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Avalia o da resistência à Xylella fastidiosa em germoplasma de tangerina e híbridos introduzidos da Itália e Córsega  [cached]
GONZáLEZ JAIMES ELENA PAOLA,SOUZA PAULO SERGIO DE,WICKERT ESTER,DONADIO LUIZ CARLOS
Revista Brasileira de Fruticultura , 2002,
Abstract: A Clorose Variegada dos Citros (CVC), causada pela bactéria Xylella fastidiosa, é atualmente uma das doen as que mais afeta a citricultura brasileira, sendo as variedades de laranja-doce as mais afetadas. Ensaio instalado na Esta o Experimental de Bebedouro (EECB), em condi es de estufa, teve como objetivo avaliar o comportamento em rela o 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 padr o. As avalia es consistiram na observa o de sintomas, teste de ELISA e PCR. Os primeiros sintomas nos materiais contaminados come aram a surgir 7 meses após a inocula o. 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 n o pelo PCR n o foram contadas como suscetíveis e, sim, como falsos positivos.
Seqüenciamento e variabilidade do fragmento gen mico de Xylella fastidiosa amplificado pelos iniciadores RST31/33  [cached]
Wendland Adriane,Daniela Truffi,Leite Júnior Rui P.,Camargo Luis E. A.
Fitopatologia Brasileira , 2003,
Abstract: Xylella fastidiosa é agente causal de diversas doen as de importancia econ mica como a clorose variegada dos citros (Citrus spp.) (CVC), mal de Pierce da videira (Vitis vinifera), escaldadura da ameixeira (Prunus salicina) e requeima do cafeeiro (Coffea arabica). A seqüência nucleotídica do fragmento gen mico, específico de X. fastidiosa, amplificado pelo par de iniciadores RST31/33 foi determinada para 38 isolados de citros e para isolados de videira, cafeeiro e ameixeira objetivando avaliar o nível de polimorfismo entre isolados e a identidade gen mica do fragmento. N o foi observado polimorfismo de seqüência nucleotídica entre isolados de citros, mas foi detectado polimorfismo entre isolados de citros e de videira, cafeeiro e ameixeira. A presen a do sítio de clivagem RsaI, que distingue isolados de citros e videira de isolados de ameixeira e outras espécies arbóreas, foi identificada em um isolado de ameixeira proveniente dos EUA mas n o em outro proveniente do Brasil.
Dinamica temporal da Clorose Variegada dos Citros em três regi es do Estado de S o Paulo  [cached]
Laranjeira Francisco F.,Bergamin Filho Armando,Amorim Lilian,Berger Richard
Fitopatologia Brasileira , 2003,
Abstract: Este trabalho objetivou esclarecer se o progresso da Clorose Variegada dos Citros (CVC) diferia entre três regi es de S o Paulo, distintas quanto à incidência da CVC em citrus (Citrus spp.). Foram avaliadas três áreas, Noroeste, Centro e Sul de S o Paulo, durante dois anos, em avalia es quinzenais, quando eram mapeadas as plantas sintomáticas. Tentou-se o ajuste de nove modelos ao progresso da doen a, além do ajuste de três modelos a segmentos das curvas originais. Foram estimadas também as diferenciais e as diferenciais secundárias de cada curva de progresso. Apenas quando as curvas foram divididas é que foram obtidos bons ajustes aos modelos de progresso. A diferencial (velocidade da doen a) e diferencial secundária (acelera o do aparecimento de novas plantas doentes) apresentaram diversos picos ao longo do tempo. Esses picos ocorreram em meses de Primavera e Ver o. Levanta-se aqui a hipótese de que os picos de diferencial - incomuns na quantidade encontrada - estejam relacionados a determinados picos de emiss o de brota es, já que as novas brota es s o o local preferido de alimenta o dos vetores de Xylella fastidiosa.
