Dataset Information

Closed-reference metatranscriptomics enables in planta profiling of putative virulence activities in the grapevine trunk disease complex.

ABSTRACT: Grapevines, like other perennial crops, are affected by so-called 'trunk diseases', which damage the trunk and other woody tissues. Mature grapevines typically contract more than one trunk disease and often multiple grapevine trunk pathogens (GTPs) are recovered from infected tissues. The co-existence of different GTP species in complex and dynamic microbial communities complicates the study of the molecular mechanisms underlying disease development, especially under vineyard conditions. The objective of this study was to develop and optimize a community-level transcriptomics (i.e. metatranscriptomics) approach that could monitor simultaneously the virulence activities of multiple GTPs in planta. The availability of annotated genomes for the most relevant co-infecting GTPs in diseased grapevine wood provided the unprecedented opportunity to generate a multi-species reference for the mapping and quantification of DNA and RNA sequencing reads. We first evaluated popular sequence read mappers using permutations of multiple simulated datasets. Alignment parameters of the selected mapper were optimized to increase the specificity and sensitivity for its application to metagenomics and metatranscriptomics analyses. Initial testing on grapevine wood experimentally inoculated with individual GTPs confirmed the validity of the method. Using naturally infected field samples expressing a variety of trunk disease symptoms, we show that our approach provides quantitative assessments of species composition, as well as genome-wide transcriptional profiling of potential virulence factors, namely cell wall degradation, secondary metabolism and nutrient uptake for all co-infecting GTPs.

SUBMITTER: Morales-Cruz A

PROVIDER: S-EPMC6638111 | biostudies-literature | 2018 Feb

REPOSITORIES: biostudies-literature

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Closed-reference metatranscriptomics enables in planta profiling of putative virulence activities in the grapevine trunk disease complex.

Morales-Cruz Abraham A Allenbeck Gabrielle G Figueroa-Balderas Rosa R Ashworth Vanessa E VE Lawrence Daniel P DP Travadon Renaud R Smith Rhonda J RJ Baumgartner Kendra K Rolshausen Philippe E PE Cantu Dario D

Molecular plant pathology 20170326 2

Grapevines, like other perennial crops, are affected by so-called 'trunk diseases', which damage the trunk and other woody tissues. Mature grapevines typically contract more than one trunk disease and often multiple grapevine trunk pathogens (GTPs) are recovered from infected tissues. The co-existence of different GTP species in complex and dynamic microbial communities complicates the study of the molecular mechanisms underlying disease development, especially under vineyard conditions. The obj ...[more]

PMID: 28218463

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Project description:Grapevine trunk diseases (GTDs) are a serious problem of grapevines worldwide. The microbiota of the grapevine endosphere comprises prokaryotic and eukaryotic endophytes, which may form varied relationships with the host plant from symbiotic to pathogenic. To explore the interaction between grapevine endophytic bacteria and GTDs, the endomicrobiome associated with grapevine wood was characterized using next-generation Illumina sequencing. Wood samples were collected from grapevine trunks with and without external symptoms of GTD (cankers) from two vineyards in the Hunter Valley and Hilltops, NSW, Australia and metagenomic characterization of the endophytic community was conducted using the 16S rRNA gene (341F/806R) and ITS (1F/2R) sequences. Among the important GTD pathogens, Phaeomoniella, Phaeoacremonium, Diplodia and Cryptovalsa species were found to be abundant in both symptomatic and asymptomatic grapevines from both vineyards. Eutypa lata and Neofusicoccum parvum, two important GTD pathogens, were detected in low numbers in Hilltops and the Hunter Valley, respectively. Interestingly, Pseudomonas dominated the bacterial community in canker-free grapevine tissues in both locations, comprising 56-74% of the total bacterial population. In contrast, the Pseudomonas population in grapevines with cankers was significantly lower, representing 29 and 2% of the bacterial community in Hilltops and the Hunter Valley, respectively. The presence of Pseudomonas in healthy grapevine tissues indicates its ability to colonize and survive in the grapevine. The potential of Pseudomonas spp. as biocontrol agents against GTD pathogens was also explored. Dual culture tests with isolated fluorescent Pseudomonas against mycelial discs of nine Botryosphaeria dieback, three Eutypa dieback, and two Esca/Petri disease pathogens, revealed antagonistic activity for 10 Pseudomonas strains. These results suggest the potential of Pseudomonas species from grapevine wood to be used as biocontrol agents to manage certain GTD pathogens.

