Insect Pathogenic Fungus Research Articles

Discovery of Gibellula floridensis from Infected Spiders and Analysis of the Surrounding Fungal Entomopathogen Community.

Characterization of fungal spider pathogens lags far behind their insect counterparts. In addition, little to nothing is known concerning the ecological reservoir and/or fungal entomopathogen community surrounding infection sites. Five infected spider cadavers were identified in the neo-tropical climate of north-central Florida, USA, from three of which viable cultures were obtained. Multi-locus molecular phylogenetic and morphological characterization identified one isolate as a new Gibellula species, here named, Gibellula floridensis, and the other isolates highly similar to Parengyodontium album. The fungal entomopathogen community surrounding infected spiders was sampled at different habitats/trophic levels, including soil, leaf litter, leaf, and twig, and analyzed using ITS amplicon sequencing. These data revealed broad but differential distribution of insect-pathogenic fungi between habitats and variation between sites, with members of genera belonging to Metarhizium and Metacordyceps from Clavicipitaceae, Purpureocillium and Polycephalomyces from Ophiocordyceps, and Akanthomyces and Simplicillium from Cordycipitaceae predominating. However, no sequences corresponding to Gibellula or Parengyodontium, even at the genera levels, could be detected. Potential explanations for these findings are discussed. These data highlight novel discovery of fungal spider pathogens and open the broader question regarding the environmental distribution and ecological niches of such host-specific pathogens.

Metarhizium caribense sp. nov., a Novel Species of Entomopathogenic Metarhizium Fungi Associated with Weevils Impairing Coffee, Sugar Cane and Sweet Potato Cultivation.

(1) Background: Insect pathogenic fungi of the genus Metarhizium are under study and in application as highly solicited, more eco-system friendly substitutes for chemical insecticides in many countries and in different agricultural contexts. In Cuba and Florida, Metarhizium strains have previously been isolated from economically important coffee and sugar cane pests. (2) Methods: Unambiguous species delineation within the Metarhizium anisopliae species complex is methodologically challenging. Recently, a species-discriminating PCR approach has been developed based on ribosomal intergenic spacer (rIGS) sequences that covered the prominent four "PARB" species within the complex. This approach is combined here with further genetic markers and is extended to a further species. (3) Results: Metarhizium isolates from Cuba, found to be more naturally associated with the coffee berry borer, Hypothenemus hampei, were morphologically, microscopically and molecular taxonomically characterized. Multilocus sequence analysis based on 5TEF, MzIGS3 and rIGS markers delineated these weevil-associated strains from all previously established Metarhizium species. (4) Conclusions: The isolates under study represent a new fungal taxon proposed to be designated Metarhizium caribense. The rIGS-based species-discriminating diagnostic PCR is a suitable tool for the identification of new Metarhizium species and can be productively combined to approaches using further genetic markers.

Root symbiotic fungi improve nitrogen transfer and morpho-physiological performance in Chenopodium quinoa.

Root-associated fungal endophytes may facilitate nitrogen (N) absorption in plants, leading to benefits in photosynthesis and growth. Here, we investigated whether endophytic insect pathogenic fungi (EIPF) are capable of transferring soil N to the crop species Chenopodium quinoa. We evaluated nutrient uptake, carbon allocation, and morpho-physiological performance in C. quinoa in symbiosis with two different EIPF (Beauveria and Metarhizium) under contrasting soil N supply. A controlled experiment was conducted using two plant groups: (1) plants subjected to low N level (5 mM urea) and (2) plants subjected to high N level (15 mM urea). Plants from each group were then inoculated with different EIPF strains, either Beauveria (EIPF1+), Metarhizium (EIPF2+) or without fungus (EIPF-). Differences in N and C content, amino acids, proteins, soluble sugars, starch, glutamine synthetase, glutamate dehydrogenase, and physiological (photosynthesis, stomatal conductance, transpiration), and morphological performance between plant groups under each treatment were examined. We found that both Beauveria and Metarhizium translocated N from the soil to the roots of C. quinoa, with positive effects on photosynthesis and plant growth. These effects, however, were differentially affected by fungal strain as well as by N level. Additionally, an improvement in root C and sugar content was observed in presence of EIPF, suggesting translocation of carbohydrates from leaves to roots. Whereas both strains were equally effective in N transfer to roots, Beauveria seemed to exert less demand in C. quinoa for photosynthesis-derived carbohydrates compared to Metarhizium. Our study revealed positive effects of EIPF on N transfer and morpho-physiological performance in crops, highlighting the potential of these fungi as an alternative to chemical fertilizers in agriculture systems.

