Hyphal Development Research Articles

The Sclerotinia sclerotiorum ADP-Ribosylation Factor 6 Plays an Essential Role in Abiotic Stress Response and Fungal Virulence to Host Plants.

The ADP-ribosylation factor 6 (Arf6), as the only member of the Arf family III protein, has been extensively studied for its diverse biological functions in animals. Previously, the Arf6 protein in Magnaporthe oryzae was found to be crucial for endocytosis and polarity establishment during asexual development. However, its role remains unclear in S. sclerotiorum. Here, we identified and characterized SsArf6 in S. sclerotiorum using a reverse genetic approach. Deletion of SsArf6 impaired hyphal growth and development and produced more branches. Interestingly, knockout of SsArf6 resulted in an augmented tolerance of S. sclerotiorum towards oxidative stress, and increased its sensitivity towards osmotic stress, indicative of the different roles of SsArf6 in various stress responses. Simultaneously, SsArf6 deletion led to an elevation in melanin accumulation. Moreover, the appressorium formation was severely impaired, and fungal virulence to host plants was significantly reduced. Overall, our findings demonstrate the essential role of SsArf6 in hyphal development, stress responses, appressorium formation, and fungal virulence to host plants.

Synergistic effects of putative Ca2+-binding sites of calmodulin in fungal development, temperature stress and virulence of Aspergillus fumigatus

ABSTRACT In pathogenic fungi, calcium-calmodulin-dependent serine-threonine-specific phosphatase calcineurin is involved in morphogenesis and virulence. Therefore, calcineurin and its tightly related protein complexes are attractive antifungal drug targets. However, there is limited knowledge available on the relationship between in vivo Ca2+-binding sites of calmodulin (CaM) and its functions in regulating stress responses, morphogenesis, and pathogenesis. In the current study, we demonstrated that calmodulin is required for hyphal growth, conidiation, and virulence in the human fungal pathogen, Aspergillus fumigatus. Site-directed mutations of calmodulin revealed that a single Ca2+-binding site mutation had no significant effect on A. fumigatus hyphal development, but multiple Ca2+-binding site mutations exhibited synergistic effects, especially when cultured at 42 °C, indicating that calmodulin function in response to temperature stress depends on its Ca2+-binding sites. Western blotting implied that mutations in Ca2+-binding sites caused highly degraded calmodulin fragments, suggesting that the loss of Ca2+-binding sites results in reduced protein stability. Moreover, normal intracellular calcium homeostasis and the nuclear translocation of the transcriptional factor CrzA are dependent on Ca2+-binding sites of AfCaM, demonstrating that Ca2+-binding sites of calmodulin are required for calcium signalling and its major transcription factor CrzA. Importantly, in situ mutations for four Ca2+-binding sites of calmodulin resulted in an almost complete loss of virulence in the Galleria mellonella wax moth model. This study shed more light on the functional characterization of putative calcium-binding sites of calmodulin in the morphogenesis and virulence of A. fumigatus, which enhances our understanding of calmodulin biological functions in cells of opportunistic fungal pathogens.

Tracing the spatial extent and lag time of carbon transfer from Picea abies to ectomycorrhizal fungi differing in host type, taxonomy, or hyphal development

We used five mature Picea abies continuously labeled with 13C-depleted CO2 in a broadleaf-dominated Swiss forest to assess the spatial extent and lag time of carbon fluxes to ectomycorrhizal fungi differing in hyphal development and host association. We traced labeled carbon into ectomycorrhizal sporocarps collected for two seasons at different distances from labeled Picea. Picea-derived photosynthate reached conifer-specific sporocarps up to 6–12 m away and reached other sporocarps only 0–6 m away. At 0–6 m, genera of lesser hyphal development acquired more Picea-derived photosynthate than those of greater hyphal development, presumably from preferential fungal colonization of inner root zones by the former genera. Correlations of sporocarp δ13C with daily solar radiation integrated for different periods indicated that carbon fluxes from Picea to sporocarps peaked 17–21 days after photosynthesis. Thus, these results provided rough estimates of the spatial extent and temporal lags of carbon transfer from Picea to ectomycorrhizal fungi.

Transcription Factors Tec1 and Tec2 Play Key Roles in the Hyphal Growth and Virulence of Mucor lusitanicus Through Increased Mitochondrial Oxidative Metabolism.

