Formalin-casein enhances water absorbency of calcium alginate beads and activity of encapsulated Metarhizium brunneum and Saccharomyces cerevisiae.

The control of root-feeding wireworms has become more challenging as synthetic soil insecticides have been progressively phased out due to environmental risk concerns. Innovative microbial control alternatives such as the so-called attract-and-kill strategy depend on the rapid and successful development of dried encapsulated microorganisms, which is initiated by rehydration. Casein is a functional additive that is already used in food or pharmaceutical industry due to its water binding capacity. Cross-linked forms such as formalin-casein (FC), exhibit altered network structures. To determine whether FC influences the rehydration of alginate beads in order to increase the efficacy of an attract-and-kill formulation for wireworm pest control, we incorporated either casein or FC in different alginate/starch formulations. We investigated the porous properties of alginate/starch beads and subsequently evaluated the activities of the encapsulated entomopathogenic fungus Metarhizium brunneum and the CO2 producing yeast Saccharomyces cerevisiae. Adding caseins altered the porous structure of beads. FC decreased the bead density from (1.0197 ± 0.0008) g/mL to (1.0144 ± 0.0008) g/mL and the pore diameter by 31%. In contrast to casein, FC enhanced the water absorbency of alginate/starch beads by 40%. Furthermore, incorporating FC quadrupled the spore density on beads containing M. brunneum and S. cerevisiae, and simultaneous venting increased the spore density even by a factor of 18. Moreover, FC increased the total CO2 produced by M. brunneum and S. cerevisiae by 29%. Thus, our findings suggest that rehydration is enhanced by larger capillaries, resulting in an increased water absorption capacity. Our data further suggest that gas exchange is improved by FC. Therefore, our results indicate that FC enhances the fungal activity of both fungi M. brunneum and S. cerevisiae, presumably leading to an enhanced attract-and-kill efficacy for pest control.

Increased and sex-selective avian predation of desert locusts Schistocerca gregaria treated with Metarhizium acridum.

The entomopathogenic fungus Metarhizium acridum in oil-based formulations (Green Muscle® (GM)) is a biopesticide for locust control lacking side-effects on biodiversity, unlike chemical insecticides. Under controlled conditions, GM-treated locusts and grasshoppers attract predators, a complementary advantage in locust control. We assessed avian predation on a population of desert locusts in northern Niger aerially sprayed operationally with GM with 107 g viable conidia ha-1. Populations of adult locusts and birds and vegetation greenness were assessed simultaneously along two transects from 12 days before until 23 days after treatment. Common kestrels Falco tinnunculus and lanners F. biarmicus were the predominant avian predators. Regurgitated pellets and prey remains were collected daily beneath "plucking posts" of kestrels. Locusts started dying five days post-spray and GM had its maximum effect one-two weeks after the spray, with 80% efficacy at day 21. After spraying, bird numbers increased significantly (P<0.05) concurrent with decreasing desert locust densities. Locust numbers decreased significantly (P<0.001) with both time since spraying and decreasing greenness. Before spraying, kestrel food remains under plucking posts accounted for 34.3 ±13.4 prey items day-1, of which 31.0 ±11.9 were adult desert locusts (90.3%), reducing post-spray to 21.1 ±7.3 prey items day-1, of which19.5 ±6.7 were adult desert locusts (92.5%), attributable to decreased use of the plucking-posts by the kestrels rather than an effect of the spray. After spraying, kestrels took significantly (P<0.05) more larger female (75-80%) than smaller male (20-25%) locusts. Avian predation probably enhanced the impact of the GM on the desert locust population, especially by removing large adult females. No direct or indirect adverse side-effects were observed on non-target organisms including locust predators such as ants and birds. These substantial ecological advantages should also be considered when choosing between conventional chemical and biopesticide-based locust control.

Chitin increases drying survival of encapsulated Metarhizium pemphigi blastospores for Ixodes ricinus control.

