A putative methyltransferase, mtrA, contributes to development, spore viability, protein secretion and virulence in the entomopathogenic fungus Beauveria bassiana

The filamentous fungus, Beauveria bassiana, is a ubiquitously distributed insect pathogen, currently used as an alternative to chemical pesticides for pest control. Conidiospores are the means by which the fungus disseminates in the environment, and these cells also represent the infectious agent most commonly used in field applications. Little, however, is known concerning the molecular basis for maintenance of spore viability, a critical feature for survival and persistence. Here, we report on the role of a putative methyltransferase, BbmtrA, in conidial viability, normal fungal growth and development, and virulence, via characterization of a targeted gene knockout strain. Loss of BbmtrA resulted in pleiotropic effects including reduced germination, growth and conidiation, with growing mycelia displaying greater branching than the WT parent. Conidial viability dramatically decreased over time, with <5 % of the cells remaining viable after 30 days as compared with >80 % of the WT. Reduced production of extracellular proteins was also observed for the ΔBbmtrA mutant, including protease/peptidases, glycoside hydrolases and the hyd1 hydrophobin. The latter was further confirmed by hyd1 gene expression analysis. Insect bioassays using the greater wax moth, Galleria mellonella, further revealed that the ΔBbmtrA strain was attenuated in virulence and failed to sporulate on host cadavers. These data support a global role for mtrA in fungal physiological processes.

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Different strategies to kill the host presented byMetarhizium anisopliaeandBeauveria bassiana

Canadian Journal of Microbiology ◽

10.1139/cjm-2017-0517 ◽

2018 ◽

Vol 64 (3) ◽

pp. 191-200 ◽

Cited By ~ 2

Author(s):

Cynthia Barbosa Rustiguel ◽

María Fernández-Bravo ◽

Luis Henrique Souza Guimarães ◽

Enrique Quesada-Moraga

Keyword(s):

Secondary Metabolites ◽

Beauveria Bassiana ◽

Entomopathogenic Fungi ◽

Metarhizium Anisopliae ◽

Galleria Mellonella ◽

Insect Pests ◽

Fungal Growth ◽

Growth Strategy

Studies conducted over the last decades have shown the potential of entomopathogenic fungi for the biocontrol of some insect pests. Entomopathogenic fungi infect their host through the cuticle, so they do not need to be ingested to be effective. These fungi also secrete secondary metabolites and proteins that are toxic to insect pests. In this context, we analyzed the pathogenicity of Metarhizium anisopliae (Metschn.) strains IBCB 384 and IBCB 425 and Beauveria bassiana (Bals.-Criv.) Vuill. strains E 1764 and E 3158 against Galleria mellonella (Linn.) larvae, during pre-invasion and post-invasion phases. The results showed M. anisopliae, especially strain IBCB 384, was most virulent in the pre-invasion phase against G. mellonella, whereas B. bassiana, especially strain E 1764, was most virulent in the post-invasion phase. During in vivo development and in the production of toxic serum, B. bassiana E 3158 was the most virulent. Different fungal growth (or toxin) strategies were observed for studied strains. Metarhizium anisopliae IBCB 425 prioritizes the growth strategy, whereas strain IBCB 384 and B. bassiana strains E 1764 and E 3158 have a toxic strategy. All strains have pathogenicity against G. mellonella, indicating their possible use for biocontrol.

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Distinct Roles of Myosins in Aspergillus fumigatus Hyphal Growth and Pathogenesis

Infection and Immunity ◽

10.1128/iai.01190-15 ◽

2016 ◽

Vol 84 (5) ◽

pp. 1556-1564 ◽

Cited By ~ 19

Author(s):

Hilary Renshaw ◽

José M. Vargas-Muñiz ◽

Amber D. Richards ◽

Yohannes G. Asfaw ◽

Praveen R. Juvvadi ◽

...