Mucormycosis is a lethal and difficult-to-treat fungal infection caused by fungi of the order Mucorales. Mucor lusitanicus, a member of Mucorales, is commonly used as a model to understand disease pathogenesis. However, transcriptional control of hyphal growth and virulence in Mucorales is poorly understood. This study aimed to investigate the role of Tec proteins, which belong to the TEA/ATTS transcription factor family, in the hyphal development and virulence of M. lusitanicus. Unlike in the genome of Ascomycetes and Basidiomycetes, which have a single Tec homologue, in the genome of Mucorales, two Tec homologues, Tec1 and Tec2, were found, except in that of Phycomyces blakesleeanus, with only one Tec homologue. tec1 and tec2 overexpression in M. lusitanicus increased mycelial growth, mitochondrial content and activity, expression of the rhizoferrin synthetase-encoding gene rfs, and virulence in nematodes and wax moth larvae but decreased cAMP levels and protein kinase A (PKA) activity. Furthermore, tec1- and tec2-overexpressing strains required adequate mitochondrial metabolism to promote the virulent phenotype. The heterotrimeric G beta subunit 1-encoding gene deletant strain (Δgpb1) increased cAMP-PKA activity, downregulation of both tec genes, decreased both virulence and hyphal development, but tec1 and tec2 overexpression restored these defects. Overexpression of allele-mutated variants of Tec1(S332A) and Tec2(S168A) in the putative phosphorylation sites for PKA increased both virulence and hyphal growth of Δgpb1. These findings suggest that Tec homologues promote mycelial development and virulence by enhancing mitochondrial metabolism and rhizoferrin accumulation, providing new information for the rational control of the virulent phenotype of M. lusitanicus.

Reduction of histone proteins dosages increases CFW sensitivity and attenuates virulence of Candida albicans

Histone proteins are important components of nucleosomes, which play an important role in regulating the accessibility of DNA and the function of genomes. However, the effect of histone proteins dosages on physiological processes is not clear in the human fungal pathogen Candida albicans. In this study, we found that the deletion of the histone protein H3 coding gene HHT21 and the histone protein H4 coding gene HHF1 resulted in a significant decrease in the expression dosage of the histone proteins H3 and H4, which had a significant impact on the localization of the histone protein H2A and plasmid maintenance. Stress sensitivity experiments showed that the mutants hht21Δ/Δ, hhf1Δ/Δ and hht21Δ/Δhhf1Δ/Δ were more sensitive to cell wall stress induced by Calcofluor White (CFW) than the wild-type strain. Further studies showed that the decrease in the dosage of the histone proteins H3 and H4 led to the change of cell wall components, increased chitin contents, and down-regulated expression of the SAP9, KAR2, and CRH11 genes involved in the cell wall integrity (CWI) pathway. Overexpression of SAP9 could rescue the sensitivity of the mutants to CFW. Moreover, the decrease in the histone protein s dosages affected the FAD-catalyzed oxidation of Ero1 protein, resulting in the obstruction of protein folding in the ER, and thus reduced resistance to CFW. It was also found that CFW induced a large amount of ROS accumulation in the mutants, and the addition of ROS scavengers could restore the growth of the mutants under CFW treatment. In addition, the reduction of the histone proteins dosages greatly weakened systemic infection and kidney fungal burden in mice, and hyphal development was significantly impaired in the mutants under macrophage treatment, indicating that the histone proteins dosages is very important for the virulence of C. albicans. This study revealed that histone proteins dosages play a key role in the cell wall stress response and pathogenicity in C. albicans.

Exogenous myristate promotes the colonization of arbuscular mycorrhizal fungi in tomato.