Interactions between migration and immunity among oriental armyworm populations infected with the insect pathogenic fungus, Beauveria bassiana.

Migration and immunity are behavioral and physiological traits that protect organisms from environmental stressors or pathogen infection. Shifting from migration to residency has become more common in some wildlife populations owing to environmental changes. However, other biological shifts, such as interactions between migration and immunity among populations within a species are largely unexplored for many agricultural migratory pests. In the field, entomopathogenic fungi infection and transmission, particularly Beauveria bassiana, can cause reduced fitness and population declines across a broad range of insect species. Here, we investigated migration-immunity interactions between migrant and resident populations of the oriental armyworm, Mythimna separata, infected with B. bassiana (the sole fungus used in this work). We found that migratory M. separata exerted stronger pathogen resistance, faster development and lower pupal weight than residents. High-dose infections (5.0 × 105 and 5.0 × 106 conidia mL-1) led to seriously decreased reproductive capacity in migrants and residents. Low-dose infections (1.0 × 104 and 5.0 × 104 conidia mL-1) led to significantly increased host flight capacities. Consecutive flight tests showed that five flight nights inhibited the reproduction of paternal infected M. separata populations. The flights also led to far-reaching transgenerational impairment of larval development and immune defense among offspring populations. By contrast, two flight nights enhanced the reproductive capacities of both M. separata populations and did not exert negative transgenerational effects on offspring populations, which may facilitate migration. This study provides insights into interactions between migration and immunity among M. separata populations. These insights will guide development of future monitoring and management technologies of this pest. © 2024 Society of Chemical Industry.

Unlocking the Siderophore Biosynthesis Pathway and Its Biological Functions in the Fungal Insect Pathogen Beauveria bassiana.

Siderophores are small molecule iron chelators. The entomopathogenic fungus Beauveria bassiana produces a plethora of siderophores under iron-limiting conditions. In this study, a siderophore biosynthesis pathway, akin to the general pathway observed in filamentous fungi, was revealed in B. bassiana. Among the siderophore biosynthesis genes (SID), BbSidA was required for the production of most siderophores, and the SidC and SidD biosynthesis gene clusters were indispensable for the production of ferricrocin and fusarinine C, respectively. Biosynthesis genes play various roles in siderophore production, vegetative growth, stress resistance, development, and virulence, in which BbSidA plays the most important role. Accordingly, B. bassiana employs a cocktail of siderophores for iron metabolism, which is essential for fungal physiology and host interactions. This study provides the initial network for the genetic modification of siderophore biosynthesis, which not only aims to improve the efficacy of biocontrol agents but also ensures the efficient production of siderophores.

Susceptibility of Dalbulus maidis to insect-pathogenic fungi: unveiling the protective role of brochosomes and self-cleaning behavior

Susceptibility of Dalbulus maidis to insect-pathogenic fungi: unveiling the protective role of brochosomes and self-cleaning behavior

The Forkhead Box Gene, MaSep1, Negatively Regulates UV- and Thermo-Tolerances and Is Required for Microcycle Conidiation in Metarhizium acridum.

Insect pathogenic fungi have shown great potential in agricultural pest control. Conidiation is crucial for the survival of filamentous fungi, and dispersal occurs through two methods: normal conidiation, where conidia differentiate from mycelium, and microcycle conidiation, which involves conidial budding. The conidiation process is related to cell separation. The forkhead box gene Sep1 in Schizosaccharomyces pombe plays a crucial role in cell separation. Nevertheless, the function of Sep1 has not been clarified in filamentous fungi. Here, MaSep1, the homolog of Sep1 in Metarhizium acridum, was identified and subjected to functional analysis. The findings revealed that conidial germination of the MaSep1-deletion strain (ΔMaSep1) was accelerated and the time for 50% germination rate of conidial was shortened by 1 h, while the conidial production of ΔMaSep1 was considerably reduced. The resistances to heat shock and UV-B irradiation of ΔMaSep1 were enhanced, and the expression of some genes involved in DNA damage repair and heat shock response was significantly increased in ΔMaSep1. The disruption of MaSep1 had no effect on the virulence of M. acridum. Interestingly, ΔMaSep1 conducted the normal conidiation on the microcycle conidiation medium, SYA. Furthermore, 127 DEGs were identified by RNA-Seq between the wild-type and ΔMaSep1 strains during microcycle conidiation, proving that MaSep1 mediated the conidiation pattern shift by governing some genes associated with conidiation, cell division, and cell wall formation.