Project description:Esca is a Grapevine Trunk Disease (GTD) caused by a broad range of taxonomically unrelated fungal pathogens. These attack grapevine wood tissues inducing necroses even in the conductive vascular tissues, thus affecting the vine physiology and potentially leading to plant death. However, the influence of Esca on leaf and whole-plant water transport disruption remains poorly understood. In this paper, a detailed analysis of xylem-related physiological parameters in grapevines that expressed Esca-foliar symptoms was carried out. The experiments were conducted in a vineyard in the Bordeaux region (France) on cv. Cabernet-Sauvignon (Vitis vinifera L.) grapevines, which were monitored for Esca-foliar symptoms over a two-year period. Heat dissipation sap-flow sensors were installed during the summer on grapevines having expressed or not Esca-foliar symptoms. Leaf water potential, stomatal conductance and leaf transpiration were also measured. Physiological monitoring showed that sap flow density and whole-plant transpiration of Esca-infected grapevines decreased significantly a week before the first foliar symptoms appeared. When atmospheric water demand (Vapour Pressure Deficit, VPD) was the highest, both parameters tended to be about twice as low in symptomatic grapevines as in asymptomatic ones. Sap flow density data at the maximum transpiration-time, was systematically 29-30% lower in Esca-infected grapevines compared to control plants before or after the appearance of Esca-foliar symptoms. This trend was observed whatever the temperatures and VPD values measured. In Esca-diseased plants, larger amounts of necrotic wood, mainly white rot, were found in the trunk and cordon of symptomatic grapevines compared to healthy ones, suggesting necroses have an influence in reducing the whole-plant hydraulic capacity. This study reveals that the use of physiological monitoring methods, together with the visual monitoring of foliar symptoms, could prove useful in providing accurate measurements of Esca disease severity.

Project description:BackgroundGrapevine trunk diseases (GTDs) is a disease complex caused by wood pathogenic fungi belonging to genera like Phaeomoniella, Phaeoacremonium, Fomitiporia, Eutypa and members of the family Botryosphaeriaceae. However, the co-occurrence of these fungi in symptomatic and asymptomatic vines at equivalent abundances has questioned their role in GTDs. Hence, we still lack a good understanding of the fungi involved in GTDs, their interactions and the factors controlling their assemblage in vines. We determined the fungal and bacterial microbiome in wood tissues of asymptomatic and symptomatic vines of three main Greek cultivars (Agiorgitiko, Xinomavro, Vidiano), each cultivated in geographically distinct viticultural zones, using amplicon sequencing.ResultsWe noted that cultivar/biogeography (lumped factor) was the strongest determinant of the wood fungal microbiome (p < 0.001, 22.7%), while GTD symptoms condition had a weaker but still significant effect (p < 0.001, 3.5%), being prominent only in the cultivar Xinomavro. Several fungal Amplicon Sequence Variants (ASVs), reported as GTD-associated pathogens like Kalmusia variispora, Fomitiporia spp., and Phaemoniella chlamydosporα (most dominant in our study), were positively correlated with symptomatic vines in a cultivar/viticultural zone dependent manner. Random Forest analysis pointed to P. chlamydosporα, K. variispora, A. alternata and Cladosporium sp., as highly accurate predictors of symptomatic vines (0% error rate). The wood bacterial microbiome showed similar patterns, with biogeography/cultivar being the main determinant (p < 0.001, 25.5%) of its composition, followed by the GTD status of vines (p < 0.001, 5.2%). Differential abundance analysis revealed a universal positive correlation (p < 0.001) of Bacillus and Streptomyces ASVs with asymptomatic vines. Network analysis identified a significant negative co-occurrence network between these bacterial genera and Phaemoniella, Phaeoacrominum and Seimatosporium. These results point to a plant beneficial interaction between Bacillus/Streptomyces and GTD pathogens.ConclusionsOur study (a) provides evidence that GTD symptomatic plants support a wood fungal microbiome, showing cultivar and biogeography-dependent patterns, that could be used as a proxy to distinguish between healthy and diseased vines, (b) points to strong interactions between the bacterial and fungal wood microbiome in asymptomatic vines that should be further pursued in the quest for discovery of novel biocontrol agents.

Dataset Information

Closed-reference metatranscriptomics enables in planta profiling of putative virulence activities in the grapevine trunk disease complex.

Publications

Closed-reference metatranscriptomics enables in planta profiling of putative virulence activities in the grapevine trunk disease complex.

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