Hyphal Development Research Articles

Anticipatory Stress Responses and Immune Evasion in Fungal Pathogens.

In certain niches, microbes encounter environmental challenges that are temporally linked. In such cases, microbial fitness is enhanced by the evolution of anticipatory responses where the initial challenge simultaneously activates pre-emptive protection against the second impending challenge. The accumulation of anticipatory responses in domesticated yeasts, which have been termed 'adaptive prediction', has led to the emergence of 'core stress responses' that provide stress cross-protection. Protective anticipatory responses also seem to be common in fungal pathogens of humans. These responses reflect the selective pressures that these fungi have faced relatively recently in their evolutionary history. Consequently, some pathogens have evolved 'core environmental responses' which exploit host signals to trigger immune evasion strategies that protect them against imminent immune attack.

A Cu-only superoxide dismutase from stripe rust fungi functions as a virulence factor deployed for counter defense against host-derived oxidative stress.

Plants quickly accumulate reactive oxygen species (ROS) to resist against pathogen invasion, while pathogens strive to escape host immune surveillance by degrading ROS. However, the nature of the strategies that fungal pathogens adopt to counteract host-derived oxidative stress is manifold and requires deep investigation. In this study, a superoxide dismutase (SOD) from Puccinia striiformis f. sp. tritici (Pst) PsSOD2 with a signal peptide (SP) and the glycophosphatidyl inositol (GPI) anchor, strongly induced during infection, was analysed for its biological characteristics and potential role in wheat-Pst interactions. The results showed that PsSOD2 encodes a Cu-only SOD and responded to ROS treatment. Heterologous complementation assays in Saccharomyces cerevisiae suggest that the SP of PsSOD2 is functional for its secretion. Transient expression in Nicotiana benthamiana leaves revealed that PsSOD2 is localized to the plasma membrane. In addition, knockdown of PsSOD2 by host-induced gene silencing reduced Pst virulence and resulted in restricted hyphal development and increased ROS accumulation. In contrast, heterologous transient assays of PsSOD2 suppressed flg22-elicited ROS production. Taken together, our data indicate that PsSOD2, as a virulence factor, was induced and localized to the plasma membrane where it may function to scavenge host-derived ROS for promoting fungal infection.

In vitro study of volatile organic compounds produced by the mutualistic fungus of leaf-cutter ants and the antagonist Escovopsis

Occasionally, the symbiosis of the fungus-growing ants is threatened by ascomycetous fungi mainly by species in the genus Escovopsis. GC-MS analyses were conducted to investigate the volatile organic compounds (VOCs) produced by Escovopsis and the mutualistic Leucocoprinus gongylophorus during dual-culture. In dual-culture Escovopsis' development was faster. Hexadecanoic acid (palmitic acid) and VOCs such as the sesquiterpenes α-cedrene, β-cedrene, thujopsene, α-santalene, α-selinene, cedrene, (+)-cuparene and δ-cadinene secreted by L. gongylophorus may be responsible for speeding the germination of Escovopsis’ conidia as well as for the faster hyphal development. Based on this study and available literature we infer that the relationship between these two species is mediated by hyphal interference (HI). Additional studies should unveil if the biological activity of some compounds produced by these fungi is attractive, repellent or toxic to the ants and inhibitory to other fungi.

Role of the Inositol Polyphosphate Kinase Vip1 in Autophagy and Pathogenesis in Candida Albicans

Aim: Inositol polyphosphate kinases are involved in regulation of many cellular processes in eukaryotic cells. In this study, we investigated the functions of the inositol polyphosphate kinase Vip1 in autophagy and pathogenicity of Candida albicans. Results: Loss of Vip1 caused significantly increased sensitivity to nitrogen source starvation, abnormal localization and degradation of autophagy protein, higher vacuolar pH and higher (rather than lower) intracellular ATP levels compared with control strains. Besides, the mutant showed attenuated hyphal development and virulence during systemic infection to mice. Conclusion: The results reveal that Vip1 is important to autophagy of C. albicans. The maintenance of vacuolar acidic pH contributed to the role of Vip1 in autophagy. Vip1 is also required for pathogenicity of C. albicans.

Phagolysosomal Survival Enables Non-lytic Hyphal Escape and Ramification Through Lung Epithelium During Aspergillus fumigatus Infection.

