Sonic restoration: acoustic stimulation enhances plant growth-promoting fungi activity.

Ecosystem restoration interventions often utilize visible elements to restore an ecosystem (e.g. replanting native plant communities and reintroducing lost species). However, using acoustic stimulation to help restore ecosystems and promote plant growth has received little attention. Our study aimed to assess the effect of acoustic stimulation on the growth rate and sporulation of the plant growth-promoting fungus Trichoderma harzianum Rifai, 1969. We played a monotone acoustic stimulus (80 dB sound pressure level (SPL) at a peak frequency of 8 kHz and a bandwidth at -10 dB from the peak of 6819 Hz-parameters determined via review and pilot research) over 5 days to T. harzianum to assess whether acoustic stimulation affected the growth rate and sporulation of this fungus (control samples received only ambient sound stimulation less than 30 dB). We show that the acoustic stimulation treatments resulted in increased fungal biomass and enhanced T. harzianum conidia (spore) activity compared to controls. These results indicate that acoustic stimulation influences plant growth-promoting fungal growth and potentially facilitates their functioning (e.g. stimulating sporulation). The mechanism responsible for this phenomenon may be fungal mechanoreceptor stimulation and/or potentially a piezoelectric effect; however, further research is required to confirm this hypothesis. Our novel study highlights the potential of acoustic stimulation to alter important fungal attributes, which could, with further development, be harnessed to aid ecosystem restoration and sustainable agriculture.

The conserved WetA protein involved in asexual development in fungi is localized to the nuclei in the asexual spores of Paecilomyces variotii.

Mutations have underpinned research into gene characterization across all domains of life. This includes the discovery of the genes involved in the development of asexual spores in filamentous fungi. Mutants in the ascomycete Paecilomyces variotii were isolated with impaired biosynthesis of the characteristic yellow pigment produced by this filamentous fungus. The affected genes were identified as pvpP, encoding the polyketide synthase that is required for synthesis of the pigment YWA1, and abaA and wetA that are two genes that encode components of the AbaA-BrlA-WetA module required for the development of asexual spores in species in the Eurotiales order. WetA was further characterized. A strain expressing a functional WetA-GFP fusion was created and used to find that WetA is expressed primarily in spores and concentrated in their nuclei, providing evidence that this conserved protein likely functions as a regulator of transcription in conidia. Analysis of the phenotypes of the P. variotii wetA mutant suggests that how this three-protein module impacts fungal biology will vary from species-to-species, despite being conserved amongst filamentous Ascomycete species.

Micro PIXE mapping proves a differential distribution and concentration of trace elements in fungal structures of Rhizophagus intraradices.

Arbuscular mycorrhizal (AM) fungi can sequester different potentially toxic elements, such as trace elements (TEs), within their structures to alleviate the toxicity for its host plant and themselves. To elucidate the role of AM fungi in TEs immobilization in the rhizosphere of host plants, it is important to know the TEs distribution in AM fungal structures. In the present study, we investigated the distribution and concentration of TEs within extraradical spores and mycelium of the AM fungus Rhizophagus intraradices, collected from the rhizosphere of Senecio bonariensis plants grown in a soil polluted with multiple TEs, by using Particle-Induced X-ray Emission with a micro-focused beam (micro PIXE). This technique enabled the simultaneous micrometric mapping of elements in a sample. The calculated values were compared with those in the polluted substrate, measured by the Wavelength Dispersive X-ray Fluorescence technique. The highest concentrations of Fe, P, Ti, Mn, Cr, Cu and Zn were found in AM fungal spores, where they were accumulated, while extraradical mycelium was enriched in Cu. Finally, we demonstrated that AM fungi can simultaneously accumulate high amounts of different TEs in their structures, thus reducing the toxicity of these elements to its host plant.

Morpho-molecular characterization of phoma-like fungi from Morus alba in northern Thailand; a novel species (Boeremia albae) and a new host record (B. maritima).