Ticks, like Ixodes ricinus, have negative impacts on human and animal health in Germany and worldwide, with almost no specific scientifically proven biological control agent commercially available. Biological control agents containing entomopathogenic fungi present many advantages over chemical acaricides but usually high doses of aerial conidia (1013-1014 conidia/ha) are required to control arthropod pests in the field. A suitable formulation containing nutrients not only makes sensitive blastospores applicable but also functions as a microfermenter to multiply the biomass and thus significantly reduce the required application dosage. For this approach, Metarhizium pemphigi X1c blastospores were encapsulated in calcium alginate beads with granular corn starch or chitin powder as nutrients to ensure formation of aerial conidia on the surface and were then dried. The highest concentration was obtained with moist beads containing chitin (4.68 ±â€¯0.71∙107 conidia∙bead-1). The highest drying survival was also obtained with chitin as the additive (14.7 ±â€¯2.18%). Newly formed aerial conidia of all formulations showed high virulence and caused 100% mortality of I. ricinus nymphs. Altogether, this study paves the way for a lower dose and cost-effective application of blastospores for the control of above ground arthropod pests.

Larvicidal toxicity of Metarhizium anisopliae metabolites against three mosquito species and non-targeting organisms.

BACKGROUND: The fungal toxin acts as effective, low-cost chemical substances for pest control worldwide and also an alternative to synthetic insecticides. This study assessed the larvicidal potential of Metarhizium anisopliae fungi derived metabolites against Aedes aegypti, Anopheles stephensi, Culex quinquefasciatus and non-targeted organisms at 24hr post treatment. METHOD: Isolation of entomopathogenic fungi M. anisopliae from natural traps confirmed by using 18s rDNA biotechnological tools. Crude extracts from M. anisopliae solvent extraction and their secondary metabolites were bio-assayed following WHO standard procedures against Ae. aegypti, An. stephensi and Cx. quinquefasciatus, Artemia nauplii, Eudrilus eugeniae, and Solanum lycopersicum after 24 hr exposure. Histopathological analysis of E. eugeniae treated with fungi metabolites toxicity compared to those treated with Monocrotophos after 24hrpost-treatment. M. anisopliae metabolites were characterized using GC-MS and FT-IR analysis. RESULTS: The larvicidal activity was recorded in highest concentration of 75µg/ml, with 85%, 97% and 89% mortality in Ae. aegypti, An. stephensi and Cx. quinquefasciatus respectively. M. anisopliae metabolites produced LC50 values in Ae. aegypti, 59.83µg/ml, in An. stephensi, 50.16µg/ml and in Cx. quinquefasciatus, 51.15µg/ml respectively. M. anisopliae metabolites produced lower toxic effects on A. nauplii, LC50 values were, 54.96µg/ml respectively. Bio-indicator toxicity results show 18% and 58% mortality was recorded in E. eugeniae and A. nauplii and also there is no phytotoxicity that was observed on S. lycopersicum L. under semi-field condition. E. eugeniae histopathological studies shows fungal metabolites showed lower sub-lethal effects compared to synthetic chemical pesticide at 24hrs of the treatment. The GC-MS and FT-IR analysis identified five major components of active ingredients. CONCLUSION: Findings of this study indicate that, M. anisopliae ethyl acetate derived secondary metabolites are effective against larvae of Ae. aegypti, An. stephensi and Cx. quinquefasciatus mosquito species, lower toxicity effects were observed on non-target organisms such as, Artemia nauplii, Eudrilus eugeniae as well as, no toxicity effect were observed on Solanum lycopersicum. Further research should be conducted in laboratory for separation of single pure molecule and be tested semifield conditions.

Polysubstituted Phenyl Glucosides Produced by the Fungus Metarrhizium anisopliae.

Metarhizosides A-G (1-7), seven new polysubstituted phenyl glucosides, were isolated from the extracts of solid rice medium of a marine-derived fungus Metarrhizium anisopliae. Compounds 1-7 all contain a polysubstituted phenyl group and the sugar unit is identified as 4'-O-methyl-ß-D-glucopyranose. Their structures were elucidated by NMR spectroscopy and chemical method. These compounds were evaluated for anti-inflammatory activity by using LPS-stimulated murine macrophage RAW 264.7 cells and the cytotoxicities against four human cancer cell lines.

