Ginseng Alternaria leaf and stem blight, caused by Alternaria panax, imposes substantial yield and economic losses to the ginseng cultivation industry. Current diagnostic methods for ginseng diseases primarily rely on pathogen isolation from infected tissues, a procedure that is laborious, time-consuming, and inherently low in sensitivity. This study has therefore developed a rapid, specific and sensitive SYBR Green-based quantitative real-time PCR (qPCR) assay for detecting A. panax in plants, seeds, and soil. The developed qPCR assay exhibited high sensitivity and repeatability, with a detection limit of 0.074 fg/μL of target amplicon DNA (0.619 ng/μL of genomic DNA) and a coefficient of variation below 2%. In artificially inoculated tissues (leaves, stems and seeds), Ct values decreased progressively with increasing incubation time, reflecting pathogen proliferation. Analysis of field-collected leaves and stems showed a strong overall correlation between Ct values and visual disease grades. Surveying of ginseng-growing areas revealed that A. panax was detected in asymptomatic leaves and stems at rates of 12.12% and 14.29%, respectively, and in 14.46% of soil samples and 23.73% of seed samples. This qPCR assay presented here provides a robust tool for forecasting early disease, tracking the primary inoculum of the pathogen and its transmission chains, and screening of both ginseng seed lots and candidate soils for ginseng Alternaria leaf and stem blight prior to planting.

Damping-off disease caused by Rhizoctonia solani is a devastating soilborne disease that poses a serious threat to sustainable ginseng (Panax ginseng) production. In an effort to develop environmentally friendly control alternatives, we investigated the volatile constituents of Asarum heterotropoides essential oil using GC-MS in 2023 and identified methyleugenol as the predominant bioactive component, accounting for 21.32% of the total volatile fraction. Mycelial growth rate assays demonstrated that methyleugenol exerted pronounced antifungal activity against R. solani, with EC50 and EC90 values of 60 mg/L and 800 mg/L, respectively. Treatment at 0.1 mg/mL resulted in 51% growth inhibition after 4 days of incubation. To elucidate the underlying mechanisms of action, R. solani mycelia exposed to 0.1 mg/mL methyleugenol were harvested at 4 and 6 days post-treatment for integrated transcriptomic and untargeted metabolomic profiling. Transcriptome analysis revealed 1444 and 1157 significantly downregulated genes at 4 and 6 days, respectively, with 836 genes consistently repressed at both time points. GO and KEGG enrichment analyses indicated that these persistently suppressed genes were predominantly associated with protein processing in the endoplasmic reticulum, protein export, tyrosine metabolism, and peroxisome biogenesis. Metabolomic profiling identified 95 differentially accumulated metabolites that were commonly reduced across both time points, with KEGG pathway enrichment highlighting significant suppression of starch and sucrose metabolism and galactose metabolism pathways central to carbon source utilization. Integrative transcriptome–metabolome correlation analysis further uncovered three key transcription factors (gene-RDB_LOCUS74145, gene-RDB_LOCUS26344, and gene-RDB_LOCUS52181) implicated in the regulation of carbohydrate metabolism and tricarboxylic acid cycle-related metabolite production. Collectively, our findings indicate that methyleugenol suppresses R. solani growth through a multi-target mechanism involving impairment of carbon source metabolism, disruption of protein processing and secretion, and attenuation of cellular energy supply. These results provide a mechanistic foundation for the development of botanical fungicide-based strategies for the green management of ginseng damping-off disease.