Xylella fastidiosa comparative genomics  [cached]
Wim D'Haeze
Genome Biology , 2002, DOI: 10.1186/gb-2002-3-12-reports0064
Abstract: The genome sequences contained 2,731,748 (XFA), 2,625,581 (XFY) and 2,434,849 (XFX) base-pairs (bp). The G+C content was about equal for the three genomes at 52%. The genomes contain 2,985 (XFA), 2,870 (XFY) and 2,681 (XFX) open reading frames (ORFs), 58%, 62% and 62% of which were assigned functions. The XFY and XFX genomes contain a plasmid, of 30,270 and 51,158 bp, respectively, with a slightly lower G+C content of about 49%. The XFY plasmid carries genes encoding a type IV secretion system, a putative transcription regulator and a nickase. Pathway analysis revealed that the genomes contain 958 (XFA), 965 (XFY) and 938 (XFX) pathways. Compared to the genome of XFA, some functions were missing in the other two genomes. These include functions involving ribosomal SSU (1 missing in XFY, 10 missing in XFX), ribosomal LSU (4 missing in XFY, 17 missing in XFX), and aminoacyl-tRNA (1 missing in XFY, 2 missing in XFX). In addition, it was suggested that all X. fastidiosa strains possess a simple aerobic respiratory complex, enabling aerobic respiration only under high oxygen concentrations. No indications of the presence of a cytochrome c oxidase or other type of quinol oxidase as terminal oxidases were noticed. Thus, because cytochrome o (bo) ubiquinone is the only terminal oxidase present in these bacteria, it would be a suitable candidate drug target. Functional reconstruction using data revealed by the genome sequence, enabled growth media to be developed on which X. fastidiosa strains were more easily cultivated.The website of the Xylella fastidiosa genome project provides updates of the project, and the complete proteome is available from EMBL-EBI databases: proteome analysis.Bhattacharyya et al. compared the genome sequences of three plant pathogenic X. fastidiosa strains and performed a functional reconstruction to deduce new growth medium compositions that may facilitate the cultivation of those bacteria. New functions were revealed that might be involved in hos
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  [PDF]
Ping Ouyang, Mohammad Arif, Jacqueline Fletcher, Ulrich Melcher, Francisco Manuel Ochoa Corona
PLOS ONE , 2013, DOI: 10.1371/journal.pone.0081647
Abstract: 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.
Nonlinear models for describing the Citrus Variegated Chlorosis in groves of two counties at northwestern Paraná state, Brazil = Modelo n o-linear para a curva de progresso de incidência da Clorose Variegada dos Citros (CVC) em dois municípios da regi o noroeste do Estado do Paraná
Clara Matiko Ueda,Akemi Yamagata Yamamoto,William Mário de Carvalho Nunes,Carlos Alberto Scapim
Acta Scientiarum : Agronomy , 2010,
Abstract: 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 Esperan a the data were best fitted to the parameterization L5, which is also a parameterization from the Logistic model. A citricultura é afetada por diversas doen as, dentre as quais a Clorose Variegada dos Citros (CVC) causada pela bactéria Xylella fastidiosa (Wells). Para a regi o noroeste do Estado do Paraná, onde foi avaliada a CVC, prop e-se encontrar modelos n o-lineares de curvas de progresso de incidência da CVC que representem o percentual de plantas acometidas pela doen a. Para avaliar o comprometimento dos pomares com rela o à doen a, foram escolhidos pomares comerciais em dois municípios, onde foi determinada a propor o de plantas doentes. Foram selecionados talh es de laranja doce (Citrus sinensis Osbeck) nas variedades “Pêra”, “Valência”, “Natal” e “Folha Murcha” e a avalia o de todas as plantas do talh o foi realizada visualmente em rela o à presen a ou à ausência de sintomas de CVC. Para estimar o modelo que melhor se ajustou aos dados de progresso da propor o da doen a em cada talh o, foram considerados modelos n o-lineares decrescimento sigmoidal, o logístico e o de Gompertz, conforme sugerido na literatura. A escolha do melhor modelo foi com base em medidas de curvatura intrínseca e paramétrica de Bates e Watts, viés de Box e medida de assimetria de Hougaard. O modelo que melhor ajustou aos dados de Loanda foi o modelo de Fermi-Dirac e aos dados de Nova Esperan a foi outra reparametriza o do Modelo Logístico, denominado de L5.