Boeremia was established to accommodate phoma-resembling fungi. Its species occur in terrestrial ecosystems as endophytes, saprobes and pathogens, except one species reported from a marine ecosystem. Boeremia species are characterized by hyaline, thin-walled, and aseptate (occasionally 1(-2)-septate) conidia that are variable in shape, and hyaline, straight or slightly curved, thick-walled, and 1-septate ascospores that are usually constricted at the septum. In the past, host associations were used to delimit Boeremia species. However, since Boeremia taxa have overlapping morphological characters and are cryptic, it renders taxonomic identification arduous. Therefore, the use of other approaches including multi-gene phylogenetic analyses are imperative. Recommended DNA markers for species delineation are the internal transcribed spacer (ITS, nuclear rDNA consisting of ITS1-5.8S-ITS2) and large subunit (28S, D1-D2 domains of nuclear 28S rDNA) loci, and the genes for actin (ACT1), beta-tubulin (TBB1), RNA polymerase 2 (RPB2) and translation elongation factor 1α (TEF1). Here, we applied morphological and molecular phylogenetic analyses to establish a new taxon (B. albae), and a new host and geographical record for B. maritima associated with leaf spots of Morus alba (Moraceae) in northern Thailand. By providing sequence data for three additional gene regions, our phylogenetic analyses impart a stable phylogenetic placement of the ex-type strain of B. maritima, as illustrated. This is the first study that reports Boeremia species from M. alba, and B. maritima from a terrestrial habitat.

Multifaceted roles of NADPH oxidases in the growth and pathogenicity of Beauveria bassiana.

Reactive oxygen species (ROS), synthesized by the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (Nox) complex, are vital molecules in biological cells, influencing various physiological processes such as fungal growth, development, and virulence. Beauveria bassiana, an entomopathogenic fungus, is a promising biopesticide for agricultural, forestry, and urban pest control. This study focuses on the characterization of NADPH oxidases (Noxs) in B. bassiana. Gene expression profiles of Noxs in B. bassiana (BbNoxs) were analysed using RT-qPCR. Knockout strains of single BbNoxA, BbNoxB, BbNoxR, and double BbNoxA and BbNoxB were constructed via homologous recombination, and their phenotypic characteristics were examined. Fungal virulence was evaluated using Galleria mellonella larvae, and infection structures formation and penetration ability were assessed on cicada wings. ROS production and actin assembly during fungal growth and infection were detected using staining and marker methods. Expression analysis revealed significant upregulation of BbNoxs during fungal growth and infection. Compared to the wild-type strain, single knockouts (ΔBbNoxA/B/R) and double knockout (ΔBbNoxAB) of BbNoxs exhibited reduced conidial yields, accelerated conidial germination rates. Deletion of BbNoxB or BbNoxR decreased fungal virulence compared to the WT strain in topical inoculation experiments. Additionally, loss of BbNoxB or BbNoxR impaired infection structures formation, penetration ability, ROS production, and actin aggregation during fungal infection. BbNoxs are crucial for fungal growth, development, and virulence in B. bassiana, playing essential roles in infection structures formation, penetration, ROS production, and actin assembly. Understanding their functions provides insights into B. bassiana's pathogenic mechanisms.

Leaf Rust Pathogens on Hypericum pseudohenryi: Describing Melampsora danbaensis sp. nov. and M. hyperici-pseudohenryi sp. nov. from China.

Based on morphological and phylogenetic evidence, two novel species of Melampsora were discovered on Hypericum pseudohenryi in China and have been thoroughly characterized. One of these species, designated as M. danbaensis, exhibits distinct features such as aecia of Uredo-type, typically appearing in gregarious or grouped arrangements, and presenting a shallowly pulvinate structure. Aeciospores exhibit tremendous variations in size, ranging in shape from globose to ellipsoidal and bearing pronounced verrucose texture. Telia resemble crusts one-spore deep, covering nearly the entire abaxial leaf surface, with sessile teliospores reaching sizes of up to 65.8 µm, and exhibiting a clavate to cylindrical shape. Another species, designated as M. hyperici-pseudohenryi, is distinguished by Uredo-type uredinia, which are hypophyllous, scattered or grouped, and interspersed with numerous paraphyses. Its urediniospores tend to be globose, ellipsoidal or obovoid, echinulate, and are accompanied by clavate to capitate paraphyses reaching lengths up to 77.6 µm. Phylogenetically, both species form a novel monophyletic clade within the Melampsora genus, with robust support demonstrated by a high Maximum likelihood bootstrap support (MLBS) value of 100% and a Bayesian posterior probability (BPP) of 1. This study enriches our understanding of the diversity and geographical distribution of Melampsora species that infect Hypericum plants in China.

