β-1,4-Endoglucanase (ENG) is a canonical effector protein secreted by the esophageal glands of plant-parasitic nematodes. Homology analysis identified a β-1,4-endoglucanase homolog in Hirschmanniella mucronata, designated as HmENG1. In situ hybridization demonstrated the specific localization of Hmeng1 transcripts within the esophageal glands of H.mucronata. Recombinant HmENG1 exhibited cellulase activity in vitro. In planta RNAi experiments confirmed that silencing Hmeng1 significantly reduced the infection capability of H.mucronata. In addition to its potential role in degrading plant cell walls, HmENG1 was found to interact with glyceraldehyde-3-phosphate dehydrogenase (OsGAPDH) in yeast and plant systems. Furthermore, analysis of Osgapdh knockout mutants indicated that OsGAPDH is involved in rice resistance against H.mucronata. Transcriptomic analysis suggested that the absence of OsGAPDH disrupted carbohydrate metabolism and jasmonic acid-related pathways. These findings demonstrate that HmENG1 is a classic cell wall-degrading enzyme that may exhibit multifunctional roles during H.mucronata infection of rice.

Phytophthora palmivoracauses significant economic losses in Colombian cacao crops, yet its population structure and pathogenicity remain poorly characterized. This study tested the hypothesis that the Colombian Andes mountain complex acts as a geographic barrier limiting gene flow among P. palmivora populations. We analyzed 73 isolates from western, central, and eastern cacao-growing regions using DArTseq genotyping and standardized pathogenicity assays. High genotypic diversity was observed (Simpson's index = 0.99), with corrected association index analysis indicating predominantly clonal reproduction, consistent with all isolates being the A2 mating type. Discriminant analysis of principal components identified seven genetic clusters, but these showed no correlation with geographic distribution, contradicting our geographic barrier hypothesis. Mantel tests confirmed no isolation-by-distance (r = 0.004, P = 0.650). Pathogenicity tests on detached CCN51 leaves revealed extensive virulence variation that was independent of genetic clustering or geography. These findings suggest that observed genetic diversity results from multiple pathogen introductions and anthropogenic dispersal rather than geographic barrier. The lack of correlation between genotype and virulence indicates that pathogenicity determinants are independent of neutral molecular markers used for population structure analysis. Management recommendations include stringent phytosanitary measures to prevent further introductions and consideration of pathogen diversity in resistance breeding programs.

Maize stalk rot, primarily caused by Fusarium spp., is a significant disease in many maize-growing areas worldwide, often leading to considerable yield losses under favorable conditions for infection. In this study, a genome-wide association study for Fusarium stalk rot resistance was carried out using 230,506 single-nucleotide polymorphism (SNP) markers on phenotypic data from 178 maize inbred lines. Four SNPs were significantly associated with stalk rot resistance, with three located within previously reported quantitative trait locus regions and one novel locus identified on chromosome 9 (9_149289012). Candidate genes within a 600-kb window around each SNP were retrieved from the MaizeGDB database, revealing 174 candidate genes in the B73_RefGen_v5 reference genome, including eight resistance genes and eight differentially expressed genes. Notably, three potential candidate genes were identified around the novel SNP 9_149289012 on chromosome 9. Additionally, transcriptome analysis identified 1,597 genes related to disease resistance, associated with GO terms involved in resistance responses, nutrient metabolism, and toxin metabolism. Overall, this study provides valuable insights into the genetic basis of Fusarium stalk rot resistance in maize, highlighting both known and novel loci along with candidate genes for further functional characterization.