Metarhizium anisopliae CQMa128 regulates antioxidant/detoxification enzymes and exerts acaricidal activity against Psoroptes ovis var. cuniculi in rabbits: A preliminary study.

The entomopathogenic fungi Metarhizium anisopliae is highly pathogenic toward arthropods. Here, we evaluated the efficacy of a commercial formulation of M. anisopliae against P. ovis var. cuniculi in vivo and in vitro and explored the acaricidal mechanism of M. anisopliae by determining the antioxidant/detoxification-related enzymes activities including glutathione S-transferase (GST), superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT) in mites. The results showed that M. anisopliae had high acaricidal activity against P. ovis var. cuniculi in vitro, in a time- and dose-dependent manner, with 83.33 % mortality at day 9 and a median lethal time (LT50) of 6.10 days after applying 6.14 × 109 conidia/ml of M. anisopliae. In vivo experiments, M. anisopliae achieved 100 % therapeutic effect after 3 days, compared with only 62.21 % for ivermectin. Enzyme assays showed that M. anisopliae significantly upregulated activities of GST, SOD and CAT in Psoroptes mites. The results indicate that M. anisopliae may be an effective biological agent for control of P. ovis var. cuniculi infestations in rabbits and the acaricidal activity may be associated with the changes of enzyme activities of the detoxification and antioxidant system in Psoroptes mites.

Adulticidal Activity of Metarhizium anisopliae s.l. (Hypocreales: Clavicipitaceae) Native Strains From Argentina Against Aedes aegypti (Diptera: Culicidae).

The adulticidal activity of six fungal strains of Metarhizium anisopliae (Metschn.) Sorokin s.l. (Hypocreales: Clavicipitaceae) against Aedes aegypti (L.) (Diptera: Culicidae) was assessed. These strains (CEP 085, CEP 087, CEP 120, CEP 350, CEP 404, and CEP 423) were isolated from soil samples or nondipteran hosts collected from areas in Argentina where Ae. aegypti is distributed. Bioassays were performed with four conidial concentrations plus a control of each fungal strain to determine the lethal concentrations (LC50/LC90), the median survival times (ST50), the mean percentage of the surviving individuals, and the mean percentage of mycotized cadavers. The strains were able to infect and kill adult Ae. aegypti. Significant differences were found among the LC50 values, with CEP 423 as the most virulent strain with the lowest LC50 (2.4 × 106 conidia/ml). At 1 × 107 conidia/ml: no significant differences were found in the Kaplan-Meier survival functions among the strains; the ST50 ranged from 5 (CEP 085) to 8 d (CEP 350); and the mean percentage of the surviving adults was between 13.3 (CEP 085, CEP 350 and CEP 423) and 46.7% (CEP 087). Significant differences were also found among strains in the mean percentage of cadavers with fungal sporulation. Strain CEP 423 produced the highest percentage of mycotized adults (70%). The concentration of the inoculum significantly affected the survival of individuals and the percentage of mycotized cadavers within each strain. Metarhizium anisopliae s.l. CEP 423 was selected as the most promising candidate for further research aiming to develop a mycoinsecticide against Ae. aegypti.

Polyketides and a Dihydroquinolone Alkaloid from a Marine-Derived Strain of the Fungus Metarhizium marquandii.

