Nonpareil’, and Y119 were used as rootstocks for almond scion ‘Sonora’. Among X.fastidiosa-inf...

  7. Plant water stress effects on the net dispersal rate of the insect vector Homalodisca vitripennis (Germar) (Hemiptera: Cicadellidae) and movement of its egg parasitoid, Gonatocerus ashmeadi Girault (Hymenoptera: Mymaridae)

    Science.gov (United States)

    Homalodisca vitripennis, one of the main vectors of Xylella fastidiosa, is associated with citrus plantings in California, USA. Infested citrus orchards act as a source of vectors to adjacent vineyards where X. fastidiosa causes Pierce’s disease (PD). An analysis of the pattern and rate of movement ...

  8. Evaluation of <em>HER2em> Gene Amplification in Breast Cancer Using Nuclei Microarray <em>in em>S>itu em>Hybridization

    Directory of Open Access Journals (Sweden)

    Xuefeng Zhang

    2012-05-01

    Full Text Available Fluorescence<em> em>>in situ em>hybridization (FISH assay is considered the “gold standard” in evaluating <em>HER2/neu (HER2em> gene status. However, FISH detection is costly and time consuming. Thus, we established nuclei microarray with extracted intact nuclei from paraffin embedded breast cancer tissues for FISH detection. The nuclei microarray FISH (NMFISH technology serves as a useful platform for analyzing <em>HER2em> gene/chromosome 17 centromere ratio. We examined <em>HER2em> gene status in 152 cases of invasive ductal carcinomas of the breast that were resected surgically with FISH and NMFISH. <em>HER2em> gene amplification status was classified according to the guidelines of the American Society of Clinical Oncology and College of American Pathologists (ASCO/CAP. Comparison of the cut-off values for <em>HER2em>/chromosome 17 centromere copy number ratio obtained by NMFISH and FISH showed that there was almost perfect agreement between the two methods (κ coefficient 0.920. The results of the two methods were almost consistent for the evaluation of <em>HER2em> gene counts. The present study proved that NMFISH is comparable with FISH for evaluating <em>HER2em> gene status. The use of nuclei microarray technology is highly efficient, time and reagent conserving and inexpensive.

  9. A New Natural Lactone from <em>Dimocarpus> <em>longan> Lour. Seeds

    Directory of Open Access Journals (Sweden)

    Zhongjun Li

    2012-08-01

    Full Text Available A new natural product named longanlactone was isolated from <em>Dimocarpus> <em>longan> Lour. seeds. Its structure was determined as 3-(2-acetyl-1<em>H>-pyrrol-1-yl-5-(prop-2-yn-1-yldihydrofuran-2(3H-one by spectroscopic methods and HRESIMS.

  10. Reference Gene Selection in the Desert Plant <em>Eremosparton songoricuem>m>

    Directory of Open Access Journals (Sweden)

    Dao-Yuan Zhang

    2012-06-01

    Full Text Available <em>Eremosparton songoricum em>(Litv. Vass. (<em>E. songoricumem> is a rare and extremely drought-tolerant desert plant that holds promise as a model organism for the identification of genes associated with water deficit stress. Here, we cloned and evaluated the expression of eight candidate reference genes using quantitative real-time reverse transcriptase polymerase chain reactions. The expression of these candidate reference genes was analyzed in a diverse set of 20 samples including various <em>E. songoricumem> plant tissues exposed to multiple environmental stresses. GeNorm analysis indicated that expression stability varied between the reference genes in the different experimental conditions, but the two most stable reference genes were sufficient for normalization in most conditions.<em> EsEFem> and <em>Esα-TUB> were sufficient for various stress conditions, <em>EsEF> and <em>EsACT> were suitable for samples of differing germination stages, and <em>EsGAPDH>and <em>Es>UBQ em>were most stable across multiple adult tissue samples. The <em>Es18Sem> gene was unsuitable as a reference gene in our analysis. In addition, the expression level of the drought-stress related transcription factor <em>EsDREB2em>> em>verified the utility of<em> E. songoricumem> reference genes and indicated that no single gene was adequate for normalization on its own. This is the first systematic report on the selection of reference genes in <em>E. songoricumem>, and these data will facilitate future work on gene expression in this species.

  11. Synthesis, Crystal Structure and Luminescent Property of Cd (II Complex with <em>N-Benzenesulphonyl-L>-leucine

    Directory of Open Access Journals (Sweden)

    Xishi Tai

    2012-09-01

    Full Text Available A new trinuclear Cd (II complex [Cd3(L6(2,2-bipyridine3] [L =<em> Nem>-phenylsulfonyl-L>-leucinato] has been synthesized and characterized by elemental analysis, IR and X-ray single crystal diffraction analysis. The results show that the complex belongs to the orthorhombic, space group<em> Pem>212121 with<em> aem> = 16.877(3 Å, <em>b> em>= 22.875(5 Å, <em>c em>= 29.495(6 Å, <em>α> em>= <emem>= <emem>= 90°, <em>V> em>= 11387(4 Å3, <em>Z> em>= 4, <em>Dc>= 1.416 μg·m−3, <emem>= 0.737 mm−1, <em>F> em>(000 = 4992, and final <em>R>1 = 0.0390, <em>ωR>2 = 0.0989. The complex comprises two seven-coordinated Cd (II atoms, with a N2O5 distorted pengonal bipyramidal coordination environment and a six-coordinated Cd (II atom, with a N2O4 distorted octahedral coordination environment. The molecules form one dimensional chain structure by the interaction of bridged carboxylato groups, hydrogen bonds and p-p interaction of 2,2-bipyridine. The luminescent properties of the Cd (II complex and <em>N-Benzenesulphonyl-L>-leucine in solid and in CH3OH solution also have been investigated.

  12. Neonatal Phosphate Nutrition Alters <em>in em>Vivo> and <em>in em>Vitro> Satellite Cell Activity in Pigs

    Directory of Open Access Journals (Sweden)

    Chad H. Stahl

    2012-05-01

    Full Text Available Satellite cell activity is necessary for postnatal skeletal muscle growth. Severe phosphate (PO4 deficiency can alter satellite cell activity, however the role of neonatal PO4 nutrition on satellite cell biology remains obscure. Twenty-one piglets (1 day of age, 1.8 ± 0.2 kg BW were pair-fed liquid diets that were either PO4 adequate (0.9% total P, supra-adequate (1.2% total P in PO4 requirement or deficient (0.7% total P in PO4 content for 12 days. Body weight was recorded daily and blood samples collected every 6 days. At day 12, pigs were orally dosed with BrdU and 12 h later, satellite cells were isolated. Satellite cells were also cultured <em>in vitroem> for 7 days to determine if PO4 nutrition alters their ability to proceed through their myogenic lineage. Dietary PO4 deficiency resulted in reduced (<em>P> < 0.05 sera PO4 and parathyroid hormone (PTH concentrations, while supra-adequate dietary PO4 improved (<em>P> < 0.05 feed conversion efficiency as compared to the PO4 adequate group. <em>In vivoem> satellite cell proliferation was reduced (<em>P> < 0.05 among the PO4 deficient pigs, and these cells had altered <em>in vitroem> expression of markers of myogenic progression. Further work to better understand early nutritional programming of satellite cells and the potential benefits of emphasizing early PO4 nutrition for future lean growth potential is warranted.

  13. Constituents from <em>Vigna em>vexillata> and Their Anti-Inflammatory Activity

    Directory of Open Access Journals (Sweden)

    Guo-Feng Chen

    2012-08-01

    Full Text Available The seeds of <em>Vigna em>genus are important food resources and there have already been many reports regarding their bioactivities. In our preliminary bioassay, the chloroform layer of methanol extracts of<em> V. vexillata em>demonstrated significant anti-inflammatory bioactivity. Therefore, the present research is aimed to purify and identify the anti-inflammatory principles of <em>V. vexillataem>. One new sterol (1 and two new isoflavones (2,3 were reported from the natural sources for the first time and their chemical structures were determined by the spectroscopic and mass spectrometric analyses. In addition, 37 known compounds were identified by comparison of their physical and spectroscopic data with those reported in the literature. Among the isolates, daidzein (23, abscisic acid (25, and quercetin (40 displayed the most significant inhibition of superoxide anion generation and elastase release.

  14. EPG waveforms of blue green sharpshooter: impedance and voltage level effects on stylet probing

    Science.gov (United States)

    Blue-green sharpshooter (BGSS), Graphocephala atropunctata, is a native California vector of Xylella fastidiosa (Xf) a foregut-borne bacterium that is the causal agent of Pierce’s disease in grapevines. A 3rd-generation, AC-DC electropenetrograph (EPG) was used to record probing behaviors of adult B...

  15. Bacterial Leaf Scorch of Amenity Trees a Wide-Spread Problem of Economic Significance to the Urban Forest

    Science.gov (United States)

    James Lashomb; Alan Iskra; Ann Brooks Gould; George Hamilton

    2003-01-01

    Bacterial leaf scorch (BLS) of amenity trees is caused by the bacterium Xylella fastidiosa, a xylem-limited pathogen that causes water stress resulting in leaf scorch, decline, and eventual death of affected trees. Recent surveys indicate that BLS is widespread throughout the eastern half of the United States. In New Jersey, BLS primarily affects red and pin oaks...

  16. Modeling deployment of Pierce’s disease resistant grapevines

    Science.gov (United States)

    Deployment of Pierce’s disease resistant grapevines is a key solution to mitigating economic losses caused by Xylella fastidiosa. While Pierce’s disease resistant grapevines under development display mild symptoms and have lower bacterial populations than susceptible varieties, all appear to remain ...

  17. Design of a candidate vibrational signal for mating disruption against the glassy-winged sharpshooter, Homalodisca Vitripennis

    Science.gov (United States)

    The glassy-winged sharpshooter (GWSS), Homalodisca vitripennis, is an important pest of grapevines due to its ability to transmit Xylella fastidiosa, the causal agent of Pierce’s disease. GWSS mating communication is based on vibrational signals; therefore, vibrational mating disruption could be an ...

  18. Playback interference of glassy-winged sharp shooter communication

    Science.gov (United States)

    Animal communication is vital to reproduction, particularly for securing a mate. Insects commonly communicate by exchanging vibrational signals that are transmitted through host plants. The glassy-winged sharpshooter (GWSS), Homalodisca vitripennis, is an important vector of Xylella fastidiosa, a pl...

  19. Effects of nymphal diet and adult feeding on allocation of resources to glassy-winged sharpshooter egg production

    Science.gov (United States)

    The glassy-winged sharpshooter is an invasive insect capable of transmitting the bacterial pathogen Xylella fastidiosa. Pre-oviposition periods of laboratory reared glassy-winged sharpshooters are variable. Here, two questions were addressed: does nymphal diet affect pre-oviposition period and how d...

  20. EXPERIMENTAL ANALYSIS OF BIOLOGICAL PARAMETERS AND VECTOR ABILITY OF GLASSY-WINGED SHARPSHOOTERS FROM ALLOPATRIC POPULATIONS IN CALIFORNIA

    Science.gov (United States)

    The glassy-winged sharpshooter (GWSS), Homalodisca vitripennis (Germar), is native to the southeastern United States and northeastern Mexico. It was detected in southern California in the late 1980s and in the San Joaquin Valley in 1999, where it transmits the bacterium Xylella fastidiosa to grapev...

  1. Glassy-winged sharpshooters (GWSS) are not GWSS: Differential Reproductive Maturity between Allopatric Californian Populations

    Science.gov (United States)

    Homalodisca vitripennis (Germar) is native to southeastern U.S. and northeastern Mexico. It was detected in southern California in the late 1980s and in the San Joaquin Valley in 1999, where it transmits the bacterium Xylella fastidiosa Wells et al. to grapevines and other crops. The reproductive ...

  2. Differential reproductive maturity between allopatric populations of Homalodisca vitripennis (Hemiptera: Cicadellidae) in California

    Science.gov (United States)

    Homalodisca vitripennis (Germar) is native to southeastern U.S. and northeastern Mexico. It was detected in southern California in the late 1980s and in the San Joaquin Valley in 1999, where it transmits the bacterium Xylella fastidiosa Wells et al. to grapevines and other crops. The reproductive ...

  3. Mating disruption of Homalodisca vitripennis (Germar) (Hemiptera: Cicadellidae) by playback of vibrational signals in vineyard trellis

    Science.gov (United States)

    BACKGROUND: Glassy-winged sharpshooter (GWSS), Homalodisca vitripennis (Germar) (Hemiptera: Cicadellidae), is an important vector of the bacterium Xylella fastidiosa, the causal agent of Pierce’s disease of grapevine. Area-wide applications of neonicotinoid insecticides have suppressed GWSS populati...

  4. Playback of natural vibrational signals in vineyard trellis for mating disruption of glassy-winged sharpshooter

    Science.gov (United States)

    The glassy-winged sharpshooter, Homalodisca vitripennis (Germar) (Hemiptera: Cicadellidae), is a vector of Xylella fastidiosa, an important bacterial pathogen of several crops in the Americas and Europe. Mating communication of this and many other cicadellid pests involves the exchange of substrate-...

  5. Potential Military Chemical/Biological Agents and Compounds

    Science.gov (United States)

    2005-01-01

    mycoides Xylella fastidiosa Other Animal Diseases/Pathogens of Concern Fungi Contagious bovine pleuropneumonia Deutrrerophoma tracheiphila Contagious...of the disease, which include lameness, low milk production, weight loss, mastitis , debilitation, and abortion. FMD occurs in many major livestock...man. o. Contagious Bovine Pleuropneumonia (CBPP).15 J-5 (1) CBPP is caused by Mycoplasma mycoides and primarily affects cattle. The principal

  6. Ecology, Diversity and Comparative Genomics of Oceanic Cyanobacterial Viruses

    Science.gov (United States)

    2004-06-01

    A. P., Terenzi, M. F., Truffi, D., Tsai, S. M., Tsuhako, M. H., Vallada, H., Van Sluys, M. A., Verjovski-Almeida, S., Vettore, A. L., Zago , M. A...L., Zago , M. A., Zatz, M., Meidanis, J., and Setubal, J. C. (2000). The genome sequence of the plant pathogen Xylella fastidiosa. The Xylelia

  7. EPG waveform library for Graphocephala atropunctata (Hemiptera: Cicadellidae): Effect of input resistor and voltage levels on waveform appearance and probing behaviors

    Science.gov (United States)

    Graphocephala atropunctata is a vector of Xylella fastidiosa (Xf), the causal agent of Pierce’s disease of grapevines. A 3rd-generation, AC-DC electropenetrograph (EPG) was used to record stylet probing and ingestion behaviors of adult G. atropunctata on healthy grapevines. This study presents a com...

  8. Propagation of Homalodisca Coagulata Virus-01 via Homalodisca Vitripennis cell culture

    Science.gov (United States)

    The glassy-winged sharpshooter (Homalodisca vitripennis) is a highly vagile and polyphagous insect found throughout the southwestern United States. These insects are the predominant vectors of Xylella fastidiosa, a xylem-limited bacterium that is the causal agent of Pierce's disease (PD) of grapevin...

  9. Characterization of the marine propionate-degrading, sulfate-reducing bacterium Desulfofaba fastidiosa sp. nov. and reclassification of Desulfomusa hansenii as Desulfofaba hansenii comb. nov.

    Science.gov (United States)

    Abildgaard, Lone; Ramsing, Niels Birger; Finster, Kai

    2004-03-01

    A rod-shaped, slightly curved sulfate reducer, designated strain P2(T), was isolated from the sulfate-methane transition zone of a marine sediment. Cells were motile by means of a single polar flagellum. The strain reduced sulfate, thiosulfate and sulfite to sulfide and used propionate, lactate and 1-propanol as electron donors. Strain P2(T) also grew by fermentation of lactate. Propionate was oxidized incompletely to acetate and CO(2). The DNA G+C content was 48.8 mol%. Sequence analysis of the small-subunit rDNA and the dissimilatory sulfite reductase gene revealed that strain P2(T) was related to the genera Desulfonema, Desulfococcus, Desulfosarcina, 'Desulfobotulus', Desulfofaba, Desulfomusa and Desulfofrigus. These genera include incomplete as well as complete oxidizers of substrates. Strain P2(T) shared important morphological and physiological traits with Desulfofaba gelida and Desulfomusa hansenii, including the ability to oxidize propionate incompletely to acetate. The 16S rRNA gene similarities of P2(T) to Desulfofaba gelida and Desulfomusa hansenii were respectively 92.9 and 91.5 %. Combining phenotypic and genotypic traits, we propose strain P2(T) to be a member of the genus Desulfofaba. The name Desulfofaba fastidiosa sp. nov. (type strain P2(T)=DSM 15249(T)=ATCC BAA-815(T)) is proposed, reflecting the limited number of substrates consumed by the strain. In addition, the reclassification of Desulfomusa hansenii as a member of the genus Desulfofaba, Desulfofaba hansenii comb. nov., is proposed. A common line of descent and a number of shared phenotypic traits support this reclassification.

  10. Momentos em freios e em embraiagens

    OpenAIRE

    Mimoso, Rui Miguel Pereira

    2011-01-01

    Dissertação para obtenção do Grau de Mestre em Mestrado Integrado em Engenharia Mecânica Nesta dissertação reúnem-se os modelos de cálculo utilizados na determinação dos momentos em freios e em embraiagens. Neste trabalho consideram-se os casos de freios e embraiagens de atrito seco e atrito viscoso. Nos freios de atrito viscoso são considerados casos em que as características dos fluidos não são induzidas, e outros em que são induzidas modificações a essas mesmas características. São a...

  11. Dermatoses em renais cronicos em terapia dialitica

    Directory of Open Access Journals (Sweden)

    Luis Alberto Batista Peres

    2014-03-01

    Full Text Available Objetivo: As desordens cutâneas e das mucosas são comuns em pacientes em hemodiálise a longo prazo. A diálise prolonga a expectativa de vida, dando tempo para a manifestação destas anormalidades. Os objetivos deste estudo foram avaliar a prevalência de problemas dermatológicos em pacientes com doença renal crônica (DRC em hemodiálise. Métodos: Cento e quarenta e cinco pacientes com doença renal crônica em hemodiálise foram estudados. Todos os pacientes foram completamente analisados para as alterações cutâneas, de cabelos, mucosas e unhas por um único examinador e foram coletados dados de exames laboratoriais. Os dados foram armazenados em um banco de dados do Microsolft Excel e analisados por estatística descritiva. As variáveis contínuas foram comparadas pelo teste t de Student e as variáveis categóricas utilizando o teste do qui-quadrado ou o teste Exato de Fischer, conforme adequado. Resultados: O estudo incluiu 145 pacientes, com idade média de 53,6 ± 14,7 anos, predominantemente do sexo masculino (64,1% e caucasianos (90,0%. O tempo médio de diálise foi de 43,3 ± 42,3 meses. As principais doenças subjacentes foram: hipertensão arterial em 33,8%, diabetes mellitus em 29,6% e glomerulonefrite crônica em 13,1%. As principais manifestações dermatológicas observadas foram: xerose em 109 (75,2%, equimose em 87 (60,0%, prurido em 78 (53,8% e lentigo em 33 (22,8% pacientes. Conclusão: O nosso estudo mostrou a presença de mais do que uma dermatose por paciente. As alterações cutâneas são frequentes em pacientes em diálise. Mais estudos são necessários para melhor caracterização e manejo destas dermatoses.

  12. Bioassay assessment of metarhizium anisopliae (metchnikoff sorokin (deuteromycota: hyphomycetes against Oncometopia facialis (signoret (hemiptera: cicadellidae Avaliação do bioensaio de Metarhizium anisopliae (metchnikoff sorokin (deuteromycota: hyphomycetes contra Oncometopia facialis (signoret (hemiptera: cicadellidae

    Directory of Open Access Journals (Sweden)

    Wolney Dalla Pria Júnior

    2008-03-01

    Full Text Available Citrus Variegated Chlorosis (CVC is an economically important, destructive disease in Brazil and is caused by Xylella fastidiosa and transmitted by sharpshooter insects. In this study, the efficacy of the fungus Metarhizium anisopliae in controlling the sharpshooter Oncometopia facialis was studied by bioassay conditions. In the bioassay, insects were sprayed with a suspension containing 5 X 10(7 conidia mL-1. Adults captured in the field were treated in groups of 10 in a total of 11 replications per treatment. Significant differences between the natural mortality and the mortality of insects treated with the fungus were observed 6 days after inoculations (PA Clorose Variegada dos Citros (CVC é uma doença economicamente importante e destrutiva no Brasil e é causada pela bactéria Xylella fastidiosa e transmitida por insetos vetores tal como Oncometopia facialis. Nesse estudo, a eficácia do fungo Metarhizium anisopliae em controlar o inseto vetor O. facialis foi estudada em condições de bioensaio. Nesse bioensaio, insetos foram pulverizados com uma suspensão de 5 X 10(7 conídio mL-1. Insetos-adultos capturados no campo foram tratados em grupos de 10, em um total de 11 replicatas por tratamento. Diferenças significativas entre a mortalidade natural e a mortalidade dos insetos tratados com o fungo foram observadas em 6 dias após a inoculação (P<0.05. Estas diferenças significativas aumentaram antes do décimo dia após o tratamento. O fungo causou uma mortalidade de 87,1%, com uma LT50 variando entre 5 e 6 dias. A LC50 foi de 1,2 X 10(6 conídio mL-1, variando de 7,7 X 10(5 a 2 X 10(6 conídio mL-1. Estes resultados mostraram que o vetor O. facialis foi susceptível a ação entomopatogênica de M. anisopliae em condições controladas durante o bioensaio.

  13. Population genomic analysis of a bacterial plant pathogen: novel insight into the origin of Pierce's disease of grapevine in the U.S.

    Directory of Open Access Journals (Sweden)

    Leonard Nunney

    Full Text Available Invasive diseases present an increasing problem worldwide; however, genomic techniques are now available to investigate the timing and geographical origin of such introductions. We employed genomic techniques to demonstrate that the bacterial pathogen causing Pierce's disease of grapevine (PD is not native to the US as previously assumed, but descended from a single genotype introduced from Central America. PD has posed a serious threat to the US wine industry ever since its first outbreak in Anaheim, California in the 1880s and continues to inhibit grape cultivation in a large area of the country. It is caused by infection of xylem vessels by the bacterium Xylella fastidiosa subsp. fastidiosa, a genetically distinct subspecies at least 15,000 years old. We present five independent kinds of evidence that strongly support our invasion hypothesis: 1 a genome-wide lack of genetic variability in X. fastidiosa subsp. fastidiosa found in the US, consistent with a recent common ancestor; 2 evidence for historical allopatry of the North American subspecies X. fastidiosa subsp. multiplex and X. fastidiosa subsp. fastidiosa; 3 evidence that X. fastidiosa subsp. fastidiosa evolved in a more tropical climate than X. fastidiosa subsp. multiplex; 4 much greater genetic variability in the proposed source population in Central America, variation within which the US genotypes are phylogenetically nested; and 5 the circumstantial evidence of importation of known hosts (coffee plants from Central America directly into southern California just prior to the first known outbreak of the disease. The lack of genetic variation in X. fastidiosa subsp. fastidiosa in the US suggests that preventing additional introductions is important since new genetic variation may undermine PD control measures, or may lead to infection of other crop plants through the creation of novel genotypes via inter-subspecific recombination. In general, geographically mixing of previously

  14. EM International. Volume 1

    Energy Technology Data Exchange (ETDEWEB)

    1993-07-01

    It is the intent of EM International to describe the Office of Environmental Restoration and Waste Management`s (EM`s) various roles and responsibilities within the international community. Cooperative agreements and programs, descriptions of projects and technologies, and synopses of visits to international sites are all highlighted in this semiannual journal. Focus on EM programs in this issue is on international collaboration in vitrification projects. Technology highlights covers: in situ sealing for contaminated sites; and remote sensors for toxic pollutants. Section on profiles of countries includes: Arctic contamination by the former Soviet Union, and EM activities with Germany--cooperative arrangements.

  15. Microsatellite Loci in the Gypsophyte <em>Lepidium subulatum em>(Brassicaceae, and Transferability to Other <em>Lepidieae>

    Directory of Open Access Journals (Sweden)

    José Gabriel Segarra-Moragues

    2012-09-01

    Full Text Available Polymorphic microsatellite markers were developed for the Ibero-North African, strict gypsophyte <em>Lepidium subulatumem> to unravel the effects of habitat fragmentation in levels of genetic diversity, genetic structure and gene flow among its populations. Using 454 pyrosequencing 12 microsatellite loci including di- and tri-nucleotide repeats were characterized in <em>L. subulatumem>. They amplified a total of 80 alleles (2–12 alleles per locus in a sample of 35 individuals of <em>L. subulatumem>, showing relatively high levels of genetic diversity, <em>H>O = 0.645, <em>H>E = 0.627. Cross-species transferability of all 12 loci was successful for the Iberian endemics <em>Lepidium cardaminesem>, <em>Lepidium stylatumem>, and the widespread, <em>Lepidium graminifoliumem> and one species each of two related genera, <em>Cardaria drabaem> and <em>Coronopus didymusem>. These microsatellite primers will be useful to investigate genetic diversity, population structure and to address conservation genetics in species of <em>Lepidium>.

  16. Rooting of healthy and CVC-affected 'Valência' sweet orange stem cuttings, through the use of plant regulators

    Directory of Open Access Journals (Sweden)

    Gustavo Habermann

    2006-01-01

    Full Text Available Citrus variegated chlorosis (CVC is a disease caused by Xylella fastidiosa. Using different concentrations of plant regulators, such as auxins (indole-3-butyric acid and gibberellic acid biosynthesis-inhibitor (paclobutrazol, physiological rooting capacity of healthy and CVC-affected stem cuttings were evaluated in order to investigate the importance of plant hormone imbalance and xylem occlusion in plants with CVC. The percentages of dead, alive and rooted cuttings, cuttings with callus and mean number of roots per cuttings did not show statistical differences in response to the distinct concentrations of synthetic plant regulators. There were differences only between healthy and CVC-affected cuttings. This showed the importance of xylem occlusion and diffusive disturbances in diseased plants, in relation to root initiation capacity and hormonal translocation in the plant tissue.Clorose variegada dos citros (CVC é uma doença causada por Xylella fastidiosa, podendo determinar oclusão do xilema e desbalanço hormonal, o que por fim está relacionado ao processo de iniciação radicial em estacas. Usando diferentes concentrações de fitorreguladores, como auxinas (ácido 3-indol butírico e inibidores da biossíntese de ácido giberélico (paclobutrazol, que são promotores do enraizamento de estacas, verificou-se a capacidade fisiológica de enraizamento de estacas sadias e com CVC, a fim de investigar a importância do desbalanço hormonal e oclusão do xilema em plantas doentes. As porcentagens de estacas mortas, vivas, enraizadas e com calo e o número médio de raízes por estaca não mostraram diferenças estatísticas em resposta às diferentes concentrações dos reguladores vegetais sintéticos. Houve diferenças apenas entre estacas sadias e doentes. Isto aponta a importância da oclusão do xilema e distúrbios difusivos em plantas doentes, em relação à capacidade de iniciação radicial e à translocação hormonal no tecido

  17. Marketing em moda

    OpenAIRE

    Leães, Sabrina Durgante

    2008-01-01

    Dissertação de mestrado em Design e Marketing O actual estado do Marketing em Moda é uma das questões ainda complexa com que se debate a sociedade global. As questões do Marketing em Moda percorrem alguns aspectos fundamentais tais como as constantes mutações do meio envolvente, a forma de como é percebida e comunicada a identidade das marcas de moda, em busca da melhor forma de segmentar o mercado e definir o seu posicionamento, bem como a reacção ao produto de moda do consumidor final. ...

  18. Emergency Medical Service (EMS) Stations

    Data.gov (United States)

    Kansas Data Access and Support Center — EMS Locations in Kansas The EMS stations dataset consists of any location where emergency medical services (EMS) personnel are stationed or based out of, or where...

