Bioremediation of mercury-polluted soil and water by the plant symbiotic fungus Metarhizium robertsii.

Fungi are central to every terrestrial and many aquatic ecosystems, but the mechanisms underlying fungal tolerance to mercury, a global pollutant, remain unknown. Here, we show that the plant symbiotic fungus Metarhizium robertsii degrades methylmercury and reduces divalent mercury, decreasing mercury accumulation in plants and greatly increasing their growth in contaminated soils. M. robertsii does this by demethylating methylmercury via a methylmercury demethylase (MMD) and using a mercury ion reductase (MIR) to reduce divalent mercury to volatile elemental mercury. M. robertsii can also remove methylmercury and divalent mercury from fresh and sea water even in the absence of added nutrients. Overexpression of MMD and MIR significantly improved the ability of M. robertsii to bioremediate soil and water contaminated with methylmercury and divalent mercury. MIR homologs, and thereby divalent mercury tolerance, are widespread in fungi. In contrast, MMD homologs were patchily distributed among the few plant associates and soil fungi that were also able to demethylate methylmercury. Phylogenetic analysis suggests that fungi could have acquired methylmercury demethylase genes from bacteria via two independent horizontal gene transfer events. Heterologous expression of MMD in fungi that lack MMD homologs enabled them to demethylate methylmercury. Our work reveals the mechanisms underlying mercury tolerance in fungi, and may provide a cheap and environmentally friendly means of cleaning up mercury pollution.

Utilization of plant-derived sugars and lipids are coupled during colonization of rhizoplane and rhizosphere by the fungus Metarhizium robertsii.

Plant-derived sugars and lipids are key nutritional sources for plant associated fungi. However, the relationship between utilization of host-derived sugars and lipids during development of the symbiotic association remains unknown. Here we show that the fungus Metarhizium robertsii also needs plant-derived lipids to develop symbiotic relationship with plants. The fatty acid binding proteins FABP1 and FABP2 are important for utilization of plant-derived lipids as the deletion of Fabp1 and Fabp2 significantly reduced the ability of M. robertsii to colonize rhizoplane and rhizosphere of maize and Arabidopsis thaliana. Deleting Fabp1 and Fabp2 increased sugar utilization by upregulating six sugar transporters, and this explains why deleting the monosaccharide transporter gene Mst1, which plays an important role in utilization of plant-derived sugars, had no impact on the ability of the double-gene deletion mutant ΔFabp1::ΔFabp2 to colonize plant roots. FABP1 and FABP2 were also found in other plant-associated Metarhizium species, and they were highly expressed in the medium using the tomato root exudate as the sole carbon and nitrogen source, suggesting that they could be also important for these species to develop symbiotic relationship with plants. In conclusion, we discovered that utilization of plant-derived sugars and lipids are coupled during colonization of rhizoplane and rhizosphere by M. robertsii.

A novel cascade allows Metarhizium robertsii to distinguish cuticle and hemocoel microenvironments during infection of insects.

Pathogenic fungi precisely respond to dynamic microenvironments during infection, but the underlying mechanisms are not well understood. The insect pathogenic fungus Metarhizium robertsii is a representative fungus in which to study broad themes of fungal pathogenicity as it resembles some major plant and mammalian pathogenic fungi in its pathogenesis. Here we report on a novel cascade that regulates response of M. robertsii to 2 distinct microenvironments during its pathogenesis. On the insect cuticle, the transcription factor COH2 activates expression of cuticle penetration genes. In the hemocoel, the protein COH1 is expressed due to the reduction in epigenetic repression conferred by the histone deacetylase HDAC1 and the histone 3 acetyltransferase HAT1. COH1 interacts with COH2 to reduce COH2 stability, and this down-regulates cuticle penetration genes and up-regulates genes for hemocoel colonization. Our work significantly advances the insights into fungal pathogenicity in insects.

A recombinant fungal photolyase autonomously enters human cell nuclei to fix UV-induced DNA lesions.