Aspergillus fumigatus is the most important mould pathogen in immunosuppressed patients. Suboptimal clearance of inhaled spores results in the colonisation of the lung airways by invasive hyphae. The first point of contact between A. fumigatus and the host is the lung epithelium. In vitro and ex vivo studies have characterised critical aspects of the interaction of invasive hyphae on the surface of epithelial cells. However, the cellular interplay between internalised A. fumigatus and the lung epithelium remains largely unexplored. Here, we use high-resolution live-cell confocal microscopy, 3D rendered imaging and transmission electron microscopy to define the development of A. fumigatus after lung epithelium internalisation in vitro. Germination, morphology and growth of A. fumigatus were significantly impaired upon internalisation by alveolar (A549) and bronchial (16HBE) lung epithelial cells compared to those growing on the host surface. Internalised spores and germlings were surrounded by the host phagolysosome membrane. Sixty per cent of the phagosomes containing germlings were not acidified at 24 h post infection allowing hyphal development. During escape, the phagolysosomal membrane was not ruptured but likely fused to host plasma membrane allowing hyphal exit from the intact host cell in an non-lytic Manner. Subsequently, escaping hyphae elongated between or through adjacent epithelial lung cells without penetration of the host cytoplasm. Hyphal tips penetrating new epithelial cells were surrounded by the recipient cell plasma membrane. Altogether, our results suggest cells of lung epithelium survive fungal penetration because the phagolysosomal and plasma membranes are never breached and that conversely, fungal spores survive due to phagosome maturation failure. Consequently, fungal hyphae can grow through the epithelial cell layer without directly damaging the host. These processes likely prevent the activation of downstream immune responses alongside limiting the access of professional phagocytes to the invading fungal hypha. Further research is needed to investigate if these events also occur during penetration of fungi in endothelial cells, fibroblasts and other cell types.

Candida tropicalis RON1 is required for hyphal formation, biofilm development, and virulence but is dispensable for N-acetylglucosamine catabolism.

In this study, we identified the role of RON1, an NDT80-like gene, in Candida tropicalis. Unlike the gene in Candida albicans, our studies showed that RON1 is a key regulator of hyphal formation, biofilm development and virulence but is dispensable for N-acetylglucosamine catabolism in C. tropicalis.

Chytrid rhizoid morphogenesis resembles hyphal development in multicellular fungi and is adaptive to resource availability.

Key to the ecological prominence of fungi is their distinctive cell biology, our understanding of which has been principally based on dikaryan hyphal and yeast forms. The early-diverging Chytridiomycota (chytrids) are ecologically important and a significant component of fungal diversity, yet their cell biology remains poorly understood. Unlike dikaryan hyphae, chytrids typically attach to substrates and feed osmotrophically via anucleate rhizoids. The evolution of fungal hyphae appears to have occurred from rhizoid-bearing lineages and it has been hypothesized that a rhizoid-like structure was the precursor to multicellular hyphae. Here, we show in a unicellular chytrid, Rhizoclosmatium globosum, that rhizoid development exhibits striking similarities with dikaryan hyphae and is adaptive to resource availability. Rhizoid morphogenesis exhibits analogous patterns to hyphal growth and is controlled by β-glucan-dependent cell wall synthesis and actin polymerization. Chytrid rhizoids growing from individual cells also demonstrate adaptive morphological plasticity in response to resource availability, developing a searching phenotype when carbon starved and spatial differentiation when interacting with particulate organic matter. We demonstrate that the adaptive cell biology and associated developmental plasticity considered characteristic of hyphal fungi are shared more widely across the Kingdom Fungi and therefore could be conserved from their most recent common ancestor.

FgHAT2 is involved in regulating vegetative growth, conidiation, DNA damage repair, DON production and virulence in Fusarium graminearum

Histone lysine acetylation is catalyzed by acetyltransferases (HATs), which is important in regulating gene expression and physiological function in eukaryotic cells. HATs can be classified into two main types: A- and B-type HATs. Recently, in Fusarium graminearum, it has been reported that A-type HATs are involved in hyphal development, conidiation, sexual reproduction and virulence. However, the biological roles of B-type HATs are unknown. Here we report the identification and characterization of two B-type HATs (FgHat1 and FgHat2) in F. graminearum. Targeted deletion of FgHAT1 did not result in any detectable phenotypes. However, ΔFghat2 mutants were severely defective in vegetative growth, conidia production and morphogenesis, deoxynivalenol (DON) biosynthesis and virulence. Interestingly, deletion of FgHAT2 resulted in significantly increased sensitivity to the DNA-damaging agent methyl methanesulfonate (MMS). Furthermore, double deletion mutants (ΔFghat1ΔFghat2) displayed similar phenotypes to the ΔFghat2 mutants. Taken together, we conclude that FgHat2 but not FgHat1 plays essential roles in regulating morphogenesis, DNA damage repair, DON production and virulence in F. graminearum.

