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

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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....

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

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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...

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

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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

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

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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.

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

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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.

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

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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

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

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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.

Chromosome-based genetic complementation system for Xylella fastidiosa.

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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.

Potential complications when developing gene deletion clones in Xylella fastidiosa.

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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.

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

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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.

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

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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.

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

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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.

Comparative genomic characterization of citrus-associated Xylella fastidiosa strains

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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

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

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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.

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

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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.

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

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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.

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

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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.

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

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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.

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

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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...

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

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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.

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

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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.

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

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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.

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

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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.

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

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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.

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.

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

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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.

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

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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

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

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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.

Functional characterization of the role of the PilG in Xylella fastidiosa

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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...

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

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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...

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

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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.

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

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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.

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

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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...

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

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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.

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

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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.

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

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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...

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 ...

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

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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.

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

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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

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

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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

Characterization of a diffusible signaling factor from Xylella fastidiosa.

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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.

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

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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...

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

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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...

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...

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.

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

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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...

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

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

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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.

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

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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.

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

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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.

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

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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.

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

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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.

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

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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...

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

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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...

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

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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 ...

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

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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.

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

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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.

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

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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.

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

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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.

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

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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.

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

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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.

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

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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...

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

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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.

Xylella fastidiosa: Host Range and Advance in Molecular Identification Techniques

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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

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

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

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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.

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

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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.

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

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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.

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

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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...

Chitin utilization by the insect-transmitted bacterium Xylella fastidiosa.

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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.

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

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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.

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

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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.

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

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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.

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

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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...

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

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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.

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

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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.

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

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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.

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

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

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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.

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

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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.

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.

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

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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

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

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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.

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

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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.

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

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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.

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

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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...

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

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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.

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.

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

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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

RpfF-dependent regulon of Xylella fastidiosa.

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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.

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

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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.

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

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

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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...

Network analysis reveals why Xylella fastidiosa will persist in Europe.

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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.

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.

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

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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.

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.

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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.

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

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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.

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

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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.

Transmission of Xylella fastidiosa to Grapevine by the Meadow Spittlebug.

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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.

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

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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...

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...

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

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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...

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

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

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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...

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

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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.

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

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

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...

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 ...

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

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

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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’ ...

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.

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.

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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.

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.

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

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

Xylella fastidiosa gene expression analysis by DNA microarrays

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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.

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

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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.

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

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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

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

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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.

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°

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

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

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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.

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

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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.

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

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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

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

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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.

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

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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...

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

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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.

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

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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...

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.

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

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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...

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

' de C. canephora, foram os tratamentos com maior severidade e, novamente no período chuvoso, as diferenças não foram observadas. Concluiu-se, portanto, que a prática da poda do tipo decote não resultou nesse experimento em um controle eficiente da X. fastidiosa em cafeeiros de C. arabica de pé-franco ou enxertados e infectados por esta moléstia.Xylella fastidiosa plant colonization causes damages to the coffee production and nowadays there is no available economic chemical control to this bacterium. Adequate cultivation management as the use of bacteria-free scions and control of the insect vector (cicadas, are practices that may attenuate the disease incidence. Pruning, that has been recommended for citrus and vines for disease control, still does not have its efficiency proved for coffee plants. This research work aimed to quantify the proportion of xylem vessel elements obstructed by the bacteria, as well as to evaluate the disease external symptoms severity after the "decote" type pruning. In June 2003 (dry season, eight months after pruning, it was observed that 4% of the petiole vessel elements were obstructed by X. fastidiosa, 2% of the central vein and 1% of the stem. In the rainy season, 14 months after pruning, the obstruction proportion of vessel elements decreased to 2% in the petiole and to 1% in central vein and stem respectively. Therefore, the pruning practice decreased slightly the xylem vessel obstruction caused by the bacteria in the dry period, once prior to pruning a 6% obstruction in the petiole was observed. The new branches sprouted during the rainy season seemed to compensate obstruction in older branches, thus decreasing vase obstruction proportion. In 2003 there were no differences in the disease symptom severity among treatments in the two periods (dry and rainy season, however, in the dry period of 2004, 'Catuaí Vermelho IAC 81' and 'Mundo Novo IAC 515-20' cultivars, grafted on C. canephora 'Apoatã IAC 2258', presented higher

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

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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.

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

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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.

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

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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...

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

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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.

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.

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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.

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

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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.

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

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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

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

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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.

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

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

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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

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

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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.

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

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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

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

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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.

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

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

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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.

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

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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.

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

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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.

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.

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

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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.

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

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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

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

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

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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.

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

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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.

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.

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

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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.

Xylella fastidiosa esterase rather than hydroxynitrile lyase.

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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.

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

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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.

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

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

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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

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

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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.

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

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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.

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

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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.

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

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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.

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

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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.

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

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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.

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

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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.

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

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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.

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

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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.

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

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

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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

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.

RNA metabolism in Xylella fastidiosa during cold adaptation and survival responses

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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...

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

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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.

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

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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.

Functional characterization of the role of rpfA in Xylella fastidiosa

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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...

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

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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.

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

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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.

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...

Genomic insights into Xylella fastidiosa interactions with plant and insect hosts

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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 ...

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

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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...

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.

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

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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.

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...

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

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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 ...

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.

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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.

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

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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.

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

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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.

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

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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.

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

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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.

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

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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...

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.

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.

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

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

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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.

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.

Plastic strain caused by contraction of pores in polycrystalline graphites

International Nuclear Information System (INIS)

Ioka, Ikuo; Yoda, Shinichi; Konishi, Takashi.

1989-01-01

The effects of porosity on mechanical properties and deformation behavior of four isotropic polycrystalline graphites were studied. The pore size distributions of the graphites were measured using a conventional mercury penetration technique. The average pore radius of ISO-88 graphite was about one-tenth of that of ISEM-1, IG-11 or IG-15 graphites. Young's modulus of the graphites decreased with increasing porosity. The stress-strain curve of each graphite was measured in its lateral and axial directions. Young's modulus of graphite decreased with increasing load. The plastic strain at a given compressive load was calculated from the stress-strain curve and the initial gradient of the unloading curve at the load. The ratio of lateral plastic strain to axial plastic strain for the graphites was less than 0.5, indicating that the volume of the graphites decreased during compressive loading. By assuming that the volume change was caused by contraction of pores, plastic strain associated with contraction of pores was calculated from the axial plastic strain and lateral plastic strain by slips along the basal planes. The plastic strain increased with increasing axial plastic strain and porosity of graphite. (author)

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.

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

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

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

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...

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

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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.

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

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

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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.

Anterior foregut microbiota of the glassy-winged sharpshooter explored using deep 16S rRNA gene sequencing from individual insects.

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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

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

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

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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.

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.

Comparative Genomics of Escherichia coli Strains Causing Urinary Tract Infections

DEFF Research Database (Denmark)

Vejborg, Rebecca Munk; Hancock, Viktoria; Schembri, Mark A.

2011-01-01

The virulence determinants of uropathogenic Escherichia coli have been studied extensively over the years, but relatively little is known about what differentiates isolates causing various types of urinary tract infections. In this study, we compared the genomic profiles of 45 strains from a range...

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

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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.

Origins of the E. coli strain causing an outbreak of hemolytic-uremic syndrome in Germany

DEFF Research Database (Denmark)

Rasko, David A; Webster, Dale R; Sahl, Jason W

2011-01-01

A large outbreak of diarrhea and the hemolytic-uremic syndrome caused by an unusual serotype of Shiga-toxin-producing Escherichia coli (O104:H4) began in Germany in May 2011. As of July 22, a large number of cases of diarrhea caused by Shiga-toxin-producing E. coli have been reported--3167 without...... the hemolytic-uremic syndrome (16 deaths) and 908 with the hemolytic-uremic syndrome (34 deaths)--indicating that this strain is notably more virulent than most of the Shiga-toxin-producing E. coli strains. Preliminary genetic characterization of the outbreak strain suggested that, unlike most of these strains......, it should be classified within the enteroaggregative pathotype of E. coli....

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.

Genomic analysis of a Streptococcus pyogenes strain causing endocarditis in a child

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M. Beye

2017-05-01

Full Text Available We sequenced the genome of Streptococcus pyogenes strain G773 that caused an infective endocarditis in a 4-year-old boy suffering from acute endocarditis. The 1.9-Mb genome exhibited a specific combination of virulence factors including a complete integrative and conjugative element, sp2905, previously described as incomplete in S. pyogenes, and five bacteriocin-coding genes. However, strain G773 lacked a CRISPR-Cas system.

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.

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...

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.

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.

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

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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

Purulent Pericarditis Caused by Streptococcus Milleri Strains; the Gained Experience from Nine Reported Cases

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Ilias A. Kouerinis

2015-12-01

Full Text Available Bacterial pericarditis caused by Streptococcus Milleri Group (SMG strains is a particularly exceptional pathology. All the eight previous reported cases were under the care of medical teams and the seven existed reports in medical journals were more or less from this perspective. Herein, we reported a unique case of a pericardial-cutaneous fistula resulting from a recurrent purulent pericardial effusion caused by SMG strains, which had been treated with open surgical drainage two months before. A thorough review of the surgical treatment options and the results has also been presented.

Whole-Genome Characterization and Strain Comparison of VT2f-Producing Escherichia coli Causing Hemolytic Uremic Syndrome

Science.gov (United States)

Michelacci, Valeria; Bondì, Roslen; Gigliucci, Federica; Franz, Eelco; Badouei, Mahdi Askari; Schlager, Sabine; Minelli, Fabio; Tozzoli, Rosangela; Caprioli, Alfredo; Morabito, Stefano

2016-01-01

Verotoxigenic Escherichia coli infections in humans cause disease ranging from uncomplicated intestinal illnesses to bloody diarrhea and systemic sequelae, such as hemolytic uremic syndrome (HUS). Previous research indicated that pigeons may be a reservoir for a population of verotoxigenic E. coli producing the VT2f variant. We used whole-genome sequencing to characterize a set of VT2f-producing E. coli strains from human patients with diarrhea or HUS and from healthy pigeons. We describe a phage conveying the vtx2f genes and provide evidence that the strains causing milder diarrheal disease may be transmitted to humans from pigeons. The strains causing HUS could derive from VT2f phage acquisition by E. coli strains with a virulence genes asset resembling that of typical HUS-associated verotoxigenic E. coli. PMID:27584691

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.

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

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...

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...

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

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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

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

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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

Acute Hepatopancreatic Necrosis Disease-Causing Vibrio parahaemolyticus Strains Maintain an Antibacterial Type VI Secretion System with Versatile Effector Repertoires.

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Li, Peng; Kinch, Lisa N; Ray, Ann; Dalia, Ankur B; Cong, Qian; Nunan, Linda M; Camilli, Andrew; Grishin, Nick V; Salomon, Dor; Orth, Kim

2017-07-01

Acute hepatopancreatic necrosis disease (AHPND) is a newly emerging shrimp disease that has severely damaged the global shrimp industry. AHPND is caused by toxic strains of Vibrio parahaemolyticus that have acquired a "selfish plasmid" encoding the deadly binary toxins PirA vp /PirB vp To better understand the repertoire of virulence factors in AHPND-causing V. parahaemolyticus , we conducted a comparative analysis using the genome sequences of the clinical strain RIMD2210633 and of environmental non-AHPND and toxic AHPND isolates of V. parahaemolyticus Interestingly, we found that all of the AHPND strains, but none of the non-AHPND strains, harbor the antibacterial type VI secretion system 1 (T6SS1), which we previously identified and characterized in the clinical isolate RIMD2210633. This finding suggests that the acquisition of this T6SS might confer to AHPND-causing V. parahaemolyticus a fitness advantage over competing bacteria and facilitate shrimp infection. Additionally, we found highly dynamic effector loci in the T6SS1 of AHPND-causing strains, leading to diverse effector repertoires. Our discovery provides novel insights into AHPND-causing pathogens and reveals a potential target for disease control. IMPORTANCE Acute hepatopancreatic necrosis disease (AHPND) is a serious disease that has caused severe damage and significant financial losses to the global shrimp industry. To better understand and prevent this shrimp disease, it is essential to thoroughly characterize its causative agent, Vibrio parahaemolyticus Although the plasmid-encoded binary toxins PirA vp /PirB vp have been shown to be the primary cause of AHPND, it remains unknown whether other virulent factors are commonly present in V. parahaemolyticus and might play important roles during shrimp infection. Here, we analyzed the genome sequences of clinical, non-AHPND, and AHPND strains to characterize their repertoires of key virulence determinants. Our studies reveal that an antibacterial type

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

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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 ...

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

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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.

Variation in genotype and higher virulence of a strain of Sporothrix schenckii causing disseminated cutaneous sporotrichosis.

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Zhang, Zhenying; Liu, Xiaoming; Lv, Xuelian; Lin, Jingrong

2011-12-01

Sporotrichosis is usually a localized, lymphocutaneous disease, but its disseminated type was rarely reported. The main objective of this study was to identify specific DNA sequence variation and virulence of a strain of Sporothrix schenckii isolated from the lesion of disseminated cutaneous sporotrichosis. We confirmed this strain to be S. schenckii by(®) tubulin and chitin synthase gene sequence analysis in addition to the routine mycological and partial ITS and NTS sequencing. We found a 10-bp deletion in the ribosomal NTS region of this strain, in reference to the sequence of control strains isolated from fixed cutaneous sporotrichosis. After inoculated into immunosuppressed mice, this strain caused more extensive system involvement and showed stronger virulence than the control strain isolated from a fixed cutaneous sporotrichosis. Our study thus suggests that different clinical manifestation of sporotrichosis may be associated with variation in genotype and virulence of the strain, independent of effects due to the immune status of the host.

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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.

ASPECTOS ESTRUTURAIS DE CAFEEIRO INFECTADO COM XYLELLA FASTIDIOSA

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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

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

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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.

ORF Alignment: NC_002488 [GENIUS II[Archive

Lifescience Database Archive (English)

Full Text Available fastidiosa (strain ... 9a5c) ... Length = 101 ... Query: 10 ... TSHRPYLLRALVEWIND...NGMTPHILVDAGIQGVQVPESAVKDGRVVLNIAGRAVMHLHIG 69 ... TSHRPYLLRALVEWINDNGMTPHILVDAGIQGVQVPESAVK...DGRVVLNIAGRAVMHLHIG Sbjct: 1 ... TSHRPYLLRALVEWINDNGMTPHILVDAGIQGVQVPESAVKDGRVVLNIAGRAVMHLHIG 60 ...

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...

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

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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.

Evaluation of strain caused by coherent precipitates in an Al alloy using TEM techniques

Energy Technology Data Exchange (ETDEWEB)

Hernandez-Rivera, J.L., E-mail: jose.hernandez@cimav.edu.mx [Centro de Investigacion en Materiales Avanzados (CIMAV), Laboratorio Nacional de Nanotecnologia, Miguel de Cervantes 120, Z.C. 31109, Chihuahua (Mexico); Universidad del Valle de Mexico, Robles 600, Fraccionamiento Jacarandas, Z. C. 78220, San Luis Potosi (Mexico); Rivera, J.J. Cruz; Garay-Reyes, C.G.; Azpeitia, M. Ramos; Zuniga-Alonso, I. [Facultad de Ingenieria-Instituto de Metalurgia, Universidad Autonoma de San Luis Potosi, Sierra Leona 550, Lomas 2a seccion, Z.C. 78210, San Luis Potosi (Mexico); Martinez-Sanchez, R. [Centro de Investigacion en Materiales Avanzados (CIMAV), Laboratorio Nacional de Nanotecnologia, Miguel de Cervantes 120, Z.C. 31109, Chihuahua (Mexico)

2012-11-15

Elastic strains, caused by GP zones in an aged Al alloy, were determined quantitatively using two techniques: Dark Field In-line Holography (DFH) and High Resolution Transmission Electron Microscopy-Geometric Phase Analysis (HRTEM-GPA). The results obtained by both techniques showed that the elastic strain was not uniform along the precipitate-matrix interface. In some areas, it was found that strain had negligible value and this was attributed to the loss of coherence between the lattices. It is suggested that a possible explanation for this fact could be a variation in the 'vacancies pump mechanism' kinetics. To obtain a better interpretation of the experimental deformation maps, a reference GP precipitate-matrix structure was built using QSTEM software. The main advantages of DFH over HRTEM-GPA were a bigger field of view and low electron dose requirements without spatial resolution loss. Another difference found was that crystalline defects such as dislocations were evidenced by HRTEM-GPA in contrast to the result obtained by DFH. However, strain measurements were affected by mask size effect in the former. - Highlights: Black-Right-Pointing-Pointer Strain around GP precipitates was not uniform and was in a more relaxed state. Black-Right-Pointing-Pointer Strain values determined by HRTEM had more artifacts compared with holography ones. Black-Right-Pointing-Pointer Strain distribution was attributed to differences in the release of vacancies. Black-Right-Pointing-Pointer Holography showed important advantages over HRTEM regarding field of view mainly.

Evaluation of strain caused by coherent precipitates in an Al alloy using TEM techniques

International Nuclear Information System (INIS)

Hernández-Rivera, J.L.; Rivera, J.J. Cruz; Garay-Reyes, C.G.; Azpeitia, M. Ramos; Zúñiga-Alonso, I.; Martínez-Sánchez, R.

2012-01-01

Elastic strains, caused by GP zones in an aged Al alloy, were determined quantitatively using two techniques: Dark Field In-line Holography (DFH) and High Resolution Transmission Electron Microscopy-Geometric Phase Analysis (HRTEM-GPA). The results obtained by both techniques showed that the elastic strain was not uniform along the precipitate–matrix interface. In some areas, it was found that strain had negligible value and this was attributed to the loss of coherence between the lattices. It is suggested that a possible explanation for this fact could be a variation in the “vacancies pump mechanism” kinetics. To obtain a better interpretation of the experimental deformation maps, a reference GP precipitate–matrix structure was built using QSTEM software. The main advantages of DFH over HRTEM-GPA were a bigger field of view and low electron dose requirements without spatial resolution loss. Another difference found was that crystalline defects such as dislocations were evidenced by HRTEM-GPA in contrast to the result obtained by DFH. However, strain measurements were affected by mask size effect in the former. - Highlights: ► Strain around GP precipitates was not uniform and was in a more relaxed state. ► Strain values determined by HRTEM had more artifacts compared with holography ones. ► Strain distribution was attributed to differences in the release of vacancies. ► Holography showed important advantages over HRTEM regarding field of view mainly.

Molecular Identification and Genetic Characterization of Macrophomina phaseolina Strains Causing Pathogenicity on Sunflower and Chickpea

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Ali N. Khan

2017-07-01

Full Text Available Macrophomina phaseolina is the most devastating pathogen which causes charcoal rot and root rot diseases in various economically important crops. Three strains M. phaseolina 1156, M. phaseolina 1160, and M. phaseolina PCMC/F1 were tested for their virulence on sunflower (Helianthus annuus L. and chickpea (Cicer arietinum L.. The strains showed high virulence on both hosts with a disease score of 2 on chickpea and sunflower. The strains also increased the hydrogen per oxide (H2O2 content by 1.4- to 1.6-fold in root as well as shoot of chickpea and sunflower. A significant increase in antioxidant enzymes was observed in fungal infected plants which indicated prevalence of oxidative stress during pathogen propagation. The M. phaseolina strains also produced hydrolytic enzymes such as lipase, amylase, and protease with solubilization zone of 5–43 mm, 5–45 mm, and 12–35 mm, respectively. The M. phaseolina strains were identified by 18S rRNA and analyzed for genetic diversity by using random amplified polymorphic DNA (RAPD markers. The findings based on RAPD markers and 18S rRNA sequence analysis clearly indicate genetic variation among the strains collected from different hosts. The genetically diverse strains were found to be pathogenic to sunflower and chickpea.

Eradication of the corrosion-causing bacterial strains Desulfovibrio vulgaris and Desulfovibrio desulfuricans using photodisinfection

Energy Technology Data Exchange (ETDEWEB)

Street, C.N.; Gibbs, A.J. [Biocorrosion Solutions Inc., Edmonton, AB (Canada)

2010-07-01

Microbiologically influenced corrosion (MIC) can cause oil and gas pipelines to fail prematurely. The free-floating bacteria collects on the inner pipeline surface to form complex adherent biofilms. This study evaluated the use of photodisinfection as a means of treating 2 sulfate-reducing bacterial strains known to contribute to MIC. The sulfate-reducing strains Desulfovibrio vulgaris and Desulfovibrio desulfuricans were studied experimentally to a concentration of 10{sup 7} colony-forming units per millimeter. Bacterial inocula was made to an optical density of 0.150 at 420 nm in order to assess biofilm growth. The study showed that photodisinfection was able to eradicate more than 99 per cent of the bacterial populations prepared in the study. The method was highly effective in removing the biofilms known to cause MIC in oil and gas pipelines. A close-loop dynamic flow system model will be prepared to evaluate the ability of photodisinfection to inhibit bacterially-influenced corrosion of steel coupons. 24 refs., 3 tabs., 1 fig.

Diarrhea, Urosepsis and Hemolytic Uremic Syndrome Caused by the Same Heteropathogenic Escherichia coli Strain

NARCIS (Netherlands)

Ang, C. Wim; Bouts, Antonia H. M.; Rossen, John W. A.; van der Kuip, Martijn; van Heerde, Marc; Bökenkamp, Arend

2016-01-01

We describe an 8-month-old girl with diarrhea, urosepsis and hemolytic uremic syndrome caused by Escherichia coli. Typing of cultured E. coli strains from urine and blood revealed the presence of virulence factors from multiple pathotypes of E. coli. This case exemplifies the genome plasticity of E.

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.

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

Increased cytotoxicity and streptolysin O activity in group G streptococcal strains causing invasive tissue infections

DEFF Research Database (Denmark)

Siemens, Nikolai; Kittang, Bård R; Chakrakodi, Bhavya

2015-01-01

Streptococcus dysgalactiae subsp. equisimilis (SDSE) has emerged as an important cause of severe skin and soft tissue infections, but little is known of the pathogenic mechanisms underlying tissue pathology. Patient samples and a collection of invasive and non-invasive group G SDSE strains (n = 6...

Frequency of resistance to penicillin and erythromycin of pneumococcal strains that caused ottis media

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Jovanović Luka

2017-01-01

Full Text Available Introduction: Streptococcus pneumoniae is an important human pathogen and the most common cause of acute otitis media (AOM, especially in children. It is also a common cause of community acquired pneumonia, sepsis and bacterial meningitis. Drug of choice in the treatment of these disease are beta lactam antibiotics, and the first alternative are macrolides. The increasing prevalence of resistance to penicillin and macrolides, among pneumococci, has considerably complicated the treatment. Aim: The aim of this study was to determine susceptibility of pneumococcal isolates from pediatric AOM in Serbia to antibiotics. Material and methods: Antimicrobial susceptibility testing of 61 pneumococcal AOM was performed, collected from December 2014 to December 2015, using disk diffusion method and E test. Macrolide resistance profile was determined by double disk diffusion test. Results: In our study, 40 strains (65.6% showed reduced sensitivity to penicillin and erythromycin. There were 9 (14.8% high resistant isolates to penicillin, while 31 (50.8% showed reduced susceptibility. The most frequent resistance phenotype was cMLS. Co-resistance to penicillin and macrolides was found in 14.8% strains. Conclusion: Our results showed high resistance rate of S. pneumoniae, which causes AOM among children, to penicillin and macrolides. Further active surveillance of pneumococcal susceptibility to antibiotics is necessary, and use of these medications in empirical therapy should be limited.

Evaluation of Trichoderma spp. strains for control yellowing pea caused by Fusarium oxysporum

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Christian Eraso Insuasty

2014-07-01

Full Text Available The yellowing of pea caused by the fungus Fusarium oxysporum f. sp. pisi is considered the most damaging disease of this crop. This study took place at the plant health laboratory and greenhouse of the Universidad de Nariño, and the experimental stage was conducted at the Granja experimental Botana. Its purpose was to evaluate the antagonistic ability of the fungi Trichoderma spp. to F. oxysporum. Isolation of F. oxysporum was made from diseased tissue; Trichoderma strains were obtained from the rhizosphere of healthy plants (collected in the towns of Potosi, Córdoba, Gualmatán, Ipiales and Puerres in the state of Nariño, Colombia, and a commercial strain from laboratory Perkins Ltda. In laboratory, unrestrictedly randomized design with 21 treatments (strains was used. Mycelial growth and inhibition zone were evaluated in dual plantings, which served as selection criteria for greenhouse test where plant height, root length, root dry matter and percentage of incidence were evaluated. In the field, a randomized block design was used to evaluate yield components, plant height and root length with the best strains. In the laboratory, C2 (Córdoba 2, C7 (Gualmatán 3, C14 (Puerres 2, C20 (Potosi 4 and C21 (Perkins Lab. showed antagonistic activity in the greenhouse, C7, C14 and C21 were the best; in field, significant differences between C14 and C21, compared to C7 and the control, were obtained. Strains C14 and C21 have consistent antagonistic capacity and can be used to control F. oxysporum in pea.

Genomic and pathogenic analysis of a Muscovy duck parvovirus strain causing short beak and dwarfism syndrome without tongue protrusion.

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Fu, Qiuling; Huang, Yu; Wan, Chunhe; Fu, Guanghua; Qi, Baomin; Cheng, Longfei; Shi, Shaohua; Chen, Hongmei; Liu, Rongchang; Chen, Zhenhai

2017-12-01

In 2008, clinical cases of short beak and dwarfism syndrome (SBDS) caused by Muscovy duck parvovirus (MDPV) infection were found in mule duck and Taiwan white duck farms in Fujian, China. A MDPV LH strain causing duck SBDS without tongue protrusion was isolated in this study. Phylogenetic analysis show that the MDPV LH strain was clustered together with other MDPV strains, but divergent from GPV isolates. Two major fragment deletions were found in the inverted terminal repeats (ITR) of MDPV LH similar to the ones in the ITR of MDPV GX5, YY and SAAS-SHNH strains. To investigate the pathogenicity of the MDPV LH strain, virus infection of young mule ducks was performed. The infected ducks showed SBDS symptoms including retard growth and shorten beaks without tongue protrusion. Atrophy of thymus, spleen and bursa of Fabricius was identified in the infected ducks. The results show that MDPV LH strain is moderately pathogenic to mule duck, leading to occurrence of SBDS. As far as we know, it is the first study showing that SBDS without tongue protrusion, and atrophy of thymus, spleen and bursa of Fabricius possibly associated with immunosuppression were found in the MDPV-infected ducks. The established duck-MDPV-SBDS system will help us to further work on the virus pathogenesis and develop efficacious vaccine against MDPV infection. Copyright © 2017. Published by Elsevier Ltd.

