Plant necrotrophic bacterial disease resistance phenotypes, QTL, and metabolites identified through integrated genetic mapping and metabolomics in Solanum species.

Most food crops are susceptible to necrotrophic bacteria that cause rotting and wilting diseases in fleshy organs and foods. All varieties of cultivated potato (Solanum tuberosum L.) are susceptible to diseases caused by Pectobacterium species, but resistance has been demonstrated in wild potato relatives including S. chacoense. Previous studies demonstrated that resistance is in part mediated by antivirulence activity of phytochemicals in stems and tubers. Little is known about the genetic basis of antivirulence traits, and the potential for inheritance and introgression into cultivated potato is unclear. Here, the metabolites and genetic loci associated with antivirulence traits in S. chacoense were elucidated by screening a sequenced S. tuberosum x S. chacoense recombinant inbred line (RIL) population for antivirulence traits of its metabolite extracts. Metabolite extracts from the RILs exhibited a quantitative distribution for two antivirulence traits that were positively correlated: quorum sensing inhibition and exo-protease inhibition, with some evidence of transgressive segregation, supporting the role of multiple loci and metabolites regulating these resistance-associated systems. Metabolomics was performed on the highly resistant and susceptible RILs that revealed 30 metabolites associated with resistance, including several alkaloids and terpenes. Specifically, several prenylated metabolites were more abundant in resistant RILs. We constructed a high-density linkage map with 795 SNPs mapped to 12 linkage groups, spanning a length of 1,507 cM and a density of 1 marker per 1.89 cM. Genetic mapping of the antivirulence and metabolite data identified five quantitative trait loci (QTLs) related to quorum sensing inhibition that explained 8-28% of the phenotypic variation and two QTLs for protease activity inhibition that explained 14-19% of the phenotypic variation. Several candidate genes including alkaloid, and secondary metabolite biosynthesis that are related to disease resistance were identified within these QTLs. Taken together, these data support that quorum sensing inhibition and exo-protease inhibition assays may serve as breeding targets to improve resistance to nectrotrophic bacterial pathogens in potato and other plants. The identified candidate genes and metabolites can be utilized in marker assisted selection and genomic selection to improve soft- rot and blackleg disease resistance.

Brassica Cover Crops and Natural Spongospora subterranea Infestation of Peat-Based Potting Mix May Increase Powdery Scab Risk on Potato.

Spongospora subterranea is a soilborne plasmodiophorid that causes powdery scab and root gall formation in potato. In this study, 18 cover crops suitable for use in dry, high-altitude potato production regions were assessed in potting mix trials to determine whether these cover crops altered S. subterranea population levels. Although S. subterranea appeared to invade roots of all plant species tested, the pathogen was unable to complete its life cycle on 11 of 18 cover crops based on postharvest qPCR and microscopy results. Buckwheat, legumes, and scarlet barley do not appear to support pathogen replication, but the pathogen may be able to complete its life cycle in some mustards. High variability occurred in the experiments and part of this may be due to the natural infestations of peat-based potting mix with S. subterranea. A tomato bioassay was used to confirm that commercial sources of peat-based potting mix were infested with S. subterranea. Dry heat and autoclaving were tested as sanitation methods and multiple rounds of autoclaving were required to reduce viable S. subterranea in potting mix. A second cover crop experiment with autoclaved potting mix was conducted and it confirmed that buckwheat, legumes, and barley do not support S. subterranea replication but that some brassica crops may be hosts of this pathogen. The results suggest that buckwheat, legumes, and barley pose the least risk as cover crops in S. subterranea infested fields and show that peat-based potting mix should not be used in seed potato production.

Protease Inhibitors from Solanum chacoense Inhibit Pectobacterium Virulence by Reducing Bacterial Protease Activity and Motility.