Arbuscular mycorrhizal fungi (AMF) can establish symbiotic associations with the roots of most terrestrial plants, thereby improving the tolerance of the host plants to biotic and abiotic stresses. Although AMF cannot synthesize lipids de novo, they can obtain lipids from the root cells for their growth and development. A recent study reveals that AMF can directly take up myristate (C14:0 lipid) from the environment and produce a large amount of hyphae in asymbiotic status; however, the effect of environmental lipids on AM symbiosis is still unclear. In this study, we inoculated tomato (Solanum lycopersicum) with AMF in an in vitro dual culture system and a sand culture system, and then applied exogenous myristate to the substrate, in order to explore the effect of exogenous lipids on the mycorrhizal colonization of AMF. We investigated the hyphae growth, development, and colonization of AMF, and examined the gene expression involved in phosphate transport, lipid biosynthesis, and transport. Results indicate that exogenous lipids significantly stimulated the growth and branching of hyphae, and significantly increased the number of hyphopodia and mycorrhizal colonization of AMF, with arbuscular abundance and intraradical spores or vesicles being the most promoted. In contrast, exogenous myristate decreased the growth range and host tropism of the germ tubes, and largely inhibited the exchange of nutrition between symbionts. As a result, exogenous myristate did not affect the plant growth. This study suggests that lipids promote mycorrhizal colonization by enhancing the growth and development of AMF hyphae and increasing their contact opportunities with plant roots. To the best of our knowledge, this is the first report that shows that lipids promote the colonization of AMF. Our study highlights the importance of better understanding the roles of environmental lipids in the establishment and maintenance of AM symbiosis and, thus, in agricultural production.

Exploration of the Biocontrol Activity of Bacillus atrophaeus Strain HF1 against Pear Valsa Canker Caused by Valsa pyri.

Valsa pyri-induced pear Valsa canker is among the most prevalent diseases to impact pear quality and yields. Biocontrol strategies to control plant disease represent an attractive alternative to the application of fungicides. In this study, the potential utility of Bacillus atrophaeus strain HF1 was assessed as a biocontrol agent against pear Valsa canker. Strain HF1 suppressed V. pyri mycelium growth by 61.20% and induced the development of malformed hyphae. Both culture filtrate and volatile organic compounds (VOCs) derived from strain HF1 were able to antagonize V. pyri growth. Treatment with strain HF1-derived culture filtrate or VOCs also induced the destruction of hyphal cell membranes. Headspace mixtures prepared from strain HF1 were analyzed, leading to the identification of 27 potential VOCs. Of the thirteen pure chemicals tested, iberverin, hexanoic acid, and 2-methylvaleraldehyde exhibited the strongest antifungal effects on V. pyri, with respective EC50 values of 0.30, 6.65, and 74.07 μL L-1. Fumigation treatment of pear twigs with each of these three compounds was also sufficient to prevent the development of pear Valsa canker. As such, these results demonstrate that B. atrophaeus strain HF1 and the volatile compounds iberverin, hexanoic acid, and 2-methylvaleraldehyde exhibit promise as novel candidate biocontrol agents against pear Valsa canker.

Histone acetyltransferase Gcn5-mediated histone H3 acetylation facilitates cryptococcal morphogenesis and sexual reproduction.

Eukaryotic gene transcription is typically regulated by a series of histone modifications, which play a crucial role in adapting to complex environmental stresses. In the ubiquitous human fungal pathogen Cryptococcus neoformans, sexual life cycle is a continuous intracellular differentiation process that strictly occurs in response to mating stimulation. Despite the comprehensive identification of the regulatory factors and genetic pathways involved in its sexual cycle, understanding of the epigenetic modifications involved in this process remains quite limited. In this research, we found that histone acetyltransferase Gcn5-mediated histone H3 acetylation plays a crucial role in completing the cryptococcal sexual cycle, including yeast-hyphae morphogenesis and the subsequent sexual reproduction. Furthermore, we demonstrated that Gcn5 participates in this process primarily through regulating the key morphogenesis regulator Znf2 and its targets. This study thus provided a comprehensive understanding of how histone acetylation modification impacts sexual life cycle in a high-risk human pathogenic fungus.

RNA-Seq-Based Transcriptome Analysis of Nitric Oxide Scavenging Response in Neurospora crassa.

While the biological role of naturally occurring nitric oxide (NO) in filamentous fungi has been uncovered, the underlying molecular regulatory networks remain unclear. In this study, we conducted an analysis of transcriptome profiles to investigate the initial stages of understanding these NO regulatory networks in Neurospora crassa, a well-established model filamentous fungus. Utilizing RNA sequencing, differential gene expression screening, and various functional analyses, our findings revealed that the removal of intracellular NO resulted in the differential transcription of 424 genes. Notably, the majority of these differentially expressed genes were functionally linked to processes associated with carbohydrate and amino acid metabolism. Furthermore, our analysis highlighted the prevalence of four specific protein domains (zinc finger C2H2, PLCYc, PLCXc, and SH3) in the encoded proteins of these differentially expressed genes. Through protein-protein interaction network analysis, we identified eight hub genes with substantial interaction connectivity, with mss-4 and gel-3 emerging as possibly major responsive genes during NO scavenging, particularly influencing vegetative growth. Additionally, our study unveiled that NO scavenging led to the inhibition of gene transcription related to a protein complex associated with ribosome biogenesis. Overall, our investigation suggests that endogenously produced NO in N. crassa likely governs the transcription of genes responsible for protein complexes involved in carbohydrate and amino acid metabolism, as well as ribosomal biogenesis, ultimately impacting the growth and development of hyphae.