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)

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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 ...

Alfalfa and pastures: sources of pests or generalist natural enemies?

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Pierce’s disease of grapevine and almond leaf scorch disease are both caused by the bacterial pathogen Xyllela fastidiosa. In the Central Valley of California, the green sharpshooter is the most common vector of X. fastidiosa. As alfalfa fields and pastures are considered source habitats for green s...

Toxin Gene Analysis of a Variant Strain of Clostridium difficile That Causes Human Clinical Disease

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Sambol, Susan P.; Merrigan, Michelle M.; Lyerly, David; Gerding, Dale N.; Johnson, Stuart

2000-01-01

A toxin variant strain of Clostridium difficile was isolated from two patients with C. difficile-associated disease (CDAD), one of whom died from extensive pseudomembranous colitis. This strain, identified by restriction endonuclease analysis (REA) as type CF2, was not detected by an immunoassay for C. difficile toxin A. Culture supernatants of CF2 failed to elicit significant enterotoxic activity in the rabbit ileal loop assay but did produce atypical cytopathic effects in cell culture assay. Southern hybridization, PCR amplification, and DNA sequence analyses were performed on the toxin A (tcdA) and toxin B (tcdB) genes of type CF2 isolate 5340. Type CF2 5340 tcdA exhibited a 1,821-bp truncation, due to three deletions in the 3′ end of the gene, and a point mutation in the 5′ end of the gene, resulting in a premature stop codon at tcdA position 139. Type CF2 5340 tcdB exhibited multiple nucleotide base substitutions in the 5′ end of the gene compared to tcdB of the standard toxigenic strain VPI 10463. Type CF2 5340 toxin gene nucleotide sequences and deduced amino acid sequences showed a strong resemblance to those of the previously described variant C. difficile strain 1470, a strain reported to have reduced pathogenicity and no association with clinical illness in humans. REA of strain 1470 identified this strain as a distinct type (CF1) within the same REA group as the closely related type CF2. A review of our clinical-isolate collection identified five additional patients infected with type CF2, three of whom had documented CDAD. PCR amplification of the 3′ end of tcdA demonstrated identical 1.8-kb deletions in all seven type CF2 isolates. REA type CF2 is a toxin variant strain of C. difficile that retains the ability to cause disease in humans but is not detected in clinical immunoassays for toxin A. PMID:10992443

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

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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.

Genetic structure and natural variation associated with host of origin in Penicillium expansum strains causing blue mould.

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Sanzani, S M; Montemurro, C; Di Rienzo, V; Solfrizzo, M; Ippolito, A

2013-07-15

Blue mould, caused by Penicillium expansum, is one of the most economically damaging postharvest diseases of pome fruits, although it may affect a wider host range, including sweet cherries and table grapes. Several reports on the role of mycotoxins in plant pathogenesis have been published, but few focussed on the influence of mycotoxins on the variation in host preference amongst producing fungi. In the present study the influence of the host on P. expansum pathogenicity/virulence was investigated, focussing mainly on the relationship with patulin production. Three P. expansum strain groups, originating from apples, sweet cherries, and table grapes (7 strains per host) were grown on their hosts of isolation and on artificial media derived from them. Strains within each P. expansum group proved to be more aggressive and produced more patulin than the other two groups under evaluation when grown on the host from which they originated. Table grape strains were the most aggressive (81% disease incidence) and strongest patulin producers (up to 554μg/g). The difference in aggressiveness amongst strains was appreciable only in the presence of a living host, suggesting that the complex pathogen-host interaction significantly influenced the ability of P. expansum to cause the disease. Incidence/severity of the disease and patulin production proved to be positively correlated, supporting the role of patulin as virulence/pathogenicity factor. The existence of genetic variation amongst isolates was confirmed by the High Resolution Melting method that was set up herein, which permitted discrimination of P. expansum from other species (P. chrysogenum and P. crustosum) and, within the same species, amongst the host of origin. Host effect on toxin production appeared to be exerted at a transcriptional level. Copyright © 2013 Elsevier B.V. All rights reserved.

Septicaemia and meningitis caused by infection of New Zealand sea lion pups with a hypermucoviscous strain of Klebsiella pneumoniae.

Science.gov (United States)

Roe, W D; Rogers, L; Pinpimai, K; Dittmer, K; Marshall, J; Chilvers, B L

2015-04-17

This study describes a syndrome of neonatal septicemia and meningitis in New Zealand sea lions, caused by a strain of Klebsiella pneumoniae that is phenotypically similar to strains causing environmentally-acquired septicemia and neuro-invasive disease in humans. Between late 2006 and early 2010, 123 pups from the Enderby Island breeding colony died of K. pneumoniae infection, with lesions including fibrinous to fibrinosuppurative meningitis, subdural hemorrhage, septic arthritis, herniation and hemorrhage of the cerebellar vermis, lymphadenitis and cellulitis. This infection was responsible for 58% of observed pup mortality over this time period, with most deaths occurring in the latter part of the breeding season (mid February onwards). The results of this study suggest that the pattern of this disease has changed since it was first described in 2002, when most deaths occurred early in the season (early to mid-January), and that it is an important and consistent cause of pup mortality in this population. In addition, a similar disease syndrome and bacterial strain was diagnosed in a single pup in a fragile recolonizing New Zealand sea lion population on mainland New Zealand, and the potential effect on this population is unknown but could have a negative impact on recolonisation at this site. Copyright © 2015 Elsevier B.V. All rights reserved.

Virulence potential of Escherichia coli strains causing asymptomatic bacteriuria during pregnancy.

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Lavigne, Jean-Philippe; Boutet-Dubois, Adeline; Laouini, Dorsaf; Combescure, Christophe; Bouziges, Nicole; Marès, Pierre; Sotto, Albert

2011-11-01

We compared the virulence properties of a collection of asymptomatic bacteriuria (ABU) Escherichia coli strains to urinary tract infection (UTI) strains isolated from pregnant women in a university hospital over 1 year. The in vitro and in vivo studies suggest that ABU strains presented a virulence behavior similar to that of strains isolated from cases of cystitis.

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

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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.

An Emerging Tick-Borne Disease of Humans Is Caused by a Subset of Strains with Conserved Genome Structure

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Barbet, Anthony F.; Al-Khedery, Basima; Stuen, Snorre; Granquist, Erik G.; Felsheim, Roderick F.; Munderloh, Ulrike G.

2013-01-01

The prevalence of tick-borne diseases is increasing worldwide. One such emerging disease is human anaplasmosis. The causative organism, Anaplasma phagocytophilum, is known to infect multiple animal species and cause human fatalities in the U.S., Europe and Asia. Although long known to infect ruminants, it is unclear why there are increasing numbers of human infections. We analyzed the genome sequences of strains infecting humans, animals and ticks from diverse geographic locations. Despite extensive variability amongst these strains, those infecting humans had conserved genome structure including the pfam01617 superfamily that encodes the major, neutralization-sensitive, surface antigen. These data provide potential targets to identify human-infective strains and have significance for understanding the selective pressures that lead to emergence of disease in new species. PMID:25437207

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.

Evidence for Increased Aggressiveness in a Recent Widespread Strain of Puccinia striiformis f. sp. tritici Causing Stripe Rust of Wheat

DEFF Research Database (Denmark)

Milus, Eugene A; Kristensen, Kristian; Hovmøller, Mogens S

2009-01-01

Stripe rust (yellow rust) of wheat, caused by Puccinia striiformis f. sp. tritici, has become more severe in eastern United States, Australia, and elsewhere since 2000. Recent research has shown that this coincided with a global spread of two closely related strains that were similar based on vir...... that wheat rust fungi can adapt to warmer temperatures and cause severe disease in previously unfavorable environments......Stripe rust (yellow rust) of wheat, caused by Puccinia striiformis f. sp. tritici, has become more severe in eastern United States, Australia, and elsewhere since 2000. Recent research has shown that this coincided with a global spread of two closely related strains that were similar based...... regimes for latent period, lesion length, lesion width, lesion area, and spore production on adult plants of a susceptible wheat cultivar with no known genes for resistance to stripe rust. "New" isolates (since 2000) were significantly more aggressive than "old" isolates (before 2000) for all variables...

Genomic Characterization of Urethritis-Associated Neisseria meningitidis Shows that a Wide Range of N. meningitidis Strains Can Cause Urethritis.

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Ma, Kevin C; Unemo, Magnus; Jeverica, Samo; Kirkcaldy, Robert D; Takahashi, Hideyuki; Ohnishi, Makoto; Grad, Yonatan H

2017-12-01

Neisseria meningitidis , typically a resident of the oro- or nasopharynx and the causative agent of meningococcal meningitis and meningococcemia, is capable of invading and colonizing the urogenital tract. This can result in urethritis, akin to the syndrome caused by its sister species, N. gonorrhoeae , the etiologic agent of gonorrhea. Recently, meningococcal strains associated with outbreaks of urethritis were reported to share genetic characteristics with the gonococcus, raising the question of the extent to which these strains contain features that promote adaptation to the genitourinary niche, making them gonococcus-like and distinguishing them from other N. meningitidis strains. Here, we analyzed the genomes of 39 diverse N. meningitidis isolates associated with urethritis, collected independently over a decade and across three continents. In particular, we characterized the diversity of the nitrite reductase gene ( aniA ), the factor H-binding protein gene ( fHbp ), and the capsule biosynthetic locus, all of which are loci previously suggested to be associated with urogenital colonization. We observed notable diversity, including frameshift variants, in aniA and fHbp and the presence of intact, disrupted, and absent capsule biosynthetic genes, indicating that urogenital colonization and urethritis caused by N. meningitidis are possible across a range of meningococcal genotypes. Previously identified allelic patterns in urethritis-associated N. meningitidis strains may reflect genetic diversity in the underlying meningococcal population rather than novel adaptation to the urogenital tract. Copyright © 2017 American Society for Microbiology.

Comparative analysis of virulence traits between a Legionella feeleii strain implicated in Pontiac fever and a strain that caused Legionnaires' disease.

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Wang, Changle; Saito, Mitsumasa; Tanaka, Tamami; Amako, Kazunobu; Yoshida, Shin-ichi

2015-12-01

Legionella strains of the same species and serogroup are known to cause Legionnaires' disease (a potentially fatal atypical pneumonia) or Pontiac fever (a mild, flu-like disease), but the bacterial factors that define these dramatic differences in pathology have not been elucidated. To gain a better understanding of these factors, we compared the characteristics of Legionella feeleii strains that were isolated from either a sample of freshwater implicated in an outbreak of Pontiac fever (ATCC 35072, serogroup 1, LfPF), or a patient with Legionnaires' disease (ATCC 38549, serogroup 2, LfLD). Growth of LfPF and LfLD in BYE broth was slower than the positive control, Legionella pneumophila strain JR32. However, LfLD grew faster than LfPF at 42 °C. After in vitro infection to J774 murine or U937 human macrophage cell lines and A549 human lung epithelial cell line, LfLD showed a higher cell infection rate, stronger internalization by host cells, and greater cytotoxicity than that of LfPF. Large amounts of IL-6 and IL-8 were secreted by human host cells after infection with LfLD, but not with LfPF. LfLD possessed mono-polar flagellum while LfPF was unflagellated. When LfLD was cultured at 25, 30 and 37 °C, the bacteria had higher motility rate at lower temperatures. Based on our results, this is the first study that showed distinct characteristics between LfPF and LfLD, which may give important leads in elucidating differences in their virulence. Copyright © 2015 Elsevier Ltd. All rights reserved.

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)

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)

A study on plastic strain accumulation caused by traveling of temperature distribution synchronizing with temperature rise

International Nuclear Information System (INIS)

Okajima, Satoshi

2016-01-01

The prevention of excessive deformation by thermal ratcheting is important in the design of high-temperature components of fast breeder reactors (FBR). This includes evaluation methods for a new type of thermal ratcheting caused by an axial traveling of temperature distribution, which corresponds to moving-up of liquid sodium surface in startup phase. Long range traveling of the axial temperature distribution brings flat plastic deformation profile in wide range. Therefore, at the center of this range, residual stress that brings shakedown behavior does not accumulate. As a result, repeating of this temperature traveling brings continuous accumulation of the plastic strain, even if there is no primary stress. In contrast, in the case with short range traveling, residual stress is caused by constraint against elastic part, and finally it results in shakedown. Because of this mechanism, we supposed that limit for the shakedown behavior depends on distance from the elastic part (i.e. half length of region with plastic deformation). In this paper, we examined characteristics of the accumulation of the plastic strain caused by realistic heat transients, namely, traveling of temperature distribution synchronizing with temperature rise. This examination was based on finite element analyses using elastic-perfectly plastic material. As a result, we confirmed that the shakedown limit depends not on the traveling range of the temperature distribution but the plastic deformation range, which was predicted by the elastic analysis. In the actual application, we can control the plastic deformation range by changing rate of the moving-up of liquid sodium surface. (author)

Neither hippurate-negative Brachyspira pilosicoli nor Brachyspira pilosicoli type strain caused diarrhoea in early-weaned pigs by experimental infection

DEFF Research Database (Denmark)

Fossi, M.; Ahlsten, K.; Pohjanvirta, T.

2005-01-01

Fossi M, Ahlsten K, Pohjanvirta T, Anttila M, Kokkonen T, Jensen TK, Boye M, Sukura A, Pelkola K, Pelkonen S: Neither hippurate-negative Brachyspira pilosicoli nor Brachyspira pilosicoli type strain caused diarrhoea in early-weaned pigs by experimental infection. Acta vet. scand. 2005, 46, 257...

Evaluation of indigenous bacterial strains for biocontrol of the frogeye leaf spot of soya bean caused by Cercospora sojina.

Science.gov (United States)

Simonetti, E; Carmona, M A; Scandiani, M M; García, A F; Luque, A G; Correa, O S; Balestrasse, K B

2012-08-01

Assessment of biological control of Cercospora sojina, causal agent of frogeye leaf spot (FLS) of soya bean, using three indigenous bacterial strains, BNM297 (Pseudomonas fluorescens), BNM340 and BNM122 (Bacillus amyloliquefaciens). From cultures of each bacterial strain, cell suspensions and cell-free supernatants were obtained and assayed to determine their antifungal activity against C. sojina. Both mycelial growth and spore germination in vitro were more strongly inhibited by bacterial cell suspensions than by cell-free supernatants. The Bacillus strains BNM122 and BNM340 inhibited the fungal growth to a similar degree (I ≈ 52-53%), while cells from P. fluorescens BNM297 caused a lesser reduction (I ≈ 32-34%) in the fungus colony diameter. The foliar application of the two Bacillus strains on soya bean seedlings, under greenhouse conditions, significantly reduced the disease severity with respect to control soya bean seedlings and those sprayed with BNM297. This last bacterial strain was not effective in controlling FLS in vivo. Our data demonstrate that the application of antagonistic bacteria may be a promising and environmentally friendly alternative to control the FLS of soya bean.   To our knowledge, this is the first report of biological control of C. sojina by using native Bacillus strains. © 2012 The Authors. Letters in Applied Microbiology © 2012 The Society for Applied Microbiology.

Clostridium difficile outbreak caused by NAP1/BI/027 strain and non-027 strains in a Mexican hospital

Directory of Open Access Journals (Sweden)

Rayo Morfin-Otero

2016-01-01

Conclusions: C. difficile NAP1/BI/027 strain and non-027 strains are established pathogens in our hospital. Accordingly, surveillance of C. difficile infections is now part of our nosocomial prevention program.

Magnetic resonance and diffusion tensor imaging analyses indicate heterogeneous strains along human medial gastrocnemius fascicles caused by submaximal plantar-flexion activity.

Science.gov (United States)

Karakuzu, Agah; Pamuk, Uluç; Ozturk, Cengizhan; Acar, Burak; Yucesoy, Can A

2017-05-24

Sarcomere length changes are central to force production and excursion of skeletal muscle. Previous modeling indicates non-uniformity of that if mechanical interaction of muscle with its surrounding muscular and connective tissues is taken into account. Hence, quantifying length changes along the fascicles of activated human muscle in vivo is crucial, but this is lacking due to technical complexities. Combining magnetic resonance imaging deformation analyses and diffusion tensor imaging tractography, the aim was to test the hypothesis that submaximal plantar flexion activity at 15% MVC causes heterogeneous length changes along the fascicles of human medial gastrocnemius (GM) muscle. A general fascicle strain distribution pattern shown for all subjects indicates that proximal track segments are shortened, whereas distal ones are lengthened (e.g., by 13% and 29%, respectively). Mean fiber direction strains of different tracts also shows heterogeneity (for up to 57.5% of the fascicles). Inter-subject variability of amplitude and distribution of fascicle strains is notable. These findings confirm the hypothesis and are solid indicators for the functionally dependent mechanics of human muscle, in vivo. Heterogeneity of fascicle strains can be explained by epimuscular myofascial force transmission. To the best of our knowledge, this is the first study, which quantified local deformations along human skeletal muscle fascicles caused by sustained submaximal activation. The present approach and indicated fascicle strain heterogeneity has numerous implications for muscle function in health and disease to estimate the muscle's contribution to the joint moment and excursion and to evaluate mechanisms of muscle injury and several treatment techniques. Copyright © 2017 Elsevier Ltd. All rights reserved.

Outbreak of candidemia caused by fluconazole resistant Candida parapsilosis strains in an intensive care unit.

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Pinhati, Henrique Marconi Sampaio; Casulari, Luiz Augusto; Souza, Ana Carolina Remondi; Siqueira, Ricardo Andreotti; Damasceno, Camila Maria Gomes; Colombo, Arnaldo Lopes

2016-08-20

Candidemia is an increasing problem in tertiary care hospitals worldwide. Here, we report the first outbreak of candidemia caused by fluconazole-resistant C. parapsilosis (FRCP) strains in Brazil. This was a cross-sectional study of clinical and microbiological data of all candidemic episodes diagnosed from July 2011 to February 2012 in a 200-bed tertiary care hospital. Initial yeast identification and susceptibility testing were performed using the VITEK 2 - System. Isolates of Candida spp. resistant to fluconazole were sent to a reference laboratory (LEMI-UNIFESP) for further molecular identification and confirmation of resistance by CLSI microdilution test. A multivariate analysis was conducted to identify factors associated with FRCP infection. We identified a total of 40 critically ill patients with candidemia (15 women) with a median age of 70 years. The incidence of candidemia was 6 cases/1,000 patients admissions, including 28 cases (70 %) of infection with C. parapsilosis, 21 of which (75 %) were resistant to fluconazole. In only 19 % of FRCP candidemia cases had fluconazole been used previously. The results of our study indicated that diabetes is a risk factor for FRCP candidemia (p = 0.002). Overall, mortality from candidemia was 45 %, and mortality from episodes of FRCP infections was 42.9 %. The clustering of incident cases in the ICU and molecular typing of strains suggest horizontal transmission of FRCP. Accurate vigilant monitoring for new nosocomial strains of FRCP is required.

First two cases of severe multifocal infections caused by Klebsiella pneumoniae in Switzerland: characterization of an atypical non-K1/K2-serotype strain causing liver abscess and endocarditis.

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Babouee Flury, Baharak; Donà, Valentina; Buetti, Niccolò; Furrer, Hansjakob; Endimiani, Andrea

2017-09-01

We describe the first two multifocal invasive infections due to Klebsiella pneumoniae recently observed in Switzerland. Phenotypic (MIC assays and string test) and molecular analyses (PCR/Sequencing for bla, virulence factor genes and whole genome sequencing for one strain) were performed to characterize the causative K. pneumoniae isolates. Both K. pneumoniae isolates (Kp1 and Kp2) were pan-susceptible to antibiotics and produced narrow-spectrum SHV β-lactamases. However, only Kp1 was string test positive. Kp1 was of ST380 and caused liver abscess as well as pneumonia and orbital phlegmon in an Eritrean patient. It belonged to the hypervirulent capsular serotype K2 and harboured the classic virulence-associated rmpA and aerobactin genes, fulfilling both the clinical and microbiological definitions for an invasive K. pneumoniae syndrome. Kp2 was of ST1043 and caused both liver abscess and endocarditis in a Swiss patient. Moreover, it did not possess the classic virulence-associated genes. Whole genome sequencing identified less well-known virulence factors in Kp2 that might have contributed to its virulence. Among these there were genes important for intestinal colonization and/or invasion, such as genes involved in adhesion (e.g., fimABCD and mrkABCD), regulation of capsule polysaccharide biosynthesis (e.g., evgS-evgA), as well as iron uptake (iroN), energy conversion, and metabolism. This report confirms the continuous dissemination of hypervirulent K. pneumoniae strains among patients of non-Asian descent in Europe. Moreover, it highlights the genetic background of an atypical hypervirulent K. pneumoniae causing a severe invasive infection despite not possessing the classical virulence characteristics of hypermucoviscous strains. Copyright © 2017 International Society for Chemotherapy of Infection and Cancer. Published by Elsevier Ltd. All rights reserved.

Haemophilus ducreyi Cutaneous Ulcer Strains Are Nearly Identical to Class I Genital Ulcer Strains.

Directory of Open Access Journals (Sweden)

Dharanesh Gangaiah

Full Text Available Although cutaneous ulcers (CU in the tropics is frequently attributed to Treponema pallidum subspecies pertenue, the causative agent of yaws, Haemophilus ducreyi has emerged as a major cause of CU in yaws-endemic regions of the South Pacific islands and Africa. H. ducreyi is generally susceptible to macrolides, but CU strains persist after mass drug administration of azithromycin for yaws or trachoma. H. ducreyi also causes genital ulcers (GU and was thought to be exclusively transmitted by microabrasions that occur during sex. In human volunteers, the GU strain 35000HP does not infect intact skin; wounds are required to initiate infection. These data led to several questions: Are CU strains a new variant of H. ducreyi or did they evolve from GU strains? Do CU strains contain additional genes that could allow them to infect intact skin? Are CU strains susceptible to azithromycin?To address these questions, we performed whole-genome sequencing and antibiotic susceptibility testing of 5 CU strains obtained from Samoa and Vanuatu and 9 archived class I and class II GU strains. Except for single nucleotide polymorphisms, the CU strains were genetically almost identical to the class I strain 35000HP and had no additional genetic content. Phylogenetic analysis showed that class I and class II strains formed two separate clusters and CU strains evolved from class I strains. Class I strains diverged from class II strains ~1.95 million years ago (mya and CU strains diverged from the class I strain 35000HP ~0.18 mya. CU and GU strains evolved under similar selection pressures. Like 35000HP, the CU strains were highly susceptible to antibiotics, including azithromycin.These data suggest that CU strains are derivatives of class I strains that were not recognized until recently. These findings require confirmation by analysis of CU strains from other regions.

Microbial dynamics during shelf-life of industrial Ricotta cheese and identification of a Bacillus strain as a cause of a pink discolouration.

Science.gov (United States)

Sattin, E; Andreani, N A; Carraro, L; Fasolato, L; Balzan, S; Novelli, E; Squartini, A; Telatin, A; Simionati, B; Cardazzo, B

2016-08-01

Dairy products are perishable and have to be preserved from spoilage during the food chain to achieve the desired shelf-life. Ricotta is a typical Italian soft dairy food produced by heat coagulation of whey proteins and is considered to be a light and healthy product. The shelf-life of Ricotta could be extended, as required by the international food trade market; however, heat resistant microflora causes spoilage and poses issues regarding the safety of the product. Next-generation sequencing (NGS) applied to the Ricotta samples defined the composition of the microbial community in-depth during the shelf-life. The analysis demonstrated the predominance of spore-forming bacteria throughout the shelf-life, mostly belonging to Bacillus, Paenibacillus and Clostridium genera. A strain involved in spoilage and causing a pink discolouration of Ricotta was isolated and characterised as Bacillus mycoides/weihenstephanensis. This is the first report of a food discolouration caused by a toxigenic strain belonging to the Bacillus cereus group that resulted the predominant strain in the community of the defective ricotta. These results suggest that the processing of raw materials to eliminate spores and residual microflora could be essential for improving the quality and the safety of the product and to extend the shelf-life of industrial Ricotta. Copyright © 2015 Elsevier Ltd. All rights reserved.

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.

Functional genomic characterization of virulence factors from necrotizing fasciitis-causing strains of Aeromonas hydrophila.

Science.gov (United States)

Grim, Christopher J; Kozlova, Elena V; Ponnusamy, Duraisamy; Fitts, Eric C; Sha, Jian; Kirtley, Michelle L; van Lier, Christina J; Tiner, Bethany L; Erova, Tatiana E; Joseph, Sandeep J; Read, Timothy D; Shak, Joshua R; Joseph, Sam W; Singletary, Ed; Felland, Tracy; Baze, Wallace B; Horneman, Amy J; Chopra, Ashok K

2014-07-01

The genomes of 10 Aeromonas isolates identified and designated Aeromonas hydrophila WI, Riv3, and NF1 to NF4; A. dhakensis SSU; A. jandaei Riv2; and A. caviae NM22 and NM33 were sequenced and annotated. Isolates NF1 to NF4 were from a patient with necrotizing fasciitis (NF). Two environmental isolates (Riv2 and -3) were from the river water from which the NF patient acquired the infection. While isolates NF2 to NF4 were clonal, NF1 was genetically distinct. Outside the conserved core genomes of these 10 isolates, several unique genomic features were identified. The most virulent strains possessed one of the following four virulence factors or a combination of them: cytotoxic enterotoxin, exotoxin A, and type 3 and 6 secretion system effectors AexU and Hcp. In a septicemic-mouse model, SSU, NF1, and Riv2 were the most virulent, while NF2 was moderately virulent. These data correlated with high motility and biofilm formation by the former three isolates. Conversely, in a mouse model of intramuscular infection, NF2 was much more virulent than NF1. Isolates NF2, SSU, and Riv2 disseminated in high numbers from the muscular tissue to the visceral organs of mice, while NF1 reached the liver and spleen in relatively lower numbers on the basis of colony counting and tracking of bioluminescent strains in real time by in vivo imaging. Histopathologically, degeneration of myofibers with significant infiltration of polymorphonuclear cells due to the highly virulent strains was noted. Functional genomic analysis provided data that allowed us to correlate the highly infectious nature of Aeromonas pathotypes belonging to several different species with virulence signatures and their potential ability to cause NF. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

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

Directory of Open Access Journals (Sweden)

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

First report of two rapid-onset fatal infections caused by a newly emerging hypervirulent K. Pneumonia ST86 strain of serotype K2 in China.