Solar ultraviolet radiations induced DNA damages in human skin cells with cyclobutane pyrimidine dimers (CPD) and (6-4) photoproducts (6-4PPs) as the most frequent lesions. CPDs are repaired much slower than 6-4PPs by the nucleotide excision repair pathway, which are thus the major lesions that interfere with key cellular processes and give rise to gene mutations, possibly resulting in skin cancer. In prokaryotes and multicellular eukaryotes other than placental mammals, CPDs can be rapidly repaired by CPD photolyases in one simple enzymatic reaction using the energy of blue light. In this study, we aim to construct recombinant CPD photolyases that can autonomously enter human cell nuclei to fix UV-induced CPDs. A fly cell penetration peptide and a viral nucleus localization signal peptide were recombined with a fungal CPD photolyase to construct a recombinant protein. This engineered CPD photolyase autonomously crosses cytoplasm and nuclear membrane of human cell nuclei, which then efficiently photo-repairs UV-induced CPD lesions in the genomic DNA. This further protects the cells by increasing SOD activity, and decreasing cellular ROSs, malondialdehyde and apoptosis.

Air Combat Intention Recognition with Incomplete Information Based on Decision Tree and GRU Network.

Battlefield information is generally incomplete, uncertain, or deceptive. To realize enemy intention recognition in an uncertain and incomplete air combat information environment, a novel intention recognition method is proposed. After repairing the missing state data of an enemy fighter, the gated recurrent unit (GRU) network, supplemented by the highest frequency method (HFM), is used to predict the future state of enemy fighter. An intention decision tree is constructed to extract the intention classification rules from the incomplete a priori knowledge, where the decision support degree of attributes is introduced to determine the node-splitting sequence according to the information entropy of partitioning (IEP). Subsequently, the enemy fighter intention is recognized based on the established intention decision tree and the predicted state data. Furthermore, a target maneuver tendency function is proposed to screen out the possible deceptive attack intention. The one-to-one air combat simulation shows that the proposed method has advantages in both accuracy and efficiency of state prediction and intention recognition, and is suitable for enemy fighter intention recognition in small air combat situations.

An inactivating mutation in the vacuolar arginine exporter gene Vae results in culture degeneration in the fungus Metarhizium robertsii.

Culture degeneration usually results in great commercial losses in the economically important filamentous fungi, but the genetic causes of the degeneration remain elusive. In the fungus Metarhizium robertsii, we found that deletion of the vacuolar arginine exporter gene Vae caused culture degeneration. Compared to the WT strain, the mutant showed increased apoptosis, reactive oxygen species (ROS) level and mitochondrial membrane potential collapse, reduced conidial yield and abnormal lipid droplet formation. The extent of the degeneration in the mutant gradually increased over the successive subculturing, which eventually became irreversible; compared to the third subculture of the mutant, the seventh subculture showed a lower conidial yield and pathogenicity to insects, stronger apoptosis, higher ROS level and a smaller number of conidial lipid droplets. Incorporation of the genomic clone of Vae could not restore the WT phenotypes in the seventh subculture, but could in the third one. Loss-of-function in Vae resulted in vacuolar arginine accumulation and reduction in the cytosolic arginine. This downregulated the expression of the regulator CAG9 of G protein signalling pathway, which accounted for most of the phenotypic changes associated with the degeneration of the mutant. We identified a deleterious mutation that causes culture degeneration in a filamentous fugus.

Horizontal gene transfer allowed the emergence of broad host range entomopathogens.

The emergence of new pathogenic fungi has profoundly impacted global biota, but the underlying mechanisms behind host shifts remain largely unknown. The endophytic insect pathogen Metarhizium robertsii evolved from fungi that were plant associates, and entomopathogenicity is a more recently acquired adaptation. Here we report that the broad host-range entomopathogen M. robertsii has 18 genes that are derived via horizontal gene transfer (HGT). The necessity of degrading insect cuticle served as a major selective pressure to retain these genes, as 12 are up-regulated during penetration; 6 were confirmed to have a role in penetration, and their collective actions are indispensable for infection. Two lipid-carrier genes are involved in utilizing epicuticular lipids, and a third (MrNPC2a) facilitates hemocoel colonization. Three proteases degraded the procuticular protein matrix, which facilitated up-regulation of other cuticle-degrading enzymes. The three lipid carriers and one of the proteases are present in all analyzed Metarhizium species and are essential for entomopathogenicity. Acquisition of another protease (MAA_01413) in an ancestor of broad host-range lineages contributed to their host-range expansion, as heterologous expression in the locust specialist Metarhizium acridum enabled it to kill caterpillars. Our work reveals that HGT was a key mechanism in the emergence of entomopathogenicity in Metarhizium from a plant-associated ancestor and in subsequent host-range expansion by some Metarhizium lineages.

