Phylogenomic analysis of 343 Xanthomonas citri pv. citri strains unravels introduction history and dispersal paths.

Xanthomonas citri pv. citri (Xcc) causes the devastating citrus canker disease. Xcc is known to have been introduced into Florida, USA in at least three different events in 1915, 1986 and 1995 with the first two claimed to be eradicated. It was questioned whether the Xcc introduction in 1986 has been successfully eradicated. Furthermore, it is unknown how Xcc has spread throughout the citrus groves in Florida. In this study, we investigated the population structure of Xcc to address these questions. We sequenced the whole genome of 343 Xcc strains collected from Florida groves between 1997 and 2016. Our analysis revealed two distinct clusters of Xcc. Our data strongly indicate that the claimed eradication of the 1986 Xcc introduction was not successful and Xcc strains from 1986 introduction were present in samples from at least 8 counties collected after 1994. Importantly, our data revealed that the Cluster 2 strains, which are present in all 20 citrus-producing counties sampled in Florida, originated from the Xcc introduction event in the Miami area in 1995. Our data suggest that Polk County is the epicenter of the dispersal of Cluster 2 Xcc strains, which is consistent with the fact that three major hurricanes passed through Polk County in 2004. As copper-based products have been extensively used to control citrus canker, we also investigated whether Xcc strains have developed resistance to copper. Notably, none of the 343 strains contained known copper resistance genes. Twenty randomly selected Xcc strains displayed sensitivity to copper. Overall, this study provides valuable insights into the introduction, eradication, spread, and copper resistance of Xcc in Florida.

Bacterial Mutation During Seasonal Epidemics.

Rapidly evolving bacterial pathogens pose a unique challenge for long-term plant disease management. In this study, we investigated the types and rate of mutations in bacterial populations during seasonal disease epidemics. Two phylogenetically distinct strains of the bacterial spot pathogen, Xanthomonas perforans, were marked, released in tomato fields, and recaptured at several time points during the growing season. Genomic variations in recaptured isolates were identified by comparative analysis of their whole-genome sequences. In total, 180 unique variations (116 substitutions, 57 insertions/deletions, and 7 structural variations) were identified from 300 genomes, resulting in the overall host-associated mutation rate of ∼0.3 to 0.9/genome/week. This result serves as a benchmark for bacterial mutation during epidemics in similar pathosystems. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Multiple acquisitions of XopJ2 effectors in populations of Xanthomonas perforans.

Type III effectors (T3Es) are major determinants of Xanthomonas virulence and targets for resistance breeding. XopJ2 (syn. AvrBsT) is a highly conserved YopJ-family T3E acquired by X. perforans, the pathogen responsible for bacterial spot disease of tomato. In this study, we characterized a new variant (XopJ2b) of XopJ2, which is predicted to have a similar 3D structure as the canonical XopJ2 (XopJ2a) despite sharing only 70% sequence identity. XopJ2b carries an acetyltransferase domain and the critical residues required for its activity, and the positions of these residues are predicted to be conserved in 3D structure of the proteins. We demonstrated that XopJ2b is a functional T3E and triggers hypersensitive response when translocated into pepper cells. Like XopJ2a, XopJ2b triggers HR in Arabidopsis that is suppressed by the deacetylase, SOBER1. We found xopJ2b in genome sequences of X. euvesicatoria, X. campestris, X. citri, X. guizotiae, and X. vasicola strains, suggesting widespread horizontal transfer. In X. perforans, xopJ2b was present in strains collected in North and South America, Africa, Asia, Australia, and Europe, whereas xopJ2a had a more narrow geographic distribution. This study expands the Xanthomonas T3E repertoire, demonstrates functional conservation in T3E evolution, and further supports the importance of XopJ2 in X. perforans fitness on tomato.

Inter-species expression of an EF-HAND CALCIUM BINDING PROTEIN in Xanthomonas perforans leads to reduced virulence and decreased immune evasion in tomato plants.