  19. <em>Angiostrongylus vasorumem> in red foxes (<em>Vulpes vulpesem> and badgers (<em>Meles melesem> from Central and Northern Italy

    Directory of Open Access Journals (Sweden)

    Marta Magi

    2010-06-01

    Full Text Available Abstract During 2004-2005 and 2007-2008, 189 foxes (<em>Vulpes vulpesem> and 6 badgers (<em>Meles melesem> were collected in different areas of Central Northern Italy (Piedmont, Liguria and Tuscany and examined for <em>Angiostrongylus vasorumem> infection. The prevalence of the infection was significantly different in the areas considered, with the highest values in the district of Imperia (80%, Liguria and in Montezemolo (70%, southern Piedmont; the prevalence in Tuscany was 7%. One badger collected in the area of Imperia turned out to be infected, representing the first report of the parasite in this species in Italy. Further studies are needed to evaluate the role played by fox populations as reservoirs of infection and the probability of its spreading to domestic dogs.
    Riassunto <em>Angiostrongylus vasorumem> nella volpe (<em>Vulpes vulpesem> e nel tasso (<em>Meles melesem> in Italia centro-settentrionale. Nel 2004-2005 e 2007-2008, 189 volpi (<em>Vulpes vulpesem> e 6 tassi (<em>Meles melesem> provenienti da differenti aree dell'Italia settentrionale e centrale (Piemonte, Liguria Toscana, sono stati esaminati per la ricerca di <em>Angiostrongylus vasorumem>. La prevalenza del nematode è risultata significativamente diversa nelle varie zone, con valori elevati nelle zone di Imperia (80% e di Montezemolo (70%, provincia di Cuneo; la prevalenza in Toscana è risultata del 7%. Un tasso proveniente dall'area di Imperia è risultato positivo per A. vasorum; questa è la prima segnalazione del parassita in tale specie in Italia. Ulteriori studi sono necessari per valutare il potenziale della volpe come serbatoio e la possibilità di diffusione della parassitosi ai cani domestici.

    doi:10.4404/hystrix-20.2-4442

  20. International EMS Systems

    DEFF Research Database (Denmark)

    Langhelle, Audun; Lossius, Hans Morten; Silfvast, Tom

    2004-01-01

    exist, however, especially within the ground and air ambulance service, and the EMS systems face several challenges. Main problems and challenges emphasized by the authors are: (1) Denmark: the dispatch centres are presently not under medical control and are without a national criteria based system......Emergency medicine service (EMS) systems in the five Nordic countries have more similarities than differences. One similarity is the involvement of anaesthesiologists as pre-hospital physicians and their strong participation for all critically ill and injured patients in-hospital. Discrepancies do....... Access to on-line medical advice of a physician is not available; (2) Finland: the autonomy of the individual municipalities and their responsibility to cover for primary and specialised health care, as well as the EMS, and the lack of supporting or demanding legislation regarding the EMS; (3) Iceland...

  1. A poliomielite em Sergipe

    Directory of Open Access Journals (Sweden)

    Hélio A. Oliveira

    1994-06-01

    Full Text Available Os autores apresentam estudo retrospectivo da poliomielite em Sergipe. Dividem o estudo em três períodos levando em consideração a taxa de notificação de casos positivos e enfatizam o período que vai de 1979 a 1989, pelo estudo de 159 casos consecutivos. As seguintes informações foram levantadas para cada caso: sexo, idade, procedência, estado vacinai anterior e evolução clínica. São também avaliadas a incidência anual, relação incidência/cobertura vacinai e distribuição geográfica. Tecem comentários sobre os surtos epidêmicos ocorridos em 1984 e 1986, sobre as alterações na circulação do poliovírus selvagem (P1 para P3 e sobre problemas relacionados à não aquisição de imunidade em crianças com esquema vacinal completo. Consideram a poliomielite controlada no Estado, mas enfatizam a necessidade de manutenção de Vigilância epidemiológica efetiva para todos os casos de paralisia aguda e flácida em menores de 14 anos de idade.

  2. EM type radioactive standards. Radioaktivni etalony EM

    Energy Technology Data Exchange (ETDEWEB)

    1981-01-01

    The standard contains technical specifications and conditions of production, testing, packing, transport and storage of EM type planar calibration standards containing radionuclides /sup 14/C, /sup 60/Co, /sup 90/Sr, /sup 137/Cs, /sup 147/Pm, /sup 204/Tl, /sup 239/Pu, /sup 241/Am and natural U. The terminology is explained, the related Czechoslovak standards and legal prescriptions given and amendments to these prescriptions presented.

  3. Purification, Characterization and Antioxidant Activities <em>in Vitroem>> em>and <em>in Vivoem> of the Polysaccharides from <em>Boletus edulisem> Bull

    Directory of Open Access Journals (Sweden)

    Yijun Fan

    2012-07-01

    Full Text Available A water-soluble polysaccharide (BEBP was extracted from <em>Boletus edulis em>Bull using hot water extraction followed by ethanol precipitation. The polysaccharide BEBP was further purified by chromatography on a DEAE-cellulose column, giving three major polysaccharide fractions termed BEBP-1, BEBP-2 and BEBP-3. In the next experiment, the average molecular weight (Mw, IR and monosaccharide compositional analysis of the three polysaccharide fractions were determined. The evaluation of antioxidant activities both <em>in vitroem> and <em>in vivo em>suggested that BEBP-3 had good potential antioxidant activity, and should be explored as a novel potential antioxidant.

  4. Sulla presenza di <em>Sorex antinoriiem>, <em>Neomys anomalusem> (Insectivora, Soricidae e <em>Talpa caecaem> (Insectivora, Talpidae in Umbria

    Directory of Open Access Journals (Sweden)

    A.M. Paci

    2003-10-01

    Full Text Available Lo scopo del contributo è di fornire un aggiornamento sulla presenza del Toporagno del Vallese <em>Sorex antinoriiem>, del Toporagno acquatico di Miller <em>Neomys anomalusem> e della Talpa cieca <em>Talpa caecaem> in Umbria, dove le specie risultano accertate ormai da qualche anno. A tal fine sono stati rivisitati i reperti collezionati e la bibliografia conosciuta. Toporagno del Vallese: elevato di recente a livello di specie da Brünner et al. (2002, altrimenti considerato sottospecie del Toporagno comune (<em>S. araneus antinoriiem>. È conservato uno di tre crani incompleti (mancano mandibole ed incisivi superiori al momento prudenzialmente riferiti a <em>Sorex> cfr. <em>antinorii>, provenienti dall?Appennino umbro-marchigiano settentrionale (dintorni di Scalocchio - PG, 590 m. s.l.m. e determinati sulla base della pigmentazione rossa degli ipoconi del M1 e M2; Toporagno acquatico di Miller: tre crani (Breda in Paci e Romano op. cit. e un esemplare intero (Paci, ined. sono stati trovati a pochi chilometri di distanza gli uni dall?altro tra i comuni di Assisi e Valfabbrica, in ambienti mediocollinari limitrofi al Parco Regionale del M.te Subasio (Perugia. In provincia di Terni la specie viene segnalata da Isotti (op. cit. per i dintorni di Orvieto. Talpa cieca: sono noti una femmina e un maschio raccolti nel comune di Pietralunga (PG, rispettivamente in una conifereta a <em>Pinus nigraem> (m. 630 s.l.m. e nelle vicinanze di un bosco misto collinare a prevalenza di <em>Quercus cerrisem> (m. 640 s.l.m.. Recentemente un terzo individuo è stato rinvenuto nel comune di Sigillo (PG, all?interno del Parco Regionale di M.te Cucco, sul margine di una faggeta a 1100 m s.l.m. In entrambi i casi l?areale della specie è risultato parapatrico con quello di <em>Talpa europaeaem>.

  5. Methyl 2-Benzamido-2-(1<em>H>-benzimidazol-1-ylmethoxyacetate

    Directory of Open Access Journals (Sweden)

    Alami Anouar

    2012-09-01

    Full Text Available The heterocyclic carboxylic α-aminoester methyl 2-benzamido-2-(1<em>H>-benzimidazol-1-ylmethoxyacetate is obtained by <em>O>-alkylation of methyl α-azido glycinate <em>N>-benzoylated with 1<em>H>-benzimidazol-1-ylmethanol.

  6. A specimen of <em>Sorex> cfr. <em>samniticus> in Barn Owl's pellets from Murge plateau (Apulia, Italy / Su di un <em>Sorex> cfr. <em>samniticus> (Insectivora, Soricidae rinvenuto in borre di <em>Tyto albaem> delle Murge (Puglia, Italia

    Directory of Open Access Journals (Sweden)

    Giovanni Ferrara

    1992-07-01

    Full Text Available Abstract In a lot of Barn Owl's pellets from the Murge plateau a specimen of <em>Sorex> sp. was detected. Thank to some morphological and morphometrical features, the cranial bones can be tentatively attributed to <em>Sorex samniticusem> Altobello, 1926. The genus <em>Sorex> was not yet included in the Apulia's fauna southwards of the Gargano district; the origin and significance of the above record is briefly discussed, the actual presence of a natural population of <em>Sorex> in the Murge being not yet proved. Riassunto Viene segnalato il rinvenimento di un esemplare di <em>Sorex> cfr. <em>samniticus> da borre di <em>Tyto albaem> delle Murge. Poiché il genere non era stato ancora segnalato nella Puglia a sud del Gargano, viene discusso il significato faunistico del reperto.

  7. Glycosylation of Vanillin and 8-Nordihydrocapsaicin by Cultured <em>Eucalyptus perrinianaem> Cells

    Directory of Open Access Journals (Sweden)

    Naoji Kubota

    2012-05-01

    Full Text Available Glycosylation of vanilloids such as vanillin and 8-nordihydrocapsaicin by cultured plant cells of <em>Eucalyptus perrinianaem> was studied. Vanillin was converted into vanillin 4-<em>O>-b-D-glucopyranoside, vanillyl alcohol, and 4-<em>O>-b-D-glucopyranosylvanillyl alcohol by <em>E. perriniana em>cells. Incubation of cultured <em>E. perrinianaem> cells with 8-nor- dihydrocapsaicin gave 8-nordihydrocapsaicin 4-<em>O>-b-D-glucopyranoside and 8-nordihydro- capsaicin 4-<em>O>-b-D-gentiobioside.

  8. Retratos em movimento.

    Directory of Open Access Journals (Sweden)

    Luiz Carlos Oliveira Junior

    Full Text Available resumo O artigo aborda aspectos da relação do cinema com a arte do retrato. Buscamos, em primeiro lugar, uma definição estética do que seria um retrato cinematográfico, sempre em tensão com os critérios formais e padrões estilísticos que historicamente constituíram o retrato pictórico. Em seguida, relacionamos essa questão com a importância que se deu à representação do close-up de rosto nas primeiras décadas do cinema, quando foi atribuído aos filmes um papel inédito no estudo da fisionomia e da expressão facial. Por fim, apresentamos exemplos de autorretratos na pintura e no cinema para expor a forma como a autorrepresentação põe em crise as noções de subjetividade e identidade em que a definição clássica do retrato se apoiava.

  9. <em>In Vivoem> Histamine Optical Nanosensors

    Directory of Open Access Journals (Sweden)

    Heather A. Clark

    2012-08-01

    Full Text Available In this communication we discuss the development of ionophore based nanosensors for the detection and monitoring of histamine levels <em>in vivoem>. This approach is based on the use of an amine-reactive, broad spectrum ionophore which is capable of recognizing and binding to histamine. We pair this ionophore with our already established nanosensor platform, and demonstrate <em>in vitroem> and <em>in vivoem> monitoring of histamine levels. This approach enables capturing rapid kinetics of histamine after injection, which are more difficult to measure with standard approaches such as blood sampling, especially on small research models. The coupling together of <em>in vivoem> nanosensors with ionophores such as nonactin provide a way to generate nanosensors for novel targets without the difficult process of designing and synthesizing novel ionophores.

  10. Study of the <em>in Vitroem> Antiplasmodial, Antileishmanial and Antitrypanosomal Activities of Medicinal Plants from Saudi Arabia

    Directory of Open Access Journals (Sweden)

    Nawal M. Al-Musayeib

    2012-09-01

    Full Text Available The present study investigated the <em>in vitroem> antiprotozoal activity of sixteen selected medicinal plants. Plant materials were extracted with methanol and screened <em>in vitroem> against erythrocytic schizonts of <em>Plasmodium falciparumem>, intracellular amastigotes of <em>Leishmania infantum em>and <em>Trypanosoma cruzi em>and free trypomastigotes of<em> T. bruceiem>. Cytotoxic activity was determined against MRC-5 cells to assess selectivity<em>. em>The criterion for activity was an IC50 < 10 µg/mL (4. Antiplasmodial activity was found in the<em> em>extracts of<em> em>>Prosopis julifloraem> and <em>Punica granatumem>. Antileishmanial activity<em> em>against <em>L. infantumem> was demonstrated in <em>Caralluma sinaicaem> and <em>Periploca aphylla.em> Amastigotes of<em> T. cruzi em>were affected by the methanol extract of<em> em>>Albizia lebbeckem>> em>pericarp, <em>Caralluma sinaicaem>,> Periploca aphylla em>and <em>Prosopius julifloraem>. Activity against<em> T. brucei em>was obtained in<em> em>>Prosopis julifloraem>. Cytotoxicity (MRC-5 IC50 < 10 µg/mL and hence non-specific activities were observed for<em> em>>Conocarpus lancifoliusem>.>

  11. Cyberbullying em adolescentes brasileiros

    OpenAIRE

    Wendt, Guilherme Welter

    2012-01-01

    O cyberbullying é entendido como uma forma de comportamento agressivo que ocorre através dos meios eletrônicos de interação (computadores, celulares, sites de relacionamento virtual), sendo realizado de maneira intencional por uma pessoa ou grupo contra alguém em situação desigual de poder e, ainda, com dificuldade em se defender. Os estudos disponíveis até o presente momento destacam que o cyberbullying é um fator de risco para o desenvolvimento de sintomas de ansiedade, depressão, ideação s...

  12. Nietzsche em voga

    OpenAIRE

    Borromeu, Carlos

    2015-01-01

    Resumo:Texto publicado em 1941, na revista de orientação católica A Ordem, no Rio de Janeiro. Seu autor considera que Nietzsche teria negado a moral tradicional, concebendo em seu lugar outra, porém imoral e brutal. Acusa o filósofo, por fim, de ser responsável pela Guerra ora e curso na Europa. Abstract:Text published in 1941 in the Catholic orientation magazine, A Ordem, in Rio de Janeiro. The author believes that Nietzsche would have denied traditional morality, conceiving another in it...

  13. Natural Products from Antarctic Colonial Ascidians of the Genera <em>Aplidium> and <em>Synoicum>: Variability and Defensive Role

    Directory of Open Access Journals (Sweden)

    Conxita Avila

    2012-08-01

    Full Text Available Ascidians have developed multiple defensive strategies mostly related to physical, nutritional or chemical properties of the tunic. One of such is chemical defense based on secondary metabolites. We analyzed a series of colonial Antarctic ascidians from deep-water collections belonging to the genera <em>Aplidium> and <em>Synoicum> to evaluate the incidence of organic deterrents and their variability. The ether fractions from 15 samples including specimens of the species <em>A.> <em>falklandicum>, <em>A.> <em>fuegiense>, <em>A.> <em>meridianum>, <em>A.> <em>millari> and <em>S.> <em>adareanum> were subjected to feeding assays towards two relevant sympatric predators: the starfish <em>Odontaster> <em>validus>, and the amphipod <em>Cheirimedon> <em>femoratus>. All samples revealed repellency. Nonetheless, some colonies concentrated defensive chemicals in internal body-regions rather than in the tunic. Four ascidian-derived meroterpenoids, rossinones B and the three derivatives 2,3-epoxy-rossinone B, 3-epi-rossinone B, 5,6-epoxy-rossinone B, and the indole alkaloids meridianins A–G, along with other minoritary meridianin compounds were isolated from several samples. Some purified metabolites were tested in feeding assays exhibiting potent unpalatabilities, thus revealing their role in predation avoidance. Ascidian extracts and purified compound-fractions were further assessed in antibacterial tests against a marine Antarctic bacterium. Only the meridianins showed inhibition activity, demonstrating a multifunctional defensive role. According to their occurrence in nature and within our colonial specimens, the possible origin of both types of metabolites is discussed.

  14. Control of Pierce's Disease by Phage.

    Directory of Open Access Journals (Sweden)

    Mayukh Das

    Full Text Available Pierce's Disease (PD of grapevines, caused by Xylella fastidiosa subsp. fastidiosa (Xf, is a limiting factor in the cultivation of grapevines in the US. There are presently no effective control methods to prevent or treat PD. The therapeutic and prophylactic efficacy of a phage cocktail composed of four virulent (lytic phages was evaluated for control of PD. Xf levels in grapevines were significantly reduced in therapeutically or prophylactically treated grapevines. PD symptoms ceased to progress one week post-therapeutic treatment and symptoms were not observed in prophylactically treated grapevines. Cocktail phage levels increased in grapevines in the presence of the host. No in planta phage-resistant Xf isolates were obtained. Moreover, Xf mutants selected for phage resistance in vitro did not cause PD symptoms. Our results indicate that phages have great potential for biocontrol of PD and other economically important diseases caused by Xylella.

  15. Clinical Relevance of <em>CDH1em> and <em>CDH13em> DNA-Methylation in Serum of Cervical Cancer Patients

    Directory of Open Access Journals (Sweden)

    Günther K. Bonn

    2012-07-01

    Full Text Available This study was designed to investigate the DNA-methylation status of <em>E>-cadherin (<em>CDH1em> and <em>H>-cadherin (<em>CDH13em> in serum samples of cervical cancer patients and control patients with no malignant diseases and to evaluate the clinical utility of these markers. DNA-methylation status of <em>CDH1em> and <em>CDH13em> was analyzed by means of MethyLight-technology in serum samples from 49 cervical cancer patients and 40 patients with diseases other than cancer. To compare this methylation analysis with another technique, we analyzed the samples with a denaturing high performance liquid chromatography (DHPLC PCR-method. The specificity and sensitivity of <em>CDH1em> DNA-methylation measured by MethyLight was 75% and 55%, and for <em>CDH13em> DNA-methylation 95% and 10%. We identified a specificity of 92.5% and a sensitivity of only 27% for the <em>CDH1em> DHPLC-PCR analysis. Multivariate analysis showed that serum <em>CDH1em> methylation-positive patients had a 7.8-fold risk for death (95% CI: 2.2–27.7; <em>p> = 0.001 and a 92.8-fold risk for relapse (95% CI: 3.9–2207.1; <em>p> = 0.005. We concluded that the serological detection of <em>CDH1em> and <em>CDH13em> DNA-hypermethylation is not an ideal diagnostic tool due to low diagnostic specificity and sensitivity. However, it was validated that <em>CDH1em> methylation analysis in serum samples may be of potential use as a prognostic marker for cervical cancer patients.

  16. Fumigant Antifungal Activity of Myrtaceae Essential Oils and Constituents from <em>Leptospermum petersoniiem> against Three <em>Aspergillus> Species

    Directory of Open Access Journals (Sweden)

    Il-Kwon Park

    2012-09-01

    Full Text Available Commercial plant essential oils obtained from 11 Myrtaceae plant species were tested for their fumigant antifungal activity against <em>Aspergillus ochraceusem>, <em>A. flavusem>, and <em>A. nigerem>. Essential oils extracted from<em> em>Leptospermum> <em>petersonii> at air concentrations of 56 × 10−3 mg/mL and 28 × 10−3 mg/mL completely inhibited the growth of the three <em>Aspergillus> species. However, at an air concentration of 14 × 10−3 mg/mL, inhibition rates of <em>L. petersoniiem> essential oils were reduced to 20.2% and 18.8% in the case of <em>A. flavusem> and <em>A. nigerem>, respectively. The other Myrtaceae essential oils (56 × 10−3 mg/mL only weakly inhibited the fungi or had no detectable affect. Gas chromatography-mass spectrometry analysis identified 16 compounds in <em>L. petersoniiem>> em>essential> em>oil.> em>The antifungal activity of the identified compounds was tested individually by using standard or synthesized compounds. Of these, neral and geranial inhibited growth by 100%, at an air concentration of 56 × 10−3 mg/mL, whereas the activity of citronellol was somewhat lover (80%. The other compounds exhibited only moderate or weak antifungal activity. The antifungal activities of blends of constituents identified in <em>L. petersoniiem> oil indicated that neral and geranial were the major contributors to the fumigant and antifungal activities.

  17. Assessment of Genetic Fidelity in <em>Rauvolfia em>s>erpentina em>Plantlets Grown from Synthetic (Encapsulated Seeds Following <em>in Vitroem> Storage at 4 °C

    Directory of Open Access Journals (Sweden)

    Mohammad Anis

    2012-05-01

    Full Text Available An efficient method was developed for plant regeneration and establishment from alginate encapsulated synthetic seeds of <em>Rauvolfia serpentinaem>. Synthetic seeds were produced using <em>in vitroem> proliferated microshoots upon complexation of 3% sodium alginate prepared in Llyod and McCown woody plant medium (WPM and 100 mM calcium chloride. Re-growth ability of encapsulated nodal segments was evaluated after storage at 4 °C for 0, 1, 2, 4, 6 and 8 weeks and compared with non-encapsulated buds. Effects of different media <em>viz>; Murashige and Skoog medium; Lloyd and McCown woody Plant medium, Gamborg’s B5 medium and Schenk and Hildebrandt medium was also investigated for conversion into plantlets. The maximum frequency of conversion into plantlets from encapsulated nodal segments stored at 4 °C for 4 weeks was achieved on woody plant medium supplement with 5.0 μM BA and 1.0 μM NAA. Rooting in plantlets was achieved in half-strength Murashige and Skoog liquid medium containing 0.5 μM indole-3-acetic acid (IAA on filter paper bridges. Plantlets obtained from stored synseeds were hardened, established successfully <em>ex vitroem> and were morphologically similar to each other as well as their mother plant. The genetic fidelity of <em>Rauvolfia em>clones raised from synthetic seeds following four weeks of storage at 4 °C were assessed by using random amplified polymorphic<em> em>DNA (RAPD and inter-simple sequence repeat<em> em>(ISSR markers. All the RAPD and ISSR profiles from generated plantlets were monomorphic and comparable<em> em>to the mother plant, which confirms the genetic<em> em>stability among the clones. This synseed protocol could be useful for establishing a particular system for conservation, short-term storage and production of genetically identical and stable plants before it is released for commercial purposes.

  18. The EM Earthquake Precursor

    Science.gov (United States)

    Jones, K. B., II; Saxton, P. T.

    2013-12-01

    Many attempts have been made to determine a sound forecasting method regarding earthquakes and warn the public in turn. Presently, the animal kingdom leads the precursor list alluding to a transmission related source. By applying the animal-based model to an electromagnetic (EM) wave model, various hypotheses were formed, but the most interesting one required the use of a magnetometer with a differing design and geometry. To date, numerous, high-end magnetometers have been in use in close proximity to fault zones for potential earthquake forecasting; however, something is still amiss. The problem still resides with what exactly is forecastable and the investigating direction of EM. After the 1989 Loma Prieta Earthquake, American earthquake investigators predetermined magnetometer use and a minimum earthquake magnitude necessary for EM detection. This action was set in motion, due to the extensive damage incurred and public outrage concerning earthquake forecasting; however, the magnetometers employed, grounded or buried, are completely subject to static and electric fields and have yet to correlate to an identifiable precursor. Secondly, there is neither a networked array for finding any epicentral locations, nor have there been any attempts to find even one. This methodology needs dismissal, because it is overly complicated, subject to continuous change, and provides no response time. As for the minimum magnitude threshold, which was set at M5, this is simply higher than what modern technological advances have gained. Detection can now be achieved at approximately M1, which greatly improves forecasting chances. A propagating precursor has now been detected in both the field and laboratory. Field antenna testing conducted outside the NE Texas town of Timpson in February, 2013, detected three strong EM sources along with numerous weaker signals. The antenna had mobility, and observations were noted for recurrence, duration, and frequency response. Next, two

  19. Classe social: conceitos e esquemas operacionais em pesquisa em saude

    Directory of Open Access Journals (Sweden)

    Rita Barradas Barata

    2013-08-01

    Full Text Available Discute-se a utilização do conceito de classe em pesquisas em saúde, as diferentes abordagens sociológicas de estratificação social e de estrutura de classes, o potencial explicativo do conceito em estudos de determinação social e desigualdades em saúde, os modelos de operacionalização elaborados para uso em pesquisas sociológicas, demográficas ou de saúde e os limites e possibilidades desses modelos. Foram destacados quatro modelos de operacionalização: de Singer para estudo da distribuição de renda no Brasil, adaptado por Barros para uso em pesquisas epidemiológicas; de Bronfman & Tuirán para o censo demográfico mexicano, adaptado por Lombardi et al para pesquisas epidemiológicas; de Goldthorpe para estudos socioeconômicos ingleses, adaptado pela Sociedade Espanhola de Epidemiologia; e o modelo de Wright para pesquisa em sociologia e ciência política, também usado em inquéritos populacionais em saúde. Em conclusão, conceitualmente cada um dos modelos apresentados é coerente com a concepção teórica que os embasam, mas não há como optar por qualquer deles, descartando os demais.

  20. Efecto de extractos vegetales de <em>Polygonum hydropiperoidesem>, <em>Solanum nigrumem> y <em>Calliandra pittieriem> sobre el gusano cogollero (<em>Spodoptera frugiperdaem>

    Directory of Open Access Journals (Sweden)

    Lizarazo H. Karol

    2008-12-01

    Full Text Available

    El gusano cogollero <em>Spodoptera frugiperdaem> es una de las plagas que más afectan los cultivos en la región de Sumapaz (Cundinamarca, Colombia. En la actualidad se controla principalmente aplicando productos de síntesis química, sin embargo la aplicación de extractos vegetales surge como una alternativa de menor impacto sobre el ambiente. Este control se emplea debido a que las plantas contienen metabolitos secundarios que pueden inhibir el desarrollo de los insectos. Por tal motivo, la presente investigación evaluó el efecto insecticida y antialimentario de extractos vegetales de barbasco <em>Polygonum hydropiperoidesem> (Polygonaceae, carbonero <em>Calliandra pittieriem> (Mimosaceae y hierba mora <em>Solanum nigrumem> (Solanaceae sobre larvas de <em>S. frugiperdaem> biotipo maíz. Se estableció una cría masiva del insecto en el laboratorio utilizando una dieta natural con hojas de maíz. Posteriormente se obtuvieron extractos vegetales utilizando solventes de alta polaridad (agua y etanol y media polaridad (diclorometano los cuales se aplicaron sobre las larvas de segundo instar. Los resultados más destacados se presentaron con extractos de <em>P. hydropiperoidesem>, obtenidos con diclorometano en sus diferentes dosis, con los cuales se alcanzó una mortalidad de 100% 12 días después de la aplicación y un efecto antialimentario representado por un consumo de follaje de maíz inferior al 4%, efectos similares a los del testigo comercial (Clorpiriphos.

  1. Poesia em Revista: Oroboro

    Directory of Open Access Journals (Sweden)

    Helena Alves Gouveia

    2008-10-01

    Full Text Available http://dx.doi.org/10.5007/1984-784x.2008v8n12p38 A serpente que engole a si mesma é uma figura curiosa do simbolismo de um processo de contínua transformação, de um movimento circular incessante, rumo à infinitude, sem traços de fim ou começo. Oroboro é um nome de origem grega que remete a esta serpente que se morde e penetra em si mesma ao engolir o próprio rabo. Mas também é o nome da revista de cultura editada em Curitiba pelos artistas-editores Ricardo Corona e Eliana Borges.

  2. Hipervitaminose D em animais

    Directory of Open Access Journals (Sweden)

    Paulo V. Peixoto

    2012-07-01

    Full Text Available Por meio de revisão da literatura, são apresentados dados referentes ao metabolismo da vitamina D, bem como aos principais aspectos toxicológicos, clínicos, bioquímicos, macroscópicos, microscópicos, ultraestruturais, imuno-histoquímicos e radiográficos de animais intoxicados natural e experimentalmente por essa vitamina, em diferentes espécies. Este estudo objetiva demonstrar a existência de muitas lacunas no conhecimento sobre mineralização fisiológica e patológica, em especial na mediação hormonal do fenômeno, bem como alertar para os riscos de ocorrência dessa intoxicação.

  3. A democracia em Cuba

    OpenAIRE

    Zaldívar, Julio César Guanche

    2011-01-01

    O triunfo revolucionário de 1959 consagrou em Cuba um novo conceito de democracia, com o intuito de garantir o acesso à vida política ativa de grandes setores da população, antes excluídos. Para isso, foi desenvolvida uma política de inclusão social com caráter universal. A prática política popular deixou as riquezas do país em mãos da população carente e gerou uma grande mobilidade social, fato que foi central para o aumento da participação popular. O contexto de agressão imperialista e o pr...

