Phylogeography, origin and population structure of the self-fertile emerging plant pathogen Phytophthora pseudosyringae.

Phytophthora pseudosyringae is a self-fertile pathogen of woody plants, particularly associated with tree species from the genera Fagus, Notholithocarpus, Nothofagus and Quercus, which is found across Europe and in parts of North America and Chile. It can behave as a soil pathogen infecting roots and the stem collar region, as well as an aerial pathogen infecting leaves, twigs and stem barks, causing particular damage in the United Kingdom and western North America. The population structure, migration and potential outcrossing of a worldwide collection of isolates were investigated using genotyping-by-sequencing. Coalescent-based migration analysis revealed that the North American population originated from Europe. Historical gene flow has occurred between the continents in both directions to some extent, yet contemporary migration is overwhelmingly from Europe to North America. Two broad population clusters dominate the global population of the pathogen, with a subgroup derived from one of the main clusters found only in western North America. Index of association and network analyses indicate an influential level of outcrossing has occurred in this preferentially inbreeding, homothallic oomycete. Outcrossing between the two main population clusters has created distinct subgroups of admixed individuals that are, however, less common than the main population clusters. Differences in life history traits between the two main population clusters should be further investigated together with virulence and host range tests to evaluate the risk each population poses to natural environments worldwide.

Validation of reference gene stability for normalization of RT-qPCR in Phytophthora capsici Leonian during its interaction with Piper nigrum L.

The selection of stable reference genes for the normalization of reverse transcription quantitative real-time PCR (RT-qPCR) is generally overlooked despite being the crucial element in determining the accuracy of the relative expression of genes. In the present study, the stability of seven candidate reference genes: actin (act), α-tubulin (atub), ß-tubulin (btub), translation elongation factor 1-α (ef1), elongation factor 2 (ef2), ubiquitin-conjugating enzyme (ubc) and 40S ribosomal protein S3A (ws21) in Phytophthora capsici has been validated. The validation was performed at six infection time points during its interaction with its susceptible host Piper nigrum, two developmental stages, and for the combined dataset. Four algorithms: geNorm, NormFinder, BestKeeper, and the ΔCt method were compared, and a comprehensive ranking order was produced using RefFinder. The overall analysis revealed that ef1, ws21, and ubc were identified as the three most stable genes in the combined dataset, ef1, ws21, and act were the most stable at the infection stages, and, ef1, btub, and ubc were most stable during the developmental stages. These findings were further corroborated by validating the P. capsici pathogenesis gene NPP1 expression. The findings are significant as this is the first study addressing the stability of reference genes for P. capsici-P. nigrum interaction studies.

Phytophthora pluvialis maintenance, spore production and detached needle assays.

Phytophthora pluvialis is an oomycete that primarily infects Pinus radiata and Pseudotsuga menziesii causing the destructive foliar disease red needle cast (RNC). Recent observations show that P. pluvialis can also infect western hemlock inducing resinous cankers. High-throughput and reproducible infection assays are integral to find key information on tree health and oomycete pathogenicity. In this protocol, we describe the propagation and spore induction of P. pluvialis, followed by detached needle assays for verification and quantification of virulence of P. pluvialis in P. radiata needles. These needle assays can be employed for high-throughput screening of tree needles with diverse genetic backgrounds. In downstream analysis, Quantitative PCR (qPCR) was utilized to assess relative gene expression, as exemplified by candidate RxLR effector protein PpR01. Additional techniques like RNA sequencing, metabolomics, and proteomics can be combined with needle assays and can offer comprehensive insights into P. pluvialis infection mechanisms.

Unveiling the microbiome of hydroponically cultivated lettuce: impact of Phytophthora cryptogea infection on plant-associated microorganisms.