Piperine Impedes Biofilm Formation and Hyphal Morphogenesis of Candida albicans.

Candida albicans is the primary etiological agent associated with the pathogenesis of candidiasis. Unrestricted growth of C. albicans in the oral cavity may lead to oral candidiasis, which can progress to systemic infections in worst scenarios. Biofilm of C. albicans encompasses yeast and hyphal forms, where hyphal formation and yeast to hyphal morphological transitions are contemplated as the key virulence elements. Current clinical repercussions necessitate the identification of therapeutic agent that can limit the biofilm formation and escalating the susceptibility of C. albicans to immune system and conventional antifungals. In the present study, a plant-derived alkaloid molecule, piperine, was investigated for the antibiofilm and antihyphal activities against C. albicans. Piperine demonstrated a concentration-dependent antibiofilm activity without exerting negative impact on growth and metabolic activity. Inhibition in the hyphal development was witnessed through confocal laser-scanning microscopy and scanning electron microscopy. Interestingly, piperine displayed a tremendous potential to inhibit the virulence-associated colony morphologies, such as filamentation and wrinkling. Furthermore, piperine regulated morphological transitions between yeast and hyphal forms by inhibiting hyphal extension and swapping hyphal phase to yeast forms yet under filamentation-inducing circumstances. Remarkably, piperine-challenged C. albicans exhibited low potential for spontaneous antibiofilm resistance development. In addition, piperine effectively reduced in vivo colonization and prolonged survival of C. albicans-infected Caenorhabditis elegans, thereby expounding the distinct antivirulent potential. Transcriptomic analysis revealed piperine significantly downregulating the expression of several biofilm related and hyphal-specific genes (ALS3, HWP1, EFG1, CPH1, etc.). Furthermore, no acute toxicity was observed in the HBECs and nematodes exposed to piperine. Altogether, results from this study reveals the potential of piperine to inhibit biofilm and hyphal morphogenesis, and its in vivo efficacy and innocuous nature to HBECs suggests that piperine may be considered as a potential candidate for the treatment of biofilm-associated C. albicans infection, especially for oral candidiasis.

Fatty Acids as Antibiofilm and Antivirulence Agents.

Fatty acids (FAs) have emerged as a potential alternative to conventional antibiotics. Since many microbes respond differently to a variety of natural and synthetic FAs, substantial efforts have been made to understand the unique features of FAs that function as antimicrobials at high doses and biofilm inhibitors at low doses. Here, we provide an overview of the emerging antibiofilm properties of FAs and their relations to virulence and quorum sensing, such as diffusible signal factors (DSFs), acyl-homoserine lactones, and autoinducer-2 systems. Several FAs mimic DSFs and control motility, fimbriae, hyphae, and biofilm development as well as virulence characteristics of diverse microbes. The findings of FAs as antibiofilm and antivirulence agents will provide a new paradigm to cope with recalcitrant pathogens.

Identifying Candida albicans Gene Networks Involved in Pathogenicity.

Candida albicans is a normal member of the human microbiome. It is also an opportunistic pathogen, which can cause life-threatening systemic infections in severely immunocompromized individuals. Despite the availability of antifungal drugs, mortality rates of systemic infections are high and new drugs are needed to overcome therapeutic challenges including the emergence of drug resistance. Targeting known disease pathways has been suggested as a promising avenue for the development of new antifungals. However, <30% of C. albicans genes are verified with experimental evidence of a gene product, and the full complement of genes involved in important disease processes is currently unknown. Tools to predict the function of partially or uncharacterized genes and generate testable hypotheses will, therefore, help to identify potential targets for new antifungal development. Here, we employ a network-extracted ontology to leverage publicly available transcriptomics data and identify potential candidate genes involved in disease processes. A subset of these genes has been phenotypically screened using available deletion strains and we present preliminary data that one candidate, PEP8, is involved in hyphal development and immune evasion. This work demonstrates the utility of network-extracted ontologies in predicting gene function to generate testable hypotheses that can be applied to pathogenic systems. This could represent a novel first step to identifying targets for new antifungal therapies.