  4. Tuberculose Infantil em Portugal

    OpenAIRE

    Carapau, João

    2014-01-01

    Dos números recentemente publicados pela Direcção Geral da Saúde / Núcleo de Tuberculose e Doenças Respiratórias relativos aos anos de 1992 e 1993 e pelo Instituto Nacional de Estatística relativos a 1994, conclui-se que os casos de Tuberculose (TB) notificados pouco têm decrescido nos últimos 15 anos: descida média anual de 6,3% para os casos em geral e 14% para os menores de 15 anos; a taxa global de incidência apurada em 1994 voltou a subir — 51,1 (52,4 no Continente). Para o autor a me...

  5. Universidade em ruinas?

    OpenAIRE

    Katuta, Ângela Massumi; UEL/CCE/Departamento de Geociências

    2010-01-01

    A Universidade, desde as suas origens no século XII, sempre esteve atrelada a instituições e setores hegemônicos da sociedade. Segundo Trindade (2000), a sua “invenção” ocorreu em plena Idade Média na Europa, sob a proteção da Igreja romana, sendo que as Universidades de Bolonha (1108) e Paris (1211) foram as primeiras a serem criadas 

  6. Rapid Development of Microsatellite Markers with 454 Pyrosequencing in a Vulnerable Fish<em>,> the Mottled Skate<em>, Raja em>pulchra>

    Directory of Open Access Journals (Sweden)

    Jung-Ha Kang

    2012-06-01

    Full Text Available The mottled skate, <em>Raja pulchraem>, is an economically valuable fish. However, due to a severe population decline, it is listed as a vulnerable species by the International Union for Conservation of Nature. To analyze its genetic structure and diversity, microsatellite markers were developed using 454 pyrosequencing. A total of 17,033 reads containing dinucleotide microsatellite repeat units (mean, 487 base pairs were identified from 453,549 reads. Among 32 loci containing more than nine repeat units, 20 primer sets (62% produced strong PCR products, of which 14 were polymorphic. In an analysis of 60 individuals from two <em>R. pulchra em>populations, the number of alleles per locus ranged from 1–10, and the mean allelic richness was 4.7. No linkage disequilibrium was found between any pair of loci, indicating that the markers were independent. The Hardy–Weinberg equilibrium test showed significant deviation in two of the 28 single-loci after sequential Bonferroni’s correction. Using 11 primer sets, cross-species amplification was demonstrated in nine related species from four families within two classes. Among the 11 loci amplified from three other <em>Rajidae> family species; three loci were polymorphic. A monomorphic locus was amplified in all three <em>Rajidae> family species and the <em>Dasyatidae> family. Two <em>Rajidae> polymorphic loci amplified monomorphic target DNAs in four species belonging to the Carcharhiniformes class, and another was polymorphic in two Carcharhiniformes species.

  7. Acarofauna em plantas ornamentais

    Directory of Open Access Journals (Sweden)

    Jania Claudia Camilo dos Santos

    2014-10-01

    Full Text Available Normal 0 21 false false false PT-BR X-NONE X-NONE O cultivo e o comercio de plantas ornamentais vem cada vez mais ganhando espaço no Brasil, pela grande variedade das espécies existentes e exuberância de suas flores, que oferecem uma maior riqueza ao local. Dessa forma, o objetivo desse trabalho foi realizar o levantamento da população de ácaros associados às plantas ornamentais no município de Arapiraca-AL, em função dos diversos problemas acarretados por essa espécie. O levantamento foi realizado entre os meses de abril a março, através de amostragens mensais de folhas coletadas da parte basal, intermediária e apical de plantas existentes em praças e jardins. Foram coletados 55 ácaros pertencentes à ordem Prostigmata em 20 famílias de plantas. As plantas com as maiores riquezas de ácaros foram as Coleus blumei L. e Bxuxus sempervirens L., que apresentaram 65% dos valores amostrais. Analisando-se as coletas realizadas, pode-se observar que houve uma maior incidência populacional de ácaros na coleta do mês de maio, cuja percentagem foi de 36% de ácaros levantados, sendo que no levantamento dos dados amostrais de março a percentagem encontrada foi de 14%, nas amostragens dos meses de abril e junho, a percentagem amostrada dos dados foi de 22 e 28%, respectivamente. O estudo do levantamento de ácaros em plantas ornamentais permitiu observar a relação entre ácaros e a relação com a planta hospedeira, facilitando posteriormente um estudo mais aprofundado sobre plantas hospedeiras, e pode-se observar que em períodos chuvosos ocorre uma menor incidência populacional.

  8. How <em>Varroa> Parasitism Affects the Immunological and Nutritional Status of the Honey Bee, <em>Apis melliferaem>

    Directory of Open Access Journals (Sweden)

    Katherine A. Aronstein

    2012-06-01

    Full Text Available We investigated the effect of the parasitic mite <em>Varroa destructorem> on the immunological and nutritional condition of honey bees, <em>Apis melliferaem>, from the perspective of the individual bee and the colony. Pupae, newly-emerged adults and foraging adults were sampled from honey bee colonies at one site in S. Texas, USA. <em>Varroa>‑infested bees displayed elevated titer of Deformed Wing Virus (DWV, suggestive of depressed capacity to limit viral replication. Expression of genes coding three anti-microbial peptides (<em>defensin1, abaecin, hymenoptaecinem> was either not significantly different between <em>Varroa>-infested and uninfested bees or was significantly elevated in <em>Varroa>-infested bees, varying with sampling date and bee developmental age. The effect of <em>Varroa> on nutritional indices of the bees was complex, with protein, triglyceride, glycogen and sugar levels strongly influenced by life-stage of the bee and individual colony. Protein content was depressed and free amino acid content elevated in <em>Varroa>-infested pupae, suggesting that protein synthesis, and consequently growth, may be limited in these insects. No simple relationship between the values of nutritional and immune-related indices was observed, and colony-scale effects were indicated by the reduced weight of pupae in colonies with high <em>Varroa> abundance, irrespective of whether the individual pupa bore <em>Varroa>.

  9. Synthesis and Spectroscopic Analysis of Novel 1<em>H-Benzo[d>]imidazoles Phenyl Sulfonylpiperazines

    Directory of Open Access Journals (Sweden)

    Amjad M. Qandil

    2012-05-01

    Full Text Available A group of benzimidazole analogs of sildenafil, 3-benzimidazolyl-4-methoxy-phenylsulfonylpiperazines 2–4 and 3-benzimidazolyl-4-methoxy-<em>N,N>-dimethyl- benzenesulfonamide (5, were efficiently synthesized. Compounds 2–5 were characterized by NMR and MS and contrary to the reported mass spectra of sildenafil, the spectra of the piperazine-containing compounds 2–4 showed a novel fragmentation pattern leading to an <em>m/z> = 316. A mechanism for the formation of this fragment was proposed.

  10. Anestesia Diploica em Endodontia

    OpenAIRE

    Macedo, Ricardo Ribeiro Veiga de

    2013-01-01

    Trabalho final do 5º ano com vista à atribuição do grau de mestre no âmbito do ciclo de estudos de Mestrado Integrado em Medicina Dentária apresentado à Faculdade de Medicina da Universidade de Coimbra. Objetivos Comparar a eficácia das técnicas de anestesia convencionais, a anestesia infiltrativa periapical, com a anestesia diploica. Metodologia Foram selecionados 32 voluntários, saudáveis, aos quais foram administradas ambas as técnicas anestésicas no dente 1.4. Numa primeira fase os...

  11. Burnout em Cuidadores Formais

    OpenAIRE

    Silva, Juliana Marisa Fernandes

    2016-01-01

    Dissertação apresentada à Universidade Fernando Pessoa como parte dos requisitos para a obtenção do grau de Mestre em Psicologia, ramo de Psicologia Clínica e da Saúde Este estudo é sobre a Síndrome de Burnout e teve como objetivo geral compreender se o Burnout está presente nos cuidadores formais da Santa Casa da Misericórdia de Castelo de Paiva e quais as variáveis socioprofissionais que poderão exercer influência no seu aparecimento. Pretendeu-se avaliar o Burnout dos cuidadores a tr...

  12. Antioxidant Profile of <em>Trifolium pratenseem> L.

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    Heidy Schwartsova

    2012-09-01

    Full Text Available In order to examine the antioxidant properties of five different extracts of <em>Trifolium pratenseem> L. (Leguminosae leaves, various assays which measure free radical scavenging ability were carried out: 1,1-diphenyl-2-picrylhydrazyl, hydroxyl, superoxide anion and nitric oxide radical scavenger capacity tests and lipid peroxidation assay. In all of the tests, only the H2O and (to some extent the EtOAc extracts showed a potent antioxidant effect compared with BHT and BHA, well-known synthetic antioxidants. In addition, <em>in vivo em>experiments were conducted with antioxidant systems (activities of GSHPx, GSHR, Px, CAT, XOD, GSH content and intensity of LPx in liver homogenate and blood of mice after their treatment with extracts of <em>T. pratenseem> leaves, or in combination with CCl4. Besides, in the extracts examined the total phenolic and flavonoid amounts were also determined, together with presence of the selected flavonoids: quercetin, luteolin, apigenin, naringenin and kaempferol, which were studied using a HPLC-DAD technique. HPLC-DAD analysis showed a noticeable content of natural products according to which the examined <em>Trifolium pratenseem> species could well be regarded as a promising new source of bioactive natural compounds, which can be used both as a food supplement and a remedy.

  13. Proximate Composition, Nutritional Attributes and Mineral Composition of <em>Peperomia> <em>pellucida> L. (Ketumpangan Air Grown in Malaysia

    Directory of Open Access Journals (Sweden)

    Maznah Ismail

    2012-09-01

    Full Text Available This study presents the proximate and mineral composition of <em>Peperomia> <em>pellucida> L., an underexploited weed plant in Malaysia. Proximate analysis was performed using standard AOAC methods and mineral contents were determined using atomic absorption spectrometry. The results indicated <em>Peperomia> <em>pellucida> to be rich in crude protein, carbohydrate and total ash contents. The high amount of total ash (31.22% suggests a high-value mineral composition comprising potassium, calcium and iron as the main elements. The present study inferred that <em>Peperomia> <em>pellucida> would serve as a good source of protein and energy as well as micronutrients in the form of a leafy vegetable for human consumption.

  14. O insight em psiquiatria

    Directory of Open Access Journals (Sweden)

    Ana Margarida P. Cardoso

    2008-12-01

    Full Text Available O sinal de que algo está a acontecer contribui para que o paciente reconheça que alguma coisa de estranho se está a passar consigo. Este reconhecimento faz com que o sujeito possa desempenhar uma função activa e seja um elemento colaborante do seu processo de recuperação. Cada doença apresenta, contudo, diferentes sintomas, uma vez que cada doença psiquiátrica consiste em diferentes perturbações com diversos efeitos sobre o funcionamento mental. Desta maneira, o fenómeno do insight que é registado em cada doença é diferente e expressa-se sob diferentes formas, não somente devido às manifestações clínicas da doença mas também devido às características individuais do sujeito.

  15. Antibioticos profilaticos em neurocirurgia

    Directory of Open Access Journals (Sweden)

    Reynaldo A. Brandt

    1979-03-01

    Full Text Available O índice de infecções pós-operatórias em pacientes neuro-cirúrgicos que receberam antibióticos profiláticos neste período foi comparado com o de pacientes que não receberam antibióticos. Infecções ocorreram em proporções significativamente maiores nos pacientes que receberam antibióticos, particularmente naqueles com afecções intracranianas graves; estas infecções foram graves e fatais na maioria dos casos. A administração de antibióticos profiláticos nestes pacientes não só foi incapaz de prevenir o aparecimento de infecções pós-operatórias, como aparentemente favoreceu o seu desenvolvimento. Tal se deveu, provavelmente, à destruição do equilíbrio microbiano no organismo, favorecendo o desenvolvimento de germes patogênicos e resistentes aos antibióticos usuais

  16. A democracia em Cuba

    Directory of Open Access Journals (Sweden)

    Julio César Guanche Zaldívar

    2011-01-01

    Full Text Available O triunfo revolucionário de 1959 consagrou em Cuba um novo conceito de democracia, com o intuito de garantir o acesso à vida política ativa de grandes setores da população, antes excluídos. Para isso, foi desenvolvida uma política de inclusão social com caráter universal. A prática política popular deixou as riquezas do país em mãos da população carente e gerou uma grande mobilidade social, fato que foi central para o aumento da participação popular. O contexto de agressão imperialista e o próprio desenvolvimento do processo produziram a consolidação de noções que limitaram a participação popular: o apogeu da burocracia, a compreensão da unidade como unanimidade e o seguimento, em certa medida, de correntes do marxismo soviético. Os desafios atuais para aprofundar a democracia em Cuba se apresentam em três planos: socializar o poder, promover a sociodiversidade e desenvolver a ideologia revolucionária.El triunfo revolucionario de 1959 consagró en Cuba un nuevo concepto de democracia, basado en garantizar acceso a la vida política activa a grandes sectores poblacionales, antes excluidos de ella. Para ello desarrolló una política de inclusión social con carácter universal. La práctica política popular puso las riquezas del país en manos de los desposeídos y generó gran movilidad social, hecho que resultó clave para el aumento de la participación popular. El contexto de agresión imperialista y el propio desarrollo del proceso produjo el afianzamiento de nociones que limitaron la participación popular: el auge de la burocracia, la comprensión de la unidad como unanimidad y el seguimiento, en parte, de corrientes del marxismo soviético. Los desafíos actuales se presentan en tres planos para profundizar la democracia en Cuba: socializar el poder, promover la sociodiversidad y desarrollar la ideología revolucionaria.The revolutionary triumph of 1959 established in Cuba a new concept of democracy, one that

  17. Identified EM Earthquake Precursors

    Science.gov (United States)

    Jones, Kenneth, II; Saxton, Patrick

    2014-05-01

    Many attempts have been made to determine a sound forecasting method regarding earthquakes and warn the public in turn. Presently, the animal kingdom leads the precursor list alluding to a transmission related source. By applying the animal-based model to an electromagnetic (EM) wave model, various hypotheses were formed, but the most interesting one required the use of a magnetometer with a differing design and geometry. To date, numerous, high-end magnetometers have been in use in close proximity to fault zones for potential earthquake forecasting; however, something is still amiss. The problem still resides with what exactly is forecastable and the investigating direction of EM. After a number of custom rock experiments, two hypotheses were formed which could answer the EM wave model. The first hypothesis concerned a sufficient and continuous electron movement either by surface or penetrative flow, and the second regarded a novel approach to radio transmission. Electron flow along fracture surfaces was determined to be inadequate in creating strong EM fields, because rock has a very high electrical resistance making it a high quality insulator. Penetrative flow could not be corroborated as well, because it was discovered that rock was absorbing and confining electrons to a very thin skin depth. Radio wave transmission and detection worked with every single test administered. This hypothesis was reviewed for propagating, long-wave generation with sufficient amplitude, and the capability of penetrating solid rock. Additionally, fracture spaces, either air or ion-filled, can facilitate this concept from great depths and allow for surficial detection. A few propagating precursor signals have been detected in the field occurring with associated phases using custom-built loop antennae. Field testing was conducted in Southern California from 2006-2011, and outside the NE Texas town of Timpson in February, 2013. The antennae have mobility and observations were noted for

  18. Anterior foregut microbiota of the glassy-winged sharpshooter explored using deep 16S rRNA gene sequencing from individual insects.

    Directory of Open Access Journals (Sweden)

    Elizabeth E Rogers

    Full Text Available The glassy-winged sharpshooter (GWSS is an invasive insect species that transmits Xylella fastidiosa, the bacterium causing Pierce's disease of grapevine and other leaf scorch diseases. X. fastidiosa has been shown to colonize the anterior foregut (cibarium and precibarium of sharpshooters, where it may interact with other naturally-occurring bacterial species. To evaluate such interactions, a comprehensive list of bacterial species associated with the sharpshooter cibarium and precibarium is needed. Here, a survey of microbiota associated with the GWSS anterior foregut was conducted. Ninety-six individual GWSS, 24 from each of 4 locations (Bakersfield, CA; Ojai, CA; Quincy, FL; and a laboratory colony, were characterized for bacteria in dissected sharpshooter cibaria and precibaria by amplification and sequencing of a portion of the 16S rRNA gene using Illumina MiSeq technology. An average of approximately 150,000 sequence reads were obtained per insect. The most common genus detected was Wolbachia; sequencing of the Wolbachia ftsZ gene placed this strain in supergroup B, one of two Wolbachia supergroups most commonly associated with arthropods. X. fastidiosa was detected in all 96 individuals examined. By multilocus sequence typing, both X. fastidiosa subspecies fastidiosa and subspecies sandyi were present in GWSS from California and the colony; only subspecies fastidiosa was detected in GWSS from Florida. In addition to Wolbachia and X. fastidiosa, 23 other bacterial genera were detected at or above an average incidence of 0.1%; these included plant-associated microbes (Methylobacterium, Sphingomonas, Agrobacterium, and Ralstonia and soil- or water-associated microbes (Anoxybacillus, Novosphingobium, Caulobacter, and Luteimonas. Sequences belonging to species of the family Enterobacteriaceae also were detected but it was not possible to assign these to individual genera. Many of these species likely interact with X. fastidiosa in the

  19. Osteoporose em caprinos

    Directory of Open Access Journals (Sweden)

    Fábio B. Rosa

    2013-04-01

    Full Text Available Foi realizado um estudo de casos de osteoporose em caprinos provenientes de uma propriedade na área de abrangência do LPV-UFSM, determinando a epidemiologia, o quadro clínico-patológico e discutindo os prováveis mecanismos patogenéticos. Cinco cabras, fêmeas, SRD, de seis meses a seis anos de idade foram afetadas. As cabras eram mantidas em campo nativo, sem suplementação com ração e sob superlotação. Os principais sinais clínicos foram emagrecimento, dificuldade de locomoção e permanência em decúbito por longos períodos. As principais alterações macroscópicas nos ossos examinados foram vistas nas superfícies de corte e caracterizavam-se por depleção do osso esponjoso (porosidade e redução acentuada da espessura do osso cortical. Havia também marcada atrofia serosa da gordura da medula óssea. Microscopicamente, nas regiões avaliadas (úmero proximal, rádio distal, fêmur distal, tíbia proximal e corpos das vértebras lombares foi observada redução moderada a acentuada do número e da espessura das trabéculas ósseas nas epífises e metáfises dos ossos longos e nos corpos vertebrais. Os achados clínico-patológicos indicaram que a osteoporose observada provavelmente foi causada pela desnutrição. As alterações ósseas (diminuição no número e na espessura das trabéculas do osso esponjoso sugerem que ambos os mecanismos, má formação óssea e reabsorção óssea aumentada, contribuíram para a ocorrência de osteoporose nos caprinos deste estudo.

  20. Four Novel Cellulose Synthase (CESA Genes from <em>Birch> (<em>Betula platyphylla em>Suk. Involved in Primary and Secondary Cell Wall Biosynthesis

    Directory of Open Access Journals (Sweden)

    Xuemei Liu

    2012-09-01

    Full Text Available Cellulose synthase (CESA, which is an essential catalyst for the generation of plant cell wall biomass, is mainly encoded by the <em>CesA> gene family that contains ten or more members. In this study; four full-length cDNAs encoding CESA were isolated from<em> Betula platyphyllaem> Suk., which is an important timber species, using RT-PCR combined with the RACE method and were named as <em>BplCesA3em>, <em>−4em>,> −7 em>and> −8em>. These deduced CESAs contained the same typical domains and regions as their <em>Arabidopsis> homologs. The cDNA lengths differed among these four genes, as did the locations of the various protein domains inferred from the deduced amino acid sequences, which shared amino acid sequence identities ranging from only 63.8% to 70.5%. Real-time RT-PCR showed that all four <em>BplCesAs> were expressed at different levels in diverse tissues. Results indicated that BplCESA8 might be involved in secondary cell wall biosynthesis and floral development. BplCESA3 appeared in a unique expression pattern and was possibly involved in primary cell wall biosynthesis and seed development; it might also be related to the homogalacturonan synthesis. BplCESA7 and BplCESA4 may be related to the formation of a cellulose synthase complex and participate mainly in secondary cell wall biosynthesis. The extremely low expression abundance of the four BplCESAs in mature pollen suggested very little involvement of them in mature pollen formation in <em>Betula>. The distinct expression pattern of the four <em>BplCesAs> suggested they might participate in developments of various tissues and that they are possibly controlled by distinct mechanisms in <em>Betula.>

  1. <em>N>-Substituted 5-Chloro-6-phenylpyridazin-3(2<em>H>-ones: Synthesis, Insecticidal Activity Against <em>Plutella xylostella em>(L. and SAR Study

    Directory of Open Access Journals (Sweden)

    Song Yang

    2012-08-01

    Full Text Available A series of <em>N>-substituted 5-chloro-6-phenylpyridazin-3(2<em>H>-one derivatives were synthesized based on our previous work; all compounds were characterized by spectral data and tested for <em>in vitroem> insecticidal activity against <em>Plutella xylostellaem>. The results showed that the synthesized pyridazin-3(2<em>H>-one compounds possessed good insecticidal activities, especially the compounds 4b, 4d, and 4h which showed > 90% activity at 100 mg/L. The structure-activity relationships (SAR for these compounds were also discussed.

  2. INDICADORES EM SANEAMENTO

    OpenAIRE

    Costa, Samuel Alves Barbi; Côrtes, Larissa Silveira; Coelho Netto, Taiana; Freitas Junior, Moacyr Moreira de

    2016-01-01

    Este artigo se propõe a analisar a evolução dos prestadores de serviços de saneamento do estado de MinasGerais entre os anos de 2005 e 2010 com base nos indicadores do Sistema Nacional de Informações em Saneamento(SNIS). Foram definidos parâmetros técnicos para a análise dos indicadores, classificados os resultados como satisfatórios(verdes) ou insatisfatórios (vermelhos). Esta categorização atende a concepção da Regulação Sunshine, trazendo à tona omonitoramento do progresso das ações no set...

  3. Em favor da talassografia

    Directory of Open Access Journals (Sweden)

    Jean-Louis Boudou

    2001-01-01

    Full Text Available A Talassografia (“descrição do mar” interessa-sepelos impactos físicos, biológicos, ecológicos... culturais da violenta antropização dos ambientes costeiros (oceânicos e continentais, caracterizados pelaexigüidade, vulnerabilidade, fragilidade e plasticidade. Como o Brasil é um “país marítimo”, os geó-grafos (os talassógrafos brasileiros são convidadosa intensificar suas pesquisas nas áreas costeiras e acriar novas estruturas para divulgá-las: Revista, Encontros, Associação, Pós-Graduação... tudo em prolda talassografia.

  4. INFLUÊNCIA ESTOICA NA CONCEPÇÃO DE <em>STATUS> E <em>DICTUM> COMO <em> QUASI RES EM> (ὡσανεì τινά EM ABERLARDO STOIC INFLUENCE IN ABELARD'S CONCEPTION OF <em>STATUS> AND <em>DICTUM> AS <em>QUASI RESem> (ὡσανεì τινά.

    Directory of Open Access Journals (Sweden)

    Guy Hamelin

    2011-09-01

    Full Text Available Na sua obra, Pedro Abelardo (1079-1142 destaca duas noções metafísicas que fundamentam sua teoria lógica: o <em style="mso-bidi-font-style: normal;">statusem> e o <em style="mso-bidi-font-style: normal;">dictum propositionisem>, ao causar, respectivamente, a imposição (<em style="mso-bidi-font-style: normal;">impositioem> dos termos universais e o valor de verdade das proposições. Trata-se de expressões que se referem a naturezas ontológicas peculiares, na medida em que não são consideradas coisas (<em style="mso-bidi-font-style: normal;">resem>, mesmo que constituem causas. Todavia, também não são nada. Abelardo as chama de ‘quase coisas’ (<em style="mso-bidi-font-style: normal;">quasi resem>. No presente artigo, explicamos, primeiro, essas duas noções essenciais da lógica abelardiana, antes de tentar, em seguida, encontrar a fonte dessa metafísica particular. Em oposição a comentadores importantes da lógica de Abelardo, que estimam que haja uma forte influência platônica sobre essa concepção específica, defendemos antes, com apoio de textos significativos e de acordo com o nominalismo abelardiano, que a maior ascendência sobre a metafísica do nosso autor é a do estoicismo, sobretudo, antigo.In his work, Peter Abelard (1079-1142 highlights two metaphysical notions, which sustain his logical theory: the <em>status> and the <em>dictum propositionisem>, causing respectively both the imposition (<em>impositio> of universal terms and the thuth-value of propositions. Both expressions refer to peculiar ontological natures, in so far as they are not considered things (<em>res>, even if they constitute causes. Nevertheless, neither are they ‘nothing’. Abelard calls them ‘quasi-things’ (<em>quasi resem>. In the present article, we expound first these two essential notions of Abelardian logic before then trying to find the source of this particular metaphysics. Contrary to some important

  5. Ploidia de DNA em astrocitomas: estudo em 66 pacientes brasileiros

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    KRUTMAN-ZVEIBIL DEBORAH

    1999-01-01

    Full Text Available A determinação do conteúdo de DNA nuclear (fração de fase S e ploidia de DNA foi realizada por meio de análise de imagem em 66 astrocitomas, a partir de material fixado em formalina e seccionado em cortes de 5 micrômetros corados pela técnica de Feulgen. Nossos resultados mostraram forte relação entre a idade do paciente, grau histológico e sobrevida , com a ploidia de DNA e o percentual de células em fase de síntese. A análise da atividade proliferativa de astrocitomas intracranianos é a nosso ver muito útil no entendimento do comportamento biológico , do prognóstico e para o planejamento terapêutico dessas lesões.

  6. Busca de estruturas em grandes escalas em altos redshifts

    Science.gov (United States)

    Boris, N. V.; Sodré, L., Jr.; Cypriano, E.

    2003-08-01

    A busca por estruturas em grandes escalas (aglomerados de galáxias, por exemplo) é um ativo tópico de pesquisas hoje em dia, pois a detecção de um único aglomerado em altos redshifts pode por vínculos fortes sobre os modelos cosmológicos. Neste projeto estamos fazendo uma busca de estruturas distantes em campos contendo pares de quasares próximos entre si em z Â3 0.9. Os pares de quasares foram extraídos do catálogo de Véron-Cetty & Véron (2001) e estão sendo observados com os telescópios: 2,2m da University of Hawaii (UH), 2,5m do Observatório de Las Campanas e com o GEMINI. Apresentamos aqui a análise preliminar de um par de quasares observado nos filtros i'(7800 Å) e z'(9500 Å) com o GEMINI. A cor (i'-z') mostrou-se útil para detectar objetos "early-type" em redshifts menores que 1.1. No estudo do par 131046+0006/J131055+0008, com redshift ~ 0.9, o uso deste método possibilitou a detecção de sete objetos candidatos a galáxias "early-type". Num mapa da distribuição projetada dos objetos para 22 escala. Um outro argumento em favor dessa hipótese é que eles obedecem uma relação do tipo Kormendy (raio equivalente X brilho superficial dentro desse raio), como a apresentada pelas galáxias elípticas em z = 0.

  7. Extraction of Dihydroquercetin<em> em>from <em>Larix gmeliniem>i> em>with Ultrasound-Assisted and Microwave-Assisted Alternant Digestion

    Directory of Open Access Journals (Sweden)

    Yuangang Zu

    2012-07-01

    Full Text Available An ultrasound and microwave assisted alternant extraction method (UMAE was applied for extracting dihydroquercetin (DHQ from <em>Larix gmeliniem>i> wood. This investigation was conducted using 60% ethanol as solvent, 1:12 solid to liquid ratio, and 3 h soaking time. The optimum treatment time was ultrasound 40 min, microwave 20 min, respectively, and the extraction was performed once. Under the optimized conditions, satisfactory extraction yield of the target analyte was obtained. Relative to ultrasound-assisted or microwave-assisted method, the proposed approach provides higher extraction yield. The effect of DHQ of different concentrations and synthetic antioxidants on oxidative stability in soy bean oil stored for 20 days at different temperatures (25 °C and 60 °C was compared. DHQ was more effective in restraining soy bean oil oxidation, and a dose-response relationship was observed. The antioxidant activity of DHQ was a little stronger than that of BHA and BHT. Soy bean oil supplemented with 0.08 mg/g DHQ exhibited favorable antioxidant effects and is preferable for effectively avoiding oxidation. The <em>L. gmeliniiem> wood samples before and after extraction were characterized by scanning electron microscopy. The results showed that the UMAE method is a simple and efficient technique for sample preparation.