Science.gov (United States)

Zhang, Yibo; Sun, Jingyong; Mi, Chenrong; Li, Wenhui; Zhao, Shengyuan; Wang, Qun; Shi, Dake; Liu, Luo; Ding, Bingyu; Chang, Yung-Fu; Guo, Hongxiong; Guo, XiaoKui; Li, Qingtian; Zhu, Yongzhang

2015-01-01

Here, we present the first report of one suspected dead case and two confirmed rapid-onset fatal infections caused by a newly emerging hypervirulent Klebsiella pneumoniae ST86 strain of serotype K2. The three cases occurred in a surgery ward during 2013 in Shanghai, China. A combination of multilocus sequence typing, pulsed-field gel electrophoresis, phenotypic and PCR tests for detecting virulence factors (VFs) was used to identify the isolates as K2 ST86 strains with common VFs, including Aerobactin and rmpA. Furthermore, the two K2 ST86 strains additionally harbored a distinct VF kfu (responsible for iron uptake system), which commonly existed in invasive K1 strains only. Thus, the unusual presence of both K1 and K2 VFs in the lethal ST86 strain might further enhance its hypervirulence and cause rapid onset of a life-threatening infection. Nevertheless, despite the administration of a combined antibiotic treatment, these three patients all died within 24 h of acute onset, thereby highlighting that the importance of early diagnosis to determine whether the ST86 strains harbor key K2 VF and unusual K1 kfu and whether patients should receive a timely and targeted antibiotic therapy to prevent ST86 induced fatal pneumonia. Finally, even though these patients are clinically improved, keeping on with oral antibiotic treatment for additional 2-3 weeks will be also vital for successfully preventing hvKP reinfection or relapse.

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...

Infection of inbred rat strains with Rift Valley fever virus: development of a congenic resistant strain and observations on age-dependence of resistance.

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Anderson, G W; Rosebrock, J A; Johnson, A J; Jennings, G B; Peters, C J

1991-05-01

A congenic rat strain (WF.LEW) was derived from the susceptible Wistar-Furth (WF) (background strain) and the resistant LEW (donor strain) inbred strains and was used to evaluate the phenotypic expression of a dominant Mendelian gene that confers resistance to fatal hepatic disease caused by the ZH501 strain of Rift Valley fever virus (RVFV). Resistance to hepatic disease developed gradually with age, with full expression at approximately 10 weeks in the WF.LEW and LEW rat strains. The ZH501 strain caused fatal hepatitis in WF rats regardless of age. However, resistance to the SA75 RVFV strain (relatively non-pathogenic for adult rats), was age- and dose-dependent in both WF and LEW rats. The resistance gene transferred to the newly derived WF.LEW congenic rat strain appears to amplify age-dependent resistance of adult rats, resulting in protection against fatal hepatic disease caused by the virulent ZH501 strain. The congenic rat strain will be a valuable asset in elucidating the mechanism of resistance to Rift Valley fever virus governed by the dominant Mendelian gene.

Large-scale lagovirus disease outbreaks in European brown hares (Lepus europaeus) in France caused by RHDV2 strains spatially shared with rabbits (Oryctolagus cuniculus)

OpenAIRE

Le Gall-Reculé, Ghislaine; Lemaitre, Evelyne; Bertagnoli, Stéphane; Hubert, Céline; Top, Sokunthea; Decors, Anouk; Marchandeau, Stéphane; Guitton, Jean-Sébastien

2017-01-01

Rabbit haemorrhagic disease virus (RHDV) is a lagovirus that causes rabbit haemorrhagic disease (RHD) in European rabbits (Oryctolagus cuniculus). In 2010, a new genotype called RHDV2 emerged in France. It exhibits a larger host range than classical RHDV strains by sporadically infecting different hare species, including the European hare (Lepus europaeus). Phylogenetic analyses revealed that closely related RHDV2 strains circulate locally in both hares and rabbits, and therefore that RHDV2 s...

Multiple-locus variant-repeat assay (MLVA) is a useful tool for molecular epidemiologic analysis of Streptococcus agalactiae strains causing bovine mastitis.

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Radtke, Andreas; Bruheim, Torkjel; Afset, Jan Egil; Bergh, Kåre

2012-06-15

Group B streptococci (GBS) were considered a major cause of mastitis in cattle until preventive measures succeeded in controlling the disease in the 1970s and 1980s. During the last 5-6 years an increasing number of cases have been observed in some Scandinavian countries. A total of 187 GBS isolates from mastitis cases were collected from 119 animals in 34 Norwegian farms in the period from April 2007 to November 2010. 133 (71%) of the isolates were from farms with automated milking systems. The strains underwent typing of capsular polysaccharides (CPS) and surface proteins, and were analyzed by multi-locus variable repeat assay (MLVA) to investigate the epidemiological relationship of strains within and between farms. The GBS strains were differentiated into 12 types by CPS and surface protein analysis, with CPS types V (54%) and IV (34%) predominating. MLVA was superior to CPS and protein typing for strain differentiation, resolving the 187 strains into 37 types. In 29 of 34 farms all GBS strains had identical MLVA profiles specific for each farm. However, in one farm represented with 48 isolates, four MLVA variants with differences in one repeat locus were observed during the almost 3-year long collection period. Similar variations were observed at four other farms. This might reflect the stability of repeat loci under in vivo conditions. Farms with automated milking systems were overrepresented in this material. In conclusion, the five-loci MLVA allowed rapid high-resolution genotyping of the bovine GBS strains within and between farms. Copyright © 2012 Elsevier B.V. All rights reserved.

A wide extent of inter-strain diversity in virulent and vaccine strains of alphaherpesviruses.

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Szpara, Moriah L; Tafuri, Yolanda R; Parsons, Lance; Shamim, S Rafi; Verstrepen, Kevin J; Legendre, Matthieu; Enquist, L W

2011-10-01

Alphaherpesviruses are widespread in the human population, and include herpes simplex virus 1 (HSV-1) and 2, and varicella zoster virus (VZV). These viral pathogens cause epithelial lesions, and then infect the nervous system to cause lifelong latency, reactivation, and spread. A related veterinary herpesvirus, pseudorabies (PRV), causes similar disease in livestock that result in significant economic losses. Vaccines developed for VZV and PRV serve as useful models for the development of an HSV-1 vaccine. We present full genome sequence comparisons of the PRV vaccine strain Bartha, and two virulent PRV isolates, Kaplan and Becker. These genome sequences were determined by high-throughput sequencing and assembly, and present new insights into the attenuation of a mammalian alphaherpesvirus vaccine strain. We find many previously unknown coding differences between PRV Bartha and the virulent strains, including changes to the fusion proteins gH and gB, and over forty other viral proteins. Inter-strain variation in PRV protein sequences is much closer to levels previously observed for HSV-1 than for the highly stable VZV proteome. Almost 20% of the PRV genome contains tandem short sequence repeats (SSRs), a class of nucleic acids motifs whose length-variation has been associated with changes in DNA binding site efficiency, transcriptional regulation, and protein interactions. We find SSRs throughout the herpesvirus family, and provide the first global characterization of SSRs in viruses, both within and between strains. We find SSR length variation between different isolates of PRV and HSV-1, which may provide a new mechanism for phenotypic variation between strains. Finally, we detected a small number of polymorphic bases within each plaque-purified PRV strain, and we characterize the effect of passage and plaque-purification on these polymorphisms. These data add to growing evidence that even plaque-purified stocks of stable DNA viruses exhibit limited sequence

Mechanical properties of biaxially strained poly(L-lactide) tubes: Strain rate and temperature dependence

DEFF Research Database (Denmark)

Løvdal, Alexandra Liv Vest; Andreasen, Jens Wenzel; Mikkelsen, Lars Pilgaard

2017-01-01

Poly(l-lactide) (PLLA) is a bioabsorbable polymer with high stiffness and strength compared to the other commercially available bioabsorbable polymers. The properties of PLLA can be improved by straining, causing deformation-mediated molecular orientation. PLLA tubes were biaxially strained above...

Phenotypic and Genotypic Evaluation of Pseudomonas syringae pv. syringae Strains, Causing Citrus Blast in the West of Mazandaran and the East of Guilan

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S. Sameie-Shirkadeh

2017-01-01

Full Text Available Introduction: P. syringae pv. syringae (P.s.s, the causal agent of blast of citrus trees, is one of the most important plant pathogens in the world. P.s.s is unique among most P. syringae pathovars according to its ability to cause disease in over 180 species of plants in several unrelated genera. Traditionally, Strains of P.s.s are identified on the basis of biochemical and nutritional tests and symptom expression in host plants. Genomic fingerprinting methods based on the polymerase chain reaction (PCR have been applied for identification and classification of plant-associated bacteria to the subspecies level. The objectives of this study were the phenotypic and molecular evaluation of P.s. pv. syringae strains causing citrus blast in the West of Mazandaran and the East of Guilan, and study of genetic diversity of P.s.s isolates of citrus by using ERIC and REP-PCR markers. Materials and Methods: During 2011 to 2012, citrus infected tissues were sampled from different orchards in the West of Mazandaran and the East of Guilan. Bacterial phenotypes were studied based on standard physiological and biochemical tests. Gram reaction was determined by potassium hydroxide solubility test (KOH test. Strains were grown on King'B medium (KB and fluorescent pigment production was evaluated. Levan formation, oxidase reaction, potato soft rot, Arginine dihydrolase and induction of the hypersensitive reaction in tobacco leaves (LOPAT tests, were done as described by Schadd et al. The standard strains of P.s. pv. syringae form IVIA were used as reference strains in this study. Pathogenicity Test was done as described by Yessad et al. Citrus seedlings were maintained in a greenhouse at 20°C. In addition, a PCR-based method was used to confirm the genus and species of bacteria by using bacterial specific primer pair’s designed for a specific gene of syringomycin B. Genetic diversity among the strains, was studied by rep-PCR fingerprinting. Genomic

Potential of Zimbabwean commercial probiotic products and strains of Lactobacillus plantarum as prophylaxis and therapy against diarrhoea caused by Escherichia coli in children.

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Chingwaru, Walter; Vidmar, Jerneja

2017-01-01

To evaluate the potential of commercial fermented products sold in the country, and strains of Lactobacillus plantarum (L. plantarum) as prophylaxis and therapy against diarrhoea in children. The antimicrobial potential of cultures of lactobacilli enriched from 4 Zimbabwean commercial food/beverage products: Dairibord Lacto sour milk (DLSM), Probrand sour milk (PSM), Kefalos Vuka cheese (KVC) and Chibuku opaque beer (COB); and four strains of L. plantarum obtained from Balkan traditional cheeses against clinical strains of Escherichia coli (E. coli) was assayed using the well diffusion method. Three commercial paediatric antidiarrhoeal drug products: Biogaia (BG), Prolife (PL) and Probio Junior (PJ) and a mutant strain of E. coli [strain 11105 (ATCC) - a vitamin B-12 auxotroph and penicillin G acylase-producing strain] were used as controls. An agar diffusion assay and a competitive exclusion assay were carried out on Mueller Hinton agar. Crude cultures of putative lactobacillus strains obtained from Zimbabwean dairy products (Probrand sour milk, Kefalos Vuka vuka cheese and Chibuku opaque beer) had significantly higher antimicrobial activities against clinical strains of E. coli than strains of L. plantarum isolated from Balkan cheeses (CLP1, CLP2 or CLP3) and crude microbial cultures from commercial paediatric probiotic products (BG, PJ and PL) of a culture of Lactobacillus rhamnosus LGG (P sour milk, Kefalos Vuka vuka cheese and Chibuku opaque beer), and three strains of L. plantarum from Balkan cheeses (CLP1, CLP2 or CLP3) exhibited high antibacterial activities that can be harnessed to control paediatric diarrhoea that is caused by pathogenic strains of E. coli. Studies to characterise the probiotic potential of the live cultures in the products and the new strains of L. plantarum are underway. Copyright © 2017 Hainan Medical University. Production and hosting by Elsevier B.V. All rights reserved.

Biological assay of attenuated strain NADL-2 and virulent strain NADL-8 of porcine parvovirus.

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Mengeling, W L; Pejsak, Z; Paul, P S

1984-11-01

Attenuated strain NADL-2 and virulent strain NADL-8 of porcine parvovirus (PPV) were titrated in vivo and in vitro under similar conditions to provide a better understanding of some of the factors involved in virulence of PPV in causing maternal reproductive failure of swine. Both strains cause fetal death when they are injected directly into fetal fluids, but only strain NADL-8 does so when administered to pregnant swine. The strains were tested for their hemagglutinating activity (HA), median cell culture infective dose (CCID50), median fetal infective dose (FID50), and median fetal lethal dose (FLD50). The FID50 and FLD50 were determined by injecting virus directly into the amniotic fluid of fetuses in utero at 44 +/- 2 days of gestation and collecting the fetuses 15 +/- 1 days later. Both strains had an HA titer of 64, suggesting that there is a similar number of virions in stock preparations. However, other measurements differed markedly. The CCID50, FID50, and FLD50 were 10(5.5), 10(3.5), and 10(0.5), respectively, for strain NADL-2, and 10(4.5), 10(7.7), and 10(6.3), respectively, for strain NADL-8. Collectively, the values indicate that more than 10,000 times as much strain NADL-2 would need to reach the conceptus transplacentally to establish infection. These observations may help to explain the different consequences of oronasal exposure of pregnant swine to these strains of PPV.

The roles of sexual and asexual reproduction in the origin and dissemination of strains causing fungal infectious disease outbreaks.

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Ashu, Eta Ebasi; Xu, Jianping

2015-12-01

Sexual reproduction commonly refers to the reproductive process in which genomes from two sources are combined into a single cell through mating and then the zygote genomes are partitioned to progeny cells through meiosis. Reproduction in the absence of mating and meiosis is referred to as asexual or clonal reproduction. One major advantage of sexual reproduction is that it generates genetic variation among progeny which may allow for faster adaptation of the population to novel and/or stressful environments. However, adaptation to stressful or new environments can still occur through mutation, in the absence of sex. In this review, we analyzed the relative contributions of sexual and asexual reproduction in the origin and spread of strains causing fungal infectious diseases outbreaks. The necessity of sex and the ability of asexual fungi to initiate outbreaks are discussed. We propose a framework that relates the modes of reproduction to the origin and propagation of fungal disease outbreaks. Our analyses suggest that both sexual and asexual reproduction can play critical roles in the origin of outbreak strains and that the rapid spread of outbreak strains is often accomplished through asexual expansion. Copyright © 2015 Elsevier B.V. All rights reserved.

drug resistant strains of Salmonella enterica

African Journals Online (AJOL)

Conclusions: The aqueous extract of Thonningia sanguinea can provide an alternative therapy for the treatment of salmonellosis, mainly for typhoid fever caused by MDR strains of S. Typhi.The extract also inhibits S.Hadar a MDR emerging strain in Ivory Coast. Keywords: Thonningia sanguinea; Salmonella, MDR strains, ...

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.

A wide extent of inter-strain diversity in virulent and vaccine strains of alphaherpesviruses.

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Moriah L Szpara

2011-10-01

Full Text Available Alphaherpesviruses are widespread in the human population, and include herpes simplex virus 1 (HSV-1 and 2, and varicella zoster virus (VZV. These viral pathogens cause epithelial lesions, and then infect the nervous system to cause lifelong latency, reactivation, and spread. A related veterinary herpesvirus, pseudorabies (PRV, causes similar disease in livestock that result in significant economic losses. Vaccines developed for VZV and PRV serve as useful models for the development of an HSV-1 vaccine. We present full genome sequence comparisons of the PRV vaccine strain Bartha, and two virulent PRV isolates, Kaplan and Becker. These genome sequences were determined by high-throughput sequencing and assembly, and present new insights into the attenuation of a mammalian alphaherpesvirus vaccine strain. We find many previously unknown coding differences between PRV Bartha and the virulent strains, including changes to the fusion proteins gH and gB, and over forty other viral proteins. Inter-strain variation in PRV protein sequences is much closer to levels previously observed for HSV-1 than for the highly stable VZV proteome. Almost 20% of the PRV genome contains tandem short sequence repeats (SSRs, a class of nucleic acids motifs whose length-variation has been associated with changes in DNA binding site efficiency, transcriptional regulation, and protein interactions. We find SSRs throughout the herpesvirus family, and provide the first global characterization of SSRs in viruses, both within and between strains. We find SSR length variation between different isolates of PRV and HSV-1, which may provide a new mechanism for phenotypic variation between strains. Finally, we detected a small number of polymorphic bases within each plaque-purified PRV strain, and we characterize the effect of passage and plaque-purification on these polymorphisms. These data add to growing evidence that even plaque-purified stocks of stable DNA viruses exhibit

Strain-based HLA association analysis identified HLA-DRB1*09:01 associated with modern strain tuberculosis.

Science.gov (United States)

Toyo-Oka, L; Mahasirimongkol, S; Yanai, H; Mushiroda, T; Wattanapokayakit, S; Wichukchinda, N; Yamada, N; Smittipat, N; Juthayothin, T; Palittapongarnpim, P; Nedsuwan, S; Kantipong, P; Takahashi, A; Kubo, M; Sawanpanyalert, P; Tokunaga, K

2017-09-01

Tuberculosis (TB) occurs as a result of complex interactions between the host immune system and pathogen virulence factors. Human leukocyte antigen (HLA) class II molecules play an important role in the host immune system. However, no study has assessed the association between HLA class II genes and susceptibility to TB caused by specific strains. This study investigated the possible association of HLA class II genes with TB caused by modern and ancient Mycobacterium tuberculosis (MTB). The study included 682 patients with TB and 836 control subjects who were typed for HLA-DRB1 and HLA-DQB1 alleles. MTB strains were classified using a large sequence polymorphism typing method. Association analysis was performed using common HLA alleles and haplotypes in different MTB strains. HLA association analysis of patients infected with modern MTB strains showed significant association for HLA-DRB1*09:01 (odds ratio [OR] = 1.82; P-value = 9.88 × 10 -4 ) and HLA-DQB1*03:03 alleles (OR = 1.76; P-value = 1.31 × 10 -3 ) with susceptibility to TB. Haplotype analysis confirmed that these alleles were in strong linkage disequilibrium and did not exert an interactive effect. Thus, the results of this study showed an association between HLA class II genes and susceptibility to TB caused by modern MTB strains, suggesting the importance of strain-specific analysis to determine susceptibility genes associated with TB. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Asymptomatic bacteriuria Escherichia coli strains

DEFF Research Database (Denmark)

Hancock, Viktoria; Nielsen, E.M.; Klemm, Per

2006-01-01

Urinary tract infections (UTIs) affect millions of people each year. Escherichia coli is the most common organism associated with asymptomatic bacteriuria (ABU) in humans. Persons affected by ABU may carry a particular E. coli strain for extended periods of time without any symptoms. In contrast...... to uropathogenic E. coli (UPEC) that cause symptomatic UTI, very little is known about the mechanisms by which these strains colonize the urinary tract. Here, we have investigated the growth characteristics in human urine as well as adhesin repertoire of nine ABU strains; the ability of ABU strains to compete...

Francisella tularensis type A Strains Cause the Rapid Encystment of Acanthamoeba castellanii and Survive in Amoebal Cysts for Three Weeks post Infection

Energy Technology Data Exchange (ETDEWEB)

El-Etr, S H; Margolis, J; Monack, D; Robison, R; Cohen, M; Moore, E; Rasley, A

2009-07-28

Francisella tularensis, the causative agent of the zoonotic disease tularemia, has recently gained increased attention due to the emergence of tularemia in geographical areas where the disease has been previously unknown, and the organism's potential as a bioterrorism agent. Although F. tularensis has an extremely broad host range, the bacterial reservoir in nature has not been conclusively identified. In this study, the ability of virulent F. tularensis strains to survive and replicate in the amoeba Acanthamoeba castellanii was explored. We observe that A. castellanii trophozoites rapidly encyst in response to F. tularensis infection and that this rapid encystment phenotype (REP) is caused by factor(s) secreted by amoebae and/or F. tularensis into the co-culture media. Further, our results indicate that in contrast to LVS, virulent strains of F. tularensis can survive in A. castellanii cysts for at least 3 weeks post infection and that induction of rapid amoeba encystment is essential for survival. In addition, our data indicate that pathogenic F. tularensis strains block lysosomal fusion in A. castellanii. Taken together, these data suggest that the interactions between F. tularensis strains and amoeba may play a role in the environmental persistence of F. tularensis.

Sequencing of Escherichia coli that cause persistent and transient Mastitis

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The genomes of two strains of Escherichia coli that cause bovine mastitis were sequenced. These strains are known to be associated with persistent and transient mastitis: strain ECA-B causes a transient infection, and ECC-M leads to a persistent infection....

Next-Generation Sequence Analysis Reveals Transfer of Methicillin Resistance to a Methicillin-Susceptible Staphylococcus aureus Strain That Subsequently Caused a Methicillin-Resistant Staphylococcus aureus Outbreak: a Descriptive Study.

Science.gov (United States)

Weterings, Veronica; Bosch, Thijs; Witteveen, Sandra; Landman, Fabian; Schouls, Leo; Kluytmans, Jan

2017-09-01

Resistance to methicillin in Staphylococcus aureus is caused primarily by the mecA gene, which is carried on a mobile genetic element, the staphylococcal cassette chromosome mec (SCC mec ). Horizontal transfer of this element is supposed to be an important factor in the emergence of new clones of methicillin-resistant Staphylococcus aureus (MRSA) but has been rarely observed in real time. In 2012, an outbreak occurred involving a health care worker (HCW) and three patients, all carrying a fusidic acid-resistant MRSA strain. The husband of the HCW was screened for MRSA carriage, but only a methicillin-susceptible S. aureus (MSSA) strain, which was also resistant to fusidic acid, was detected. Multiple-locus variable-number tandem-repeat analysis (MLVA) typing showed that both the MSSA and MRSA isolates were MT4053-MC0005. This finding led to the hypothesis that the MSSA strain acquired the SCC mec and subsequently caused an outbreak. To support this hypothesis, next-generation sequencing of the MSSA and MRSA isolates was performed. This study showed that the MSSA isolate clustered closely with the outbreak isolates based on whole-genome multilocus sequence typing and single-nucleotide polymorphism (SNP) analysis, with a genetic distance of 17 genes and 44 SNPs, respectively. Remarkably, there were relatively large differences in the mobile genetic elements in strains within and between individuals. The limited genetic distance between the MSSA and MRSA isolates in combination with a clear epidemiologic link supports the hypothesis that the MSSA isolate acquired a SCC mec and that the resulting MRSA strain caused an outbreak. Copyright © 2017 American Society for Microbiology.

Urinary Tract Infection Caused by a Capnophilic Proteus mirabilis Strain

NARCIS (Netherlands)

Trapman, M.; Ingen, J. van; Keijman, J.; Swanink, C.M.A.

2015-01-01

From a urine sample from a patient with a urinary tract infection, a carbon dioxide-dependent Proteus mirabilis strain was isolated. It is important to perform urine cultures in 5% carbon dioxide and an anaerobic atmosphere if bacteria prominent in Gram stains do not grow on routine media in ambient

Comparative analyses of Legionella species identifies genetic features of strains causing Legionnaires' disease.

Science.gov (United States)

Gomez-Valero, Laura; Rusniok, Christophe; Rolando, Monica; Neou, Mario; Dervins-Ravault, Delphine; Demirtas, Jasmin; Rouy, Zoe; Moore, Robert J; Chen, Honglei; Petty, Nicola K; Jarraud, Sophie; Etienne, Jerome; Steinert, Michael; Heuner, Klaus; Gribaldo, Simonetta; Médigue, Claudine; Glöckner, Gernot; Hartland, Elizabeth L; Buchrieser, Carmen

2014-01-01

The genus Legionella comprises over 60 species. However, L. pneumophila and L. longbeachae alone cause over 95% of Legionnaires’ disease. To identify the genetic bases underlying the different capacities to cause disease we sequenced and compared the genomes of L. micdadei, L. hackeliae and L. fallonii (LLAP10), which are all rarely isolated from humans. We show that these Legionella species possess different virulence capacities in amoeba and macrophages, correlating with their occurrence in humans. Our comparative analysis of 11 Legionella genomes belonging to five species reveals highly heterogeneous genome content with over 60% representing species-specific genes; these comprise a complete prophage in L. micdadei, the first ever identified in a Legionella genome. Mobile elements are abundant in Legionella genomes; many encode type IV secretion systems for conjugative transfer, pointing to their importance for adaptation of the genus. The Dot/Icm secretion system is conserved, although the core set of substrates is small, as only 24 out of over 300 described Dot/Icm effector genes are present in all Legionella species. We also identified new eukaryotic motifs including thaumatin, synaptobrevin or clathrin/coatomer adaptine like domains. Legionella genomes are highly dynamic due to a large mobilome mainly comprising type IV secretion systems, while a minority of core substrates is shared among the diverse species. Eukaryotic like proteins and motifs remain a hallmark of the genus Legionella. Key factors such as proteins involved in oxygen binding, iron storage, host membrane transport and certain Dot/Icm substrates are specific features of disease-related strains.

Hydrogen production from microbial strains

Science.gov (United States)

Harwood, Caroline S; Rey, Federico E

2012-09-18

The present invention is directed to a method of screening microbe strains capable of generating hydrogen. This method involves inoculating one or more microbes in a sample containing cell culture medium to form an inoculated culture medium. The inoculated culture medium is then incubated under hydrogen producing conditions. Once incubating causes the inoculated culture medium to produce hydrogen, microbes in the culture medium are identified as candidate microbe strains capable of generating hydrogen. Methods of producing hydrogen using one or more of the microbial strains identified as well as the hydrogen producing strains themselves are also disclosed.

Oliveria decumbens essential oil: Chemical compositions and antimicrobial activity against the growth of some clinical and standard strains causing infection.

