Search Results (104)

Glomerella leaf spot (GLS), mainly caused by Colletotrichum fructicola, is a destructive disease of apple. However, the underlying pathogenesis mechanisms of GLS are still largely obscure. Previous infection transcriptome analysis showed that transcription factor CfGti1 was induced during leaf infection. The present study confirms that the CfGti1 gene is strongly expressed in conidia and early infection. To identify functions performed, we generated gene deletion mutant ΔCfGti1 by homologous recombination. Phenotypic analysis revealed that ΔCfGti1 lost pathogenicity to apple leaves by blocking appressorium-mediated host penetration, although penetration pegs still developed on cellophane. In addition, ΔCfGti1 colonization and hyphal extension in wounded apple fruit were dramatically decreased. The ΔCfGti1 mutant exhibited defects in growth and development of hyphae, which may be partly responsible for its inability to colonize apple. Comparative transcriptome and qRT-PCR analyses suggested that CfGti1 regulated appressorium-mediated host penetration by modulating genes related to metabolism of appressorial lipid droplets. Interestingly, CfGti1 also regulated the expression of ybtS and AKT1 or AFT1-1 related to biosynthesis of AK and AF host-specific toxins. This study demonstrated that CfGti1 is a pivotal regulator for apple GLS pathogenesis in C. fructicola. Full article

Biological control using beneficial microbes offers a sustainable alternative to chemical fungicides for managing postharvest diseases. This study reports the isolation and characterization of Bacillus amyloliquefaciens AsL-1 from the latex of Alstonia scholaris (L.) R. Br., unconventional ecological niche. The cell-free supernatant (CFS) of AsL-1 showed strong antifungal activity, inhibiting the growth of Colletotrichum musae (48.89 ± 0.57%), Glomerella cingulata (52.22 ± 0.00%), Fusarium graminearum (47.78 ± 0.57%), and Colletotrichum gloeosporioides (47.78 ± 0.00%) in vitro. Microscopy revealed that the CFS disrupted fungal development by blocking conidial germination and appressorium formation, and in C. gloeosporioides caused melanization defects linked to reduced virulence. In vivo tests on mango fruit confirmed that AsL-1 significantly decreased anthracnose lesion size and disease incidence. Protein analyses (SDS-PAGE, gel overlay, and LC-MS/MS) identified two antifungal proteins (24 and 16 kDa), corresponding to β-1,3-1,4-glucanase and flagellin. The detected β-1,3-1,4-glucanase activity indicates its role in degrading fungal cell walls and interfering with melanin biosynthesis pathways essential for pathogenicity. Overall, these findings highlight B. amyloliquefaciens AsL-1 as a promising protein-based biocontrol agent and show that latex-associated microbes may serve as valuable sources of new antifungal strategies. Full article

The rate of appressoria formation following conidial germination in Beauveria bassiana is closely associated with its pathogenicity. This study investigated the effects of insect cuticular compounds on the formation and metabolism in B. bassiana through the addition of insect cuticle analogues. Results indicate that both the fatty acid compound carnitine C3:0 and the organic acid compound Thiamine Pyrophosphate (TPP) exert dose-dependent bell-shaped effects on B. bassiana spore germination and appressorium formation at different concentrations. Both low and high concentrations inhibit spore germination and appendage formation. At a concentration of 0.10 mg/mL, spore germination and appendage formation rates peaked at all time points, being significantly higher than the control (p ≤ 0.05). Compounds in the benzene and its derivatives class, such as enilconazole and disulfide bis(2-hydroxy-3-methylpropan-2-yl) (DSBA), significantly reduced spore germination and appressorium formation in B. bassiana (p ≤ 0.05), with stronger inhibition becoming more pronounced at higher concentrations. In contrast, amino acids and their metabolites (e.g., glycylmethionine) and glycerophospholipid compounds like 1,2-Dioleoyl-sn-Glycero-3-Phosphocholine (DOPC) had no significant effects on spore germination or appressorium formation at any tested concentration (p > 0.05). LC-MS analysis revealed that the insect cuticular fatty acyl compound carnitine C3:0 broadly modulated the secondary metabolism of B. bassiana. Following appressorium formation, 146 metabolites with significant changes in abundance were identified. Before appressorium formation, carnitine C3:0 promoted the activation of B. bassiana signaling pathways, such as Rap1, and stimulated antibiotic biosynthesis (penicillin and cephalosporin), thereby suppressing competing microorganisms and facilitating initial attachment. After appressorium formation, carnitine C3:0 activated pathways related to metabolite synthesis (e.g., arginine and nucleotides biosynthesis) and population regulation (ferroptosis), thereby enhancing appressorium function and structural stability. Thus, carnitine C3:0 enhances B. bassiana’s ability to establish infection sites before appressorium formation through antibiotic clearance and signal activation, and maintain infection structures after formation via metabolic reinforcement and population regulation. This study lays a theoretical foundation for further investigations into B. bassiana infection mechanisms, pathogenicity, and the role of its conidiophores. Full article