How Metarhizium robertsii's mycelial consciousness gets its conidia Zen-ready for stress.

This memoir takes a whimsical ride through my professional adventures, spotlighting my fungal stress research on the insect-pathogenic fungus Metarhizium robertsii, which transformed many of my wildest dreams into reality. Imagine the magic of fungi meeting science and me, a happy researcher, arriving at Utah State University ready to dive deep into studies with the legendary insect pathologist, my advisor Donald W. Roberts, and my co-advisor Anne J. Anderson. From my very first "Aha!" moment in the lab, I plunged into a vortex of discovery, turning out research like a mycelium on a mission. Who knew 18 h/day, seven days a week, could be so exhilarating? I was fueled by an insatiable curiosity, boundless creativity, and a perhaps slightly alarming level of motivation. Years later, I managed to bring my grandest vision to life: the International Symposium on Fungal Stress-ISFUS. This groundbreaking event has attracted 162 esteemed speakers from 29 countries to Brazil, proving that fungi can be both fun and globally fascinating. ISFUS is celebrating its fifth edition in 2024, a decade after its 2014 debut.

Adaptative responses of Neurospora crassa by histidine kinases upon the attack of the arthropod Sinella curviseta.

Histidine kinases (HKs) are important sensor proteins in fungi and play an essential role in environmental adaptation. However, the mechanisms by which fungi sense and respond to fungivores attack via HKs are not fully understood. In this study, we utilized Neurospora crassa to investigate the involvement of HKs in responding to fungivores attack. We found that the 11 HKs in N. crassa not only affected the growth and development, but also led to fluctuations in antioxidant production. Ten mutants in the genes encoding HKs (except ∆phy1) showed increased production of reactive oxygen species (ROS), especially upon Sinella curviseta attack. The ROS burst triggered changes in conidia and perithecial beaks formation, as well as accumulation of ß-glucan, ergothioneine, ergosterol, and carotenoids. ß-glucan was increased in ∆hk9, ∆os1, ∆hcp1, ∆nik2, ∆sln1, ∆phy1 and ∆phy2 mutants compared to the wild-type strain. In parallel, ergothioneine accumulation was improved in ∆phy1 and ∆hk16 mutants and further increased upon attack, except in ∆os1 and ∆hk16 mutants. Additionally, fungivores attack stimulated ergosterol and dehydroergosterol production in ∆hk9 and ∆os1 mutants. Furthermore, deletion of these genes altered carotenoid accumulation, with wild-type strain, ∆hk9, ∆os1, ∆hcp1, ∆sln1, ∆phy2, and ∆dcc1mutants showing an increase in carotenoids upon attack. Taken together, HKs are involved in regulating the production of conidia and antioxidants. Thus, HKs may act as sensors of fungivores attack and effectively improve the adaptive capacity of fungi to environmental stimuli.

Fusarium graminearum spores disrupt gut microbiota and metabolome via the lung-gut axis in mice.

Fusarium graminearum, the primary pathogen responsible for wheat Fusarium head blight, can induce pulmonary damage through its spores. However, the detailed mechanism by which these spores cause intestinal injury is not yet fully understood. This study aimed to investigate the impact of exposure to fungal spores on the intestinal microbiota using a mice model that mimics the effects of fusarium graminearum spores on the gut microbiota and its metabolic profile. The study utilized 16S rRNA sequencing and metabolomics methodologies to analyze the contents of the cecum and feces in mice. The results showed that exposure to fungal spores led to significant changes in the composition of the intestinal microbiota in mice, characterized by an increase in Akkermansia and Staphylococcus populations. A non-targeted metabolomics analysis identified 316 metabolites associated with various metabolic pathways, particularly galactose metabolism. Pre-exposure to antibiotics before fungal spore exposure resulted in a decrease in the metabolic capacity of the intestinal microbiota in mice. This research demonstrates that fusarium graminearum spores can disrupt the intestinal microbiota and metabolome via the lung-gut axis. These findings provide valuable insights into the intestinal damage caused by fungal spores and offer important support for the development of therapeutic strategies for intestinal diseases.

Acptp2,3 participates in the regulation of spore production, stress response, and pigments synthesis in Aspergillus cirstatus.

