Gray mold, caused by the fungus Botrytis cinerea Pers., is one of the main diseases affecting strawberry, representing a problem during cultivation and post-harvest. The intensive use of chemical fungicides has favored the selection of resistant isolates to the products currently available on the market. This study aimed to establish a baseline sensitivity of Botrytis cinerea isolates from Brazil and the USA to mefentrifluconazole, assess potential cross-resistance with other DMI fungicides, identify mutations in the CYP51 gene associated with resistance, and evaluate the occurrence of practical resistance by comparing the efficacy of mefentrifluconazole with other commercially available fungicides. To build the baseline, the EC 50 of 155 isolates (70 isolates from Parana, Brazil and 85 from Florida, USA) was determined. Isolates from Parana were less sensitive than those from Florida, with two isolates presenting EC 50 values above 1.00 µg/ml. Average EC 50 of isolates from Parana was 0.169 µg/ml (0.046 to 1.303 µg/ml), while isolates from Florida averaged 0.084 µg/ml (0.030 to 0.384 µg/ml). Twenty out of 155 isolates were selected for a cross-resistance study, and their EC 50 values were determined for difenoconazole and triflumizole. Pearson's correlation analysis suggested the presence of cross-resistance between fungicides. DNA was extracted for three isolates with the highest and three with the lowest EC 50 for mefentrifluconazole, followed by PCR and sequencing of the CYP51 gene. All three high-EC 50 isolates carried mutations: two had the G461S mutation, and one had the R464K mutation. The efficacy of mefentrifluconazole was compared with cyprodinil and fludioxonil on detached fruit assays using three isolates (one sensitive, one with the G461S mutation and one with R464K). Mefentrifluconazole was effective against sensitive isolates, performing similarly to fludioxonil and cyprodinil, and represents a valuable option for gray mold management.

Fusarium wilt of strawberry is a widespread disease controlled through genetic resistance to Fusarium oxysporum f. sp. fragariae (Fof) race 1 conferred by the FW1 locus. In 2022, a novel race called CA Fof race 2 was identified in Oxnard, California (CA) that can overcome FW1 resistance and cannot be identified using published molecular diagnostic assays. To understand the prevalence and enable early detection of the novel CA Fof race 2, this research developed strain-specific qPCR and recombinase polymerase amplification (RPA) diagnostic assays and applied them to survey CA Fof race 2 in strawberry. A candidate locus was identified through alignment of reads from off-target isolates to genome assemblies of two virulent CA Fof race 2 isolates. Diagnostic qPCR and RPA tests were validated using target and off-target DNA. The RPA was validated in two laboratories to confirm reproducibility and used to test 2,945 diagnostic samples collected in strawberry-growing areas of the United States of America, Canada, and Mexico. Both assays showed 100% specificity based on laboratory screening of DNA. The limit of detection was 100 fg of DNA for the qPCR assay and 1 pg for the RPA assay. The RPA assay identified 19 out of 2,945 samples as positive for CA Fof race 2, all from six fields around Oxnard, CA; these results were confirmed by subsequent isolations and qPCR. Outcomes of this research provide essential diagnostic tools for quantifying and identifying CA Fof race 2 and highlight the limited geographic distribution of this novel resistance-breaking strawberry pathogen.

Tar spot, caused by Phyllachora maydis Maubl, has significantly impacted U.S. corn production since its first detection in 2015. The elusive nature of this fungus has hindered advancement in disease characterization, especially pertaining to its spatiotemporal development. This study provides an in-depth analysis of field-scale epidemics by considering both temporal and vertical dynamics to ascertain the spatiotemporal intensification of this disease. The analysis involved generating high-resolution severity data across four growing seasons (2021 to 2024) in two production-style corn fields in northwest Indiana. We then applied population growth models and Markov chains to parameterize the temporal and vertical dynamics of tar spot progression within the corn canopy. From this, we found three distinct epidemiological phases: an establishment phase characterized by sporadic onset with <0.5% severity, a lag phase with widespread low severity (0.5 to 1%), and an exponential phase with rapid severity increases exceeding 40% in some cases. Beyond the conventional "bottom-up" paradigm, we observed diverse infection-like patterns influenced by canopy position, onset timing, and corn growth stage. Exponential growth models described severity intensification with an average apparent infection rate of 0.20/day, which was applicable across canopy positions, locations, and years. An aggregated Markov chain model, built from the 2021-2023 data, accurately estimated the 2024 epidemic's vertical-temporal progression and thus validated the framework developed from this study. Ultimately, this approach supports improved surveillance for targeted management by establishing a foundation for real-time detection and probabilistic modeling to aid in the enhancement of crop production.