Colletotrichum gloeosporioides causes rubber tree anthracnose and leads to serious loss in natural rubber production. Autophagy is a highly conserved process to maintain nutrient recycling and plays important roles in growth, development and pathogenicity in plant pathogenic fungi. The process of autophagy is modulated by a series of autophagy-related (ATG) genes. ATG16 is a subunit of the ATG12-ATG5-ATG16 complex which functions in a manner analogous to an E3-like enzyme which is essential for autophagosome formation. However, the function of the ATG16 homolog in C. gloeosporioides remains unknown. In this study, the ATG16 homolog of C. gloeosporioides was identified and named as CgATG16. The expression level of CgATG16 was particularly higher in conidium, germination, appressorium, and the early stage of infection, and significantly induced by nutritional deficiency. Absence of CgATG16 led to slower colony growth, decreased conidia production and germination rate, longer germ tube cells, lower appressorium formation rate and impaired pathogenicity to rubber tree leaves. Absence of CgATG16 resulted in lower melanin content with decreased expression of polyketide synthase gene CgPKS1 and scytalone dehydratase gene CgSCD1. Moreover, absence of CgATG16 also led to the universal autophagy marker ATG8-GFP failing to enter into the vacuoles in mycelium and during appressorium development with a significantly reduced autophagosome number. Both rapamycin and cyclic adenosine monophosphate (cAMP) partially restored the appressorium formation ability in CgATG16 knockout mutant. Absence of CgATG16 increased the activity of target of rapamycin (TOR) kinase and decreased the content of cAMP. These data suggest that CgATG16 contributes to the pathogenicity of C. gloeosporioides to the rubber tree by regulating the mycelium growth, melanin synthesis and the formation of invasion structure, and this process is related to autophagy mediated by TOR and cAMP signaling. Full article

Sclerotinia sclerotiorum, a soil-borne phytopathogenic fungus with a broad host range, often leads to severe disease and significant economic losses in agricultural production. The guanine exchange factor EFA6 of ADP-ribosylation factor 6 (ARF6) has been extensively studied in animals, but its function in fungi is seldom reported. Here, reverse genetics methods were employed to explore the effects of SsEFA6 in the process of pathogenicity of S. sclerotiorum. Knockout of SsEFA6 hindered appressoria formation and sclerotia production. However, it did not affect the secretion of oxalic acid, the sensitivity to cell wall inhibitors, or hyperosmotic stress. Nevertheless, SsEFA6 deletion did result in a significant decrease in mutant virulence, indicative of its indispensability in virulence. Therefore, SsEFA6 plays an essential role in appressoria formation, sclerotia production, and fungal virulence in S. sclerotiorum. Full article

Sclerotinia sclerotiorum is a devastating soilborne fungal pathogen that causes Sclerotinia stem rot in many economically important crops. It forms sclerotia, resilient dormant structures that can persist in soil for years. Understanding the molecular mechanism of sclerotia formation is crucial for developing effective control strategies, but only a limited number of signaling components have been uncovered in this process. Through independent forward genetic screens, we identified SsLae1 and SsVel1, two core components of the conserved fungal velvet complex, as essential regulators of sclerotia formation and virulence in S. sclerotiorum. Disruption of either gene abolished sclerotia formation, impaired compound appressorium development, and significantly reduced virulence. Further RNA-seq analysis using the Ssvel1 mutant revealed widespread downregulation of known developmental and virulence regulators. Collectively, these findings establish the velvet complex as a master regulator for both sclerotia development and virulence in S. sclerotiorum. Full article