Nuclear Ssr4 Is Required for the In Vitro and In Vivo Asexual Cycles and Global Gene Activity of Beauveria bassiana.

Ssr4 serves as a cosubunit of chromatin-remodeling SWI/SNF and RSC complexes in yeasts but remains functionally uncharacterized due to its essentiality for yeast viability. Here, we report pleiotropic effects of the deletion of the ssr4 ortholog nonessential for cell viability in Beauveria bassiana, an asexual insect mycopathogen. The deletion of ssr4 resulted in severe growth defects on different carbon/nitrogen sources, increased hyphal hydrophilicity, blocked hyphal differentiation, and 98% reduced conidiation capacity compared to a wild-type standard. The limited Δssr4 conidia featured an impaired coat with disordered or obscure hydrophobin rodlet bundles, decreased hydrophobicity, increased size, and lost insect pathogenicity via normal cuticle infection and 90% of virulence via intrahemocoel injection. The expression of genes required for hydrophobin biosynthesis and assembly of the rodlet layer was drastically repressed in more hydrophilic Δssr4 cells. Transcriptomic analysis revealed 2,517 genes differentially expressed in the Δssr4 mutant, including 1,505 downregulated genes and 1,012 upregulated genes. The proteins encoded by hundreds of repressed genes were involved in metabolism and/or transport of carbohydrates, amino acids, and lipids, inorganic ion transport and energy production or conversion, including dozens involved in DNA replication, transcription, translation, and posttranslational modifications. However, purified Ssr4 samples showed no DNA-binding activity, implying that the role of Ssr4 in genome-wide gene regulation could rely upon its acting as a cosubunit of the two complexes. These findings provide the first insight into an essential role of Ssr4 in the asexual cycle in vitro and in vivo of B. bassiana and highlights its importance for the filamentous fungal lifestyle.IMPORTANCE Ssr4 is known to serve as a cosubunit of chromatin-remodeling SWI/SNF and RSC complexes in yeasts but has not been functionally characterized in fungi. This study unveils for the first time the pleiotropic effects caused by deletion of ssr4 and its role in mediating global gene expression in a fungal insect pathogen. Our findings confirm an essential role of Ssr4 in hydrophobin biosynthesis and assembly required for growth, differentiation, and development of aerial hyphae for conidiation and conidial adhesion to insect surface and its essentiality for insect pathogenicity and virulence-related cellular events. Importantly, Ssr4 can regulate nearly one-fourth of all genes in the fungal genome in direct and indirect manners, including dozens involved in gene activity and hundreds involved in metabolism and/or transport of carbohydrates, amino acids, lipids, and/or inorganic ions. These findings highlight a significance of Ssr4 for filamentous fungal lifestyle.

Lactoferrin Is Broadly Active against Yeasts and Highly Synergistic with Amphotericin B.

Lactoferrin (LF) is a multifunctional milk protein with antimicrobial activity against a range of pathogens. While numerous studies report that LF is active against fungi, there are considerable differences in the level of antifungal activity and the capacity of LF to interact with other drugs. Here we undertook a comprehensive evaluation of the antifungal spectrum of activity of three defined sources of LF across 22 yeast and 24 mold species and assessed its interactions with six widely used antifungal drugs. LF was broadly and consistently active against all yeast species tested (MICs, 8 to 64 μg/ml), with the extent of activity being strongly affected by iron saturation. LF was synergistic with amphotericin B (AMB) against 19 out of 22 yeast species tested, and synergy was unaffected by iron saturation but was affected by the extent of LF digestion. LF-AMB combination therapy significantly prolonged the survival of Galleria mellonella wax moth larvae infected with Candida albicans or Cryptococcus neoformans and decreased the fungal burden 12- to 25-fold. Evidence that LF directly interacts with the fungal cell surface was seen via scanning electron microscopy, which showed pore formation, hyphal thinning, and major cell collapse in response to LF-AMB synergy. Important virulence mechanisms were disrupted by LF-AMB treatment, which significantly prevented biofilms in C. albicans and C. glabrata, inhibited hyphal development in C. albicans, and reduced cell and capsule size and phenotypic diversity in Cryptococcus Our results demonstrate the potential of LF-AMB as an antifungal treatment that is broadly synergistic against important yeast pathogens, with the synergy being attributed to the presence of one or more LF peptides.

A Cryptic Alternative for the Evolution of Hyphae.