Many phytopathogenic bacteria require a type three secretion system (TTSS) to activate effector triggered immunity (ETI). We identified a calcium binding protein, EfhXXfa, in the citrus pathogen, X. citri subsp. aurantifolii, that does not require a TTSS to activate reactive oxygen species (ROS) and elicit a hypersensitive reaction (HR) in tomato leaves following infection. Purified, recombinant EfhXXfa was shown to bind two moles of calcium per mole of protein, whereas mutation of the first of two EF-hands did not bind calcium . EfhXXfa expression was determined to be inducible in hrp-inducing medium. Additionally, growth of X. perforans transconjugants with and without the efhXXfa gene in hrp-inducing medium differed in intracellular calcium concentration; the transconjugant without efhXXfa yielded higher cell pellet masses and higher increased intracellular calcium concentrations relative to cells expressing EfhXXfa. An EfhXXfa homolog, EfhXXe, present in the pepper pathogen, X. euvesicatoria, when expressed in the tomato pathogen, X. perforans, triggered ROS production and an HR in tomato leaves and is a host-limiting factor. Interestingly, all tested X. perforans and X. euvesicatoria strains pathogenic on tomato contain a stop codon immediately upstream of the first EF-hand domain in the efhXXe gene, whereas most X. euvesicatoria strains pathogenic on pepper do not.

Phylodynamic Insights into Global Emergence and Diversification of the Tomato Pathogen Xanthomonas hortorum pv. gardneri.

The emergence of plant pathogens is often associated with waves of unique evolutionary and epidemiological events. Xanthomonas hortorum pv. gardneri is one of the major pathogens causing bacterial spot disease of tomatoes. After its first report in the 1950s, there were no formal reports on this pathogen until the 1990s, despite active global research on the pathogens that cause tomato and pepper bacterial spot disease. Given the recently documented global distribution of X. hortorum pv. gardneri, our objective was to examine genomic diversification associated with its emergence. We sequenced the genomes of X. hortorum pv. gardneri strains collected in eight countries to examine global population structure and pathways of emergence using phylodynamic analysis. We found that strains isolated post-1990 group by region of collection and show minimal impact of recombination on genetic variation. A period of rapid geographic expansion in X. hortorum pv. gardneri is associated with acquisition of a large plasmid conferring copper tolerance by horizontal transfer and coincides with the burgeoning hybrid tomato seed industry through the 1980s. The ancestry of X. hortorum pv. gardneri is consistent with introduction to hybrid tomato seed production and dissemination during the rapid increase in trade of hybrid seeds.

Insights into bs5 resistance mechanisms in pepper against Xanthomonas euvesicatoria through transcriptome profiling.

BACKGROUND: Bacterial spot of pepper (BSP), caused by four different Xanthomonas species, primarily X. euvesicatoria (Xe), poses a significant challenge in pepper cultivation. Host resistance is considered the most important approach for BSP control, offering long-term protection and sustainability. While breeding for resistance to BSP for many years focused on dominant R genes, introgression of recessive resistance has been a more recent focus of breeding programs. The molecular interactions underlying recessive resistance remain poorly understood. RESULTS: In this study, transcriptomic analyses were performed to elucidate defense responses triggered by Xe race P6 infection by two distinct pepper lines: the Xe-resistant line ECW50R containing bs5, a recessive resistance gene that confers resistance to all pepper Xe races, and the Xe-susceptible line ECW. The results revealed a total of 3357 upregulated and 4091 downregulated genes at 0, 1, 2, and 4 days post-inoculation (dpi), with the highest number of differentially expressed genes (DEGs) observed at 2 dpi. Pathway analysis highlighted DEGs in key pathways such as plant-pathogen interaction, MAPK signaling pathway, plant hormone signal transduction, and photosynthesis - antenna proteins, along with cysteine and methionine metabolism. Notably, upregulation of genes associated with PAMP-Triggered Immunity (PTI) was observed, including components like FLS2, Ca-dependent pathways, Rboh, and reactive oxygen species (ROS) generation. In support of these results, infiltration of ECW50R leaves with bacterial suspension of Xe led to observable hydrogen peroxide accumulation without a rapid increase in electrolyte leakage, suggestive of the absence of Effector-Triggered Immunity (ETI). Furthermore, the study confirmed that bs5 does not disrupt the effector delivery system, as evidenced by incompatible interactions between avirulence genes and their corresponding dominant resistant genes in the bs5 background. CONCLUSION: Overall, these findings provide insights into the molecular mechanisms underlying bs5-mediated resistance in pepper against Xe and suggest a robust defense mechanism in ECW50R, primarily mediated through PTI. Given that bs5 provides early strong response for resistance, combining this resistance with other dominant resistance genes will enhance the durability of resistance to BSP.