Understanding the complex interactions between plants and their associated microorganisms is crucial for optimizing plant health and productivity. While microbiomes of soil-bound cultivated crops are extensively studied, microbiomes of hydroponically cultivated crops have received limited attention. To address this knowledge gap, we investigated the rhizosphere and root endosphere of hydroponically cultivated lettuce. Additionally, we sought to explore the potential impact of the oomycete pathogen Phytophthora cryptogea on these microbiomes. Root samples were collected from symptomatic and nonsymptomatic plants in three different greenhouses. Amplicon sequencing of the bacterial 16S rRNA gene revealed significant alterations in the bacterial community upon P. cryptogea infection, particularly in the rhizosphere. Permutational multivariate analysis of variance (perMANOVA) revealed significant differences in microbial communities between plants from the three greenhouses, and between symptomatic and nonsymptomatic plants. Further analysis uncovered differentially abundant zero-radius operational taxonomic units (zOTUs) between symptomatic and nonsymptomatic plants. Interestingly, members of Pseudomonas and Flavobacterium were positively associated with symptomatic plants. Overall, this study provides valuable insights into the microbiome of hydroponically cultivated plants and highlights the influence of pathogen invasion on plant-associated microbial communities. Further research is required to elucidate the potential role of Pseudomonas and Flavobacterium spp. in controlling P. cryptogea infections within hydroponically cultivated lettuce greenhouses.

Diversity and distribution of Phytophthora species across different types of riparian vegetation in Italy with the description of Phytophthora heteromorpha sp. nov.

Riparian formations encompass a diverse suite of transitional zones between terrestrial and aquatic ecosystems. During the last decades, these formations have been impacted by several emerging diseases. The first outbreaks were detected on alder formations, but have progressively also been observed on other plant species such as Betula pubescens, Nerium oleander, Populus alba, Salix alpina, Salix purpurea and Tamarix gallica. Declining plants showed a plethora of symptoms (leaf spot, shoot blight, bleeding cankers and root rot) indicative of Phytophthora infections. Since there is little information about the aetiology of these pathosystems, from November 2019 to March 2023, an in-depth study was conducted in 46 riparian ecosystems spanning from the Mediterranean to Alpine regions. Overall, 744 symptomatic samples (stem bleeding cankers and root with rhizosphere) from 27 host species were collected for Phytophthora isolation. Based on morphology and DNA sequence data, 20 known Phytophthora species belonging to seven phylogenetic clades have been identified: P. plurivora (202 isolates), P. gonapodyides (156), P. pseudosyringae (84), P. lacustris (57), P. acerina (31), P. idaei (30), P. alpina (20), P. pseudocryptogea (19), P. cambivora (13), P. pseudotsugae (13), P. cactorum (9), P. honggalleglyana (6), P. pseudogregata (6), P. debattistii (4), P. multivora (4), P. cinnamomi (3), P. bilorbang (2) P. crassamura (2), P. ilicis (2) and P. inundata (2). In addition, 26 isolates of a new putative species obtained from Alnus incana and Pinus sylvestris are described here as Phytophthora heteromorpha sp. nov. The new species proved to be pathogenic on grey alder causing symptoms congruent with field observations. This study represents the most comprehensive investigation on the Phytophthora species associated with declining riparian vegetation in Italy and highlights that the polyphagous pathogen P. plurivora represents a growing threat to Mediterranean, temperate and alpine ecosystems.

Cacao pod transcriptome profiling of seven genotypes identifies features associated with post-penetration resistance to Phytophthora palmivora.