Three new natural products (1-3), including two butenolide derivatives (1 and 2) and one dihydroquinolone derivative (3), together with nine known natural products were isolated from a marine-derived strain of the fungus Metarhizium marquandii. The structures of the new compounds were unambiguously deduced by spectroscopic means including HRESIMS and 1D/2D NMR spectroscopy, ECD, VCD, OR measurements, and calculations. The absolute configuration of marqualide (1) was determined by a combination of modified Mosher's method with TDDFT-ECD calculations at different levels, which revealed the importance of intramolecular hydrogen bonding in determining the ECD features. The (3R,4R) absolute configuration of aflaquinolone I (3), determined by OR, ECD, and VCD calculations, was found to be opposite of the (3S,4S) absolute configuration of the related aflaquinolones A-G, suggesting that the fungus M. marquandii produces aflaquinolone I with a different configuration (chiral switching). The absolute configuration of the known natural product terrestric acid hydrate (4) was likewise determined for the first time in this study. TDDFT-ECD calculations allowed determination of the absolute configuration of its chirality center remote from the stereogenic unsaturated γ-lactone chromophore. ECD calculations aided by solvent models revealed the importance of intramolecular hydrogen bond networks in stabilizing conformers and determining relationships between ECD transitions and absolute configurations.

Fungal volatile organic compounds show promise as potent molluscicides.

BACKGROUND: Slugs and snails constitute major crop pests. Withdrawal of metaldehyde has prompted a search for more environmentally friendly yet fast acting molluscicides. This study investigated the response of representative molluscs to conidia and volatile organic compounds (VOCs) of the insect pathogenic fungus Metarhizium brunneum Petch. RESULTS: Conidia of M. brunneum had antifeedant/repellent properties with repellency being dependent upon the fungal strain and conidia concentration. Three commonly produced fungal VOCs, 1-octene, 3-octanone and 1-octen-3-ol, were repellent at low doses (1-5 µL) but could kill slugs and snails on contact or fumigation. At the highest dose tested (10 µL), 100% mortality was achieved for Cornu aspersum Muller (garden snail) and Derocerus reticulatum Muller (grey field slug) within 1 h post-treatment with the first deaths being recorded in <11 min. Aqueous formulations (20% v/v) of the most potent VOCs, 3-octanone and 1-octen-3-ol, could be sprayed onto plants to kill or drive the pest of the crop with no phytotoxic effects. CONCLUSION: The sensitivity of terrestrial molluscs to 3-octanone and 1-octen-3-ol and the ephemeral nature of these compounds makes these excellent candidates for development as mollusc repellents or molluscicides. © 2019 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Metarhizium rileyi biopesticide to control Spodoptera frugiperda: Stability and insecticidal activity under glasshouse conditions.

The insect-pathogenic fungus Metarhizium rileyi is highly sensitive to nutritional and environmental conditions which makes it difficult to produce as a stable biopesticide. In this study, a Colombian isolate of this fungus was produced in bulk, and conidia were formulated as an emulsifiable concentrate (EC). The stability of formulated conidia was studied. Conidial viability was maintained at >85 % viability for 12 m under refrigeration and for >three months at 18 °C. The pH values were stable, while contaminant content was significantly reduced. The efficacy of the EC to control Spodoptera frugiperda (Smith) was correlated with the storage time using different mathematical models, and conservative values of six and 12 months at 8 °C and 18 °C respectively, were established. Finally, the EC was evaluated in maize plants under glasshouse conditions. The LC50 and LC90 were estimated to be 1.17 × 104 and 4.03 × 106 conidia/mL respectively and a 57 % reduction in recent damage of plants was achieved. This study demonstrated the potential of M. rileyi formulated as EC to control S. frugiperda in maize. Therefore, it is necessary to continue developing this biopesticide, in order to deliver a new tool to be integrated in pest management programs.

The effect of entomopathogenic fungal culture filtrate on the immune response and haemolymph proteome of the large pine weevil, Hylobius abietis.