Polyphyllin I, a strong antifungal compound against Candida albicans.

This study was performed to explore the antifungal and antibiofilm effects of polyphyllin I (PPI) on Candida albicans. Microdilution assay was performed to determine the minimal inhibitory concentrations (MIC) of PPI against Candida species. Adhesion assay, hyphal growth assay, biofilm formation, and development were used to test the impacts of PPI on C. albicans virulence factors. Propidium iodide staining was performed to test whether the permeability of cell membrane was influenced by PPI. PPI showed significant antifungal activities against several Candida species, with MIC below or equal to 6.25 μM. PPI also inhibited the adhesion to polystyrene surfaces, hyphal growth, and biofilm formation. PPI significantly increased the permeability of C. albicans cell membrane. In sum, PPI can suppress the planktonic growth and biofilm of C. albicans and its mechanism involves the increased permeability of cell membrane.

Artemisinins inhibit oral candidiasis caused by Candida albicans through the repression on its hyphal development

Candida albicans is the most abundant fungal species in oral cavity. As a smart opportunistic pathogen, it increases the virulence by switching its forms from yeasts to hyphae and becomes the major pathogenic agent for oral candidiasis. However, the overuse of current clinical antifungals and lack of new types of drugs highlight the challenges in the antifungal treatments because of the drug resistance and side effects. Anti-virulence strategy is proved as a practical way to develop new types of anti-infective drugs. Here, seven artemisinins, including artemisinin, dihydroartemisinin, artemisinic acid, dihydroartemisinic acid, artesunate, artemether and arteether, were employed to target at the hyphal development, the most important virulence factor of C. albicans. Artemisinins failed to affect the growth, but significantly inhibited the hyphal development of C. albicans, including the clinical azole resistant isolates, and reduced their damage to oral epithelial cells, while arteether showed the strongest activities. The transcriptome suggested that arteether could affect the energy metabolism of C. albicans. Seven artemisinins were then proved to significantly inhibit the productions of ATP and cAMP, while reduced the hyphal inhibition on RAS1 overexpression strain indicating that artemisinins regulated the Ras1-cAMP-Efg1 pathway to inhibit the hyphal development. Importantly, arteether significantly inhibited the fungal burden and infections with no systemic toxicity in the murine oropharyngeal candidiasis models in vivo caused by both fluconazole sensitive and resistant strains. Our results for the first time indicated that artemisinins can be potential antifungal compounds against C. albicans infections by targeting at its hyphal development.

Effector secretion and stability in the maize anthracnose pathogen Colletotrichum graminicola requires N-linked protein glycosylation and the ER chaperone pathway.

N-linked protein glycosylation is a conserved and essential modification mediating protein processing and quality control in the endoplasmic reticulum (ER), but how this contributes to the infection cycle of phytopathogenic fungi is largely unknown. In this study, we discovered that inhibition of protein N-glycosylation severely affected vegetative growth, hyphal tip development, conidial germination, appressorium formation, and, ultimately, the ability of the maize (Zea mays) anthracnose pathogen Colletotrichum graminicola to infect its host. Quantitative proteomics analysis showed that N-glycosylation can coordinate protein O-glycosylation, glycosylphosphatidylinositol anchor modification, and endoplasmic reticulum quality control (ERQC) by directly targeting the proteins from the corresponding pathway in the ER. We performed a functional study of the N-glycosylation pathway-related protein CgALG3 and of the ERQC pathway-related protein CgCNX1, which demonstrated that N-glycosylation of ER chaperone proteins is essential for effector stability, secretion, and pathogenicity of C. graminicola. Our study provides concrete evidence for the regulation of effector protein stability and secretion by N-glycosylation.