The oomycete Phytophthora palmivora infects the fruit of cacao trees (Theobroma cacao) causing black pod rot and reducing yields. Cacao genotypes vary in their resistance levels to P. palmivora, yet our understanding of how cacao fruit respond to the pathogen at the molecular level during disease establishment is limited. To address this issue, disease development and RNA-Seq studies were conducted on pods of seven cacao genotypes (ICS1, WFT, Gu133, Spa9, CCN51, Sca6 and Pound7) to better understand their reactions to the post-penetration stage of P. palmivora infection. The pod tissue-P. palmivora pathogen assay resulted in the genotypes being classified as susceptible (ICS1, WFT, Gu133 and Spa9) or resistant (CCN51, Sca6 and Pound7). The number of differentially expressed genes (DEGs) ranged from 1625 to 6957 depending on genotype. A custom gene correlation approach identified 34 correlation groups. De novo motif analysis was conducted on upstream promoter sequences of differentially expressed genes, identifying 76 novel motifs, 31 of which were over-represented in the upstream sequences of correlation groups and associated with gene ontology terms related to oxidative stress response, defense against fungal pathogens, general metabolism and cell function. Genes in one correlation group (Group 6) were strongly induced in all genotypes and enriched in genes annotated with defense-responsive terms. Expression pattern profiling revealed that genes in Group 6 were induced to higher levels in the resistant genotypes. An additional analysis allowed the identification of 17 candidate cis-regulatory modules likely to be involved in cacao defense against P. palmivora. This study is a comprehensive exploration of the cacao pod transcriptional response to P. palmivora spread after infection. We identified cacao genes, promoter motifs, and promoter motif combinations associated with post-penetration resistance to P. palmivora in cacao pods and provide this information as a resource to support future and ongoing efforts to breed P. palmivora-resistant cacao.

Transcriptome-based analysis of candidate gene markers associated with resistance mechanism to Phytophthora melonis that causes root and crown rot in pumpkin.

Root and crown rot incited by an oomycete, Phytophthora melonis , causes significant yield losses in commercial pumpkin (Cucurbita pepo ) production worldwide. Currently, resistant cultivars and knowledge of molecular mechanism of C. pepo against P. melonis are scarce. Here, we analysed the quantitative gene expression changes of 10 candidate gene markers (bHLH87, ERF014, HSF, MYB, PR-1, WRKY21, CPI, POD, PSK, SGT ) in pumpkin roots and leaves at three time points (h post-inoculation, hpi) following inoculation with P. melonis in two resistant (Ghelyani and Tanbal), and two susceptible (Marmari and Khoreshti) varieties of pumpkin. Gene expression using quantitative real time PCR along a time course revealed the strongest transcriptomic response at 48 and 72hpi in resistant genotypes, 1.1-2.7-fold in roots and leaves, respectively, with a high significant correlation (r =0.857**-0.974**). We also found that CPI , PSK, SGT1 and POD act as a dual regulator that similarly modulate immunity not only against P. melonis , but also against other diseases such as early blight (Alternaria cucumerina) , powdery mildew (Podosphaera xanthii ), downy mildews (Pseudoperonospora cubensis ), and pathogenic plant nematodes (Meloidogyne javanica ). Furthermore, significantly higher activities of the ROS scavenging defence enzymes, catalase (1.6-fold increase) and peroxidase (6-fold increase) were observed in the roots of resistant cultivars at different hpi compared with non-inoculated controls. In addition, the biomass growth parameters including leaf and root length, stem and root diameter, root fresh weight and volume were significantly different among studied genotypes. Cumulatively, the transcriptome data provide novel insights into the response of pumpkins for improving pumpkin breeding to P. melonis .

Unveiling Vacuolar H+-ATPase Subunit a as the Primary Target of the Pyridinylmethyl-Benzamide Fungicide, Fluopicolide.

An estimated 240 fungicides are presently in use, but the direct targets for the majority remain elusive, constraining fungicide development and efficient resistance monitoring. In this study, we found that Pcα-actinin knockout did not influence the sensitivity of Phytophthora capsici to fluopicolide, which is a notable oomycete inhibitor. Using a combination of Bulk Segregant Analysis Sequencing and Drug Affinity Responsive Target Stability (DARTS) assays, the vacuolar H+-ATPase subunit a (PcVHA-a) was pinpointed as the target protein of fluopicolide. We also confirmed four distinct point mutations in PcVHA-a responsible for fluopicolide resistance in P. capsici through site-directed mutagenesis. Molecular docking, ATPase activity assays, and a DARTS assay suggested a fluopicolide-PcVHA-a interaction. Sequence analysis and further molecular docking validated the specificity of fluopicolide for oomycetes or fish. These findings support the claim that PcVHA-a is the target of fluopicolide, proposing vacuolar H+-ATPase as a promising target for novel fungicide development.