Ginseng Alternaria leaf and stem blight (GALSB), caused by Alternaria alternata, poses a severe threat to ginseng cultivation. Although azoxystrobin is a cornerstone fungicide for GALSB management, the emergence of widespread adaptive resistance has severely curtailed its field efficacy. This study integrated molecular, transcriptomic, and genetic approaches to unravel the underlying resistance mechanisms. Targeted gene sequencing and molecular docking revealed that resistant strains harbor a conserved G143A point mutation in the AaCyt b protein. This mutation weakens the azoxystrobin–AaCyt b protein binding affinity by decreasing the binding energy from −8.31 to −7.08 kcal/mol. Additionally, comparative transcriptomics and RT-qPCR demonstrated pronounced upregulation of the alternative oxidase gene (AaAOX) and core energy metabolism pathways in resistant strain TYC8-2, with AaAOX expression increasing 4.45–6.91-fold. Fungicidal inhibition of AOX via salicylhydroxamic acid (SHAM) restored fungal sensitivity, increasing azoxystrobin sensitivity by 11.66-fold. Crucially, genetic knockout of AaAOX enhanced sensitivity by approximately 2.7 × 104-fold. Phenotypic assays further established AaAOX as a multifunctional regulator; the AaAOX mutant exhibited attenuated virulence on ginseng leaves and increased sensitivity to oxidative and osmotic stresses (NaCl, H2O2, NaAc). We conclude that the G143A mutation in AaCyt b and the transcriptional overexpression of AaAOX act independently to drive azoxystrobin resistance in A. alternata. These findings provide comprehensive mechanistic insights to guide resistance monitoring, optimize fungicide applications, and develop precision strategies for GALSB management.

Anthracnose, primarily caused by Colletotrichum panacicola, is a major above-ground disease affecting Panax species. However, a rapid and accurate quantitative molecular detection method has been lacking. To address this, we developed a specific qPCR assay for the first time. Primers (q11329F/q11329R) were designed based on pan-genomic analysis of sequence of C. panacicola from NCBI, and a standard curve was established using recombinant plasmids. The assay successfully detected and quantified C. panacicola in artificially infected ginseng leaves and seeds, with monitoring of pathogen DNA accumulation during infection. It effectively identified C. panacicola in both symptomatic and asymptomatic naturally infected seeds. This qPCR method demonstrated superior efficiency, accuracy, and with greater sensitivity compared to conventional PCR. Furthermore, the assay was validated on symptomatic seed samples collected in 2024 from 16 ginseng-producing areas across Jilin and Liaoning provinces of China. This study provides a robust tool for the early diagnosis and quantification of C. panacicola, facilitating real-time disease surveillance and management.

Ginseng Alternaria leaf and stem blight (GALSB), caused by Alternaria spp., is a significant aboveground disease that severely impacts both the yield and quality of ginseng. Tebuconazole has been widely used for many years as a registered fungicide to control GALSB; however, its efficacy has declined in recent years. To elucidate the resistance mechanism of A. alternata to tebuconazole, this study assessed the sensitivity of A. alternata isolates using the mycelial growth rate method, evaluated resistance levels and risk, and investigated resistance mechanisms. The results showed a baseline sensitivity of A. alternata to tebuconazole at 3.90 μg/ml, with a low resistance frequency of 11.30%, and no moderately or highly resistant isolates were detected. Three genetically stable resistant mutants were obtained through fungicide adaptation. Compared with their parental strains, these mutants exhibited reduced mycelial growth, conidial production and germination, and pathogenicity, yet showed enhanced tolerance to temperature fluctuations and osmotic stress. The resistance risk was classified as low, and the mutants showed weak competitiveness under field conditions. A G462S point mutation in the CYP51 gene was identified in the resistant strains. Molecular docking analysis confirmed that this mutation reduces the binding affinity between tebuconazole and its target protein (CYP51). Expression analysis revealed significant upregulation of the CYP51 gene in the resistant mutants compared with the parental strain. In summary, A. alternata exhibits a low level and risk of resistance to tebuconazole; point mutation and overexpression of the CYP51 gene are the primary mechanisms contributing to the development of resistance. These findings provide a scientific basis for the rational use of tebuconazole in controlling GALSB. Monitoring these resistance mechanisms will facilitate the development of effective resistance management strategies, such as rotation and combined application of fungicides, to achieve sustainable and environmentally friendly disease control.