Background: Aspergillus cristatus was a filamentous fungus that produced sexual spores under hypotonic stress and asexual spores under hypertonic stress. It could be useful for understanding filamentous fungi's sporulation mechanism. Previously, we conducted functional studies on Achog1, which regulated the hyperosmotic glycerol signaling (HOG) pathway and found that SI65_02513 was significantly downregulated in the transcriptomics data of ΔAchog1 knockout strain. This gene was located at multiple locations in the HOG pathway, indicating that it might play an important role in the HOG pathway of A. cristatus. Furthermore, the function of this gene had not been identified in Aspergillus fungi, necessitating further investigation. This gene's conserved domain study revealed that it has the same protein tyrosine phosphatases (PTPs) functional domain as Saccharomyces cerevisiae, hence SI65_02513 was named Acptp2,3. Methods: The function of this gene was mostly validated using gene knockout and gene complementation approaches. Knockout strains exhibited sexual and asexual development, as well as pigments synthesis. Morphological observations of the knockout strain were carried out under several stress conditions (osmotic stress, oxidative stress, Congo Red, and sodium dodecyl sulfate (SDS). Real-time fluorescence polymerase chain reaction (PCR) identified the expression of genes involved in sporulation, stress response, and pigments synthesis. Results: The deletion of Acptp2,3 reduced sexual and asexual spore production by 4.4 and 4.6 times, demonstrating that Acptp2,3 positively regulated the sporulation of A. cristatus. The sensitivity tests to osmotic stress revealed that ΔAcptp2,3 strains did not respond to sorbitol-induced osmotic stress. However, ΔAcptp2.3 strains grew considerably slower than the wild type in high concentration sucrose medium. The ΔAcptp2,3 strains grew slower than the wild type on media containing hydrogen peroxide, Congo red, and SDS. These findings showed that Acptp2,3 favorably controlled osmotic stress, oxidative stress, and cell wall-damaging chemical stress in A. cristatus. Deleting Acptp2,3 resulted in a deeper colony color, demonstrating that Apctp2,3 regulated pigment synthesis in A. cistatus. The expression levels of numerous stress-and pigments-related genes matched the phenotypic data. Conclusion: According to our findings, Acptp2,3 played an important role in the regulation of sporulation, stress response, and pigments synthesis in A. cristatus. This was the first study on the function of PTPs in Aspergillus fungi.

Disseminated emergomyces orientalis infection in a patient with systemic lupus erythematosus.

A case of Eimonosis orientalis was reported in a 52-year-old male farmer who presented with cough, phlegm, fever, headache, and nausea for more than 4 days. Haemophilic cells and fungal spores were identified in the bone marrow smear and confirmed as Aemon orientalis by culture. The same bacteria were also isolated from blood cultures.

Accumulated melanin in molds provides wavelength-dependent UV tolerance.

Fungal contamination poses a serious threat to public health and food safety because molds can grow under stressful conditions through melanin accumulation. Although ultraviolet (UV) irradiation is popular for inhibiting microorganisms, its effectiveness is limited by our insufficient knowledge about UV tolerance in melanin-accumulating molds. In this study, we first confirmed the protective effect of melanin by evaluating the UV sensitivity of young and mature spores. Additionally, we compared UV sensitivity between spores with accumulated melanin and spores prepared with melanin biosynthesis inhibitors. We found that mature spores were less UV-sensitive than young spores, and that reduced melanin accumulation by inhibitors led to reduced UV sensitivity. These results suggest that melanin protects cells against UV irradiation. To determine the most effective wavelength for inhibition, we evaluated the wavelength dependence of UV tolerance in a yeast (Rhodotorula mucilaginosa) and in molds (Aspergillus fumigatus, Cladosporium halotolerans, Cladosporium sphaerospermum, Aspergillus brasiliensis, Penicillium roqueforti, and Botrytis cinerea). We assessed UV tolerance using a UV-light emitting diode (LED) irradiation system with 13 wavelength-ranked LEDs between 250 and 365 nm, a krypton chlorine (KrCl) excimer lamp device, and a low pressure (LP) Hg lamp device. The inhibition of fungi peaked at around 270 nm, and most molds showed reduced UV sensitivity at shorter wavelengths as they accumulated pigment. Absorption spectra of the pigments showed greater absorption at shorter wavelengths, suggesting greater UV protection at these wavelengths. These results will assist in the development of fungal disinfection systems using UV, such as closed systems of air and water purification.