MoScd2 is involved in appressorium formation and pathogenicity via the Pmk1 MAPK pathway in Magnaporthe oryzae

Rice blast, known as rice “cancer”, is caused by Magnaporthe oryzae and is particularly serious in Asian and African rice regions. China is also a frequently occurring region of rice blast. Rice blast not only seriously threatens the yield and quality of rice but also affects food security in China. In M. oryzae, the Mst11-Mst7-Pmk1 MAPK signaling pathway mediates pathogenicity by regulating the formation of appressorium and the development of infection hyphae. Stomatal cytokinesis defective 2 (Scd2, also called Ral3 or Bem1) is a component of the Scd complex, which has been proven to be closely related to the MAPK signaling pathway. However, its biological roles in M. oryzae remain elusive. Here, we identified MoScd2, a homologous protein of Schizosaccharomyces pombe Scd2, and preliminarily revealed its role in the development of rice blast fungus. We found that MoScd2 was involved in colony growth, sporulation, spore morphology, spore germination, appressorium formation, turgor in appressoria, mobilization of glycogen from spores to appressoria and pathogenicity. The deletion of MoScd2 resulted in a reduction in Pmk1 and Mps1 phosphorylation levels. In addition, MoScd2 was confirmed to interact with MoMst50, which is a key component of the MAPK signaling pathway in M. oryzae. In summary, MoScd2 was involved in the MAPK signaling pathway of M. oryzae via interaction with MoMst50 to participate in the influence of pathogenicity. In addition, MoScd2 also influences M. oryzae pathogenicity by participating in autophagy.

Bioinspired ferromagnetic CoFe2O4 nanoparticles: Potential pharmaceutical and medical applications

Abstract The primary goal of this work was to develop a cost-effective, non-toxic, eco-friendly, and simple approach for the green synthesis of CoFe2O4 nanoparticles (NPs) using Aloe vera leaf extract by the sol–gel auto-combustion method. In order to figure out their structural, morphological, and magnetic properties, the synthesized NPs were characterized using X-ray diffraction (XRD), Fourier transform infrared (FTIR), scanning electron microscope (SEM), energy-dispersive X-ray spectroscopy (EDX), transmission electron microscope (TEM), dynamic light scattering (DLS), zeta potential, and vibrating-sample magnetometer (VSM). XRD analysis showed that particles had a single-phase spinel crystalline structure with an average crystalline size of 33.5 nm. Under VSM studies, the produced NPs exhibit a soft ferromagnetic property. SEM revealed that the as-prepared NPs were agglomerated due to their magnetic behavior. To the best of our knowledge, the anticandidal, antibiofilm, antibacterial, and anticancer activities of CoFe2O4 NPs toward drug-resistant gram-positive and gram-negative bacteria, as well as fungal strains, have been comprehensively investigated for the first time. The synthesized NPs had a minimal inhibitory concentration of 0.25–0.75 mg/ml against the tested pathogens. CoFe2O4 NPs inhibited the biofilm formation by 37.3–61.8% in selected strains at concentrations of 0.125–0.5 mg/ml. It was observed that the NPs not only suppress biofilm formation but also eradicate established mature biofilms by 50.9–64.49% that was further supported by SEM. SEM analysis shows that NPs significantly inhibit the colonization and aggregation of tested biofilm strains. Light microscopic analysis revealed that NPs completely inhibit the development of hyphae and filaments in Candida albicans, which significantly attenuates their pathogenicity. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays and 4′,6-diamidino-2-phenylindole (DAPI) staining demonstrate that NPs significantly inhibit the proliferation of HCT-116 and HeLa cells. Furthermore, the SEM images of treated cells showed wrinkled and damaged cell walls, indicating the disruption and disorganization of the membrane. This study showed that the synthesized NPs were effective in inhibiting the growth of drug-resistant bacteria, candida, and their preformed biofilms as well. Thus, these NPs with broad-spectrum applications could be exploited in medical settings to diminish biofilm-based infections caused by these pathogenic strains.