  8. Preferencia alimenticia del ácaro depredador <em>Balaustium> sp. en condiciones controladas

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    Muñoz Karen

    2009-04-01

    Full Text Available

    Se evaluó la preferencia de presas de <em>Balaustium> sp., enemigo natural de diferentes artrópodos plaga, y el cual es nativo de la Sabana de Bogotá. En unidades experimentales construidas con foliolos de plantas de rosa se colocaron independientemente individuos de <em>Balaustium> sp. y se registró el número de presas consumidas. De esta manera se determinó la preferencia de los tres estados móviles del ácaro depredador <em>Balaustium> sp. por diferentes edades de tres presas. Las especies y edades de las presas estudiadas fueron: huevos, ninfas y adultos de <em>Trialeurodes vaporariorumem>, huevos, ninfas y adultos de <em>Tetranychus urticaeem>, y larvas de primer y segundo instar y adultos de <em>Frankliniella occidentalisem>. Los estados menos desarrollados fueron preferidos, aunque se observó que los adultos del depredador tienen gran habilidad para consumir adultos de <em>T. vaporariorumem>. La presa preferida por las larvas de <em>Balaustium> sp. fue los huevos de <em>T. urticaeem> con una proporción de consumo de 0,54 de los huevos que se ofrecieron de esta presa; las deutoninfas del depredador eligieron huevos de <em>T. vaporariorumem> (0,537 o de <em>T. urticaeem> (0,497 y los adultos de <em>Balaustium> sp. prefrieron los huevos de <em>T. vaporariorumem> (0,588.

  9. Conjuntivite em animais de companhia

    OpenAIRE

    Gonçalves, Mariana Inês Pereira Coelho

    2015-01-01

    Dissertação de Mestrado Integrado em Medicina Veterinária, Ciências Veterinárias A conjuntivite é a doença ocular mais frequentemente diagnosticada em Medicina Veterinária, tanto em cães como em gatos. O facto de a conjuntiva ser uma membrana mucosa tão exposta a agentes externos faz com que esteja bastante suscetível a sofrer lesões, com consequente inflamação. Nesta dissertação abordam-se os diferentes tipos de conjuntivite, que são classificados de acordo com a sua etiologia. No Hospita...

  10. Conhecimento e interesse em economia

    Directory of Open Access Journals (Sweden)

    João Antonio de Paula

    2003-09-01

    Full Text Available O artigo argumenta em favor da necessidade de preservar o pluralismo em economia. Discute: i a natureza do conhecimento e de sua produção; ii as especificidades da produção do conhecimento em economia e os condicionamentos histórico-culturais do conhecimento; iii as implicações da hegemonia de certa concepção de método em economia.The article argues the need to preserve the pluralism in economics. The article examines: i the nature of the knowledge and of its production; ii the specificity of the knowledge production in economics and the historical and cultural conditionings of the knowledge; iii the implications of the hegemony of a certain methodological conception in economics.

  11. Activity-Guided Isolation of Antioxidant Compounds from <em>Rhizophora apiculataem>

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    Hongbin Xiao

    2012-09-01

    Full Text Available <em>Rhizophora apiculataem> (<em>R. apiculataem> contains an abundance of biologically active compounds due its special salt-tolerant living surroundings. In this study, the total phenolic content and antioxidant activities of various extract and fractions of stem of <em>R. apiculataem> were investigated. Results indicated that butanol fraction possesses the highest total phenolic content (181.84 mg/g GAE/g dry extract with strongest antioxidant abilities. Following <em>in vitroem> antioxidant activity-guided phytochemical separation procedures, lyoniresinol-3α-<em>O>-β-arabinopyranoside (1, lyoniresinol-3α-<em>O>-β-rhamnoside (2, and afzelechin-3-<em>O>-L-rhamno-pyranoside (3 were separated from the butanol fraction. These compounds showed more noticeable antioxidant activity than a BHT standard in the DPPH, ABTS and hydroxyl radical scavenging assays. HPLC analysis results showed that among different plant parts, the highest content of 13 was located in the bark (0.068%, 0.066% and 0.011%, respectively. The results imply that the <em>R. apiculataem> might be a potential source of natural antioxidants and 13 are antioxidant ingredients in <em>R. apiculataem>.

  12. Regional Suppression of <em>Bactrocera> Fruit Flies (Diptera: Tephritidae in the Pacific through Biological Control and Prospects for Future Introductions into Other Areas of the World

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    Roger I. Vargas

    2012-08-01

    Full Text Available <em>Bactrocera> fruit fly species are economically important throughout the Pacific. The USDA, ARS U.S. Pacific Basin Agricultural Research Center has been a world leader in promoting biological control of <em>Bactrocera> spp. that includes classical, augmentative, conservation and IPM approaches. In Hawaii, establishment of <em>Bactrocera> <em>cucurbitae> (Coquillett in 1895 resulted in the introduction of the most successful parasitoid, <em>Psyttalia> <em>fletcheri> (Silvestri; similarly, establishment of <em>Bactrocera> <em>dorsalis> (Hendel in 1945 resulted in the introduction of 32 natural enemies of which <em>Fopius> <em>arisanus> (Sonan, <em>Diachasmimorpha> <em>longicaudata> (Ashmead and <em>Fopius> <em>vandenboschi> (Fullaway were most successful. Hawaii has also been a source of parasitoids for fruit fly control throughout the Pacific region including Australia, Pacific Island Nations, Central and South America, not only for <em>Bactrocera> spp. but also for <em>Ceratitis> and <em>Anastrepha> spp. Most recently, in 2002, <em>F.> <em>arisanus> was introduced into French Polynesia where <em>B.> <em>dorsalis> had invaded in 1996. Establishment of <em>D.> <em>longicaudata> into the new world has been important to augmentative biological control releases against <em>Anastrepha> spp. With the rapid expansion of airline travel and global trade there has been an alarming spread of <em>Bactrocera> spp. into new areas of the world (<em>i.e.>, South America and Africa. Results of studies in Hawaii and French Polynesia, support parasitoid introductions into South America and Africa, where <em>B.> <em>carambolae> and <em>B.> <em>invadens>, respectively, have become established. In addition, <em>P.> <em>fletcheri> is a candidate for biological control of <em>B.> <em>cucurbitae> in Africa. We review past and more

  13. <em>Trichoderma harzianumem> como promotor del crecimiento vegetal del maracuyá (<em>Passiflora edulisem> var. <em>flavicarpa> Degener

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    Cubillos-Hinojosa Juan

    2009-04-01

    Full Text Available

    Se realizó un experimento en condiciones de laboratorio e invernadero, con el propósito de evaluar el efecto de la cepa nativa TCN-014 y la cepa comercial TCC-005 de <em>Trichoderma harzianumem> sobre la germinación y el crecimiento temprano del maracuyá. Se adecuaron inóculos de 104, 106 y 108 conidias/mL para cada cepa y se aplicaron a semillas de maracuyá; se evaluó el número de semillas germinadas durante 15 días; se calculó el porcentaje de germinación, el índice de velocidad de germinación y el tiempo medio de germinación. Posteriormente las semillas germinadas se llevaron a condiciones de invernadero y transcurridos dos meses se midió la altura de las plántulas, el grosor del tallo, el número de hojas, la longitud de la raíz y el peso seco total. Todos los tratamientos estimularon la germinación de las semillas y el desarrollo de las plántulas; sin embargo la cepa nativa en concentraciones 106 y 108 conidias/mL mostró resultados superiores frente a la cepa comercial. Los resultados sugieren una acción efectiva de <em>T. harzianumem> como promotor de crecimiento vegetal, mostrando que tiene potencial para la elaboración de un bioproducto útil para el manejo ecológico del cultivo de maracuyá.

  14. Tratamento da dor em queimados

    Directory of Open Access Journals (Sweden)

    Rodrigo José Alencar de Castro

    2013-02-01

    Full Text Available JUSTIFICATIVA E OBJETIVOS: Apesar dos avanços, ainda é observado manejo analgésico inadequado dos pacientes com queimaduras. O objetivo desta revisão foi coletar dados sobre tratamento da dor em queimados. CONTEÚDO: Foi feita revisão sobre os mecanismos de dor, avaliação do paciente com queimadura e o tratamento farmacológico e não farmacológico. CONCLUSÕES: O manejo da dor em pacientes vítimas de queimaduras ainda é um desafio por parte da equipe multiprofissional. A avaliação frequente e contínua da resposta apresentada pelo paciente é muito importante, tendo em vista os vários momentos por que passa o paciente internado em decorrência de uma queimadura, além de uma terapêutica combinada com medicações analgésicas e medidas não farmacológicas. Entender a complexidade de alterações fisiopatológicas, psicológicas e bioquímicas apresentadas por um paciente em tratamento de queimadura é o primeiro passo para alcançar o sucesso no seu manejo analgésico.

  15. Objetos mediadores em museus

    Directory of Open Access Journals (Sweden)

    Inês Ferreira

    2015-02-01

    Full Text Available Um museu participativo incentiva a que a experiência da visita seja criativa e pessoal. Facilita a participação biográfica do visitante - com a sua experiência, conhecimento e interesses. Muitas exposições, porém, não promovem esse envolvimento pessoal. Neste artigo apresentamos uma revisão da literatura que indicia que a mediação com objetos mediadores facilita o envolvimento pessoal, a construção de conhecimento e a criatividade. O problema teórico que colocamos é se os museus mais tradicionais se podem abrir à participação e, neste contexto, o nosso objetivo é perceber que papel podem ter os objetos mediadores nesse processo. Discutimos o recurso a objetos mediadores como forma de adicionar múltiplos níveis de relação e modos de ver, potencial que consideramos subaproveitado em muitos museus. Começamos por apresentar o museu como espaço potencialmente participativo, que convida a um envolvimento ativo e crítico – ativo, porque o visitante é desafiado a fazer coisas; crítico, porque é estimulado a pensar, optar, discutir e refletir. Discutimos depois diferentes formas de usar objetos mediadores para incentivar o visitante a um envolvimento com os artefactos, recorrendo à memória, experiência, conhecimento e sentimentos. A participação biográfica potencia o cruzamento entre o mundo interior do visitante e os artefactos e conteúdos do museu. Esse espaço de cruzamento - inbetween - é o espaço da construção de conhecimento. Apresentamos diversas formas de um museu, mesmo não sendo interativo, se tornar mais criativo e participativo, o que pode contribuir para uma mudança nas práticas dos museus, nomeadamente na montagem e remodelação de exposições permanentes.

  16. Acessibilidade web em redes sociais

    OpenAIRE

    Loureiro, Janaína Rolan

    2014-01-01

    A acessibilidade tem sido uma preocupação em diversas áreas nos últimos anos e, em relação à Web, trata-se de um direito garantido por lei às pessoas com deficiência. Muitos estudos visando avaliar e melhorar a Acessibilidade Web foram realizados desde que este direito foi instituído, culminando com o surgimento de tecnologias assistivas, ferramentas de avaliação automatizadas e diretrizes para padronizar o desenvolvimento de sites acessíveis. Porém, alguns domínios ainda c...

  17. Efeito da enxertia em mandioca

    OpenAIRE

    Bomfim, Nayra Nascimento

    2011-01-01

    Enxertos de duas espécies silvestres (M. glaziovii e M. fortalezensis) sobre M. esculenta, mandioca, foram estudados morfologicamente, anatomicamente e citogeneticamente e a fim de se avaliar possíveis efeitos devido à interação entre enxerto e porta-enxerto. Foram feitas análises anatômicas das raízes dos porta-enxertos UnB 201 e UnB 122 (variedades de M. esculenta), em comparação com as variedades não enxertadas. Analisou-se a meiose em enxerto da espécie M. glaziovii que apresentava morfol...

  18. NAVIER-STOKES EM GPU

    OpenAIRE

    ALEX LAIER BORDIGNON

    2006-01-01

    Nesse trabalho, mostramos como simular um fluido em duas dimensões em um domínio com fronteiras arbitrárias. Nosso trabalho é baseado no esquema stable fluids desenvolvido por Joe Stam. A implementação é feita na GPU (Graphics Processing Unit), permitindo velocidade de interação com o fluido. Fazemos uso da linguagem Cg (C for Graphics), desenvolvida pela companhia NVidia. Nossas principais contribuições são o tratamento das múltiplas fronteiras, o...

  19. Factores de risco em implantologia

    OpenAIRE

    Sousa, Inês Silveira e Luz Nunes de

    2015-01-01

    A Implantologia tem ganho popularidade na medicina dentária pelos resultados previsíveis em reabilitações de áreas edêntulas, permitindo minimizar as consequências funcionais, fonéticas e estéticas que advém da perda de dentes através de uma solução fixa que satizfaz os pacientes. Esta é uma revisão da literatura existente sobre factores de risco, fracassos e complicações inerentes ao tratamento com implantes dentários. Categorizam-se os factores de risco em implantologia co...

  20. Thermal Studies of Zn(II, Cd(II and Hg(II Complexes of Some <em>N-Alkyl-N>-Phenyl-Dithiocarbamates

    Directory of Open Access Journals (Sweden)

    Peter A. Ajibade

    2012-07-01

    Full Text Available The thermal decomposition of Zn(II, Cd(II and Hg(II complexes of <em>N-ethyl-N>-phenyl and <em>N-butyl-N>-phenyl dithiocarbamates have been studied using thermogravimetric analysis (TGA and differential scanning calorimetry (DSC. The products of the decomposition, at two different temperatures, were further characterized by scanning electron microscopy (SEM and energy-dispersive X-ray spectroscopy (EDX. The results show that while the zinc and cadmium complexes undergo decomposition to form metal sulphides, and further undergo oxidation forming metal oxides as final products, the mercury complexes gave unstable volatiles as the final product.

  1. Evaluation of Antioxidant Activities of Aqueous Extracts and Fractionation of Different Parts of <em>Elsholtzia em>ciliata>

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    Yuangang Zu

    2012-05-01

    Full Text Available The aim of this study was to investigate the antioxidant and free-radical scavenging activity of extract and fractions from various parts of <em>Elsholtzia ciliataem>. The inflorescences, leaves, stems and roots of <em>E. ciliataem> were extracted separately and two phenolic component enrichment methods: ethyl acetate-water liquid-liquid extraction and macroporous resin adsorption-desorption, were adopted in this study. The antioxidant activities of water extracts and fractions of <em>E. ciliataem> were examined using different assay model systems <em>in vitroem>. The fraction root E (purified by HPD300 macroporous resin exhibited the highest total phenolics content (497.2 ± 24.9 mg GAE/g, accompanied with the highest antioxidant activity against various antioxidant systems <em>in vitroem> compared to other fractions. On the basis of the results obtained, <em>E. ciliataem> extracts can be used potentially as a ready accessible and valuable bioactive source of natural antioxidants.

  2. A Novel Apoptosis Correlated Molecule: Expression and Characterization of Protein Latcripin-1 from <em>Lentinula em>edodes> C91–3

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    Min Huang

    2012-05-01

    Full Text Available An apoptosis correlated molecule—protein Latcripin-1 of <em>Lentinula> edodesem> C91-3—was expressed and characterized in <em>Pichia pastorisem> GS115. The total RNA was obtained from <em>Lentinula edodesem> C91–3. According to the transcriptome, the full-length gene of Latcripin-1 was isolated with 3'-Full Rapid Amplification of cDNA Ends (RACE and 5'-Full RACE methods. The full-length gene was inserted into the secretory expression vector pPIC9K. The protein Latcripin-1 was expressed in <em>Pichia pastorisem> GS115 and analyzed by Sodium Dodecylsulfonate Polyacrylate Gel Electrophoresis (SDS-PAGE and Western blot. The Western blot showed that the protein was expressed successfully. The biological function of protein Latcripin-1 on A549 cells was studied with flow cytometry and the 3-(4,5-Dimethylthiazol-2-yl-2,5-Diphenyl-tetrazolium Bromide (MTT method. The toxic effect of protein Latcripin-1 was detected with the MTT method by co-culturing the characterized protein with chick embryo fibroblasts. The MTT assay results showed that there was a great difference between protein Latcripin-1 groups and the control group (<em>p em>< 0.05. There was no toxic effect of the characterized protein on chick embryo fibroblasts. The flow cytometry showed that there was a significant difference between the protein groups of interest and the control group according to apoptosis function (<em>p em>< 0.05. At the same time, cell ultrastructure observed by transmission electron microscopy supported the results of flow cytometry. The work demonstrates that protein Latcripin-1 can induce apoptosis of human lung cancer cells A549 and brings new insights into and advantages to finding anti-tumor proteins.

  3. Action of Chitosan Against <em>Xanthomonas> Pathogenic Bacteria Isolated from <em>Euphorbia pulcherrimaem>

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    Yanli Wang

    2012-06-01

    Full Text Available The antibacterial activity and mechanism of two kinds of chitosan were investigated against twelve <em>Xanthomonas> strains recovered from <em>Euphorbia pulcherrimaem>. Results indicated that both chitosans markedly inhibited bacterial growth based on OD loss. Furthermore, the release of DNA and RNA from three selected strains was increased by both chitosans. However, the release of intracellular proteins was inhibited by both chitosans at different concentration and incubation times, except chitosan A at 0.1 mg/mL for 0.5 h incubation and 0.2 mg/mL for 2.0 h incubation increased the release of proteins, indicating the complexity of the interaction and cell membranes, which was affected by incubation time, bacterial species, chitosan type and concentration. Transmission electron microscopic observations revealed that chitosan caused changes in protoplast concentration and surface morphology. In some cells, the membranes and walls were badly distorted and disrupted, while other cells were enveloped by a thick and compact ribbon-like layer. The contrary influence on cell morphology may explain the differential effect in the release of material. In addition, scanning electron microscope and biofilm formation test revealed that both chitosans removed biofilm biomass. Overall, this study showed that membrane and biofilm play an important role in the antibacterial mechanism of chitosan.

  4. Allometric and Isometric variations in the Italian <em>Apodemus sylvaticusem> and <em>Apodemus flavicollisem> with respect to the conditions of allopatry and sympatry / Variazioni allometriche e isometriche in <em>Apodemus sylvaticusem> e <em>Apodemus flavicollisem> italiani, rispetto alle condizioni di allopatria e simpatria

    Directory of Open Access Journals (Sweden)

    Giovanni Amori

    1986-12-01

    Full Text Available Abstract In Italy there are two species of <em>Apodemus> (<em>Sylvaemus>: <em>Apodemus sylvaticusem> on the mainland and the main island, and <em>Apodemus flavicollisem> only on the mainland. The trend of some morphometric characters of the skull (incisive foramen length - FI; interorbital breadth = IO; length of palatal bridge = PP; upper alveolar length = $M^1M^3$ was analized and some theoretical models verified for <em>A. sylvaticusem>. If one considers the sympatric population of <em>A. sylvaticusem> and <em>A. flavicollisem> simultaneously the characters PP, IO and $M^1M^3$ appear significantly isometric being directly correlated ($P leq O.O1$, while FI character results allometric with respect to the previous ones, as expected. If one considers the sympatric populations of each of the species separately, the scenario is different. For <em>A. sylvaticusem> only PP and $M^1M^3$ are isometric ($P leq 0.05$. For <em>A. flavicollisem> only $M^1M^3$ and FI appear to be correlated, although not as significantly as for <em>A. sylvaticusem> ($P le 0.05$; one tail. The insular populations of <em>A. sylvaticusem> do not show significant correlations, except for FI and $M^1M^3$ ($P le 0.05$. On the contrary, considering all populations, sympatric and allopatric, of <em>A. sylvaticusem> at the same time are significant correlations ($P le 0.05$ in all combinations of characters, except for those involving the IO. We suggest that the isometric relations in sympatric assemblages are confined within a morphological range available to the genus <em>Apodemus>. In such a space, the two species are split in two different and innerly homogeneous distributions. We found no evidence to confirm the niche variation hypothesis. On the contrary, the variability expressed as SO or CV's appears higher in the sympatric populations than in the allopatric ones, for three of the four characters, confirming previous results

  5. Low Diversity Bacterial Community and the Trapping Activity of Metabolites from Cultivable Bacteria Species in the Female Reproductive System of the Oriental Fruit Fly, <em>Bactrocera dorsalisem> Hendel (Diptera: Tephritidae

    Directory of Open Access Journals (Sweden)

    Hongyu Zhang

    2012-05-01

    Full Text Available Our goal was to identify the bacteria inhabiting the reproductive system of the female oriental fruit fly, <em>Bactrocera dorsalisem> (Hendel, and evaluate the chemotaxis of <em>B>. <em>dorsalis> to the metabolites produced by the bacteria. Based on 16S rRNA-based polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE, 18 operational taxonomic units (OTUs were assigned to the five bacterial classes <em>Betaproteobacteria>, <em>Alphaproteobacteria>, <em>Gammaproteobacteria>, <em>Bacilli> and <em>Actinobacteria>. Nine OTUs were assigned to <em>Gammaproteobacteria>, which was the most highly represented class. <em>Enterobacteriaceae> constituted the dominant family, and within this family, three genera and five species were identified, including <em>Enterobacter sakazakiiem>, <em>Klebsiella> <em>oxytoca>, <em>Klebsiella> <em>pneumoniae,> Raoultellaem> <em>terrigena em>and <em>Enterobacter> <em>amnigenus>. In this set, the first two species were the dominant components, and the latter three species were the minor ones. Finally, we found that the metabolites produced by <em>R.> terrigenaem>, <em>K. oxytocaem> and <em>K.> pneumoniae em>were attractive to the <em>B.> dorsalisem> adults, and in field studies, <em>B.> dorsalisem> adults were most attracted to <em>K.> oxytocaem>. Collectively, our results suggest that the female reproductive system plays an important role in the transfer of enterobacteria from the gut to fruit. Our data may prompt the development of a female-targeted population control strategy for this fly.

  6. Parasitic zoonoses: survey in foxes (<em>Vulpes vulpesem> in the northern Apennines / Zoonosi parassitarie: indagini in volpi (<em>Vulpes vulpesem> dell'Appennino settentrionale

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    Vittorio Guberti

    1991-07-01

    Full Text Available Abstract A parasitological survey on 153 foxes was carried out in the northern Apennines, during the period 1984-1987. The following parasites were identified: <em>Toxocara canisem> (46.4%, <em>Taenia> sp. (17%, <em>Uncinaria stenocephalaem> (11.8%, <em>Mesocestoides lineatusem> (11.1%, <em>Ancylostoma caninumem> (3.9%, <em>Taenia hydatigenaem> (3.3%, <em>Trichuris vulpisem> (3.3%, <em>Dipylidium caninumem> (2.6%, <em>Taenia crassicepsem> (2%. All foxes were negative for <em>Trichinella> sp. A statistical analysis was performed to evaluate differences in the parasitic fauna according to the sex and age classes of the hosts. The role that the fox could have as a reservoir of helminthic zoonoses is discussed. The results are compared with those of similar studies carried out in Italy. Riassunto Nel periodo 1984-1987 è stata condotta un'indagine parassitologica su 153 volpi abbattute nell'Appennino romagnolo. Sono stati reperiti i seguenti parassiti: <em>Toxocara canisem> (46,4%, <em>Taenia> sp. (17%, <em>Uncinaria stenocephalaem> (11,8%, <em>Mesocestoides lineatusem> (11,1%, <em>Ancylostoma caninumem> (3,9%, <em>Taenia hydatigenaem> (3,3%, <em>Trichuris vulpisem> (3,3%, <em>Dipylidium caninumem> (2,6%, <em>Taenia crassicepsem> (2%. Tutte le volpi esaminate sono risultate negative per <em>Trichinella> sp. È stata effettuata l'analisi statistica dei dati per evidenziare eventuali differenze della fauna parassitaria in relazione al sesso e all'età delle volpi. Sulla base dei dati ottenuti viene discussa l'importanza che la Volpe può assumere come serbatoio di zoonosi elmintiche. I risultati acquisiti sono inoltre comparati con quelli ottenuti in analoghe ricerche condotte in Italia.

  7. Fatores de risco cardiovascular em alcoolistas em tratamento

    Directory of Open Access Journals (Sweden)

    Mayla Cardoso Fernandes Toffolo

    2013-06-01

    Full Text Available OBJETIVO: Avaliar a presença de fatores de risco cardiovascular, com ênfase na hipertensão e na adiposidade corporal, em alcoolistas abstinentes ou não abstinentes em tratamento. MÉTODO: Trata-se de um estudo transversal com 65 pacientes alcoolistas em tratamento no CAP-Sad. O grau de dependência do álcool foi avaliado pelo SADD (Short Alcohol Dependence Data e o uso de outras drogas, pelo ASSIST (Alcohol Smoking and Substance Involvement Screening Test. Foram avaliados o perfil bioquímico e o antropométrico dos usuários. RESULTADOS: Participaram do estudo 42 homens e 23 mulheres. A maioria dos participantes (67,74% apresentou dependência alcoólica grave, com uso de álcool associado principalmente a cigarro (66,15%. A média da circunferência da cintura (CC foi significativamente maior entre os abstinentes, em comparação aos não abstinentes (AB: 88,15 ± 15,95 x NA: 81,04 ± 9,86; p = 0,03. Pacientes abstinentes há mais tempo tiveram maior sobrepeso/obesidade e adiposidade abdominal (CC do que os não abstinentes e abstinentes recentes, com razão de chances de 5,25. Os abstinentes apresentaram razão de chances de 3,38 para %GC acima da média, independente do tempo de abstinência. CONCLUSÃO: Pacientes alcoolistas abstinentes apresentam mais sobrepeso/ obesidade, adiposidade corporal (%GC e abdominal (CC do que os não abstinentes. É importante o acompanhamento multiprofissional no tratamento de alcoolistas com abordagem para fatores de risco cardiovasculares, principalmente evitando o ganho de peso.

  8. Enfermagem em cardiologia baseada em evidência

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    Eneida Rejane Rabelo da Silva

    2007-10-01

    Full Text Available Neste artigo apresentamos, dentro do cenário cardiovascular, estudos recentes do nosso grupo. Buscando melhores evidências de enfermagem investigamos por meio de ensaios clínicos randomizados (ECR estratégias de educação e acompanhamento destes pacientes (pctes. O primeiro ECR avaliou o impacto da educação individual e da orientação em grupo no conhecimento sobre doença e autocuidado em pctes com insuficiência cardíaca (IC. No grupo intervenção, houve uma melhora do conhecimento dos cuidados essenciais (controle de sal e da ingesta hídrica, peso diário, realização de atividade física P=0,008, demonstrando que o atendimento em grupo é semelhante ao individual. No ambiente hospitalar, segundo ECR, comparamos o efeito da educação de enfermagem e seguimento telefônico após alta versus educação sem a monitorização por telefone. Demonstramos que intervenção educativa durante a internação melhora o conhecimento da doença, autocuidado e qualidade de vida (P

  9. Conservazione e gestione della Lepre italica (<em>Lepus corsicanusem>

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    Francesco Riga

    2003-10-01

    Full Text Available Il recente riconoscimento dello <em>status> specifico della Lepre italica (<em>Lepus corsicanusem> e l?accertamento dell?areale distributivo rappresentano le azioni più importanti per la conservazione di un <em>taxon endemicoem> che si era creduto estinto. Nella penisola la specie presenta un areale discontinuo, il cui limite settentrionale è dato dal comune di Manciano (GR, sul versante tirrenico e da una linea che dalla provincia de L'Aquila arriva al Gargano. In Sicilia la distribuzione è relativamente continua anche in aree non protette. Dati genetici hanno permesso di confermare la presenza in Corsica. Al contrario, nell?Isola d'Elba, a seguito di estese ricerche, sono stati identificati solo esemplari di <em>L. europaeusem>. Nell?Italia peninsulare <em>L. corsicanusem> è spesso presente in simpatria con popolazioni di <em>L. europaeusem>, mentre in Sicilia la lepre europea non ha originato popolazioni stabili, nonostante l?immissione di molte migliaia di individui. La distribuzione ecologica di <em>L. corsicanusem> ed analisi ambientali specifiche, suggeriscono l?adattamento prevalente agli ambienti a clima mediterraneo, benché essa sia presente anche a quote elevate (> 1.500 m s.l.m.. Dati preliminari di abbondanza relativa hanno evidenziato una situazione diversificata tra la penisola e la Sicilia e tra aree a diverso regime di gestione; un confronto tra le aree protette ha evidenziato rispettivamente valori di 5,54 e 11,73 ind./km². La riduzione quali-quantitativa e la frammentazione dell?<em>habitat> delle lepri è un fenomeno potenzialmente pericoloso per la sopravvivenza delle popolazioni, determinando fenomeni di estinzione locale dovuti alle basse densità di popolazione, inducendo fenomeni di erosione della variabilità genetica e di riduzione della <em>fitness> degli individui. L?introduzione di <em>L. europaeusem> può costituire un importante fattore limitante sia per la possibile competizione

  10. Identification of <em>Colletotrichum> species causing anthracnose on Tahiti lime, tree tomato and mango

    Directory of Open Access Journals (Sweden)

    Martínez Erika P.

