Insect fungal pathogens secrete a cell wall-associated glucanase that acts to help avoid recognition by the host immune system.

Fungal insect pathogens have evolved diverse mechanisms to evade host immune recognition and defense responses. However, identification of fungal factors involved in host immune evasion during cuticular penetration and subsequent hemocoel colonization remains limited. Here, we report that the entomopathogenic fungus Beauveria bassiana expresses an endo-ß-1,3-glucanase (BbEng1) that functions in helping cells evade insect immune recognition/ responses. BbEng1 was specifically expressed during infection, in response to host cuticle and hemolymph, and in the presence of osmotic or oxidative stress. BbEng1 was localized to the fungal cell surface/ cell wall, where it acts to remodel the cell wall pathogen associated molecular patterns (PAMPs) that can trigger host defenses, thus facilitating fungal cell evasion of host immune defenses. BbEng1 was secreted where it could bind to fungal cells. Cell wall ß-1,3-glucan levels were unchanged in ΔBbEng1 cells derived from in vitro growth media, but was elevated in hyphal bodies, whereas glucan levels were reduced in most cell types derived from the BbEng1 overexpressing strain (BbEng1OE). The BbEng1OE strain proliferated more rapidly in the host hemocoel and displayed higher virulence as compared to the wild type parent. Overexpression of their respective Eng1 homologs or of BbEng1 in the insect fungal pathogens, Metarhizium robertsii and M. acridum also resulted in increased virulence. Our data support a mechanism by which BbEng1 helps the fungal pathogen to evade host immune surveillance by decreasing cell wall glucan PAMPs, promoting successful fungal mycosis.

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

BACKGROUND: Response 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. RESULTS: Three 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. CONCLUSIONS: These 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(â ¡)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.

A novel transcription factor negatively regulates antioxidant response, cell wall integrity and virulence in the fungal insect pathogen, Beauveria bassiana.

Genomic data have identified a class of fungal specific transcription factors (FsTFs) that are thought to regulate unique aspects of fungal gene expression, although the functions of many of these proteins remain unknown. Here, a novel FsTF (BbStf1), which features a leucine zipper dimerization domain and a fungal transcription factor regulatory middle homology region, was characterized in Beauveria bassiana, a filamentous insect fungal pathogen. Transcriptional activation and nuclear localization were experimentally confirmed for BbStf1. Disruption of Bbstf1 resulted in increased tolerance to oxidative stress and cell wall perturbation, accompanied by increased peroxidase (POD) and superoxide dismutase (SOD) activities and ratio of reduced/oxidized glutathione (GSH/GSSG), and by thickened cell wall and altered composition. Gene expression profile analysis revealed that transcription patterns of antioxidant enzyme and cell wall integrity-involved genes were altered in the ∆Bbstf1, including some BbStf1-targeted genes clarified with evidence. The ∆Bbstf1 strain displayed greater virulence to Galleria mellonella in the bioassays through both topical infection and intrahaemocoel injection due to more rapid proliferation in the haemocoel as compared to the wild-type strain. Altogether, BbStf1 acts as a negative regulator of antioxidant response, cell wall integrity and virulence in B. bassiana.

Multifunctional role of a fungal pathogen-secreted laccase 2 in evasion of insect immune defense.

Laccases are widely present in bacteria, fungi, plants and invertebrates and involved in a variety of physiological functions. Here, we report that Beauveria bassiana, an economic important entomopathogenic fungus, secretes a laccase 2 (BbLac2) during infection that detoxifies insect immune response-generated reactive oxygen species (ROS) and interferes with host immune phenoloxidase (PO) activation. BbLac2 is expressed in fungal cells during proliferation in the insect haemocoel and can be found to distribute on the surface of haemolymph-derived in vivo fungal hyphal bodies or be secreted. Targeted gene-knockout of BbLac2 increased fungal sensitivity to oxidative stress, decreased virulence to insect, and increased host PO activity. Strains overexpressing BbLac2 showed increased virulence, with reduced host PO activity and lowered ROS levels in infected insects. In vitro assays revealed that BbLac2 could eliminate ROS and oxidize PO substrates (phenols), verifying the enzymatic functioning of the protein in detoxification of cytotoxic ROS and interference with the PO cascade. Moreover, BbLac2 acted as a cell surface protein that masked pathogen associated molecular patterns (PAMPs), enabling the pathogen to evade immune recognition. Our data suggest a multifunctional role for fungal pathogen-secreted laccase 2 in evasion of insect immune defenses.