Science.gov (United States)

Alizadeh Behbahani, Behrooz; Tabatabaei Yazdi, Farideh; Vasiee, Alireza; Mortazavi, Seyed Ali

2018-01-01

Oliveria decumbens as a valuable medicinal plant is extensively used in traditional medicine. clinical and standard strains causing infection resistance to antimicrobial agents, is one of the important problems in medicine. The aim of this study was to investigate the antibacterial activities and phytochemical analysis of Oliveria decumbens essential oil on the growth of some clinical and standard strains causing infection (Pseudomonas aerogenes, Escherichia coli, Streptococcus pyogenes and Staphylococcus epidermidis). Oliveria decumbens essential oil composition was identified by gas chromatography/mass spectrometry. Phytochemical analysis (alkaloids, saponins, flavone and phenolic) essential oil of the Oliveria decumbens were appraised based on qualitative methods. Several methods (disk diffusion, minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC)) were used to appraise the antibacterial activity of the Oliveria decumbens essential oil. Thymol (28.45%) was the major compound of Oliveria decumbens essential oil. The total phenolics content (TPC) of the essential oil positively correlated with antioxidant activity (AA). The TPC and AA of Oliveria decumbens essential oil was equal to 92.45 ± 0.70 μg GAE/mg and 164.45 ± 1.20 μg/ml, respectively. The MIC of Oliveria decumbens essential oil ranged from 1 to 8 mg/ml depending on the type of bacteria (clinical and standard strains). The MBC of Oliveria decumbens essential oil varied from 1 mg/ml to 16 mg/ml. The smallest inhibition zone diameter (IZD) on different Oliveria decumbens essential oil concentrations on P. aeruginosa. Results indicate that Oliveria decumbens essential oil can prove to be an important source of AA and antibacterial and may be used for the treatment of infection diseases. Copyright © 2017 Elsevier Ltd. All rights reserved.

Case of Meningitis in a Neonate Caused by an Extended-Spectrum-Beta-Lactamase-Producing Strain of Hypervirulent Klebsiella pneumoniae

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Khalit S. Khaertynov

2017-08-01

Full Text Available Klebsiella pneumoniae is one of the most important infectious agents among neonates. This pathogen has a potential to develop an increased antimicrobial resistance and virulence. The classic non-virulent strain of K. pneumoniae, producing an extended-spectrum beta-lactamases (ESBL, is associated with nosocomial infection mainly in preterm neonates. Hypervirulent K. pneumoniae strains are associated with invasive infection among previously healthy ambulatory patients, and most of them exhibit antimicrobial susceptibility. During the last few years, several cases of diseases caused by hypervirulent K. pneumoniae producing ESBL have been registered in different geographical regions of the world. However, reports of such cases in neonates are rare. Here, we reported that this pathogen can cause pyogenic meningitis in full-term neonate with poor prognosis. A previously healthy, full-term, 12-day-old neonate was admitted to the infectious diseases hospital with suspected meningitis. The clinical symptoms included loss of appetite, irritability, fever, seizures, and a bulging anterior fontanelle. The analysis of the cerebrospinal fluid confirmed the diagnosis of meningitis. Blood and cerebrospinal fluid cultures were positive for K. pneumoniae, producing ESBL. K. pneumoniae isolates were resistant to aminopenicillins, 3rd generation cephalosporins but were sensitive to imipenem and meropenem. The “string test” was positive. The study of the virulence factors of K. pneumoniae by PCR revealed the presence of the rmpA gene. A combination of K. pneumoniae virulence and drug resistance complicated by cerebral oedema led to the death of the neonate. We concluded that both the risk of developing severe forms of infection and the outcome of the disease due to K. pneumonia are associated with the phenotypic features of the pathogen such as its antibiotic susceptibility and virulence factors. Emergence of the ESBL-producing strain of hypervirulent K

Fatal case of bacteremia caused by an atypical strain of Corynebacterium mucifaciens

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Vlademir Vicente Cantarelli

Full Text Available Corynebacterium species have often been considered normal skin flora or contaminants; however, in recent years they have been increasingly implicated in serious infections. Moreover, many new species have been discovered and old species renamed, especially after molecular biology techniques were introduced. Corynebacterium mucifaciens is mainly isolated from blood and from other normally-sterile body fluids; it forms slightly yellow, mucoid colonies on blood agar. We report a fatal case of bacteremia due to an atypical strain of C. mucifaciens. This strain had atypical colony morphology; analysis of the 16S rRNA gene was used to define the species.

Control of Pierce's Disease by Phage.

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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.

Strain concentration at structural discontinuities and its quantification by elastic follow-up parameter

International Nuclear Information System (INIS)

Kasahara, Naoto; Takasho, Hideki

1998-12-01

Elevated temperature structural design codes pay attention to strain concentration at structural discontinuities due to creep and plasticity, since it causes to enlarge creep-fatigue damage of material. One of the difficulties to predict strain concentration is its dependency on loading, constitutive equations, and relaxation time. This study investigated fundamental mechanism of strain concentration and its main factors. It was clarified that strain concentration was caused from strain redistribution between elastic and inelastic regions, which can be quantified by the elastic follow-up parameter. As a function of inelastic strain, the elastic follow-up parameter can describe variation of strain concentration during incremental loading and relaxation process, caused by transition of strain distribution from peak strain concentration to secondary stress redistribution. Structures have their own elastic follow-up characteristics as a function of inelastic strain, which is insensitive to constitutive equations. It means that application of inelastic analysis is not difficult to obtain elastic follow-up characteristics. (author)

Variation in the strain anisotropy of Zircaloy with temperature and strain

International Nuclear Information System (INIS)

Hindle, E.D.; Worswick, D.

1984-01-01

The strong crystallographic texture which is developed during the fabrication of zirconium-based alloys causes pronounced anisotropy in their mechanical properties, particularly deformation. The tendency for circular-section tension specimens with a high concentration of basal poles in one direction to become elliptical when deformed in tension has been used in this study to provide quantitative data on the effects of both strain and temperature on strain anisotropy. Tension tests were carried out over a temperature range of 293 to 1193 K on specimens machined from Zircaloy-2 plate. The strain anisotropy was found to increase markedly at temperatures over 923 K, reaching a maximum in the region of 1070 K. The strain anisotropy increased with increasing strain in this temperature region. The study was extended to Zircaloy-4 pressurized-water reactor fuel cladding by carrying out tube swelling tests and evaluating the axial deformation produced. Although scatter in the test results was higher than that exhibited in the tension tests, the general trend in the data was similar. The effects of the strain anisotropy observed are discussed in relation to the effects of temperature on the ductility of Zircaloy fuel cladding tubes during postulated largebreak loss-of-coolant accidents

Haemophilus ducreyi Cutaneous Ulcer Strains Diverged from Both Class I and Class II Genital Ulcer Strains: Implications for Epidemiological Studies.

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Dharanesh Gangaiah

2016-12-01

Full Text Available Haemophilus ducreyi has emerged as a major cause of cutaneous ulcers (CU in yaws-endemic regions of the tropics in the South Pacific, South East Asia and Africa. H. ducreyi was once thought only to cause the genital ulcer (GU disease chancroid; GU strains belong to 2 distinct classes, class I and class II. Using whole-genome sequencing of 4 CU strains from Samoa, 1 from Vanuatu and 1 from Papua New Guinea, we showed that CU strains diverged from the class I strain 35000HP and that one CU strain expressed β-lactamase. Recently, the Center for Disease Control and Prevention released the genomes of 11 additional CU strains from Vanuatu and Ghana; however, the evolutionary relationship of these CU strains to previously-characterized CU and GU strains is unknown.We performed phylogenetic analysis of 17 CU and 10 GU strains. Class I and class II GU strains formed two distinct clades. The class I strains formed two subclades, one containing 35000HP and HD183 and the other containing the remainder of the class I strains. Twelve of the CU strains formed a subclone under the class I 35000HP subclade, while 2 CU strains formed a subclone under the other class I subclade. Unexpectedly, 3 of the CU strains formed a subclone under the class II clade. Phylogenetic analysis of dsrA-hgbA-ncaA sequences yielded a tree similar to that of whole-genome phylogenetic tree.CU strains diverged from multiple lineages within both class I and class II GU strains. Multilocus sequence typing of dsrA-hgbA-ncaA could be reliably used for epidemiological investigation of CU and GU strains. As class II strains grow relatively poorly and are relatively more susceptible to vancomycin than class I strains, these findings have implications for methods to recover CU strains. Comparison of contemporary CU and GU isolates would help clarify the relationship between these entities.

Haemophilus ducreyi Cutaneous Ulcer Strains Diverged from Both Class I and Class II Genital Ulcer Strains: Implications for Epidemiological Studies.

Science.gov (United States)

Gangaiah, Dharanesh; Spinola, Stanley M

2016-12-01

Haemophilus ducreyi has emerged as a major cause of cutaneous ulcers (CU) in yaws-endemic regions of the tropics in the South Pacific, South East Asia and Africa. H. ducreyi was once thought only to cause the genital ulcer (GU) disease chancroid; GU strains belong to 2 distinct classes, class I and class II. Using whole-genome sequencing of 4 CU strains from Samoa, 1 from Vanuatu and 1 from Papua New Guinea, we showed that CU strains diverged from the class I strain 35000HP and that one CU strain expressed β-lactamase. Recently, the Center for Disease Control and Prevention released the genomes of 11 additional CU strains from Vanuatu and Ghana; however, the evolutionary relationship of these CU strains to previously-characterized CU and GU strains is unknown. We performed phylogenetic analysis of 17 CU and 10 GU strains. Class I and class II GU strains formed two distinct clades. The class I strains formed two subclades, one containing 35000HP and HD183 and the other containing the remainder of the class I strains. Twelve of the CU strains formed a subclone under the class I 35000HP subclade, while 2 CU strains formed a subclone under the other class I subclade. Unexpectedly, 3 of the CU strains formed a subclone under the class II clade. Phylogenetic analysis of dsrA-hgbA-ncaA sequences yielded a tree similar to that of whole-genome phylogenetic tree. CU strains diverged from multiple lineages within both class I and class II GU strains. Multilocus sequence typing of dsrA-hgbA-ncaA could be reliably used for epidemiological investigation of CU and GU strains. As class II strains grow relatively poorly and are relatively more susceptible to vancomycin than class I strains, these findings have implications for methods to recover CU strains. Comparison of contemporary CU and GU isolates would help clarify the relationship between these entities.

Large-scale lagovirus disease outbreaks in European brown hares (Lepus europaeus) in France caused by RHDV2 strains spatially shared with rabbits (Oryctolagus cuniculus).

Science.gov (United States)

Le Gall-Reculé, Ghislaine; Lemaitre, Evelyne; Bertagnoli, Stéphane; Hubert, Céline; Top, Sokunthea; Decors, Anouk; Marchandeau, Stéphane; Guitton, Jean-Sébastien

2017-10-28

Rabbit haemorrhagic disease virus (RHDV) is a lagovirus that causes rabbit haemorrhagic disease (RHD) in European rabbits (Oryctolagus cuniculus). In 2010, a new genotype called RHDV2 emerged in France. It exhibits a larger host range than classical RHDV strains by sporadically infecting different hare species, including the European hare (Lepus europaeus). Phylogenetic analyses revealed that closely related RHDV2 strains circulate locally in both hares and rabbits, and therefore that RHDV2 strains infecting hares do not belong to a lineage that has evolved only in this species. We showed that RHDV2 is widely distributed in France and that it was responsible for more than a third of cases of lagovirus disease in European hare populations in 2015. The oldest RHDV2 positive hare was sampled in November 2013 and we reported two hares co-infected by EBHSV and RHDV2. All together, our results raise important epidemiological and evolutionary issues. In particular, along with the potential emergence of recombinant EBHSV/RHDV2 strains in hares, the enlargement of the host range changes the host population structure of RHDV2 and may alter the impact of the virus on rabbit and hare populations.

The new strains Brucella inopinata BO1 and Brucella species 83-210 behave biologically like classic infectious Brucella species and cause death in murine models of infection.

Science.gov (United States)

Jiménez de Bagüés, María P; Iturralde, María; Arias, Maykel A; Pardo, Julián; Cloeckaert, Axel; Zygmunt, Michel S

2014-08-01

Recently, novel atypical Brucella strains isolated from humans and wild rodents have been reported. They are phenotypically close to Ochrobactrum species but belong to the genus Brucella, based on genetic relatedness, although genetic diversity is higher among the atypical Brucella strains than between the classic species. They were classified within or close to the novel species Brucella inopinata. However, with the exception of Brucella microti, the virulence of these novel strains has not been investigated in experimental models of infection. The type species B. inopinata strain BO1 (isolated from a human) and Brucella species strain 83-210 (isolated from a wild Australian rodent) were investigated. A classic infectious Brucella reference strain, B. suis 1330, was also used. BALB/c, C57BL/6, and CD1 mice models and C57BL/6 mouse bone-marrow-derived macrophages (BMDMs) were used as infection models. Strains BO1 and 83-210 behaved similarly to reference strain 1330 in all mouse infection models: there were similar growth curves in spleens and livers of mice and similar intracellular replication rates in BMDMs. However, unlike strain 1330, strains BO1 and 83-210 showed lethality in the 3 mouse models. The novel atypical Brucella strains of this study behave like classic intracellular Brucella pathogens. In addition, they cause death in murine models of infection, as previously published for B. microti, another recently described environmental and wildlife species. © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Co-infection of sweet orange with severe and mild strains of citrus tristeza virus is overwhelmingly dominated by the severe strain on both the transcriptional and biological levels

Science.gov (United States)

Citrus tristeza is one of the most destructive citrus diseases and is caused by the phloem-restricted Closterovirus, Citrus tristeza virus. Mild strain CTV-B2 does not cause obvious symptoms on indicators whereas severe strain CTV-B6 causes symptoms, including stem pitting, cupping, yellowing and s...

Brucella abortus strain 2308 Wisconsin genome: importance of the definition of reference strains

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Marcela Suárez-Esquivel

2016-09-01

Full Text Available Brucellosis is a bacterial infectious disease affecting a wide range of mammals and a neglected zoonosis caused by species of the genetically homogenous genus Brucella. As in most studies on bacterial diseases, research in brucellosis is carried out by using reference strains as canonical models to understand the mechanisms underlying host pathogen interactions. We performed whole genome sequencing (WGS analysis of the reference strain Brucella abortus 2308 routinely used in our laboratory, including manual curated annotation accessible as an editable version at www.wikipedia.Comparison of this genome with two publically available 2308 genomes showed significant differences, particularly indels related to insertional elements, suggesting variability related to the transposition of these elements within the same strain. Considering the outcome of high resolution genomic techniques in the bacteriology field, the conventional concept of strain definition needs to be revised.

The asymptomatic bacteriuria Escherichia coli strain 83972 outcompetes uropathogenic E. coli strains in human urine

DEFF Research Database (Denmark)

Hancock, Viktoria; Ulett, G.C.; Schembri, M.A.

2006-01-01

Escherichia coli is the most common organism associated with asymptomatic bacteriuria (ABU). In contrast to uropathogenic E. coli (UPEC), which causes symptomatic urinary tract infections (UTI), very little is known about the mechanisms by which these strains colonize the human urinary tract....... The prototype ABU E. coli strain 83972 was originally isolated from a girl who had carried it asymptomatically for 3 years. Deliberate colonization of UTI-susceptible individuals with E. coli 83972 has been used successfully as an alternative approach for the treatment of patients who are refractory...... to conventional therapy. Colonization with strain 83972 appears to prevent infection with UPEC strains in such patients despite the fact that this strain is unable to express the primary adhesins involved in UTI, viz. P and type 1 fimbriae. Here we investigated the growth characteristics of E. coli 83972 in human...

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

The prevalence of O serogroups of Escherichia coli strains causing acute urinary tract infection in children in Iran

Directory of Open Access Journals (Sweden)

Fatemeh Emamghorashi

2011-01-01

Full Text Available The aim of present study was to determine the prevalence of O serogroups of Escherichia coli (E. coli strains that cause community-acquired urinary tract infections (UTI in children. In this study, 96 children with UTI referred to two Jahrom University-affiliated Hospitals in Iran were enrolled, during the period from August 2005 to August 2006. Drug sensitivity was tested by disk diffusion method and serotyping done by slide agglutination method. A total of 96 E. coli strains were isolated from urine samples of the study children whose age ranged from one month to 14 years. Cystitis was diagnosed in 49.2% and pyelonephritis in 50.8% of the study patients. Maximum drug resistance was seen with ampicilin (80.2% and the least with imipenem (1.1%. The most common type of O antigen was O1 (12.2%. There was significant correlation between the presence of O antigens and sensitivity to nalidixic acid and gentamicin (P < 0.05. This is the first report of E. coli serotyping in children with UTI from the south of Iran and their relation to antibiotic resistance and clinical presentation. Further studies from other parts of Iran and on other serotypes are recommended.

Immunization by intrabronchial administration to 1-week-old foals of an unmarked double gene disruption strain of Rhodococcus equi strain 103+.

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Pei, Yanlong; Nicholson, Vivian; Woods, Katharine; Prescott, John F

2007-11-15

Rhodococcus equi causes fatal granulomatous pneumonia in foals and immunocompromised animals and humans. However, there is no effective vaccine against this infection. In this study, the chromosomal genes isocitrate lyase (icl) and cholesterol oxidase (choE) were chosen as targets for mutation and assessment of the double mutant as an intrabronchial vaccine in 1-week-old foals. Using a modification of a suicide plasmid previously developed in this laboratory, we developed a choE-icl unmarked deletion mutant of R. equi strain 103+. Five 1-week-old foals were infected intrabronchially with the mutant and challenged intrabronchially with the parent, virulent, strain 2 weeks later. Three of the foals were protected against pneumonia caused by the virulent strain, but the other two foals developed pneumonia caused by the mutant strain during the post-challenge period. Since infection of 3-week-old foals by an icl mutant in an earlier study had shown complete attenuation of the strain, we conclude that a proportion of foals in the 1st week or so of life are predisposed to developing R. equi pneumonia because of an inability to mount an effective immune response. This has been suspected previously but this is the first time that this has been demonstrated experimentally.

Draft genome sequence of Bacillus velezensis 2A-2B strain: a rhizospheric inhabitant of Sporobolus airoides (Torr.) Torr., with antifungal activity against root rot causing phytopathogens.

Science.gov (United States)

Martínez-Raudales, Inés; De La Cruz-Rodríguez, Yumiko; Alvarado-Gutiérrez, Alejandro; Vega-Arreguín, Julio; Fraire-Mayorga, Ahuitz; Alvarado-Rodríguez, Miguel; Balderas-Hernández, Victor; Fraire-Velázquez, Saúl

2017-01-01

A Bacillus velezensis strain from the rhizosphere of Sporobolus airoides (Torr.) Torr . , a grass in central-north México, was isolated during a biocontrol of phytopathogens scrutiny study. The 2A-2B strain exhibited at least 60% of growth inhibition of virulent isolates of phytopathogens causing root rot. These phytopathogens include Phytophthora capsici , Fusarium solani , Fusarium oxysporum and Rhizoctonia solani . Furthermore, the 2A-2B strain is an indolacetic acid producer, and a plant inducer of PR1, which is an induced systemic resistance related gene in chili pepper plantlets. Whole genome sequencing was performed to generate a draft genome assembly of 3.953 MB with 46.36% of GC content, and a N50 of 294,737. The genome contains 3713 protein coding genes and 89 RNA genes. Moreover, comparative genome analysis revealed that the 2A-2B strain had the greatest identity (98.4%) with Bacillus velezensis.

Genome analysis of Mycoplasma synoviae strain MS-H, the most common M. synoviae strain with a worldwide distribution.

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Zhu, Ling; Shahid, Muhammad A; Markham, John; Browning, Glenn F; Noormohammadi, Amir H; Marenda, Marc S

2018-02-02

The bacterial pathogen Mycoplasma synoviae can cause subclinical respiratory disease, synovitis, airsacculitis and reproductive tract disease in poultry and is a major cause of economic loss worldwide. The M. synoviae strain MS-H was developed by chemical mutagenesis of an Australian isolate and has been used as a live attenuated vaccine in many countries over the past two decades. As a result it may now be the most prevalent strain of M. synoviae globally. Differentiation of the MS-H vaccine from local field strains is important for epidemiological investigations and is often required for registration of the vaccine. The complete genomic sequence of the MS-H strain was determined using a combination of Illumina and Nanopore methods and compared to WVU-1853, the M. synoviae type strain isolated in the USA 30 years before the parent strain of MS-H, and MS53, a more recent isolate from Brazil. The vaccine strain genome had a slightly larger number of pseudogenes than the two other strains and contained a unique 55 kb chromosomal inversion partially affecting a putative genomic island. Variations in gene content were also noted, including a deoxyribose-phosphate aldolase (deoC) fragment and an ATP-dependent DNA helicase gene found only in MS-H. Some of these sequences may have been acquired horizontally from other avian mycoplasma species. MS-H was somewhat more similar to WVU-1853 than to MS53. The genome sequence of MS-H will enable identification of vaccine-specific genetic markers for use as diagnostic and epidemiological tools to better control M. synoviae.

Strain quantification in epitaxial thin films

International Nuclear Information System (INIS)

Cushley, M

2008-01-01

Strain arising in epitaxial thin films can be beneficial in some cases but devastating in others. By altering the lattice parameters, strain may give a thin film properties hitherto unseen in the bulk material. On the other hand, heavily strained systems are prone to develop lattice defects in order to relieve the strain, which can cause device failure or, at least, a decrease in functionality. Using convergent beam electron diffraction (CBED) and high-resolution transmission electron microscopy (HRTEM), it is possible to determine local strains within a material. By comparing the results from CBED and HRTEM experiments, it is possible to gain a complete view of a material, including the strain and any lattice defects present. As well as looking at how the two experimental techniques differ from each other, I will also look at how results from different image analysis algorithms compare. Strain in Si/SiGe samples and BST/SRO/MgO capacitor structures will be discussed.

Brucella abortus Strain 2308 Wisconsin Genome: Importance of the Definition of Reference Strains

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Suárez-Esquivel, Marcela; Ruiz-Villalobos, Nazareth; Castillo-Zeledón, Amanda; Jiménez-Rojas, César; Roop II, R. Martin; Comerci, Diego J.; Barquero-Calvo, Elías; Chacón-Díaz, Carlos; Caswell, Clayton C.; Baker, Kate S.; Chaves-Olarte, Esteban; Thomson, Nicholas R.; Moreno, Edgardo; Letesson, Jean J.; De Bolle, Xavier; Guzmán-Verri, Caterina

2016-01-01

Brucellosis is a bacterial infectious disease affecting a wide range of mammals and a neglected zoonosis caused by species of the genetically homogenous genus Brucella. As in most studies on bacterial diseases, research in brucellosis is carried out by using reference strains as canonical models to understand the mechanisms underlying host pathogen interactions. We performed whole genome sequencing analysis of the reference strain B. abortus 2308 routinely used in our laboratory, including manual curated annotation accessible as an editable version through a link at https://en.wikipedia.org/wiki/Brucella#Genomics. Comparison of this genome with two publically available 2308 genomes showed significant differences, particularly indels related to insertional elements, suggesting variability related to the transposition of these elements within the same strain. Considering the outcome of high resolution genomic techniques in the bacteriology field, the conventional concept of strain definition needs to be revised. PMID:27746773

Amplification and sequencing of varicella zoster virus (VZV) gene 4: point mutation in a VZV strain causing chickenpox during pregnancy

International Nuclear Information System (INIS)

Chow, V.T.K.; Lim, K.P.

1997-01-01

The varicella-zoster virus (VZV) causes chickenpox (varicella) as the primary disease and shingles (zoster) as a recurrent manifestation of infection, both being generality benign and self-limiting. While these infections may be severe in adults and even life-threatening in immunosuppressed individuals, they may be amenable to effective antiviral drugs or varicella-zoster immune globulin, provided the treatment is administered early. The prompt diagnosis of VZV infections may be accelerated by rapid, sensitive and specific molecular techniques such as amplification by polymerase chain reaction (PCR) compared with slower and more cumbersome tissue culture and serological procedures. Based on the VZV gene 4 which encodes a transcriptional activator, primers were designed for use in PCR to amplify a target fragment of 381 bp. Distinct diagnostic bands were observed by agarose gel electrophoresis of PCR products of VZV strains isolated from II varicella and 7 zoster patients in Singapore, as well as of the Japanese vaccine Oka strain. The detection sensitivity of this PCR assay was determined to be 1 pg of purified VZV DNA equivalent to about 7,000 viral DNA copies. No target bands were amplified from negative control templates from five related human herpes-viruses and from human DNA. The specificity of the PCR products was ensured by direct cycle DNA sequencing, which revealed complete identity of the 18 VZV isolates with the published European Dumas strain. The strong sequence conservation of the target fragment renders this PCR assay highly reliable for detecting the VZV sequence. Only one VZV strain isolated from a patient with varicella during pregnancy exhibited a Gaga to GAA point mutation at codon 46 of gene 4, culminating in the non-conservative substitution of Ser with Phe. The predicted secondary structure of the mutant polypeptide portrayed a radical alteration, which may influence its function in transcriptional activation. (authors)

Cell Vacuolation Caused by Vibrio cholerae Hemolysin

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Figueroa-Arredondo, Paula; Heuser, John E.; Akopyants, Natalia S.; Morisaki, J. Hiroshi; Giono-Cerezo, Silvia; Enríquez-Rincón, Fernando; Berg, Douglas E.

2001-01-01

Non-O1 strains of Vibrio cholerae implicated in gastroenteritis and diarrhea generally lack virulence determinants such as cholera toxin that are characteristic of epidemic strains; the factors that contribute to their virulence are not understood. Here we report that at least one-third of diarrhea-associated nonepidemic V. cholerae strains from Mexico cause vacuolation of cultured Vero cells. Detailed analyses indicated that this vacuolation was related to that caused by aerolysin, a pore-forming toxin of Aeromonas; it involved primarily the endoplasmic reticulum at early times (∼1 to 4 h after exposure), and resulted in formation of large, acidic, endosome-like multivesicular vacuoles (probably autophagosomes) only at late times (∼16 h). In contrast to vacuolation caused by Helicobacter pylori VacA protein, that induced by V. cholerae was exacerbated by agents that block vacuolar proton pumping but not by endosome-targeted weak bases. It caused centripetal redistribution of endosomes, reflecting cytoplasmic alkalinization. The gene for V. cholerae vacuolating activity was cloned and was found to correspond to hlyA, the structural gene for hemolysin. HlyA protein is a pore-forming toxin that causes ion leakage and, ultimately, eukaryotic cell lysis. Thus, a distinct form of cell vacuolation precedes cytolysis at low doses of hemolysin. We propose that this vacuolation, in itself, contributes to the virulence of V. cholerae strains, perhaps by perturbing intracellular membrane trafficking or ion exchange in target cells and thereby affecting local intestinal inflammatory or other defense responses. PMID:11179335

Cell vacuolation caused by Vibrio cholerae hemolysin.