Potato is a major staple crop, and necrotrophic bacterial pathogens such as Pectobacterium spp. are a major threat to global food security. Most lines of cultivated potato (Solanum tuberosum) are susceptible to Pectobacterium spp., but some lines of wild potato are resistant, including Solanum chacoense M6. Despite the discovery of resistance in wild potatoes, specific resistance genes are yet to be discovered. Crude protein extract from M6 had a global effect on Pectobacterium brasiliense Pb1692 (Pb1692) virulence phenotypes. Specifically, M6 protein extracts resulted in reduced Pectobacterium exo-protease activity and motility, induced cell elongation, and affected bacterial virulence and metabolic gene expression. These effects were not observed from protein extracts of susceptible potato S. tuberosum DM1. A proteomics approach identified protease inhibitors (PIs) as candidates for S. chacoense resistance, and genomic analysis showed higher abundance and diversity of PIs in M6 than in DM1. We cloned five PIs that are unique or had high abundance in M6 compared with DM1 and purified the proteins (g18987, g28531, g39249, g40384, g6571). Four of the PIs significantly reduced bacterial protease activity, with strongest effects from g28531 and g6571. Three PIs (g18987, g28531, g6571) inhibited disease when co-inoculated with Pectobacterium pathogens into potato tubers. Two PIs (g28531, g6571) also significantly reduced Pb1692 motility and are promising as resistance genes. These results show that S. chacoense PIs contribute to bacterial disease resistance by inhibiting exo-proteases, motility, and tuber maceration and by modulating cell morphology and metabolism. [Formula: see text] Copyright © 2022 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Species of Dickeya and Pectobacterium Isolated during an Outbreak of Blackleg and Soft Rot of Potato in Northeastern and North Central United States.

An outbreak of bacterial soft rot and blackleg of potato has occurred since 2014 with the epicenter being in the northeastern region of the United States. Multiple species of Pectobacterium and Dickeya are causal agents, resulting in losses to commercial and seed potato production over the past decade in the Northeastern and North Central United States. To clarify the pathogen present at the outset of the epidemic in 2015 and 2016, a phylogenetic study was made of 121 pectolytic soft rot bacteria isolated from symptomatic potato; also included were 27 type strains of Dickeya and Pectobacterium species, and 47 historic reference strains. Phylogenetic trees constructed based on multilocus sequence alignments of concatenated dnaJ, dnaX and gyrB fragments revealed the epidemic isolates to cluster with type strains of D. chrysanthemi, D. dianthicola, D. dadantii, P. atrosepticum, P. brasiliense, P. carotovorum, P. parmentieri, P. polaris, P. punjabense, and P. versatile. Genetic diversity within D. dianthicola strains was low, with one sequence type (ST1) identified in 17 of 19 strains. Pectobacterium parmentieri was more diverse, with ten sequence types detected among 37 of the 2015-2016 strains. This study can aid in monitoring future shifts in potato soft rot pathogens within the U.S. and inform strategies for disease management.

Pangenomic Analysis of Dickeya dianthicola Strains Related to the Outbreak of Blackleg and Soft Rot of Potato in the United States.

Dickeya dianthicola has caused an outbreak of blackleg and soft rot of potato in the eastern half of the United States since 2015. To investigate genetic diversity of the pathogen, a comparative analysis was conducted on genomes of D. dianthicola strains. Whole genomes of 16 strains from the United States outbreak were assembled and compared with 16 previously sequenced genomes of D. dianthicola isolated from potato or carnation. Among the 32 strains, eight distinct clades were distinguished based on phylogenomic analysis. The outbreak strains were grouped into three clades, with the majority of the strains in clade I. Clade I strains were unique and homogeneous, suggesting a recent incursion of this strain into potato production from alternative hosts or environmental sources. The pangenome of the 32 strains contained 6,693 genes, 3,377 of which were core genes. By screening primary protein subunits associated with virulence from all U.S. strains, we found that many virulence-related gene clusters, such as plant cell wall degrading enzyme genes, flagellar and chemotaxis related genes, two-component regulatory genes, and type I/II/III secretion system genes, were highly conserved but that type IV and type VI secretion system genes varied. The clade I strains encoded two clusters of type IV secretion systems, whereas the clade II and III strains encoded only one cluster. Clade I and II strains encoded one more VgrG/PAAR spike protein than did clade III. Thus, we predicted that the presence of additional virulence-related genes may have enabled the unique clade I strain to become predominant in the U.S. outbreak.