Analysis of the biofilm proteome of Xylella fastidiosa
Mariana S Silva, Alessandra A De Souza, Marco A Takita, Carlos A Labate, Marcos A Machado
Proteome Science , 2011, DOI: 10.1186/1477-5956-9-58
Abstract: 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.We observed overexpression of proteins related to quorum sensing, proving the existence of communication between cells, and thus the development of structuring the biofilm (mature biofilm) leading to obstruction of vessels and development of disease. This paper reports a first proteomic analysis of mature biofilm of X. fastidiosa, opening new perspectives for understanding the biochemistry of mature biofilm growth in a plant pathogen.Xylella fastidiosa is a slow growing Gram-negative bacterium involved in many economically important plant diseases, such as citrus variegated chlorosis (CVC) in sweet orange, Pierce's disease (PD) in grapevine and other species such as coffee and plum. In all cases, X. fastidiosa is transmitted by leafhoppers into the xylem vessel where it colonizes and blocks the movement of water and nutrients, causing typical disease symptoms according to the host.It is generally accepted that microbial populations use cell attachment to adhere to solid supports, surfaces and particles where they grow
Morphological evidence for phages in Xylella fastidiosa
Jianchi Chen, Edwin L Civerolo
Virology Journal , 2008, DOI: 10.1186/1743-422x-5-75
Abstract: Xylella fastidiosa [1] is a Gram negative plant pathogen causing many economically important diseases including Pierce's disease (PD) of grapevine, almond leaf scorch disease and citrus variegated chlorosis disease. Because of nutritional fastidiousness, many biological aspects of the bacterium including the occurrence of phages are difficult to study. Analyses of whole genome sequences of X. fastidosa strains identified many prophage sequences [2-5], including putative Siphoviridae [2,4], Podoviridae [6] and Inoviridae [3] phages. Yet, physical evidence for the presence of phage particles in X. fastidiosa is very limited. Lauzon and Miller [7] reported the association of particles resembling phages in the families Microviridae and Podoviridae with X. fastidiosa. However, only limited details regarding the origin(s) or nature of these particles were provided. Chen et al. [6] reported a phage DNA sequence of 547 bp from the genome of a PD strain isolated in Florida. The sequence shared high similarity to that of an integrase gene in the Podoviridae phage family. Interestingly, this sequence is absent in the whole genome sequence of a California PD strain Temecula-1, but is present in other California PD strains. In this paper, we report our observations of presumptive phage particles in a X. fastidiosa PD strain through transmission electron microscopy (TEM).Phage observations were first made with intact bacterial cells. X. fastidosa strain Temecula-1 was cultured in 30 ml of PW broth medium [8] for 30 days at 28 C. Before bacterial cell collection, a loop of bacterial culture was streaked on PW plate and incubated at 28 C to check for possible contamination based on culture characteristics (slow growing opalescent colonies with entire smooth margin) as well as PCR [9]. Bacterial cells were then collected by centrifugation at 3,000 g for 30 minutes. Cell pellets were suspended in 1 ml of TE (10 mM Tris-HCl, pH 8.0 and 50 mM EDTA) buffer, transferred to a 1.5 ml micro
Xylella fastidiosa gene expression analysis by DNA microarrays
Travensolo, Regiane F.;Carareto-Alves, Lucia M.;Costa, Maria V.C.G.;Lopes, Tiago J.S.;Carrilho, Emanuel;Lemos, Eliana G.M.;
Genetics and Molecular Biology , 2009, DOI: 10.1590/S1415-47572009005000038
Abstract: 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.
The Bacterial Pathogen Xylella fastidiosa Affects the Leaf Ionome of Plant Hosts during Infection  [PDF]
Leonardo De La Fuente, Jennifer K. Parker, Jonathan E. Oliver, Shea Granger, Phillip M. Brannen, Edzard van Santen, Paul A. Cobine
PLOS ONE , 2013, DOI: 10.1371/journal.pone.0062945
Abstract: 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.
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