Science.gov (United States)

Figueroa-Arredondo, P; Heuser, J E; Akopyants, N S; Morisaki, J H; Giono-Cerezo, S; Enríquez-Rincón, F; Berg, D E

2001-03-01

Non-O1 strains of Vibrio cholerae implicated in gastroenteritis and diarrhea generally lack virulence determinants such as cholera toxin that are characteristic of epidemic strains; the factors that contribute to their virulence are not understood. Here we report that at least one-third of diarrhea-associated nonepidemic V. cholerae strains from Mexico cause vacuolation of cultured Vero cells. Detailed analyses indicated that this vacuolation was related to that caused by aerolysin, a pore-forming toxin of Aeromonas; it involved primarily the endoplasmic reticulum at early times (approximately 1 to 4 h after exposure), and resulted in formation of large, acidic, endosome-like multivesicular vacuoles (probably autophagosomes) only at late times (approximately 16 h). In contrast to vacuolation caused by Helicobacter pylori VacA protein, that induced by V. cholerae was exacerbated by agents that block vacuolar proton pumping but not by endosome-targeted weak bases. It caused centripetal redistribution of endosomes, reflecting cytoplasmic alkalinization. The gene for V. cholerae vacuolating activity was cloned and was found to correspond to hlyA, the structural gene for hemolysin. HlyA protein is a pore-forming toxin that causes ion leakage and, ultimately, eukaryotic cell lysis. Thus, a distinct form of cell vacuolation precedes cytolysis at low doses of hemolysin. We propose that this vacuolation, in itself, contributes to the virulence of V. cholerae strains, perhaps by perturbing intracellular membrane trafficking or ion exchange in target cells and thereby affecting local intestinal inflammatory or other defense responses.

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.

Mechanical properties of seabed deposits of sand with strain history caused by waves

International Nuclear Information System (INIS)

Nishi, Koichi; Kanatani, Mamoru

1989-01-01

The research project on floating nuclear power plants, which was taken up as one of new siting technologies for the future, has been advanced by the Central Research Institute of Electric Power Industry. In this case, it is very important to cope with the stability problems of breakwaters, revetments, artificial islands and the foundation of mooring against strong earthquake motion and storm wave force. Accordingly it is necessary to evaluate accurately the stability, and to sufficiently understand the mechanical properties of seabed as the foundation ground of these offshore structures. Since seabed has the inherent strain history induced by the action of wave force, it is important to take such characteristics into account in the evaluation of the mechanical properties. In this report, the experimental results about the effect of the strain history on the strength-deformation properties of sand deposited on seabed are described, in order to contribute to the establishment of the method for precisely evaluating the properties of seabed. The computation method for shearing strain history in seabed and the method of estimating the strength-deformation characteristics of seabed are reported. (K.I.)

Engineering and preclinical evaluation of attenuated nontyphoidal Salmonella strains serving as live oral vaccines and as reagent strains.

Science.gov (United States)

Tennant, Sharon M; Wang, Jin-Yuan; Galen, James E; Simon, Raphael; Pasetti, Marcela F; Gat, Orit; Levine, Myron M

2011-10-01

While nontyphoidal Salmonella (NTS) has long been recognized as a cause of self-limited gastroenteritis, it is becoming increasingly evident that multiple-antibiotic-resistant strains are also emerging as important causes of invasive bacteremia and focal infections, resulting in hospitalizations and deaths. We have constructed attenuated Salmonella enterica serovar Typhimurium and Salmonella enterica serovar Enteritidis strains that can serve as live oral vaccines and as "reagent strains" for subunit vaccine production in a safe and economical manner. Prototype attenuated vaccine strains CVD 1921 and CVD 1941, derived from the invasive wild-type strains S. Typhimurium I77 and S. Enteritidis R11, respectively, were constructed by deleting guaBA, encoding guanine biosynthesis, and clpP, encoding a master protease regulator. The clpP mutation resulted in a hyperflagellation phenotype. An additional deletion in fliD yielded reagent strains CVD 1923 and CVD 1943, respectively, which export flagellin monomers. Oral 50% lethal dose (LD₅₀) analyses showed that the NTS vaccine strains were all highly attenuated in mice. Oral immunization with CVD 1921 or CVD 1923 protected mice against lethal challenge with wild-type S. Typhimurium I77. Immunization with CVD 1941 but not CVD 1943 protected mice against lethal infection with S. Enteritidis R11. Immune responses induced by these strains included high levels of serum IgG anti-lipopolysaccharide (LPS) and anti-flagellum antibodies, with titers increasing progressively during the immunization schedule. Since S. Typhimurium and S. Enteritidis are the most common NTS serovars associated with invasive disease, these findings can pave the way for development of a highly effective, broad-spectrum vaccine against invasive NTS.

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...

Causes, prevention and treatment of Escherichia coli infections.

Science.gov (United States)

Gould, Dinah

Escherichia coli is a normal inhabitant of the human gastrointestinal tract and can cause healthcare-associated infections. The organism is most frequently responsible for urinary tract infections and it is the bacterium most often implicated in the cause of diarrhoea in people travelling overseas. In recent years, a strain called Ecoli O157 has gained notoriety for causing foodborne infection, which can have severe health consequences, especially in young children. This article describes the range of different infections caused by Ecoli in healthcare settings and the community and discusses the characteristics of the different strains of the bacteria that explain variations in their pathogenicity.

School District Fiscal Strain: Implications for State and Federal Financial Assistance.

Science.gov (United States)

Hentschke, Guilbert; Yagielski, John

1982-01-01

Uses a model portraying school district decision makers as "consumers" to analyze fiscal strain's causes (enrollment decline, input price increases, and changes in input mix) as variants of the general consumer model. Measures the impact of each cause of fiscal strain and discusses implications for state and federal aid. (Author/RW)

Genome Sequences of Three Vaccine Strains and Two Wild-Type Canine Distemper Virus Strains from a Recent Disease Outbreak in South Africa.

Science.gov (United States)

Loots, Angelika K; Du Plessis, Morné; Dalton, Desiré Lee; Mitchell, Emily; Venter, Estelle H

2017-07-06

Canine distemper virus causes global multihost infectious disease. This report details complete genome sequences of three vaccine and two new wild-type strains. The wild-type strains belong to the South African lineage, and all three vaccine strains to the America 1 lineage. This constitutes the first genomic sequences of this virus from South Africa. Copyright © 2017 Loots et al.

Differential Delivery of Genomic Double-Stranded RNA Causes Reovirus Strain-Specific Differences in Interferon Regulatory Factor 3 Activation.

Science.gov (United States)

Stuart, Johnasha D; Holm, Geoffrey H; Boehme, Karl W

2018-05-01

signaling pathways, leading to the activation of interferon regulatory factor 3 (IRF3) and NF-κB, key transcription factors required for IFN-I induction. Serotype 3 (T3) reoviruses induce significantly more IFN-I than serotype 1 (T1) strains. In this work, we found that differences in IFN-I production by T1 and T3 reoviruses correlate with differential IRF3 activation. Differences in IRF3 activation are not caused by a blockade of the IRF3 activation by a T1 strain. Rather, differences in events during the late stages of viral entry determine the capacity of reovirus to activate host IFN-I responses. Together, our work provides insight into mechanisms of IFN-I induction by nonenveloped viruses. Copyright © 2018 American Society for Microbiology.

Generating strain signals under consideration of road surface profiles

Science.gov (United States)

Putra, T. E.; Abdullah, S.; Schramm, D.; Nuawi, M. Z.; Bruckmann, T.

2015-08-01

The current study aimed to develop the mechanism for generating strain signal utilising computer-based simulation. The strain data, caused by the acceleration, were undertaken from a fatigue data acquisition involving car movements. Using a mathematical model, the measured strain signals yielded to acceleration data used to describe the bumpiness of road surfaces. The acceleration signals were considered as an external disturbance on generating strain signals. Based on this comparison, both the actual and simulated strain data have similar pattern. The results are expected to provide new knowledge to generate a strain signal via a simulation.

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 ...

Comparison of the immune responses associated with experimental bovine mastitis caused by different strains of Escherichia coli.

Science.gov (United States)

Blum, Shlomo E; Heller, Elimelech D; Jacoby, Shamay; Krifucks, Oleg; Leitner, Gabriel

2017-05-01

We studied the mammary immune response to different mammary pathogenic Escherichia coli (MPEC) strains in cows, hypothesising that the dynamics of response would differ. E. coli is a major aetiologic agent of acute clinical bovine mastitis of various degrees of severity with specific strains being associated with persistent infections. We compared challenge with three distinct pathogenic MPEC strains (VL2874, VL2732 and P4), isolated from different forms of mastitis (per-acute, persistent and acute, respectively). A secondary objective was to verify the lack of mammary pathogenicity of an environmental isolate (K71) that is used for comparison against MPEC in genomic and phenotypic studies. Twelve cows were challenged by intra-mammary infusion with one of the strains. Cellular and chemokine responses and bacterial culture follow-up were performed for 35 d. All cows challenged by any of the MPEC strains developed clinical mastitis. Differences were found in the intensity and duration of response, in somatic cell count, secreted cytokines (TNF-α, IL-6 and IL-17) and levels of milk leucocyte membrane Toll-like receptor 4 (TLR4). A sharp decrease of TLR4 on leucocytes was observed concomitantly to peak bacterial counts in milk. Intra-mammary infusion of strain K71 did not elicit inflammation and bacteria were not recovered from milk. Results suggest some differences in the mammary immune response to distinct MPEC strains that could be correlated to their previously observed pathogenic traits. This is also the first report of an E. coli strain that is non-pathogenic to the bovine mammary gland.

Genetic analysis of Streptococcus agalactiae strains isolated from neonates and their mothers.

NARCIS (Netherlands)

Melchers, W.J.G.; Bakkers, J.M.J.E.; Toonen, M.; Kuppeveld, F.J.M. van; Trijbels-Smeulders, M.J.A.M.; Hoogkamp-Korstanje, J.A.A.

2003-01-01

Streptococcus agalactiae or group B streptococcus (GBS) is the most common cause of neonatal sepsis and meningitis in neonates. One of the major questions is whether the GBS strains able to cause neonatal invasive disease have peculiar genetic features. A collection of S. agalactiae strains,

Clinical and Molecular Characteristics of Human Rotavirus G8P[8] Outbreak Strain, Japan, 2014.

Science.gov (United States)

Kondo, Kenji; Tsugawa, Takeshi; Ono, Mayumi; Ohara, Toshio; Fujibayashi, Shinsuke; Tahara, Yasuo; Kubo, Noriaki; Nakata, Shuji; Higashidate, Yoshihito; Fujii, Yoshiki; Katayama, Kazuhiko; Yoto, Yuko; Tsutsumi, Hiroyuki

2017-06-01

During March-July 2014, rotavirus G8P[8] emerged as the predominant cause of rotavirus gastroenteritis among children in Hokkaido Prefecture, Japan. Clinical characteristics were similar for infections caused by G8 and non-G8 strains. Sequence and phylogenetic analyses suggest the strains were generated by multiple reassortment events between DS-1-like P[8] strains and bovine strains from Asia.

Differential stress transcriptome landscape of historic and recently emerged hypervirulent strains of Clostridium difficile strains determined using RNA-seq.

Directory of Open Access Journals (Sweden)

Joy Scaria

Full Text Available C. difficile is the most common cause of nosocomial diarrhea in North America and Europe. Genomes of individual strains of C. difficile are highly divergent. To determine how divergent strains respond to environmental changes, the transcriptomes of two historic and two recently isolated hypervirulent strains were analyzed following nutrient shift and osmotic shock. Illumina based RNA-seq was used to sequence these transcriptomes. Our results reveal that although C. difficile strains contain a large number of shared and strain specific genes, the majority of the differentially expressed genes were core genes. We also detected a number of transcriptionally active regions that were not part of the primary genome annotation. Some of these are likely to be small regulatory RNAs.

The Genetically Remote Pathogenic Strain NVH391-98 of the Bacillus cereus Group Represents the Cluster of Thermophilic Strains

Energy Technology Data Exchange (ETDEWEB)

Auger, Sandrine; Galleron, Nathalie; Bidnenko, Elena; Ehrlich, S. Dusko; Lapidus, Alla; Sorokin, Alexei

2007-10-02

Bacteria of the Bacillus cereus group are known to cause food poisoning. A rare phylogenetically remote strain, NVH391-98, was recently characterized to encode a particularly efficient cytotoxin K presumably responsible for food poisoning. This pathogenic strain and its close relatives can be phenotypically distinguished from other strains of the B. cereus group by the inability to grow at temperatures below 17 degrees C and by the ability to grow at temperatures from 48 to 53 degrees C. A temperate phage, phBC391A2, residing in the genome of NVH391-98 allows us to distinguish the three known members of this thermophilic strain cluster.

Failure strains and proposed limit strains for an reactor pressure vessel under severe accident conditions

International Nuclear Information System (INIS)

Krieg, R.

2005-01-01

The local failure strains of essential design elements of a reactor vessel are investigated. The size influence of the structure is of special interest. Typical severe accident conditions including elevated temperatures and dynamic loads are considered. The main part of work consists of test families with specimens under uniaxial and biaxial load. Within one test family the specimen geometry and the load conditions are similar, but the size is varied up to reactor dimensions. Special attention is given to geometries with a hole or a notch causing non-uniform stress and strain distributions typical for the reactor vessel. A key problem is to determine the local failure strain. Here suitable methods had to be developed including the so-called 'vanishing gap method', and the 'forging die method'. They are based on post-test geometrical measurements of the fracture surfaces and reconstructions of the related strain fields using finite element models. The results indicate that stresses versus dimensionless deformations are approximately size independent up to failure for specimens of similar geometry under similar load conditions. Local failure strains could be determined. The values are rather high and size dependent. Statistical evaluation allow the proposal of limit strains which are also size dependent. If these limit strains are not exceeded, the structures will not fracture

Evaluation of pathogenesis caused in cattle and guinea pig by a Mycobacterium bovis strain isolated from wild boar

Directory of Open Access Journals (Sweden)

Di Rienzo Julio

2011-07-01

Full Text Available Abstract Background In many regions of the world, wild mammals act as reservoir of Mycobacterium bovis, a situation that prevents the eradication of bovine tuberculosis. In order to observe whether a strain isolated from a wild boar, previously tested as highly virulent in a mice model, is also virulent in cattle, we performed cattle experimental inoculation with this strain Results Groups of Friesian calves were either infected with the wild boar strain M. bovis 04-303 or with the bovine strain NCTC10772 as a control. We found that antigen-specific IFN-γ release in whole blood samples occurred earlier in animals infected with M. bovis 04-303. Both M. bovis strains resulted in a positive skin test, with animals infected with the wild boar isolate showing a stronger response. These results and the presence of more severe organ lesions, with granuloma and pneumonic areas in cattle demonstrate that the wild boar isolate is more virulent than the NCTC10772 strain. Additionally, we tested the infectivity of the M. bovis strains in guinea pigs and found that M. bovis 04-303 had the highest pathogenicity. Conclusions M. bovis strains isolated from wild boars may be pathogenic for cattle, producing TB lesions.

Quantitative strain analysis for advanced CMOS technology by Nano Beam Diffraction

KAUST Repository

Wang, Qingxiao

2010-07-01

Nano Beam Diffraction has been used to analyze the local strain distribution in MOS transistors. The influence of wafer process on the channel strain has been systematically analyzed in this paper. The source/drain implantation can cause a little strain loss but the silicidation step is the key process in which dramatic strain loss has been found. © 2010 IEEE.

Quantitative strain analysis for advanced CMOS technology by Nano Beam Diffraction

KAUST Repository

Wang, Qingxiao; Zhu, Jinmin; Du, Anyan; Liu, Jinping; Hua, YouNan

2010-01-01

Nano Beam Diffraction has been used to analyze the local strain distribution in MOS transistors. The influence of wafer process on the channel strain has been systematically analyzed in this paper. The source/drain implantation can cause a little strain loss but the silicidation step is the key process in which dramatic strain loss has been found. © 2010 IEEE.

Histophilus somni IbpA Fic cytotoxin is conserved in disease strains and most carrier strains from cattle, sheep and bison.

Science.gov (United States)

Zekarias, B; O'Toole, D; Lehmann, J; Corbeil, L B

2011-04-21

Histophilus somni causes bovine pneumonia, septicemia, myocarditis, thrombotic meningoencephalitis and arthritis, as well as a genital or upper respiratory carrier state in normal animals. However, differences in virulence factors among strains are not well studied. The surface and secreted immunoglobulin binding protein A (IbpA) Fic motif of H. somni causes bovine alveolar type 2 (BAT2) cells to retract, allowing virulent bacteria to cross the alveolar monolayer. Because H. somni IbpA is an important virulence factor, its presence was evaluated in different strains from cattle, sheep and bison to define whether there are syndrome specific markers and whether antigenic/molecular/functional conservation occurs. A few preputial carrier strains lacked IbpA by Western blotting but all other tested disease or carrier strains were IbpA positive. These positive strains had either both IbpA DR1/Fic and IbpA DR2/Fic or only IbpA DR2/Fic by PCR. IbpA Fic mediated cytotoxicity for BAT2 cells and sequence analysis of IbpA DR2/Fic from selected strains revealed conservation of sequence and function in disease and IbpA positive carrier strains. Passive protection of mice against H. somni septicemia with antibody to IbpA DR2/Fic, along with previous data, indicates that the IbpA DR1/Fic and/or DR2/Fic domains are candidate vaccine antigens for protection against many strains of H. somni. Since IbpA DR2/Fic is conserved in most carrier strains, they may be virulent if introduced to susceptible animals at susceptible sites. Conservation of the protective IbpA antigen in all disease isolates tested is encouraging for development of protective vaccines and diagnostic assays. Copyright © 2010 Elsevier B.V. All rights reserved.

Identification of Diarrheagenic Escherichia coli Strains from Avian Organic Fertilizers

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Juan Puño-Sarmiento

2014-08-01

Full Text Available The Brazilian poultry industry generates large amounts of organic waste, such as chicken litter, which is often used in agriculture. Among the bacteria present in organic fertilizer are members of the Enterobacteriaceae family. The objective of this study was to detect the presence of diarrheagenic Escherichia coli (DEC strains in avian organic fertilizer, and assess the potential damage they can cause in humans due to antimicrobial resistance. The presence of DEC pathotypes and phylogenetic groups were detected by multiplex-PCR. Phenotypic assays, such as tests for adhesion, cytotoxicity activity, biofilm formation and especially antimicrobial susceptibility, were performed. Fifteen DEC strains from 64 E. coli were isolated. Among these, four strains were classified as enteropathogenic (EPEC; 6.2%, three strains as Shiga toxin-producing (STEC; 4.7%, 10 strains as enteroaggregative (EAEC; 12.5%, but two of these harbored the eaeA gene too. The low number of isolated strains was most likely due to the composting process, which reduces the number of microorganisms. These strains were able to adhere to HEp-2 and HeLa cells and produce Shiga-toxins and biofilms; in addition, some of the strains showed antimicrobial resistance, which indicates a risk of the transfer of resistance genes to human E. coli. These results showed that DEC strains isolated from avian organic fertilizers can cause human infections.

Identification of diarrheagenic Escherichia coli strains from avian organic fertilizers.

Science.gov (United States)

Puño-Sarmiento, Juan; Gazal, Luis Eduardo; Medeiros, Leonardo P; Nishio, Erick K; Kobayashi, Renata K T; Nakazato, Gerson

2014-08-28

The Brazilian poultry industry generates large amounts of organic waste, such as chicken litter, which is often used in agriculture. Among the bacteria present in organic fertilizer are members of the Enterobacteriaceae family. The objective of this study was to detect the presence of diarrheagenic Escherichia coli (DEC) strains in avian organic fertilizer, and assess the potential damage they can cause in humans due to antimicrobial resistance. The presence of DEC pathotypes and phylogenetic groups were detected by multiplex-PCR. Phenotypic assays, such as tests for adhesion, cytotoxicity activity, biofilm formation and especially antimicrobial susceptibility, were performed. Fifteen DEC strains from 64 E. coli were isolated. Among these, four strains were classified as enteropathogenic (EPEC; 6.2%), three strains as Shiga toxin-producing (STEC; 4.7%), 10 strains as enteroaggregative (EAEC; 12.5%), but two of these harbored the eaeA gene too. The low number of isolated strains was most likely due to the composting process, which reduces the number of microorganisms. These strains were able to adhere to HEp-2 and HeLa cells and produce Shiga-toxins and biofilms; in addition, some of the strains showed antimicrobial resistance, which indicates a risk of the transfer of resistance genes to human E. coli. These results showed that DEC strains isolated from avian organic fertilizers can cause human infections.

Effect of various Portland cement paste compositions on early-age strain

Science.gov (United States)

Guzzetta, Alana G.

Early-age strain in paste, mortar, and concrete mixtures was investigated using a new method where the specimen shape was a cone frustum. Strain of the specimen from both the horizontal and vertical directions was captured by height change measurement. The volumetric strain was then calculated as a function of the height change and was plotted versus time. A correlation was found between the slopes of the volumetric strain curve resulting from this test method and the initial setting time of the tested material. An initial evaluation of the repeatability of this innovative test method was conducted. The early-age strain effects of aggregate volume, shrinkage reducing admixture, water-cementitious ratio (w/cm), and partial cement replacement with supplementary cementitious materials were tested and individually compared. From these comparisons, it was observed that ambient temperature, bleed water development, and rheological properties had a significant impact on the volumetric strain results. Data showed increased strain as aggregate volume was reduced and as the w/cm was changed from 0.25 up to 0.50. The addition of shrinkage reducing admixture generally caused an increase in the 36-hour volumetric strain value. In most of the mixtures, cement replacement with 20% fly ash or 10% metakaolin reduced the measured volumetric strain when the w/cm was 0.30. Replacement of cement with 10% silica fume caused an insignificant change in volumetric strain results.

Analysis of invasive pneumonia-causing strains of Streptococcus pneumoniae: serotypes and antimicrobial susceptibility.

Science.gov (United States)

Yoshioka, Cristina R M; Martinez, Marina B; Brandileone, Maria C C; Ragazzi, Selma B; Guerra, Maria L L S; Santos, Silvia R; Shieh, Huei H; Gilio, Alfredo E

2011-01-01

To identify the most common pneumococcal serotypes in children hospitalized with invasive pneumonia, correlate isolated serotypes with those included in conjugate vaccines, and ascertain the sensitivity of the isolated pneumococcal strains to penicillin and other antibiotics. From January 2003 to October 2008, a retrospective study of hospitalized children with a diagnosis of Streptococcus pneumoniae pneumonia was conducted at the university hospital of Universidade de São Paulo. Criteria for inclusion were: age greater than 29 days and less than 15 years, radiological and clinical diagnosis of pneumonia, and isolation of Streptococcus pneumoniae in blood cultures and/or pleural effusion. The study included 107 children. The most common serotypes were 14 (36.5%), 1 (16%), 5 (14.6%), 6B (6.3%) and 3 (4.2%). The proportion of identified serotypes contained in the heptavalent, 10-valent and 13-valent conjugate vaccines was 53.1, 86.5, and 96.9%, respectively. Pneumococcal strains were sensitive to penicillin (minimum inhibitory concentration, MIC ≤ 2 µg/mL) in 100 cases (93.5%) and displayed intermediate resistance (MIC = 4 µg/mL) in 7 cases (6.5%). No strains were penicillin-resistant (MIC ≥ 8 µg/mL) according to the Clinical and Laboratory Standards Institute 2008 standards. Tested isolates were highly sensitive to vancomycin, rifampicin, ceftriaxone, clindamycin, erythromycin, and chloramphenicol. Our results confirm a significant potential impact of conjugate vaccines, mainly 10-valent and 13-valent, on invasive pneumonia. Furthermore, susceptibility testing results show that penicillin is still the treatment of choice for invasive pneumonia in our setting.

Nanocomposite Strain Gauges Having Small TCRs

Science.gov (United States)

Gregory, Otto; Chen, Ximing

2009-01-01

Ceramic strain gauges in which the strain-sensitive electrically conductive strips made from nanocomposites of noble metal and indium tin oxide (ITO) are being developed for use in gas turbine engines and other power-generation systems in which gas temperatures can exceed 1,500 F (about 816 C). In general, strain gauges exhibit spurious thermally induced components of response denoted apparent strain. When temperature varies, a strain-gauge material that has a nonzero temperature coefficient of resistance (TCR) exhibits an undesired change in electrical resistance that can be mistaken for the change in resistance caused by a change in strain. It would be desirable to formulate straingauge materials having TCRs as small as possible so as to minimize apparent strain. Most metals exhibit positive TCRs, while most semiconductors, including ITO, exhibit negative TCRs. The present development is based on the idea of using the negative TCR of ITO to counter the positive TCRs of noble metals and of obtaining the benefit of the ability of both ITO and noble metals to endure high temperatures. The noble metal used in this development thus far has been platinum. Combinatorial libraries of many ceramic strain gauges containing nanocomposites of various proportions of ITO and platinum were fabricated by reactive co-sputtering from ITO and platinum targets onto alumina- and zirconia-based substrates mounted at various positions between the targets.

Mechanical control over valley magnetotransport in strained graphene

Energy Technology Data Exchange (ETDEWEB)

Ma, Ning, E-mail: maning@stu.xjtu.edu.cn [Department of Physics, MOE Key Laboratory of Advanced Transducers and Intelligent Control System, Taiyuan University of Technology, Taiyuan 030024 (China); Department of Applied Physics, MOE Key Laboratory for Nonequilibrium Synthesis and Modulation of Condensed Matter, Xi' an Jiaotong University, Xi' an 710049 (China); Zhang, Shengli, E-mail: zhangsl@mail.xjtu.edu.cn [Department of Applied Physics, MOE Key Laboratory for Nonequilibrium Synthesis and Modulation of Condensed Matter, Xi' an Jiaotong University, Xi' an 710049 (China); Liu, Daqing, E-mail: liudq@cczu.edu.cn [School of Mathematics and Physics, Changzhou University, Changzhou 213164 (China)

2016-05-06

Recent experiments report that the graphene exhibits Landau levels (LLs) that form in the presence of a uniform strain pseudomagnetic field with magnitudes up to hundreds of tesla. We further reveal that the strain removes the valley degeneracy in LLs, and leads to a significant valley polarization with inversion symmetry broken. This accordingly gives rise to the well separated valley Hall plateaus and Shubnikov–de Haas oscillations. These effects are absent in strainless graphene, and can be used to generate and detect valley polarization by mechanical means, forming the basis for the new paradigm “valleytronics” applications. - Highlights: • We explore the mechanical strain effects on the valley magnetotransport in graphene. • We analytically derive the dc collisional and Hall conductivities under strain. • The strain removes the valley degeneracy in Landau levels. • The strain causes a significant valley polarization with inversion symmetry broken. • The strain leads to the well separated valley Hall and Shubnikov–de Haas effects.