Laboratory Assays Used to Rank Potato Cultivar Tolerance to Blackleg Showed That Tuber Vacuum Infiltration Results Correlate With Field Observations.

Seed potato certification data collected in Colorado from 2012 to 2016 was used to rank potato cultivar tolerance to blackleg. Five cultivars with different tolerance levels to blackleg ('Chipeta' ≈ 'Alegria' ≈ 'Lamoka' < 'Classic Russet' < 'Yukon Gem') were tested to determine whether we could validate field data with laboratory assays. A strain isolated from Colorado, Pectobacterium atrosepticum CW1-4, and the P. atrosepticum type strain, 33260, were used to inoculate plants through vacuum infiltration of tubers or stem inoculation. Disease was assessed with time-lapse video and by measuring lesion length and disease incidence. After vacuum infiltration of tubers with P. atrosepticum CW1-4 and 33260, cultivars varied in tolerance to soft rot ('Lamoka' < 'Classic Russet' < 'Alegria' ≈ 'Yukon Gem' < 'Chipeta'). Blackleg tolerance of the five cultivars also varied after vacuum infiltration ('Lamoka' < 'Alegria' ≈ 'Chipeta' < 'Classic Russet' ≈ 'Yukon Gem'). All cultivars were susceptible after stem inoculation with either strain. In this assay, 'Chipeta' had the longest lesions, and 'Lamoka' had the smallest lesions. Time-lapse video was used to assess 'Classic Russet' and 'Yukon Gem.' 'Yukon Gem' developed disease symptoms faster than 'Classic Russet,' but the difference was not significant. These results indicate that relative susceptibility of the five cultivars to P. atrosepticum depends on the assay used and that laboratory and greenhouse results differed from field observations.

Genotyping Dickeya dianthicola Causing Potato Blackleg and Soft Rot Outbreak Associated With Inoculum Geography in the United States.

An outbreak of blackleg and soft rot of potato, caused primarily by the bacterial pathogen Dickeya dianthicola, has resulted in significant economic losses in the northeastern United States since 2015. The spread of this seedborne disease is highly associated with seed distribution; therefore, the pathogen likely spread with seed tubers. To describe the blackleg epidemic and track inoculum origins, a total of 1,183 potato samples were collected from 11 states associated with blackleg outbreak from 2015 to 2019. Of these samples, 39.8% tested positive for D. dianthicola. Seventeen isolates of D. dianthicola were recovered from these samples and the genetic diversity of these isolates was examined. Fingerprinting with BOX-A1R-based repetitive extragenic palindromic PCR and phylogenetic analysis based on sequences of the 16S rRNA and gapA genes indicated that D. dianthicola isolates were divided into three genotypes, denoted types I, II, and III. Ninety-five percent of samples from Maine were type I. Type II was found in Maine only in 2015 and 2018. Type II was present throughout the 5 years in some states at a lower percentage than type I. Type III was found in Pennsylvania, New Jersey, and Massachusetts, but not in Maine. Therefore, type I appears to be associated with Maine, but type II appeared to be distributed throughout the northeastern United States. The type II and rarer type III strains were closer to the D. dianthicola type strain isolated from the United Kingdom. This work provides evidence that the outbreak of blackleg of potato in the northeastern United States was caused by multiple strains of D. dianthicola. The geographic origins of these strains remain unknown.

Metabolites from Wild Potato Inhibit Virulence Factors of the Soft Rot and Blackleg Pathogen Pectobacterium brasiliense.