The large pine weevil Hylobius abietis L. is a major forestry pest in 15 European countries, where it is a threat to 3.4 million hectares of forest. A cellular and proteomic analysis of the effect of culture filtrate of three entomopathogenic fungi (EPF) species on the immune system of H. abietis was performed. Injection with Metarhizium brunneum or Beauvaria bassiana culture filtrate facilitated a significantly increased yeast cell proliferation in larvae. Larvae co-injected with either Beauvaria caledonica or B. bassiana culture filtrate and Candida albicans showed significantly increased mortality. Together these results suggest that EPF culture filtrate has the potential to modulate the insect immune system allowing a subsequent pathogen to proliferate. Injection with EPF culture filtrate was shown to alter the abundance of protease inhibitors, detoxifing enzymes, antimicrobial peptides and proteins involved in reception/detection and development in H. abietis larvae. Larvae injected with B. caledonica culture filtrate displayed significant alterations in abundance of proteins involved in cellulolytic and other metabolic processes in their haemolymph proteome. Screening EPF for their ability to modulate the insect immune response represents a means of assessing EPF for use as biocontrol agents, particularly if the goal is to use them in combination with other control agents.

X-ray and cryo-EM structures of the mitochondrial calcium uniporter.

Mitochondrial calcium uptake is critical for regulating ATP production, intracellular calcium signalling, and cell death. This uptake is mediated by a highly selective calcium channel called the mitochondrial calcium uniporter (MCU). Here, we determined the structures of the pore-forming MCU proteins from two fungi by X-ray crystallography and single-particle cryo-electron microscopy. The stoichiometry, overall architecture, and individual subunit structure differed markedly from those described in the recent nuclear magnetic resonance structure of Caenorhabditis elegans MCU. We observed a dimer-of-dimer architecture across species and chemical environments, which was corroborated by biochemical experiments. Structural analyses and functional characterization uncovered the roles of key residues in the pore. These results reveal a new ion channel architecture, provide insights into calcium coordination, selectivity and conduction, and establish a structural framework for understanding the mechanism of mitochondrial calcium uniporter function.

iTRAQ-based quantitative proteomic analysis of conidia and mycelium in the filamentous fungus Metarhizium robertsii.

Metarhizium robertsii is widely applied in biological control via conidia application. To clarify the proteomic differences between conidia and mycelia and explore the underlying mechanisms of conidia as a unit responsible for dispersal and environmental stress, we carried out an iTRAQ (isobaric tags for relative and absolute quantitation)-based quantitative proteomic analysis for two developmental stages from M. robertsii. A total of 2052 proteins were detected, and 90 showed differential protein abundance between the conidia and mycelia. These 90 proteins were primarily associated with stress resistance, amino acid and protein metabolism, and energy metabolism. Further bioinformatics analysis showed that these proteins could be mapped to 52 pathways, five of which were significantly enriched after mapping to KEGG pathways. Interestingly, many proteins involved in the significantly enriched pathway of peroxisome, biosynthesis of secondary metabolites and glyoxylate and dicarboxylate metabolism, including catalase, peroxisomal membrane anchor protein, formate dehydrogenase and isocitrate lyase, were identified with higher abundance in conidia. The results deepened our understanding of the conidia proteome in M. robertsii and provide a basis for further exploration for improving the efficiency of the fungi as biocontrol agents.

Biogenic nanosilver synthesized in Metarhizium robertsii waste mycelium extract - As a modulator of Candida albicans morphogenesis, membrane lipidome and biofilm.

Due to low efficacy of classic antimicrobial drugs, finding new active preparations attracts much attention. In this study an innovative, cost-effective and environmentally friendly method was applied to produce silver nanoparticles (AgNPs) using filamentous fungi Metarhizium robertsii biomass waste. It was shown that these NPs possess prominent antifungal effects against C. albicans, C. glabrata and C. parapsilosis reference strains. Further detailed studies were performed on C. albicans ATCC 90028. AgNPs kill curve (CFU method and esterase-mediated reduction of fluorescein diacetate); fractionally inhibitory concentration index (FICI) with fluconazole (FLC); effect on fungal cell membrane permeability (propidium iodide (PI) staining), membrane lipids profile (HPLC-MS), yeast morphotypes and intracellular reactive oxygen species level (H2DCFDA probe) were investigated. Anti-adhesive and anti-biofilm properties of AgNPs (alone and in combination with FLC) were also tested. Biosafety of AgNPs use was assessed in vitro in cytotoxicity tests against L929 fibroblasts, pulmonary epithelial A549 cell line, and red blood cells. Significant reduction in the viability of yeast cells treated with AgNPs was shown within 6 h. The proportion of C. albicans PI-positive cells increased in a dose and time-dependent manner. Changes in the qualitative and quantitative profile of cell membrane lipids, including significant decline in the quantity of most phospholipid species containing C18:2 and an increase in the amount of phospholipids containing C18:1 acyl species were observed after yeast exposure to AgNPs. CLSM images showed an enhancement in ROS intracellular accumulation in C. albicans treated with biogenic nanosilver. C. albicans transformation from yeast to hyphal forms was also reduced. AgNPs decreased adhesion of yeast to abiotic surfaces, as well as acted synergistically with FLC against sessile population. At fungicidal and fungistatic concentrations, they were non-toxic to mammalian cells. Obtained results confirm suitability of our "green synthesis" method to produce AgNPs with therapeutic potential against fungal infections.