Epidemiology and Outcomes of Acute Kidney Injury in COVID-19 Patients with Acute Respiratory Distress Syndrome: A Multicenter Retrospective Study.

BACKGROUND: Acute kidney injury (AKI) is associated with increased mortality in patients with acute respiratory distress syndrome (ARDS). However, the epidemiological features and outcomes of AKI among COVID-19 patients with ARDS are unknown. METHODS: We retrospectively recruited consecutive adult COVID-19 patients who were diagnosed with ARDS according to Berlin definition from 13 designated intensive care units in the city of Wuhan, China. Potential risk factors of AKI as well as the relation between AKI and in-hospital mortality were investigated. RESULTS: A total of 275 COVID-19 patients with ARDS were included in the study, and 49.5% of them developed AKI during their hospital stay. In comparison with patients without AKI, patients who developed AKI were older, tended to have chronic kidney disease, had higher Sepsis-Related Organ Failure Assessment score on day 1, and were more likely to receive invasive ventilation and develop acute organ dysfunction. Multivariate analysis showed that age, history of chronic kidney disease, neutrophil-to-lymphocyte ratio, and albumin level were independently associated with the occurrence of AKI. Importantly, increasing AKI severity was associated with increased in-hospital mortality when adjusted for other potential variables: odds ratio of stage 1 = 5.374 (95% CI: 2.147-13.452; p < 0.001), stage 2 = 6.216 (95% CI: 2.011-19.210; p = 0.002), and stage 3 = 34.033 (95% CI: 9.723-119.129; p < 0.001). CONCLUSION: In this multicenter retrospective study, we found that nearly half of COVID-19 patients with ARDS experienced AKI during their hospital stay. The coexistence of AKI significantly increased the mortality of these patients.

The fungal mitochondrial membrane protein, BbOhmm, antagonistically controls hypoxia tolerance.

Adaptation to low-oxygen (LO) environment in host tissues is crucial for microbial pathogens, particularly fungi, to successfully infect target hosts. However, the underlying mechanisms responsible for hypoxia tolerance in most pathogens are poorly understood. A mitochondrial protein, BbOhmm, is demonstrated to limit oxidative stress resistance and virulence in the insect fungal pathogen, Beauveria bassiana. Here, we found that BbOhmm negatively affected hypoxic adaptation in the insect haemocoel while regulating respiration-related events, heme synthesis and mitochondrial iron homeostasis. A homologue of the mammalian sterol regulatory element-binding proteins (SREBPs), BbSre1, was shown to be involved in BbOhmm-mediated LO adaptation. Inactivation of BbSre1 resulted in a significant increase in sensitivity to hypoxic and oxidative stress. Similar to ΔBbOhmm, ΔBbSre1 or the ΔBbOhmmΔBbSre1 double mutant accumulated high levels of heme and mitochondrial iron, regulating the similar pathways during hypoxic stress. BbSre1 transcriptional activity and nuclear import were repressed in ΔBbOhmm cells and affected by intracellular reactive oxygen species (ROS) and oxygen levels. These findings have led to a new model in which BbOhmm affects ROS homeostasis in combination with available oxygen to control the transcriptional activity of BbSre1, which in turn mediates LO adaptation by regulating mitochondrial iron homeostasis, heme synthesis and respiration-implicated genes.

Upside-down InAs/InAs1-xSbx type-II superlattice-based nBn mid-infrared photodetectors with an AlGaAsSb quaternary alloy barrier.