To tackle the QoI fungicide resistance of Alternaria spp. (the causal agent of ginseng Alternaria leaf and stem blight (GALSB)), this study systematically assessed the antifungal activity and biochemical effects of fluopyram·tebuconazole on Alternaria spp. via the mycelial growth rate method and conidia germination assay. It also evaluated protective and curative efficacy of fluopyram·tebuconazole (FT) 35% suspension concentrate (SC) against GALSB under in vitro and field conditions. Results demonstrated that FT significantly inhibited mycelial growth in three Alternaria species (A. alternata, A. tenuissima, and A. panax) and conidial germination in two Alternaria species (A. alternata and A. tenuissima). Notably, its inhibitory activity against mycelial growth was more potent than that against conidial germination. FT treatment disrupted mycelial growth, ergosterol/DNA/protein biosynthesis, increased cell membrane permeability, damaged membrane integrity, and downregulated resistance-related genes (AOX, P450) and activity of AOX. No positive cross-resistance was observed between FT and other eight commonly used GALSB fungicides (including azoxystrobin). Under in vitro conditions, FT 35% SC at medium-to-high doses achieved protective efficacy (86.75-95.81%) and curative efficacy (88.61-98.35%) against GALSB, outperforming azoxystrobin at equivalent doses. One-year field trials at three sites showed medium-high FT 35% SC doses provided 83.38-85.05% (7 days), 79.59-83.42% (15 days), and 70.44-76.87% (30 days) average disease control after final application. These values exceeded those of control fungicides pyraclostrobin, azoxystrobin, kresoxim-methyl by 10.96-34.38%, 12.51-45.18%, and 19.93-36.29%, respectively. Notably, FT 35% SC retained >70% efficacy 30 days after the final spray, indicating prolonged residual activity. Additionally, FT 35% SC increased ginseng yield by 10.09-16.71%. In summary, FT exhibits potent antifungal and biochemical activity against Alternaria spp., making it a viable alternative to QoI fungicides for GALSB control and resistance management.

Ginseng rusty root rot, caused by Ilyonectria robusta, is the most severe soil-borne disease affecting ginseng cultivation. A synthetic microbial community (SynCom) was developed for the first time using antagonistic and growth-promoting endophytic and rhizosphere bacterial strains to control ginseng rusty root rot, and its synergistic mechanisms had been clarified. The 89 out of 98 biocontrol bacterial groups exhibited synergistic inhibitory effects against I. robusta, while 40 consortia showed both growth-promoting and synergistic effects. Among these, the 24 consortia inhibited mycelial growth of I. robusta by more than 80%, and seven consortia suppressed spore growth by over 60%. The inhibition rates were increased by 19%-131% and 5%-137%, respectively, compared with single strain treatment. The biocontrol consortia achieved superior control efficacy against ginseng rusty root rot in the field, which six consortia exhibited control efficacy exceeding 50%, with the consortium CL95 showing the highest efficacy at 66.67% and significantly promoted ginseng growth. The content of I. robusta in ginseng roots decreased by 46.97% following CL95 treatment compared to the singlestrain NJ13. Mechanistic studies indicated that the colonization efficiency of the consortia in ginseng tissues and rhizosphere soil was significantly higher than that of single strains within 63 days. The colonization of CL95 in ginseng leaves, stems, epidermis, root core, lateral root, fibrous root and soil was 33%-215% higher than that of single strain NJ13, respectively. In addition, compared with NJ13, the four soil enzyme activities were increased by 100%-150%, and the expression levels of five defense enzyme genes in three different growth periods of ginseng were increased by 75%-167%, among which CL95 had the most obvious effect. It was also found that biocontrol consortia significantly increased the diversity of microbial communities and the abundance of beneficial fungi and bacteria in ginseng rhizosphere soil, while significantly reducing the abundance of pathogenic fungi. Correlation analysis showed that the consortia were significantly positively correlated with root weight and soil enzyme activity, promoting ginseng growth. This study lays a theoretical foundation for overcoming the limitations of single-strain biocontrol agents and highlight the potential for industrial application of biocontrol consortia in sustainable agriculture.

Abstract Ginseng rusty root rot caused by Ilyonectria robusta threatens ginseng production. This study screened synergistic combinations of biocontrol agents and chemical fungicides. Combined treatments inhibited mycelial growth, yield, and conidial production and germination. Among nine combinations, five achieved over 50% control efficacy; NJF 5:1 reached 81.48% field efficacy and reduced I. robusta in roots and soil. Biocontrol strain colonization increased versus singlestrain treatments. Ginseng defense enzyme genes and soil enzyme activities were enhanced. Three-stage soil microbiomics showed improved diversity, increased beneficial Granulicella and Pseudogymnoascus, and decreased Ilyonectria pathogen, which positively correlated with disease index, while beneficial genera correlated with soil enzyme activity. Seven lipopeptide biosynthesis genes were upregulated by the combination versus single strain. Metabolomics and transcriptomics confirmed that fludioxonil promoted the accumulation of antifungal p-hydroxybenzaldehyde and 4-methylphenol in NJ13. This study presents a novel synergistic interaction model reducing chemical usage and clarifies multiple synergistic effects against ginseng rusty root rot.