The underground world of plant disease: Rhizosphere dysbiosis reduces above-ground plant resistance to bacterial leaf spot and alters plant transcriptome.

Just as the human gut microbiome is colonized by a variety of microbes, so too is the rhizosphere of plants. An imbalance in this microbial community, known as dysbiosis, can have a negative impact on plant health. This study sought to explore the effect of rhizosphere dysbiosis on the health of tomato plants (Solanum lycopersicum L.), using them and the foliar bacterial spot pathogen Xanthomonas perforans as model organisms. The rhizospheres of 3-week-old tomato plants were treated with either streptomycin or water as a control, and then spray-inoculated with X. perforans after 24 h. Half of the plants that were treated with both streptomycin and X. perforans received soil microbiome transplants from uninfected plant donors 48 h after the streptomycin was applied. The plants treated with streptomycin showed a 26% increase in disease severity compared to those that did not receive the antibiotic. However, the plants that received the soil microbiome transplant exhibited an intermediate level of disease severity. The antibiotic-treated plants demonstrated a reduced abundance of rhizobacterial taxa such as Cyanobacteria from the genus Cylindrospermum. They also showed a down-regulation of genes related to plant primary and secondary metabolism, and an up-regulation of plant defence genes associated with induced systemic resistance. This study highlights the vital role that beneficial rhizosphere microbes play in disease resistance, even against foliar pathogens.

Amylase-associated genetic pattern in Xanthomonas euvesicatoria on pepper.

Bacterial leaf spot of pepper (BSP), primarily caused by Xanthomonas euvesicatoria (Xe), poses a significant challenge to pepper production worldwide. Despite its impact, the genetic diversity of this pathogen remains underexplored, which limits our understanding of its population structure. To bridge this knowledge gap, we conducted a comprehensive analysis using 103 Xe strains isolated from pepper in southwest Florida to characterize genomic and type III effector (T3E) variation in this population. Phylogenetic analysis of core genomes revealed a major distinct genetic lineage associated with amylolytic activity. This amylolytic lineage was represented in Xe strains globally. Molecular clock analysis dated the emergence of amylolytic strains in Xe to around 1972. Notably, non-amylolytic strains possessed a single base pair frameshift deletion in the ⍺-amylase gene yet retained a conserved C-terminus. GUS assay revealed the expression of two open reading frames in non-amylolytic strains, one at the N-terminus and another that starts 136 base pairs upstream of the ⍺-amylase gene. Analysis of T3Es in the Florida Xe population identified variation in 12 effectors, including two classes of mutations in avrBs2 that prevent AvrBs2 from triggering a hypersensitive response in Bs2-resistant pepper plants. Knowledge of T3E variation could be used for effector-targeted disease management. This study revealed previously undescribed population structure in this economically important pathogen.IMPORTANCEBacterial leaf spot (BSP), a significant threat to pepper production globally, is primarily caused by Xanthomonas euvesicatoria (Xe). Limited genomic data has hindered detailed studies on its population diversity. This study analyzed the whole-genome sequences of 103 Xe strains from peppers in southwest Florida, along with additional global strains, to explore the pathogen's diversity. The study revealed two major distinct genetic groups based on their amylolytic activity, the ability to break down starch, with non-amylolytic strains having a mutation in the ⍺-amylase gene. Additionally, two classes of mutations in the avrBs2 gene were found, leading to susceptibility in pepper plants with the Bs2 resistance gene, a commercially available resistance gene for BSP. These findings highlight the need to forecast the emergence of such strains, identify genetic factors for innovative disease management, and understand how this pathogen evolves and spreads.

New Genotypes of Phytophthora nicotianae Found on Strawberry in Florida.