Keyword(s):

Aspergillus Fumigatus ◽

Galleria Mellonella ◽

Fungal Growth ◽

Hyphal Growth ◽

Lavage Fluid ◽

Class Ii ◽

Necrosis Factor Alpha ◽

Class V ◽

Content Type ◽

Conidial Viability

Myosins are a family of actin-based motor proteins found in many organisms and are categorized into classes based on their structures. Class II and V myosins are known to be important for critical cellular processes, including cytokinesis, endocytosis, exocytosis, and organelle trafficking, in the model fungiSaccharomyces cerevisiaeandAspergillus nidulans. However, the roles of myosins in the growth and virulence of the pathogenAspergillus fumigatusare unknown. We constructed single- and double-deletion strains of the class II and class V myosins inA. fumigatusand found that while the class II myosin (myoB) is dispensable for growth, the class V myosin (myoE) is required for proper hyphal extension; deletion ofmyoEresulted in hyperbranching and loss of hyphal polarity. BothmyoBandmyoEare necessary for proper septation, conidiation, and conidial germination, but onlymyoBis required for conidial viability. Infection with the ΔmyoEstrain in the invertebrateGalleria mellonellamodel and also in a persistently immunosuppressed murine model of invasive aspergillosis resulted in hypovirulence, while analysis of bronchoalveolar lavage fluid revealed that tumor necrosis factor alpha (TNF-α) release and cellular infiltration were similar compared to those of the wild-type strain. The ΔmyoEstrain showed fungal growth in the murine lung, while the ΔmyoBstrain exhibited little fungal burden, most likely due to the reduced conidial viability. These results show, for the first time, the important role these cytoskeletal components play in the growth of and disease caused by a known pathogen, prompting future studies to understand their regulation and potential targeting for novel antifungal therapies.

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A response of entomopathogenic fungi to botanical fungicides and synthetic fungicides in vitro

Acta Agrobotanica ◽

10.5586/aa.2005.016 ◽

2012 ◽

Vol 58 (1) ◽

pp. 101-111

Author(s):

Anna Sapieha-Waszkiewicz ◽

Barbara Marjańska-Cichoń ◽

Ryszard Miętkiewski ◽

Mieczysław Żurek

Keyword(s):

Beauveria Bassiana ◽

Entomopathogenic Fungi ◽

Metarhizium Anisopliae ◽

Galleria Mellonella ◽

Morphological Changes ◽

Fungal Growth ◽

Apple Orchard ◽

Paecilomyces Fumosoroseus ◽

Arable Field

The growth colonies' morphology and sporulation of entomopathogenic fungi was estimated on Sabouraud medium containing botanical fungicides (Bioczos liquid and Biosept 33 SL), and synthetic ones (Teldor 500 SC and Sumilex 500 SC). Entomopathogenic fungi (Beauveria bassiana (Balls.) Vuill., Metarhizium anisopliae (Metsch.) Sorok, and Paecilomyces fumosoroseus (Wize) Brown et Smith.), were isolated from soil by means of Galleria mellonella larvae as baits. Isolates Bb I, Ma I, Pf I derivered from herbicide fallow from apple orchard and isolates Bb II, Ma II and Pf II from arable field adjacent to orchard. Fungicides were added to Sabouraud medium at the following concentrations: A-recommended dose, B-10-times lower than the recommended, C-100-times lower than the recommended. The growth of colonies their morphological changes and sporulations were estimated after 5 and 20 days. The same parametrs were observed when fungi were transfered from the medium containing fungicides on the medium with out fungicides. From botanical fungicides Bioczos liquid inhibited fungal growth more than Biosept 33 SL. Both of fungicides were most toxic to fungi at concentration A. Both of P. fumosoroseus isolates were the most sensitive to Bioczos liquid and M. anisopliae to Biosept 33 SL. All concentrations of synthetic fungicides (Sumilex 500 SC, Teldor 500 SC) restricted growth of B. bassiana and M. anisopliae isolates more from arable soil than from herbicide fallow but opposite reaction was found with respect to P. fumosoroseus.

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A Multiomic Approach to Understand How Pleurotus eryngii Transforms Non-Woody Lignocellulosic Material

Journal of Fungi ◽

10.3390/jof7060426 ◽

2021 ◽

Vol 7 (6) ◽

pp. 426

Author(s):

Ander Peña ◽

Rashid Babiker ◽

Delphine Chaduli ◽

Anna Lipzen ◽

Mei Wang ◽

...