Potato (Solanum tuberosum L.) is the primary vegetable crop consumed worldwide and is largely affected by bacterial pathogens that can cause soft rot and blackleg disease. Recently, resistance to these diseases has been identified in the wild potato S. chacoense, and the mechanism of resistance is unknown. Here, it was hypothesized that S. chacoense stems or tubers have unique chemistry that confers resistance to the pathogen Pectobacterium brasiliense through bactericidal, bacteriostatic, or antivirulence activity. Stem and tuber metabolite extracts were collected from S. chacoense and tested for effects on Pectobacterium bacterial multiplication rates, and activity and expression of known exoenzymes and virulence genes using S. tuberosum extracts as a comparative control. Comparatively, the S. chacoense extracts did not affect bacterial multiplication rate; however, they did reduce pectinase, cellulase, and protease activities. The chemical extracts were profiled using a bioassay-guided fractionation, and a nontargeted metabolomics comparison of S. chacoense and S. tuberosum stems and tubers was performed. The data showed that selected alkaloids, phenolic amines, phenols, amines, and peptides are integrative chemical sources of resistance against the bacteria.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

The Role of ATP-Binding Cassette Transporters in Bacterial Phytopathogenesis.

Bacteria use selective membrane transporting strategies to support cell survival in different environments. Of the membrane transport systems, ATP-binding cassette (ABC) transporters, which utilize the energy of ATP hydrolysis to deliver substrate across the cytoplasmic membrane, are the largest and most diverse superfamily. These transporters import nutrients, export molecules, and are required for diverse cell functions, including cell division and morphology, gene regulation, surface motility, chemotaxis, and interspecies competition. Phytobacterial pathogens encode numerous ABC transporter homologs compared with related nonphytopathogens, with up to 160 transporters per genome, suggesting that plant pathogens must be able to import or respond to a greater number of molecules compared with saprophytes or animal pathogens. Despite their importance, ABC transporters have been little examined in plant pathogens. To understand bacterial phytopathogenesis and evolution, we need to understand the roles that ABC transporters play in plant-microbe interactions. In this review, we outline a multitude of roles that bacterial ABC transporters play, using both plant and animal pathogens as examples, to emphasize the importance of exploring these transporters in phytobacteriology.

Evaluation of Effects of Chemical Soil Treatments and Potato Cultivars on Spongospora subterranea Soil Inoculum and Incidence of Powdery Scab and Potato Mop-Top Virus in Potato.

Spongospora subterranea is a soilborne plasmodiophorid that causes powdery scab in potato. It also transmits potato mop-top virus (PMTV), which causes necrotic arcs (spraing) in potato tubers. Three field experiments were conducted in naturally S. subterranea-infested soil to investigate the effects of two chemicals, Omega 500F (fluazinam) and FOLI-R-PLUS RIDEZ (biological extract), on powdery scab, PMTV, and changes in S. subterranea inoculum with six different potato cultivars. The efficacy of soil treatment with these two chemicals on tuber lesions, root galling, and pathogen population was also assessed in greenhouse trials. The chemical treatments did not reduce powdery scab, root gall formation, or S. subterranea inoculum in the field or greenhouse trials. Postharvest S. subterranea soil inoculum in fields varied across farms and among potato cultivars but the pathogen population consistently increased by the end of the growing season. The evaluated russet cultivars were more tolerant to powdery scab than the yellow- or red-skinned cultivars but all were susceptible to PMTV. In the field, powdery scab indices and soil inoculum changes were positively correlated, while postharvest S. subterranea inoculum was positively correlated with root galling in both greenhouse trials. Powdery scab and PMTV occurred in noninoculated potting mix, indicating that peat-based potting mix is a source for both pathogens. These results demonstrate that chemical management methods currently used by farmers are ineffective, that S. subterranea and PMTV in potting mix can cause severe epidemics in greenhouses, and that potato cultivar choices impact inoculum increases in soil.

Direct Binding of Salicylic Acid to Pectobacterium N-Acyl-Homoserine Lactone Synthase.