The appressorium is a specialised infection structure formed by Beauveria bassiana during host invasion. This study used sulforaphane to regulate the formation rate of B. bassiana appressoria, evaluated the correlation between appressorium formation and fungal pathogenicity, and explored its impact on insect cuticular metabolism. The results showed that sulforaphane significantly modulated appressorium formation. Spore suspensions with varying appressorium formation rates were injected into Opisina arenosella and Bombyx mori larvae. As the appressorium formation rate increased, B. bassiana exhibited enhanced pathogenicity, leading to accelerated larval mortality. A significant positive correlation (p ≤ 0.05) was observed between appressorium formation and pathogenicity. LC-MS analysis revealed that, prior to appressorium development, larvae activated defence mechanisms involving secondary metabolites, hormone signalling, and toxin metabolism pathways. Following appressorium formation, 61 unique cuticular compounds were identified, along with activation of host lipid metabolism (notably glycerophospholipid degradation), programmed cell death pathways (ferroptosis, necroptosis), and enhanced energy metabolism via the citric acid cycle—collectively indicating disruption of the epidermal defence barrier. Overall, appressorium development by B. bassiana significantly reshapes the metabolic landscape of the larval cuticle, thereby enhancing fungal virulence. This study provides a theoretical foundation for understanding the pathogenic mechanisms of B. bassiana. Full article

Protein phosphatases are crucial enzymes that regulate key cellular processes such as the cell cycle, gene transcription, and translation in eukaryotes. Seven PP2C protein phosphatases have been identified in Magnaporthe oryzae. However, their synergistic roles in the pathology and physiology of M. oryzae remain poorly investigated. By qRT-PCR analysis, we found that PTC1 and PTC2 are significantly upregulated in the PTC5 deletion mutant. The double deletion of the MoPTC5/MoPTC1 and MoPTC5/MoPTC2 genes significantly reduced hyphal growth, conidiophore formation, sporulation, and virulence in M. oryzae. In addition, the double-knockout mutants were increasingly sensitive to different osmotic, oxidative, and cell wall stresses. Western blot analysis revealed that MoPtc5 plays a synergistic function with MoPtc1 and MoPtc2 in the regulation of MoMps1 and MoOsm1 phosphorylation levels. Lastly, appressorium formation and turgor generation were remarkably affected in the ΔMoptc5ΔMoptc1 and ΔMoptc5ΔMoptc2 double-deletion mutants. These findings demonstrate the overlapping roles of PP2c protein phosphatase in the fungal development and pathogenesis of M. oryzae. Full article

Nitrogen is an essential nutrient that frequently determines the growth rate of fungi. Nitrate transporter proteins (Nrts) play a crucial role in the cellular absorption of nitrate from the environment. Entomopathogenic fungi (EPF) have shown their potential in the biological control of pests. Thus, comprehending the mechanisms that govern the pathogenicity and stress tolerance of EPF is helpful in improving the effectiveness and practical application of these fungal biocontrol agents. In this study, we utilized homologous recombination to create MaNrtB deletion mutants and complementation strains. We systematically investigated the biological functions of the nitrate transporter protein gene MaNrtB in M. acridum. Our findings revealed that the disruption of MaNrtB resulted in delayed conidial germination without affecting conidial production. Stress tolerance assays demonstrated that the MaNrtB disruption strain was more vulnerable to UV-B irradiation, hyperosmotic stress, and cell wall disturbing agents, yet it exhibited increased heat resistance compared to the wild-type strain. Bioassays on the locust Locusta migratoria manilensis showed that the disruption of MaNrtB impaired the fungal virulence owing to the reduced appressorium formation on the insect cuticle and the attenuated growth in the locust hemolymph. These findings provide new perspectives for understanding the pathogenesis of EPF. Full article