Phytophthora cactorum is the most common causal agent of Phytophthora crown rot and leather rot of strawberry, but P. nicotianae is also responsible for the disease in Florida. Studies of P. nicotianae populations have suggested that different groups of genotypes are associated with different hosts; however, it is not yet clear how many lineages exist globally and how they are related to different production systems. The aim of this study was to determine the genetic relationships of P. nicotianae isolates from Florida strawberry with genotypes reported from other hosts, quantify the genetic variation on strawberry, and test for an association with nursery source. A total of 49 isolates of P. nicotianae were collected from strawberry plants originating from multiple nursery sources during six seasons of commercial fruit production in Florida. Microsatellite genotyping identified 28 multilocus genotypes on strawberry that were distinct among 208 isolates originating from various hosts and locations. Based on STRUCTURE analysis, two genetic groups were identified: one consisting of isolates from strawberry, and the other comprising samples from different hosts. Multilocus genotypes were shared among nursery sources, and populations defined by nursery were not differentiated. Both mating types were found among the isolates from North Carolina- and California-origin plants and in most strawberry seasons; however, a predominance of A1 was observed, and regular sexual reproduction was not supported by the data. This study reveals a unique genetic population of P. nicotianae associated with strawberry and emphasizes the vital role of nursery monitoring in mitigating disease spread.

Whole-Genome-Sequence-Based Classification of Xanthomonas euvesicatoria pv. eucalypti and Computational Analysis of the Type III Secretion System.

Xanthomonas spp. infect a wide range of annual and perennial plants. Bacterial blight in young seedlings of Eucalyptus spp. in Indonesia was originally identified as X. perforans. However, these strains failed to elicit a hypersensitive response (HR) on either tomatoes or peppers. Two of the strains, EPK43 and BCC 972, when infiltrated into tomato and pepper leaves, failed to grow to significant levels in comparison with well-characterized X. euvesicatoria pv. perforans (Xp) strains. Furthermore, spray inoculation of 'Bonny Best' tomato plants with a bacterial suspension of the Eucalyptus strains resulted in no obvious symptoms. We sequenced the whole genomes of eight strains isolated from two Eucalyptus species between 2007 and 2015. The strains had average nucleotide identities (ANIs) of at least 97.8 with Xp and X. euvesicatoria pv. euvesicatoria (Xeu) strains, both of which are causal agents of bacterial spot of tomatoes and peppers. A comparison of the Eucalyptus strains revealed that the ANI values were >99.99% with each other. Core genome phylogeny clustered all Eucalyptus strains with X. euvesicatoria pv. rosa. They formed separate clades, which included X. euvesicatoria pv. alangii, X. euvesicatoria pv. citrumelonis, and X. euvesicatoria pv. alfalfae. Based on ANI, phylogenetic relationships, and pathogenicity, we designated these Eucalyptus strains as X. euvesicatoria pv. eucalypti (Xee). Comparative analysis of sequenced strains provided unique profiles of type III secretion effectors. Core effector XopD, present in all pathogenic Xp and Xeu strains, was absent in the Xee strains. Comparison of the hrp clusters of Xee, Xp, and Xeu genomes revealed that HrpE in Xee strains was very different from that in Xp and Xeu. To determine if it was functional, we deleted the gene and complemented with the Xee hrpE, confirming it was essential for secretion of type III effectors. HrpE has a hypervariable N-terminus in Xanthomonas spp., in which the N-terminus of Xee strains differs significantly from those of Xeu and Xp strains.

Xanthomonas as a Model System for Studying Pathogen Emergence and Evolution.

In this review, we highlight studies in which whole-genome sequencing, comparative genomics, and population genomics have provided unprecedented insights into past and ongoing pathogen evolution. These include new understandings of the adaptive evolution of secretion systems and their effectors. We focus on Xanthomonas pathosystems that have seen intensive study and improved our understanding of pathogen emergence and evolution, particularly in the context of host specialization: citrus canker, bacterial blight of rice, and bacterial spot of tomato and pepper. Across pathosystems, pathogens appear to follow a pattern of bursts of evolution and diversification that impact host adaptation. There remains a need for studies on the mechanisms of host range evolution and genetic exchange among closely related but differentially host-specialized species and to start moving beyond the study of specific strain and host cultivar pairwise interactions to thinking about these pathosystems in a community context.