    2009-08-01

    Full Text Available

    In Colombia, citrus, tree tomato and mango crops are likely to suffer considerable losses from anthracnose caused by several <em>Colletotrichum> species, which were identified by the present study on infected organs of the three fruit crops, sampled in different regions of the country. Identification was based on their morphological and molecular characteristics, as well as on fungicide (benomyl and copper hydroxide sensitivity and pathogenicity tests. The latter assessed infectivity on both the original hosting crop and the other two crops (crossed infection, by putting the fungi in contact with organs taken from the three fruit crops. Molecular identification of the <em>Colletotrichum> species was carried out through amplification of rDNA ITS regions by means of <em>C. gloeosporioidesem> (CgInt and <em>C. acutatumem> (CaInt2 specific primer PCR combining the use of ITS4 universal primer. The results indicate that <em>C. acutatumem> is the infectious agent in Tahiti lime and tree tomato, and so is <em>C. gloeosporioidesem> in mango. Although <em>C. acutatumem> is the infectious agent in two diferent fruit species, the strains proved to be specific of their original hosts.

  11. <em>Ipomoea aquaticaem> Extract Shows Protective Action Against Thioacetamide-Induced Hepatotoxicity

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    A. Hamid A. Hadi

    2012-05-01

    Full Text Available In the Indian system of traditional medicine (Ayurveda it is recommended to consume <em>Ipomoea em>aquatica> to mitigate disorders like jaundice. In this study, the protective effects of ethanol extract of <em>I. aquaticaem> against liver damage were evaluated in thioacetamide (TAA-induced chronic hepatotoxicity in rats. There was no sign of toxicity in the acute toxicity study, in which Sprague-Dawley (SD rats were orally fed with <em>I. aquaticaem> (250 and 500 mg/kg for two months along with administration of TAA (i.p injection 200 mg/kg three times a week for two months. The results showed that the treatment of <em>I. aquaticaem> significantly lowered the TAA-induced serum levels of hepatic enzyme markers (ALP, ALT, AST, protein, albumin, bilirubin and prothrombin time. The hepatic content of activities and expressions SOD and CAT that were reduced by TAA were brought back to control levels by the plant extract supplement. Meanwhile, the rise in MDA level in the TAA receiving groups also were significantly reduced by <em>I. aquaticaem> treatment. Histopathology of hepatic tissues by H&E and Masson trichrome stains displayed that <em>I. aquaticaem> has reduced the incidence of liver lesions, including hepatic cells cloudy swelling, infiltration, hepatic necrosis, and fibrous connective tissue proliferation induced by TAA in rats. Therefore, the results of this study show that the protective effect of <em>I. aquaticaem> in TAA-induced liver damage might be contributed to its modulation on detoxification enzymes and its antioxidant and free radical scavenger effects. Moreover, it confirms a scientific basis for the traditional use of <em>I. aquaticaem> for the treatment of liver disorders.

  12. Culturas orais em sociedades letradas

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    Geraldi João Wanderley

    2000-01-01

    Full Text Available Na relação entre linguagem e cultura, um aspecto essencial diz respeito às diferentes funções da escrita e às formas de sua apropriação social em sociedades letradas. Tomando como inspiração a forma de um uso específico da escrita pelo povo indígena Jarawara e narrativas de ribeirinhos do rio Madeira do mito amazônico Cobra Norato, são discutidas as diferentes relações entre oralidade e escrita que, atravessadas pelas relações de poder, transformam o alfabeto, de artefato histórico construído para o registro da oralidade, em espaço social de construção de distinções, clivagens e exclusões.

  13. Paratuberculose em ruminantes no Brasil

    OpenAIRE

    Yamasaki,Elise M.; Brito,Marilene F.; Mota,Rinaldo A.; McIntosh,Douglas; Tokarnia,Carlos H.

    2013-01-01

    A paratuberculose ou doença de Johne é uma enterite granulomatosa causada por Mycobacterium avium subsp. paratuberculosis (Map) e comumente afeta ruminantes domésticos, no entanto, pode infectar várias espécies de mamíferos. Está presente nos cinco continentes e é considerada endêmica em algumas regiões pela Organização Internacional de Epizootias (OIE). Pertence à lista de enfermidades notificáveis, que compreende as doenças transmissíveis de importância sócio-econômica e/ou em saúde-pública...

  14. COMPETÊNCIA EM MEDICINA

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    Hélio Teixeira MD.

    2005-01-01

    Full Text Available Competência no exercício da medicina depende de um processo evolutivo ao longo da vida do profissional, processo este fundamentado em eficiência, experiência e em princípios éticos. A eficiência depende da cultura médica, de habilidades técnicas e relacionamento interpessoal com o paciente. A experiência ao longo da vida vai refinando o médico, favorecendo o evolver do conhecimento e da sabedoria vivencial. Finalmente, a ética médica se fundamenta na justiça, compaixão e amor ao próximo. Didaticamente, distinguimos 3 fases da evolução profissional do médico: a infância profissional ou visão linear: o médico se restringe em geral ao atendimento da doença, negligenciando o doente. É quase exclusivamente técnico e tem percepção acanhada da medicina-arte. b Maturidade profissional ou visão humanística: resultado de evolução da personalidade, da cultura e da experiência do médico, agora voltado para o doente com sua doença. c Excelência profissional ou visão holística: estágio máximo que se pode alcançar, quando o médico vislumbra a si e ao seu cliente em todas suas dimensões integradas e atua como mestre, procurando despertar no paciente condições próprias de cura, oriundas de sua essência. Conclui-se que exercer medicina com competência significa desempenhar adequadamente a ciência e a arte médica.

  15. Phytochemical Composition, Antioxidant and Xanthine Oxidase Inhibitory Activities of <em>Amaranthus cruentusem> L. and <em>Amaranthus hybridusem> L. Extracts

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    Jeanne F. Millogo

    2012-06-01

    Full Text Available This paper describes a preliminary assessment of the nutraceutical value of <em>Amaranthus cruentus (A. cruentusem> and <em>Amaranthus hybridus (A. hybridusem>, two food plant species found in Burkina Faso. Hydroacetonic (HAE, methanolic (ME, and aqueous extracts (AE from the aerial parts were screened for <em>in vitroem> antioxidant and xanthine oxidase inhibitory activities. Phytochemical analyses revealed the presence of polyphenols, tannins, flavonoids, steroids, terpenoids, saponins and betalains. Hydroacetonic extracts have shown the most diversity for secondary metabolites. The TLC analyses of flavonoids from HAE extracts showed the presence of rutin and other unidentified compounds. The phenolic compound contents of the HAE, ME and AE extracts were determined using the Folin–Ciocalteu method and ranged from 7.55 to 10.18 mg Gallic acid equivalent GAE/100 mg. Tannins, flavonoids, and flavonols ranged from 2.83 to 10.17 mg tannic acid equivalent (TAE/100 mg, 0.37 to 7.06 mg quercetin equivalent (QE /100 mg, and 0.09 to 1.31 mg QE/100 mg, respectively. The betacyanin contents were 40.42 and 6.35 mg Amaranthin Equivalent/100 g aerial parts (dry weight in <em>A. cruentusem> and <em>A. hybridusem>, respectively. Free-radical scavenging activity expressed as IC50 (DPPH method and iron reducing power (FRAP method ranged from 56 to 423 µg/mL and from 2.26 to 2.56 mmol AAE/g, respectively. Xanthine oxidase inhibitory activities of extracts of <em>A. cruentus em>and <em>A. hybridusem> were 3.18% and 38.22%, respectively.<em> em>The> A. hybridusem> extract showed the best antioxidant and xanthine oxidase inhibition activities. The results indicated that the phytochemical contents of the two species justify their traditional uses as nutraceutical food plants.

  16. Existe sujeito em Michel Maffesoli?

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    Marli Appel da Silva

    2010-06-01

    Full Text Available Este ensaio discute a concepção de sujeito na abordagem teórica de Michel Maffesoli. As ideias desse autor estão em voga em alguns meios acadêmicos no Brasil e são difundidas por algumas mídias de grande circulação nacional. Entretanto, ao longo de suas obras, os pressupostos que definem quem é o sujeito maffesoliano se encontram pouco clarificados. Portanto, para alcançar o objetivo a que se propõe, este ensaio desenvolve uma análise da epistemologia e da ontologia maffesoliana com a finalidade de compreender as origens dos pressupostos desse autor, ou seja, as teorias e os autores em que Maffesoli se baseou para desenvolver uma visão de sujeito. Com essa compreensão, pretende-se responder à questão: existe sujeito na abordagem teórica de Maffesoli.

  17. Complaints against an EMS system.

    Science.gov (United States)

    Colwell, Christopher B; Pons, Peter T; Pi, Randy

    2003-11-01

    Complaints against Emergency Medical Services (EMS) agencies represent a concerning and potentially time-consuming problem for all involved in the delivery of prehospital emergency medical care. The objective of this study was to identify the source of complaints against an EMS system to help focus quality and performance improvement and customer service efforts. We conducted a retrospective review of complaints filed against a busy urban EMS agency over a 6-year period. All complaints were included, totaled by season and by year, and categorized by originator and nature of the complaint. A total of 286 complaints were registered during the 6-year period, with an average of 48 per year and 9.3 per 10,000 responses. The most common originators of complaints were patients (53%) followed by medical personnel (19%) and family members or friends (12%). Rude behavior accounted for 23% of the complaints registered, followed by technical skills (20%), transport problems (18%), and loss of belongings (13%). The identification of areas of dissatisfaction will allow focused quality and performance improvement programs directed at customer service and risk management.

  18. Adding large EM stack support

    KAUST Repository

    Holst, Glendon

    2016-12-01

    Serial section electron microscopy (SSEM) image stacks generated using high throughput microscopy techniques are an integral tool for investigating brain connectivity and cell morphology. FIB or 3View scanning electron microscopes easily generate gigabytes of data. In order to produce analyzable 3D dataset from the imaged volumes, efficient and reliable image segmentation is crucial. Classical manual approaches to segmentation are time consuming and labour intensive. Semiautomatic seeded watershed segmentation algorithms, such as those implemented by ilastik image processing software, are a very powerful alternative, substantially speeding up segmentation times. We have used ilastik effectively for small EM stacks – on a laptop, no less; however, ilastik was unable to carve the large EM stacks we needed to segment because its memory requirements grew too large – even for the biggest workstations we had available. For this reason, we refactored the carving module of ilastik to scale it up to large EM stacks on large workstations, and tested its efficiency. We modified the carving module, building on existing blockwise processing functionality to process data in manageable chunks that can fit within RAM (main memory). We review this refactoring work, highlighting the software architecture, design choices, modifications, and issues encountered.

  19. Paratuberculose em ruminantes no Brasil

    Directory of Open Access Journals (Sweden)

    Elise M. Yamasaki

    2013-02-01

    Full Text Available A paratuberculose ou doença de Johne é uma enterite granulomatosa causada por Mycobacterium avium subsp. paratuberculosis (Map e comumente afeta ruminantes domésticos, no entanto, pode infectar várias espécies de mamíferos. Está presente nos cinco continentes e é considerada endêmica em algumas regiões pela Organização Internacional de Epizootias (OIE. Pertence à lista de enfermidades notificáveis, que compreende as doenças transmissíveis de importância sócio-econômica e/ou em saúde-pública, cujo controle é necessário para o comércio internacional de animais e alimentos de origem animal. A importância da doença de Johne não se restringe somente aos prejuízos econômicos causados à indústria animal, mas também na possível participação do Map na íleocolite granulomatosa que afeta seres humanos, conhecida como doença de Crohn. No Brasil, a paratuberculose já foi descrita em diversas espécies de ruminantes e em vários estados. Embora os relatos naturais da enfermidade sejam pontuais, acredita-se na possibilidade da transmissão interespecífica e na disseminação do agente através da compra e venda de animais infectados. O objetivo deste artigo foi reunir as informações disponíveis referentes aos aspectos epidemiológicos, clínico-patológicos e laboratoriais da paratuberculose em bovinos, bubalinos, caprinos e ovinos no Brasil, e salientar a necessidade de implementação de medidas de controle sanitário da enfermidade no país, o que possibilitaria a melhoria da qualidade e valorização dos produtos de origem animal no mercado internacional.

  20. Ultrasound-Assisted Extraction of Carnosic Acid and Rosmarinic Acid Using Ionic Liquid Solution from <em>Rosmarinus officinalisem>>

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    Chunjian Zhao

    2012-09-01

    Full Text Available Ionic liquid based, ultrasound-assisted extraction was successfully applied to the extraction of phenolcarboxylic acids, carnosic acid and rosmarinic acid, from <em>Rosmarinus officinalisem>. Eight ionic liquids, with different cations and anions, were investigated in this work and [C8mim]Br was selected as the optimal solvent. Ultrasound extraction parameters, including soaking time, solid–liquid ratio, ultrasound power and time, and the number of extraction cycles, were discussed by single factor experiments and the main influence factors were optimized by response surface methodology. The proposed approach was demonstrated as having higher efficiency, shorter extraction time and as a new alternative for the extraction of carnosic acid and rosmarinic acid from <em>R. officinalisem>> em>compared with traditional reference extraction methods. Ionic liquids are considered to be green solvents, in the ultrasound-assisted extraction of key chemicals from medicinal plants, and show great potential.

  1. Trophic systems and chorology: data from shrews, moles and voles of Italy preyed by the barn owl / Sistemi trofici e corologia: dati su <em>Soricidae>, <em>Talpidae> ed <em>Arvicolidae> d'Italia predati da <em>Tyto albaem> (Scopoli 1769

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    Longino Contoli

    1986-12-01

    Full Text Available Abstract In small Mammals biogeography, available data are up to now by far too scanty for elucidate the distribution of a lot of taxa, especially with regard to the absence from a given area. In this respect, standardized quantitative sampling techniques, like Owl pellets analysis can enable not only to enhance faunistic knowledges, but also to estimate the actual absence probability of a given taxon "m", lacking from the diet of an individual raptor. For the last purpose, the relevant frequencies of "m" in the other ecologically similar sites of the same raptor species diets are averaged ($f_m$ : the relevant standard error (multiplicated by a coefficient, according to the desired degree of accuracy, in relation of the integral of probabilities subtracted ($overline{F}_m - a E$: then, the probability that a single specimen is not pertaining to "m" is obtained ($P_0 = 1 - F_m + a E$; lastly, the desiderate accuracy probability ($P_d$ is chosen. Now, "$N_d$" (the number of individuals of all prey species in a single site needed for obtain, with the desired probability, a specimen at least of "m" is obtained through $$N = frac{ln P_d}{ln P_0}$$ Obviously, every site-diet with more than "N" preyed individuals and without any "i" specimen is considered to be lacking of such taxon. A "usefulness index" for the above purposes is outlined and checked about three raptors. Some exanples about usefulness of the Owl pellet analysis method in biogeography are given, concerning <em>Tyto albaem> diets in peninsular Italy about: - <em>Sorex minutusem>, lacking in some quite insulated areas; - <em>Sorex araneusem> (sensu stricto, after GRAF et al., 1979, present also in lowland areas in Emilia-Romagna; - <em>Crocidura suaveolensem> and - <em>Suncus etruscusem>, present also in the southermost part of Calabria (Reggio province; - <em>Talpa caecaem>, present also in the Antiapennines of Latium (Cimini mounts; - <em>Talpa romanaem

  2. Tumores em bovinos encontrados em abatedouros frigoríficos

    Directory of Open Access Journals (Sweden)

    Bianca Tessele

    2016-03-01

    Full Text Available Resumo: Numa pesquisa em abatedouros a procura de lesões em bovinos, realizada de janeiro de 2011 a julho de 2014, 544 foram encontradas, das quais 65 eram neoplasmas. Quarenta e dois porcento desses tumores eram de origem mesenquimal; 37% eram epiteliais; 14,5% eram derivados da crista neural; 5% eram tumores do cordão sexual; e 1,5 eram tumores originários do sistema nervoso periférico. O tumor mais frequentemente encontrado foi o linfoma (28% de todos os tumores, a maioria dos casos como parte do complexo leucose bovina enzoótica. O carcinoma de células escamosas foi o segundo tumor mais frequente (15% de todos os tumores. É chamada a atenção para a frequência desses tumores e para a sua importância no diagnóstico diferencial no abatedouro frigorífico entre eles e outras lesões importantes, incluindo as lesões granulomatosas da tuberculose. Houve uma ocorrência significativa do feocromocitoma adrenal (13% de todos os tumores. Papilomatose representou apenas 3% de todos os tumores. Como papilomas são comuns em bovinos, seu baixo número nesse estudo pode ser explicado pelo fato de que eles não são usualmente diagnosticados no exame post mortem (quando o couro já foi retirado da carcaça, mas sim no exame ante mortem, como ocorreu na maioria dos casos deste estudo. Tumores encontrados com menor frequência (cada um perfazendo entre 1.5 e 3% de todos os tumores incluíram adenocarcinoma apócrino misto da cauda, adenocarcinoma mamário, adenocarcinoma uterino, carcinoma de células renais, fibroma interdigital, hemangiossarcoma, leiomioma uterino, lipoma, lipossarcoma, mesotelioma, neurofibroma, tumor de células da granulosa, tumor estromal gastrointestinal, tumores hepáticos, tumores melanocíticos, e tumores pulmonares primários. Pretende-se que os resultados dessa pesquisa possam ajudar na identificação das lesões na inspeção oficial em matadouros frigoríficos.

  3. Variabilità morfologica ed ecologica in <em>Neomys fodiensem> e <em>Neomys anomalusem> nell'Appennino settentrionale

    Directory of Open Access Journals (Sweden)

    Dino Scaravelli

    2003-10-01

    Full Text Available I due <em>Neomys> italiani sono ancora da chiarire dal punto di vista della loro caratterizzazione morfologica e ecologica. Il lavoro prende in considerazione un campione di entrambe le specie proveniente da habitat forestali dell?Appennino settentrionale per i quali sono stati identificati i principali parametri ambientali. Vengono quindi descritte la variabilità dei tratti morfologici delle due specie in aree localizzate nel Parco Nazionale Foreste Casentinesi, Monte Falterona e Campigna nell?Appennino tosco-romagnolo. Risultano di sicuro effetto discriminatorio la maschera facciale, il rapporto piede posteriore/coda e i caratteri cranici. Sulla base dei criteri identificativi si sono realizzate rilevazioni di misure corporee per le due specie e una comparazione degli habitat utilizzati. <em>N. fodiensem> appare unica specie nelle faggete-abetine e dominante nei castagneti, mentre nell?Ontaneta e nelle zone aperte e termofile si registra la sola presenza di <em>N. anomalusem>. Mancano entrambi nei prati cespugliati, nella pecceta e nella cerreta. I gradienti presenti sono quindi illustrati. Non appare una differenza altitudinale nel campione esaminato, posto in stazioni tra i 400 e i 1300 m, ma per entrambe vi sono maggiori riscontri nella fascia tra 700 e 850 m. Nell?analisi multivariata rispetto alle altre specie e alle variabili ambientali si riscontra sempre una discreta correlazione con la presenza di acqua di una certa ampiezza, che comunque è significativa solo per <em>N. fodiensem>, mentre risulta di interesse la positiva correlazione di <em>N. anomalusem> con <em>Apodemus sylvaticusem>.

  4. Morphological identification of the Soprano Pipistrelle (<em>Pipistrellus pygmaeusem> Leach, 1825 in Croatia.

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    Igor Pavlinić

    2008-07-01

    Full Text Available Abstract After the discovery of two different phonic types within the common pipistrelle (<em>Pipistrellus pipistrellusem>, mtDNA analysis confirmed the existence of two separate species named as common pipistrelle (<em>P. pipistrellusem> and soprano pipistrelle (<em>P. pygmaeusem>. The discrimination of these two cryptic species using external characters and measures has proved to be somewhat problematic. We examined two colonies of soprano pipistrelle from Donji Miholjac, Croatia. As a result, only two characters proved to be of help for field identification: wing venation (89% of cases and penis morphology and colour for males. The difference in length between the 2nd and 3rd phalanxes of the 3rd finger should be discarded as diagnostic trait between <em>P. pipistrellusem> and <em>P. pygmaeusem> in Croatia. Riassunto Identificazione su basi morfologiche del pipistrello pigmeo (<em>Pipistrellus pygmeausem>, Leach, 1825 in Croazia. A seguito della descrizione di due differenti "tipi fonici" nel pipistrello nano (<em>Pipistrellus pipistrellusem> e della successiva conferma su basi genetiche dell'esistenza di due specie distinte, designate come pipistrello nano (<em>P. pipistrellusem> e pipistrello pigmeo (<em>P. pygmaeusem>, la distinzione delle due specie in base a caratteristiche morfologiche esterne si è dimostrata un problema di difficile soluzione. Sulla base delle caratteristiche distintive e delle differenze biometriche proposte da altri Autori, sono state esaminate due colonie di pipistrello pigmeo a Donji Miholjac, in Croazia. I risultati ottenuti evidenziano che, tra tutti i potenziali caratteri sinora proposti, solo due risultano utili per un'identificazione diretta sul campo: la venatura delle ali, risultata utile alla discriminazione nell'89% degli esemplari analizzati, e la colorazione e morfologia del pene nei maschi. La

  5. Historicidade da filosofia em Hegel e Heidegger

    OpenAIRE

    Christian Iber

    2017-01-01

    Gostaria de discutir, em forma de teses, a temática da historicidade da filosofia em contraste e na sua divergência em Hegel e Heidegger. Diferente do historicismo, que não soluciona o problema da historicidade da filosofia, porque ele dissolve tendencialmente a filosofia na história da filosofia e não coloca em especial a questão da verdade, Hegel e Heidegger desenvolvem uma teoria genuinamente filosófica da história da filosofia do interesse filosófico pela verdade. Esse interesse genuiname...

  6. Encapsulation-Induced Stress Helps <em>Saccharomyces cerevisiae em>Resist Convertible Lignocellulose Derived Inhibitors

    Directory of Open Access Journals (Sweden)

    Johan O. Westman

    2012-09-01

    Full Text Available The ability of macroencapsulated <em>Saccharomyces cerevisiae em>CBS8066<em> em>to withstand readily and not readily <em>in situem> convertible lignocellulose-derived inhibitors was investigated in anaerobic batch cultivations. It was shown that encapsulation increased the tolerance against readily convertible furan aldehyde inhibitors and to dilute acid spruce hydrolysate, but not to organic acid inhibitors that cannot be metabolized anaerobically. Gene expression analysis showed that the protective effect arising from the encapsulation is evident also on the transcriptome level, as the expression of the stress-related genes <em>YAP1em>, <em>ATR1em> and <em>FLR1em> was induced upon encapsulation. The transcript levels were increased due to encapsulation already in the medium without added inhibitors, indicating that the cells sensed low stress level arising from the encapsulation itself. We present a model, where the stress response is induced by nutrient limitation, that this helps the cells to cope with the increased stress added by a toxic medium, and that superficial cells in the capsules degrade convertible inhibitors, alleviating the inhibition for the cells deeper in the capsule.

  7. A Favela posta em Cena

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    Fabiana Valdoski Ribeiro

    2010-08-01

    As publicações do Observatório das Favelas nos instigam na construção de um espaço transgressor Possível como necessidade de uma ação urgente, contudo a perspectiva do Impossível como projeto nos promove a revelar os limites, as normas e as ideologias que perpassam as ações internas ao Estado, já que este se constitui agente fundamental da engrenagem da produção de desigualdades e em sua força legitimadora realiza a ideologia mais eficiente.

  8. Maquinas virtuais em ambientes seguros

    OpenAIRE

    Arthur Bispo de Castro

    2006-01-01

    Resumo: Desde o início da computação a idéia de máquinas virtuais vem sendo aplicada para estender o multiprocessamento, multi-programação e multi-acesso, tornando os sistemas multi-ambiente. O contínuo aumento no poder de processamento dos computadores fez com que máquinas muito rápidas estivessem ao alcance de qualquer usuário, surgindo PCs com processamento, espaço em disco e memória suficiente para comportar mais de um sistema compartilhando o mesmo hardware. Basicamente, o objetivo das m...

  9. A metrologia em diferentes equipamentos

    OpenAIRE

    Gomes, Andreia Oliveira

    2017-01-01

    A seleção de um equipamento metrológico pode tornar-se uma difícil tarefa, dado que hoje em dia existe uma grande variedade. Quando se pretende adquirir ou até mesmo apenas saber qual o melhor equipamento para o controlo dimensional de um ou mais objetos, a opção pode não ser clara devido às inúmeras características associadas. Este trabalho tem como objetivo o desenvolvimento de uma metodologia que permita a qualquer pessoa com conhecimentos mínimos, saber o que há-de escolher, com bas...

  10. A melancolia em Al Berto

    OpenAIRE

    Camila Emanuele Martins de Souza

    2015-01-01

    Este trabalho objetiva estudar, sob a perspectiva da melancolia, os livros Salsugem (1984) e Horto de Incêndio (1997), de Al Berto. O autor português produz escritos em que a temática da melancolia atravessa a palavra, o que justifica o debruçar-se sobre as construções textuais da poética albertiana, a fim de traçar-lhe novas possibilidades de leitura por esse ângulo. Para isso, com o intuito de compreender o discurso melancólico segue-se o embasamento teórico: de Klibansky, Panosfsky e Saxl ...

  11. Altitudinal distribution of the common longeared bat <em>Plecotus auritusem> (Linnaeus, 1758 and grey long-eared bat <em>Plecotus austriacusem> (J. B. Fischer, 1829 (Chiroptera, Vespertilionidae in the Tatra mountains (southern Poland

    Directory of Open Access Journals (Sweden)

    Krzysztof Piksa

    2006-03-01

    Full Text Available Riassunto Distribuzione altitudinale di Orecchione bruno (<em>Plecotus auritusem> e Orecchione meridionale (<em>Plecotus austriacusem> nei Monti Tatra (Polonia meridionale. Vengono riportati nuovi dati relativi alla distribuzione altitudinale nei Monti Tatra (Polonia meridionale di <em>Plecotus auritusem> e <em>P. austriacusem>. Tali segnalazioni incrementano le conoscenze relative alla presenza di questi chirotteri a quote elevate, in particolare per la Polonia. In inverno <em>P. auritusem> è stato rinvenuto a 1921 m s.l.m. mentre in estate è stato rinvenuto a 2250 m s.l.m.; in aggiunta, sono stati ritrovati resti ossei a 1929 m s.l.m. <em>P. austriacusem> è stato segnalato in ibernazione a 1294 m s.l.m.

  12. Estratégia e estrutura: em busca do alinhamento organizacional em um clube social esportivo

    OpenAIRE

    Cruz Junior,Augusto de Toledo; Carvalho,Marly Monteiro de; Laurindo,Fernando José Barbin

    2005-01-01

    A literatura descreve tipologias de formas organizações, que podem ser divididas em organização baseada em trabalho e organização baseada em conhecimento. A forma de representação mais utilizada é o organograma. Contudo, esta forma de representação não consegue ilustrar adequadamente a dinâmica de funcionamento da organização, sobretudo em organizações em que o conhecimento e a experiência são os ativos-chave. Com o objetivo de investigar o processo de alinhamento entre estratégia e estrutura...

  13. Dualismos em duelo Dueling dualism

    Directory of Open Access Journals (Sweden)

    Anne Fausto-Sterling

    2002-01-01

    Full Text Available Os modos europeus e norte-americanos de entender como funciona o mundo dependem em grande parte do uso de dualismos - pares de conceitos, objetos ou sistemas de crenças opostos. Este ensaio enquadra especialmente três deles: sexo/gênero, natureza/criação e real/construído. Embora este texto verse sobre gênero, discuto regularmente o modo como as idéias de raça e gênero surgem a partir de supostos subjacentes sobre a natureza física do corpo. Entender como operam raça e gênero - em conjunto e independentemente - nos ajuda a compreender melhor como o social se torna corporificado.Euro-American ways of understanding how the world works depend heavily on the use of dualisms - pairs of opposing concepts, objects, or belief sytems. This text focuses especially on three of these: sex/gender, nature/nurture, and real/constructed. And although this text focuses on gender, I regularly discuss the ways in which the ideas of both race and gender emerge from underlying assumptions about the body's physical nature. Understanding how race and gender work - together and independently - help us learn more about how the social becomes embodied.