Ga-free InAs/InAsSb type-II superlattices (T2SLs) are emerging as candidate materials for high temperature operation of mid-infrared photodetectors, which are critical for infrared technology with an aim to provide low-cost and compact detection systems. In this work, by utilizing upside-down device structure, a closely lattice-matched Al0.83Ga0.17AsSb quaternary alloy as electron barrier was pre-grown before the growth of InAs/InAsSb T2SLs absorber in a nBn device. Based on this design, we have demonstrated 5-µm cut-off mid-wavelength infrared (MWIR) photodetectors that exhibited a dark current density of 1.55 × 10-4 A/cm2 at an operation bias 400mV at 150K. A saturated quantum efficiency at ∼4.0 µm reaches 37.5% with a 2 µm absorber and the peak responsivity reaches 1.2 A/W, which yields a peak specific detectivity as high as ∼1.82 × 1011 cm·H z1/2/W at a forward bias of 400mV.

A young male with chronic nonproductive cough diagnosed with blastomycosis in China: a case report.

BACKGROUND: Blastomycosis is a fungal infectious disease prevalent in North America and rarely reported in Asia. Misdiagnosis of malignancy and other infectious diseases were reported. CASE PRESENTATION: A 24-years-old male patient presented with chronic non-productive cough of 4 months duration. He had been diagnosed with Mycobacterium tuberculosis infection and lung malignancy elsewhere and presented to us as the symptoms persisted. We offered him the biopsy under endobronchial ultrasound-guide sheath-transbronchial lung biopsy and sample specimen were sent for next generation sequencing analysis, returned as Blastomyces Dermatitidis infection. The patient was treated by itraconazole for 6 months, his symptoms decreased significantly and the CT scan showed resolution of the lesion. CONCLUSION: We shared a case of blastomycosis with delayed and difficult diagnosis and reviewed the knowledge regarding differential diagnosis and next generation sequencing technologies.

Participation of a MADS-box transcription factor, Mb1, in regulation of the biocontrol potential in an insect fungal pathogen.

The cell wall is crucial for fungal growth, proliferation and interaction with the environment and host. Understanding the regulatory mechanism of cell wall integrity may help with improvement of fungal biocontrol agents. Here, a putative target of the cell wall integrity pathway-involved Slt2 MAP kinase, Mb1, an orthologue of MADS-box transcription factor Rlm1, was characterized in an economically important insect fungal pathogen, Beauveria bassiana. Mb1 disruption mutant (ΔMb1) displayed reduced growth and increased conidial production on minimal medium but not on rich-nutrient media, which is different from ΔSlt2 to a great extent. Loss of Mb1 resulted in a significant increase in sensitivity to cell wall-perturbing agents (Congo red and calcofluor white), with alteration in cell composition that was inconsistent with ΔSlt2 strain, including increased chitin content and reduced chitin-binding ß-1, 3/1,6-glucan levels in the absence of any stress. Transcription levels of 15 chitin synthesis and metabolism-associated and 17 Pkc1-Slt2 CWI (cell wall integrity) pathway, glucan synthesis, and cell wall remodeling enzyme synthesis-involved genes were significantly increased and repressed in ΔMb1 strain, respectively, some of which were verified to be the targets of Mb1. Insect bioassays revealed decreased virulence for the ΔMb1 strain in both topical and intrahemocoel injection assays. Our results demonstrated that Mb1 control fungal biocontrol potential-associated traits, including growth, conidiation and cell wall integrity, in B. bassiana. The difference of Mb1 and Slt2 in contribution to cell wall integrity is discussed.

MADS-box transcription factor Mcm1 controls cell cycle, fungal development, cell integrity and virulence in the filamentous insect pathogenic fungus Beauveria bassiana.