Salicylic acid (SA) is a hormone that mediates systemic acquired resistance in plants. We demonstrated that SA can interfere with group behavior and virulence of the soft-rot plant pathogen Pectobacterium spp. through quorum sensing (QS) inhibition. QS is a population density-dependent communication system that relies on the signal molecule acyl-homoserine lactone (AHL) to synchronize infection. P. parmentieri mutants, lacking the QS AHL synthase (expI-) or the response regulator (expR-), were used to determine how SA inhibits QS. ExpI was expressed in DH5α, the QS negative strain of Escherichia coli, revealing direct interference of SA with AHL synthesis. Docking simulations showed SA is a potential ExpI ligand. This hypothesis was further confirmed by direct binding of SA to purified ExpI, shown by isothermal titration calorimetry and microscale thermophoresis. Computational alanine scanning was employed to design a mutant ExpI with predicted weaker binding affinity to SA. The mutant was constructed and displayed lower affinity to the ligand in the binding assay, and its physiological inhibition by SA was reduced. Taken together, these data support a likely mode of action and a role for SA as potent inhibitor of AHL synthase and QS.

Detection and Quantification of Spongospora subterranea Sporosori in Soil by Quantitative Real-Time PCR.

Powdery scab on potato tubers is caused by the obligate soilborne biotroph Spongospora subterranea and is known to cause substantial losses in potato production. The pathogen also infects roots of susceptible hosts, forming galls which can negatively affect root function. S. subterranea is also the vector of Potato mop-top virus, which causes a tuber necrosis disease that can, depending on temperature and cultivar, render potato tubers unmarketable. In this study, we adapted a published protocol to develop a sensitive and robust quantitative real-time PCR (qPCR) assay using specific primers and probes for detecting and quantifying S. subterranea sporosori in soil types that differ in physical properties, including organic matter content and soil pH. For the first time, an external control was utilized and applied directly to the soil prior to DNA extraction, which facilitated normalization of S. subterranea sporosori soil levels from sample to sample. The duplex qPCR protocol was demonstrated to be highly sensitive, capable of detecting and quantifying as few as 1 sporosorus/g of soil, with consistently high qPCR efficiency and the coefficient of determination (R2) values ranging from 94 to 99% and 0.98 to 0.99, respectively. The protocol was successfully implemented in enumerating S. subterranea sporosori in naturally infested field soil collected from several states and in artificial potting mixes with high organic matter content ranging from 64 to 71%. The qPCR method developed can be useful for potato growers to avoid agricultural soils highly infested with S. subterranea and in the development of risk assessment models in the future that incorporate cultivar susceptibility to powdery scab and soil infestation levels.

Development of the Automated Primer Design Workflow Uniqprimer and Diagnostic Primers for the Broad-Host-Range Plant Pathogen Dickeya dianthicola.

Uniqprimer, a software pipeline developed in Python, was deployed as a user-friendly internet tool in Rice Galaxy for comparative genome analyses to design primer sets for PCRassays capable of detecting target bacterial taxa. The pipeline was trialed with Dickeya dianthicola, a destructive broad-host-range bacterial pathogen found in most potato-growing regions. Dickeya is a highly variable genus, and some primers available to detect this genus and species exhibit common diagnostic failures. Upon uploading a selection of target and nontarget genomes, six primer sets were rapidly identified with Uniqprimer, of which two were specific and sensitive when tested with D. dianthicola. The remaining four amplified a minority of the nontarget strains tested. The two promising candidate primer sets were trialed with DNA isolated from 116 field samples from across the United States that were previously submitted for testing. D. dianthicola was detected in 41 samples, demonstrating the applicability of our detection primers and suggesting widespread occurrence of D. dianthicola in North America.

Optimization of an isothermal recombinase polymerase amplification method for real-time detection of Potato virus Y O and N types in potato.

