80 research outputs found
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Horizontal gene transfer through natural competence and recombination in the generalist plant pathogen Xylella fastidiosa
Full text linkHorizontal gene transfer has been implicated as a contributing factor towards the diversity and adaptation of pathogens, and the emergence of new diseases. For naturally competent bacteria, DNA acquired through transformation and recombined into the genome could provide a means for this genetic transfer to occur. The work presented here illustrates that Xylella fastidiosa, a bacterial pathogen responsible for several important plant diseases, is naturally competent and able to homologously recombine acquired DNA into its genome. Xylella fastidiosa is vector-transmitted and often exists in natural environments as an endophyte, but causes disease when it multiplies to high levels inside the xylem vessels of its host plants, causing symptoms such as leaf scorching and stunted growth. Several factors were identified that affect the competence of X. fastidiosa, including nutrient availability, growth stage, and methylation state and size of transforming DNA. Recombination efficiencies for X. fastidiosa were at least 1000-fold higher when cells were grown in a defined nutrient medium compared to cells grown in a rich medium. In addition, surface-attached cells transformed and recombined DNA at efficiencies approximately two orders of magnitude higher than their planktonic counterparts. Maximum recombination efficiencies, defined as the number of recombinant cells recovered divided by the total number of cells, were approximately 10-3 when high concentrations of exogenous plasmid DNA were added to cells, and 10-5 when strains harboring different antibiotic markers were co-cultured on solid medium. For planktonic cells, maximum recombination efficiencies were only approximately 10-5 when DNA was added and 10-7 when different strains were co-cultured. Cells appeared most competent while undergoing exponential growth. For planktonic cells, competence peaked after two days of growth and then rapidly declined, with no recombination observed after 8 days. In biofilms, however, cells remained highly competent for at least five days, with recombination events observed even after 21 days of growth. The transformation mechanism in X. fastidiosa is likely similar to that of other naturally competent bacteria, with mutations in type IV pili, competence-related genes (com genes), and cell-cell signaling genes impacting competence. It was also experimentally determined that flanking homologous sequences as short as 96bp in transforming DNA is sufficient to initiate recombination, with efficiencies increasing exponentially with length of the homologous region up to 1kb. In addition, recombination efficiencies decreased exponentially with the size of non-homologous insert. Integration of up to 4kb of non-homologous DNA was observed experimentally. An in silico analysis of genomic sequences confirmed that the experimental data was consistent with events detected in natural populations, with an estimated mean size of recombination events of 1,906 bp. Each recombination event also modified, on average, 1.79% of the nucleotides in the recombined region. Based on sequence similarity of shared coding regions, it appears that recombination between different subspecies of X. fastidiosa could frequently occur. Originally, it was thought that X. fastidiosa was primarily clonal, but recent studies have suggested that recombination plays a significant role in generating genetic diversity in this bacterium. The work presented here illustrates that X. fastidiosa is naturally competent and that DNA acquired through natural transformation could be a substantial source of donor DNA for recombination. Understanding how this process is regulated and what factors affect its efficiency could provide insight into the genetic diversity of this organism and how new diseases emerge
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Dissecting Novel Grapevine-Mealybug-Virus Interactions
Full text linkThe biological mechanisms underlying vector transmission of grapevine leafroll-associated virus 3 (GLRaV-3) remain poorly understood due to limitations of a technically challenging host- pathogen system in Vitis vinifera. GLRaV-3 was able to infect the model organism Nicotiana benthamiana by insect-vector mediated transmission using the vine mealybug, Planococcus ficus. Working with GLRaV-3 infected N. benthamiana revealed distinct advantages in comparison with its natural host Vitis vinifera, yielding both higher viral protein and virion concentrations in western blot and transmission electron microscopy (TEM) observations, respectively. Immunogold labelling of thin sections through N. benthamiana petioles revealed filamentous particles in the phloem