MADS-box transcription factor Mcm1 plays crucial roles in regulating mating processes and pathogenesis in some fungi. However, its roles are varied in fungal species, and its function remains unclear in entomopathogenic fungi. Here, Mcm1 orthologue, Bbmcm1, was characterized in a filamentous entomopathogenic fungus, Beauveria bassiana. Disruption of Bbmcm1 resulted in a distinct reduction in growth with abnormal conidiogenesis, and a significant decrease in conidial viability with abnormal germination. ΔBbmcm1 displayed impaired cell integrity, with distorted cell wall structure and altered cell wall component. Abnormal cell cycle was detected in ΔBbmcm1 with longer G2 /M phase but shorter G1 /G0 and S phases in unicellular blastospores, and sparser septa in multicellular hyphae, which might be responsible for defects in development and differentiation due to the regulation of cell cycle-involved genes, as well as the corresponding cellular events-associated genes. Dramatically decreased virulence was examined in ΔBbmcm1, with impaired ability to escape haemocyte encapsulation, which was consistent with markedly reduced cuticle-degrading enzyme production by repressing their coding genes, and downregulated fungal effector protein-coding genes, suggesting a novel role of Mcm1 in interaction with host insect. These data indicate that Mcm1 is a crucial regulator of development, cell integrity, cell cycle and virulence in insect fungal pathogens.

The Beauveria bassiana Gas3 ß-Glucanosyltransferase Contributes to Fungal Adaptation to Extreme Alkaline Conditions.

Fungal ß-1,3-glucanosyltransferases are cell wall-remodeling enzymes implicated in stress response, cell wall integrity, and virulence, with most fungal genomes containing multiple members. The insect-pathogenic fungus Beauveria bassiana displays robust growth over a wide pH range (pH 4 to 10). A random insertion mutant library screening for increased sensitivity to alkaline (pH 10) growth conditions resulted in the identification and mapping of a mutant to a ß-1,3-glucanosyltransferase gene (Bbgas3). Bbgas3 expression was pH dependent and regulated by the PacC transcription factor, which activates genes in response to neutral/alkaline growth conditions. Targeted gene knockout of Bbgas3 resulted in reduced growth under alkaline conditions, with only minor effects of increased sensitivity to cell wall stress (Congo red and calcofluor white) and no significant effects on fungal sensitivity to oxidative or osmotic stress. The cell walls of ΔBbgas3 aerial conidia were thinner than those of the wild-type and complemented strains in response to alkaline conditions, and ß-1,3-glucan antibody and lectin staining revealed alterations in cell surface carbohydrate epitopes. The ΔBbgas3 mutant displayed alterations in cell wall chitin and carbohydrate content in response to alkaline pH. Insect bioassays revealed impaired virulence for the ΔBbgas3 mutant depending upon the pH of the media on which the conidia were grown and harvested. Unexpectedly, a decreased median lethal time to kill (LT50, i.e., increased virulence) was seen for the mutant using intrahemocoel injection assays using conidia grown at acidic pH (5.6). These data show that BbGas3 acts as a pH-responsive cell wall-remodeling enzyme involved in resistance to extreme pH (>9).IMPORTANCE Little is known about adaptations required for growth at high (>9) pH. Here, we show that a specific fungal membrane-remodeling ß-1,3-glucanosyltransferase gene (Bbgas3) regulated by the pH-responsive PacC transcription factor forms a critical aspect of the ability of the insect-pathogenic fungus Beauveria bassiana to grow at extreme pH. The loss of Bbgas3 resulted in a unique decreased ability to grow at high pH, with little to no effects seen with respect to other stress conditions, i.e., cell wall integrity and osmotic and oxidative stress. However, pH-dependent alternations in cell wall properties and virulence were noted for the ΔBbgas3 mutant. These data provide a mechanistic insight into the importance of the specific cell wall structure required to stabilize the cell at high pH and link it to the PacC/Pal/Rim pH-sensing and regulatory system.

The PacC transcription factor regulates secondary metabolite production and stress response, but has only minor effects on virulence in the insect pathogenic fungus Beauveria bassiana.