  14. Laddering em pesquisa de marketing

    Directory of Open Access Journals (Sweden)

    Tânia Modesto Veludo-de-Oliveira

    Full Text Available O artigo discute a técnica de pesquisa laddering em marketing, considerando a proposta de renovação de conhecimentos sobre metodologia na área. Para isso investiga diversos estudos, artigos e pesquisas que tratam do assunto em nível nacional e internacional. O conceito de cadeias meios-fins é abordado pela estrutura teórica que fundamenta a técnica, ligando atributos, conseqüências percebidas do consumo e valores pessoais relativos a um produto. O artigo segue com a explicação dos procedimentos de campo, como a entrevista e a análise dos dados. A laddering é um instrumento de pesquisa qualitativa bastante útil e poderoso, mas ainda pouco utilizado tanto por acadêmicos como por profissionais de mercado, no Brasil, provavelmente por desconhecimento e pouca divulgação. Cabe, portanto, uma maior disseminação de seu uso.

  15. Dois funcionalistas franceses em confronto

    Directory of Open Access Journals (Sweden)

    Rafael Eugenio Hoyos-Andrade

    2001-02-01

    Full Text Available

    Comparamos, neste artigo, de modo aproximativo e provisório, dois modelos de análise sintática e dois métodos de representação gráfica dessas análises: trata-se dos modelos e métodos de André Martinet e Lucien Tesnière. Depois de algumas considerações introdutórias relativas ao funcionalismo dos autores em causa, arrolamos um conjunto de características que aproximam ou opõem os seus respectivos modelos. Apresentamos, em seguida, no intuito de beneficiar os estudantes universitários de sintaxe, as representações gráficas de um mesmo enunciado segundo o método estemático de Tesnière e segundo o método de visualização das relações sintáticas de Martinet, para concluir que os dois modelos/métodos, embora diferentes, poderiam eventualmente ser integrados num único sistema de análise e representação.

  16. Compositions and Methods for the Treatment of Pierce's Disease

    Energy Technology Data Exchange (ETDEWEB)

    Gupta, Goutam (Santa Fe, NM)

    2008-10-07

    Chimeric anti-microbial proteins, compositions, and methods for the therapeutic and prophylactic treatment of plant diseases caused by the bacterial pathogen Xylella fastidiosa are provided. The anti-microbial proteins of the invention generally comprise a surface recognition domain polypeptide, capable of binding to a bacterial membrane component, fused to a bacterial lysis domain polypeptide, capable of affecting lysis or rupture of the bacterial membrane, typically via a fused polypeptide linker. In particular, methods and compositions for the treatment or prevention of Pierce's disease of grapevines are provided. Methods for the generation of transgenic Vitus vinefera plants expressing xylem-secreted anti-microbial chimeras are also provided.

  17. PEG Enhancement for EM1 and EM2+ Missions

    Science.gov (United States)

    Von der Porten, Paul; Ahmad, Naeem; Hawkins, Matt

    2018-01-01

    NASA is currently building the Space Launch System (SLS) Block-1 launch vehicle for the Exploration Mission 1 (EM-1) test flight. The next evolution of SLS, the Block-1B Exploration Mission 2 (EM-2), is currently being designed. The Block-1 and Block-1B vehicles will use the Powered Explicit Guidance (PEG) algorithm. Due to the relatively low thrust-to-weight ratio of the Exploration Upper Stage (EUS), certain enhancements to the Block-1 PEG algorithm are needed to perform Block-1B missions. In order to accommodate mission design for EM-2 and beyond, PEG has been significantly improved since its use on the Space Shuttle program. The current version of PEG has the ability to switch to different targets during Core Stage (CS) or EUS flight, and can automatically reconfigure for a single Engine Out (EO) scenario, loss of communication with the Launch Abort System (LAS), and Inertial Navigation System (INS) failure. The Thrust Factor (TF) algorithm uses measured state information in addition to a priori parameters, providing PEG with an improved estimate of propulsion information. This provides robustness against unknown or undetected engine failures. A loft parameter input allows LAS jettison while maximizing payload mass. The current PEG algorithm is now able to handle various classes of missions with burn arcs much longer than were seen in the shuttle program. These missions include targeting a circular LEO orbit with a low-thrust, long-burn-duration upper stage, targeting a highly eccentric Trans-Lunar Injection (TLI) orbit, targeting a disposal orbit using the low-thrust Reaction Control System (RCS), and targeting a hyperbolic orbit. This paper will describe the design and implementation of the TF algorithm, the strategy to handle EO in various flight regimes, algorithms to cover off-nominal conditions, and other enhancements to the Block-1 PEG algorithm. This paper illustrates challenges posed by the Block-1B vehicle, and results show that the improved PEG

  18. Terapia breve em hospital fechado

    Directory of Open Access Journals (Sweden)

    Paulo Cesar Sandler

    1978-09-01

    Full Text Available O presente estudo cuida de pacientes "difíceis" da clínica psiquiátrica e psicoterápica: os pacientes que apresentam o fenômeno da atuação, cuja aproximação terapêutica tem se demonstrado problemática, mesmo com o advento das modernas técnicas de psicoterapia e das comunidades terapêuticas. Os diagnósticos psiquiátricos destes pacientes variam: são neuróticos, apresentam transtornos de pesonalidade, são psicóticos e quase sempre com graves doenças secundárias: alcoolismo e toxicomania (Códigos Classificação Internacional de Doenças 295 a 299, 300. 301, 303 e 304. Propõe-se, à luz da compreensão psicanalítica destas pessoas, um tratamento psicoterápico breve, focal, com o auxílio de uma hospitalização em regime fechado. Para a avaliação dos resultados do método psicoterápico diádico, criaram-se critérios de cura coerentes com a proposta terapêutica, e equiparam-se estes critérios clínicos a valores numéricos. Utilizando 36 pacientes internados, verificou-se até que ponto eles alcançavam estes critérios. Cada paciente recebeu um escore - valor numérico - que expressou o seu desempenho durante o tratamento em regime de internação, e sua conduta pós-alta hospitalar, durante um seguimento médio de 21 meses. Estes 36 pacientes puderam ser divididos em dois grupos: os de evolução favorável (obtivemos 36,1% e os de evolução desfavorável. Por meio de uma amostra sistemática composta por 41 pacientes que não foram submetidos ao método, internados no mesmo período de tempo, no mesmo hospital, e pareados com nossa casuística, fêz-se uma comparação estatística entre os percentis obtidos nas evoluções dos dois grupos. O valor do teste estatístico, o chi quadrado, não foi significante. O exame das proporções correspondentes a cada patologia em separado sugeriu fortemente que o tratamento estaria indicado nas neuroses e nas psicoses, mas que seria realmente ineficaz nos transtornos de

  19. X-FIDO: An Effective Application for Detecting Olive Quick Decline Syndrome with Deep Learning and Data Fusion

    Science.gov (United States)

    Cruz, Albert C.; Luvisi, Andrea; De Bellis, Luigi; Ampatzidis, Yiannis

    2017-01-01

    We have developed a vision-based program to detect symptoms of Olive Quick Decline Syndrome (OQDS) on leaves of Olea europaea L. infected by Xylella fastidiosa, named X-FIDO (Xylella FastIdiosa Detector for O. europaea L.). Previous work predicted disease from leaf images with deep learning but required a vast amount of data which was obtained via crowd sourcing such as the PlantVillage project. This approach has limited applicability when samples need to be tested with traditional methods (i.e., PCR) to avoid incorrect training input or for quarantine pests which manipulation is restricted. In this paper, we demonstrate that transfer learning can be leveraged when it is not possible to collect thousands of new leaf images. Transfer learning is the re-application of an already trained deep learner to a new problem. We present a novel algorithm for fusing data at different levels of abstraction to improve performance of the system. The algorithm discovers low-level features from raw data to automatically detect veins and colors that lead to symptomatic leaves. The experiment included images of 100 healthy leaves, 99 X. fastidiosa-positive leaves and 100 X. fastidiosa-negative leaves with symptoms related to other stress factors (i.e., abiotic factors such as water stress or others diseases). The program detects OQDS with a true positive rate of 98.60 ± 1.47% in testing, showing great potential for image analysis for this disease. Results were obtained with a convolutional neural network trained with the stochastic gradient descent method, and ten trials with a 75/25 split of training and testing data. This work shows potential for massive screening of plants with reduced diagnosis time and cost. PMID:29067037

  20. X-FIDO: An Effective Application for Detecting Olive Quick Decline Syndrome with Deep Learning and Data Fusion

    Directory of Open Access Journals (Sweden)

    Albert C. Cruz

    2017-10-01

    Full Text Available We have developed a vision-based program to detect symptoms of Olive Quick Decline Syndrome (OQDS on leaves of Olea europaea L. infected by Xylella fastidiosa, named X-FIDO (Xylella FastIdiosa Detector for O. europaea L.. Previous work predicted disease from leaf images with deep learning but required a vast amount of data which was obtained via crowd sourcing such as the PlantVillage project. This approach has limited applicability when samples need to be tested with traditional methods (i.e., PCR to avoid incorrect training input or for quarantine pests which manipulation is restricted. In this paper, we demonstrate that transfer learning can be leveraged when it is not possible to collect thousands of new leaf images. Transfer learning is the re-application of an already trained deep learner to a new problem. We present a novel algorithm for fusing data at different levels of abstraction to improve performance of the system. The algorithm discovers low-level features from raw data to automatically detect veins and colors that lead to symptomatic leaves. The experiment included images of 100 healthy leaves, 99 X. fastidiosa-positive leaves and 100 X. fastidiosa-negative leaves with symptoms related to other stress factors (i.e., abiotic factors such as water stress or others diseases. The program detects OQDS with a true positive rate of 98.60 ± 1.47% in testing, showing great potential for image analysis for this disease. Results were obtained with a convolutional neural network trained with the stochastic gradient descent method, and ten trials with a 75/25 split of training and testing data. This work shows potential for massive screening of plants with reduced diagnosis time and cost.

  1. X-FIDO: An Effective Application for Detecting Olive Quick Decline Syndrome with Deep Learning and Data Fusion.

    Science.gov (United States)

    Cruz, Albert C; Luvisi, Andrea; De Bellis, Luigi; Ampatzidis, Yiannis

    2017-01-01

    We have developed a vision-based program to detect symptoms of Olive Quick Decline Syndrome (OQDS) on leaves of Olea europaea L. infected by Xylella fastidiosa , named X-FIDO ( Xylella FastIdiosa Detector for O. europaea L.). Previous work predicted disease from leaf images with deep learning but required a vast amount of data which was obtained via crowd sourcing such as the PlantVillage project. This approach has limited applicability when samples need to be tested with traditional methods (i.e., PCR) to avoid incorrect training input or for quarantine pests which manipulation is restricted. In this paper, we demonstrate that transfer learning can be leveraged when it is not possible to collect thousands of new leaf images. Transfer learning is the re-application of an already trained deep learner to a new problem. We present a novel algorithm for fusing data at different levels of abstraction to improve performance of the system. The algorithm discovers low-level features from raw data to automatically detect veins and colors that lead to symptomatic leaves. The experiment included images of 100 healthy leaves, 99 X. fastidiosa -positive leaves and 100 X. fastidiosa -negative leaves with symptoms related to other stress factors (i.e., abiotic factors such as water stress or others diseases). The program detects OQDS with a true positive rate of 98.60 ± 1.47% in testing, showing great potential for image analysis for this disease. Results were obtained with a convolutional neural network trained with the stochastic gradient descent method, and ten trials with a 75/25 split of training and testing data. This work shows potential for massive screening of plants with reduced diagnosis time and cost.

  2. Bioassay-Guided Antidiabetic Study of <em>Phaleria macrocarpaem> Fruit Extract

    Directory of Open Access Journals (Sweden)

    Mohd Z. Asmawi

    2012-04-01

    Full Text Available An earlier anti-hyperglycemic study with serial crude extracts of <em>Phaleria macrocarpaem> (PM fruit indicated methanol extract (ME as the most effective. In the present investigation, the methanol extract was further fractionated to obtain chloroform (CF, ethyl acetate (EAF, <em>n>-butanol (NBF and aqueous (AF fractions, which were tested for antidiabetic activity. The NBF reduced blood glucose (<em>p> < 0.05 15 min after administration, in an intraperitoneal glucose tolerance test (IPGTT similar to metformin. Moreover, it lowered blood glucose in diabetic rats by 66.67% (<em>p> em>< 0.05, similar to metformin (51.11%, glibenclamide (66.67% and insulin (71.43% after a 12-day treatment, hence considered to be the most active fraction. Further fractionation of NBF yielded sub-fractions I (SFI and II (SFII, and only SFI lowered blood glucose (<em>p em>< 0.05, in IPGTT similar to glibenclamide. The ME, NBF, and SFI correspondingly lowered plasma insulin (<em>p em>< 0.05 and dose-dependently inhibited glucose transport across isolated rat jejunum implying an extra-pancreatic mechanism. Phytochemical screening showed the presence of flavonoids, terpenes and tannins, in ME, NBF and SFI, and LC-MS analyses revealed 9.52%, 33.30% and 22.50% mangiferin respectively. PM fruit possesses anti-hyperglycemic effect, exerted probably through extra-pancreatic action. Magniferin, contained therein may be responsible for this reported activity.

  3. OCCURENCE OF Salmonella IN CHICKEN CARCASSES AND CUTS COMMERCIALIZED IN GOIÂNIA-GO OCORRÊNCIA DE Salmonella EM CARCAÇAS E CORTES DE FRANGOS COMERCIALIZADOS EM GOIÂNIA-GO

    Directory of Open Access Journals (Sweden)

    Iolanda Aparecida Nunes

    2007-09-01

    Full Text Available

    Its was analised 53 chicken carcasses and commercial cuts acquired in supermarkets of Goiânia-GO aiming to isolate <em>Salmonella>. The contamination rate was 13.2% and serotypes <em>S. brandenburgem>, <em>S. typhimuriumem>, <em>S. agonaem>, <em>S. derbyem> and <em>S. hadorem> were identified.

    KEY-WORDS: <em>Salmonella>; chicken; chicken carcasses; commercial cuts.

    No presente estudo foram analisadas, com vistas ao isolamento de <em>Salmonella>, 53 amostras de carcaças e cortes comerciais de frangos colhidas em supermercados e casas de carnes de Goiânia - GO. Observou-se freqüência média do patógeno de 13,2%, sendo identificados os sorotipos <em>E. brandenburgem>, <em>S. typhimuriumem>, <em>S.> <em>agona>, <em>S. derbyem> e <em>S. Hadarem>.

    PALAVRAS-CHAVE: <em>Salmonella>; frango de corte; carcaça; cortes comerciais.

  4. Alimentazione di <em>Marmota marmotaem> in praterie altimontane delle dolomiti bellunesi

    Directory of Open Access Journals (Sweden)

    Alessandro Rudatis

    2006-03-01

    Full Text Available Abstract The diet of <em>Marmota marmotaem> in the mountain prairie of south-eastern Italian Alps. Diet composition of two family groups of alpine marmots was investigated in two areas of the Agordino’s Dolomites (Italian Alps in June-September 2001, by means of microscopic analysis of faeces and of direct observation of feeding activity. During the whole period of activity, a high consume of Angiosperms was confirmed, especially plants in flower; among them the “graminoids” seemed to play an important role only during the initial part of the active period. Generally vegetative parts predominated over flowers. The ingestion of animal preys was not confirmed by the analysis of droppings. Comparing diet composition of the two groups, Graminaceae (<em>Poa>, <em>Phleum>, Compositae (<em>Achillea>, Cyperaceae/Juncaceae, Leguminosae (<em>Anthyllis>, Rosaceae, and Labiatae (<em>Prunella>, <em>Stachys> formed the bulk of marmot diet in the study areas. Diet showed low diversity considering the abundance of plant species in the surrounding environment. Food resources were probably used in relation to their easy digestibility, with a high content in proteins, sugar and water. The knowledge of vegetation features in relation to marmot trophic habits can represent a useful tool for the management of this species. Riassunto Il regime alimentare di due gruppi di Marmotta alpina è stato studiato in giugno-settembre 2001 in due aree delle Dolomiti agordine (SE Italia, attraverso l’analisi microscopica delle feci e l’osservazione diretta dell’attività alimentare. Durante tutto il periodo di attività si è notato un forte consumo di Angiosperme, specialmente piante a fiore, mentre le ”graminoidi” sembra giochino un ruolo importante all’inizio della stagione. In generale le parti vegetali predominano sui fiori. L’ingestione di prede animali non è stata

  5. Soybean breeding with EMS mutagenesis

    International Nuclear Information System (INIS)

    Gu Aiqiu; Geng Yuxuan; Zhu Baogo

    1990-01-01

    Full text: 'Yudou No. 2' is a good soybean variety grown in the Honan Province. EMS was applied to seeds and valuable mutants were selected among the descendants. In a short period, several genetically stable strains were obtained. In the M 2 population, the early-maturing mutants were the most frequent, followed by short culm mutants. Other mutations altered leaf shape, grain size, habit of pod bearing, number of pods etc. One of the best strains is '86-180'. It is highly disease-resistant and ripens 19 days earlier than the original 'Yodou No. 2'. It bears more pods, although the seeds are a little bit smaller and is highly productive (4110kg/ha). Another good strain is '86-223'. It is also disease-resistant and highly productive (3390kg/ha). (author)

  6. Mastites em ruminantes no Brasil

    OpenAIRE

    Acosta, Atzel Candido; Silva, Leonildo Bento Galiza da; Medeiros, Elizabeth Sampaio; Pinheiro-Júnior, José Wilton; Mota, Rinaldo Aparecido

    2016-01-01

    Resumo: A mastite é uma doença complexa e considerada uma das principais causas de perdas à indústria leiteira mundial. Objetivou-se com esta revisão compilar informações dos últimos dez anos sobre a mastite em ruminantes no Brasil. A prevalência da mastite subclínica chega a 48,64% na espécie bovina, 30,7% na espécie caprina, 31,45% na espécie ovina e 42,2% na espécie bubalina, destacando-se a etiologia por Staphylococcus spp. Os fatores de risco associados à ocorrência de mastite estão rela...

  7. O "Popular" em Egberto Gismonti

    Directory of Open Access Journals (Sweden)

    Rúrion Soares Melo

    2007-07-01

    Full Text Available Nossa música, rica e moderna, não pode ser integralmente enquadrada em tipologias. Este artigo destaca a importância do material musical e da autonomia nos procedimentos composicionais, e propõe que a compreensão das obras de Egberto Gismonti não pode se restringir aos elementos propriamente "brasileiros" do "nacional-popular".Brazilian music, rich and modern, cannot be integrally fit in typologies. This article detaches the importance of the musical material and the autonomy in the compositional procedures, and considers that the understanding of the work of Egberto Gismonti cannot be restricted to strictly "Brazilian" elements of the "nacional-popular".

  8. Soybean breeding with EMS mutagenesis

    Energy Technology Data Exchange (ETDEWEB)

    Aiqiu, Gu; Yuxuan, Geng; Baogo, Zhu [Institute of Genetics, Academia Sinica, Beijing (China)

    1990-01-01

    Full text: 'Yudou No. 2' is a good soybean variety grown in the Honan Province. EMS was applied to seeds and valuable mutants were selected among the descendants. In a short period, several genetically stable strains were obtained. In the M{sub 2} population, the early-maturing mutants were the most frequent, followed by short culm mutants. Other mutations altered leaf shape, grain size, habit of pod bearing, number of pods etc. One of the best strains is '86-180'. It is highly disease-resistant and ripens 19 days earlier than the original 'Yodou No. 2'. It bears more pods, although the seeds are a little bit smaller and is highly productive (4110kg/ha). Another good strain is '86-223'. It is also disease-resistant and highly productive (3390kg/ha). (author)

  9. Medidas e forma em geometria

    OpenAIRE

    Edjan Fernandes dos Santos

    2015-01-01

    O trabalho traz inicialmente uma abordagem histÃrica, da GrÃcia (com os pitagÃricos), com o matemÃtico Eudoxo, fazendo referÃncia a talvez à maior obra matemÃtica, os livros de Euclides. Em seguida, trazemos definiÃÃes e construÃÃes sobre os nÃmeros reais com um corpo completo, os conceitos de Ãnfimo, supremo, sequÃncias infinitas com destaque as convergentes, sequÃncia de Cauchy e os trÃs teoremas fundamentais para o curso de cÃlculo, o teorema do anulamento, do valor intermediÃrio e de Weie...

  10. HAICAIS EM MANOEL DE BARROS

    Directory of Open Access Journals (Sweden)

    Profª Drª Nery Reiner

    2012-09-01

    Full Text Available O objetivo deste artigo, voltado para a criação poética de Manoel de Barros, é focalizar o conceito da obra de arte como jogo. Escolhemos Manoel de Barros, porque o lúdico, a brincadeira e o humor estão sempre presentes em sua obra. A escolha dos dados, o encaixe de palavras escolhidas a dedo, no eixo paradigmático, formam um caleidoscópio, mostrando novas estruturas, deixando explícito o desejo de brincar com os elementos linguísticos e da natureza. Para este trabalho, escolhemos, para análise, pequenos poemas semelhantes aos haicais japoneses e outros menores ainda, com dois ou até um verso apenas. Contaremos, nesta pesquisa, com a ajuda de teóricos como Johan Huizinga, Ezra Pound, Octavio Paz, Lévi-Strauss, Olga Savary e Alfredo Bosi.

  11. Synthesis and Antibacterial Evaluation of New <em>N>-acylhydrazone Derivatives from Dehydroabietic Acid

    Directory of Open Access Journals (Sweden)

    Wen Gu

    2012-04-01

    Full Text Available A series of new <em>N>-acylhydrazone derivatives were synthesized in good yields through the reactions of dehydroabietic acid hydrazide with a variety of substituted arylaldehydes. The structures of the synthesized compounds were confirmed by IR, 1H- and 13C-NMR, ESI-MS, elemental analysis and single crystal X-ray diffraction. From the crystal structure of compound 4l, the C=N double bonds of these <em>N>-acylhydrazones showed (<em>E>-configuration, while the NMR data of compounds 4aq indicated the existence of two rotamers for each compound in solution. The target compounds were evaluated for their antibacterial activities against four microbial strains. The result suggested that several compounds exhibited pronounced antibacterial activities. Particularly, compound 4p exhibited good antibacterial activity against <em>Staphylococcus aureusem> and <em>Bacillus subtilisem> comparable to positive control. The possible antibacterial metabolism and the strategy for further optimization of this compound were also discussed.

  12. Peat soils stabilization using Effective Microorganisms (EM)

    Science.gov (United States)

    Yusof, N. Z.; Samsuddin, N. S.; Hanif, M. F.; Syed Osman, S. B.

    2018-04-01

    Peat soil is known as geotechnical problematic soil since it is the softest soil having highly organic and moisture content which led to high compressibility, low shear strength and long-term settlement. The aim of this study was to obtain the stabilized peat soils using the Effective Microorganisms (EM). The volume of EM added and mixed with peat soils varied with 2%, 4%, 6%, 8% and 10% and then were cured for 7, 14 and 21 days. The experiment was done for uncontrolled and controlled moisture content. Prior conducting the main experiments, the physical properties such as moisture content, liquid limit, specific gravity, and plastic limit etc. were measure for raw peat samples. The Unconfined Compressive Strength (UCS) test was performed followed by regression analysis to check the effect of EM on the soil strength. Obtained results have shown that the mix design for controlled moisture contents showed the promising improvement in their compressive strength. The peat soil samples with 10% of EM shows the highest increment in UCS value and the percentage of increments are in the range of 44% to 65% after curing for 21 days. The regression analysis of the EM with the soil compressive strength showed that in controlled moisture conditions, EM significantly improved the soil stability as the value of R2 ranged between 0.97 – 0.78. The results have indicated that the addition of EM in peat soils provides significant improving in the strength of the soil as well as the other engineering properties.

  13. Cytotoxicity and Glycan-Binding Properties of an 18 kDa Lectin Isolated from the Marine Sponge <em>Halichondria em>okadai>

    Directory of Open Access Journals (Sweden)

    Yasuhiro Ozeki

    2012-04-01

    Full Text Available A divalent cation-independent lectin—HOL-18, with cytotoxic activity against leukemia cells, was purified from a demosponge, <em>Halichondria okadaiem>. HOL-18 is a 72 kDa tetrameric lectin that consists of four non-covalently bonded 18 kDa subunits. Hemagglutination activity of the lectin was strongly inhibited by chitotriose (GlcNAcβ1-4GlcNAcβ1-4GlcNAc, fetuin and mucins from porcine stomach and bovine submaxillary gland. Lectin activity was stable at pH 4–12 and temperatures lower than 60 °C. Frontal affinity chromatography with 16 types of pyridylaminated oligosaccharides indicated that the lectin had an affinity for <em>N>-linked complex-type and sphingolipid-type oligosaccharides with <em>N>-acetylated hexosamines and neuramic acid at the non-reducing termini. The lectin killed Jurkat leukemia T cells and K562 erythroleukemia cells in a dose- and carbohydrate-dependent manner.

  14. Echolocation calls and morphology in the Mehelyi’s (<em>Rhinolophus mehelyiem> and mediterranean (<em>R. euryaleem> horseshoe bats: implications for resource partitioning

    Directory of Open Access Journals (Sweden)

    Egoitz Salsamendi

    2006-03-01

    Full Text Available Abstract <em>Rhinolophus euryaleem> and <em>R. mehelyiem> are morphologically very similar species and their distributions overlap extensively in the Mediterranean basin. We modelled their foraging behaviour using echolocation calls and wing morphology and, assuming niche segregation occurs between the two species, we explored how it is shaped by these factors. Resting frequency of echolocation calls was recorded and weight, forearm length, wing loading, aspect ratio and wing tip shape index were measured. <em>R. mehelyiem> showed a significantly higher resting frequency than <em>R. euryaleem>, but differences are deemed insufficient for dietary niche segregation. Weight and forearm length were significantly larger in <em>R. mehelyiem>. The higher values of aspect ratio and wing loading and a lower value of wing tip shape index in <em>R. melehyiem> restrict its flight manoeuvrability and agility. Therefore, the flight ability of <em>R. mehelyiem> may decrease as habitat complexity increases. Thus, the principal mechanism for resource partitioning seems to be based on differing habitat use arising from differences in wing morphology. Riassunto Ecolocalizzazione e morfologia nei rinolofi di Mehely (<em>Rhinolophus mehelyiem> e euriale (<em>R. euryaleem>: implicazioni nella segregazione delle risorse trofiche. <em>Rhinolophus euryaleem> e <em>R. mehelyiem> sono specie morfologicamente molto simili, la cui distribuzione risulta largamente coincidente in area mediterranea. Il comportamento di foraggiamento delle due specie è stato analizzato in funzione delle caratteristiche dei segnali di ecolocalizzazione e della morfologia alare, ed è stata valutata l’incidenza di questi fattori nell’ipotesi di una segregazione delle nicchie. È stata rilevata la frequenza a riposo dei segnali ultrasonori, così come il peso, la lunghezza dell’avambraccio, il carico alare, e due

  15. Training for Defense? From Stochastic Traits to Synchrony in Giant Honey Bees (<em>Apis dorsataem>

    Directory of Open Access Journals (Sweden)

    Gerald Kastberger

    2012-08-01

    Full Text Available In Giant Honey Bees, abdomen flipping happens in a variety of contexts. It can be either synchronous or cascaded, such as in the collective defense traits of shimmering and rearing-up, or it can happen as single-agent behavior. Abdomen flipping is also involved in flickering behavior, which occurs regularly under quiescent colony state displaying singular or collective traits, with stochastic, and (semi- synchronized properties. It presumably acts via visual, mechanoceptive, and pheromonal pathways and its goals are still unknown. This study questions whether flickering is preliminary to shimmering which is subject of the <em>fs em>(flickering-shimmering-transition> hypothesis? We tested the respective prediction that trigger sites (<em>ts> at the nest surface (where shimmering waves had been generated show higher flickering activity than the alternative non-trigger sites (<em>nts>. We measured the flickering activity of <em>ts>- and <em>nts>-surface bees from two experimental nests, before and after the colony had been aroused by a dummy wasp. Arousal increased rate and intensity of the flickering activity of both <em>ts>- and <em>nts> cohorts (P < 0.05, whereby the flickering intensity of <em>ts>-bees were higher than that of <em>nts>-bees (P < 0.05. Under arousal, the colonies also increased the number of flickering-active <em>ts>- and <em>nts>-cohorts (P < 0.05. This provides evidence that cohorts which are specialist at launching shimmering waves are found across the quiescent nest zone. It also proves that arousal may reinforce the responsiveness of quiescent curtain bees for participating in shimmering, practically by recruiting additional trigger site bees for expanding repetition of rate and intensity of shimmering waves. This finding confirms the <em>fs-transition> hypothesis and constitutes evidence that flickering is part of a basal colony-intrinsic information system

  16. Mastite em pequenos ruminantes no Brasil

    OpenAIRE

    Peixoto,Rodolfo de M.; Mota,Rinaldo Aparecido; Costa,Mateus M. da

    2010-01-01

    Este artigo objetivou revisar as informações recentes sobre mastite em pequenos ruminantes, abrangendo etiologia, epidemiologia, aspectos de controle e profilaxia. Houve a preocupação em reunir resultados de estudos desenvolvidos no Brasil, uma vez que a mastite tem a interferência de uma série de fatores, como fatores ambientais e outros decorrentes dos sistemas de manejo empregados, condições essas determinantes para etiologia e epidemiologia da enfermidade. A prevalência da mastite em capr...