Antagonism of Trichoderma spp. strains against pea (Pisum sativum L. Fusarium wilt caused by Fusarium oxysporum f. sp. pisi.

Directory of Open Access Journals (Sweden)

Oscar Eduardo Checa Coral

2017-07-01

Full Text Available The antagonistic effectiveness of native strains of Trichoderma spp. on Fusarium oxysporum f. sp. pisi. in vitro, greenhouse and field conditions, were evaluated. in vitro conditions, the antagonistic capacity of 12 strains of Trichoderma spp., C2, C7, C12 and C21 strains, exhibited a better behavior measured by the following variables: inhibition halo and mycelial growth. In greenhouse conditions, the four strains, which showed the best in vitro antagonistic behavior, were evaluated using a DIA experimental design with factorial arrangement for three factors, which corresponded to strain, concentration and dose. The results of this evaluation, showed that C12 and C21 strains at doses of 20 mL, and at concentrations of 108 and 106 conidia.mL-1, respectively. The best antagonistic response was determined by variables as follows: plant height, fresh root weight and incidence. Under field conditions, the evaluations were carried out in the municipalities of Ipiales, Pupiales and Gualmatán, in the department of Nariño, Colombia. In each location, a BCA experimental design was used with four treatments and five replicates, treatments were as follows: C12 strains at 108 concentration, C21 at 106 concentration, chemical control and absolute control. In Gualmatan location, C12 and C21 strains, showed no antagonistic capacity, whereas in Ipiales and Pupiales locations, strain C12, presented a lower incidence of F. oxysporum than the control, but with no effect on yields. In Pupiales location, C21 strain surpassed in performance to the control treatment, even though the two treatments had similar incidence.

Strain Rate Effect on Tensile Behavior for a High Specific Strength Steel: From Quasi-Static to Intermediate Strain Rates

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Wei Wang

2017-12-01

Full Text Available The strain rate effect on the tensile behaviors of a high specific strength steel (HSSS with dual-phase microstructure has been investigated. The yield strength, the ultimate strength and the tensile toughness were all observed to increase with increasing strain rates at the range of 0.0006 to 56/s, rendering this HSSS as an excellent candidate for an energy absorber in the automobile industry, since vehicle crushing often happens at intermediate strain rates. Back stress hardening has been found to play an important role for this HSSS due to load transfer and strain partitioning between two phases, and a higher strain rate could cause even higher strain partitioning in the softer austenite grains, delaying the deformation instability. Deformation twins are observed in the austenite grains at all strain rates to facilitate the uniform tensile deformation. The B2 phase (FeAl intermetallic compound is less deformable at higher strain rates, resulting in easier brittle fracture in B2 particles, smaller dimple size and a higher density of phase interfaces in final fracture surfaces. Thus, more energy need be consumed during the final fracture for the experiments conducted at higher strain rates, resulting in better tensile toughness.

Strain engineering on transmission carriers of monolayer phosphorene.

Science.gov (United States)

Zhang, Wei; Li, Feng; Hu, Junsong; Zhang, Ping; Yin, Jiuren; Tang, Xianqiong; Jiang, Yong; Wu, Bozhao; Ding, Yanhuai

2017-11-22

The effects of uniaxial strain on the structure, band gap and transmission carriers of monolayer phosphorene were investigated by first-principles calculations. The strain induced semiconductor-metal as well as direct-indirect transitions were studied in monolayer phosphorene. The position of CBM which belonged to indirect gap shifts along the direction of the applied strain. We have concluded the change rules of the carrier effective mass when plane strains are applied. In band structure, the sudden decrease of band gap or the new formation of CBM (VBM) causes the unexpected change in carrier effective mass. The effects of zigzag and armchair strain on the effective electron mass in phosphorene are different. The strain along zigzag direction has effects on the electrons effective mass along both zigzag and armchair direction. By contrast, armchair-direction strain seems to affect only on the free electron mass along zigzag direction. For the holes, the effective masses along zigzag direction are largely affected by plane strains while the effective mass along armchair direction exhibits independence in strain processing. The carrier density of monolayer phosphorene at 300 K is calculated about [Formula: see text] cm -2 , which is greatly influenced by the temperature and strain. Strain engineering is an efficient method to improve the carrier density in phosphorene.

Kinases of two strains of Mycoplasma hyopneumoniae and a strain of Mycoplasma synoviae: an overview

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Alexandre Melo Bailão

2007-01-01

Full Text Available Mycoplasma synoviae and Mycoplasma hyopneumoniae are wall-less eubacteria belonging to the class of Mollicutes. These prokaryotes have a reduced genome size and reduced biosynthetic machinery. They cause great losses in animal production. M. synoviae is responsible for an upper respiratory tract disease of chickens and turkeys. M. hyopneumoniae is the causative agent of enzootic pneumonia in pigs. The complete genomes of these organisms showed 17 ORFs encoding kinases in M. synoviae and 15 in each of the M. hyopneumoniae strain. Four kinase genes were restricted to the avian pathogen while three were specific to the pig pathogen when compared to each other. All deduced kinases found in the non pathogenic strain (J[ATCC25934] were also found in the pathogenic M. hyopneumoniae strain. The enzymes were classified in nine families composing five fold groups.

Isolation and purification of Gallid herpesvirus 2 strains currently distributed in Japan.

Science.gov (United States)

Machida, Yuka; Murata, Shiro; Matsuyama-Kato, Ayumi; Isezaki, Masayoshi; Taneno, Akira; Sakai, Eishi; Konnai, Satoru; Ohashi, Kazuhiko

2017-01-20

Gallid herpesvirus 2 (GaHV-2) causes malignant lymphomas in chickens (Marek's disease, MD). Although MD is controlled through vaccination efforts, field isolates of GaHV-2 have increased in virulence worldwide and even cause MD in vaccinated chickens. GaHV-2 strains are classified into four categories (mild, virulent, very virulent and very virulent +) based on the virulence exhibited in experimental infection in unvaccinated or MD-vaccinated susceptible chickens. Although MD cases are sporadically reported in Japan, the recent field strains of GaHV-2 in Japan have not been characterized. During isolation of recent field strains by using primary chicken kidney cell cultures, a method classically used for GaHV-2 isolation, vaccine strains were simultaneously isolated. Therefore, it is necessary to separate vaccine strains to characterize the virulence and pathogenicity of the GaHV-2 strains currently distributed in Japan. In this study, we prepared cell suspensions from the spleens of MD-symptomatic chickens, inoculated day-old-chicks and isolated GaHV-2 strains by primary chicken kidney cell cultures at 2-3 weeks post inoculation. The isolated strains were passaged several times on chicken embryo fibroblast cells, and PCR analysis revealed that the isolated strains were not contaminated with vaccine strains. Moreover, the contaminant vaccine strains were completely removed by the purification of plaques observed in chicken kidney cells. These procedures are necessary to isolate GaHV-2 field strains from vaccine strains in order to carry out future studies to characterize these strains and glean insights into GaHV-2 virulence and pathogenicity.

Finite Element Model of the Strain Gauge For Determining Uniaxial Tension

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Vladimír GOGA

2013-12-01

Full Text Available Strain gauge is device used to measure the mechanical strains of solid bodies. Deformation of the strain gauge element causes changes its electrical resistance. This resistance change, usually measured using a Wheatstone bridge, is related to the strain by the quantity known as the gauge factor. When the stains are known, it is possible to determined state of stress at a point of measured body using generalized Hooke`s law and Mohr`s circle. Finite element analysis of strain gauge measurement using ANSYS software is subject of this article.

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 ...

Quantification of gravity-induced skin strain across the breast surface.

Science.gov (United States)

Sanchez, Amy; Mills, Chris; Haake, Steve; Norris, Michelle; Scurr, Joanna

2017-12-01

Quantification of the magnitude of skin strain in different regions of the breast may help to estimate possible gravity-induced damage whilst also being able to inform the selection of incision locations during breast surgery. The aim of this study was to quantify static skin strain over the breast surface and to estimate the risk of skin damage caused by gravitational loading. Fourteen participants had 21 markers applied to their torso and left breast. The non-gravity breast position was estimated as the mid-point of the breast positions in water and soybean oil (higher and lower density than breast respectively). The static gravity-loaded breast position was also measured. Skin strain was calculated as the percentage extension between adjacent breast markers in the gravity and non-gravity loaded conditions. Gravity induced breast deformation caused peak strains ranging from 14 to 75% across participants, with potentially damaging skin strain (>60%) in one participant and skin strains above 30% (skin resistance zone) in a further four participants. These peak strain values all occurred in the longitudinal direction in the upper region of the breast skin. In the latitudinal direction, smaller-breasted participants experienced greater strain on the outer (lateral) breast regions and less strain on the inner (medial) breast regions, a trend which was reversed in the larger breasted participants (above size 34D). To reduce tension on surgical incisions it is suggested that preference should be given to medial latitudinal locations for smaller breasted women and lateral latitudinal locations for larger breasted women. Copyright © 2017 Elsevier Ltd. All rights reserved.

Specific selection for virulent urinary tract infectious Escherichia coli strains during catheter-associated biofilm formation

DEFF Research Database (Denmark)

Ferrieres, Lionel; Hancock, Viktoria; Klemm, Per

2007-01-01

microorganisms can attach. Urinary tract infectious (UTI) Escherichia coli range in pathogenicity and the damage they cause - from benign asymptomatic bacteriuria (ABU) strains, which inflict no or few problems to the host, to uropathogenic E. coli (UPEC) strains, which are virulent and often cause severe...... for and promote biofilm formation of the most virulent group of UTI E. coli strains, hardly a desirable situation for the catheterized patient....

[Virulence markers of Escherichia coli O1 strains].

Science.gov (United States)

Makarova, M A; Kaftyreva, L A; Grigor'eva, N S; Kicha, E V; Lipatova, L A

2011-01-01

To detect virulence genes in clinical isolates of Escherichia coli O1 using polymerase chain reaction (PCR). One hundred and twenty strains of E.coli O1 strains isolated from faeces of patients with acute diarrhea (n = 45) and healthy persons (n = 75) were studied. PCR with primers for rfb and fliC genes, which control synthesis of O- and H- antigens respectively, was used. Fourteen virulence genes (pap, aaf, sfa, afa, eaeA, bfpA, ial, hly, cnf, stx1, stx2, lt, st, and aer) were detected by PCR primers. K1-antigen was determined by Pastorex Meningo B/E. coli O1 kit (Bio-Rad). rfb gene controlling O-antigen synthesis in serogroup O1 as well as fliC gene controlling synthesis of H7 and K1 antigens were detected in all strains. Thus all E. coli strains had antigenic structure O1:K1 :H-:F7. Virulence genes aafl, sfa, afa, eaeA, bfpA, ial, hly, cnf, stx1, stx2, lt, and st were not detected. All strains owned pap and aer genes regardless of the presence of acute diarrhea symptoms. It was shown that E. coli O1:KI:H-:F7 strains do not have virulence genes which are characteristic for diarrhea-causing Escherichia. In accordance with the presence of pap and aer genes they could be attributed to uropathogenic Escherichia (UPEC) or avian-pathogenic Escherichia (APEC). It is necessary to detect virulence factors in order to determine E. coli as a cause of intestinal infection.

Laser-induced photo-thermal strain imaging

Science.gov (United States)

Choi, Changhoon; Ahn, Joongho; Jeon, Seungwan; Kim, Chulhong

2018-02-01

Vulnerable plaque is the one of the leading causes of cardiovascular disease occurrence. However, conventional intravascular imaging techniques suffer from difficulty in finding vulnerable plaque due to limitation such as lack of physiological information, imaging depth, and depth sensitivity. Therefore, new techniques are needed to help determine the vulnerability of plaque, Thermal strain imaging (TSI) is an imaging technique based on ultrasound (US) wave propagation speed that varies with temperature of medium. During temperature increase, strain occurs in the medium and its variation tendency is depending on the type of tissue, which makes it possible to use for tissue differentiation. Here, we demonstrate laser-induced photo-thermal strain imaging (pTSI) to differentiate tissue using an intravascular ultrasound (IVUS) catheter and a 1210-nm continuous-wave laser for heating lipids intensively. During heating, consecutive US images were obtained from a custom-made phantom made of porcine fat and gelatin. A cross correlation-based speckle-tracking algorithm was then applied to calculate the strain of US images. In the strain images, the positive strain produced in lipids (porcine fat) was clearly differentiated from water-bearing tissue (gelatin). This result shows that laser-induced pTSI could be a new method to distinguish lipids in the plaque and can help to differentiate vulnerability of plaque.

The plane strain shear fracture of the advanced high strength steels

International Nuclear Information System (INIS)

Sun, Li

2013-01-01

The “shear fracture” which occurs at the high-curvature die radii in the sheet metal forming has been reported to remarkably limit the application of the advanced high strength steels (AHSS) in the automobile industry. However, this unusual fracture behavior generally cannot be predicted by the traditional forming limit diagram (FLD). In this research, a new experimental system was developed in order to simulate the shear fracture, especially at the plane strain state which is the most common state in the auto-industry and difficult to achieve in the lab due to sample size. Furthermore, the system has the capability to operate in a strain rate range from quasi-static state to the industrial forming state. One kinds of AHSS, Quenching-Partitioning (QP) steels have been performed in this test and the results show that the limiting fracture strain is related to the bending ratio and strain rate. The experimental data support that deformation-induced heating is an important cause of “shear fracture” phenomena for AHSS: a deformation-induced quasi-heating caused by smaller bending ratio and high strain rate produce a smaller limiting plane strain and lead a “shear fracture” in the component

[Determination of genetic bases of auxotrophy in Yersinia pestis ssp. caucasica strains].

Science.gov (United States)

Odinokov, G N; Eroshenko, G A; Kukleva, L M; Shavina, N Iu; Krasnov, Ia M; Kutyrev, V V

2012-04-01

Based on the results of computer analysis of nucleotide sequences in strains Yersinia pestis and Y. pseudotuberculosis recorded in the files of NCBI GenBank database, differences between genes argA, aroG, aroF, thiH, and thiG of strain Pestoides F (subspecies caucasica) were found, compared to other strains of plaque agent and pseudotuberculosis microbe. Using PCR with calculated primers and the method of sequence analysis, the structure of variable regions of these genes was studied in 96 natural Y. pestis and Y. pseudotuberculosis strains. It was shown that all examined strains of subspecies caucasica, unlike strains of plague-causing agent of other subspecies and pseudotubercolosis microbe, had identical mutations in genes argA (integration of the insertion sequence IS100), aroG (insertion of ten nucleotides), aroF (inserion of IS100), thiH (insertion of nucleotide T), and thiG (deletion of 13 nucleotides). These mutations are the reason for the absence in strains belonging to this subspecies of the ability to synthesize arginine, phenylalanine, tyrosine, and vitamin B1 (thiamine), and cause their auxotrophy for these growth factors.

Root causes investigation of catastrophic optical bulk damage in high-power InGaAs-AlGaAs strained QW lasers

Science.gov (United States)

Sin, Yongkun; Lingley, Zachary; Ayvazian, Talin; Brodie, Miles; Ives, Neil

2018-02-01

High-power single-mode (SM) and multi-mode (MM) InGaAs-AlGaAs strained quantum well (QW) lasers are critical components for both terrestrial and space satellite communications systems. Since these lasers predominantly fail by catastrophic and sudden degradation due to COD, it is especially crucial for space satellite applications to investigate reliability, failure modes, and degradation mechanisms of these lasers. Our group reported a new failure mode in MM and SM InGaAs-AlGaAs strained QW lasers in 2009 and 2016, respectively. Our group also reported in 2017 that bulk failure due to catastrophic optical bulk damage (COBD) is the dominant failure mode of both SM and MM lasers that were subject to long-term life-tests. For the present study, we report root causes investigation of COBD by performing long-term lifetests followed by failure mode analysis (FMA) using various micro-analytical techniques including electron beam induced current (EBIC), time-resolved electroluminescence (EL), focused ion beam (FIB), high-resolution transmission electron microscopy (TEM), and deep level transient spectroscopy (DLTS). Our life-tests with accumulated test hours of over 25,000 hours for SM lasers and over 35,000 hours for MM lasers generated a number of COBD failures with various failure times. EBIC techniques were employed to study dark line defects (DLDs) generated in SM COBD failures stressed under different test conditions. FIB and high-resolution TEM were employed to prepare cross sectional and plan view TEM specimens to study DLD areas (dislocations) in post-aged SM lasers. Time-resolved EL techniques were employed to study initiation and progressions of dark spots and dark lines in real time as MM lasers were aged. Lastly, to investigate precursor signatures of failure and degradation mechanisms responsible for COBD in both SM and MM lasers, we employed DLTS techniques to study a role that electron traps (non-radiative recombination centers) play in degradation of these

Endemic and Epidemic Lineages of Escherichia coli that Cause Urinary Tract Infections

Science.gov (United States)

Tabor, Helen; Tellis, Patricia; Vincent, Caroline; Tellier, Pierre-Paul

2008-01-01

Women with urinary tract infections (UTIs) in California, USA (1999–2001), were infected with closely related or indistinguishable strains of Escherichia coli (clonal groups), which suggests point source dissemination. We compared strains of UTI-causing E. coli in California with strains causing such infections in Montréal, Québec, Canada. Urine specimens from women with community-acquired UTIs in Montréal (2006) were cultured for E. coli. Isolates that caused 256 consecutive episodes of UTI were characterized by antimicrobial drug susceptibility profile, enterobacterial repetitive intergenic consensus 2 PCR, serotyping, XbaI and NotI pulsed-field gel electrophoresis, multilocus sequence typing, and phylogenetic typing. We confirmed the presence of drug-resistant, genetically related, and temporally clustered E. coli clonal groups that caused community-acquired UTIs in unrelated women in 2 locations and 2 different times. Two clonal groups were identified in both locations. Epidemic transmission followed by endemic transmission of UTI-causing clonal groups may explain these clusters of UTI cases. PMID:18826822

Genetic Diversity of Tick-Borne Rickettsial Pathogens; Insights Gained from Distant Strains

Directory of Open Access Journals (Sweden)

Sebastián Aguilar Pierlé

2014-01-01

Full Text Available The ability to capture genetic variation with unprecedented resolution improves our understanding of bacterial populations and their ability to cause disease. The goal of the pathogenomics era is to define genetic diversity that results in disease. Despite the economic losses caused by vector-borne bacteria in the Order Rickettsiales, little is known about the genetic variants responsible for observed phenotypes. The tick-transmitted rickettsial pathogen Anaplasma marginale infects cattle in tropical and subtropical regions worldwide, including Australia. Genomic analysis of North American A. marginale strains reveals a closed core genome defined by high levels of Single Nucleotide Polymorphisms (SNPs. Here we report the first genome sequences and comparative analysis for Australian strains that differ in virulence and transmissibility. A list of genetic differences that segregate with phenotype was evaluated for the ability to distinguish the attenuated strain from virulent field strains. Phylogenetic analyses of the Australian strains revealed a marked evolutionary distance from all previously sequenced strains. SNP analysis showed a strikingly reduced genetic diversity between these strains, with the smallest number of SNPs detected between any two A. marginale strains. The low diversity between these phenotypically distinct bacteria presents a unique opportunity to identify the genetic determinants of virulence and transmission.

Strain and defect microstructure in ion-irradiated GeSi/Si strained layers as a function of annealing temperature

International Nuclear Information System (INIS)

Glasko, J.M.; Elliman, R.G.; Zou, J.; Cockayne, D.J.H.; Fitz Gerald, J.D.

1998-01-01

High energy (1 MeV), ion irradiation of GeSi/Si strained layers at elevated temperatures can cause strain relaxation. In this study, the effect of subsequent thermal annealing was investigated. Three distinct annealing stages were identified and correlated with the evolution of the defect microstructure. In the temperature range from 350 to 600 deg C, a gradual recovery of strain is observed. This is believed to result from the annealing of small defect clusters and the growth of voids. The voids are visible at annealing temperatures in excess of 600 deg C, consistent with an excess vacancy concentration in the irradiated alloy layer. The 600 to 750 deg C range is marked by pronounced maximal recovery of strain, and is correlated with the dissolution of faulted loops in the substrate. At temperatures in the range 750-1000 deg C, strain relaxation is observed and is correlated with the growth of intrinsic dislocations within the alloy layer. These dislocations nucleate at the alloy-substrate interface and grow within the alloy layer, towards the surface. (authors)

Influence of Environmental Parameters on Trichoderma Strains with Biocontrol Potential

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Zsuzsanna Antal

2003-01-01

Full Text Available Several mycoparasitic strains belonging to the filamentous fungal genus Trichoderma are promising candidates for the biological control of plant pathogenic fungi. When planning the application of antagonistic Trichoderma strains for the purposes of biological control, it is very important to consider the environmental parameters affecting the biocontrol agents in the soil. A series of abiotic and biotic environmental parameters has an influence on the biocontrol efficacy of Trichoderma. Some important parameters to be considered are the effects of temperature, water potential and pH, and the presence of pesticides, metal ions and antagonistic bacteria in the soil. Most of the Trichoderma strains are mesophilic. Low temperatures in winter may cause a problem during biological control by influencing the activity of the biocontrol agents. Another problem emerging during the application of Trichoderma strains as biocontrol agents is that they cannot tolerate dry conditions, however, we may need biocontrol agents against plant pathogenic fungi which are able to grow and cause disease even in dry soils. The pH characteristics of the soil also belong to the most important environmental parameters affecting the activities of mycoparasitic Trichoderma strains. Within the frames of a complex integrated plant protection strategy, we may have to combine Trichoderma strains with chemical pesticides or metal compounds, therefore it is important to collect information about the effects of pesticides and metal ions on the biocontrol strains. Antagonistic soil bacteria may also have negative effects on the biocontrol abilities of Trichoderma strains, therefore it may be advantageous if a biocontrol strain possesses bacterium- degrading abilities as well. This review will discuss the literature about the influence of temperature, water potential, pH, pesticides, metal ions and antagonistic bacteria on mycoparasitic Trichoderma strains including the results of our

Uniaxial Strain Induced Critical Current Degradation of Ag-Sheathed Bi-2212 Round Wire

NARCIS (Netherlands)

Dai, Chao; Qin, Jinggang; Liu, Bo; Liu, Peihang; Wu, Yu; Nijhuis, Arend; Zhou, Chao; Li, Chenshan; Hao, Qingbin; Liu, Sheng

2018-01-01

The critical current degradation of Bi-2212 Ag-sheathed round wire subjected to uniaxial strain was studied at 4.2 K in 14 T background field. The strains applied on the sample are both tension and compression. The additional tensile strain caused by the difference in thermal expansion between the

Characterization of a mouse-adapted Staphylococcus aureus strain.

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Silva Holtfreter

Full Text Available More effective antibiotics and a protective vaccine are desperately needed to combat the 'superbug' Staphylococcus aureus. While in vivo pathogenicity studies routinely involve infection of mice with human S. aureus isolates, recent genetic studies have demonstrated that S. aureus lineages are largely host-specific. The use of such animal-adapted S. aureus strains may therefore be a promising approach for developing more clinically relevant animal infection models. We have isolated a mouse-adapted S. aureus strain (JSNZ which caused a severe outbreak of preputial gland abscesses among male C57BL/6J mice. We aimed to extensively characterize this strain on a genomic level and determine its virulence potential in murine colonization and infection models. JSNZ belongs to the MLST type ST88, rare among human isolates, and lacks an hlb-converting phage encoding human-specific immune evasion factors. Naive mice were found to be more susceptible to nasal and gastrointestinal colonization with JSNZ than with the human-derived Newman strain. Furthermore, naïve mice required antibiotic pre-treatment to become colonized with Newman. In contrast, JSNZ was able to colonize mice in the absence of antibiotic treatment suggesting that this strain can compete with the natural flora for space and nutrients. In a renal abscess model, JSNZ caused more severe disease than Newman with greater weight loss and bacterial burden. In contrast to most other clinical isolates, JSNZ can also be readily genetically modified by phage transduction and electroporation. In conclusion, the mouse-adapted strain JSNZ may represent a valuable tool for studying aspects of mucosal colonization and for screening novel vaccines and therapies directed at preventing colonization.

Comparative genomics of transport proteins in probiotic and pathogenic Escherichia coli and Salmonella enterica strains.

Science.gov (United States)

Do, Jimmy; Zafar, Hassan; Saier, Milton H

2017-06-01

Escherichia coli is a genetically diverse species that can be pathogenic, probiotic, commensal, or a harmless laboratory strain. Pathogenic strains of E. coli cause urinary tract infections, diarrhea, hemorrhagic colitis, and pyelonephritis, while the two known probiotic E. coli strains combat inflammatory bowel disease and play a role in immunomodulation. Salmonella enterica, a close relative of E. coli, includes two important pathogenic serovars, Typhi and Typhimurium, causing typhoid fever and enterocolitis in humans, respectively, with the latter strain also causing a lethal typhoid fever-like disease in mice. In this study, we identify the transport systems and their substrates within seven E. coli strains: two probiotic strains, two extracellular pathogens, two intracellular pathogens, and K-12, as well as the two intracellular pathogenic S. enterica strains noted above. Transport systems characteristic of each probiotic or pathogenic species were thus identified, and the tabulated results obtained with all of these strains were compared. We found that the probiotic and pathogenic strains generally contain more iron-siderophore and sugar transporters than E. coli K-12. Pathogens have increased numbers of pore-forming toxins, protein secretion systems, decarboxylation-driven Na + exporters, electron flow-driven monovalent cation exporters, and putative transporters of unknown function compared to the probiotic strains. Both pathogens and probiotic strains encode metabolite transporters that reflect their intracellular versus extracellular environments. The results indicate that the probiotic strains live extracellularly. It seems that relatively few virulence factors can convert a beneficial or commensal microorganism into a pathogen. Taken together, the results reveal the distinguishing features of these strains and provide a starting point for future engineering of beneficial enteric bacteria. Copyright © 2017 Elsevier Ltd. All rights reserved.