Cross-pathogenicity of Phytophthora palmivora associated with bud rot disease of oil palm and development of biomarkers for detection.

Phytophthora palmivora has caused disease in many crops including oil palm in the South America region. The pathogen has had a significant economic impact on oil palm cultivation in Colombia, and therefore poses a threat to oil palm cultivation in other regions of the World, especially in Southeast Asia, the largest producer of the crop. This study aimed to look at the ability of isolates from Malaysia, Colombia, and other regions to cross-infect Malaysian oil palm, durian, and cocoa and to develop specific biomarkers and assays for identification, detection, and diagnosis of P. palmivora as a key component for the oil palm biosecurity continuum in order to contain the disease especially at the ports of entry. We have developed specific molecular biomarkers to identify and detect Phytophthora palmivora using polymerase chain reaction (PCR) and real-time loop mediated isothermal amplification (rt-LAMP) in various sample types such as soil and plants. The limit of detection (DNA template, pure culture assay) for the PCR assay is 5.94 × 10-2 ng µl-1 and for rt-LAMP is 9.28 × 10-4 ng µl-1. Diagnosis using rt-LAMP can be achieved within 30 min of incubation. In addition, PCR primer pair AV3F/AV3R developed successfully distinguished the Colombian and Malaysian P. palmivora isolates.

A rapid molecular diagnostic tool to discriminate alleles of avirulence genes and haplotypes of Phytophthora sojae using high-resolution melting analysis.

Effectors encoded by avirulence genes (Avr) interact with the Phytophthora sojae resistance gene (Rps) products to generate incompatible interactions. The virulence profile of P. sojae is rapidly evolving as a result of the large-scale deployment of Rps genes in soybean. For a successful exploitation of Rps genes, it is recommended that soybean growers use cultivars containing the Rps genes corresponding to Avr genes present in P. sojae populations present in their fields. Determination of the virulence profile of P. sojae isolates is critical for the selection of soybean cultivars. High-resolution melting curve (HRM) analysis is a powerful tool, first applied in medicine, for detecting mutations with potential applications in different biological fields. Here, we report the development of an HRM protocol, as an original approach to discriminate effectors, to differentiate P. sojae haplotypes for six Avr genes. An HRM assay was performed on 24 P. sojae isolates with different haplotypes collected from soybean fields across Canada. The results clearly confirmed that the HRM assay discriminated different virulence genotypes. Moreover, the HRM assay was able to differentiate multiple haplotypes representing small allelic variations. HRM-based prediction was validated by phenotyping assays. This HRM assay provides a unique, cost-effective and efficient tool to predict virulence pathotypes associated with six different Avr (1b, 1c, 1d, 1k, 3a and 6) genes from P. sojae, which can be applied in the deployment of appropriate Rps genes in soybean fields.

Phytophthora sojae Effector PsCRN108 Targets CAMTA2 to Suppress HSP40 Expression and ROS Burst.

Oomycete pathogens secrete numerous crinkling and necrosis proteins (CRNs) to manipulate plant immunity and promote infection. However, the functional mechanism of CRN effectors is still poorly understood. Previous research has shown that the Phytophthora sojae effector PsCRN108 binds to the promoter of HSP90s and inhibits their expression, resulting in impaired plant immunity. In this study, we found that in addition to HSP90, PsCRN108 also suppressed other Heat Shock Protein (HSP) family genes, including HSP40. Interestingly, PsCRN108 inhibited the expression of NbHSP40 through its promoter, but did not directly bind to its promoter. Instead, PsCRN108 interacted with NbCAMTA2, a negative regulator of plant immunity. NbCAMTA2 was a negative regulator of NbHSP40 expression, and PsCRN108 could promote such inhibition activity of NbCAMTA2. Our results elucidated the multiple roles of PsCRN108 in the suppression of plant immunity and revealed a new mechanism by which the CRN effector hijacked transcription factors to affect immunity. [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.