Analytical strategy for determination of known and unknown destruxins using hybrid quadrupole-Orbitrap high-resolution mass spectrometry.

An analytical strategy based on a hybrid quadrupole-Orbitrap mass spectrometry was proposed for the simultaneous screening of known destruxins and characterization of potential members of this class of secondary metabolites, in order to evaluate the metabolite production of entomopathogenic fungi used as biocontrol agents. Initially, the fragmentation pathway of the known and commercially available destruxin A was established combining high resolution mass spectrometry (HRMS) and multiple stage MS data in order to obtain the strategy for the characterization of other destruxins for which reference standards were not available. Nineteen known destruxins including A, B, C, D, Ed, F, A1, B1, Ed1, A2, B2, D2, A3, DesmA, DesmB, DesmC, DesmB2, and two chloro-derivatives (Cl and E2 chlorohydrin) were unequivocally identified in Metarhizium brunneum using the proposed strategy. In addition, four unknown destruxins, namely C1, Ed2, G, and G1, were structurally elucidated and characterized for the first time in this fungal strain.

Carbon translocation from a plant to an insect-pathogenic endophytic fungus.

Metarhizium robertsii is a common soil fungus that occupies a specialized ecological niche as an endophyte and an insect pathogen. Previously, we showed that the endophytic capability and insect pathogenicity of Metarhizium are coupled to provide an active method of insect-derived nitrogen transfer to a host plant via fungal mycelia. We speculated that in exchange for this insect-derived nitrogen, the plant would provide photosynthate to the fungus. By using 13CO2, we show the incorporation of 13C into photosynthate and the subsequent translocation of 13C into fungal-specific carbohydrates (trehalose and chitin) in the root/endophyte complex. We determined the amount of 13C present in root-associated fungal biomass over a 21-day period by extracting fungal carbohydrates and analysing their composition using nuclear magnetic resonance (NMR) spectroscopy. These findings are evidence that the host plant is providing photosynthate to the fungus, likely in exchange for insect-derived nitrogen in a tripartite, and symbiotic, interaction.

New natural products isolated from Metarhizium robertsii ARSEF 23 by chemical screening and identification of the gene cluster through engineered biosynthesis in Aspergillus nidulans A1145.

To rapidly identify novel natural products and their associated biosynthetic genes from underutilized and genetically difficult-to-manipulate microbes, we developed a method that uses (1) chemical screening to isolate novel microbial secondary metabolites, (2) bioinformatic analyses to identify a potential biosynthetic gene cluster and (3) heterologous expression of the genes in a convenient host to confirm the identity of the gene cluster and the proposed biosynthetic mechanism. The chemical screen was achieved by searching known natural product databases with data from liquid chromatographic and high-resolution mass spectrometric analyses collected on the extract from a target microbe culture. Using this method, we were able to isolate two new meroterpenes, subglutinols C (1) and D (2), from an entomopathogenic filamentous fungus Metarhizium robertsii ARSEF 23. Bioinformatics analysis of the genome allowed us to identify a gene cluster likely to be responsible for the formation of subglutinols. Heterologous expression of three genes from the gene cluster encoding a polyketide synthase, a prenyltransferase and a geranylgeranyl pyrophosphate synthase in Aspergillus nidulans A1145 afforded an α-pyrone-fused uncyclized diterpene, the expected intermediate of the subglutinol biosynthesis, thereby confirming the gene cluster to be responsible for the subglutinol biosynthesis. These results indicate the usefulness of our methodology in isolating new natural products and identifying their associated biosynthetic gene cluster from microbes that are not amenable to genetic manipulation. Our method should facilitate the natural product discovery efforts by expediting the identification of new secondary metabolites and their associated biosynthetic genes from a wider source of microbes.