Fusarium root rot on ginseng is an important root disease that seriously affects ginseng (Panax ginseng) yield and quality. However, the species categories and distribution of Fusarium causing ginseng root rot in China have not been systematically examined. A total of 571 pure Fusarium isolates were obtained from 2018 to 2019 from 14 ginseng-producing regions in Jilin, Liaoning, and Heilongjiang provinces, China. Based on multilocus sequence analysis of ITS-tef1-rpb2 and morphological characteristics, the 571 Fusarium isolates were identified as F. oxysporum (accounting for 47.46% of the total isolates), F. solani (35.38%), F. equiseti (5.78%), F. proliferatum (2.80%), F. cerealis (1.75%), F. semitectum (1.75%), F. acuminatum (1.75%), F. redolens (1.58%), F. verticillioides (1.05%), and F. graminearum (0.70%). Among them, F. oxysporum and F. solani were the dominant species, and F. graminearum and F. verticillioides were first recorded on ginseng in China. The fungicides captan and hymexazol were tested in vitro for their inhibitory activities against 10 Fusarium species. All Fusarium species displayed enhanced sensitivity to captan compared with hymexazol. The protective effects of captan against ginseng root rot caused by F. oxysporum ranged from 94.07 to 97.78% at concentrations of 200, 400, and 600 μg·ml-1; however, its curative effects were much lower, ranging from 29.63 to 35.56%. In comparison, hymexazol exhibited protective effects of only 24.00 to 52.00% and curative effects of 16.00 to 45.33% at concentrations of 600 to 1,000 μg·ml-1. In addition, microconidia were highly sensitive to both fungicides, compared with mycelia and macroconidia.

Bacillus mycoides JA20-1 was screened and identified as a biocontrol bacterium with a high capacity for producing volatile organic compounds(VOCs) in the laboratory. This strain had significant inhibitory effects on various postharvest disease pathogens in crops, such as Botrytis cinerea, as well as soil-borne disease pathogens in ginseng, such as Sclerotinia ginseng. In order to accelerate its industrialization process, in this study, single-factor experiments and response surface optimization methods were used. The fermentation medium and fermentation conditions in the shake flask of strain JA20-1 were systematically optimized by using cell production volume as the response variable. Meanwhile, the biocontrol effect of JA20-1 on B. cinerea of ginseng during the storage period was evaluated by using the method of fumigation in a dry dish in vitro. The results indicated that the optimal fermentation medium formulation for strain JA20-1 was as follows: 1% yeast paste, 1% soluble starch, 0.25% K_2HPO_4·3H_2O, and 0.2% NaCl. The optimal fermentation conditions in the shake flask were vaccination size of 3%, culture volume of 50 mL in a 250 mL Erlenmeyer flask, pH of 6.2, fermentation temperature of 34 ℃, shaking speed of 180 r·min~(-1), and incubation time of 18 hours. The bacteria count in the fermentation broth under these conditions reached 2.17 × 10~8 CFU·mL~(-1), which was 6.58 times higher than before. The average control efficacy of the fermentation broth on Botrytis cinerea of ginseng under in vitro fumigation reached 61.70% and 84.04% respectively, when 20 mL and 30 mL per dish were used. The research provided theoretical support and technical foundation for the development and utilization of Bacillus mycoides JA20-1 and the biocontrol of soil-borne diseases in ginseng and postharvest diseases in crops.