A growing awareness of a subsurface fossil record of mostly hyphal fungi organisms stretching back through the Phanerozoic to ≈400 megaannum (Ma) and possibly earlier, provides an alternative view on hyphal development. Parallel with the emergence of hyphal fungi during Ordovician-Devonian times when plants colonized the land, which is the traditional notion of hyphal evolution, hyphae-based fungi existed in the deep biosphere. New insights suggest that the fundamental functions of hyphae may have evolved in response to an ancient subsurface endolithic life style and might have been in place before the colonization of land. To address the gaps in the current understanding of hyphal evolution a strategy based on research prospects involving investigations of uncharted geological material, new diagnostics, and comparisons to live species is proposed.

Lack of the Histone Methyltransferase Gene Ash2 Results in the Loss of Citrinin Production in Monascus purpureus

Absent, small, or homeotic discs 2 (Ash2), a histone H3K4 methyltransferase complex, has been implicated in the control of hyphal development and secondary metabolism in many kinds of filamentous fungi. We constructed an Ash2 deletion mutant (ΔAsh2) by using an Agrobacterium-mediated gene knockout method to investigate the function of the Ash2 gene in the mold Monascus purpureus. Lack of the Ash2 gene resulted in the formation of a lower colony phenotype with fluffy aerial hyphae that autolyzed as the colony grew on potato dextrose agar at 30°C. The production of pigments and the number of conidia were significantly lower in the ΔAsh2 than in the wild type. Citrinin production by the ΔAsh2 was not detected during 15 days of fermentation. Relative expression levels of secondary metabolite regulatory genes PigR and CTNR, secondary metabolite synthesizing genes PKSPT and CTN, key genes of mitogen-activated protein kinase pathway Spk1 and its downstream gene mam2, the conidium development control gene BrlA, and global regulatory genes LaeA and VeA were detected by the quantitative real-time PCR. These results indicate that the Ash2 gene is involved in conidial germination, pigment production, and citrinin production and plays a key role in development and secondary metabolism in M. purpureus.

Magnaporthe oryzae fimbrin organizes actin networks in the hyphal tip during polar growth and pathogenesis.

Magnaporthe oryzae causes rice blast disease, but little is known about the dynamic restructuring of the actin cytoskeleton during its polarized tip growth and pathogenesis. Here, we used super-resolution live-cell imaging to investigate the dynamic organization of the actin cytoskeleton in M. oryzae during hyphal tip growth and pathogenesis. We observed a dense actin network at the apical region of the hyphae and actin filaments originating from the Spitzenkörper (Spk, the organizing center for hyphal growth and development) that formed branched actin bundles radiating to the cell membrane. The actin cross-linking protein Fimbrin (MoFim1) helps organize this actin distribution. MoFim1 localizes to the actin at the subapical collar, the actin bundles, and actin at the Spk. Knockout of MoFim1 resulted in impaired Spk maintenance and reduced actin bundle formation, preventing polar growth, vesicle transport, and the expansion of hyphae in plant cells. Finally, transgenic rice (Oryza sativa) expressing RNA hairpins targeting MoFim1 exhibited improved resistance to M. oryzae infection, indicating that MoFim1 represents an excellent candidate for M. oryzae control. These results reveal the dynamics of actin assembly in M. oryzae during hyphal tip development and pathogenesis, and they suggest a mechanism in which MoFim1 organizes such actin networks.

A new epiphyllous fly-speck fungus from the Early Cretaceous Potomac Group of Virginia (125–112 Ma): Protographum luttrellii, gen. et sp. nov.

ABSTRACT Fly-speck fungi reproduce via thyriothecia that consist of sporogenous tissue appressed to cuticle surfaces of plant leaves and covered by a shield-like scutellum. Thyriothecial scutella likely evolved repeatedly in Dothideomycetes (Ascomycota), and their morphology varies by lineage. Fly-speck fungi have an exceptionally good fossil record that begins in the Mesozoic. The interpretation of scutellum characters in fossils may provide insights into origins of Dothideomycetes and help calibrate the timing of ascomycete evolution. From sediments of the Lower Cretaceous (125–112 Ma) Potomac Group of Virginia, from Dutch Gap Canal, lower Zone 1, we found scutella similar to those of extant Aulographaceae (Dothideomycetes), attached to a single piece of dispersed coniferous cuticle. We analyze hyphae and scutellum development among four extant Aulographaceae species for comparison with the fossil. The excellent preservation of fungi on the leaf cuticle surface allows us to infer a developmental sequence for the fossil. Scutellum development begins with coordinated growth of multiple neighboring generator hyphae and continues with hyphae producing two-dimensional pseudomonopodial, dichotomous, radial growth. Asci and ascospores were not found. We coded states for seven morphological characters using direct observations of the fossil and eight extant taxa, and using the literature for 28 others. We inferred a phylogeny using nuclear 18S and 28S rDNA of 36 extant taxa, 34 Dothideomycetes and two Arthoniomycetes. The phylogeny includes newly determined sequences from five species, two from Aulographaceae. With a branch-and-bound search, we inferred the most parsimonious placements of the fossil given the molecular tree topology. The parsimony analysis constrained by the rDNA phylogeny places the fossil taxon among stem lineages near Aulographaceae or among the known living members of Aulographaceae. We describe the fossil morphotype as Protographum luttrellii, gen. et sp. nov. The fossil provides the oldest evidence of morphological characters restricted among extant fungi to Aulographaceae.