  17. Psicologia feminista em Portugal : dificuldades e conquistas

    OpenAIRE

    Saavedra, Luísa

    2010-01-01

    Pretende-se, com este artigo, fazer um balanço do que tem sido a Psicologia Feminista em Portugal. Tendo em conta que cada ciência comporta evoluções e posições epistemológicas e metodológicas distintas, começar-se-á por traçar em linhas gerais alguns marcos da psicologia no que diz respeito à forma de abordar as questões das mulheres e homens e definindo o que se entende por psicologia feminista. Seguidamente, serão evidenciados alguns aspectos políticos e sociais que condicionaram o desenvo...

  18. Ondas de spin em redes decoradas

    OpenAIRE

    WanÃssa FaÃanha da Silva

    2014-01-01

    Sistema de baixa dimensionalidade tÃm atraÃdo uma grande atenÃÃo ultimamente devido a sistemas como grafeno e nanotubos de carbono. Tais sistemas tÃm grandes possibilidades de aplicaÃÃes tecnolÃgicas, em particular na criaÃÃo de dispositivos eletrÃnicos, devido Ãs suas propriedades eletrÃnicas especÃficas. Nesse sentido, o estudos de outros sistemas em baixa dimensÃo se torna urgente. Mais especificamente, o estudo de propriedades magnÃticas de materiais de materiais em baixa dimensionalidade...

  19. Metabolic Profiling of <em>Lactococcus lactisem> Under Different Culture Conditions

    Directory of Open Access Journals (Sweden)

    Normah Mohd Noor

    2012-07-01

    Full Text Available Gas chromatography mass spectrometry (GC-MS and headspace gas chromatography mass spectrometry (HS/GC-MS were used to study metabolites produced by <em>Lactococcus lactisem> subsp. <em>cremoris> MG1363 grown at a temperature of 30 °C with and without agitation at 150 rpm, and at 37 °C without agitation. It was observed that <em>L. lactisem> produced more organic acids under agitation. Primary alcohols, aldehydes, ketones and polyols were identified as the corresponding trimethylsilyl (TMS derivatives, whereas amino acids and organic acids, including fatty acids, were detected through methyl chloroformate derivatization. HS analysis indicated that branched-chain methyl aldehydes, including 2-methylbutanal, 3-methylbutanal, and 2-methylpropanal are degdradation products of isoleucine, leucine or valine. Multivariate analysis (MVA using partial least squares discriminant analysis (PLS-DA revealed the major differences between treatments were due to changes of amino acids and fermentation products.

  20. Pré-resfriamento em água de lichia 'B3' mantida em armazenamento refrigerado

    OpenAIRE

    del Aguila,Juan Saavedra; Hofman,Peter; Campbell,Terrence; Marques,José Roberto; Heiffig-del Aguila,Lília Sichmann; Kluge,Ricardo Alfredo

    2009-01-01

    Este experimento visou a estudar o efeito de diferentes temperaturas e tempos de pré-resfriamento em água sobre a qualidade de lichias 'B3', após a frigoconservação. Os tratamentos foram: T1 = controle (sem pré-resfriamento); T2 = imersão por cinco minutos em água a 2,5°C; T3 = imersão por 10 minutos em água a 2,5°C; T4 = imersão por 20 minutos em água a 2,5°C; T5 = imersão por sete minutos em água a 6°C; T6 = imersão por 20 minutos em água a 8°C e T7 = imersão por 10 minu...

  1. Feeding habits of the genet <em>Genetta genettaem> in an Iberian continental wetland

    Directory of Open Access Journals (Sweden)

    Maite Sánchez

    2009-02-01

    Full Text Available Abstract This study quantified the diet of the genet <em>Genetta genettaem> in an Iberian continental wetland, the Galachos Nature Reserve, northern Spain, based on the frequency with which items appeared in five monthly-surveyed latrines during 2004-2005. Prey types were identified <em>de visuem> using identification keys and expert advice. the genet was confirmed as an opportunistic and generalist predator, its diet including mammals (95.0%, plants (68.3% and arthropods (60.0% as main prey. With the exception of a newly available prey species, the alien crayfish <em>Procambarus clarkiiem>, the genet probably consumed arthropods because of chance encounters rather than active search. The consumption of fruits and small mammals varied seasonally. Riassunto Alimentazione della genetta <em>Genetta genettaem> in un'area umida continentale della penisola iberica La dieta della genetta è stata investigata in un'area umida interna della Spagna settentrionale, la riserva Naturale dei Galachos. La frequenza di ciascuna categoria alimentare è stata espressa come numero di occorrenze mansili in cinque latrine monitorate per il periodo 2004-2005. Le categorie sono state identificate <em>de visuem> tramite chiavi per il riconoscimento o l'intervento di singoli esperti. I risultati confermano il comportamento alimentare generalista e opportunista della genetta. La dieta ha incluso principalmente mammiferi (95%, vegetali (68.3% e artropodi (60%. Questi ultimi, con l'eccezione del gambero <em>Procambarus clarkiiem>, introdotto e di recente presenza, sono probabilmente utilizzati in modo opportunistico, piuttosto che in seguito a ricerca attiva. L'uso dei frutti e dei micromammiferi varia stagionalmente.

  2. Over-Expression of CYP2E1 mRNA and Protein: Implications of Xenobiotic Induced Damage in Patients with <em>De Novoem> Acute Myeloid Leukemia with inv(16(p13.1q22; <em>CBFβ>-MYH11em>

    Directory of Open Access Journals (Sweden)

    Carlos E. Bueso-Ramos

    2012-08-01

    Full Text Available Environmental exposure to benzene occurs through cigarette smoke, unleaded gasoline and certain types of plastic. Benzene is converted to hematotoxic metabolites by the hepatic phase-I enzyme CYP2E1, and these metabolites are detoxified by the phase-II enzyme NQO1. The genes encoding these enzymes are highly polymorphic and studies of these polymorphisms have shown different pathogenic and prognostic features in various hematological malignancies. The potential role of different cytochrome p450 metabolizing enzymes in the pathogenesis of acute myeloid leukemia (AML in an area of active interest. In this study, we demonstrate aberrant CYP2E1 mRNA over-expression by quantitative real-time polymerase chain reaction in 11 cases of <em>de novoem> AML with inv(16; CBFβ-MYH11. CYP2E1 mRNA levels correlated with <em>CBFβ>-MYH11 em>transcript levels and with bone marrow blast counts in all cases. CYP2E1 over-expression correlated positively with NQO1 mRNA levels (R2 = 0.934, n = 7. By immunohistochemistry, CYP2E1 protein was more frequently expressed in AML with inv(16 compared with other types of AML (<em>p> < 0.001. We obtained serial bone marrow samples from two patients with AML with inv(16 before and after treatment. CYP2E1 mRNA expression levels decreased in parallel with <em>CBFβ>-MYH11 em>transcript levels and blast counts following chemotherapy. In contrast, CYP1A2 transcript levels did not change in either patient. This is the first study to demonstrate concurrent over-expression of CYP2E1 and NQO1 mRNA in AML with inv(16. These findings also suggest that a balance between CYP2E1 and NQO1 may be important in the pathogenesis of AML with inv(16.

  3. Synthesis and Sensory Evaluation of <em>ent>-Kaurane Diterpene Glycosides

    Directory of Open Access Journals (Sweden)

    Venkata Sai Prakash Chaturvedula

    2012-07-01

    Full Text Available Catalytic hydrogenation of the three <em>ent>-kaurane diterpene glycosides isolated from <em>Stevia rebaudianaem>, namely rubusoside, stevioside, and rebaudioside-A has been carried out using Pd(OH2 and their corresponding dihydro derivatives have been isolated as the products. Synthesis of reduced steviol glycosides was performed using straightforward chemistry and their structures were characterized on the basis of 1D and 2D NMR spectral data and chemical studies. Also, we report herewith the sensory evaluation of all the reduced compounds against their corresponding original steviol glycosides and sucrose for the sweetness property of these molecules.

  4. Complete BWR--EM LOCA analysis using the WRAP--EM system

    International Nuclear Information System (INIS)

    Beckmeyer, R.R.; Gregory, M.V.; Buckner, M.R.

    1979-01-01

    The Water Reactor Analysis Package, Evaluation Model (WRAP--EM), provides a complete analysis of postulated loss-of-coolant accidents (LOCA's) in light--water nuclear power reactors. The system is being developed at the Savannah River Laboratory (SRL) for use by the Nuclear Regulatory Commission (NRC) to interpret and evaluate reactor vendor, evaluation model (EM) analyses. The initial version of the WRAP--EM system for analysis of boiling water reactors (BWR's) is operational. To demonstrate the complete capability of the WRAP--BWR--EM system, a LOCA analysis has been performed for the Hope Creek Plant

  5. Monitores de diagnóstico em Mamografia : (Os 5MP em Portugal)

    OpenAIRE

    Timóteo, Maria Teresa

    2012-01-01

    O carcinoma da mama é o mais comum nas mulheres, em Portugal morrem cerca de 1500 mulheres por ano devido a esta patologia. Serviu o presente estudo para investigar se os Hospitais da Região de Lisboa e Vale do Tejo estão equipados, com monitores de diagnóstico em mamografia, que correspondam com as guidelines europeias. O serviço de Mamografia, em Portugal rege-se por estas guidelines, em que é referenciado que para um correcto diagnóstico, com toda a sensibilidade e especific...

  6. Osteocondromatose em gato: relato de caso

    Directory of Open Access Journals (Sweden)

    S. Winter

    Full Text Available RESUMO A osteocondromatose é caracterizada por nódulos únicos ou múltiplos decorrentes de um crescimento ósseo excessivo benigno. É encontrada em cães, gatos, equinos e humanos. Em felinos, tem maior incidência dos dois aos quatro anos de idade. A etiologia em gatos está relacionada ao vírus da leucemia felina, e também já foi encontrada relação com o fibrossarcoma. A manifestação clínica depende do local acometido e do tamanho da lesão. O diagnóstico definitivo é por meio de histopatologia e o prognóstico é desfavorável, pois ocorrem muitas recidivas. Este relato de caso objetiva descrever a apresentação dessa enfermidade em um felino jovem.

  7. Probióticos em avicultura

    Directory of Open Access Journals (Sweden)

    Gil de los Santos João Rodrigo

    2005-01-01

    Full Text Available Durante os últimos anos, o incremento de toxinfecções alimentares em humanos pela ingestão de produtos avícolas contaminados com bactérias, sugeriu que produtos oriundos de aves portadoras dessas bactérias podem veiculá-las ao consumidor, provocando doenças entéricas em humanos. Essas observações e a restrição, por parte do mercado consumidor, ao uso de antimicrobianos na produção animal, aumentaram o interesse mundial pelos probióticos e, conseqüentemente, o número de pesquisas realizadas. Esta revisão mostra os avanços e a situação em que se encontram os probióticos em relação à indústria avícola.

  8. Comments on theoretical foundation of "EM Drive"

    Science.gov (United States)

    Wu, C.-W.

    2018-03-01

    The concept of EM Drive has attracted much attention and groups of work have been conducted to prove or verify it, of which the published experimental outcome is criticized in great details while the theoretical foundation has not been discussed. The present essay investigates on the theoretical derivations of the net thrust in the "EM drive" and reveals the self-contradiction arising at the very start, when the law of conservation of momentum was utilized and opposed simultaneously.

  9. Isolamento de Corynebacterium aquaticum em leite bubalino

    Directory of Open Access Journals (Sweden)

    Andréa Alice da Fonseca Oliveira

    2005-08-01

    Full Text Available Estudou-se 548 quartos mamários de búfalas, realizando-se exame clínico, CMT para detecção de mastite e coleta de amostras para isolamento bacteriano. Houve crescimento em duas amostras de Corynebacterium aquaticum caracterizadas bioquimicamente. Relata-se a participação do agente como colonizador do úbere e possível causador de mastites em bubalinos.

  10. Resumos de artigos em Português

    OpenAIRE

    Bogdan Patrut

    2010-01-01

    Resumos de artigos em Português
    BRAIN. Broad Research in Artificial Intelligence and Neuroscience
    CÉREBRO. Ampla pesquisa em Inteligência Artificial e Neurociências
    ISSN 2067-3957
    Volume 1, Número 2
    Abril de 2010, "Feliz Verão 2010!"
    www.brain.edusoft.ro
    Editor - chefe: Bogdan Pătruţ

  11. Feocromocitoma em criança

    OpenAIRE

    Herdy,Gesmar Volga Haddad; Olivaes,Maria Cecília; Lopes,Vânia Glória Silami; Pontes,Carlos Adriano Gazanego; Ormond Filho,João Batista; Fonseca,Eliene C.

    2005-01-01

    Feocromocitoma é uma neoplasia de células cromafins que causa sintomas e sinais decorrentes da libertação de catecolminas e é muito rara em crianças. Relatamos um caso feocromocitoma em criança, com difícil manejo clínico, confirmado também pela anatomia-patológica e curado após a ressecção cirúrgica.

  12. NOVOS PARASITÓIDES EM OVOS DE Dirphia Araucariae (LEPIDOPTERA, ATTACIDAE NEW PARASITOIDS IN EGGS OF Dirphia araucariae (LEP.; ATTACIDAE

    Directory of Open Access Journals (Sweden)

    Jácomo Divino Borges

    2007-09-01

    Full Text Available

    Foram observados novos parasitóides nos ovos de <em>Dirphia> <em>araucariae> Jones, 1908 (<em>Lepidoptera>, <em>Attacidae>. A porcentagem do parasitismo em ovos do hospedeiro, coletados na Fazenda Marabá, Município de Congonhinhas (PR foi de 20,71%, e as espécies de parasitóides identificadas e suas freqüências do: <em>Anastatus> <em>sp> (0,45%; <em>A. charitos em>De Santis, 1982 (1,08%; A. (<em>Proanastatus> <em>excavatus> (De Santis, 1952 (1,97%; <em>Ooencyrtus> <em>sp> (17,86%; <em>O.> <em>blastothricoide> De Santis, 1988 (43,65%; e <em>O. dirphiaeem> De Santis, 1988 (34,99%.

    PALAVRAS-CHAVE: <em>Anastatus> <em>spp>; <em>Araucaria> <em>angustifolia>; controle biológico; lagarta-da-araucária; <em>Ooencyrtus sppem>; pinheiro-do-Paraná.

    The parasitism in Dirphia araucariae Jones, 1908 (<em>Lepidoptera>, <em>Attacidae> was studied and the parasitoids observed on its eggs collected in 'Fazenda Marabá’, Municipality of Congonhinhas, State of Paraná, Brazil, were the following: <em>Anastatus> <em>sp>.; <em>A>. <em>charitos> De Santis, 1982; <em>A.> (<em>Proanastatus> <em>excavatus> (De Santis, 1952; <em>Ooencyrtus> <em>sp.>; <em>O. blastothricoideem> De Santis, 1988, and <em>O. dirphiaeem> De Santis, 1988, with a frequency of 0.45%, 1.08%, 1.97%, 17.86%, 43.65% and 34.99%, respectively.

    KEY-WORDS: Biological control; Paraná-pine.

  13. Development of 101 Gene-based Single Nucleotide Polymorphism Markers in Sea Cucumber, <em>Apostichopus japonicusem>>

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    Wei Lu

    2012-06-01

    Full Text Available Single nucleotide polymorphisms (SNPs are currently the marker of choice in a variety of genetic studies. Using the high resolution melting (HRM genotyping approach, 101 gene-based SNP markers were developed for <em>Apostichopus japonicusem>, a sea cucumber species with economic significance for the aquaculture industry in East Asian countries. HRM analysis revealed that all the loci showed polymorphisms when evaluated using 40 <em>A. japonicusem> individuals collected from a natural population. The minor allele frequency ranged from 0.035 to 0.489. The observed and expected heterozygosities ranged from 0.050 to 0.833 and 0.073 to 0.907, respectively. Thirteen loci were found to depart significantly from Hardy–Weinberg equilibrium (HWE after Bonferroni corrections. Significant linkage disequilibrium (LD was detected in one pair of markers. These SNP markers are expected to be useful for future quantitative trait loci (QTL analysis, and to facilitate marker-assisted selection (MAS in <em>A. japonicusem>.

  14. Effect of β,β-Dimethylacrylshikonin on Inhibition of Human Colorectal Cancer Cell Growth <em>in Vitro em>and <em>in Vivoem>

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    Ting Feng

    2012-07-01

    Full Text Available In traditional Chinese medicine, shikonin and its derivatives, has been used in East Asia for several years for the prevention and treatment of several diseases, including cancer. We previously identified that β,β-dimethylacrylshikonin (DA could inhibit hepatocellular carcinoma growth. In the present study, we investigated the inhibitory effects of DA on human colorectal cancer (CRC cell line HCT-116 <em>in vitroem> and <em>in vivoem>. A viability assay showed that DA could inhibit tumor cell growth in a time- and dose-dependent manner. Flow cytometry showed that DA blocks the cell cycle at G0/G1 phase. Western blotting results demonstrated that the induction of apoptosis by DA correlated with the induction of pro-apoptotic proteins Bax, and Bid, and a decrease in the expression of anti-apoptotic proteins Bcl-2 and Bcl-xl. Furthermore, treatment of HCT-116 bearing nude mice with DA significantly retarded the growth of xenografts. Consistent with the results <em>in vitroem>, the DA-mediated suppression of HCT-116 xenografts correlated with Bax and Bcl-2. Taken together, these results suggest that DA could be a novel and promising approach to the treatment of CRC.

  15. Dermatite multifatorial em um canino

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    C.B. Matos

    2012-12-01

    Full Text Available Relatou-se um caso clínico de dermatite multifatorial em cão, envolvendo Demodex canis, Sporothrix schenckii e bactérias dos gêneros Staphylococcus e Streptococcus. O animal, macho, da raça Pit Bull, com um ano e três meses de idade, apresentou lesões úmidas, hiperêmicas, descamativas e ulcerativas nas regiões do pescoço e do tórax. Exames laboratoriais confirmaram o diagnóstico de infecção concomitante por fungo, bactéria e parasito. A terapia antimicrobiana com moxidectina, itraconazol e amoxicilina foi efetiva, com remissão dos sinais clínicos após 30 dias de tratamento. Este relato salienta a importância do diagnóstico de certeza, que foi determinante para estabelecimento da terapêutica adequada e recuperação do animal.

  16. Monorquidismo em um equino marchador

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    Jéssica Fontana de Magalhães

    2015-06-01

    Full Text Available O monorquidismo é um quadro raramente descrito em equinos, que não apresenta descrições prévias na literatura nacional. Trata-se de um equino adulto brasileiro, com oito anos de idade, sem raça definida, porém com características de raças de origem nacional. Por abordagem inguinal do lado esquerdo, localizou-se a túnica vaginal intacta, que, aberta, continha a cauda de um epidídimo mal formado, o ducto deferente e um delgado pedículo vascular, mas sem indícios de testículo. Exames histológicos e hormonais confirmaram a ausência de tecido testicular. Conclui-se que os exames histológicos e hormonais são essenciais para o diagnóstico de monorquidismo que, apesar de raro, também pode acometer equinos nacionais

  17. Self: um conceito em desenvolvimento

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    Lídia Suzana Rocha de Macedo

    2012-08-01

    Full Text Available Diversas definições de self coexistem nas teorias e práticas psicológicas. Essa variedade é resultante de bases epistemológicas a partir das quais se adotam estratégias diferentes para abordar e demarcar os limites do objeto em questão e descrevê-lo. Este estudo teve como objetivo oferecer uma revisão dos conceitos de self e uma reflexão sobre como esse conceito se articula nas diferentes abordagens teóricas da psicologia. Destaca-se que dilemas centrais à psicologia do desenvolvimento atravessam o conceito de self. Por essa razão, apresenta-se a tese de que, ao analisar cada definição de self, deve-se buscar responder como cada teoria colocou-se diante das dicotomias estabilidade versus transformação, específico versus universal e mundo interno versus mundo externo.

  18. Mastites em ruminantes no Brasil

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    Atzel Candido Acosta

    Full Text Available Resumo: A mastite é uma doença complexa e considerada uma das principais causas de perdas à indústria leiteira mundial. Objetivou-se com esta revisão compilar informações dos últimos dez anos sobre a mastite em ruminantes no Brasil. A prevalência da mastite subclínica chega a 48,64% na espécie bovina, 30,7% na espécie caprina, 31,45% na espécie ovina e 42,2% na espécie bubalina, destacando-se a etiologia por Staphylococcus spp. Os fatores de risco associados à ocorrência de mastite estão relacionados a problemas no saneamento ambiental e ao manejo dos animais. As bactérias isoladas do leite mastítico apresentam maior percentual de resistência a penicilina, ampicilina, amoxicilina e neomicina e a utilização de técnicas moleculares no diagnóstico dos agentes causadores de mastites no país, ainda é escassa o que dificulta a obtenção de um diagnóstico mais rápido, sensível e específico.

  19. Literatura e(m hipertexto

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    Bellei, Sérgio Luiz Prado

    2010-01-01

    Full Text Available A crescente hegemonia do modo de produção digital de textos literários a serem disponibilizados em rede torna urgente o questionamento a respeito das possíveis mudanças que ocorrem na produção de hipertextos eletrônicos a partir de textos literários. O presente ensaio desenvolve uma reflexão crítica sobre o problema a partir de uma distinção do duplo significado de tecnologia enquanto, de um lado, instrumental para a produção de objetos e, de outro, ambiente estruturante que não só produz objetos mas, principalmente, molda os sujeitos que deles fazem uso, reconstituindo no processo hábitos e procedimentos. Se levarmos a sério o conceito de tecnologia enquanto ambiente estruturante, então é bem possível que a literatura traduzida ou produzida diretamente no formato hipertextual sofra modificações profundas quando comparadas à produção literária no meio impresso

  20. Identification of Volatile Components of Liverwort (<em>Porella cordaeanaem> Extracts Using GC/MS-SPME and Their Antimicrobial Activity

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    Maria Elisabetta Guerzoni

    2012-06-01

    Full Text Available Chemical constituents of liverwort (<em>Porella cordaeanaem> extracts have been identified using solid-phase microextraction-gas chromatography mass spectrometry (SPME-GC/MS. The methanol, ethanol and ethyl acetate extracts were rich in terpenoids such as sesquiterpene hydrocarbons (53.12%, 51.68%, 23.16%, and monoterpene hydrocarbons (22.83%, 18.90%, 23.36%, respectively. The dominant compounds in the extracts were β-phellandrene (15.54%, 13.66%, 12.10% and β-caryophyllene (10.72%, 8.29%, 7.79%, respectively. The antimicrobial activity of the extracts was evaluated against eleven food microorganisms using the microdilution and disc diffusion methods. The minimum inhibitory concentration (MIC varied from 0.50 to 2.00 mg/mL for yeast strains (<em>Saccharomyces cerevisiae 635em>, <em>Zygosacharomyces bailii 45em>, <em>Aerobasidium pullulans L6Fem>, <em>Pichia membranaefaciens OC 71em>, <em>Pichia membranaefaciens OC 70em>, <em>Pichia anomalaem> <em>CBS 5759em>, <em>Pichia anomala DBVPG 3003em> and<em> em>>Yarrowia lipolytica RO13em>, and from 1.00 to 3.00 mg/mL for bacterial strains<em> em>(Salmonella> <em>enteritidis 155em>, <em>Escherichia coli 555em> and <em>Listeria monocytogenes 56Lyem>. Methanol extract showed better activity in comparison with ethanol and ethyl acetate extracts. High percentages of monoterpene and sesquiterpene hydrocarbons could be responsible for the better antimicrobial activity.

  1. Revivendo experiências: em foco a leitura em comunidades

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    Marlene Edite Pereira Rezende

    Full Text Available Este artigo teve como objetivo recuperar, de forma destacada, duas experiências de trabalho de extensão na UFMG: o carro-biblioteca da ECI\\UFMG e a implantação das Bibliotecas Públicas e Biblioteca Móvel do COLTEC\\UFMG. A estas experiências somaram-se ainda as informações sobre as atividades de extensão, através de Carros-Biblioteca, realizadas no âmbito da Biblioteca Pública Estadual Luiz de Bessa e pelo SESC. Revendo estas experiências torna-se patente a importância destas ações para as comunidades, mas, revelam-se também as dificuldades/limitações para a continuidade e ampliação destes serviços que são de caráter significativo para a população. Destaca-se também que os projetos visam permitir o acesso à leitura em comunidades com carências socioculturais, mas, demostram, de forma crítica, a realidade e os desafios dos municípios de Caparaó e Alto Caparaó na administração e manutenção das bibliotecas lá instaladas. Argumenta-se ainda, como aspecto significativo, a necessidade de articulação das comunidades, de forma a reivindicarem junto aos gestores públicos o atendimento aos direitos da população.

  2. Teores proteicos em dietas para vacas Holandesas leiteiras em confinamento

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    V.R. Paiva

    2013-08-01

    Full Text Available Avaliou-se o efeito de dietas com níveis de proteína bruta ‒ PB (10, 12, 14 e 16% na matéria seca ‒ MS em vacas Holandesas mestiças com produção de leite médio (PL de 13kg/dia. Utilizou-se silagem de milho como volumoso, e a relação volumoso:concentrado foi de 75:25. Houve aumento linear para o consumo de todos os componentes da dieta, exceto para o consumo de carboidratos totais e carboidratos não fibrosos (CNF, que reduziram linearmente. O consumo dos nutrientes digestíveis totais (NDT apresentou comportamento quadrático, com valor máximo estimado de 10,13kg/dia, na dose de 15,62% de PB na dieta. As digestibilidades de PB, EE, FDN e FDNcp aumentaram linearmente. Já os valores de digestibilidade de MS, MO, CNF e NDT apresentaram efeito quadrático, estimando-se ponto de máximo de 65,09; 67,23; 78,35 e 67,92 nas doses de 15,39; 15,22; 15,62 e 15,68% de PB, respectivamente. A PL, sem e com correção para 3,5% de gordura, a variação de peso corporal e a composição do leite aumentaram linearmente, exceto para lactose, que não apresentou efeito. O nível de 14% de PB na dieta é o mais indicado para alimentação de vacas com produção média de 13kg/dia de leite.

  3. <em>Helicobacter pyloriem> Disrupts Host Cell Membranes, Initiating a Repair Response and Cell Proliferation

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    Hsueh-Fen Juan

    2012-08-01

    Full Text Available <em>Helicobacter pyloriem> (<em>H. pyloriem>, the human stomach pathogen, lives on the inner surface of the stomach and causes chronic gastritis, peptic ulcer, and gastric cancer. Plasma membrane repair response is a matter of life and death for human cells against physical and biological damage. We here test the hypothesis that <em>H. pyloriem> also causes plasma membrane disruption injury, and that not only a membrane repair response but also a cell proliferation response are thereby activated. Vacuolating cytotoxin A (VacA and cytotoxin-associated gene A (CagA have been considered to be major <em>H. pyloriem> virulence factors. Gastric cancer cells were infected with <em>H. pyloriem> wild type (vacA+/cagA+, single mutant (ΔvacA or ΔcagA or double mutant (ΔvacA/ΔcagA strains and plasma membrane disruption events and consequent activation of membrane repair components monitored. <em>H. pyloriem> disrupts the host cell plasma membrane, allowing localized dye and extracellular Ca2+ influx. Ca2+-triggered members of the annexin family, A1 and A4, translocate, in response to injury, to the plasma membrane, and cell surface expression of an exocytotic maker of repair, LAMP-2, increases. Additional forms of plasma membrane disruption, unrelated to <em>H.> pylori em>exposure, also promote host cell proliferation. We propose that <em>H. pyloriem> activation of a plasma membrane repair is pro-proliferative. This study might therefore provide new insight into potential mechanisms of <em>H. pyloriem>-induced gastric carcinogenesis.