Gray mold, caused by Botrytis cinerea, is a globally devastating plant disease that severely threatens the economic sustainability of ginseng cultivation in Northeast China. Our previous field studies demonstrated that Bacillus amyloliquefaciens FS6 (BaFS6) exhibits strong biocontrol efficacy against ginseng gray mold. However, the underlying mechanism of its antifungal activity remained unclear. In this study, we identified an antifungal polypeptide (AFP) from Ba. amyloliquefaciens FS6 through comparative proteomic analysis. Prokaryotic expression of the afp gene in Escherichia coli yielded a recombinant protein (RP BaAFP) that significantly inhibited Bo. cinerea hyphal growth, conidial germination, germ tube elongation and disruption of cell wall integrity. Notably, RP BaAFP exhibited exceptional stability across a broad pH range (4-9), tolerated 2-8 h of UV irradiation, and retained activity at elevated temperatures (40-80 degrees C), outperforming the reference protein bovine serum albumin (BSA) in stability. Fermentation broth (FB) from Ba. amyloliquefaciens FS6 strain overexpressing afp (FS6-BaAfpOE) demonstrated superior antifungal activity compared to the wild-type Ba. amyloliquefaciens FS6 FB. Both RP BaAFP and the FS6-BaAfpOE cell-free FB effectively protected detached ginseng tissues and whole plants in greenhouse trials, significantly reducing Bo. cinerea infection. These findings elucidate the functional role of AFP in Ba. amyloliquefaciens FS6-mediated biocontrol and highlight its potential, along with the FS6 strain, for sustainable management of ginseng gray mold.

Sclerotinia root rot is one of the most destructive soilborne diseases in ginseng (Asian ginseng) production regions in China. To identify the causal pathogens, 409 Sclerotinia isolates were recovered from ginseng plants at 11 plantations in Jilin, Heilongjiang, and Liaoning Provinces in China from 2018 to 2021. Based on morphological characteristics, sequence analyses of the internal transcribed spacer region and the beta-tubulin gene, as well as a phylogenetic analysis, the isolates were identified as S. nivalis (n = 268), S. ginseng (n = 114), and S. sclerotiorum (n = 27). S. nivalis and S. sclerotiorum were found on fields growing Panax ginseng for the first time in China. Bacillus amyloliquefaciens FS6 (CGMCC 9538) exerted significant antifungal effects on all three Sclerotinia spp. by inhibiting in vitro mycelial growth by 85%, resulting in deformed and broken hyphae and dissolving sclerotia; on average, >73% inhibition (sclerotia formation) was achieved by FS6. Notably, the germination of sclerotia was inhibited for up to 20 days post-treatment in the fermentation broth (FB) of FS6 at concentrations of 4 × 106 to 4 × 108 CFU·mL-1. Using those FS6 FB concentrations in vitro resulted in control efficacy of 43.48% to 100.00% against Sclerotinia root rot. Field trials in 2021 and 2022 showed that 107 to 108 CFU·mL-1 FS6 FB sprayed on ginseng leaves three times every 7 days exhibited a promising control effect against S. nivalis, with a control efficiency ranging from 53.00% to 73.51% 15 days after the third application. This study clarified that Sclerotinia root rot affecting Asian ginseng in Northeast China is caused by three species of Sclerotinia, for which B. amyloliquefaciens FS6 is a promising biocontrol agent.IMPORTANCESclerotinia root rot typically causes substantial losses in ginseng yield. This study aims to identify the types of pathogens that cause ginseng sclerotinia rot and their distribution in various production areas. This study is the first to report S. nivalis as a dominant species causing sclerotinia root rot in Asian ginseng cultivated in China. The most effective method for the prevention and control of sclerotinia root rot is biological control. Our findings suggest that B. amyloliquefaciens FS6 could be a promising way to control sclerotinia disease prevalent in commercial ginseng plantations in China.