Glyoxal oxidase-mediated detoxification of reactive carbonyl species contributes to virulence, stress tolerance, and development in a pathogenic fungus.

Reactive carbonyl and oxygen species (RCS/ROS), often generated as metabolic byproducts, particularly under conditions of pathology, can cause direct damage to proteins, lipids, and nucleic acids. Glyoxal oxidases (Gloxs) oxidize aldehydes to carboxylic acids, generating hydrogen peroxide (H2O2). Although best characterized for their roles in lignin degradation, Glox in plant fungal pathogens are known to contribute to virulence, however, the mechanism underlying such effects are unclear. Here, we show that Glox in the insect pathogenic fungus, Metarhizium acridum, is highly expressed in mycelia and during formation of infection structures (appressoria), with the enzyme localizing to the cell membrane. MaGlox targeted gene disruption mutants showed RCS and ROS accumulation, resulting in cell toxicity, induction of apoptosis and increased autophagy, inhibiting normal fungal growth and development. The ability of the MaGlox mutant to scavenge RCS was significantly reduced, and the mutant exhibited increased susceptibility to aldehydes, oxidative and cell wall perturbing agents but not toward osmotic stress, with altered cell wall contents. The ΔMaGlox mutant was impaired in its ability to penetrate the host cuticle and evade host immune defense resulting in attenuated pathogenicity. Overexpression of MaGlox promoted fungal growth and conidial germination, increased tolerance to H2O2, but had little to other phenotypic effects. Transcriptomic analyses revealed downregulation of genes related to cell wall synthesis, conidiation, stress tolerance, and host cuticle penetration in the ΔMaGlox mutant. These findings demonstrate that MaGlox-mediated scavenging of RCS is required for virulence, and contributes to normal fungal growth and development, stress resistance.

Light enhances the production of conidia and influences their hydrophobicity in Tolypocladium inflatum

Insect fungal pathogens such as Beauveria bassiana, Metarhizium robertsii, and Tolypocladium inflatum have been used as insect biocontrol agents. Their infection mechanism involves non-specific adhesion to the host cuticle, which is controlled by hydrophobins, small proteins that form an amphipathic monolayer with rodlet morphology on diverse fungal structures. Light is an abiotic factor that may influence a wide range of cellular processes, including conidiogenesis, stress tolerance, and metabolite biosynthesis. Although its effects have been studied in many fungi, little is known about the effects of light on the hydrophobic properties of conidia. The aim of this work was to investigate the influence of visible light on the conidial hydrophobicity of three entomopathogenic fungal species. For this, conidia of B. bassiana, M. robertsii, and T. inflatum were grown either under light or in the dark, drop profiles of water and diiodomethane on conidial surfaces were analyzed, and conidial hydrophobicity was estimated from contact angle measurements. Moreover, conidial production was determined, and their genome was screened with sequences for hydrophobins. Conidia of B. bassiana and M. robertsii are more hydrophobic than conidia of T. inflatum. The light modified the surface tension of T. inflatum; therefore, conidia of T. inflatum became hydrophilic. However, light did not affect the conidial hydrophobicity of B. bassiana and M. robertsii. In addition, light modified the conidial production of B. bassiana and T. inflatum cultures, but it had no effect on the conidial production of M. robertsii. The T. inflatum genome contains two predicted proteins whose sequence is akin to that of proven class II hydrophobins from other ascomycetes. Presumably, these proteins are responsible for the conidial hydrophobicity properties in this fungus. Our study helps elucidate how light affects the conidial hydrophobicity of entomopathogenic fungi.

Photoreactivation Activities of Rad5, Rad16A and Rad16B Help Beauveria bassiana to Recover from Solar Ultraviolet Damage.