  4. Net Energy, CO2 Emission and Land-Based Cost-Benefit Analyses of <em>Jatropha> em>Biodiesel: A Case Study of the Panzhihua Region of Sichuan Province in China

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    Xiangzheng Deng

    2012-06-01

    Full Text Available Bioenergy is currently regarded as a renewable energy source with a high growth potential. Forest-based biodiesel, with the significant advantage of not competing with grain production on cultivated land, has been considered as a promising substitute for diesel fuel by many countries, including China. Consequently, extracting biodiesel from <em>Jatropha> curcasem> has become a growing industry. However, many key issues related to the development of this industry are still not fully resolved and the prospects for this industry are complicated. The aim of this paper is to evaluate the net energy, CO2 emission, and cost efficiency of <em>Jatropha> biodiesel as a substitute fuel in China to help resolve some of the key issues by studying data from this region of China that is well suited to growing <em>Jatropha>. Our results show that: (1 <em>Jatropha> biodiesel is preferable for global warming mitigation over diesel fuel in terms of the carbon sink during <em>Jatropha> tree growth. (2 The net energy yield of <em>Jatropha> biodiesel is much lower than that of fossil fuel, induced by the high energy consumption during <em>Jatropha> plantation establishment and the conversion from seed oil to diesel fuel step. Therefore, the energy efficiencies of the production of <em>Jatropha> and its conversion to biodiesel need to be improved. (3 Due to current low profit and high risk in the study area, farmers have little incentive to continue or increase <em>Jatropha> production. (4 It is necessary to provide more subsidies and preferential policies for <em>Jatropha> plantations if this industry is to grow. It is also necessary for local government to set realistic objectives and make rational plans to choose proper sites for <em>Jatropha> biodiesel development and the work reported here should assist that effort. Future research focused on breading high-yield varieties, development of efficient field

  5. Local feeding specialization of the red fox (<em>Vulpes vulpesem> in response to eastern cottontail (<em>Sylvilagus floridanusem> introduction (NW Italy

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    Alessandro Balestrieri

    2006-03-01

    Full Text Available Abstract To appreciate the influence of the introduction of the Eastern cottontail (<em>Sylvilagus floridanusem> on the food habits of the red fox (Vulpes vulpes, between June 1998 and February 2000 fox diet was investigated by means of scat analysis (N=115 in a 250 ha wide Natural Reserve of NW Italy, and compared with data collected in the same area prior to cottontail colonization (1988-1989. Comparison included also the diet of badgers (<em>Meles melesem>, considered as potential competitors for food resources. Alien lagomorphs (mean percent volume, Vm% = 68% represented by far the most exploited resource, only three other food items reaching values of mean percent volume barely higher than 5%. Cottontails frequency of occurrence did not vary according either to season or to their reproductive cycle (II-IX vs. X-I, whilst diet niche breadth varied inversely proportional to the use of this key-resource. Overall fox trophic niche breadth varied from 0.64 in 1988-89 to 0.31 in 1998-00 (B, Levin’s index. These findings led us to consider the feeding habits of the fox in the study area as a result of local specialization of a typical generalist carnivore, according to the predictions of optimal foraging theory. No variation occurred in the badger niche breadth since cottontail introduction, whilst niche overlap between foxes and badgers decreased from 0.59 to 0.13 (O, Pianka’s index, possibly reducing competition for food in summer. Riassunto Specializzazione alimentare a livello locale della Volpe <em>Vulpes vulpesem> in risposta all’introduzione del Silvilago <em>Sylvilagus floridanusem> (Italia nord occidentale. Per valutare gli effetti dell’introduzione del Silvilago (<em>Sylvilagus floridanusem> sul comportamento alimentare della volpe (<em>Vulpes vulpesem>, nel periodo giugno 1998-febbraio 2000, la dieta del carnivoro è stata definita tramite l’analisi di 115

  6. Antibacterial Secondary Metabolites from the Cave Sponge <em>Xestospongia em>sp.

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    Sridevi Ankisetty

    2012-05-01

    Full Text Available Chemical investigation of the cave sponge <em>Xestospongia em>sp. resulted in the isolation of three new polyacetylenic long chain compounds along with two known metabolites. The structures of the new metabolites were established by NMR and MS analyses. The antibacterial activity of the new metabolites was also evaluated.

  7. Refração por retinoscopia com luz em faixa em cães

    OpenAIRE

    Ranzani,J.J.T.; Mobricci,L.A.L.; Carvalho,L.R.; Brandão,C.V.S.

    2008-01-01

    A refração ocular por retinoscopia com luz em faixa em 10 cães normais, sem raça definida, demonstrou que a maioria dos animais, independentemente do porte, apresentou tendência à hipermetropia, além de anisometropia e astigmatismo.

  8. Enzymatic Properties and Mutational Studies of Chalcone Synthase from <em>Physcomitrella patensem>

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    Mahiran Basri

    2012-08-01

    Full Text Available <em>Pp>CHS is a member of the type III polyketide synthase family and catalyses the synthesis of the flavonoid precursor naringenin chalcone from <em>p>-coumaroyl-CoA. Recent research reports the production of pyrone derivatives using either hexanoyl-CoA or butyryl-CoA as starter molecule. The Cys-His-Asn catalytic triad found in other plant chalcone synthase predicted polypeptides is conserved in <em>Pp>CHS. Site directed mutagenesis involving these amino acids residing in the active-site cavity revealed that the cavity volume of the active-site plays a significant role in the selection of starter molecules as well as product formation. Substitutions of Cys 170 with Arg and Ser amino acids decreased the ability of the <em>Pp>CHS to utilize hexanoyl-CoA as a starter molecule, which directly effected the production of pyrone derivatives (products. These substitutions are believed to have a restricted number of elongations of the growing polypeptide chain due to the smaller cavity volume of the mutant’s active site.

  9. <em>In Vitro em>Phytotoxicity and Antioxidant Activity of Selected Flavonoids

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    Rita Patrizia Aquino

    2012-05-01

    Full Text Available The knowledge of flavonoids involved in plant-plant interactions and their mechanisms of action are poor and, moreover, the structural characteristics required for these biological activities are scarcely known. The objective of this work was to study the possible <em>in vitro em>phytotoxic effects of 27 flavonoids on the germination and early radical growth of <em>Raphanus sativus em>L.> and <em>Lepidium sativumem> L., with the aim to evaluate the possible structure/activity relationship. Moreover, the antioxidant activity of the same compounds was also evaluated. Generally, in response to various tested flavonoids, germination was only slightly affected, whereas significant differences were observed in the activity of the various tested flavonoids against radical elongation. DPPH test confirms the antioxidant activity of luteolin, quercetin, catechol, morin, and catechin. The biological activity recorded is discussed in relation to the structure of compounds and their capability to interact with cell structures and physiology. No correlation was found between phytotoxic and antioxidant activities.

  10. Home range dynamics of mountain hare (<em>Lepus timidusem> in the Swiss Alps

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    Anne-Sophie Genini-Gamboni

    2009-02-01

    Full Text Available Abstract Little is known on the ecology and behaviour of alpine mountain hare (<em>Lepus timidusem>. Between 1996 and 1997 we analysed by radiotracking the pattern of space use of 8 mountain hares from the Swiss Alps. We estimated home range size using both the kernel density estimator and the minimum convex polygon. We found smaller ranges (38 ha compared to those reported for the species in boreal or arctic habitats, but similar to ranges in Scotland. Hares did not use a centre of major activity (core area and showed high home range overlap, confirming their non-territorial behaviour. Smaller ranges were used during winter compared to the other seasons, whilst no difference in size was found between sexes. Riassunto Dinamica dell'uso dello spazio della lepre bianca (<em>Lepus timidusem> nelle Alpi Svizzere Le informazioni relative all'ecologia e al comportamento della lepre alpina (<em>Lepus timidusem> sono ad oggi scarse. In questo studio abbiamo analizzato l'utilizzo dello spazio di una popolazione di lepre bianca sulle Alpi Svizzere. Tra il 1996 e il 1997 sono stati marcati con redio collare 8 individui di lepre alpina. L'<em>home rangeem> è stato calcolato utilizzando lo stimatore di densità kernel (KDE ed il metodo del minimo poligono convesso (MCP. L'ampiezza degli <em>home rangeem> (38 ha è risultata inferiore a quella riportata per la specie in habitat boreali ed artici. ma simile a quella riscontrata in Scozia. All'interno dell <em>home rangeem> non è stato rilevato alcun centro di maggiore attività (<em>core areaem> ed è stata evidenziata una notevole sovrapposizione tra gli stessi, confermando la non territorialità della specie. Le aree frequentate in inverno sono risultate più piccole rispetto alle altre stagioni e non sono state riscontrate differenze tra i sessi.

  11. The EM technology development strategy

    International Nuclear Information System (INIS)

    Frank, C.W.; Barainca, M.; Kubo, A.S.

    1992-01-01

    The Office of Technology Development (TD) supports research and development of technologies that will lower cost, reduce risk, improve safety, and accelerate cleanup of the Nuclear Weapons Complex and provide solutions to currently untractable environmental problems. The TD strategic plan outlines Applied Research, Development, Demonstration, Testing, and Evaluation (RDDT and E) that will provide needed technology products to be used by Environmental Restoration and Waste Management operations (i.e., our customers). The TD strategic plan is derived from EM Goals, Objectives, and Strategy and is incorporated into DOE'S Five-Year Plan for Environmental Restoration and Waste Management. The TD strategic plan is developed based on integrating customer requirements, and is complemented by a top-down, bottom-up analysis of Site Specific Technology Needs and environmental problems. The execution of TD's strategic plan is implemented largely through Integrated Programs (IP) and Integrated Demonstrations (ID). IDs have proven to be a cost-effective method of managing technology development, testing and evaluation, and implementation of successful technology systems into the DOE Environmental Restoration and Waste Management Programs. The Savannah River ID for Volatile Organic Compounds (VOCs) in Saturated Soils resulted in a 51 percent cost savings over stand-alone demonstrations, saving over $8 million. The IPs and IDs are selected based on customer needs, technical complexity, and complex-wide regulatory and compliance agreements. New technology systems are selected for incorporation into an IP or ID from offerings of the DOE laboratories, industry, and the universities. A major TD initiative was announced in August 1991, with the release of a Program Research and Development Announcement (PRDA) requesting industry and universities to propose innovative new technologies to clean up the Weapons Complex. (author)

  12. Are EM's communication tools effective? Evaluation research of two EM publications

    International Nuclear Information System (INIS)

    Wight, Evelyn; Gardner, Gene; Harvey, Tony

    1992-01-01

    As a reflection of its growing culture of openness, and in response to the public's need for accurate information about its activities, the U.S. Department of Energy (DOE) Office of the Assistant Secretary for Environmental Restoration and Waste Management (EM) has increased the amount of information available to the public through communication tools such as brochures, fact sheets, and a travelling exhibit with an interactive computer display. Our involvement with this effort has been to design, develop, and critique booklets, brochures, fact sheets and other communication tools for EM. This paper presents an evaluation of the effectiveness of two communication tools we developed: the EM Booklet and the EM Fact Sheets. We measured effectiveness using non-parametric testing. This paper describes DOE's culture change, EM's communication tools and their context within DOE'S new open culture, our research, test methods and results, the significance of our research, and our plans for future research. (author)

  13. Consumo de energia em escalonadores de transações em sistemas de memória transacional em software

    OpenAIRE

    Marques Junior, Ademir [UNESP

    2016-01-01

    O conceito de Memória Transacional foi criado para simplificar a sincronização de dados em memória, necessária para evitar a computação de dados inconsistentes por processadores multinúcleos, que se tornaram padrão devido às limitações encontradas em processadores de um núcleo. Em evolução constante pela busca de desempenho, os escalonadores de transação foram criados como alternativa aos gerenciadores de contenção presentes nos Sistemas de Memória Transacional. O consumo de energia é preocup...

  14. Infracommunities of intestinal helminths of the Red Fox <em>Vulpes vulpesem> (Linnaeus, 1758 from Italian Alps

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    Anna Rita Di Cerbo

    2003-10-01

    Full Text Available <em>Vulpes vulpesem> (Linnaeus, 1758 is one of the most common carnivore in Italy and its spread includes almost the whole national territory. The species shows an high ecological plasticity and a variable diet composition connected in part to human sources. This high adaptability permits the red foxes to colonize different habitats like the suburbs of large cities as well as the small villages located in mountain areas. On the other hand, the tourism pressure seems to assume a great importance in the Alps, also in those areas where the foxes live. So, indirect interactions could take place between these animals and the humans. The role of <em>V. vulpesem> in the zoonoses has not to be understated since this carnivore could transmit parasitic diseases that are able to cause serious pathologies in humans. This study aims just to investigate on intestinal communities of helmiths of <em>V. vulpesem> in order to make progress in current knowledge on epidemiological situation in Italian Alps. During 1998-2003, we have examined 450 foxes from Trentino Alto Adige, Veneto, Lombardia and Valle d'Aosta. The specimens collected were found dead or have been hunted (according to national law n. 157/92 in localities situated between 170 and 2200 m a.s.l. The carcasses were carried to the provincial sections of Zooprofilattici Institutes, where the intestine was drawn by each sample and all the material was sent to the Faculty of Veterinary Medicine of Milan. Parasitological examination of the small intestines was performed by the analysis of the whole sediment and counting technique (SCT. Parasites were preserved in alcohol 70° before to be clarified or stained and identified by microscope (Zeiss Axioscop. Mean abundance, mean intensity and prevalence were calculated for each taxon of helminth. Dates of sampling were grouped within the four seasons. Statistic tests were performed with software package SPSS rel. 11.5 and spatial analysis with the

  15. Prevalência de comportamentos alimentares disfuncionais em mulheres idosas

    OpenAIRE

    Gomes, Fabiana Vanessa Silva

    2015-01-01

    Dissertação de mestrado em Psicologia Aplicada (área de especialização em Psicologia Clínica e da Saúde) Comportamentos alimentares disfuncionais são frequentes em jovens, mas pouco se sabe sobre a sua prevalência em mulheres idosas. Este estudo consiste em estudar a prevalência pontual de comportamentos alimentares disfuncionais em mulheres com 65 anos ou mais. Estudo de prevalência constituído por um momento de avaliação de duas fases com 342 mulheres entre os 65 e 94 anos...

  16. EM International, July 1994, Volume 2

    International Nuclear Information System (INIS)

    1994-01-01

    The Office of Environmental Management (EM) at the Department of Energy (DOE) is seeking out and leveraging foreign technology, data, and resources in keeping with EM's mandate to protect public health and the environment through the safe and cost-effective remediation of the Department's nuclear weapons sites. EM works closely with foreign governments, industry, and universities to obtain innovative environmental technologies, scientific and engineering expertise, and operations experience that will support EM's objectives. Where appropriate, these international resources are used to manage the more urgent risks at our sites, secure a safe workplace, help build consensus on critical issues, and strengthen our technology development program. Through international agreements EM engages in cooperative exchange of information, technology, and individuals. Currently, we are managing agreements with a dozen countries in Europe, Latin America, and Asia. These agreements focus on environmental restoration, waste management, transportation of radioactive wastes, and decontamination and decommissioning. This publication contains the following articles: in situ remediation integrated program; in-situ characterization and inspection of tanks; multimedia environmental pollutant assessment system (MEPAS); LLNL wet oxidation -- AEA technology. Besides these articles, this publication covers: EU activities with Russia; technology transfer activities; and international organization activities

  17. NECESSIDADE E LIBERDADE EM HANNAH ARENDT

    Directory of Open Access Journals (Sweden)

    Odílio Alves Aguiar

    2015-07-01

    Full Text Available O presente artigo visa refletir sobre o pensamento de Hannah Arendt à luz do par conceitual necessidade-liberdade. Esse caminho é produtivo, pois dá conta tanto da obra mais conhecida e reconhecida da autora quanto das suas reflexões anteriores à Origens do Totalitarismo, que se dirigiam, na sua maioria, à comunidade dos Judeus e tinham a questão judaica como ponto central. As meditações arendtianas sobre o judaísmo, consideradas no presente artigo, foram publicadas em duas coletâneas na França: La tradition Cachée (1987 e Auschwitz et Jérusalem (1991. O artigo gravita em torno da defesa arendtiana sobre a inerência entre política e liberdade e sua substituição pela compreensão da política como submetida à esfera da necessidade. Essa perspectiva, segundo a autora, fez-se presente na tradição assimilacionista entre os judeus, mas já tinha obtido a hegemonia na cultura ocidental. Isso sucedeu em razão da primazia da vita contemplativa sobre a vita activa, do trabalho e da questão social em detrimento da ação e, por fim, da supremacia da filosofia da história em prejuízo da Filosofia Política.

  18. Synthesis and Cytotoxicity Evaluation of 13-<em>n>-Alkyl Berberine and Palmatine Analogues as Anticancer Agents

    Directory of Open Access Journals (Sweden)

    Lei Zhang

    2012-09-01

    Full Text Available By introducing long carbon-chain alkyl groups at the C-13 position of berberine and palmatine, 13-<em>n>-hexyl/13-<em>n>-octyl berberine and palmatine chloride analogues 4ad were synthesized and examined by MTT assays for cytotoxic activity in seven human cancer cell lines (7701QGY, SMMC7721, HepG2, CEM, CEM/VCR, KIII, Lewis, yielding IC50 values of 0.02 ± 0.01–13.58 ± 2.84 μM. 13-<em>n>-Octyl palmatine (compound 4d gave the most potent inhibitor activity, with an IC50 of 0.02 ± 0.01 μM for SMMC7721. In all cases, the 13-<em>n>-alkyl berberine and palmatine analogues 4ad were more cytotoxic than berberine and palmatine. In addition, compounds 4ad also exhibited more potent cytotoxicity than berberine and palmatine in mice with S180 sarcoma xenografted<em> in vivoem>. The primary screening results indicated that the 13-<em>n>-hexyl/13-<em>n>-octyl berberine and palmatine analogues might be valuable source for new potent anticancer drug candidates.

  19. Ceratomicose em equinos Equine keratomycosis

    Directory of Open Access Journals (Sweden)

    Paula Diniz Galera

    2012-07-01

    Full Text Available O cavalo, dado o seu meio ambiente, está sujeito a afecções frequentes da córnea e da conjuntiva, tecidos oculares bastante expostos a bactérias e fungos, principalmente Aspergillus spp. e Fusarium spp. As ceratites ulcerativas bacterianas e fúngicas, bem como as ceratites fúngicas não ulcerativas, caracterizadas principalmente pelo abscesso estromal, são frequentes nessa espécie. Ocorrida a lesão inicial, perpetua-se um ciclo vicioso, com liberação de citocinas inflamatórias, que desencadeiam uma rápida e severa infiltração corneal por células polimorfonucleares. A córnea torna-se sujeita à destruição por enzimas proteolíticas liberadas pelos micro-organismos e por células inflamatórias, capazes de desencadear a dissolução estromal e a perfuração do bulbo ocular. O tratamento clínico para a resolução da doença corneal e o controle da uveíte reflexa deve ser agressivo e associado, muitas das vezes, à terapia cirúrgica. Este artigo discorre sobre a fisiopatologia e o tratamento da ceratomicose em equinos.Environmental and behavioral factors make horses susceptible to corneal and conjunctival lesions, since these structures are constantly exposed to bacteria and fungi specially Aspergillus spp. and Fusarium spp. Bacterial and fungal ulcerative keratitis, as well as non-ulcerative fungal keratitis such as stromal abscess, are frequent in horses. A "cascade" effect follows the initial lesion which triggers the release of inflammatory cytokines followed by an acute and severe infiltrate of polymorphonuclear cells in the cornea. The cornea becomes susceptible to the activity of proteolytic enzymes released by microorganisms and polymorphonuclear cells, resulting in stromal degradation and ocular perforation. The medical treatment targeting the corneal disease and the controlling of reflexive uveitis should be aggressive and surgical therapy should be associated in most of the cases. This paper reviews the

  20. Foucault e a pesquisa educacional em arte

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    Marcilio de Souza Vieira

    2008-09-01

    Full Text Available Este texto objetiva compreender como a obra de Foucault pode contribuir para a pesquisa educacional reconhecendo essas contribuições para o conhecimento na área da educação. Medéia e Hamlet, respectivamente dos tragediógrafos Eurípides e Shakespeare são possibilidades de exploração da loucura, da sexualidade e da linguagem que podemos encontrar nas obras de Foucault. Pensamos esta abordagem como lições que norteiam o pensamento de Foucault acerca da pesquisa educacional em Arte, embora o filósofo não tenha escrito sobre educação, mas exemplifica em suas obras, artistas e obras de arte que nos ajudam a compreender a loucura, a linguagem e a sexualidade e discuti-las nas aulas de Arte, em especial, nas aulas de teatro.

  1. Intense EM filamentation in relativistic hot plasmas

    Energy Technology Data Exchange (ETDEWEB)

    Hu, Qiang-Lin [Department of Physics, Jinggangshan University, Ji' an, Jiangxi 343009 (China); Chen, Zhong-Ping [Department of Physics and Institute for Fusion Studies, The University of Texas at Austin, Austin, TX 78712 (United States); Mahajan, Swadesh M., E-mail: mahajan@mail.utexas.edu [Department of Physics and Institute for Fusion Studies, The University of Texas at Austin, Austin, TX 78712 (United States); Department of Physics, School of Natural Sciences, Shiv Nadar University, Uttar Pradesh 201314 (India)

    2017-03-03

    Highlights: • Breaking up of an intense EM pulse into filaments is a spectacular demonstration of the nonlinear wave-plasma interaction. • Filaments are spectacularly sharper, highly extended and longer lived at relativistic temperatures. • EM energy concentration can trigger new nonlinear phenomena with absolute consequences for high energy density matter. - Abstract: Through 2D particle-in-cell (PIC) simulations, we demonstrate that the nature of filamentation of a high intensity electromagnetic (EM) pulse propagating in an underdense plasma, is profoundly affected at relativistically high temperatures. The “relativistic” filaments are sharper, are dramatically extended (along the direction of propagation), and live much longer than their lower temperature counterparts. The thermally boosted electron inertia is invoked to understand this very interesting and powerful phenomenon.

  2. Preditores do Comportamento Antissocial em Adolescentes

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    Fernanda Lüdke Nardi

    Full Text Available RESUMO Este estudo investigou preditores do comportamento antissocial em 142 adolescentes em medida socioeducativa de internação (G1 e 691 estudantes de escolas públicas (G2, que responderam a um questionário. Foram observadas diferenças significativas entre os grupos quanto ao comportamento antissocial, violência intra e extrafamiliar, uso de drogas e eventos estressores, com médias mais altas em G1. Dentre as variáveis investigadas, grupo, uso de drogas e eventos estressores, juntamente com a covariável sexo, explicaram 66,5% da variável dependente. Um modelo de path analysis demonstrou que as variáveis sexo e ambiente estressor, computado pelos eventos estressores, qualidade do relacionamento familiar e violência intra e extrafamiliar predizem o uso de drogas e comportamentos antissociais.

  3. The DOE/EM facility transition program

    International Nuclear Information System (INIS)

    Bixby, W.

    1994-01-01

    The mission of EM-60 is to plan, implement, and manage receipt of surplus facilities resulting from downsizing of the DOE Weapons Complex facilities and DOE operating program offices to EM, and to ensure prompt deactivation of such facilities in order to reach a minimum surveillance and maintenance condition. The revised organizational structure of EM-60 into four offices (one at headquarters, and the other three at field sites), reflects increased operating functions associated with deactivation, surveillance, and maintenance of facilities. EM-60's deactivation and transition role concerns technical, socioeconomic, institutional, and administrative issues. The primary objective of the deactivation process is to put facilities in the lowest surveillance and maintenance condition safely and quickly by driving down the open-quotes mortgageclose quotes costs of maintaining them until final disposition. EM-60's three key activities are: (1) Inventory of surplus facilities - The 1993 Surplus Facility Inventory and Assessment (SFIA) serves as a planning tool to help the Department and EM-60 determine optimal transition phasing, with safety and cost-effectiveness remaining a priority. (2) Management of accelerated facility life cycle transition - Transitions currently underway illustrate site issues. These include addressing the interests of federal and state regulatory agencies as well as interests of local stakeholders, safe management of large amounts of production residues, and options for treatment, storage, transportation, and disposal. Of equal importance in the transition process is planning the optimal transition of the labor force. (3) Economic development - to address the socio-economic impacts on affected communities of the severe and rapid downsizing of the DOE Weapons Complex, DOE is pursuing an approach that uses the land, equipment, technology assets, and highly skilled local workforces as a basis for alternative economic development

  4. Isojacareubin from the Chinese Herb <em>Hypericum japonicumem>: Potent Antibacterial and Synergistic Effects on Clinical Methicillin-Resistant <em>Staphylococcus aureusem> (MRSA

    Directory of Open Access Journals (Sweden)

    Gen-Chun Wang

    2012-07-01

    Full Text Available Through bioassay-guided fractionation of the extracts from the aerial parts of the Chinese herb <em>Hypericum japonicumem> Thunb. Murray, Isojacareubin (ISJ was characterized as a potent antibacterial compound against the clinical methicillin-resistant S<em>taphylococcus aureusem> (MRSA. The broth microdilution assay was used to determine the minimal inhibitory concentrations (MICs and minimal bactericidal concentrations (MBCs of ISJ alone. The results showed that its MICs/MBCs ranged from 4/16 to 16/64 μg/mL, with the concentrations required to inhibit or kill 50% of the strains (MIC50/MBC50 at 8/16 μg/mL. Synergistic evaluations of this compound with four conventional antibacterial agents representing different types were performed by the chequerboard and time-kill tests. The chequerboard method showed significant synergy effects when ISJ was combined with Ceftazidime (CAZ, Levofloxacin (LEV and Ampicillin (AMP, with the values of 50% of the fractional inhibitory concentration indices (FICI50 at 0.25, 0.37 and 0.37, respectively. Combined bactericidal activities were also observed in the time-kill dynamic assay. The results showed the ability of ISJ to reduce MRSA viable counts by log10CFU/mL at 24 h of incubation at a concentration of 1 × MIC were 1.5 (LEV, additivity, 0.92 (CAZ, indifference and 0.82 (AMP, indifference, respectively. These <em>in vitroem> anti-MRSA activities of ISJ alone and its synergy with conventional antibacterial agents demonstrated that ISJ enhanced their efficacy, which is of potential use for single and combinatory therapy of patients infected with MRSA.

  5. AMIDO RESISTENTE EM FARINHAS DE BANANA VERDE

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    Dayana Portes RAMOS

    2010-03-01

    Full Text Available

    Este trabalho teve por objetivo avaliar o teor de amido resistente (AR em farinhas de banana verde produzidas a partir de treze genótipos de bananeira. Para a produção da farinha foram separadas a 1ª, 3ª e 5ª pencas de cada genótipo, na qual cada penca correspondeu a uma repetição. Os frutos de cada penca no estádio 1 (casca completamente verde de maturação foram descascados manualmente, cortados em fatias circulares de 0,5 cm e desidratados em estufa com circulação de ar a 40ºC por 48 horas, sendo em seguida moídos. A análise de AR consistiu em um processo enzimático, calculando-se o conteúdo final pela concentração de glicose liberada. Os resultados foram submetidos à análise estatística e mostraram diferenças significativas para o teor de AR nas farinhas obtidas dos genótipos de bananeira, sendo que a farinha com maior teor de AR foi a produzida a partir do cultivar ‘Nam’ (40,25% e a menor pelo híbrido ‘Fhia 01’ (10,01%. Pode-se concluir que o conteúdo de AR varia em relação ao genótipo utilizado para a confecção da farinha e que a banana pode ser uma boa opção de estudo de alimento funcional.

  6. Roost selection by barbastelle bats (<em>Barbastella barbastellusem>, Chiroptera: Vespertilionidae in beech woodlands of central Italy

    Directory of Open Access Journa