Management of the reniform nematode (Rotylenchulus reniformis Linford and Oliveira) in upland cotton (Gossypium hirsutum L.) continues to be a challenge primarily due to inconsistent performance of resistant genotypes. We hypothesize that diversity in reproduction and virulence in R. reniformis has contributed to the inconsistent performance of resistant cotton genotypes. Rotylenchulus reniformis isolates from Tennessee (TN), Alabama (AL), Mississippi (MS), Texas (TX), Louisiana (LA), Arkansas (AR), Florida (FL), South Carolina (SC) and Georgia (GA) were assessed for their reproduction and virulence on a resistant cultivar DP 2141NR B3XF and a susceptible cultivar DP 2317 B3TXF. The AR isolate had the greatest reproduction factor (Rf) of 19.7, while the AL isolate had the least Rf of 3.3. Rfs of other isolates were intermediate. Up to 85% of reproductions were suppressed by the resistant cultivar relative to the susceptible cultivar. While plants inoculated with nematodes had lower root biomass than the non-inoculated control, the degree of root biomass reduction varied by isolate, suggesting differing levels of virulence among R. reniformis isolates. The AR isolate reproduced the most and was the most virulent. The AL isolate reproduced the least and was less virulent. The AR, MS, TN and LA isolates were the top four reproducing isolates, probably indicating a greater suitability of the Delta region for nematode reproduction. The correlation analysis indicated that up to 85% variation in plant biomasses was explained by nematode reproduction. These results suggest that resistance breeding programs in cotton must consider the diversity in R. reniformis.

This study systematically evaluated cultivation requirements, host range, and chemical control options for Cercospora citrullina causing watermelon spot disease. Among 11 chemically defined media tested, Corn Meal Agar (CMA) medium supported optimal mycelial growth of C. citrullina, with an average radial growth rate of 56.72 ± 1.45 mm under controlled conditions (25°C, darkness). Host range determination via artificial inoculation of 19 plant species confirmed the host range of strain UNL090101 is limited to the Cucurbitaceae species tested., with watermelon (Citrullus lanatus) exhibiting the highest susceptibility, followed by melon (Cucumis melo), and cucumber (Cucumis sativus). Fungicide screening of 17 commercial formulations identified 40% iminoctadine tris (albesilate) WP (EC₅₀ = 2.82 μg·L⁻¹) and 64% mancozeb + 8% cymoxanil (WS) (EC₅₀ = 48.75 μg·L⁻¹) as the most effective treatments, achieving control efficacies of 74.47% and 58.62%, respectively. These findings provide actionable guidelines for optimizing crop rotation, inter-cropping strategies, and fungicide selection in watermelon production systems.

Plasmodiophora brassicae, an obligate Chromist pathogen that causes clubroot disease, occurs wherever brassica crops are grown. This review synthesizes recent advances in clubroot research, with an emphasis on clubroot management on canola (Brassica napus) in Canada. Fields infested with P. brassicae occur across the 8 M ha of annual canola production in Canada. Clubroot on canola is currently being managed effectively with a combination of genetic resistance and extended crop rotation, but virulent pathotypes are emerging rapidly and replacing resident pathotypes. Erosion of even a subset of available resistance genes could increase disease risk and impact across the region. New and diversified sources of genetic resistance are being developed and incorporated into the next generation of canola cultivars, supported by studies of host physiology, pathogen variability and resistance durability. However, the rapid change in pathotypes has resulted in concerns about the sustainability of a management strategy that relies almost exclusively on genetic resistance. This concern has stimulated studies in epidemiology, population biology, resistance stewardship, biological control and cultural management approaches such as soil amendments and enhanced surveillance. Genomic analyses of field collections and single resting spores of P. brassicae have demonstrated a predominance of clonal reproduction shaped by balancing selection and have enabled development of improved reference genomes. Pathotype identification methods, critical for matching canola cultivars with effective resistance profiles, continue to progress but more improvement is required. Continued innovation and integration of management strategies will be necessary to maintain effective long-term clubroot management.