The PacC transcription factor is an important component of the fungal ambient pH-responsive regulatory system. Loss of pacC in the insect pathogenic fungus Beauveria bassiana resulted in an alkaline pH-dependent decrease in growth and pH-dependent increased susceptibility to osmotic (salt, sorbitol) stress and SDS. Extreme susceptibility to Congo Red was noted irrespective of pH, and ΔBbpacC conidia showed subtle increases in UV susceptibility. The ΔBbPacC mutant showed a reduced ability to acidify media during growth due to failure to produce oxalic acid. The ΔBbPacC mutant also did not produce the insecticidal compound dipicolinic acid, however, production of a yellow-colored compound was noted. The compound, named bassianolone B, was purified and its structure determined. Despite defects in growth, stress resistance, and oxalate/insecticidal compound production, only a small decrease in virulence was seen for the ΔBbpacC strain in topical insect bioassays using larvae from the greater waxmoth, Galleria mellonella or adults of the beetle, Tenebrio molitor. However, slightly more pronounced decreases were seen in virulence via intrahemcoel injection assays (G. mellonella) and in assays using T. molitor larvae. These data suggest important roles for BbpacC in mediating growth at alkaline pH, regulating secondary metabolite production, and in targeting specific insect stages.

Correlation of cell surface proteins of distinct Beauveria bassiana cell types and adaption to varied environment and interaction with the host insect.

The insect fungal pathogen Beauveria bassiana produces a number of distinct cell types that include aerial conidia, blastospores and haemolymph-derived cells, termed hyphal bodies, to adapt varied environment niches and within the host insect. These cells display distinct biochemical properties and surface structures, and a highly ordered outermost brush-like structure uniquely present on hyphal bodies, but not on any in vitro cells. Here, we found that the outermost structure on the hyphal bodies mainly consisted of proteins associated to structural wall components in that most of it could be removed by dithiothreitol (DTT) or proteinase K. DTT-treatment also caused delayed germination, decreased tolerance to ultraviolet irradiation and virulence of conidia or blastospores, with decreased adherence and alternated carbohydrate epitopes, suggesting involvement in fungal development, stress responses and virulence. To characterize these cell surface molecules, proteins were released from the living cells using DTT, and identified and quantitated using label-free quantitative mass spectrometry. Thereafter, a series of bioinformatics programs were used to predict cell surface-associated proteins (CSAPs), and 96, 166 and 54 CSAPs were predicted from the identified protein pools of conidia, blastospores and hyphal bodies, respectively, which were involved in utilization of carbohydrate, nitrogen, and lipid, detoxification, pathogen-host interaction, and likely other cellular processes. Thirteen, sixty-nine and six CSAPs were exclusive in conidia, blastospores and hyphal bodies, respectively, which were verified by eGFP-tagged proteins at their N-terminus. Our data provide a crucial cue to understand mechanism of B. bassiana to adapt to varied environment and interaction with insect host.

Dissection of the contributions of cyclophilin genes to development and virulence in a fungal insect pathogen.

Cyclophilins are ubiquitous proteins found in all domains of life, catalyzing peptidyl-prolyl cis-trans isomerization (PPIase activity) and functioning in diverse cellular processes. The filamentous insect pathogenic fungus, Beauveria bassiana, contains 11 cyclophilin genes whose roles were probed via individual gene knockouts, construction of over-expression strains, and a simultaneous gene knockdown strategy using tandem SiRNA. Mutants were examined for effects on conidiation, hyphal growth, cyclosporine and stress resistance, and insect virulence. BbCypA was found to be the most highly expressed cyclophilin during growth and purified recombinant BbCypA displayed cyclosporine sensitive PPIase activity. Except for ΔBbCypA, targeted gene knockouts or overexpression of any cyclophilin resulted in temperature sensitivity (TS). Specific cyclophilin mutants showed impaired hyphal growth and differential effects on conidiation and cyclosporine resistance. Insect bioassays revealed decreased virulence for two cyclophilins (ΔBbCypE and ΔBbCyp6) and the simultaneous gene knockdown mutant constructs (SiRNA30). The BbSiRNA30 strains were unaffected in growth, conidiation, or under osmotic or cell wall perturbing stress, but did show increased resistance to cyclosporine and a TS phenotype. These results revealed common and unique roles for cyclophilins in B. bassiana and validate a method for examining the effects of multi-gene families via simultaneous gene knockdown.

Bbmsn2 acts as a pH-dependent negative regulator of secondary metabolite production in the entomopathogenic fungus Beauveria bassiana.