Plant volatile organic compound (E)-2-hexenal facilitates Botrytis cinerea infection of fruits by inducing sulfate assimilation.

Investigation into plant-fungal pathogen interactions is one of the most interesting fields in plant sciences. However, the roles of plant volatile organic compounds in the arms race are still largely unknown. Based on precise quantification of plant volatiles, we discovered that the plant volatile organic compound (E)-2-hexenal, at concentrations that were similar to or lower than those in tissues of strawberry and tomato fruits, upregulates sulfate assimilation in spores and hyphae of the phytopathogenic fungus Botrytis cinerea. This upregulation is independent of the types of sulfur sources in the plant and can be achieved in the presence of inorganic sulfate and organic sulfur sources. Using the fungal deletion mutants, we further found that sulfate assimilation is involved in the infection of tomato and strawberry fruits by B. cinerea, and that the severity of the disease is proportional to the sulfate content in the fruits. Both before and during the infection, (E)-2-hexenal induced utilisation of plant sulfate by B. cinerea facilitates its pathogenesis through enhancing its tolerance to oxidative stress. This work provides novel insights into the role of plant volatiles in plant-fungal pathogen interaction and highlights the importance of sulfur levels in the host in the prevention of grey mould disease.

Duplication of a Pks gene cluster and subsequent functional diversification facilitate environmental adaptation in Metarhizium species.

The ecological importance of the duplication and diversification of gene clusters that synthesize secondary metabolites in fungi remains poorly understood. Here, we demonstrated that the duplication and subsequent diversification of a gene cluster produced two polyketide synthase gene clusters in the cosmopolitan fungal genus Metarhizium. Diversification occurred in the promoter regions and the exon-intron structures of the two Pks paralogs (Pks1 and Pks2). These two Pks genes have distinct expression patterns, with Pks1 highly expressed during conidiation and Pks2 highly expressed during infection. Different upstream signaling pathways were found to regulate the two Pks genes. Pks1 is positively regulated by Hog1-MAPK, Slt2-MAPK and Mr-OPY2, while Pks2 is positively regulated by Fus3-MAPK and negatively regulated by Mr-OPY2. Pks1 and Pks2 have been subjected to positive selection and synthesize different secondary metabolites. PKS1 is involved in synthesis of an anthraquinone derivative, and contributes to conidial pigmentation, which plays an important role in fungal tolerance to UV radiation and extreme temperatures. Disruption of the Pks2 gene delayed formation of infectious structures and increased the time taken to kill insects, indicating that Pks2 contributes to pathogenesis. Thus, the duplication of a Pks gene cluster and its subsequent functional diversification has increased the adaptive flexibility of Metarhizium species.

Two-stage inventory management with financing under demand updates.

Considered is a retailer (she) facing non-stationary stochastic demand. Demand can be fully observed and backlogged, consequently the retailer can update the initial demand information using a Bayesian approach. To alleviate the demand risk, the retailer may use a secondary opportunity to replenish through an option contract. In addition, the retailer also has access to an immediate loan if she faces capital constraints and to a risk-free investment if she has surplus funds. The paper presents a recourse approach to solve the two-stage optimization problem and derive the optimal inventory/financing policies. The results show that the option procurement policy has a two-threshold base-stock structure depending on the first procurement, demand update and also the retailer's financial state. The initial procurement can be computed subsequently. A sufficiently large initial demand will induce the retailer to seize the secondary procurement opportunity. Finally, a series of numerical examples demonstrate the resulting policy under various inventory/financial situations. This research incorporates the financial and operational decisions into demand updates, and brings new managerial results and insights.

MrSt12 implicated in the regulation of transcription factor AFTF1 by Fus3-MAPK during cuticle penetration by the entomopathogenic fungus Metarhizium robertsii.