Potato virus Y (PVY) is a global challenge for potato production and the leading cause of seed crop downgrading and rejection for certification. Accurate and timely diagnosis is key to effective control of PVY. Here we optimized the isothermal recombinase polymerase amplification (RPA) for accurate detection of different PVY O and N types that were tested, present in different tissues of potato plants including tubers with a primer set that specifically targets the highly conserved pipo region within the viral genome. Combined with a simplified preparation of the template by tissue homogenization, we established a rapid RPA procedure, which can allow real time detection in less than 10 min with a fluorescent probe. Specificity of the reaction was determined by the lack of cross-reactivity with other common potato viruses. Although RPA reagents remain more expensive than PCR reagents, RPA technology is equivalent in that results can be visualized by gel electrophoresis or with a fluorescent probe with greater sensitivity; and it is superior to the common PCR-based assay in its versatility, speed, and lack of need for a highly purified RNA template.

Complete Genome Sequence of Dickeya dianthicola ME23, a Pathogen Causing Blackleg and Soft Rot Diseases of Potato.

In 2014, an outbreak of potato blackleg and soft rot disease emerged in North America and continues to impact potato production. Here, we report the annotated genome sequence of Dickeya dianthicola ME23, a strain hypothesized to be representative of the bacterial population responsible for this disease outbreak.

Colorado Seed Potato Certification Data Analysis Shows Mosaic and Blackleg are Major Diseases of Seed Potato and Identifies Tolerant Potato Varieties.

Seed potato certification programs aim to limit disease incidence in planting material to levels below a threshold that causes significant losses to seed potato buyers. Records maintained for seed potato certification can be analyzed for trends in seed potato varietal mixture and disease incidences over time. We used logistic regression models to determine effects of year, potato variety, and their interaction on the incidences of potato diseases and disorders based on seed potato certification data collected in Colorado from 2012 to 2016. The effect of seed generation of important varieties on the incidence of common potato diseases was also quantified. Among the documented diseases, mosaic, caused primarily by Potato virus Y, is a persistent problem leading to high percentage of seed lot rejections in both summer and winter inspections, while blackleg, caused by Dickeya spp. and Pectobacterium spp., was a common disease causing summer rejections. The model demonstrated year, potato variety, and their interaction were key factors contributing to incidence of varietal mixture, or one or more potato diseases and disorders. For summer inspections, our models identified 53, 42, and six varieties sensitive to mosaic, blackleg, and leafroll, respectively. There were 17, 15, and six varieties that were tolerant to mosaic, blackleg, and leafroll regardless of pathogen pressures in the environment. For winter inspections, 51 varieties were sensitive to mosaic and three to leafroll, whereas 45 and one were relatively tolerant to mosaic and leafroll. The pattern of seed generation effects of selected potato varieties on mosaic and blackleg incidence was inconsistent across inspection years. In addition, we observed a significant negative correlation between the relative abundance of the green peach aphid, Myzus persicae, and mosaic in winter inspections. Mosaic incidences in summer and leafroll incidences were not influenced by common aphid species captured or total aphids in the valley. These results identify mosaic and blackleg as major causes of seed potato rejections and downgrades, sensitive and tolerant varieties, and provide suggestions for improving integrated crop management practices in Colorado.

The Changing Face of Bacterial Soft-Rot Diseases.

Bacterial soft rot is a disease complex caused by multiple genera of gram-negative and gram-positive bacteria, with Dickeya and Pectobacterium being the most widely studied soft-rot bacterial pathogens. In addition to soft rot, these bacteria also cause blackleg of potato, foot rot of rice, and bleeding canker of pear. Multiple Dickeya and Pectobacterium species cause the same symptoms on potato, complicating epidemiology and disease resistance studies. The primary pathogen species present in potato-growing regions differs over time and space, further complicating disease management. Genomics technologies are providing new management possibilities, including improved detection and biocontrol methods that may finally allow effective disease management. The recent development of inbred diploid potato lines is also having a major impact on studying soft-rot pathogens because it is now possible to study soft-rot disease in model plant species that produce starchy vegetative storage organs. Together, these new discoveries have changed how we face diseases caused by these pathogens.