In budding yeast, Rad5 and Rad7-Rad16 play respective roles in the error-free post-replication repair and nucleotide excision repair of ultraviolet-induced DNA damage; however, their homologs have not yet been studied in non-yeast fungi. In the fungus Beauveria bassiana, a deficiency in the Rad7 homolog, Rad5 ortholog and two Rad16 paralogs (Rad16A/B) instituted an ability to help the insect-pathogenic fungus to recover from solar UVB damage through photoreactivation. The fungal lifecycle-related phenotypes were not altered in the absence of rad5, rad16A or rad16B, while severe defects in growth and conidiation were caused by the double deletion of rad16A and rad16B. Compared with the wild-type and complemented strains, the mutants showed differentially reduced activities regarding the resilience of UVB-impaired conidia at 25 °C through a 12-h incubation in a regime of visible light plus dark (L/D 3:9 h or 5:7 h for photoreactivation) or of full darkness (dark reactivation) mimicking a natural nighttime. The estimates of the median lethal UVB dose LD50 from the dark and L/D treatments revealed greater activities of Rad5 and Rad16B than of Rad16A and additive activities of Rad16A and Rad16B in either NER-dependent dark reactivation or photorepair-dependent photoreactivation. However, their dark reactivation activities were limited to recovering low UVB dose-impaired conidia but were unable to recover conidia impaired by sublethal and lethal UVB doses as did their photoreactivation activities at L/D 3:9 or 5:7, unless the night/dark time was doubled or further prolonged. Therefore, the anti-UV effects of Rad5, Rad16A and Rad16B in B. bassiana depend primarily on photoreactivation and are mechanistically distinct from those for their yeast homologs.

Entomophthorales Fungi Parasitizing Sucking Insects in Egypt

Entomophthorales are insect pathogenic fungi significant biological control potentials due to their high insect toxicity. This review focuses on the survey and morphological descriptions of entomophthoralean species attacking insect pests in Egypt. Until now 10 species of Entomophthorales fungi, belonging to three families (Entomophthoraceae, Neozygitaceae and Ancylistaceace) have been reported to suck insects as their hosts. These fungi are widely distributed in various climatic conditions in several Governorates, representing Lower and Upper Egypt. The fungi are the only pathogens that regularly and effectively control sucking insect populations in the natural ecosystems and agroecosystems. The present review emphasizes more studies and isolations of Entomophthorales species by using modern identification techniques so that their epidemiology and control potentials can be predicated on their role against insect pests under variable climatic conditions in Egypt. The possible relationship between population densities of sucking insect pests and Entomophthorales can be further studied to explore their effective applications under variable climatic conditions in the country.

Advances in biosynthesis and metabolic engineering strategies of cordycepin.

Cordyceps militaris, also called as bei-chong-cao, is an insect-pathogenic fungus from the Ascomycota phylum and the Clavicipitaceae family. It is a valuable filamentous fungus with medicinal and edible properties that has been utilized in traditional Chinese medicine (TCM) and as a nutritious food. Cordycepin is the bioactive compound firstly isolated from C. militaris and has a variety of nutraceutical and health-promoting properties, making it widely employed in nutraceutical and pharmaceutical fields. Due to the low composition and paucity of wild resources, its availability from natural sources is limited. With the elucidation of the cordycepin biosynthetic pathway and the advent of synthetic biology, a green cordycepin biosynthesis in Saccharomyces cerevisiae and Metarhizium robertsii has been developed, indicating a potential sustainable production method of cordycepin. Given that, this review primarily focused on the metabolic engineering and heterologous biosynthesis strategies of cordycepin.

Expression of a viral ecdysteroid UDP-glucosyltransferase enhanced the insecticidal activity of the insect pathogenic fungus Beauveria bassiana.