Metarhizium robertsii is a versatile fungus with multifactorial lifestyles, and it is an emerging fungal model for investigating the mechanisms of multiple lifestyle transitions that involve trans-kingdom host jumping. Penetration of the insect cuticle is the necessary step for the transition from saprophytic or symbiotic to pathogenic lifestyle. Previously, we found the transcription factor AFTF1 plays an important role in cuticle penetration, which is precisely regulated by Fus3-MAPK, Slt2-MAPK, and the membrane protein Mr-OPY2. Here, we identified a transcription factor (MrSt12) that directly regulated the transcription of Aftf1 by physically interacting with the cis-acting element (ATGAAACA) in the promoter of Aftf1. The deletion mutant of MrSt12 failed to form the infection structure appressorium and was thus nonpathogenic. We further found that the regulation of Aftf1 by MrSt12 was directly controlled by the Fus3-MAPK. In conclusion, we found a new signaling cascade containing Fus3-MAPK, MrSt12, and AFTF1, which regulates cuticle penetration by M. robertsii.

Stress-induced pyruvate accumulation contributes to cross protection in a fungus.

It is commonly observed that microorganisms subjected to a mild stress develop tolerance not only to higher doses of the same stress but also to other stresses - a phenomenon called cross protection. The mechanisms for cross protection have not been fully revealed. Here, we report that heat shock induced cross protection against UV, oxidative and osmotic/salt stress conditions in the cosmopolitan fungus Metarhizium robertsii. Similarly, oxidative and osmotic/salt stresses also induced cross protection against multiple other stresses. We found that oxidative and osmotic/salt stresses produce an accumulation of pyruvate that scavenges stress-induced reactive oxygen species and promotes fungal growth. Thus, stress-induced pyruvate accumulation contributes to cross protection. RNA-seq and qRT-PCR analyses showed that UV, osmotic/salt and oxidative stress conditions decrease the expression level of pyruvate consumption genes in the trichloroacetic acid cycle and fermentation pathways leading to pyruvate accumulation. Our work presents a novel mechanism for cross protection in microorganisms.

Genome-wide identification of pathogenicity, conidiation and colony sectorization genes in Metarhizium robertsii.

Metarhizium robertsii occupies a wide array of ecological niches and has diverse lifestyle options (saprophyte, insect pathogen and plant symbiont), that renders it an unusually effective model for studying genetic mechanisms for fungal adaptation. Here over 20,000 M. robertsii T-DNA mutants were screened in order to elucidate genetic mechanism by which M. robertsii replicates and persists in diverse niches. About 287 conidiation, colony sectorization or pathogenicity loci, many of which have not been reported in other fungi were identified. By analysing a series of conidial pigmentation mutants, a new fungal pigmentation gene cluster, which contains Mr-Pks1, Mr-EthD and Mlac1 was identified. A conserved conidiation regulatory pathway containing Mr-BrlA, Mr-AbaA and Mr-WetA regulates expression of these pigmentation genes. During conidiation Mr-BlrA up-regulates Mr-AbaA, which in turn controls Mr-WetA. It was found that Hog1-MAPK regulates fungal conidiation by controlling the conidiation regulatory pathway, and that all three pigmentation genes exercise feedback regulation of conidiation. This work provided the foundation for deeper understanding of the genetic processes behind M. robertsii adaptive phenotypes, and advances our insights into conidiation and pigmentation in this fungus.

Metarhizium robertsii produces indole-3-acetic acid, which promotes root growth in Arabidopsis and enhances virulence to insects.

The plant root colonizing insect-pathogenic fungus Metarhizium robertsii has been shown to boost plant growth, but little is known about the responsible mechanisms. Here we show that M. robertsii promotes lateral root growth and root hair development of Arabidopsis seedlings in part through an auxin [indole-3-acetic acid (IAA)]-dependent mechanism. M. robertsii, or its auxin-containing culture filtrate promoted root proliferation, activated IAA-regulated gene expression and rescued the root hair defect of the IAA-deficient rhd6 Arabidopsis mutant. Substrate feeding assays suggest that M. robertsii possesses tryptamine (TAM) and indole-3-acetamide tryptophan (Trp)-dependent auxin biosynthetic pathways. Deletion of Mrtdc impaired M. robertsii IAA production by blocking conversion of Trp to TAM but the reduction was not sufficient to affect plant growth enhancement. We also show that M. robertsii secretes IAA on insect cuticle. ∆Mrtdc produced fewer infection structures and was less virulent to insects than the wild-type, whereas M. robertsii spores harvested from culture media containing IAA were more virulent. Furthermore, exogenous application of IAA increased appressorial formation and virulence. Together, these results suggest that auxins play an important role in the ability of M. robertsii to promote plant growth, and the endogenous pathways for IAA production may also be involved in regulating entomopathogenicity. Auxins were also produced by other Metarhizium species and the endophytic insect pathogen Beauveria bassiana suggesting that interplay between plant- and fungal-derived auxins has important implications for plant-microbe-insect interactions.