Biocontrol potential of Bacillus velezensis HG-8-2 against postharvest anthracnose on chili pepper caused by Colletotrichum scovillei.

Anthracnose caused by Colletotrichum scovillei is a significant disease of pepper, including in postharvest stage. Bacillus species represent a potential microbial resource for controlling postharvest plant diseases. Here, a strain HG-8-2 was obtained and identified as Bacillus velezensis through morphological, biochemical, physiological, and molecular analyses. The culture filtrate showed highly antifungal activity against C. scovillei both in vitro and on pepper fruit. Crude lipopeptide extracts, which had excellent stability, could effectively inhibit mycelial growth of C. scovillei with an EC50 value of 28.48 ± 1.45 µg mL-1 and inhibited conidial germination. Pretreatment with the extracts reduced the incidence and lesion size of postharvest anthracnose on pepper fruit. Analysis using propidium iodide staining, malondialdehyde content detection and scanning electron microscope observation suggested that the crude lipopeptide extracts harbored antifungal activity by damaging cell membranes and mycelial structures. The RNA-seq analysis conducted on C. scovillei samples treated with the extracts, as compared to untreated samples, revealed significant alterations in the expression of multiple genes involved in protein biosynthesis. Overall, these results demonstrated that B. velezensis HG-8-2 and its crude lipopeptide extracts exhibit highly antagonistic ability against C. scovillei, thereby offering an effective biological agent for the control of anthracnose in pepper fruit.

Innovative sustainable bioreactor-in-a-granule formulation of Trichoderma asperelloides.

The advancement of fungal biocontrol agents depends on replacing cereal grains with low-cost agro-industrial byproducts for their economical mass production and development of stable formulations. We propose an innovative approach to develop a rice flour-based formulation of the beneficial biocontrol agent Trichoderma asperelloides CMAA1584 designed to simulate a micro-bioreactor within the concept of full biorefinery process, affording in situ conidiation, extended shelf-life, and effective control of Sclerotinia sclerotiorum, a devastating pathogen of several dicot agricultural crops worldwide. Rice flour is an inexpensive and underexplored byproduct derived from broken rice after milling, capable of sustaining high yields of conidial production through our optimized fermentation-formulation route. Conidial yield was mainly influenced by nitrogen content (0.1% w/w) added to the rice meal coupled with the fermentor type. Hydrolyzed yeast was the best nitrogen source yielding 2.6 × 109 colony-forming units (CFU)/g within 14 days. Subsequently, GControl, GLecithin, GBreak-Thru, GBentonite, and GOrganic compost+Break-Thru formulations were obtained by extrusion followed by air-drying and further assessed for their potential to induce secondary sporulation in situ, storage stability, and efficacy against Sclerotinia. GControl, GBreak-Thru, GBentonite, and GOrganic compost+Break-Thru stood out with the highest number of CFU after sporulation upon re-hydration on water-agar medium. Shelf-life of formulations GControl and GBentonite remained consistent for > 3 months at ambient temperature, while in GBentonite and GOrganic compost+Break-Thru formulations remained viable for 24 months during refrigerated storage. Formulations exhibited similar efficacy in suppressing the myceliogenic germination of Sclerotinia irrespective of their concentration tested (5 × 104 to 5 × 106 CFU/g of soil), resulting in 79.2 to 93.7% relative inhibition. Noteworthily, all 24-month-old formulations kept under cold storage successfully suppressed sclerotia. This work provides an environmentally friendly bioprocess method using rice flour as the main feedstock to develop waste-free granular formulations of Trichoderma conidia that are effective in suppressing Sclerotinia while also improving biopesticide shelf-life. KEY POINTS: • Innovative "bioreactor-in-a-granule" system for T. asperelloides is devised. • Dry granules of aerial conidia remain highly viable for 24 months at 4 °C. • Effective control of white-mold sclerotia via soil application of Trichoderma-based granules.

Engineering Entomopathogenic Fungi Using Thermal-Responsive Polymer to Boost Their Resilience against Abiotic Stresses.