Entomopathogenic fungi, such as Beauveria bassiana, hold promise as biological control agents against insect pests. However, the efficacy of these fungi can be hindered by insect immune responses. One strategy to enhance fungal virulence is to manipulate host immune by targeting key regulatory molecules like 20-hydroxyecdysone (20E). In this study, we engineered B. bassiana strains to constitutively express the enzyme ecdysteroid UDP-glucosyltransferase (EGT), which inactivates 20E, a crucial insect molting hormone. The engineered strain Bb::EGT-1 exhibited robust expression of EGT, leading to a significant reduction in insect 20E levels upon infection. Moreover, infection with Bb::EGT-1 resulted in accelerated larval mortality. Immune responses analysis revealed repression of insect immune response genes and decreased phenoloxidase (PO) activity in larvae infected with Bb::EGT-1. Microbiome analysis indicated alterations in bacterial composition within infected insects, with increased abundance observed during infection with Bb::EGT-1. Additionally, the presence of bacteria hindered hyphal emergence from insect cadavers, suggesting a role for microbial competition in fungal dissemination. Constitutive expression of EGT in B. bassiana enhances fungal virulence by reducing insect 20E levels, suppressing immune responses, and altering the insect microbiome. These findings highlighted the potential of engineered fungi as effective biocontrol agents against insect pests and provide insights into the complex interactions between entomopathogenic fungi, their hosts, and associated microbes. © 2024 Society of Chemical Industry.

Comparative transcriptome analysis reveals disruption of Plutella xylostella immune system by fungal peptide cyclosporin C

The diamondback moth (Plutella xylostella), a major threat to crucifers across the globe, has developed resistance against the majority of insecticides enhancing the need for alternate control measures against this pest. Recently cyclosporin C, a secondary metabolite produced by the insect pathogenic fungus Purpeocillium lilacinum, has been reported to induce lethal and sub-lethal effects against P. xylostella. To date, little is known about the molecular mechanisms of interaction between cyclosporin C and P. xylostella immune systems. This study reports the transcriptome-based immune response of P. xylostella to cyclosprin C treatment. Our results showed differential expression of 322, 97, and 504 differentially expressed genes (DEGS) in P. xylostella treated with cyclosporin C compared to control 24, 48, and 72 h post-treatment, respectively. Thirteen DEGs were commonly expressed at different time intervals in P. xylostella larvae treated with cyclosporin C compared to control. Cyclosporin C treatment induced the down-regulated expression of majority of immune-related genes related to pattern recognition responses, signal modulation, Toll and IMD pathways, antimicrobial peptides and antioxidant responses confirming the ability to suppress immune response of P. xylostella. These results will further improve our knowledge of the infection mechanism and complex biochemical processes involved in interaction between cyclosporin C and insect immune systems.

Genome-Wide Identification of SNARE Family Genes and Functional Characterization of an R-SNARE Gene BbSEC22 in a Fungal Insect Pathogen Beauveria bassiana.

Soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) are central components of the machinery mediating cell membrane fusion and intracellular vesicular trafficking in eukaryotic cells, and have been well-documented to play critical roles in growth, development, and pathogenesis in the filamentous fungal plant pathogens. However, little is known about the contributions of SNAREs to the physiology and biocontrol potential in entomopathogenic filamentous fungi. Here, a genome-wide analysis of SNARE genes was performed taking advantage of the available whole genome sequence of Beauveria bassiana, a classical entomopathogenic fungus. Based on the compared genomic method, 22 genes encoding putative SNAREs were identified from the whole genome of B. bassiana, and were classified into four groups (7 Qa-, 4 Qb-, 6 Qc-, and 5 R-SNAREs) according to the conserved structural features of their encoding proteins. An R-SNARE encoding gene BbSEC22 was further functionally characterized by gene disruption and complementation. The BbSEC22 null mutant showed a fluffy appearance in mycelial growth and an obvious lag in conidial germination. The null mutant also exhibited significantly increased sensitivity to oxidative stress and cell wall perturbing agents and reduced the yield of conidia production by 43.1% compared with the wild-type strain. Moreover, disruption of BbSEC22 caused a significant decrease in conidial virulence to Spodoptera litura larvae. Overall, our results provide an overview of vesicle trafficking in B. bassiana and revealed that BbSec22 was a multifunctional protein associated with mycelial growth, sporulation, conidial germination, stress tolerance, and insecticidal virulence.

A transcription factor-mediated regulatory network controls fungal pathogen colonization of insect body cavities.