Unveiling the mechanism by which microsporidian parasites prevent locust swarm behavior.

Locusts are infamous for their ability to aggregate into gregarious migratory swarms that pose a major threat to food security. Aggregation is elicited by an interplay of visual, tactile, and chemical stimuli, but the aggregation pheromone in feces is particularly important. Infection by the microsporidian parasite Paranosema (Nosema) locustae is known to inhibit aggregation of solitary Locusta migratoria manilensis and to induce gregarious locusts to shift back to solitary behavior. Here we suggest that P. locustae achieves this effect by acidifying the hindgut and modulating the locust immune response, which suppresses the growth of the hindgut bacteria that produce aggregation pheromones. This in turn reduces production of the neurotransmitter serotonin that initiates gregarious behavior. Healthy L. migratoria manilensis exposed to olfactory stimuli from parasite-infected locusts also produced significantly less serotonin, reducing gregarization. P. locustae also suppresses biosynthesis of the neurotransmitter dopamine that maintains gregarization. Our findings reveal the mechanisms by which P. locustae reduces production of aggregation pheromone and blocks the initiation and maintainence of gregarious behavior.

MAPK cascade-mediated regulation of pathogenicity, conidiation and tolerance to abiotic stresses in the entomopathogenic fungus Metarhizium robertsii.

Metarhizium robertsii has been used as a model to study fungal pathogenesis in insects, and its pathogenicity has many parallels with plant and mammal pathogenic fungi. MAPK (Mitogen-activated protein kinase) cascades play pivotal roles in cellular regulation in fungi, but their functions have not been characterized in M. robertsii. In this study, we identified the full complement of MAPK cascade components in M. robertsii and dissected their regulatory roles in pathogenesis, conidiation and stress tolerance. The nine components of the Fus3, Hog1 and Slt2-MAPK cascades are all involved in conidiation. The Fus3- and Hog1-MAPK cascades are necessary for tolerance to hyperosmotic stress, and the Slt2- and Fus3-MAPK cascades both mediate cell wall integrity. The Hog1 and Slt2-MAPK cascades contribute to pathogenicity; the Fus3-MAPK cascade is indispensable for fungal pathogenesis. During its life cycle, M. robertsii experiences multiple microenvironments as it transverses the cuticle into the haemocoel. RNA-seq analysis revealed that MAPK cascades collectively play a major role in regulating the adaptation of M. robertsii to the microenvironmental change from the cuticle to the haemolymph. The three MAPKs each regulate their own distinctive subset of genes during penetration of the cuticle and haemocoel colonization, but they function redundantly to regulate adaptation to microenvironmental change.

Host-to-pathogen gene transfer facilitated infection of insects by a pathogenic fungus.

Metarhizium robertsii is a plant root colonizing fungus that is also an insect pathogen. Its entomopathogenicity is a characteristic that was acquired during evolution from a plant endophyte ancestor. This transition provides a novel perspective on how new functional mechanisms important for host switching and virulence have evolved. From a random T-DNA insertion library, we obtained a pathogenicity defective mutant that resulted from the disruption of a sterol carrier gene (Mr-npc2a). Phylogenetic analysis revealed that Metarhizium acquired Mr-npc2a from an insect by horizontal gene transfer (HGT). Mr-NPC2a binds to cholesterol, an animal sterol, rather than the fungal sterol ergosterol, indicating it retains the specificity of insect NPC2 proteins. Mr-NPC2a is an intracellular protein and is exclusively expressed in the hemolymph of living insects. The disruption of Mr-npc2a reduced the amount of sterol in cell membranes of the yeast-like hyphal bodies that facilitate dispersal in the host body. These were consequently more susceptible to insect immune responses than the wild type. Transgenic expression of Mr-NPC2a increased the virulence of Beauveria bassiana, an endophytic insect-pathogenic fungus that lacks a Mr-NPC2a homolog.

Overexpression of a Metarhizium robertsii HSP25 gene increases thermotolerance and survival in soil.