The Gti1/Pac2 protein family, which is highly conserved across fungi, is pivotal in processes such as fungal development, spore formation, protein export, toxin production, and virulence. Despite its importance, the precise functions of Pac2 within entomopathogenic fungi have yet to be fully understood. In our study, the MaPac2 gene from M. acridum was identified, and its functions were explored. Studying the domain of the protein showed that MaPac2 comprises 422 amino acids with a characteristic Gti1/Pac2 family domain (Pfam09729). Additionally, MaPac2 is predicted to have an N-terminal protein kinase A phosphorylation site and a potential cyclin-dependent kinase phosphorylation site, highlighting its potential regulatory roles in the fungus. Our findings indicate that the inactivation of MaPac2 resulted in faster germination of conidia and a marked reduction in conidial production. Furthermore, stress tolerance tests revealed that the absence of MaPac2 significantly bolstered the fungal resilience to UV-B radiation, heat shock, SDS exposure, and stresses induced by hyperosmotic conditions and oxidative challenges. Virulence assessments through bioassays indicated no substantial differences among the WT, MaPac2-disrupted strain, and CP strains in the topical inoculation trials. Interestingly, deletion of MaPac2 increased the fungal virulence by intrahemocoel injection. Furthermore, we found that disruption of MaPac2 impaired fungal cuticle penetration due to the diminished appressorium formation but increased the fungal growth in locust hemolymph. These findings provide further insights into the roles played by Gti1/Pac2 in insect pathogenic fungi. Full article

Unique genes refer to genes specific to a particular organism and play crucial roles in the biological functions, evolutionary processes, and adaptations to external environments. However, the roles of unique genes in plant pathogenic fungi remain largely unexplored. In this study, we identified a novel unique gene in the rice blast fungus Magnaporthe oryzae, named MoUNG (M. oryzae unique gene), through T-DNA insertion mutagenesis. The disruption of the MoUNG promoter region in the T-DNA insertion mutant (T30-104) led to an almost loss of MoUNG expression. MoUNG has no functional domains and lacks homologues in other organism. It is highly expressed during the early-infection stage between 16 and 32 h post-inoculation (HPI), in contrast to its expression in mycelia and at the later infection stage of 48 HPI. Notably, attempts to knock out MoUNG were unsuccessful, so we examined the T30-104 mutant and found it showed significantly reduced growth, conidiation, and pathogenicity. Introducing the full-length MoUNG with its promoter into T30-104 restored these phenotypic defects. Additionally, subcellular localization assays revealed that MoUNG exhibits a dot-like distribution within the cytoplasm of mycelium, conidium, appressorium, and invasive hypha. Furthermore, knock-down of MoUNG produced results similar to those observed with the insertion mutation. In conclusion, we identified a novel unique gene MoUNG in M. oryzae and demonstrated its involvement in growth, conidiation, and pathogenicity. Full article

The present study aimed to evaluate the effect of silver nanoparticles (AgNPs) on the development of Colletotrichum lindemuthianum, the progression of anthracnose symptoms, and the growth of common bean plants. For this purpose, the fungal mycelial growth and conidial germination were assessed at AgNP concentrations of 0, 10, 30, and 50 mg·L⁻¹ after seven days of incubation, as well as at 0, 0.1, 0.5, 1, 10, 30, and 50 mg·L⁻¹ after 72 h, respectively. Bean plants of the IPR Uirapuru cultivar were sprayed at the V3 growth stage with AgNPs at 0, 10, 30, or 50 mg·L⁻¹, either two days before, on the day of, or two days after inoculation. Conidial germination and appressoria melanization were measured on the leaf discs collected 24, 48, and 72 h after inoculation, and disease severity was assessed at 7 and 12 days post-inoculation. Another set of bean plants grown under the same conditions was used to evaluate growth promotion by AgNPs. For this, the plants were sprayed twice (with a seven-day interval), starting at the V3 growth stage, with AgNPs at 0, 10, 30, or 50 mg·L⁻¹. Seven days after the second treatment, plant length and the fresh and dry weights of shoots and roots were measured, and the foliar pigments were quantified. The AgNPs did not reduce mycelial growth but completely inhibited the germination of C. lindemuthianum conidia. The severity of anthracnose decreased with the AgNPs in a dose- and application time-dependent manner, with the highest reduction (90%) observed when applied on the same day as an inoculation at 50 mg·L⁻¹. This was strongly linked to a 70% decline in conidia germination and appressorium melanization on bean leaves. AgNPs at 50 mg·L⁻¹ promoted plant growth by increasing the total length by 3%, as well as the fresh weights of bean shoots and roots by 17% and 90%, respectively, but did not affect the content of leaf pigments. Full article