Entomopathogenic fungi offer an ecologically sustainable and highly effective alternative to chemical pesticides for managing plant pests. However, the efficacy of mycoinsecticides in pest control suffers from environmental abiotic stresses, such as solar UV radiation and temperature fluctuations, which seriously hinder their practical application in the field. Herein, we discovered that the synthetic amphiphilic thermal-responsive polymers are able to significantly enhance the resistance of Metarhizium robertsii conidia against thermal and UV irradiation stresses. The thermosensitive polymers with extremely low cytotoxicity and good biocompatibility can be engineered onto the M. robertsii conidia surface by anchoring hydrophobic alkyl chains. Further investigations revealed that polymer supplementation remarkably augmented the capacity for penetration and the virulence of M. robertsii under heat and UV stresses. Notably, broad-spectrum entomopathogenic fungi can be protected by the polymers. The molecular mechanism was elucidated through exploring RNA sequencing and in vivo/vitro enzyme activity assays. This work provides a novel avenue for fortifying the resilience of entomopathogenic fungi, potentially advancing their practical application as biopesticides.

Antifungal effect of cinnamon essential oil against Penicillium oxalicum on rice noodles.

Plant essential oils have been extensively investigated for their application in food industry due to their broad antimicrobial spectrum and safety. However, rare studies investigated their application in decontaminating rice noodles from fungal contamination. In this study, the cinnamon essential oil was screened out among 12 species of plant essential oils, and its antifungal activity against Penicillium oxalicum isolated from rice noodles was investigated. Our study revealed that cinnamon essential oil inhibited the spore germination in a concentration-dependent manner, and a dosage of 0.025% (v/v) could entirely disable the spore germination. The disruption of the fungal plasma membrane was evidenced by the change of plasma membrane permeability and the leakage of cellular components. The cinnamon essential oil in vapor phase (0.00625% [v/v]) could totally inhibit the growth of fungi inoculated on rice noodles. In addition to the potential application in inactivating fungi germination on rice noodles, this study also demonstrated the feasibility of cinnamon essential as an environmental disinfectant. This study is the first report that cinnamon essential oil has been studied for decontaminating rice noodles from fungal contamination with P. oxalicum, which not only broadens the application field of plant essential oil but also provides an alternative approach for rice noodle preservation.

Impact of metal ions on PMS/Cl- disinfection efficacy: Enhancing or impeding microbial inactivation?

Peroxymonosulfate (PMS) is an eco-friendly disinfectant gaining attention. This study examined the influence of metal ions (Co(II), Cu(II), Fe(II)) on PMS disinfection with chloride ions (Cl-) against waterborne microorganisms, encompassing both bacteria and fungal spores. The findings elucidated that metal ions augment the inactivation of bacteria in the PMS/Cl- system while concurrently impeding the inactivation of fungal spores. Specifically, the PMS/Co(II)/Cl- process increased E. coli inactivation rates by 2.25 and 2.75 times compared to PMS/Co(II) and PMS/Cl-, respectively. Conversely, PMS/Me(II)/Cl- generally exhibited a diminished inactivation capacity against the three fungal spores compared to PMS/Cl-, albeit surpassing the efficacy of PMS/Me(II). For instance, the inactivation levels of A. niger by PMS/Cl-, PMS/Cu(II)/Cl-, and PMS/Cu(II) are 4.47-log, 1.92-log, and 0.11-log, respectively. Notably, fungal spores demonstrated a substantially higher resistance to disinfectants compared to bacteria. Differences in microbial susceptibility were linked to cell wall structure, composition, antioxidant defenses, and reactive species generation, such as hydroxyl radicals (•OH), sulfate radicals (SO4•-), and reactive chlorine species (RCS). This study demonstrated the novel and unique phenomenon of metal ions' dual role in modulating the PMS/Cl- disinfection process, which has not been reported before and has important implications for the field of water treatment.

The Fungal Transcription Factor BcTbs1 from Botrytis cinerea Promotes Pathogenicity via Host Cellulose Degradation.