Induction apparatus monitoring structural strains in liquid-metal-cooled nuclear reactor

International Nuclear Information System (INIS)

Dean, S.A.; Evans, R.A.

1981-01-01

An improved method of monitoring induced torsional and linear strains in the internal structures of liquid metal cooled nuclear reactors is described. An electrical induction apparatus indicates the variation of magnetic coupling caused by a ferromagnetic member of the apparatus being subjected to such strains. (U.K.)

Foliar Application of Extract from an Azalomycin-Producing Streptomyces malaysiensis Strain MJM1968 Suppresses Yam Anthracnose Caused by Colletotrichum gloeosporioides.

Science.gov (United States)

Arunachalam Palaniyandi, Sasikumar; Yang, Seung Hwan; Suh, Joo-Woh

2016-06-28

Yam anthracnose caused by Colletotrichum gloeosporioides (C.g) is the most devastating disease of yam (Dioscorea sp.). In the present study, we evaluated the culture filtrate extract (CFE) of azalomycin-producing Streptomyces malaysiensis strain MJM1968 for the control of yam anthracnose. MJM1968 showed strong antagonistic activity against C.g in vitro. Furthermore, the MJM1968 CFE was tested for inhibition of spore germination in C.g, where it completely inhibited spore germination at a concentration of 50 μg/ml. To assess the in planta efficacy of the CFE and spores of MJM1968 against C.g, a detached leaf bioassay was conducted, which showed both the treatments suppressed anthracnose development on detached yam leaves. Furthermore, a greenhouse study was conducted to evaluate the CFE from MJM1968 as a fungicide for the control of yam anthracnose. The CFE non-treated plants showed a disease severity of >92% after 90 days of artificial inoculation with C.g, whereas the disease severity of CFE-treated and benomyl-treated yam plants was reduced to 26% and 15%, respectively, after 90 days. Analysis of the yam tubers from the CFE-treated and non-treated groups showed that tubers from the CFE-treated plants were larger than that of non-treated plants, which produced abnormal smaller tubers typical of anthracnose. This study demonstrated the utility of the CFE from S. malaysiensis strain MJM1968 as a biofungicide for the control of yam anthracnose.

Highly sensitive strain sensors based on fragmentized carbon nanotube/polydimethylsiloxane composites

Science.gov (United States)

Gao, Yang; Fang, Xiaoliang; Tan, Jianping; Lu, Ting; Pan, Likun; Xuan, Fuzhen

2018-06-01

Wearable strain sensors based on nanomaterial/elastomer composites have potential applications in flexible electronic skin, human motion detection, human–machine interfaces, etc. In this research, a type of high performance strain sensors has been developed using fragmentized carbon nanotube/polydimethylsiloxane (CNT/PDMS) composites. The CNT/PDMS composites were ground into fragments, and a liquid-induced densification method was used to fabricate the strain sensors. The strain sensors showed high sensitivity with gauge factors (GFs) larger than 200 and a broad strain detection range up to 80%, much higher than those strain sensors based on unfragmentized CNT/PDMS composites (GF composite particles during mechanical deformation, which causes significant resistance change in the strain sensors. The strain sensors can differentiate mechanical stimuli and monitor various human body motions, such as bending of the fingers, human breathing, and blood pulsing.

Characterization of Brucella abortus mutant strain Δ22915, a potential vaccine candidate.

Science.gov (United States)

Bao, Yanqing; Tian, Mingxing; Li, Peng; Liu, Jiameng; Ding, Chan; Yu, Shengqing

2017-04-04

Brucellosis, caused by Brucella spp., is an important zoonosis worldwide. Vaccination is an effective strategy for protection against Brucella infection in livestock in developing countries and in wildlife in developed countries. However, current vaccine strains including S19 and RB51 are pathogenic to humans and pregnant animals, limiting their use. In this study, we constructed the Brucella abortus (B. abortus) S2308 mutant strain Δ22915, in which the putative lytic transglycosylase gene BAB_RS22915 was deleted. The biological properties of mutant strain Δ22915 were characterized and protection of mice against virulent S2308 challenge was evaluated. The mutant strain Δ22915 showed reduced survival within RAW264.7 cells and survival in vivo in mice. In addition, the mutant strain Δ22915 failed to escape fusion with lysosomes within host cells, and caused no observable pathological damage. RNA-seq analysis indicated that four genes associated with amino acid/nucleotide transport and metabolism were significantly upregulated in mutant strain Δ22915. Furthermore, inoculation of ∆22915 at 10 5 colony forming units induced effective host immune responses and long-term protection of BALB/c mice. Therefore, mutant strain ∆22915 could be used as a novel vaccine candidate in the future to protect animals against B. abortus infection.

Reduction of residual strains in weldments in explosion welding

International Nuclear Information System (INIS)

Tsemakhovich, B.D.

1985-01-01

Peculiarities of large-size item strains in explosion cladding have been investigated. Causes of explosion-welded item destruction under the action of reflected waves have been established by means of the acoustic analysis of the item-support system. It is suggested to use grit as a support which permits to decrease 3-4 times and stabilize residual strains and reduce to minimum the probability of crack appearance

Dynamic strain measurements in a sliding microstructured contact

International Nuclear Information System (INIS)

Bennewitz, Roland; David, Jonathan; Lannoy, Charles-Francois de; Drevniok, Benedict; Hubbard-Davis, Paris; Miura, Takashi; Trichtchenko, Olga

2008-01-01

A novel experiment is described which measures the tangential strain development across the contact between a PDMS (polydimethylsiloxane) block and a glass surface during the initial stages of sliding. The surface of the PDMS block has been microfabricated to take the form of a regular array of pyramidal tips at 20 μm separation. Tangential strain is measured by means of light scattering from the interface between the block and surface. Three phases are observed in all experiments: initial shear deformation of the whole PDMS block, a pre-sliding tangential compression of the tip array with stepwise increase of the compressive strain, and sliding in stick-slip movements as revealed by periodic variation of the strain. The stick-slip sliding between the regular tip array and the randomly rough counter surface always takes on the periodicity of the tip array. The fast slip can cause either a sudden increase or a sudden decrease in compressive strain

Flinders Island spotted fever rickettsioses caused by "marmionii" strain of Rickettsia honei, Eastern Australia.

Science.gov (United States)

Unsworth, Nathan B; Stenos, John; Graves, Stephen R; Faa, Antony G; Cox, G Erika; Dyer, John R; Boutlis, Craig S; Lane, Amanda M; Shaw, Matthew D; Robson, Jennifer; Nissen, Michael D

2007-04-01

Australia has 4 rickettsial diseases: murine typhus, Queensland tick typhus, Flinders Island spotted fever, and scrub typhus. We describe 7 cases of a rickettsiosis with an acute onset and symptoms of fever (100%), headache (71%), arthralgia (43%), myalgia (43%), cough (43%), maculopapular/petechial rash (43%), nausea (29%), pharyngitis (29%), lymphadenopathy (29%), and eschar (29%). Cases were most prevalent in autumn and from eastern Australia, including Queensland, Tasmania, and South Australia. One patient had a history of tick bite (Haemaphysalis novaeguineae). An isolate shared 99.2%, 99.8%, 99.8%, 99.9%, and 100% homology with the 17 kDa, ompA, gltA, 16S rRNA, and Sca4 genes, respectively, of Rickettsia honei. This Australian rickettsiosis has similar symptoms to Flinders Island spotted fever, and the strain is genetically related to R. honei. It has been designated the "marmionii" strain of R. honei, in honor of Australian physician and scientist Barrie Marmion.

Evaluation of local strain in Si using UV-Raman spectroscopy

Energy Technology Data Exchange (ETDEWEB)

Ogura, Atsushi [School of Science and Technology, Meiji University, 1-1-1 Higashimita, Tama-ku, Kawasaki, Kanagawa 214-8571 (Japan)], E-mail: a_ogura@isc.meiji.ac.jp; Kosemura, Daisuke; Takei, Munehisa [School of Science and Technology, Meiji University, 1-1-1 Higashimita, Tama-ku, Kawasaki, Kanagawa 214-8571 (Japan); Uchida, Hidetsugu; Hattori, Nobuyoshi [Semiconductor Technology Academic Research Center, 3-17-2 Shinyokohama, Kouhoku-ku, Yokohama 220-0033 (Japan); Yoshimaru, Masaki [Semiconductor Business Group, Sony Corporation, Atsugi Tec., 4-14-1 Asahi-cho, Atsugi-shi, Kanagawa 243-0014 (Japan); Mayuzumi, Satoru [School of Science and Technology, Meiji University, 1-1-1 Higashimita, Tama-ku, Kawasaki, Kanagawa 214-8571 (Japan); Semiconductor Business Group, Sony Corporation, Atsugi Tec., 4-14-1 Asahi-cho, Atsugi-shi, Kanagawa 243-0014 (Japan); Wakabayashi, Hitoshi [Semiconductor Business Group, Sony Corporation, Atsugi Tec., 4-14-1 Asahi-cho, Atsugi-shi, Kanagawa 243-0014 (Japan)

2009-03-15

'Strained-Si', in which intentional strain is introduced in Si crystal to improve carrier mobility by using a modulated band structure, is recognized as one of the most important technologies in post-scaling-generation LSIs. Strain-evaluation technology to probe strain in shallow surfaces that correspond to the channels of MOSFETs is crucial to achieving strained-Si technology. In this paper, we introduce the results we obtained by evaluating strain with the new UV-Raman spectroscopy we developed. Quasi-line shape illumination enabled Raman measurements with 200-nm intervals on the sample. The local-strain mechanism caused by SiN stressors covering a MOSFET was clarified by measuring one-dimensional strain profiles induced by patterned SiN film on Si. We also demonstrated that the induced strain was proportional to the inner stresses of SiN film and that it is more effective to introduce strain in SOI substrates than in bulk substrates. In the evaluation of a actual device fabricated by using the gate-last process in which strain was significantly enhanced after the dummy gate was removed, the size effect, i.e., an increase in induced strain with a decrease in gate length, was confirmed through one-dimensional strain-profile measurements with various gate lengths.

Evaluation of local strain in Si using UV-Raman spectroscopy

International Nuclear Information System (INIS)

Ogura, Atsushi; Kosemura, Daisuke; Takei, Munehisa; Uchida, Hidetsugu; Hattori, Nobuyoshi; Yoshimaru, Masaki; Mayuzumi, Satoru; Wakabayashi, Hitoshi

2009-01-01

'Strained-Si', in which intentional strain is introduced in Si crystal to improve carrier mobility by using a modulated band structure, is recognized as one of the most important technologies in post-scaling-generation LSIs. Strain-evaluation technology to probe strain in shallow surfaces that correspond to the channels of MOSFETs is crucial to achieving strained-Si technology. In this paper, we introduce the results we obtained by evaluating strain with the new UV-Raman spectroscopy we developed. Quasi-line shape illumination enabled Raman measurements with 200-nm intervals on the sample. The local-strain mechanism caused by SiN stressors covering a MOSFET was clarified by measuring one-dimensional strain profiles induced by patterned SiN film on Si. We also demonstrated that the induced strain was proportional to the inner stresses of SiN film and that it is more effective to introduce strain in SOI substrates than in bulk substrates. In the evaluation of a actual device fabricated by using the gate-last process in which strain was significantly enhanced after the dummy gate was removed, the size effect, i.e., an increase in induced strain with a decrease in gate length, was confirmed through one-dimensional strain-profile measurements with various gate lengths.

Respiration shutoff in Escherichia coli K12 strains is induced by far ultraviolet radiations and by mitomycin C

International Nuclear Information System (INIS)

Swenson, P.A.; Norton, I.L.

1984-01-01

Near ultraviolet radiations (UV) cause respiration to shutoff in Escherichia coli B/r. It has been reported that E. coli K12 strains do not shut off respiration after UV. It is also reported that mitomycin C did not cause this 'SOS' response. In this paper it is reported that higher UV fluences than were previously used will cause respiration shutoff in K12 strain W3110 and that cyclic AMP increases the sensitivity of respiration shutoff of irradiated cell suspensions. Also mitomycin C shuts off respiration in this strain. Neither UV nor mitomycin C causes respiration shutoff in the recA56 derivative of W3110. Thus respiration shutoff is a recA dependent response to UV and mitomycin C in E. coli K12 strains. (Auth.)

Comminution of solids caused by kinetic energy of high shear strain rate, with implications for impact, shock, and shale fracturing.

Science.gov (United States)

Bazant, Zdenek P; Caner, Ferhun C

2013-11-26

Although there exists a vast literature on the dynamic comminution or fragmentation of rocks, concrete, metals, and ceramics, none of the known models suffices for macroscopic dynamic finite element analysis. This paper outlines the basic idea of the macroscopic model. Unlike static fracture, in which the driving force is the release of strain energy, here the essential idea is that the driving force of comminution under high-rate compression is the release of the local kinetic energy of shear strain rate. The density of this energy at strain rates >1,000/s is found to exceed the maximum possible strain energy density by orders of magnitude, making the strain energy irrelevant. It is shown that particle size is proportional to the -2/3 power of the shear strain rate and the 2/3 power of the interface fracture energy or interface shear stress, and that the comminution process is macroscopically equivalent to an apparent shear viscosity that is proportional (at constant interface stress) to the -1/3 power of this rate. A dimensionless indicator of the comminution intensity is formulated. The theory was inspired by noting that the local kinetic energy of shear strain rate plays a role analogous to the local kinetic energy of eddies in turbulent flow.

Distribution of strain type and antimicrobial susceptibility of Escherichia coli isolates causing meningitis in a large urban setting in Brazil.

Science.gov (United States)

Berman, Hillary; Barberino, Maria Goreth; Moreira, Edson Duarte; Riley, Lee; Reis, Joice N

2014-05-01

The clinical management of meningitis caused by Escherichia coli is greatly complicated when the organism becomes resistant to broad-spectrum antibiotics. We sought to characterize the antimicrobial susceptibilities, sequence types (ST), and presence of known drug resistance genes of E. coli isolates that caused meningitis between 1996 and 2011 in Salvador, Brazil. We then compared these findings to those for E. coli isolates from community-acquired urinary tract infections (UTI) that occurred during the same time period and in the same city. We found that 19% of E. coli isolates from cases of meningitis and less than 1% of isolates from UTI were resistant to third-generation cephalosporins. The sequence types of E. coli isolates from cases of meningitis included ST131, ST69, ST405, and ST62, which were also found among isolates from UTI. Additionally, among the E. coli isolates that were resistant to third-generation cephalosporins, we found genes that encode the extended-spectrum beta-lactamases CTX-M-2, CTX-M-14, and CTX-M-15. These observations demonstrate that compared to E. coli strains isolated from cases of community-acquired UTI, those isolated from cases of meningitis are more resistant to third-generation cephalosporins, even though the same sequence types are shared between the two forms of extraintestinal infections.

Establishment of a nanoparticle-assisted RT-PCR assay to distinguish field strains and attenuated strains of porcine epidemic diarrhea virus.

Science.gov (United States)

Zhu, Yu; Wang, Gui-Hua; Cui, Yu-Dong; Cui, Shang-Jin

2016-09-01

Porcine epidemic diarrhea virus (PEDV) can cause serious disease and even death in neonatal piglets, resulting in serious damage to the swine industry worldwide. Open reading frame 3 (ORF3) is the only accessory gene in the PEDV genome. Previous studies have indicated that PEDV vaccine strains have a partial deletion in ORF3. In this study, a nanoparticle-assisted polymerase chain reaction (nanoparticle-assisted RT-PCR) assay targeting the ORF3 of PEDV was developed to distinguish PEDV field strains from attenuated strains by using a specific pair of primers. The PCR products of field strains and attenuated strains were 264 bp and 215 bp in length, respectively. The sensitivity and specificity of this assay were also assessed. The nanoparticle-assisted RT-PCR assay was 10-100 times more sensitive than the conventional RT-PCR assay, with no cross-reactions when amplifying porcine pseudorabies virus (PRV), porcine circovirus type 2 (PCV2), classical swine fever virus (CSFV), porcine parvovirus (PPV), porcine reproductive and respiratory syndrome virus (PRRSV), porcine rotavirus (RV), and porcine transmissible gastroenteritis virus (TGEV). The nanoparticle-assisted RT-PCR assay we describe here can be used to distinguish field strains from vaccine strains of PEDV, and it shows promise for reducing economic loss due to PEDV infection.

Biopesticide activity of sugarcane associated rhizobacteria: Ochrobactrum intermedium strain NH-5 and Stenotrophomonas maltophilia strain NH-300 against red rot under field conditions

Directory of Open Access Journals (Sweden)

Muhammad Nadeem HASSAN

2014-09-01

Full Text Available Colletotrichum falcatum is the major fungal pathogen causing sugarcane red rot. Four antagonistic bacterial strains exhibiting biocontrol activity against this pathogen in greenhouse conditions were characterized for production of different antifungal metabolites and biocontrol determinants to elucidate the mechanism of action involved in their antagonistic activity. The strains were also evaluated under field conditions to assess their biocontrol potential. All the strains produced hydrogen cyanide (HCN, and volatile and diffusible antibiotics. In addition, the Ochrobactrum intermedium strain NH-5 produced siderophores and the broad spectrum antibiotic 2, 4-diacetylphloroglucinol (2,4-DAPG; Pseudomonas sp. NH-203 produced siderophores, and Pseudomonas sp. NH-276 produced protease. Two strains, Ochrobactrum intermedium NH-5 and Stenotrophomonas maltophilia NH-300, exhibited good biocontrol activity, suppressing red rot by 44–52% on two sugarcane varieties, SPF-234 and Co-1148, in field experiments. The strains gave consistent results in three consecutive years and showed potential to be used as biopesticides.

Genomic Comparison among Lethal Invasive Strains of Streptococcus pyogenes Serotype M1

Directory of Open Access Journals (Sweden)

Gabriel R. Fernandes

2017-10-01

Full Text Available Streptococcus pyogenes, also known as group A Streptococcus (GAS, is a human pathogen that causes diverse human diseases including streptococcal toxic shock syndrome (STSS. A GAS outbreak occurred in Brasilia, Brazil, during the second half of the year 2011, causing 26 deaths. Whole genome sequencing was performed using Illumina platform. The sequences were assembled and genes were predicted for comparative analysis with emm type 1 strains: MGAS5005 and M1 GAS. Genomics comparison revealed one of the invasive strains that differ from others isolates and from emm 1 reference genomes. Also, the new invasive strain showed differences in the content of virulence factors compared to other isolated in the same outbreak. The evolution of contemporary GAS strains is strongly associated with horizontal gene transfer. This is the first genomic study of a Streptococcal emm 1 outbreak in Brazil, and revealed the rapid bacterial evolution leading to new clones. The emergence of new invasive strains can be a consequence of the injudicious use of antibiotics in Brazil during the past decades.

Complete genome sequence of Kytococcus sedentarius type strain (strain 541T)

Energy Technology Data Exchange (ETDEWEB)

Sims, David; Brettin, Thomas; Detter, John C.; Han, Cliff; Lapidus, Alla; Copeland, Alex; Glavina Del Rio, Tijana; Nolan, Matt; Chen, Feng; Lucas, Susan; Tice, Hope; Cheng, Jan-Fang; Bruce, David; Goodwin, Lynne; Pitluck, Sam; Ovchinnikova, Galina; Pati, Amrita; Ivanova, Natalia; Mavrommatis, Konstantinos; Chen, Amy; Palaniappan, Krishna; D' haeseleer, Patrick; Chain, Patrick; Bristow, James; Eisen, Jonathan A.; Markowitz, Victor; Hugenholtz, Philip; Schneider, Susanne; Goker, Markus; Pukall, Rudiger; Kyrpides, Nikos C.; Klenk, Hans-Peter

2009-05-20

Kytococcus sedentarius (ZoBell and Upham 1944) Stackebrandt et al. 1995 is the type strain of the species, and is of phylogenetic interest because of its location in the Dermacoccaceae, a poorly studied family within the actinobacterial suborder Micrococcineae. K. sedentarius is known for the production of oligoketide antibiotics as well as for its role as an opportunistic pathogen causing valve endocarditis, hemorrhagic pneumonia, and pitted keratolysis. It is strictly aerobic and can only grow when several amino acids are provided in the medium. The strain described in this report is a free-living, nonmotile, Gram-positive bacterium, originally isolated from a marine environment. Here we describe the features of this organism, together with the complete genome sequence, and annotation. This is the first complete genome sequence of a member of the family Dermacoccaceae and the 2,785,024 bp long single replicon genome with its 2639 protein-coding and 64 RNA genes is part of the Genomic Encyclopedia of Bacteria and Archaea project.

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

Highly sensitive strain sensors based on fragmentized carbon nanotube/polydimethylsiloxane composites.

Science.gov (United States)

Gao, Yang; Fang, Xiaoliang; Tan, Jianping; Lu, Ting; Pan, Likun; Xuan, Fuzhen

2018-06-08

Wearable strain sensors based on nanomaterial/elastomer composites have potential applications in flexible electronic skin, human motion detection, human-machine interfaces, etc. In this research, a type of high performance strain sensors has been developed using fragmentized carbon nanotube/polydimethylsiloxane (CNT/PDMS) composites. The CNT/PDMS composites were ground into fragments, and a liquid-induced densification method was used to fabricate the strain sensors. The strain sensors showed high sensitivity with gauge factors (GFs) larger than 200 and a broad strain detection range up to 80%, much higher than those strain sensors based on unfragmentized CNT/PDMS composites (GF sensors is ascribed to the sliding of individual fragmentized-CNT/PDMS-composite particles during mechanical deformation, which causes significant resistance change in the strain sensors. The strain sensors can differentiate mechanical stimuli and monitor various human body motions, such as bending of the fingers, human breathing, and blood pulsing.

Necrotizing Fasciitis Caused by Hypermucoviscous Klebsiella pneumoniae in a Filipino Female in North America

Directory of Open Access Journals (Sweden)

Ng, Daniel

2014-12-01

Full Text Available Necrotizing fasciitis caused by Klebsiella pneumoniae has been described in Southeast Asia, but has only recently begun to emerge in North America. The hypermucoviscous strain of K. pneumoniae is a particularly virulent strain known to cause devastatingly invasive infections, including necrotizing fasciitis. Here we present the first known case of necrotizing fasciitis caused by hypermucoviscous K. pneumoniae in North America. [West J Emerg Med. 2015;16(1:165–168.

Abortive Intestinal Infection With an Escherichia coli-Shigella flexneri Hybrid Strain

Science.gov (United States)

Formal, Samuel B.; LaBrec, E. H.; Kent, T. H.; Falkow, S.

1965-01-01

Formal, Samuel B., (Walter Reed Army Institute of Research, Washington, D.C.), E. H. LaBrec, T. H. Kent, and S. Falkow. Abortive intestinal infection with an Escherichia coli-Shigella flexneri hybrid strain. J. Bacteriol. 89:1374–1382. 1965.—The mechanism of the apparent loss of virulence of an Escherichia coli-Shigella flexneri hybrid strain was studied. The parent Shigella strain caused a fatal enteric infection when fed to starved guinea pigs, and signs of dysentery followed its oral administration to monkeys. The hybrid strain failed to produce any apparent symptoms when fed to either of these species. The parent strain was shown to invade the intestinal mucosa of starved guinea pigs. This caused a severe inflammatory reaction in the lamina propria, which progressed to ulceration of the intestinal epithelium and resulted in death of the animal. The hybrid strain also invaded the intestinal mucosa and produced an inflammatory reaction. In this case, the inflammatory reaction subsided, the intestine returned to normal within 4 days after challenge, and the animal survived. Both fluorescent-antibody techniques and in vivo growth studies have shown that the hybrid strain can not maintain itself in the intestinal mucosa. Preliminary studies have indicated that a similar situation also exists in the monkey. It is concluded that the virulence of dysentery bacilli rests not only in the capacity to reach the lamina propria, but also in the ability to multiply in this region. Images PMID:14293011

Determining the cause of recurrent Clostridium difficile infection using whole genome sequencing.

Science.gov (United States)

Sim, James Heng Chiak; Truong, Cynthia; Minot, Samuel S; Greenfield, Nick; Budvytiene, Indre; Lohith, Akshar; Anikst, Victoria; Pourmand, Nader; Banaei, Niaz

2017-01-01

Understanding the contribution of relapse and reinfection to recurrent Clostridium difficile infection (CDI) has implications for therapy and infection prevention, respectively. We used whole genome sequencing to determine the relation of C. difficile strains isolated from patients with recurrent CDI at an academic medical center in the United States. Thirty-five toxigenic C. difficile isolates from 16 patients with 19 recurrent CDI episodes with median time of 53.5days (range, 13-362) between episodes were whole genome sequenced on the Illumina MiSeq platform. In 84% (16) of recurrences, the cause of recurrence was relapse with prior strain of C. difficile. In 16% (3) of recurrent episodes, reinfection with a new strain of C. difficile was the cause. In conclusion, the majority of CDI recurrences at our institution were due to infection with the same strain rather than infection with a new strain. Copyright © 2016 Elsevier Inc. All rights reserved.

Effects of static strain aging on residual stress stability and alternating bending strength of shot peened AISI 4140

Energy Technology Data Exchange (ETDEWEB)

Menig, R.; Schulze, V.; Voehringer, O. [Inst. fuer Werkstoffkunde 1, Univ. Karlsruhe (TH), Karlsruhe (Germany)

2002-07-01

Increases of residual stress stability and alternating bending strength of shot peened AISI 4140 are obtained by successive annealing treatments. This is caused by static strain aging effects, which lead to pinning of dislocations by carbon atoms and very small carbides. It will be shown that by well directed annealing of a quenched and tempered AISI 4140 it is possible to maximize the positive effects of static strain aging, without causing extended thermal residual stress relaxation. The amount of yield stress increases caused by static strain aging is quantified using tensile tests. Static strain aging is also found to be responsible for an increase of the quasi static and cyclic surface yield strength present after shot peening. (orig.)

Characterization of a new pathovar of Agrobacterium vitis causing banana leaf blight in China.

Science.gov (United States)

Huang, Siliang; Long, Mengling; Fu, Gang; Lin, Shanhai; Qin, Liping; Hu, Chunjin; Cen, Zhenlu; Lu, Jie; Li, Qiqin

2015-01-01

A new banana leaf blight was found in Nanning city, China, during a 7-year survey (2003-2009) of the bacterial diseases on banana plants. Eight bacterial strains were isolated from affected banana leaves, and identified as an intraspecific taxon of Agrobacterium vitis based on their 16S rDNA sequence similarities with those of 37 randomly selected bacterial strains registered in GenBank database. The representative strain Ag-1 was virulent on banana leaves and shared similar growth and biochemical reactions with the reference strain IAM14140 of A. vitis. The strains causing banana leaf blight were denominated as A. vitis pv. musae. The traditional A. vitis strains virulent to grapevines were proposed to be revised as A. vitis pv. vitis. This is the first record of a new type of A. vitis causing banana leaf blight in China. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

[A prophylactic program for strain urinary incontinence].