Characterizing the Dynamics of Virulence and Fungicide Resistance of Phytophthora capsici in Michigan Vegetable Fields Reveals Loci Associated with Virulence.

The oomycete Phytophthora capsici is a destructive pathogen infecting more than 50 plant species and is one of the most serious threats to cucurbit production. Phytophthora blight caused by Phytophthora capsici can affect all plant growth stages, and fungicides and cultural controls are used to limit losses. Dissecting pathogen virulence and fungicide resistance can provide insights into pathogenic mechanisms and inform effective management practices to control P. capsici. In this study, we assessed virulence, mefenoxam sensitivity, and genetic diversity of nine P. capsici populations collected from Cucurbitaceae, Solanaceae, and Fabaceae host families in Michigan from 2002 to 2016. We developed 992 simple sequence repeats (SSRs) in the P. capsici genome and identified 60 SSRs located within or close to RXLR-class (Arginine-any amino acid-Leucine-Arginine) effectors and 29 SSRs within or close to effector CRN (CRinkling and Necrosis) family protein, which represent 62 RXLR and 34 putative CRNs. Population structure analysis shows that mefenoxam resistance was not associated with the year of collection, host type, or location, but there were significant differences in virulence among the populations. Using the general linear model and mixed linear model-based association analyses with all effector-related SSR markers, we identified four SSR markers significantly associated with at least one of the virulence-related parameters. Of these, one (Pce_SC18) was in a predicted CRN effector and had high identity with the putative PhCRN37 effector in the pathogen Plasmopara halstedii, which can be further verified for virulence identification in P. capsici.

Discovery and Identification of Viruses Infecting Oomycetes.

This chapter describes protocols suitable for the detection and identification of RNA viruses infecting oomycetes (so-called water molds of Kingdom Heterokonta, Stramenopila), focusing on species of Phytophthora and exemplified by P. fragariae. The protocol includes laboratory procedures for oomycete cultivation and total RNA extraction from harvested mycelia, followed by instructions on suitable parameters given for sequencing companies on ribosomal RNA depletion, cDNA library preparation, and total RNA-sequencing (RNA-Seq). We also describe the bioinformatics steps needed for de novo assembly of raw reads into contigs, removal of host-associated contigs, and virus identification by database searches, as well as host validation by RT-PCR. All steps are described using an exemplar RNA-Seq library containing a yet undescribed fusagravirus hosted by a P. fragariae isolate.

Characteristics of famoxadone-resistant mutants of Phytophthora litchii and their effect on lychee fruit quality.

Lychee downy blight (LDB), a common disease caused by the oomycete Phytophthora litchii, poses a significant threat to both pre- and post-harvest stages, leading to substantial economic losses. Famoxadone, a quinone outside inhibitor fungicide, was registered for controlling LDB in China in 2002. However, limited information is available regarding the risk, mechanism, and impact on lychee fruit quality associated with famoxadone resistance. In this study, we determined the sensitivity of 133 P. litchii isolates to famoxadone, yielding a mean EC50 value of 0.46 ± 0.21 µg/mL. Through fungicide adaption, we derived resistant mutants with M124I and Y131C substitutions in PlCyt b (Cytochrome b in P. litchii) from wild-type isolates. In vitro assessments revealed that the fitness of the resistant mutants was significantly lower compared to the parental isolates. These laboratory findings demonstrate a moderate resistance risk of P. litchii to famoxadone. Molecular docking analyses indicated that the M124I and Y131C alterations disrupted hydrogen bonds and weakened the binding energy between famoxadone and PlCyt b. This indicates that the M124I and Y131C changes do indeed confer famoxadone resistance in P. litchii. Infection caused by famoxadone-resistant mutants exhibited a decreased or comparable impact on the characteristic traits of lychee fruit compared to the sensitive isolate. For future detection of famoxadone-resistant strains, AS-PCR primers were designed based on the M124I substitution.