Molecular Genetics of Secondary Chemistry in Metarhizium Fungi.

As with many microbes, entomopathogenic fungi from the genus Metarhizium produce a plethora of small molecule metabolites, often referred to as secondary metabolites. Although these intriguing compounds are a conspicuous feature of the biology of the producing fungi, their roles in pathogenicity and other interactions with their hosts and competing microbes are still not well understood. In this review, secondary metabolites that have been isolated from Metarhizium are cataloged along with the history of their discovery and structural elucidation and the salient biological activities attributed to them. Newly available genome sequences revealed an abundance of biosynthetic pathways and a capacity for producing SMs by Metarhizium species that far exceeds the known chemistry. Secondary metabolism genes identified in nine sequenced Metarhizium species are analyzed in detail and classified into distinct families based on orthology, phylogenetic analysis, and conservation of the gene organization around them. This analysis led to the identification of seven hybrid polyketide/nonribosomal peptide synthetases (M-HPNs), two inverted hybrid nonribosomal peptide/polyketide synthetases (M-IHs), 27 nonribosomal peptide synthetases (M-NRPSs), 14 nonribosomal peptide synthetase-like (M-NPL) pathways, 32 polyketide synthases, and 44 terpene biosynthetic genes having a nonuniform distribution and largely following established phylogenetic relationships within the genus Metarhzium. This systematization also identified candidate pathways for known Metarhizium chemistries and predicted the presence of unknown natural products for this genus by drawing connections between these pathways and natural products known to be produced by other fungi.

A natural fungal infection of a sylvatic cockroach with Metarhizium blattodeae sp. nov., a member of the M. flavoviride species complex.

A wild, forest-dwelling cockroach from the subfamily Ectobiidae (order Blattodea) in a nature reserve in Cavalcante, in the state of Goiás, Brazil, was found to be infected by a new, genetically distinct species in the Metarhizium flavoviride species complex that we describe here as Metarhizium blattodeae. The status of this fungus as a new species is supported by both multigenic sequence comparisons and protein profiles generated by MALDI-TOF (matrix-assisted laser desorption/ionization time-of-flight) mass spectrometry. This is one of the first reports of a naturally occurring fungal pathogen affecting any sylvatic (forest-dwelling) cockroach from any part of the world. M. blattodeae caused up to 96 % mortality of Periplaneta americana nymphs (a serious peridomestic cockroach species) after 10 d.

New antibacterial isocoumarin glycosides from a wetland soil derived fungal strain Metarhizium anisopliae.

Eight new isocoumarin glycosides (1-8) were obtained from the solid culture of the wetland soil-derived fungus Metarhizium anisopliae (No. DTH12-10). Their chemical structures were elucidated by analyses of HR ESI-TOF MS, (1)H, (13)C NMR, (1)H-(1)H COSY, HSQC, and HMBC spectra. The absolute configurations were determined by single crystal X-ray diffraction, circular dichroism (CD) spectrum, and chemical derivatization methods. In addition, inhibition of the biofilm formation and the secretion of virulence factor of the new isocoumarin glycosides against Pseudomonas aeruginosa strain PAOA (clinical isolates) were evaluated. The result revealed that compound 1 showed antibacterial activity comparable with (Z)-4-bromo-5-(bromomethylene)-2(5H)-furanone (BF).