Forty percent prothioconazole-tebuconazole suspension concentrate(SC)has been registered to control multiple crop diseases in China.This study aims to evaluate its efficacy for the control of Alternaria leaf spot of ginseng,so as to lay the foundation for the management of resistance of Alternaria spp.causing Alternaria leaf spot of ginseng to fungicides.The study first detected its antifungal activity against three species(A.alternata,A.panax,and A.tenuissima)and biochemical activity against A.alternata and then determined its efficacy against Alternaria leaf spot of ginseng in vitro.Field experiments lasting for one year at three locations were conducted to evaluate its field efficacy against Alternaria leaf spot of ginseng.Spearman's correlation analysis was used to determine the cross-resistance of 40％prothioconazole-tebuconazole SC with eight commonly used fungicides.The results showed that prothioconazole-tebuconazole had a good inhibitory effect on mycelial growth and spore germination of the three Alternaria species,and the inhibitory effect on mycelial growth was stronger than that on spore germination.The treatment with prothioconazole-tebuconazole at the concentrations of 0.1,1.0,and 10.0 μg·mL-1 resulted in deformity of mycelia,shortened germ tubes of conidia,reduced content of DNA and soluble proteins,and decreased permeability of cell membrane of A.alternata.Under in vitro conditions,the protective and curative effects of 40％prothioconazole-tebuconazole SC in the concentration range of 50-150 μg·mL-1 on Alternaria leaf spot of ginseng were 65.45％-100.00％and 65.58％-96.36％,respectively,which were significantly better than those of tebuconazole at the same concentration.There was no cross-resistance between prothioconazole-tebuconazole and the eight fungicides(difenoconazole,propiconazole,tebuconazole,iprodione,pyraclostrobin,azoxystrobin,kresoxim-methyl,and mancozeb).These eight fungicides were widely used to prevent and control Alternaria leaf spot of ginseng,and A.panax causing Alternaria leaf spot of ginseng had resistance to them.The results of the field experiments at three locations showed that 40％prothioconazole-tebuconazole SC was more effective than difenoconazole,propiconazole,tebuconazole,and iprodione 7,15,and 30 d after its last application and had long effective duration.In conclusion,40％prothioconazole-tebuconazole SC had high antifungal activity and biochemical activity against Alternaria spp.and better efficacy for the prevention and control of Alternaria leaf spot of ginseng in the field than the commonly used fungicides to which Alternaria spp.were resistant,and no cross-resistance was found between 40％prothioconazole-tebuconazole SC and the fungicides.It can be used for resistance management of Alternaria spp.

Introduction:Fusarium solani is a widespread plant pathogen known to damage numerous crops, including causing severe root rot in Panax ginseng. In this study, we identified a novel ourmia-like mycovirus in F. solani, named "Fusarium solani ourmia-like virus 1" (FsoOLV1). We demonstrated that FsoOLV1 confers hypovirulence in its host F. solani and three other ginseng root rot pathogens, F. oxysporum, F. proliferatum, and F. verticillioides. Additionally, we verified its horizontal and vertical transmission capabilities. Methods:FsoOLV1 was discovered in F. solani strain SJH 2-4 using high-throughput sequencing. The genome sequence of FsoOLV1 was obtained through RT-PCR and RACE. Virus elimination was conducted to assess the effect of FsoOLV1 on fungal virulence. Protoplast transfection experiments were performed to evaluate the impact of the virus on other ginseng root rot pathogens. Horizontal and vertical transmission studies were also carried out to examine the spread of the virus. Results:The genome of FsoOLV1 is 2,801 nucleotides (nt) in length with a GC content of 47.05%. It encodes a 750 amino acid RNA-dependent RNA polymerase (RdRp), with a molecular weight of approximately 84.84 kDa. Phylogenetic analysis indicated that FsoOLV1 clusters within a clade containing the Magoulivirus genus in the Botourmiaviridae family. Curing FsoOLV1 from the fungal host strain revealed that the mycovirus plays a role in reducing the virulence of the F. solani strain SJH 2-4. Furthermore, protoplast transfection revealed that FsoOLV1 can significantly reduce the virulence of other ginseng root rot pathogens, including F. oxysporum, F. proliferatum, and F. verticillioides. Additionally, FsoOLV1 is capable of horizontal transmission between F. solani strains and vertical transmission to the next generation via conidia. Discussion:This study presents the first hypovirulent ourmia-like virus, FsoOLV1, which reduces the virulence of F. solani and other ginseng root rot pathogens. Our findings suggest that FsoOLV1 could serve as a promising biological agent for controlling ginseng root rot. This research not only expands the diversity of known hypovirulent mycoviruses but also provides a potential candidate for controlling Fusarium diseases in ginseng cultivation.