Genetic diversity of Actinidia spp. shapes the oomycete pattern associated with Kiwifruit Vine Decline Syndrome (KVDS).

Kiwifruit Vine Decline Syndrome (KVDS) is an important soil-borne disease for the Italian kiwifruit industry, causing €300,000 in economic losses in 2020 alone. So far, the organisms recognized as involved in the aetiology of KVDS mainly belong to the Oomycota. As no effective management strategies exist, a promising approach to overcoming KVDS is the use of resistant species as rootstocks or for inclusion in breeding programs. Several Actinidia genotypes showing different level of resistance to KVDS were grown in disease-promoting soils. A metabarcoding approach was set up to identify KVDS-associated oomycetes and investigate whether the main species involved may vary according to plant genotype. Our results clearly showed significant differences between the genotypes in terms of oomycetes present in both plant rhizosphere and endosphere, which were strongly correlated with the symptoms displayed. We found out that the resistance of Actinidia macrosperma to KVDS is related to its ability to shape the pathobiome, particularly as far as the endosphere is concerned. In our conditions, Phytophthora sp. was predominantly found in sensitive genotypes, whilst Globisporangium intermedium was mainly detected in asymptomatic plants, suggesting that the latter species could compete with the recruitment of Phytophthora sp. in plants with different levels of resistance, consequently, explaining the onset of symptoms and the resistance condition.

A global-temporal analysis on Phytophthora sojae resistance-gene efficacy.

Plant disease resistance genes are widely used in agriculture to reduce disease outbreaks and epidemics and ensure global food security. In soybean, Rps (Resistance to Phytophthora sojae) genes are used to manage Phytophthora sojae, a major oomycete pathogen that causes Phytophthora stem and root rot (PRR) worldwide. This study aims to identify temporal changes in P. sojae pathotype complexity, diversity, and Rps gene efficacy. Pathotype data was collected from 5121 isolates of P. sojae, derived from 29 surveys conducted between 1990 and 2019 across the United States, Argentina, Canada, and China. This systematic review shows a loss of efficacy of specific Rps genes utilized for disease management and a significant increase in the pathotype diversity of isolates over time. This study finds that the most widely deployed Rps genes used to manage PRR globally, Rps1a, Rps1c and Rps1k, are no longer effective for PRR management in the United States, Argentina, and Canada. This systematic review emphasizes the need to widely introduce new sources of resistance to P. sojae, such as Rps3a, Rps6, or Rps11, into commercial cultivars to effectively manage PRR going forward.

Phyllosphere microbial diversity and specific taxa mediate within-cultivar resistance to Phytophthora palmivora in cacao.

The oomycete pathogen Phytophthora palmivora, which causes black pod rot (BPR) on cacao (Theobroma cacao L.), is responsible for devastating yield losses worldwide. Genetic variation in resistance to Phytophthora spp. is well documented among cacao cultivars, but variation has also been observed in the incidence of BPR even among trees of the same cultivar. In light of evidence that the naturally occurring phyllosphere microbiome can influence foliar disease resistance in other host-pathogen systems, it was hypothesized that differences in the phyllosphere microbiome between two field accessions of the cultivar Gainesville II 164 could be responsible for their contrasting resistance to P. palmivora. Bacterial alpha diversity was higher but fungal alpha diversity was lower in the more resistant accession MITC-331, and the accessions harbored phyllosphere microbiomes with distinct community compositions. Six bacterial and 82 fungal amplicon sequence variants (ASVs) differed in relative abundance between MITC-333 and MITC-331, including bacterial putative biocontrol agents and a high proportion of fungal pathogens, and nine fungal ASVs were correlated with increased lesion development. The roles of contrasting light availability and host mineral nutrition, particularly potassium, are also discussed. Results of this preliminary study can be used to guide research into microbiome-informed integrated pest management strategies effective against Phytophthora spp. in cacao. IMPORTANCE Up to 40% of the world's cacao is lost each year to diseases, the most devastating of which is black pod rot, caused by Phytophthora palmivora. Though disease resistance is often attributed to cacao genotypes (i.e., disease-resistant rootstocks), this study highlights the role of the microbiome in contributing to differences in resistance even among accessions of the same cacao cultivar. Future studies of plant-pathogen interactions may need to account for variation in the host microbiome, and optimizing the cacao phyllosphere microbiome could be a promising new direction for P. palmivora resistance research.