Fungal secondary metabolites are chemical compounds important for development, environmental adaptation and for potential biotechnological and pharmaceutical applications. Oosporein, a red-pigmented benzoquinone, produced by many fungal insect pathogenic Beauveria spp., shows remarkable functional diversity, displaying antimicrobial, antiviral and even anti-proliferative activities. A homologue of the msn2/seb1 transcription factor was identified in a Beauveria bassiana random T-DNA insertion library. Targeted gene-knockout of Bbmsn2 resulted in reduced growth and increased sensitivity to Calcofluor White, H2 O2 and Congo Red. However, when normalized to growth at 26°C, the ΔBbmsn2 mutant was more tolerant to high temperature (32°C) than the wild type parent. The ΔBbmsn2 mutant also displayed a pH-dependent growth phenotype, with little growth seen at pH < 5.0 but, better growth at alkaline conditions (pH > 8.0). Unexpectedly, a pH-dependent deregulation of a red pigment, identified as oosporein, was seen in the ΔBbmsn2 mutant. The ΔBbmsn2 strain was impaired in virulence in both topical and intrahaemocoel injection bioassays against Galleria mellonella. ΔBbmsn2 proliferation in the host haemolymph and conidiation on the host cadaver was reduced. These data indicate that Bbmsn2 acts as a negative regulator of oosporein production and contributes to virulence and growth in response to external pH in B. bassiana.

Improving mycoinsecticides for insect biological control.

The desire for decreased reliance on chemical pesticides continues to fuel interest in alternative means for pest control including the use of naturally occurring microbial insect pathogens. Insects, as vectors of disease causing agents or as agricultural pests, are responsible for millions of deaths and significant economic losses worldwide, placing stresses on productivity (GDP) and human health and welfare. In addition, alterations in climate change are likely to affect insect ranges, expanding their access to previously constrained geographic areas, a potentially worrisome outcome. Metarhizium anisopliae and Beauveria bassiana, two cosmopolitan fungal pathogens of insects found in almost all ecosystems, are the most commonly applied mycoinsecticides for a variety of insect control purposes. The availability of the complete genomes for both organisms coupled to robust technologies for their transformation has led to several advances in engineering these fungi for greater efficacy and/or utility in pest control applications. Here, we will provide an overview of the fungal-insect and fungal-plant interactions that occur and highlight recent advances in the genetic engineering of these fungi. The latter work has resulted in the development of strains displaying (1) increased resistance to abiotic stress, (2) increased cuticular targeting and degradation, (3) increased virulence via expression of insecticidal protein/peptide toxins, (4) the ability to block transmission of disease causing agents, and (5) the ability to target specific insect hosts, decrease host fecundity, and/or alter insect behaviors.

Ablation of the creA regulator results in amino acid toxicity, temperature sensitivity, pleiotropic effects on cellular development and loss of virulence in the filamentous fungus Beauveria bassiana.

For most organisms, carbon and nitrogen uptake are essential for growth, development and, where applicable, pathogenesis. The role of the carbon catabolite repressor transcription factor homologue BbcreA in the entomopathogenic fungus Beauveria bassiana was investigated. Deletion of BbcreA resulted in pleiotropic effects, including nutrient toxicity, leading to a novel cell lytic phenotype. Fungal growth in rich media and minimal media containing select amino acids/peptides was severely compromised, with microscopic examination revealing conidial-base germ tube degeneration and cell lysis occurring during growth, a phenomenon exacerbated at higher temperatures (32°C). Depending upon nutrient conditions, growth, pigment and aerial mycelium production, sporulation and dimorphic transition to blastospore production were also impaired in the ΔBbcreA strain. Although loss of BbcreA resulted in de-repression of secreted protease and lipase, enzymes critical in mediating pathogenesis, insect bioassays indicated severe defects in virulence using both topical and intra-haemocoel injection assays, with eruption and subsequent sporulation on host cadavers greatly reduced in the mutant. These data suggest that BbcreA functions as more than a carbon repressor and plays important roles in nutrient utilization, cell homeostasis and virulence. In particular, BbcreA is required for proper assimilation of select amino acids and peptides, including asparagine, arginine and proline.