ANTIFUNGAL ACTIVITY OF REUTERIN AGAINST AFLATOXIGENIC ASPERGILLUS FLAVUS

Reuterin is a secondary metabolite of the lactic acid bacteria Limosilactobacillus reuteri having broad antimicrobial activity. This study aimed to determine the antifungal activity of reuterin against aflatoxigenic Aspergillus flavus (A. flavus). The aflatoxigenic A. flavus BIO 33212 isolated from Indonesian nutmeg was used in this study. The minimum inhibitory concentration (MIC) of reuterin based on radial growth and colony-forming assays in solid and liquid fat-enriched media against A. flavus was determined. The effect of reuterin on the morphological structure of A. flavus was investigated by scanning electron microscopy (SEM). The aflatoxin B1 (AFB1) produced by A. flavus was evaluated by enzyme-linked immunosorbent assay. Reuterin effectively prolonged the lag phase and decreased the growth rate of A. flavus. At 2 and 4 mM, reuterin demonstrated radial growth inhibition by 5.59% and 35.40%, respectively, while the reductions in colony numbers were 18.36% and 69.51%, respectively. At higher concentrations (6 and 8 mM), the fungal growth was inhibited and reduced completely (100%). SEM revealed the disruption of hyphae and conidiophore development and conidial damage. However, a contradictory phenomenon was found when reuterin at < 6 mM triggered a higher production of AFB1.

Evaluation of the Antifungal Activity of Polysubstituted Cyclic 1,2-Diketones against Colletotrichum gloeosporioides

(1) Anthracnose caused by Colletotrichum damages crops, ornamentals, and forest trees severely, resulting in enormous economic losses to agricultural and forestry systems. Overusing traditional chemical fungicides leads to fungicide resistance, environmental pollution, and potential risks to public health. Therefore, priorities should be given to developing efficient and environmentally friendly approaches to pest management, including anthracnose. (2) In this study, the antifungal activity of botanical derivative polysubstituted cyclic 1,2-diketones (FPL001) against C. gloeosporioides was examined. (3) FPL001 significantly inhibited the vegetative growth of C. gloeosporioides with an EC50 of 160.23 µg/mL. When the concentration of FPL001 reached 30 µg/mL, the conidial germination and appressorium formation of C. gloeosporioides were completely inhibited. FPL001 also significantly suppressed the invasive hyphae development and plant infection of C. gloeosporioides. FPL001 did not exhibit toxicity to model organisms such as alfalfa and silkworm larvae. (4) These results indicate that compound FPL001 is a potential and efficient agent for green control of C. gloeosporioides.

The C2H2 transcription factor SltA is required for germination and hyphal development in Aspergillus fumigatus.

Germination of inhaled Aspergillus fumigatus conidia is a necessary sequitur for infection. Germination of conidia starts with the breaking of dormancy, which is initiated by an increase of the cellular perimeter in a process termed isotropic growth. This swelling phase is followed by polarized growth, resulting in the formation of a germ tube. The multinucleate tubular cells exhibit tip growth from the hyphae, after which lateral branches emerge to form the mycelial network. The regulatory mechanisms governing conidial germination are not well defined. In this study, we identified a novel role for the transcription factor SltA in the orchestration of germination and hyphal development. Conidia lacking sltA fail to appropriately regulate isotropic growth and begin to swell earlier and subsequently switch to polarized growth faster. Additionally, hyphal development is distorted in a ∆sltA isolate as hyphae are hyper-branching and wider, and show branching at the apical tip. ∆sltA conidia are more tolerant to cell wall stressors on minimal medium compared to the wild-type (WT) strain. A transcriptome analysis of different stages of early growth was carried out to assess the regulatory role of SltA. Null mutants generated for three of the most dysregulated genes showed rapid germ tube emergence. Distinct from the phenotype observed for ∆sltA, conidia from these strains lacked defects in isotropic growth, but switched to polarized growth faster. Here, we characterize and describe several genes in the regulon of SltA, highlighting the complex nature of germination.IMPORTANCEAspergillus fumigatus is the main human fungal pathogen causing aspergillosis. For this fungus, azoles are the most commonly used antifungal drugs for treatment of aspergillosis. However, the prevalence of azole resistance is alarmingly increasing and linked with elevated mortality. Germination of conidia is crucial within its asexual life cycle and plays a critical role during the infection in the human host. Precluding germination could be a promising strategy considering the role of germination in Aspergillus spp. pathogenicity. Here, we identify a novel role for SltA in appropriate maintenance of dormancy, germination, and hyphal development. Three genes in the regulon of SltA were also essential for appropriate germination of conidia. With an expanding knowledge of germination and its different morphotypes, more advances can be made toward potential anti-germination targets for therapy.

The MAP Kinase PvMK1 Regulates Hyphal Development, Autophagy, and Pathogenesis in the Bayberry Twig Blight Fungus Pestalotiopsis versicolor.