Temperature extremes are an important adverse factor limiting the effectiveness of microbial pest control agents. They reduce virulence and persistence in the plant root-colonizing insect pathogen Metarhizium robertsii. Small heat shock proteins have been shown to confer thermotolerance in many organisms. In this study, we report on the cloning and characterization of a small heat shock protein gene hsp25 from M. robertsii. hsp25 expression was upregulated when the fungus was grown at extreme temperatures (4, 35, and 42 °C) or in the presence of oxidative or osmotic agents. Expression of hsp25 in Escherichia coli increased bacterial thermotolerance confirming that hsp25 encodes a functional heat shock protein. Overexpressing hsp25 in M. robertsii increased fungal growth under heat stress either in nutrient-rich medium or on locust wings and enhanced the tolerance of heat shock-treated conidia to osmotic stress. In addition, overexpression of hsp25 increased the persistence of M. robertsii in rhizospheric soils in outdoor microcosms, though it did not affect survival in bulk soil, indicating that M. robertsii's survival in soil is dependent on interactions with plant roots.

The plant beneficial effects of Metarhizium species correlate with their association with roots.

Metarhizium species have recently been found to be plant rhizosphere associates as well as insect pathogens. Because of their abundance, rhizospheric Metarhizium could have enormous environmental impact, with co-evolutionary implications. Here, we tested the hypothesis that some Metarhizium spp. are multifactorial plant growth promoters. In two consecutive years, corn seeds were treated with entomopathogenic Metarhizium spp. and field tested at the Beltsville Facility in Maryland. Seed treatments included application of green fluorescent protein (GFP)-tagged strains of Metarhizium brunneum, Metarhizium anisopliae, Metarhizium robertsii, and M. robertsii gene disruption mutants that were either avirulent (Δmcl1), unable to adhere to plant roots (Δmad2), or poorly utilized root exudates (Δmrt). Relative to seeds treated with heat-killed conidia, M. brunneum, M. anisopliae, and M. robertsii significantly increased leaf collar formation (by 15, 14, and 13 %), stalk length (by 16, 10, and 10 %), average ear biomass (by 61, 56, and 36 %), and average stalk and foliage biomass (by 46, 36, and 33 %). Their major impact on corn yield was during early vegetative growth by allowing the plants to establish earlier and thereby potentially outpacing ambient biotic and abiotic stressors. Δmcl1 colonized roots and promoted plant growth to a similar extent as the parent wild type, showing that Metarhizium populations are plant growth promoters irrespective of their role as insect pathogens. In contrast, rhizospheric populations and growth promotion by Δmrt were significantly reduced, and Δmad2 failed to colonize roots or impact plant growth, suggesting that colonization of the root is a prerequisite for most, if not all, of the beneficial effects of Metarhizium.

Genome sequencing and comparative transcriptomics of the model entomopathogenic fungi Metarhizium anisopliae and M. acridum.

Metarhizium spp. are being used as environmentally friendly alternatives to chemical insecticides, as model systems for studying insect-fungus interactions, and as a resource of genes for biotechnology. We present a comparative analysis of the genome sequences of the broad-spectrum insect pathogen Metarhizium anisopliae and the acridid-specific M. acridum. Whole-genome analyses indicate that the genome structures of these two species are highly syntenic and suggest that the genus Metarhizium evolved from plant endophytes or pathogens. Both M. anisopliae and M. acridum have a strikingly larger proportion of genes encoding secreted proteins than other fungi, while ~30% of these have no functionally characterized homologs, suggesting hitherto unsuspected interactions between fungal pathogens and insects. The analysis of transposase genes provided evidence of repeat-induced point mutations occurring in M. acridum but not in M. anisopliae. With the help of pathogen-host interaction gene database, ~16% of Metarhizium genes were identified that are similar to experimentally verified genes involved in pathogenicity in other fungi, particularly plant pathogens. However, relative to M. acridum, M. anisopliae has evolved with many expanded gene families of proteases, chitinases, cytochrome P450s, polyketide synthases, and nonribosomal peptide synthetases for cuticle-degradation, detoxification, and toxin biosynthesis that may facilitate its ability to adapt to heterogeneous environments. Transcriptional analysis of both fungi during early infection processes provided further insights into the genes and pathways involved in infectivity and specificity. Of particular note, M. acridum transcribed distinct G-protein coupled receptors on cuticles from locusts (the natural hosts) and cockroaches, whereas M. anisopliae transcribed the same receptor on both hosts. This study will facilitate the identification of virulence genes and the development of improved biocontrol strains with customized properties.