Melon severe mosaic virus (MSMV) was identified in a pumpkin plant collected in Mississippi during the fall of 2022; the plant exhibited mosaic symptoms and frilly edges on leaves, symptoms typical of cucurbit plants infected with widely prevalent cucurbit-infecting viruses in the United States (U.S.). High-throughput sequencing was used for virus identification after routine assays for five widely prevalent mosaic-inducing viruses were negative. Screening of stored nucleic acid extracts from 53 additional cucurbit plants collected from Mississippi in 2021 and 2022 using RT-PCR assays revealed two additional positives for MSMV from 2022. MSMV was confirmed in the three positive samples using RT-qPCR and RT-PCR with amplicon sequencing. These results indicate that MSMV was present in the U.S. at least one year prior to its first report in the country. This example in cucurbits illustrates the importance of seeking diagnoses for plants exhibiting symptoms typical of commonly observed virus diseases as well as the need for routine virus monitoring with diagnostic assays capable of detecting multiple viruses, including exotic viruses. This approach allows for more rapid detection of virus introductions and, with appropriate action, could prevent or limit their establishment and spread in any given location or country.

Cyst nematodes, such as Heterodera schachtii (HS) and H. trifolii (HT), are major pathogens affecting kimchi cabbage production in Korea. Accurate species identification is essential for effective management, as these nematodes exhibit similar morphological features but differ in host range and pathogenicity. In this study, we developed a multiplex real-time TaqMan qPCR assay for the simultaneous detection of HS and HT. Species-specific primers and fluorescent probes targeting the coxI gene (HS) and a hypothetical protein gene (HT) were designed using MIG-seq-based polymorphism data. The assay demonstrated high amplification efficiency (HS: 89.3%, HT: 121.7%) and strong linearity (R² > 0.98) across serial DNA dilutions (1 ng to 100 fg). Specificity testing against 8 non-target plant-parasitic nematodes, including H. glycines, H. sojae, Meloidogyne incognita, and Pratylenchus penetrans, confirmed that no cross-reactivity occurred. Notably, the assay permits direct use of crushed cyst sap as a template, eliminating DNA purification. Detection was consistent from a single cyst, and reproducible across mixtures with varying ratios of HS and HT. The multiplex format reliably identified both species in co-infected samples without interference. This assay provides a rapid, sensitive, and field-adaptable molecular tool for the diagnosis of HS and HT, contributing to early detection and improved management of cyst nematodes in cabbage-growing regions.

Fusarium oxysporum is a representative soil-borne fungal pathogen that causes strawberry wilt, a disease characterized by an extremely high mortality rate that poses a severe threat to the sustainable development of the global strawberry industry. However, traditional detection methods are often time-consuming and dependent on specialized laboratory equipment, while existing soil nucleic acid extraction protocols are highly susceptible to interference from inhibitors in complex matrices, leading to low detection efficiency or high false-negative rates. To address these limitations, this study developed a novel on-site detection platform based on in-situ biological enrichment and purification-free nucleic acid release. A specialized enrichment rod targeting Fusarium oxysporum was developed to leverage the tropic growth characteristics of the pathogen, achieving physical separation from the soil matrix and effectively eliminating interference from complex soil inhibitors such as humic acids. The enriched pathogens release nucleic acids via a rapid lysis buffer, which are then neutralized and used directly as templates for RPA-CRISPR/Cas12b isothermal detection, enabling visual field identification through lateral flow strips. This method requires no specialized instrumentation, achieving a detection limit of 8.5 CFU/g and a sensitivity of 70 copies. After completing 48 hours of in situ enrichment, the entire process from rod retrieval to detection completion requires only 40 minutes (with hands-on operation time <10 minutes). By effectively circumventing soil inhibitor interference and simplifying complex nucleic acid extraction into a rapid, integrated protocol, this platform provides a critical technical solution for the on-site monitoring and precise control of soil-borne pathogens.

Brown root rot, caused by Phellinus noxius, is a major threat to rubber tree cultivation, resulting in substantial economic losses. Traditional control methods, such as root irrigation with fungicides, are labor-intensive, water-consuming, and inefficient, particularly in regions with limited water resources. This study introduces fluorescent mesoporous silica nanoparticles (FL-MSNs) as a novel delivery platform for tebuconazole to target P. noxius infection. The nanoparticles, with a high fungicide loading efficiency (43.81%), exhibit controlled release under varying pH conditions, providing sustained tebuconazole delivery. Fluorescent labeling allowed tracking of the nanoparticles, confirming effective transport from leaves to roots via phloem. Greenhouse trials demonstrated that foliar application of Teb@FL-MSNs resulted in an 84% survival rate of infected seedlings, significantly outperforming untreated controls, which exhibited 100% mortality. The nanoparticle formulation also reduced lesion size, highlighting its potential to improve disease control while minimizing fungicide use. These findings emphasize the advantages of Teb@FL-MSNs over traditional methods, offering a more efficient, environmentally friendly, and sustainable approach to managing brown root rot in rubber plantations.