Bayberry twig blight caused by the ascomycete fungus Pestalotiopsis versicolor is a devastating disease threatening worldwide bayberry production. However, the molecular basis underlying the pathogenesis of P. versicolor is largely unknown. Here, we identified and functionally characterized the MAP kinase PvMk1 in P. versicolor through genetic and cellular biochemical approaches. Our analysis reveals a central role of PvMk1 in regulating P. versicolor virulence on bayberry. We demonstrate that PvMk1 is involved in hyphal development, conidiation, melanin biosynthesis, and cell wall stress responses. Notably, PvMk1 regulates P. versicolor autophagy and is essential for hyphal growth under nitrogen-depleting conditions. These findings suggest the multifaceted role of PvMk1 in regulating P. versicolor development and virulence. More remarkably, this evidence of virulence-involved cellular processes regulated by PvMk1 has paved a fundamental way for further understanding the impact of P. versicolor pathogenesis on bayberry.

Streptococcus mutans sigX-inducing peptide inhibits the virulence of Candida albicans and oral candidiasis through the Ras1-cAMP-Efg1 pathway

Oral candidiasis is the most common fungal infectious disease in the human oral cavity, and Candida albicans is the major pathogenic agent. Increasing drug resistance and the lack of new types of antifungals greatly increase the challenges for treating fungal infections. Targeting hyphal transition provides a promising strategy to inhibit the virulence of C. albicans and overcome drug resistance. This study aimed to investigate the effects and mechanisms of sigX-inducing peptide (XIP), a quorum-sensing signal peptide secreted by Streptococcus mutans, on C. albicans hyphal development and biofilm formation in vitro and oropharyngeal candidiasis in vivo. XIP significantly inhibited C. albicans yeast-to-hypha transition and biofilm formation in a dose-dependent manner from 0.01 to 0.1 µM. XIP significantly downregulated expression of genes from the Ras1-cAMP-Efg1 pathway (RAS1, CYR1, TPK2, EFG1 and UME6), a key pathway to regulate C. albicans hyphal development. Importantly, XIP reduced the levels of key molecules cAMP and ATP from this pathway, while the addition of exogenous cAMP and overexpression of RAS1 restored the hyphal development inhibited by XIP. XIP also lost its hyphal inhibitory effects on ras1Δ/Δ and efg1Δ/Δ strains. These results further confirmed that XIP inhibited hyphal development through downregulation of the Ras1-cAMP-Efg1 pathway. A murine oropharyngeal candidiasis model was employed to evaluate the therapeutic effects of XIP on oral candidiasis. XIP effectively reduced the infected epithelial area, fungal burden, hyphal invasion and inflammatory infiltrates. These results revealed the antifungal effects of XIP, and highlighted that XIP can be a potential antifungal peptide against C. albicans infection.

A transcription factor and a phosphatase regulate temperature-dependent morphogenesis in the fungal plant pathogen Zymoseptoria tritici

Naturally fluctuating temperatures provide a constant environmental stress that requires adaptation. Some fungal pathogens respond to heat stress by producing new morphotypes that maximize their overall fitness. The fungal wheat pathogen Zymoseptoria tritici responds to heat stress by switching from its yeast-like blastospore form to hyphae or chlamydospores. The regulatory mechanisms underlying this switch are unknown. Here, we demonstrate that a differential heat stress response is ubiquitous in Z. tritici populations around the world. We used QTL mapping to identify a single locus associated with the temperature-dependent morphogenesis and we found two genes, the transcription factor ZtMsr1 and the protein phosphatase ZtYvh1, regulating this mechanism. We find that ZtMsr1 regulates repression of hyphal growth and induces chlamydospore formation whereas ZtYvh1 is required for hyphal growth. We next showed that chlamydospore formation is a response to the intracellular osmotic stress generated by the heat stress. This intracellular stress stimulates the cell wall integrity (CWI) and high-osmolarity glycerol (HOG) MAPK pathways resulting in hyphal growth. If cell wall integrity is compromised, however, ZtMsr1 represses the hyphal development program and may induce the chlamydospore-inducing genes as a stress-response survival strategy. Taken together, these results suggest a novel mechanism through which morphological transitions are orchestrated in Z. tritici – a mechanism that may also be present in other pleomorphic fungi.