Dimethylaminododecyl methacrylate inhibits Candida albicans and oropharyngeal candidiasis in a pH-dependent manner

The prevalence of stomatitis, especially that caused by Candida albicans, has highlighted the need for new antifungal agents. We previously found that a type of quaternary ammonium salts, dimethylaminododecyl methacrylate (DMADDM), incorporated in dental materials inhibited the growth and hyphal development of C. albicans. However, how the quaternary ammonium salts inhibited the fungal pathogens and whether the oral condition, such as salivary pH variation under different diseases, can affect the antimicrobial capacity of quaternary ammonium salts is unknown. This study evaluated the antifungal effects of DMADDM at different pH in vitro and in vivo. A pH-dependent antifungal effect of DMADDM was observed in planktonic and biofilm growth. DMADDM enhanced antifungal activity at alkaline pH. Two pH-regulated genes (PHR1/PHR2) of C. albicans were correlated with the pH-dependent antifungal effects of DMADDM. The PHR1/PHR2 genes and pH values regulated the zeta potential of C. albicans, which then influenced the binding between C. albicans cells and DMADDM. The pH-dependent antifungal activity of DMADDM was then substantiated in a murine oropharyngeal candidiasis model. We directly demonstrated that the antifungal abilities of quaternary ammonium salts relied on the cell zeta potential which affected the binding between fungal cells and quaternary ammonium salts. These findings suggest a new antifungal mechanism of quaternary ammonium under different pH and that DMADDM can be a potential antifungal agent applied in dental materials and stomatitis therapy.Key Points • DMADDM has stronger antifungal activity in alkaline than in acidic pH conditions. • The pH values and pH-regulated genes can affect the zeta potential of fungal cells. • Zeta potential of fungal cells directly affect the binding between DMADDM and cells. Graphical abstract Schematic diagram of the antifungal activities of DMADDM at different pH values.

Cyclic di-GMP in Streptomycetes: A New Conformation, New Binding Mode, New Receptor, and a New Mechanism to Control Cell Development.

A recent paper by Gallagher etal. (2020) demonstrates that c-di-GMP controls spore formation in Streptomyces venezuelae through sequestering the sporulation sigma factor σWhiG and presents the crystal structure of a ternary complex between c-di-GMP, σWhiG, and its anti-sigma factor, RsiG.

The C3HC type zinc-finger protein (ZFC3) interacting with Lon/MAP1 is important for mitochondrial gene regulation, infection hypha development and longevity of Magnaporthe oryzae

BackgroundThe rice blast is a typical fungal disease caused by Magnaporthe oryzae, and the mitochondrial ATP-dependent Lon protease (MAP1) has been proven to be involved in blast development. We previously screened a C3HC type Zinc-finger domain protein (ZFC3), which is interacted with MAP1. The purpose of this research was to study the biological function of ZFC3 protein in M. oryzae.ResultsWe first confirmed that the ZFC3-RFP fusion protein is localized within the mitochondria. The deleted mutant strains of ZFC3 (∆ZFC3) showed the enhanced expression level of mtATP6, particularly mtATP8, and almost unchanged nATP9. ΔZFC3 produces more conidia and more tolerance to multiple stressors. The knock-out strain shows more melanin accumulation suggests the susceptibility to aging. ΔZFC3 displays faster early-stage hypha infiltration involved in MAP1-mediated pathogenicity in host rice.ConclusionThese results support the view that ZFC3 is a key regulator involved in gene regulation, stress response, cell wall integrity, longevity, conidiation, infection hypha development and MAP1-mediated pathogenicity in M. oryzae.