Successful host tissue colonization is crucial for fungal pathogens to cause mycosis and complete the infection cycle, in which fungal cells undergo a series of morphological transition-included cellular events to combat with hosts. However, many transcription factors (TFs) and their mediated networks regulating fungal pathogen colonization of host tissue are not well characterized. Here, a TF (BbHCR1)-mediated regulatory network was identified in an insect pathogenic fungus, Beauveria bassiana, that controlled insect hemocoel colonization. BbHCR1 was highly expressed in fungal cells after reaching insect hemocoel and controlled the yeast (in vivo blastospores)-to-hyphal morphological switch, evasion of immune defense response, and fungal virulence. Comparative analysis of RNA sequencing and chromatin immunoprecipitation sequencing identified a core set of BbHCR1 target genes during hemocoel colonization, in which abaA and brlA were targeted to limit the rapid switch from blastospores to hyphae and fungal virulence. Two targets encoding hypothetical proteins, HP1 and HP2, were activated and repressed by BbHCR1, respectively, which acted as a virulence factor and repressor, respectively, suggesting that BbHCR1 activated virulence factors but repressed virulence repressors during the colonization of insect hemocoel. BbHCR1 tuned the expression of two dominant hemocoel colonization-involved metabolite biosynthetic gene clusters, which linked its regulatory role in evasion of immune response. Those functions of BbHCR1 were found to be collaboratively regulated by Fus3- and Hog1-MAP kinases via phosphorylation. These findings have drawn a regulatory network in which Fus3- and Hog1-MAP kinases phosphorylate BbHCR1, which in turn controls the colonization of insect body cavities by regulating fungal morphological transition and virulence-implicated genes.IMPORTANCEFungal pathogens adopt a series of tactics for successful colonization in host tissues, which include morphological transition and the generation of toxic and immunosuppressive molecules. However, many transcription factors (TFs) and their linked pathways that regulate tissue colonization are not well characterized. Here, we identified a TF (BbHCR1)-mediated regulatory network that controls the insect fungal pathogen, Beauveria bassiana, colonization of insect hemocoel. During these processes, BbHCR1 targeted the fungal central development pathway for the control of yeast (blastospores)-to-hyphae morphological transition, activated virulence factors, repressed virulence repressors, and tuned the expression of two dominant hemocoel colonization-involved immunosuppressive and immunostimulatory metabolite biosynthetic gene clusters. The BbHCR1 regulatory function was governed by Fus3- and Hog1-MAP kinases. These findings led to a new regulatory network composed of Fus3- and Hog1-MAP kinases and BbHCR1 that control insect body cavity colonization by regulating fungal morphological transition and virulence-implicated genes.

A network of transcription factors in complex with a regulating cell cycle cyclin orchestrates fungal oxidative stress responses

BackgroundResponse to oxidative stress is universal in almost all organisms and the mitochondrial membrane protein, BbOhmm, negatively affects oxidative stress responses and virulence in the insect fungal pathogen, Beauveria bassiana. Nothing further, however, is known concerning how BbOhmm and this phenomenon is regulated.ResultsThree oxidative stress response regulating Zn2Cys6 transcription factors (BbOsrR1, 2, and 3) were identified and verified via chromatin immunoprecipitation (ChIP)-qPCR analysis as binding to the BbOhmm promoter region, with BbOsrR2 showing the strongest binding. Targeted gene knockout of BbOsrR1 or BbOsrR3 led to decreased BbOhmm expression and consequently increased tolerances to free radical generating compounds (H2O2 and menadione), whereas the ΔBbOsrR2 strain showed increased BbOhmm expression with concomitant decreased tolerances to these compounds. RNA and ChIP sequencing analysis revealed that BbOsrR1 directly regulated a wide range of antioxidation and transcription-associated genes, negatively affecting the expression of the BbClp1 cyclin and BbOsrR2. BbClp1 was shown to localize to the cell nucleus and negatively mediate oxidative stress responses. BbOsrR2 and BbOsrR3 were shown to feed into the Fus3-MAPK pathway in addition to regulating antioxidation and detoxification genes. Binding motifs for the three transcription factors were found to partially overlap in the promoter region of BbOhmm and other target genes. Whereas BbOsrR1 appeared to function independently, co-immunoprecipitation revealed complex formation between BbClp1, BbOsrR2, and BbOsrR3, with BbClp1 partially regulating BbOsrR2 phosphorylation.ConclusionsThese findings reveal a regulatory network mediated by BbOsrR1 and the formation of a BbClp1-BbOsrR2-BbOsrR3 complex that orchestrates fungal oxidative stress responses.