Keyword(s):

Extracellular Enzymes ◽

Fungal Growth ◽

2D Nmr ◽

Pleurotus Eryngii ◽

Glycoside Hydrolases ◽

Oxidative Enzymes ◽

Initial Growth ◽

Lignocellulosic Material ◽

Polysaccharide Lyases ◽

Polysaccharide Monooxygenases

Pleurotus eryngii is a grassland-inhabiting fungus of biotechnological interest due to its ability to colonize non-woody lignocellulosic material. Genomic, transcriptomic, exoproteomic, and metabolomic analyses were combined to explain the enzymatic aspects underlaying wheat–straw transformation. Up-regulated and constitutive glycoside–hydrolases, polysaccharide–lyases, and carbohydrate–esterases active on polysaccharides, laccases active on lignin, and a surprisingly high amount of constitutive/inducible aryl–alcohol oxidases (AAOs) constituted the suite of extracellular enzymes at early fungal growth. Higher enzyme diversity and abundance characterized the longer-term growth, with an array of oxidoreductases involved in depolymerization of both cellulose and lignin, which were often up-regulated since initial growth. These oxidative enzymes included lytic polysaccharide monooxygenases (LPMOs) acting on crystalline polysaccharides, cellobiose dehydrogenase involved in LPMO activation, and ligninolytic peroxidases (mainly manganese-oxidizing peroxidases), together with highly abundant H2O2-producing AAOs. Interestingly, some of the most relevant enzymes acting on polysaccharides were appended to a cellulose-binding module. This is potentially related to the non-woody habitat of P. eryngii (in contrast to the wood habitat of many basidiomycetes). Additionally, insights into the intracellular catabolism of aromatic compounds, which is a neglected area of study in lignin degradation by basidiomycetes, were also provided. The multiomic approach reveals that although non-woody decay does not result in dramatic modifications, as revealed by detailed 2D-NMR and other analyses, it implies activation of the complete set of hydrolytic and oxidative enzymes characterizing lignocellulose-decaying basidiomycetes.

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Evaluation of the Entomopathogenic Fungi Beauveria bassiana, Metarhizium anisopliae, and Isaria fumosorosea for Control of Sitophilus oryzae

Journal of Food Protection ◽

10.4315/0362-028x.jfp-13-196 ◽

2014 ◽

Vol 77 (1) ◽

pp. 87-93 ◽

Cited By ~ 10

Author(s):

NICKOLAS G. KAVALLIERATOS ◽

CHRISTOS G. ATHANASSIOU ◽

MARIA M. AOUNTALA ◽

DEMETRIUS C. KONTODIMAS

Keyword(s):

Beauveria Bassiana ◽

Entomopathogenic Fungi ◽

Metarhizium Anisopliae ◽

Galleria Mellonella ◽

Sitophilus Oryzae ◽

Exposure Period ◽

Isaria Fumosorosea ◽

Stored Grain ◽

Stored Grain Pest ◽

Petri Dishes

The entomopathogenic fungi Beauveria bassiana, Metarhizium anisopliae, and Isaria fumosorosea were tested against the stored-grain pest Sitophilus oryzae. The fungi were isolated from the soil (from three locations in Attica, Greece: B. bassiana from Tatoion, M. anisopliae from Marathon, and I. fumosorosea from Aghios Stefanos) using larvae of Galleria mellonella as bait. Suspensions of 2.11 × 107 and 2.11 × 108, 1.77 × 107 and 1.77 × 108, and 1.81 × 107 and 1.81 × 108 conidia per ml of B. bassiana, M. anisopliae, and I. fumosorosea, respectively, were applied by three treatments: (i) sprayed on food and set in petri dishes with adults of S. oryzae, (ii) sprayed on adults of S. oryzae and set in petri dishes without food, and (iii) sprayed on adults of S. oryzae and set in petri dishes with food. The observed mortality of S. oryzae adults during the overall exposure period for the lowest, as well as for the highest, concentrations of B. bassiana, M. anisopliae, and I. fumosorosea ranged from 0 to 100%. Concentration was, in most of the cases tested, a critical parameter that determined the “speed of kill” of the exposed insect species for B. bassiana and M. anisopliae. Conversely, concentration was not that critical for I. fumosorosea, and survival was high in some of the combinations tested, even after 14 days of exposure. Both in the highest and the lowest concentrations of fungi, the mortality of S. oryzae adults was higher when the fungi were applied on adults than when they were applied on food. Higher mortality was observed when food was absent than when food was present, in most of the cases tested. The high efficacy levels recorded in the current study indicate that the tested fungi could be effective biocontrol agents against S. oryzae.

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Three Related Enzymes in Candida albicans Achieve Arginine- and Agmatine-Dependent Metabolism That Is Essential for Growth and Fungal Virulence

mBio ◽

10.1128/mbio.01845-20 ◽

2020 ◽

Vol 11 (4) ◽

Author(s):

Katja Schaefer ◽

Jeanette Wagener ◽

Ryan M. Ames ◽

Stella Christou ◽

Donna M. MacCallum ◽

...