CRISPR/Cas12a-based approaches for efficient and accurate detection of Phytophthora ramorum.

Introduction: Phytophthora ramorum is a quarantine pathogen that causes leaf blight and shoot dieback of the crown, bark cankers and death on a number of both ornamental and forest trees, especially in North America and northern Europe, where it has produced severe outbreaks. Symptoms caused by P. ramorum can be confused with those by other Phytophthora and fungal species. Early and accurate detection of the causal pathogen P. ramorum is crucial for effective prevention and control of Sudden Oak Death. Methods: In this study, we developed a P. ramorum detection technique based on a combination of recombinase polymerase amplification (RPA) with CRISPR/Cas12a technology (termed RPACRISPR/ Cas12a). Results: This novel method can be utilized for the molecular identification of P. ramorum under UV light and readout coming from fluorophores, and can specifically detect P. ramorum at DNA concentrations as low as 100 pg within 25 min at 37°C. Discussion: We have developed a simple, rapid, sensitive, unaided-eye visualization, RPA CRISPR/Cas12a-based detection system for the molecular identification of P. ramorum that does not require technical expertise or expensive ancillary equipment. And this system is sensitive for both standard laboratory samples and samples from the field.

Comparative Transcriptome Analysis Reveals Novel Candidate Resistance Genes Involved in Defence against Phytophthora cactorum in Strawberry.

Crown rot, caused by Phytophthora cactorum, is a devastating disease of strawberry. While most commercial octoploid strawberry cultivars (Fragaria × ananassa Duch) are generally susceptible, the diploid species Fragaria vesca is a potential source of resistance genes to P. cactorum. We previously reported several F. vesca genotypes with varying degrees of resistance to P. cactorum. To gain insights into the strawberry defence mechanisms, comparative transcriptome profiles of two resistant genotypes (NCGR1603 and Bukammen) and a susceptible genotype (NCGR1218) of F. vesca were analysed by RNA-Seq after wounding and subsequent inoculation with P. cactorum. Differential gene expression analysis identified several defence-related genes that are highly expressed in the resistant genotypes relative to the susceptible genotype in response to P. cactorum after wounding. These included putative disease resistance (R) genes encoding receptor-like proteins, receptor-like kinases, nucleotide-binding sites, leucine-rich repeat proteins, RPW8-type disease resistance proteins, and 'pathogenesis-related protein 1'. Seven of these R-genes were expressed only in the resistant genotypes and not in the susceptible genotype, and these appeared to be present only in the genomes of the resistant genotypes, as confirmed by PCR analysis. We previously reported a single major gene locus RPc-1 (Resistance to Phytophthora cactorum 1) in F. vesca that contributed resistance to P. cactorum. Here, we report that 4-5% of the genes (35-38 of ca 800 genes) in the RPc-1 locus are differentially expressed in the resistant genotypes compared to the susceptible genotype after inoculation with P. cactorum. In particular, we identified three defence-related genes encoding wall-associated receptor-like kinase 3, receptor-like protein 12, and non-specific lipid-transfer protein 1-like that were highly expressed in the resistant genotypes compared to the susceptible one. The present study reports several novel candidate disease resistance genes that warrant further investigation for their role in plant defence against P. cactorum.