The Bbotf1 Zn(Ⅱ)2Cys6 transcription factor contributes to antioxidant response, fatty acid assimilation, peroxisome proliferation and infection cycles in insect pathogenic fungus Beauveria bassiana

The abilities to withstand oxidation and assimilate fatty acids are critical for successful infection by many pathogenic fungi. Here, we characterized a Zn(II)2Cys6 transcription factor Bbotf1 in the insect pathogenic fungus Beauveria bassiana, which links oxidative response and fatty acid assimilation via regulating peroxisome proliferation. The null mutant ΔBbotf1 showed impaired resistance to oxidants, accompanied by decreased activities of antioxidant enzymes including CATs, PODs and SODs, and down-regulated expression of many antioxidation-associated genes under oxidative stress condition. Meanwhile, Bbotf1 acts as an activator to regulate fatty acid assimilation, lipid and iron homeostasis as well as peroxisome proliferation and localization, and the expressions of some critical genes related to glyoxylate cycle and peroxins were down-regulated in ΔBbotf1 in presence of oleic acid. In addition, ΔBbotf1 was more sensitive to osmotic stressors, CFW, SDS and LDS. Insect bioassays revealed that insignificant changes in virulence were seen between the null mutant and parent strain when conidia produced on CZP plates were used for topical application. However, propagules recovered from cadavers killed by ΔBbotf1 exhibited impaired virulence as compared with counterparts of the parent strain. These data offer a novel insight into fine-tuned aspects of Bbotf1 concerning multi-stress responses, lipid catabolism and infection cycles.

Frequent horizontal chromosome transfer between asexual fungal insect pathogens

Entire chromosomes are typically only transmitted vertically from one generation to the next. The horizontal transfer of such chromosomes has long been considered improbable, yet gained recent support in several pathogenic fungi where it may affect the fitness or host specificity. To date, it is unknown how these transfers occur, how common they are, and whether they can occur between different species. In this study, we show multiple independent instances of horizontal transfers of the same accessory chromosome between two distinct strains of the asexual entomopathogenic fungus Metarhizium robertsii during experimental co-infection of its insect host, the Argentine ant. Notably, only the one chromosome-but no other-was transferred from the donor to the recipient strain. The recipient strain, now harboring the accessory chromosome, exhibited a competitive advantage under certain host conditions. By phylogenetic analysis, we further demonstrate that the same accessory chromosome was horizontally transferred in a natural environment between M. robertsii and another congeneric insect pathogen, Metarhizium guizhouense. Hence, horizontal chromosome transfer is not limited to the observed frequent events within species during experimental infections but also occurs naturally across species. The accessory chromosome that was transferred contains genes that may be involved in its preferential horizontal transfer or support its establishment. These genes encode putative histones and histone-modifying enzymes, as well as putative virulence factors. Our study reveals that both intra- and interspecies horizontal transfer of entire chromosomes is more frequent than previously assumed, likely representing a not uncommon mechanism for gene exchange.

Essential roles of ferric reductase-like proteins in growth, development, stress response, and virulence of the filamentous entomopathogenic fungus Beauveria bassiana

In yeasts, ferric reductase catalyzes reduction of ferric ion to ferrous form, which is essential for the reductive iron assimilation system. However, the physiological roles of ferric reductases remain largely unknown in the filamentous fungi. In this study, genome-wide annotation revealed thirteen ferric reductase-like (Fre) proteins in the filamentous insect pathogenic fungus Beauveria bassiana, and all their functions were genetically characterized. Ferric reductase family proteins exhibit different sub-cellular distributions (e.g., cell periphery and vacuole), which was due to divergent domain architectures. Fre proteins had a synergistic effect on fungal virulence, which was ascribed to their distinct functions in different physiologies. Ten Fre proteins were not involved in reduction of ferric ion in submerged mycelia, but most proteins contributed to blastospore development. Only two Fre proteins significantly contributed to B. bassiana vegetative growth under the chemical-induced iron starvation, but most Fre proteins were involved in resistance to osmotic and oxidative stresses. Notably, a bZIP-type transcription factor HapX bound to the promoter regions of all FRE genes in B. bassiana, and displayed varying roles in the transcription activation of these genes. This study reveals the important role of BbFre family proteins in development, stress response, and insect pathogenicity, as well as their distinctive role in the absorption of ferric iron from the environment.