A Neopestalotiopsis spp. has recently emerged as an invasive fungal pathogen threatening strawberry (Fragaria × ananassa) production in North Carolina by causing leaf spots, fruit rot, and crown rot, resulting in significant production challenges. Understanding pathogen diversity and virulence variability is vital for developing effective disease management strategies and improving host resistance. Sixty-five Neopestalotiopsis isolates were collected from multiple strawberry cultivars across diverse production regions in North Carolina and characterized using a high-resolution melting quantitative real-time PCR (HRM-qPCR) assay targeting a partial β-tubulin (β-tub) gene sequence. HRM-qPCR analysis differentiated the isolates into two distinct genetic groups, comprising 28 and 37 isolates, respectively, indicating substantial genetic diversity within the pathogen population. To establish reliable disease screening methods, four inoculation techniques were evaluated: foliar spray inoculation with spore suspensions and inoculation with Neopestalotiopsis-infested oatmeal grain, both of which consistently produced disease symptoms and were selected for virulence assessments. To further investigate pathogenic variability, 10 arbitrarily selected isolates were evaluated by spray inoculatingon across 15 strawberry cultivars. Significant effects of isolate, cultivar, and isolate × cultivar interactions were observed, suggesting potential pathogenic specialization among isolates. Although all isolates were pathogenic across the cultivars tested, notable variability in virulence was observed among isolate-cultivar combinations. Among the cultivars evaluated, AC Valley Sunset, Jewel, and Malwina consistently exhibited lower disease susceptibility, suggesting greater tolerance to Neopestalotiopsis. These findings improve understanding of the pathogen diversity and host-pathogen interactions and provide valuable information for cultivar selection, breeding programs, and integrated disease management strategies against this emerging pathogen.

Phloem-feeding insects, including the Asian citrus psyllid (Diaphorina citri), navigate one of the most osmotically challenging diets in nature, the sugar-rich phloem sap. Maintaining osmotic homeostasis is critical for survival, development, and vector competence, particularly in transmitting 'Candidatus Liberibacter asiaticus', the causal agent of citrus greening disease or huanglongbing (HLB). This review synthesizes current knowledge on the physiological, biochemical, and behavioral adaptations that enable D. citri to cope with extreme osmotic pressures. Key strategies include sucrose hydrolysis and conversion into glucose, fructose, and trehalose, as well as redistribution via the haemolymph, and efficient excretion of honeydew, which varies structurally and chemically across different developmental stages and sexes. These adaptations support prolonged phloem feeding, facilitate dual-lifestyle pathogen survival in both phloem and haemolymph, and directly enhance 'Ca. L. asiaticus' acquisition and transmission efficiency. HI also highlight honeydew as an indicator for feeding behavior, host suitability, and susceptibility to insecticides is also highlighted. Finally, emerging approaches to disrupt osmoregulation, including RNAi-mediated interference with sugar metabolism and water transport, which may provide innovative avenues for integrated management of D. citri and mitigation of HLB spread, will be discussed.

Biotrophic rust fungi secrete effector proteins through haustoria to promote successful colonization. Consequently, the functional analysis of haustorial effector proteins has become central to understanding rust pathogenicity. In previous work, we generated the haustorial transcriptome of Gymnosporangium yamadae, the causal agent of apple rust disease, and predicted numerous candidate effector genes. Here, we systematically characterized the functions of 13 haustorially expressed candidate effectors of G. yamadae using a heterologous system to assess their modulation of plant immunity and subcellular localization. Confocal microscopy revealed diverse and distinct localization patterns for both full-length and signal peptide-truncated effector constructs fused with a GFP tag at either the N- or C-terminus, including targeting to the nucleus, nuclear speck, chloroplast, and punctate cytosolic structures. Among them, eight effector genes exhibited elevated expression during early colonization and during the spermogonia and aecia stages of fungal development. Our findings highlight the multifunctional roles of G. yamadae haustorial effectors in modulating plant immunity and potentially other cellular processes. This study addresses the lack of functional analyses of haustorial effectors of forest tree rust pathogens and provides new insights into rust-host interaction mechanisms.

Bacterial wilt caused by Ralstonia pseudosolanacearum poses a significant threat to turmeric (Curcuma longa L.) production in many growing regions. Although plant-associated microbial communities may contribute to disease suppression, the ecological roles of rhizosphere and endosphere microbiomes in turmeric are still underexplored. Here, we characterized rhizosphere and endosphere bacterial communities using 16S rRNA gene amplicon sequencing and evaluated their antagonistic activity against bacterial wilt pathogen of turmeric R. pseudosolanacearum strain RalsTur1. Microbiome profiling revealed compartment-specific patterns in turmeric-associated bacterial communities, with rhizosphere communities strongly structured by geographic location and endosphere communities comparatively stable across field sites. The endosphere core microbiome was dominated by bacterial members in the family Enterobacteriaceae, particularly Enterobacter, along with Pseudomonas, whereas rhizosphere communities included diverse taxa such as Bacillus and members of the Allorhizobium-Neorhizobium-Pararhizobium-Rhizobium group. Interbacterial competition assays showed that several turmeric-associated isolates reduced RalsTur1 populations in vitro. However, in planta assays using tissue-cultured turmeric plants revealed that only the endosphere-derived bacterial community, including Chryseobacterium gleum (ED4), Pseudomonas laurentiana (ED4-21), and Pantoea sp. (WEH1), significantly reduced pathogen populations, resulting in a two-log reduction in pathogen abundance. These findings suggest that colonization within plant tissues may contribute to suppression of vascular pathogens and highlight endophytic bacteria as candidates for further investigation in microbiome-based management of bacterial wilt.