Zn(II)2Cys6 proteins constitute the largest group of fungal-specific transcription factors. However, little is known about their functions in the crop killer Botrytis cinerea. In this work, a T-DNA insertion strain M13448 was identified which was inserted into the Zn(II)2Cys6 TF-encoding gene BcTBS1. Knockout of BcTBS1 did not affect mycelia growth, appressorium formation, and sclerotium germination, but impaired fungal conidiation, conidial morphogenesis, conidial germination, infection cushion development, and sclerotial formation. Accordingly, ΔBctbs1 mutants showed reduced virulence in its host plants. Further study proved that BcTBS1, BCIN_15g03870, and BCIN_12g06630 were induced by cellulose. Subsequent cellulase activity assays revealed that the loss of BcTBS1 significantly decreased cellulase activity. In addition, we verified that the BCIN_15g03870 and BCIN_12g06630 genes were positive regulated by BcTBS1 by quantitative real-time reverse-transcription-polymerase chain reaction (qRT-PCR). Taken together, these results suggested that BcTBS1 can promote pathogenicity by modulating cellulase-encoding genes that participate in host cellulose degradation.

The SUMO gene MrSmt3 is involved in SUMOylation, conidiation and stress response in Metarhizium robertsii.

Smt3, as a small ubiquitin-like modifier (SUMO), play an essential role in the regulation of protein SUMOylation, and thus this process can affect various important biological functions. Here, we investigated the roles of MrSmt3 (yeast SUMO/Smt3 homologs) in the entomopathogenic fungus Metarhizium robertsii. Our results of subcellular localization assays demonstrated that MrSmt3 was present in the cytoplasm and nucleus, whereas MrSmt3 was largely localized in the nucleus during oxidative stress. Importantly, disruption of MrSmt3 significantly decreased the level of protein SUMOylation under heat stress. Deletion of MrSmt3 led to a significant decrease in conidial production, and increased sensitivity to various stresses, including heat, oxidative, and cell wall-disturbing agents. However, bioassays of direct injection and topical inoculation demonstrated that deletion of MrSmt3 did not affect fungal virulence. Furthermore, RNA-seq analysis identified 1,484 differentially expressed genes (DEGs) of the WT and ΔMrSmt3 during conidiation, including 971 down-regulated DEGs and 513 up-regulated DEGs, and further analysis showed that the expression level of several classical conidiation-associated genes, such as transcription factor AbaA (MAA_00694), transcription factor bZIP (MAA_00888) and transcription factor Ste12 (MAA_10450), was down-regulated in the ΔMrSmt3 mutant. Specifically, the major downregulated DEGs were mainly associated with a variety of metabolic regulatory processes including metabolic process, organic substance metabolic process and primary metabolic process. Collectively, our findings highlight the important roles of the SUMO gene MrSmt3 in modulating SUMOylation, conidiation and stress response in M. robertsii.

Evolutionarily diverse fungal zoospores show contrasting swimming patterns specific to ultrastructure.

Zoosporic fungi, also called chytrids, produce single-celled motile spores with flagellar swimming tails (zoospores).1,2 These fungi are key components of aquatic food webs, acting as pathogens, saprotrophs, and prey.3,4,5,6,7,8 Little is known about the swimming behavior of fungal zoospores, a crucial factor governing dispersal, biogeographical range, ecological function, and infection dynamics.6,9 Here, we track the swimming patterns of zoospores from 12 evolutionarily divergent species of zoosporic fungi from across seven orders of the Chytridiomycota and the Blastocladiomycota. We report two major swimming patterns that correlate with the cytoskeletal ultrastructure of these zoospores. Specifically, we show that species without major cytoplasmic tubulin components swim in a circular fashion, while species with prominent cytoplasmic tubulin structures swim in a pattern akin to a random walk (move-stop-redirect-move). We confirm cytoskeletal architecture by performing fluorescence confocal microscopy across all 12 species. We then treat representative species with variant swimming behaviors and cytoplasmic-cytoskeletal arrangements with tubulin-stabilizing (Taxol) and depolymerizing (nocodazole) pharmacological compounds. We observed that when treating the "random walk" species with nocodazole, their swimming behavior changed to a circular-swimming pattern. Confocal imaging of the nocodazole-treated zoospores demonstrates that these cells maintain flagellum tubulin structures but lack their characteristic cytoplasmic tubulin structures. Our data demonstrate that the capability of zoospores to perform "complex" random-walk movement is linked to the presence of prominent cytoplasmic tubulin structures and suggest a link between cytology, sensory systems, and swimming behavior in a diversity of zoosporic fungi.