Ginseng Alternaria leaf and stem blight (GALSB), caused by Alternaria spp., poses a serious threat to ginseng production. Alternaria spp. has developed resistance to currently used fungicides. To evaluate the efficacy of pyraclostrobin & sdot;tebuconazole (PT) 30 % SC for the control of GALSB disease, and lay the foundation for the management of resistance of Alternaria spp., the study was conducted to evaluate the effectiveness of PT in controlling GALSB on detached ginseng leaves and in one-year field experiments, based on its inhibitory and biochemical activities against A. alternata, A. panax, and A. tenuissima. The cross-resistance of PT with eight widely used fungicides was ascertained using Spearman's correlation analysis. PT has a better inhibitory effect on the mycelium growth and conidial germination of Alternaria spp., while having no observable effect on mycelium deformity. However, PT significantly shortened the germ tube, reduced the DNA and soluble protein contents, and decreased the permeability of the cell membrane of Alternaria spp. PT displayed significant protective (66.26 %-98.92 %) and curative (60.08 %-99.31 %) effects against GALSB on detached leaves at 50-150 mu g/mL, outperforming tebuconazole (p < 0.05). Field trials confirmed that PT had a superior control efficacy at medium to high doses compared to difenoconazole, propiconazole, tebuconazole, and iprodione at seven, 15, and 30 days following the last application. Crucially, PT had no cross-resistance with the other evaluated fungicides. These findings highlight that PT provides effective protection and treatment against GALSB and, importantly, is a candidate for mitigating fungicide resistance development in Alternaria spp. associated with GALSB control, with potential as a rotation or replacement fungicide in resistance management programs.

To manage the developing resistance of Alternaria spp. [the causal fungi of ginseng Alternaria leaf and stem blight (GALSB)] to QoIs fungicides, the toxicity and biochemical activity of pyrimidine nucleoside antibiotics (PNA) against Alternaria spp., cross-resistance between PNA and eight other fungicides currently used to control GALSB disease, and the efficacy of PNA for controlling GALSB in vitro and in vivo were investigated. The distributions of EC50 values of PNA against the mycelial growth (115 isolates) and conidia germination (89 isolates) of A. alternata were unimodal, with mean EC50 values of 10.192 ± 4.961 μg/mL and 0.828 ± 0.101 μg/mL, respectively. There were no significant correlations between the sensitivity of A. alternata to PNA and eight other fungicides (p < 0.05). PNA caused morphological changes in A. alternata mycelia and germ tubes, increased cell membrane permeability, and reduced intracellular DNA and protein levels. On detached ginseng leaves, 300 μg/mL PNA achieved mean protective and curative effects of 87.93% and 94.77% against A. alternata 7 days post-inoculation, outperforming that of 300 μg/mL kresoxim-methyl. Field trial results showed that PNA (180 g a.i./hm2) achieved mean efficacies of 85.63%, 84.07%, and 72.55% at three sites 7, 15, and 30 days after the last spray, which were 5.28–37.74% higher than those of control fungicides pyraclostrobin, azoxystrobin, and kresoxim-methyl at corresponding time points. Overall, our findings indicate that PNA are effective agents for the management of Alternaria spp. resistance to QoIs fungicides.

Fusarium wilt of tobacco (FWT) is a globally significant disease responsible for major losses in tobacco yield and quality. A total of 320 Fusarium oxysporum species complex (FOSC) isolates derived from five tobacco-growing regions in Jilin province, China, have been confirmed as the predominant pathogens causing FWT, including F. tobaccum, F. cugenangense, F. odoratissimum and F. foetens. In this study, the results indicated that differences in the virulence of the representative isolates from tobacco from the same Fusarium species were significant. Aside from infecting tobacco, the virulence of these representative FOSC isolates differed on chilli and other crops, and they were not pathogenic or were weakly pathogenic to a further 20 crops. Six strains of formae speciales (ff. sp.) of F. oxysporum had the strongest virulence on their original hosts and were not or weakly pathogenic to tobacco. Joint analysis of the pg1-pg5-pgx1-pgx4 genes indicated that 21 isolates of F. cugenangense were divided into four branches, which were clustered with different ff. sp. of F. oxysporum; five isolates of F. foetens were clustered in a branch with f. sp. fragariae, which were not pathogenic to strawberries; six isolates of F. tobaccum and five isolates of F. odoratissimum comprised independent branches and did not cluster with any other f. sp. of F. oxysporum. In sum, none of the FOSC isolates causing FWT in Jilin exhibited tobacco specificity. Our findings further confirmed that FOSC isolates from tobacco could be distinguished into F. tobaccum, F. cugenangense, F. odoratissimum and F. foetens.