Slt2 is an important component of the Slt2-MAPK pathway and plays critical regulatory roles in growth, cell wall integrity, melanin biosynthesis, and pathogenicity of plant fungi. AaSlt2, an ortholog of the Saccharomyces cerevisiae Slt2 gene, was identified from A. alternata in this study, and its function was clarified by knockout of the gene. The ΔAaSlt2 strain of A. alternata was found to be defective in spore morphology, vegetative growth, and sporulation. Analysis of gene expression showed that expression of the AaSlt2 gene was significantly up-regulated during infection structure formation of A. alternata on hydrophobic and pear wax extract-coated surfaces. Further tests on onion epidermis confirmed that spore germination was reduced in the ΔAaSlt2 strain, together with decreased formation of appressorium and infection hyphae. Moreover, the ΔAaSlt2 strain was sensitive to cell wall inhibitors, and showed significantly reduced virulence on pear fruit. Furthermore, cell wall degradation enzyme (CWDE) activities, melanin accumulation, and toxin biosynthesis were significantly lower in the ΔAaSlt2 strain. Overall, the findings demonstrate the critical involvement of AaSlt2 in growth regulation, stress adaptation, infection structure formation, and virulence in A. alternata. Full article

Alternaria alternata, a causal agent of pear black spot, can recognize and respond to physicochemical signals from fruit surfaces through an intricate signaling network to initiate infection. Crz1 is an important transcription factor downstream of the calcium signaling pathway. In this study, we first investigated the infection structure formation process of the wild type (WT) and ΔAaCrz1 strains induced by the cuticular wax of the “Zaosu” pear by microscopic observation. We found that the infection process was delayed and the rate of appressorium formation and infection hyphae formation was significantly decreased in the ΔAaCrz1 strain. RNA-seq of WT and ΔAaCrz1 strains was analyzed after 6 h of induction with pear wax. A total of 893 up-regulated and 534 down-regulated genes were identified. Among them, genes related to cell wall degrading enzymes, ABC transporters, and ion homeostasis were down-regulated, and the autophagy pathway was induced and activated. In addition, disruption to the intracellular antioxidant system was also found after AaCrz1 knockdown. In summary, this study provides new information on the mechanism of the transcription factor AaCrz1 in the regulation of infection structure formation of A. alternata induced by pear peel wax signal, which can be used to develop new strategies for controlling fungal diseases in the future. Full article

Poplar is an economically and ecologically valuable tree species. Anthracnose, which severely affects poplar tree growth, is mainly caused by Colletotrichum gloeosporioides. In the infestation cycle of poplar anthracnose, the entry of C. gloeosporioides into the host tissue depends on the formation of an appressorium. The subsequent development of the appressorium determines the pathogenesis of poplar anthracnose and the degree of damage. Previous studies have found that the transcription factor CgSte12 affects appressorium formation and development by regulating the expression of a series of genes, including the sterol-synthesis-related gene CgCYP51, which influences appressorium formation and development. In this study, knockout and functional analyses of CgCYP51 revealed decreases in differentiation, darkening rate, and turgor pressure of appressoria in mutants. Additionally, compared with the wild-type appressorium, mutant appressoria secreted less mucus and exhibited abnormal penetration pore formation, ultimately leading to decreased pathogenicity. Moreover, CgCyp51 affected the sensitivity of C. gloeosporioides to sterol biosynthesis inhibitors. Considered together, the study findings indicate CgCYP51 is a key CgSte12-regulated gene that affects C. gloeosporioides appressorium formation and development. Furthermore, the study data provide new insights into the molecular basis of C. gloeosporioides appressorium formation and development. Full article