Phenotypic and Genetic Diversity of Xanthomonads Isolated from Pepper (Capsicum spp.) in Taiwan from 1989 to 2019.

Bacterial spot caused by Xanthomonas spp. is an economically important disease of pepper causing significant yield losses in Taiwan. Monitoring the pathogen population on a continuous basis is necessary for developing disease management strategies. We analyzed a collection of xanthomonad strains isolated from pepper in Taiwan between 1989 and 2019. Among the sequenced genomes, 65 were identified as Xanthomonas euvesicatoria, and 10 were X. perforans. Thirty-five X. euvesicatoria and 10 X. perforans strains were copper tolerant, whereas only four X. euvesicatoria and none of the X. perforans strains were tolerant to streptomycin. Nine X. euvesicatoria strains were amylolytic, which is considered an unusual characteristic for X. euvesicatoria. Bayesian analysis of the population structure based on core gene single-nucleotide polymorphisms clustered the strains into five clusters for X. euvesicatoria and three clusters for X. perforans. One X. perforans cluster, designated as TP-2019, appears to be a novel genetic cluster based on core genes, accessory gene content, and effector profile. This knowledge of pathogen diversity with whole genomic information will be useful in future comparative studies and in improving breeding programs to develop disease-resistant cultivars and other disease management options.

Population Genomics Reveals an Emerging Lineage of Xanthomonas perforans on Pepper.

Xanthomonas perforans-the dominant causal agent of bacterial leaf spot of tomato-is an emerging pathogen of pepper, indicative of a potential host expansion across the southeastern United States. However, studies of the genetic diversity and evolution of X. perforans from pepper remain limited. In this study, the whole-genome sequences of 35 X. perforans strains isolated from pepper from four fields and two transplant facilities across southwest Florida between 2019 and 2021 were used to compare genomic divergence, evolution, and variation in type III secreted effectors. Phylogenetic analysis based on core genes revealed that all 35 X. perforans strains formed one genetic cluster with pepper and tomato strains from Alabama and Turkey and were closely related to strains isolated from tomato in Indiana, Mexico, and Louisiana. The in planta population growth of tomato strains isolated from Indiana, Mexico, Louisiana, and Turkey in pepper leaf mesophyll was on par with pepper X. perforans and X. euvesicatoria strains. Molecular clock analysis of the 35 Florida strains dated their emergence to approximately 2017. While strains varied in copper tolerance, all sequenced strains harbored the avrHah1 transcription activation-like effector located on a conjugative plasmid, not previously reported in Florida. Our findings suggest that there is a geographically distributed lineage of X. perforans strains on tomato that has the genetic background to cause disease on pepper. Moreover, this study clarifies potential adaptive variants of X. perforans on pepper that could help forecast the emergence of such strains and enable immediate or preemptive intervention.

Open Access and Reproducibility in Plant Pathology Research: Guidelines and Best Practices.

The landscape of scientific publishing is experiencing a transformative shift toward open access, a paradigm that mandates the availability of research outputs such as data, code, materials, and publications. Open access provides increased reproducibility and allows for reuse of these resources. This article provides guidance for best publishing practices of scientific research, data, and associated resources, including code, in The American Phytopathological Society journals. Key areas such as diagnostic assays, experimental design, data sharing, and code deposition are explored in detail. This guidance aligns with that observed by other leading journals. We hope the information assembled in this paper will raise awareness of best practices and enable greater appraisal of the true effects of biological phenomena in plant pathology.

Unexpected Diversity of Pseudomonads Associated with Bacterial Leaf Spot of Cucurbits in the Southeastern United States.