The objective of this study was to identify effective agents for the prevention and control of ginseng Sclerotinia root rot disease caused by Sclerotinia nivalis. The inhibitory effects of 16 chemical fungicides and 10 biocontrol agents (strains) on mycelial growth and sclerotium formation in S. nivalis were determined using a plate confrontation essay. The results showed that the best chemical agents for inhibiting the mycelial growth and sclerotium formation of S. nivalis were fluconazole and fludioxonil, while Bacillus amyloliquefaciens FS6 and B. subtilis (Kono) were the best biocontrol agents (strains). The results of field trials in 2022 and 2023 showed that the control effects of fluconazole and fludioxonil on ginseng Sclerotinia root rot disease were 90.60–98.16%, and those of the biocontrol agents B. amyloliquefaciens FS6 and B. subtilis (Kono) were 94.80–97.24%, respectively. Chemical agents produced abnormal and twisted mycelia, while the biocontrol agents increased mycelial branching, dilated the mycelium tip, and revealed an abnormal balloon. All of the fungicides decreased the ergosterol content, changed the cell membrane permeability, and increased the protein and nucleic acid permeability. These results suggest that these are potential agents for controlling ginseng Sclerotinia root rot disease, and their biochemical mechanisms of chemical and biocontrol of this disease were demonstrated.

A novel mitovirus, tentatively designated as "Fusarium oxysporum mitovirus 2" (FoMV2), was isolated from the pathogenic Fusarium oxysporum f. sp. ginseng strain 0414 infecting Panax ginseng. The complete genome of FoMV2 is 2388 nt in length with a GC content of 30.57%. It contains a large open reading frame (ORF) encoding a putative RNA-dependent RNA polymerase (RdRp) of 713 amino acids with a molecular weight of 83.05 kDa. The sequence identity between FoMV2 and Botrytis cinerea mitovirus 8 and Fusarium verticillioides mitovirus 1 was 87.94% and 77.85%, respectively. Phylogenetic analysis showed that FoMV2 belongs to the genus Unuamitovirus in the family Mitoviridae. To the best of our knowledge, this is the first report of an unuamitovirus isolated from F. oxysporum f. sp. ginseng causing ginseng root rot.

ABSTRACT Fusarium wilt of tobacco (FWT), caused by Fusarium spp., has emerged as a severe threat to tobacco production in China. In all, 132 isolates of Fusarium were isolated from tobacco and pathogenic to tobacco-causing FWT in Jilin Province, China. In this study, we identified 7 of 132 isolates as a novel species Fusarium tobaccum sp. nov. Zhao Xie & Jie Gao, using multi-gene phylogenetic analyses of translation elongation factor ( tef1 ), β-tubulin ( tub2 ), calmodulin ( cmdA ), and RNA polymerase II second largest subunit ( rpb2 ) genes, along with subtle morphological differences. Isolates of F. tobaccum sp. nov. were clustered in a distinct branch in the maximum parsimony phylogenetic tree generated from the sequences of tef1-rpb2-tub2-cmdA and can be distinguished from closely related species F. cugenangense , F. callistephi , and F. elaeidis . The morphological characteristics of F. tobaccum sp. nov. are distinct from other Fusarium species. F. tobaccum sp. nov. exhibits abundant aerial mycelia and pigment production on potato dextrose agar (PDA), microconidia with 0–1 septa, and macroconidia with 2–5 septa on carnation leaf-piece agar (CLA), and produces abundant chlamydospores on Spezieller Nährstoffarmer agar (SNA) and CLA. The mycelia of F. tobaccum exhibited optimal growth at a pH of 7.1 and a temperature of 23.6°C. Sucrose and NaNO 3 significantly promoted the mycelial growth of F. tobaccum . PD medium was optimal for total sporulation. However, the sporulation ratios of the macrospores of F. tobaccum in PD, SN, and CMC were relatively low (0.48%, 2.51%, and 2.16%, respectively). These findings provided valuable insights into the morphological and biological characteristics of F. tobaccum . IMPORTANCE Fusarium wilt of tobacco (FWT) is a prevalent issue in tobacco-growing regions globally, leading to significant losses in yield and quality. This study identified F. tobaccum sp. nov., a novel species of Fusarium causing FWT in China. The identification was based on multi-gene phylogenetic analyses and morphological characteristics. The effects of temperature, pH, carbon source, nitrogen source, medium, and light on the mycelial growth of F. tobaccum sp. nov. were determined. These findings might contribute to future research on the pathogenic mechanisms of this novel species and the development of strategies to control FWT.