Science.gov (United States)

Stadnicka, Grazyna; Iwanowicz-Palus, Grazyna J; Bień, Agnieszka M

2002-01-01

The aim of the study was to work out a prophylactic program for strain urinary incontinence. Analysis of literature on the subject and results of own investigations presented in the first part of the paper indicate that the program of prophylaxis of strain urinary incontinence should primarily include: (1) Preparation of the medical staff (nurses, midwives) for propagating health education among women on prevention of strain urinary incontinence. (2) Preparation of adequate educational materials in the form of brochures, leaflets, information posters about symptoms, causes and prophylaxis of urinary incontinence indicating health care institutions available to all women when the disease is suspected or already present. (3) Propagation of problems connected with strain urinary incontinence in the mass media providing information to a wide audience in order to make people realize the significance of this social problem and break stereotypes associated with this disease of "shame". (4) Preparation of sets of exercises for the muscles of the base of the pelvis to be performed during pregnancy, confinement and menopause to maintain their proper function. (5) Indicating factors predisposing to strain urinary incontinence with focus on possibilities of their reduction or elimination.

Strain features and condition assessment of orthotropic steel deck cable-supported bridges subjected to vehicle loads by using dense FBG strain sensors

Science.gov (United States)

Wei, Shiyin; Zhang, Zhaohui; Li, Shunlong; Li, Hui

2017-10-01

Strain is a direct indicator of structural safety. Therefore, strain sensors have been used in most structural health monitoring systems for bridges. However, until now, the investigation of strain response has been insufficient. This paper conducts a comprehensive study of the strain features of the U ribs and transverse diaphragm on an orthotropic steel deck and proposes a statistical paradigm for crack detection based on the features of vehicle-induced strain response by using the densely distributed optic fibre Bragg grating (FBG) strain sensors. The local feature of strain under vehicle load is highlighted, which enables the use of measurement data to determine the vehicle loading event and to make a decision regarding the health status of a girder near the strain sensors via technical elimination of the load information. Time-frequency analysis shows that the strain contains three features: the long-term trend item, the short-term trend item, and the instantaneous vehicle-induced item (IVII). The IVII is the wheel-induced strain with a remarkable local feature, and the measured wheel-induced strain is only influenced by the vehicle near the FBG sensor, while other vehicles slightly farther away have no effect on the wheel-induced strain. This causes the local strain series, among the FBG strain sensors in the same transverse locations of different cross-sections, to present similarities in shape to some extent and presents a time delay in successive order along the driving direction. Therefore, the strain series induced by an identical vehicle can be easily tracked and compared by extracting the amplitude and calculating the mutual ratio to eliminate vehicle loading information, leaving the girder information alone. The statistical paradigm for crack detection is finally proposed, and the detection accuracy is then validated by using dense FBG strain sensors on a long-span suspension bridge in China.

Identification of resistance and virulence factors in an epidemic Enterobacter hormaechei outbreak strain

NARCIS (Netherlands)

Paauw, A.; Caspers, M.P.M.; Leverstein-van Hall, M.A.; Schuren, F.H.J.; Montijn, R.C.; Verhoef, J.; Fluit, A.C.

2009-01-01

Bacterial strains differ in their ability to cause hospital outbreaks. Using comparative genomic hybridization, Enterobacter cloacae complex isolates were studied to identify genetic markers specific for Enterobacter cloacae complex outbreak strains. No outbreak-specific genes were found that were

Incorrect strain information for mouse cell lines: sequential influence of misidentification on sublines.

Science.gov (United States)

Uchio-Yamada, Kozue; Kasai, Fumio; Ozawa, Midori; Kohara, Arihiro

2017-03-01

Misidentification or cross-contamination of cell lines can cause serious issues. Human cell lines have been authenticated by short tandem repeat profiling; however, mouse cell lines have not been adequately assessed. In this study, mouse cell lines registered with the JCRB cell bank were examined by simple sequence length polymorphism (SSLP) analysis to identify their strains. Based on comparisons with 7 major inbred strains, our results revealed their strains in 80 of 90 cell lines. However, 12 of the 80 cell lines (15%) were found to differ from registered information. Of them, 4 cell lines originated from the same mouse, which had been generated through mating between two different inbred strains. The genotype of the mouse sample had not been examined after the backcross, leading to strain misidentification in those cell lines. Although 8 other cell lines had been established as sublines of a BALB/c cell line, their SSLP profiles are similar to a Swiss cell line. This affects differences in genotypes between inbred and outbred strains. Because the use of inbred samples and interbreeding between strains are not involved in human materials, our results suggest that the cause and influence of misidentification in mouse cell lines are different from those in human.

Incorporation of Listeria monocytogenes strains in raw milk biofilms.

Science.gov (United States)

Weiler, Christiane; Ifland, Andrea; Naumann, Annette; Kleta, Sylvia; Noll, Matthias

2013-02-01

Biofilms develop successively on devices of milk production without sufficient cleaning and originate from the microbial community of raw milk. The established biofilm matrices enable incorporation of pathogens like Listeria monocytogenes, which can cause a continuous contamination of food processing plants. L. monocytogenes is frequently found in raw milk and non-pasteurized raw milk products and as part of a biofilm community in milk meters and bulk milk tanks. The aim of this study was to analyze whether different L. monocytogenes strains are interacting with the microbial community of raw milk in terms of biofilm formation in the same manner, and to identify at which stage of biofilm formation a selected L. monocytogenes strain settles best. Bacterial community structure and composition of biofilms were analyzed by a cloning and sequencing approach and terminal restriction fragment length polymorphism analysis (T-RFLP) based on the bacterial 16S rRNA gene. The chemical composition of biofilms was analyzed by Fourier transform infrared spectroscopy (FTIR), while settled L. monocytogenes cells were quantified by fluorescence in situ hybridization (FISH). Addition of individual L. monocytogenes strains to raw milk caused significant shifts in the biofilm biomass, in the chemical as well as in the bacterial community composition. Biofilm formation and attachment of L. monocytogenes cells were not serotype but strain specific. However, the added L. monocytogenes strains were not abundant since mainly members of the genera Citrobacter and Lactococcus dominated the bacterial biofilm community. Overall, added L. monocytogenes strains led to a highly competitive interaction with the raw milk community and triggered alterations in biofilm formation. Copyright © 2012 Elsevier B.V. All rights reserved.

Strain rate effects on reinforcing steels in tension

Science.gov (United States)

Cadoni, Ezio; Forni, Daniele

2015-09-01

It is unquestionable the fact that a structural system should be able to fulfil the function for which it was created, without being damaged to an extent disproportionate to the cause of damage. In addition, it is an undeniable fact that in reinforced concrete structures under severe dynamic loadings, both concrete and reinforcing bars are subjected to high strain-rates. Although the behavior of the reinforcing steel under high strain rates is of capital importance in the structural assessment under the abovementioned conditions, only the behaviour of concrete has been widely studied. Due to this lack of data on the reinforcing steel under high strain rates, an experimental program on rebar reinforcing steels under high strain rates in tension is running at the DynaMat Laboratory. In this paper a comparison of the behaviour in a wide range of strain-rates of several types of reinforcing steel in tension is presented. Three reinforcing steels, commonly proposed by the European Standards, are compared: B500A, B500B and B500C. Lastly, an evaluation of the most common constitutive laws is performed.

Susceptibilities of Mycoplasma bovis, Mycoplasma dispar, and Ureaplasma diversum strains to antimicrobial agents in vitro.

OpenAIRE

ter Laak, E A; Noordergraaf, J H; Verschure, M H

1993-01-01

The purpose of this study was to determine the susceptibility of various strains of Mycoplasma bovis, Mycoplasma dispar, and Ureaplasma diversum, which are prevalent causes of pneumonia in calves, to 16 antimicrobial agents in vitro. The MICs of the antimicrobial agents were determined by a serial broth dilution method for 16 field strains and the type strain of M. bovis, for 19 field strains and the type strain of M. dispar, and for 17 field strains of U. diversum. Final MICs for M. bovis an...

Sepsis puerperalis caused by a genotypically proven cat-derived Pasteurella multocida strain.

Science.gov (United States)

Voss, A; van Zwam, Y H; Meis, J F; Melchers, W; Steegers, E A

1998-01-01

We report a disseminated intrauterine Pasteurella multocida infection in a puerperal woman who could not remember any traumatic exposure to her cat. An oral swab taken from the cat, just 2 days after the patient's admission, grew Pasteurella multocida, with an PCR-fingerprinting pattern identical to the patient's isolate. Hand-washing after every contact with cats and dogs and if feasible separation of in-house pets from mother and infant should be applied to prevent this uncommon but serious occurrence of post-partum infections. To our knowledge this is the first case of Pasteurella multocida 'child-bed fever', with a genotypically identical strain isolated from the in-house cat.

A piezoelectric transducer for measurement of dynamic strain in pipes

Energy Technology Data Exchange (ETDEWEB)

Lannes, Daniel P.; Gama, Antonio L. [Universidade Federal Fluminense (UFF), Niteroi, RJ (Brazil). Dept. de Engenharia Mecanica

2009-07-01

This work presents a new strain transducer developed mainly for the inspection and evaluation of piping systems with excessive vibration. Vibration is one of the most common causes of piping failures. These failures could be avoided if the vibration problems were identified and quickly evaluated. Procedures for evaluation of piping vibration are usually based on pipe velocity or displacement. Although simple and fast, these procedures do not provide precise information on the risk of piping fatigue failure. Through the measurement of pipe dynamic strains the risk of failure due to vibration can be determined more accurately. The measurement of strain is usually performed using the conventional strain gauge method. Although efficient and accurate, the implementation of the conventional strain gauge technique may become a difficult task in certain industrial scenarios. Motivated by the need of a simple and rapid method for pipe dynamic strain measurement, a piezoelectric dynamic strain transducer was developed. This work presents a description of the piezoelectric strain transducer and the preliminary results of pipe strain measurements. The transducer can be applied directly to the pipe through magnetic bases allowing for the quick measurement of the dynamic strains in many points of the pipe. The transducer signal can be read with the same commercial data collectors used for accelerometers. (author)

Temperature affects the morphology and calcification of Emiliania huxleyi strains

Science.gov (United States)

Rosas-Navarro, Anaid; Langer, Gerald; Ziveri, Patrizia

2016-05-01

The global warming debate has sparked an unprecedented interest in temperature effects on coccolithophores. The calcification response to temperature changes reported in the literature, however, is ambiguous. The two main sources of this ambiguity are putatively differences in experimental setup and strain specificity. In this study we therefore compare three strains isolated in the North Pacific under identical experimental conditions. Three strains of Emiliania huxleyi type A were grown under non-limiting nutrient and light conditions, at 10, 15, 20 and 25 °C. All three strains displayed similar growth rate versus temperature relationships, with an optimum at 20-25 °C. Elemental production (particulate inorganic carbon (PIC), particulate organic carbon (POC), total particulate nitrogen (TPN)), coccolith mass, coccolith size, and width of the tube element cycle were positively correlated with temperature over the sub-optimum to optimum temperature range. The correlation between PIC production and coccolith mass/size supports the notion that coccolith mass can be used as a proxy for PIC production in sediment samples. Increasing PIC production was significantly positively correlated with the percentage of incomplete coccoliths in one strain only. Generally, coccoliths were heavier when PIC production was higher. This shows that incompleteness of coccoliths is not due to time shortage at high PIC production. Sub-optimal growth temperatures lead to an increase in the percentage of malformed coccoliths in a strain-specific fashion. Since in total only six strains have been tested thus far, it is presently difficult to say whether sub-optimal temperature is an important factor causing malformations in the field. The most important parameter in biogeochemical terms, the PIC : POC ratio, shows a minimum at optimum growth temperature in all investigated strains. This clarifies the ambiguous picture featuring in the literature, i.e. discrepancies between PIC : POC

Sequence analysis of measles virus strains collected during the pre- and early-vaccination era in Denmark reveals a considerable diversity of ancient strains

DEFF Research Database (Denmark)

Christensen, Laurids Siig; Schöller, S.; Schierup, M. H.

2002-01-01

A total of 199 serum samples from patients with measles collected in Denmark, Greenland and the Faroe Islands from 1964 to 1983 were analysed by PCR. Measles virus (MV) RNA could be detected in 38 (19%) of the samples and a total of 18 strains were subjected to partial sequence analysis of the he......A total of 199 serum samples from patients with measles collected in Denmark, Greenland and the Faroe Islands from 1964 to 1983 were analysed by PCR. Measles virus (MV) RNA could be detected in 38 (19%) of the samples and a total of 18 strains were subjected to partial sequence analysis...... of the hemagglutinin gene. The strains exhibited a considerable genomic diversity, which is at odds with the assumption that one genome type prevailed among globally circulating MV strains prior to the advent of live-attenuated vaccines. Our data indicate that the similarity of the various vaccine strains...... is attributed to their having originated from the same primary isolate. Consequently, it is implied that a small number of clinical manifestations of MV worldwide from which strains similar to the vaccine strain were identified were vaccine related rather than being caused by members of a persistently...

Residual strain sensor using Al-packaged optical fiber and Brillouin optical correlation domain analysis.

Science.gov (United States)

Choi, Bo-Hun; Kwon, Il-Bum

2015-03-09

We propose a distributed residual strain sensor that uses an Al-packaged optical fiber for the first time. The residual strain which causes Brillouin frequency shifts in the optical fiber was measured using Brillouin optical correlation domain analysis with 2 cm spatial resolution. We quantified the Brillouin frequency shifts in the Al-packaged optical fiber by the tensile stress and compared them for a varying number of Al layers in the optical fiber. The Brillouin frequency shift of an optical fiber with one Al layer had a slope of 0.038 MHz/με with respect to tensile stress, which corresponds to 78% of that for an optical fiber without Al layers. After removal of the stress, 87% of the strain remained as residual strain. When different tensile stresses were randomly applied, the strain caused by the highest stress was the only one detected as residual strain. The residual strain was repeatedly measured for a time span of nine months for the purpose of reliability testing, and there was no change in the strain except for a 4% reduction, which is within the error tolerance of the experiment. A composite material plate equipped with our proposed Al-packaged optical fiber sensor was hammered for impact experiment and the residual strain in the plate was successfully detected. We suggest that the Al-packaged optical fiber can be adapted as a distributed strain sensor for smart structures, including aerospace structures.

Nutritional deficiency in citrus with symptoms of citrus variegated chlorosis disease

Directory of Open Access Journals (Sweden)

ME. Silva-Stenico

Full Text Available It is well known that citrus plants that have been infected by Xylella fastidiosa display nutritional deficiencies, probably caused by production of extracellular polymers by the bacteria that block normal nutrient flow through the xylem. The aim of this work was to study the mineral composition of specific foliar areas in different stages of infection in citrus. Thus, the concentrations of macro and micronutrients in leaves of citrus infected by X. fastidiosa were measured. Samples from four infected citrus orchards in the State of São Paulo, Brazil, were respectively collected from Santa Rita do Passa Quatro, Neves Paulista, Gavião Peixoto and Paraíso counties. The presence of X. fastidiosa in leaves was confirmed by polymerase chain reaction (PCR using specific PCR primers. To understand the variation in leaf-nutrient content in citrus plants, we used foliar nutrient values from control (non-symptomatic plants as a reference. Chemometric analysis showed that the deficiency of P and K in symptomatic trees for all orchards and high concentrations of Fe, Mn and Zn were observed in chlorotic areas, although other studies revealed deficiency of zinc in leaves. This is the first report showing that a correlation between chlorotic citrus leaf and higher concentrations of Fe, Mn and Zn are observed when infected and healthy plants were compared.

Management of digital eye strain.

Science.gov (United States)

Coles-Brennan, Chantal; Sulley, Anna; Young, Graeme

2018-05-23

Digital eye strain, an emerging public health issue, is a condition characterised by visual disturbance and/or ocular discomfort related to the use of digital devices and resulting from a range of stresses on the ocular environment. This review aims to provide an overview of the extensive literature on digital eye strain research with particular reference to the clinical management of symptoms. As many as 90 per cent of digital device users experience symptoms of digital eye strain. Many studies suggest that the following factors are associated with digital eye strain: uncorrected refractive error (including presbyopia), accommodative and vergence anomalies, altered blinking pattern (reduced rate and incomplete blinking), excessive exposure to intense light, closer working distance, and smaller font size. Since a symptom may be caused by one or more factors, a holistic approach should be adopted. The following management strategies have been suggested: (i) appropriate correction of refractive error, including astigmatism and presbyopia; (ii) management of vergence anomalies, with the aim of inducing or leaving a small amount of heterophoria (~1.5 Δ Exo); (iii) blinking exercise/training to maintain normal blinking pattern; (iv) use of lubricating eye drops (artificial tears) to help alleviate dry eye-related symptoms; (v) contact lenses with enhanced comfort, particularly at end-of-day and in challenging environments; (vi) prescription of colour filters in all vision correction options, especially blue light-absorbing filters; and (vii) management of accommodative anomalies. Prevention is the main strategy for management of digital eye strain, which involves: (i) ensuring an ergonomic work environment and practice (through patient education and the implementation of ergonomic workplace policies); and (ii) visual examination and eye care to treat visual disorders. Special consideration is needed for people at a high risk of digital eye strain, such as computer

Exploring the Genomic Traits of Non-toxigenic Vibrio parahaemolyticus Strains Isolated in Southern Chile

DEFF Research Database (Denmark)

Castillo Bermúdez, Daniel Elías; Pérez-Reytor, Diliana; Plaza, Nicolás

2018-01-01

Vibrio parahaemolyticus is the leading cause of seafood-borne gastroenteritis worldwide. As reported in other countries, after the rise and fall of the pandemic strain in Chile, other post-pandemic strains have been associated with clinical cases, including strains lacking the major toxins TDH...

Characterizing large strain crush response of redwood

International Nuclear Information System (INIS)

Cramer, S.M.; Hermanson, J.C.

1996-12-01

Containers for the transportation of hazardous and radioactive materials incorporate redwood in impact limiters. Redwood is an excellent energy absorber, but only the most rudimentary information exists on its crush properties. The objectives of the study were to fill the information gap by collecting triaxial load-deformation data for redwood; to use these data to characterize redwood crush, assess current wood failure theories, provide developments toward a complete stress-strain theory for redwood; and to review the literature on strain-rate effects on redwood crush performance. The load-deformation responses of redwood at temperature conditions corresponding to ambient (70 degrees F), 150 degrees F, and -20 degrees F conditions were measured in approximately 100 confined compression tests for crush levels leading to material densification. Data analysis provided a more complete description of redwood crush performance and a basis for assessing proposed general orthotropic stress-strain relationships for redwood. A review of existing literature indicated that strain-rate effects cause at most a 20 percent increase in crush stress parallel to grain

International Clostridium difficile animal strain collection and large diversity of animal associated strains

DEFF Research Database (Denmark)

Janezic, Sandra; Zidaric, Valerija; Pardon, Bart

2014-01-01

Background: Clostridium difficile is an important cause of intestinal infections in some animal species and animals might be a reservoir for community associated human infections. Here we describe a collection of animal associated C. difficile strains from 12 countries based on inclusion criteria...... of one strain (PCR ribotype) per animal species per laboratory. Results: Altogether 112 isolates were collected and distributed into 38 PCR ribotypes with agarose based approach and 50 PCR ribotypes with sequencer based approach. Four PCR ribotypes were most prevalent in terms of number of isolates...... as well as in terms of number of different host species: 078 (14.3% of isolates; 4 hosts), 014/020 (11.6%; 8 hosts); 002 (5.4%; 4 hosts) and 012 (5.4%; 5 hosts). Two animal hosts were best represented; cattle with 31 isolates (20 PCR ribotypes; 7 countries) and pigs with 31 isolates (16 PCR ribotypes; 10...

Estimation of strain from piezoelectric effect and domain switching in morphotropic PZT by combined analysis of macroscopic strain measurements and synchrotron X-ray data

International Nuclear Information System (INIS)

Kungl, Hans; Theissmann, Ralf; Knapp, Michael; Baehtz, Carsten; Fuess, Hartmut; Wagner, Susanne; Fett, Theo; Hoffmann, Michael J.

2007-01-01

Morphotropic PZT ceramics are State of the art materials for ferroelectric actuators. Essential performance parameters for these materials are strain and hysteresis. On a microscopic scale the strain provided by an electric field is due to two different mechanisms. The piezoelectric effect causes an elongation of the unit cells, whereas domain switching changes their crystallographic orientation by aligning the polarization axis towards the field direction. A method is outlined to estimate the contribution of the two mechanisms to total strain by combining macroscopic strain measurements and X-ray diffraction (XRD) data. Results from macroscopic measurements of remanent and unipolar strain with the corresponding data on texture, derived from in situ synchrotron radiation XRD patterns, are analyzed and evaluated by a semi-empirical approach. The method was applied to six morphotropic, LaSr doped PZT materials of different Zr/Ti ratios. Results are discussed with respect to the differences between the materials

Genetic and virulence variability among Erwinia tracheiphila strains recovered from different cucurbit hosts.

Science.gov (United States)

Rojas, E Saalau; Dixon, P M; Batzer, J C; Gleason, M L

2013-09-01

The causal agent of cucurbit bacterial wilt, Erwinia tracheiphila, has a wide host range in the family Cucurbitaceae, including economically important crops such as muskmelon (Cucumis melo), cucumber (C. sativus), and squash (Cucurbita spp.). Genetic variability of 69 E. tracheiphila strains was investigated by repetitive-element polymerase chain reaction (rep-PCR) using BOXA1R and ERIC1-2 primers. Fingerprint profiles revealed significant variability associated with crop host; strains isolated from Cucumis spp. were clearly distinguishable from Cucurbita spp.-isolated strains regardless of geographic origin. Twelve E. tracheiphila strains isolated from muskmelon, cucumber, or summer squash were inoculated onto muskmelon and summer squash seedlings, followed by incubation in a growth chamber. Wilt symptoms were assessed over 3 weeks, strains were reisolated, and rep-PCR profiles were compared with the inoculated strains. Wilting occurred significantly faster when seedlings were inoculated with strains that originated from the same crop host genus (P<0.001). In the first run of the experiment, cucumber and muskmelon strains caused wilting on muskmelon seedlings at a median of 7.8 and 5.6 days after inoculation (dai), respectively. Summer squash seedlings wilted 18.0, 15.7, and 5.7 dai when inoculated with muskmelon-, cucumber-, and squash-origin strains, respectively. In a second run of the experiment, cucumber and muskmelon strains caused wilting on muskmelon at 7.0 and 6.9 dai, respectively, whereas summer squash seedlings wilted at 23.6, 29.0 and 9.0 dai when inoculated with muskmelon-, cucumber-, and squash-origin strains, respectively. Our results provide the first evidence of genetic diversity within E. tracheiphila and suggest that strain specificity is associated with plant host. This advance is a first step toward understanding the genetic and population structure of E. tracheiphila.

Role strain among male RNs in the critical care setting: Perceptions of an unfriendly workplace.

Science.gov (United States)

Carte, Nicholas S; Williams, Collette

2017-12-01

Traditionally, nursing has been a female-dominated profession. Men employed as registered nurses have been in the minority and little is known about the experiences of this demographic. The purpose of this descriptive, quantitative study was to understand the relationship between the variables of demographics and causes of role strain among male nurses in critical care settings. The Sherrod Role Strain Scale assesses role strain within the context of role conflict, role overload, role ambiguity and role incongruity. Data analysis of the results included descriptive and inferential statistics. Inferential statistics involved the use of repeated measures ANOVA testing for significant difference in the causes of role strain between male nurses employed in critical care settings and a post hoc comparison of specific demographic data using multivariate analyses of variance (MANOVAs). Data from 37 male nurses in critical care settings from the northeast of the United States were used to calculate descriptive statistics standard deviation, mean of the data analysis and results of the repeated ANOVA and the post hoc secondary MANOVA analysis. The descriptive data showed that all participants worked full-time. There was an even split from those participants who worked day shift (46%) vs. night shift (43%), most the participants indicated they had 15 years or more experience as an registered nurse (54%). Significant findings of this study include two causes of role strain in male nurses employed in critical care settings which are: role ambiguity and role overload based on ethnicity. Consistent with previous research findings, the results of this study suggest that male registered nurses employed in critical care settings do experience role strain. The two main causes of role strain in male nurses are role ambiguity and role overload. Copyright © 2017. Published by Elsevier Ltd.

Genome sequencing and analysis of BCG vaccine strains.

Directory of Open Access Journals (Sweden)

Wen Zhang

Full Text Available BACKGROUND: Although the Bacillus Calmette-Guérin (BCG vaccine against tuberculosis (TB has been available for more than 75 years, one third of the world's population is still infected with Mycobacterium tuberculosis and approximately 2 million people die of TB every year. To reduce this immense TB burden, a clearer understanding of the functional genes underlying the action of BCG and the development of new vaccines are urgently needed. METHODS AND FINDINGS: Comparative genomic analysis of 19 M. tuberculosis complex strains showed that BCG strains underwent repeated human manipulation, had higher region of deletion rates than those of natural M. tuberculosis strains, and lost several essential components such as T-cell epitopes. A total of 188 BCG strain T-cell epitopes were lost to various degrees. The non-virulent BCG Tokyo strain, which has the largest number of T-cell epitopes (359, lost 124. Here we propose that BCG strain protection variability results from different epitopes. This study is the first to present BCG as a model organism for genetics research. BCG strains have a very well-documented history and now detailed genome information. Genome comparison revealed the selection process of BCG strains under human manipulation (1908-1966. CONCLUSIONS: Our results revealed the cause of BCG vaccine strain protection variability at the genome level and supported the hypothesis that the restoration of lost BCG Tokyo epitopes is a useful future vaccine development strategy. Furthermore, these detailed BCG vaccine genome investigation results will be useful in microbial genetics, microbial engineering and other research fields.

Elliptical posts allow for detailed control of non-equibiaxial straining of cell cultures

DEFF Research Database (Denmark)

Olesen, Christian Gammelgaard; Pennisi, Cristian Pablo; de Zee, Mark