Bacterial leaf spot of cucurbits (BLS) is an emerging disease in the southeastern United States that is capable of causing widespread outbreaks under conducive conditions. Historically attributed solely to the bacterium Pseudomonas syringae pv. lachrymans, recent studies have identified additional P. syringae pathovars as causal agents of the disease. To further investigate the identity and diversity of P. syringae strains associated with BLS in the southeastern United States, 47 bacterial isolates were recovered from symptomatic cucurbits from Florida, Alabama, and Georgia. Strains were characterized using the LOPAT testing scheme, fluorescence, and pathogenicity to watermelon and squash seedlings. Thirty-eight fluorescent isolates underwent whole-genome sequencing and were further characterized with 16S rRNA, four gene multilocus sequence analysis (MLSA) phylogeny, and average nucleotide identity analysis. Thirty-four isolates were identified as members of the P. syringae species complex, including P. syringae sensu stricto (12), P. alliivorans (12), P. capsici (nine), and P. viridiflava (one). An additional four isolates were found to belong to the Pseudomonas genus outside of the syringae species complex, though they did not share 95% or greater average nucleotide identity to any validly published species and are believed to belong to three novel Pseudomonas species. These results reveal an unpredicted level of diversity of Pseudomonas strains associated with BLS in the region and show the benefits of whole-genome sequencing for strain identification. Identification of P. capsici, which is capable of causing disease at higher temperatures than P. syringae, as a causal agent of BLS may also affect management strategies in the future.

Strength in Numbers: Density-Dependent Volatile-Induced Antimicrobial Activity by Xanthomonas perforans.

For most of the 20th century, Xanthomonas euvesicatoria was the only known bacterium associated with bacterial spot of tomato in Florida. X. perforans quickly replaced X. euvesicatoria, mainly because of production of three bacteriocins (BCNs) against X. euvesicatoria; however, X. perforans outcompeted X. euvesicatoria even when the three known BCNs were deleted. Surprisingly, we observed antimicrobial activity against X. euvesicatoria in the BCN triple mutant when the triple mutant was grown in Petri plates containing multiple spots but not in Petri plates containing only one spot. We determined that changes in the headspace composition (i.e., volatiles) rather than a diffusible signal in the agar were required for induction of the antimicrobial activity. Other Xanthomonas species also produced volatile-induced antimicrobial compounds against X. euvesicatoria and elicited antimicrobial activity by X. perforans. A wide range of plant pathogenic bacteria, including Clavibacter michiganensis subsp. michiganensis, Pantoea stewartii, and Pseudomonas cichorii, also elicited antimicrobial activity by X. perforans when multiple spots of the species were present. To identify potential antimicrobial compounds, we performed liquid chromatography with high-resolution mass spectrometry of the agar surrounding the spot in the high cell density Petri plates where the antimicrobial activity was present compared with agar surrounding the spot in Petri plates with one spot where antimicrobial activity was not observed. Among the compounds identified in the zone of inhibition were N-butanoyl-L-homoserine lactone and N-(3-hydroxy-butanoyl)-homoserine lactone, which are known quorum-sensing metabolites in other bacteria.

Exploring Diversity of Bacterial Spot Associated Xanthomonas Population of Pepper in Southwest Florida.

Bacterial spot caused by Xanthomonas spp. is a significant disease that challenges pepper growers worldwide and is particularly severe in a hot and humid environment. Understanding the pathogen's population biology is critical for sustainable disease management. The goal of this study was to characterize the species, race, and bactericide sensitivity of bacterial spot-associated Xanthomonas collected from pepper in Florida. A survey of pepper production fields in southwest Florida between 2019 and 2021-covering two counties, eight farms, and two transplant facilities-resulted in the isolation of 542 Xanthomonas euvesicatoria and 35 Xanthomonas perforans strains. Four races were identified on pepper, of which most strains were race P1 (42%), race P6 (26%), race P3 (24%), and less common was race P4 (8%). All X. perforans strains were characterized as race P1 and showed a compatible reaction on tomato. Sixty-two and 96% of strains were sensitive to copper sulfate and streptomycin, respectively. One farm that did not use copper to manage the disease contained only copper-sensitive strains and was the only farm with race P3 strains. Strains were assayed for starch hydrolysis activity of which a third of X. euvesicatoria strains were strongly amylolytic, a characteristic not typically observed in X. euvesicatoria. All X. perforans strains produced bacteriocins against X. euvesicatoria in vitro. The Xanthomonas population causing bacterial spot on pepper in southwest Florida is diverse and dynamic; thus, regular monitoring provides pertinent information to plant breeders and growers for designing disease management strategies.