cells of GLRaV-3 positive plants. Comparison of assembled whole genomes from GLRaV-3 infected V. vinifera vs. N. benthamiana revealed identical sequences. High throughput sequencing was used to compare host response to GLRaV-3 infection between V. vinifera and N. benthamiana. General families of differentially expressed genes (DEGs) common in both hosts followed similar expression changes with six upregulated, seven downregulated, and two stably expressed genes in common. Overall, both hosts have many DEGs unique to each host as well as responses in common to GLRaV-3 infection. The vine mealybug, Planococcus ficus, fed through a membrane feeding system on GLRaV-3 viral purifications from both V. vinifera and N. benthamiana, and transmitted the virus to test plants.An immunofluorescence approach was used to localize virions to two retention sites in P. ficus mouthparts. Assays testing molecules capable of blocking virus transmission demonstrated that GLRaV-3 transmission by P. ficus can be disrupted. Our results indicate that our membrane feeding system and transmission blocking assays are a valid approach and can be used to screen other candidate blocking molecules. GLRaV-3 continues to impact grape-growing regions worldwide and the lack of knowledge surrounding virus-vector interactions remains limiting to the field. Elucidating the transmission biology of this important virus contributes to the eventual goal of blocking of transmission in insect vectors and the development of improved control strategies in vineyards
Glassy-winged sharpshooter transmission of Xylella fastidiosa to plants
Full text linkXylella fastidiosa is a xylem limited bacterium that causes disease in many plants. Insect dissemination of X. fastidiosa is only possible by xylem-feeding vectors belonging to the subfamily Cicadellinae (Hemiptera, Cicadellidae; sharpshooter leafhoppers) and the family Cercopidae (Hemiptera; spittlebugs). The best studied vector of X. fastidiosa in relation to pathogen transmission is Graphocephala atropunctata Signoret (blue-green sharpshooter). However, in recent years another sharpshooter, Homalodisca coagulata Say (glassy-winged sharpshooter), has been the focus of much interest. This insect has been recently introduced into tropical islands in the Pacific, notably Tahiti and Oahu in Hawaii. Understanding X. fastidiosa transmission parameters will help determine the potential threat of this pest
Exploiting a chitinase to suppress Xylella fastidiosa colonization of plants and insects
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How to test if disrupting Xylella fastidiosa-vector interactions can control disease spread?
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Population structure and adaptation of a bacterial pathogen in California grapevines
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Data from: Museum specimen data reveal emergence of a plant disease may be linked to increases in the insect vector population
No full textThe emergence rate of new plant diseases is increasing due to novel introductions, climate change, and changes in vector populations, posing risks to agricultural sustainability. Assessing and managing future disease risks depends on understanding the causes of contemporary and historical emergence events. Since the mid-1990s, potato growers in the western United States, Mexico, and Central America have experienced severe yield loss from Zebra Chip disease and have responded by increasing insecticide use to suppress populations of the insect vector, the potato psyllid, Bactericera cockerelli (Hemiptera: Triozidae). Despite the severe nature of Zebra Chip outbreaks, the causes of emergence remain unknown. We tested the hypotheses that 1) B. cockerelli occupancy has increased over the last century in California and 2) such increases are related to climate change, specifically warmer winters. We compiled a dataset of 87,000 museum specimen occurrence records across the order Hemiptera collected between 1900 and 2014. We then analyzed changes in B. cockerelli distribution using a hierarchical occupancy model using changes in background species lists to correct for collecting effort. We found evidence that B. cockerelli occupancy has increased over the last century. However, these changes appear to be unrelated to climate changes, at least at the scale of our analysis. To the extent that species occupancy is related to abundance, our analysis provides the first quantitative support for the hypothesis that B. cockerelli population abundance has increased, but further work is needed to link B. cockerelli population dynamics to Zebra Chip epidemics. Finally, we demonstrate how this historical macro-ecological approach provides a general framework for comparative risk assessment of future pest and insect vector outbreaks
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