Phylogenetic and Morphological Identification of the Novel Pathogen of Rheum palmatum Leaf Spot in Gansu, China.

A new leaf spot disease was observed on leaves of Rheum palmatum (Chinese rhubarb) in Northwest China (Gansu Province) starting in 2005. A Septoria-like fungus was isolated and completion of Koch's postulates confirmed that the fungus was the casual agent of the leaf spot disease. Morphology and molecular methods were combined to identify the pathogen. The fungus produced conidiomata pycnidia and the conidia were 2~5 septate, 61.2~134.1 µm in length and 3.53~5.3 µm in width, which is much larger than the known Spetoria species that infects Polygonaceae species. Phylogenic analysis of the internal transcribed spacer region confirmed that this Spetoria-like fungus is within the Spetoria genus but distinct from known Spetoria species. Together, these morphological and phylogenetic data support that the R. palmatum infecting Spetoria strain is a newly-described plant pathogenic species.

Perspectives on the Transition From Bacterial Phytopathogen Genomics Studies to Applications Enhancing Disease Management: From Promise to Practice.

The advent of genomics has advanced science into a new era, providing a plethora of "toys" for researchers in many related and disparate fields. Genomics has also spawned many new fields, including proteomics and metabolomics, furthering our ability to gain a more comprehensive view of individual organisms and of interacting organisms. Genomic information of both bacterial pathogens and their hosts has provided the critical starting point in understanding the molecular bases of how pathogens disrupt host cells to cause disease. In addition, knowledge of the complete genome sequence of the pathogen provides a potentially broad slate of targets for the development of novel virulence inhibitors that are desperately needed for disease management. Regarding plant bacterial pathogens and disease management, the potential for utilizing genomics resources in the development of durable resistance is enhanced because of developing technologies that enable targeted modification of the host. Here, we summarize the role of genomics studies in furthering efforts to manage bacterial plant diseases and highlight novel genomics-enabled strategies heading down this path.

Identification of host-microbe interaction factors in the genomes of soft rot-associated pathogens Dickeya dadantii 3937 and Pectobacterium carotovorum WPP14 with supervised machine learning.

BACKGROUND: A wealth of genome sequences has provided thousands of genes of unknown function, but identification of functions for the large numbers of hypothetical genes in phytopathogens remains a challenge that impacts all research on plant-microbe interactions. Decades of research on the molecular basis of pathogenesis focused on a limited number of factors associated with long-known host-microbe interaction systems, providing limited direction into this challenge. Computational approaches to identify virulence genes often rely on two strategies: searching for sequence similarity to known host-microbe interaction factors from other organisms, and identifying islands of genes that discriminate between pathogens of one type and closely related non-pathogens or pathogens of a different type. The former is limited to known genes, excluding vast collections of genes of unknown function found in every genome. The latter lacks specificity, since many genes in genomic islands have little to do with host-interaction. RESULT: In this study, we developed a supervised machine learning approach that was designed to recognize patterns from large and disparate data types, in order to identify candidate host-microbe interaction factors. The soft rot Enterobacteriaceae strains Dickeya dadantii 3937 and Pectobacterium carotovorum WPP14 were used for development of this tool, because these pathogens are important on multiple high value crops in agriculture worldwide and more genomic and functional data is available for the Enterobacteriaceae than any other microbial family. Our approach achieved greater than 90% precision and a recall rate over 80% in 10-fold cross validation tests. CONCLUSION: Application of the learning scheme to the complete genome of these two organisms generated a list of roughly 200 candidates, many of which were previously not implicated in plant-microbe interaction and many of which are of completely unknown function. These lists provide new targets for experimental validation and further characterization, and our approach presents a promising pattern-learning scheme that can be generalized to create a resource to study host-microbe interactions in other bacterial phytopathogens.