Ralstonia Strains from Potato-Growing Regions of Kenya Reveal Two Phylotypes and Epidemic Clonality of Phylotype II Sequevar 1 Strains.

Bacterial wilt, caused by the Ralstonia solanacearum species complex (RSSC), is the most destructive potato disease in Kenya. Studies were conducted to (i) determine the molecular diversity of RSSC strains associated with bacterial wilt of potato in Kenya, (ii) generate an RSSC distribution map for epidemiological inference, and (iii) determine whether phylotype II sequevar 1 strains exhibit epidemic clonality. Surveys were conducted in 2018 and 2019, in which tubers from wilting potato plants and stem samples of potential alternative hosts were collected for pathogen isolation. The pathogen was phylotyped by multiplex PCR and 536 RSSC strains typed at a sequevar level. Two RSSC phylotypes were identified, phylotype II (98.4%, n = 506 [sequevar 1 (n = 505) and sequevar 2 (n = 1)]) and phylotype I (1.6%, n = 30 [sequevar 13 (n = 9) and a new sequevar (n = 21)]). The phylotype II sequevar 1 strains were haplotyped using multilocus tandem repeat sequence typing (TRST) schemes. The TRST scheme identified 51 TRST profiles within the phylotype II sequevar 1 strains with a modest diversity index (HGDI = 0.87), confirming the epidemic clonality of RSSC phylotype II sequevar 1 strains in Kenya. A minimum spanning tree and mapping of the TRST profiles revealed that TRST27 '8-5-12-7-5' is the primary founder of the clonal complex of RSSC phylotype II sequevar 1 and is widely distributed via latently infected seed tubers. [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.

How Do Pathogens Evolve Novel Virulence Activities?

This article is part of the Top 10 Unanswered Questions in MPMI invited review series.We consider the state of knowledge on pathogen evolution of novel virulence activities, broadly defined as anything that increases pathogen fitness with the consequence of causing disease in either the qualitative or quantitative senses, including adaptation of pathogens to host immunity and physiology, host species, genotypes, or tissues, or the environment. The evolution of novel virulence activities as an adaptive trait is based on the selection exerted by hosts on variants that have been generated de novo or arrived from elsewhere. In addition, the biotic and abiotic environment a pathogen experiences beyond the host may influence pathogen virulence activities. We consider host-pathogen evolution, host range expansion, and external factors that can mediate pathogen evolution. We then discuss the mechanisms by which pathogens generate and recombine the genetic variation that leads to novel virulence activities, including DNA point mutation, transposable element activity, gene duplication and neofunctionalization, and genetic exchange. In summary, if there is an (epi)genetic mechanism that can create variation in the genome, it will be used by pathogens to evolve virulence factors. Our knowledge of virulence evolution has been biased by pathogen evolution in response to major gene resistance, leaving other virulence activities underexplored. Understanding the key driving forces that give rise to novel virulence activities and the integration of evolutionary concepts and methods with mechanistic research on plant-microbe interactions can help inform crop protection.[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.

Need for speed: bacterial effector XopJ2 is associated with increased dispersal velocity of Xanthomonas perforans.

Bacterial spot caused by Xanthomonas perforans (Xp) is an economically important disease in tomato. Previous studies have shown that the recently isolated Xp strains have acquired and retained the effector gene, xopJ2, which has been reported to increase fitness of the pathogen in the field. To elucidate the fitness benefit of xopJ2, we quantified the effect of xopJ2 on the dispersal and evolution of Xp populations on tomato. We compared movement of two wild-type Xp strains expressing xopJ2 to their respective xopJ2 mutants when co-inoculated in the field. We developed a binary logistic model to predict the presence of Xp over spatial and temporal dimensions with or without xopJ2. Based on the model, wild-type bacteria were dispersed approximately three times faster than the xopJ2 mutants. In a simulation experiment, the selective advantage due to increased dispersal velocity led to an increase in the frequency of xopJ2 gene in the Xp population and its apparent fixation within 10 to 12 cropping seasons of the tomato crop. Our results show that the presence of a single gene can affect the dispersal of a bacterial pathogen and significantly alter its population dynamics.