Alternaria brassicicola is the predominant causal agent of Alternaria leaf blight and head rot (ABHR) in organic broccoli production in the Northeastern United States. A prior regional study indicated a high level of genetic diversity within individual fields, which we sought to investigate here to understand important routes of spread and mechanisms of reproduction. We genotyped 171 isolates collected from three certified organic farms in Connecticut across two growing seasons (2022 and 2023) using microsatellite markers. Results revealed high genotypic diversity and significant linkage disequilibrium, indicating a clonal population structure. Discriminant analysis of principal components (DAPC) and hierarchical analysis of molecular variance (AMOVA) showed low but significant genetic differentiation among fields and years, with most variation occurring within populations. Shared multilocus genotypes (MLGs) across fields and years suggest that diversity is primarily driven by multiple introductions, likely via contaminated seed, rather than recombination. These findings underscore the importance of seed source in pathogen dispersal and highlight the need for improved seed testing and cultural management practices to mitigate ABHR in organic broccoli systems.

Phytophthora sansomeana was detected in several soybean root rot surveys in the United States alongside another well studied oomycete, P. sojae. We compared the transcriptomic response among three P. sansomeana isolates that differed in aggressiveness and P. sojae isolate OH25 to to the most aggressive P. sansomeana isolate in a moderately susceptible soybean using dual host-pathogen RNA-Seq. At each time point, relative pathogen read abundance was higher following inoculation with P. sojae than P. sansomeana. Differentially expressed genes (DEGs) relative to mycelial samples was more pronounced for P. sojae than P. sansomeana. More than a hundred genes were significantly upregulated across all three P. sansomeana isolates for all 3 time points. These included genes associated with cell wall degradation, jacalin-like lectin, NmrA-like family, one RxLR, and amino acid auxin permease family. For P. sansomeana, more genes annotated as CRN were downregulated and fewer Necrosis inducing peptides (NPL) were upregulated compared to P. sojae, which could explain in part the difference in necrosis symptom development between the two species. Additionally, P. sojae had more DEGs associated with pathogenicity than the most aggressive P. sansomeana isolate. Several key defense pathways were upregulated earlier in response to the three P. sansomeana isolates than to P. sojae, including glucosinolate and glutathione biosynthesis, sulfur and tropane, piperidine and pyridine alkaloid metabolism, which may also explain the reduced lesion sizes. These results provide a framework for using effector screening to select for effective host resistance against populations of both P. sansomeana and P. sojae.

Plants restrict microbial entry into their leaves by closing their stomata upon recognition of conserved microbe-associated molecular patterns (MAMPs). The hydathode pore is a stomata-like opening on leaf margins which are thought to lack MAMP recognition and to be an entry point for microbial pathogens. Here, we observe marginal hydathode pore closure in response to abscisic acid, and the MAMPs chitin and flg22 in Arabidopsis thaliana leaves. Hydathode pore closure occurs within 3-9 hours of chitin exposure and pores reopen after 12 hours. Under conditions when hydathode pores are open, external fluids enter approximately 80% of the marginal hydathodes within a leaf. After hydathode pores close in response to ABA, chitin or flg22, external fluids accumulate in under 20% of hydathodes with in a leaf. MAMP-induced hydathode restriction was similar for dye or fluorescent Pseudomonas syringae pv. tomato bacteria and was dependent on pattern recognition receptors including CERK1 for chitin and FLS2 for flg22. Chitin-induced hydathode limitation was also dependent on the NADPH oxidase RBOHF and was partially hampered in rbohD, lyk4 lyk5, bak1-5 bkk1, slac1-3 slah3-1 knockout mutants. Together, this work indicates that MAMP recognition regulates entry into the hydathode and induces transient closure of the hydathode pores.