Keyword(s):

Candida Albicans ◽

Amino Acid ◽

Galleria Mellonella ◽

Fungal Growth ◽

Open Reading Frames ◽

Mammalian Host ◽

No Production ◽

Triple Mutant ◽

Fungal Virulence ◽

Content Type

ABSTRACT Amino acid metabolism is crucial for fungal growth and development. Ureohydrolases produce amines when acting on l-arginine, agmatine, and guanidinobutyrate (GB), and these enzymes generate ornithine (by arginase), putrescine (by agmatinase), or GABA (by 4-guanidinobutyrase or GBase). Candida albicans can metabolize and grow on arginine, agmatine, or guanidinobutyrate as the sole nitrogen source. Three related C. albicans genes whose sequences suggested that they were putative arginase or arginase-like genes were examined for their role in these metabolic pathways. Of these, Car1 encoded the only bona fide arginase, whereas we provide evidence that the other two open reading frames, orf19.5862 and orf19.3418, encode agmatinase and guanidinobutyrase (Gbase), respectively. Analysis of strains with single and multiple mutations suggested the presence of arginase-dependent and arginase-independent routes for polyamine production. CAR1 played a role in hyphal morphogenesis in response to arginine, and the virulence of a triple mutant was reduced in both Galleria mellonella and Mus musculus infection models. In the bloodstream, arginine is an essential amino acid that is required by phagocytes to synthesize nitric oxide (NO). However, none of the single or multiple mutants affected host NO production, suggesting that they did not influence the oxidative burst of phagocytes. IMPORTANCE We show that the C. albicans ureohydrolases arginase (Car1), agmatinase (Agt1), and guanidinobutyrase (Gbu1) can orchestrate an arginase-independent route for polyamine production and that this is important for C. albicans growth and survival in microenvironments of the mammalian host.

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Environmental Tolerance of Entomopathogenic Fungi: A New Strain of Cordyceps javanica Isolated from a Whitefly Epizootic Versus Commercial Fungal Strains

Insects ◽

10.3390/insects11100711 ◽

2020 ◽

Vol 11 (10) ◽

pp. 711

Author(s):

Shaohui Wu ◽

Michael D. Toews ◽

Camila Oliveira-Hofman ◽

Robert W. Behle ◽

Alvin M. Simmons ◽

...

Keyword(s):

Environmental Factors ◽

Pest Management ◽

Beauveria Bassiana ◽

Entomopathogenic Fungi ◽

Galleria Mellonella ◽

Uv Light ◽

Variable Temperature ◽

Environmental Tolerance ◽

Southern Georgia ◽

Lower Tolerance

A new strain of Cordyceps javanica (wf GA17) was observed causing widespread epizootics among whiteflies in Southern Georgia in 2017. The tolerance of conidia to environmental factors including variable temperature and ultraviolet (UV) light was compared between this strain and three commercial strains of entomopathogenic fungi (Metarhizium brunneum F52, Cordyceps fumosorosea Apopka97, and Beauveria bassiana GHA). Under 10–30 °C, C. javanica wf GA17 responded similarly to other fungi, with the highest virulence against Galleria mellonella at 25 °C, followed by 20, 30, and 15 °C; lowest virulence was observed at 10 °C. At 35 °C and 40 °C, C. javanica wf GA17 had lower tolerance than M. brunneum F52 and B. bassiana GHA, but was superior to C. fumosorosea Apopka97 in conidia viability and post-treatment virulence. After exposure to −20 °C for 56 d, C. javanica wf GA17 exhibited lower germination than M. brunneum F52 and lower virulence than M. brunneum F52 and B. bassiana GHA, but higher germination and virulence than C. fumosorosea Apopka97. Following exposure to strong UV light, viability and virulence of all fungi were reduced with increasing exposure periods. Increased environmental tolerance of C. javanica wf GA17 over C. fumosorosea Apopka97 suggests that the new strain could have applicability for commercial pest management.

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Interplay between Fungal Infection and Bacterial Associates in the Wax Moth Galleria mellonella under Different Temperature Conditions

Journal of Fungi ◽

10.3390/jof6030170 ◽

2020 ◽

Vol 6 (3) ◽

pp. 170

Author(s):

Vadim Yu Kryukov ◽

Elena Kosman ◽

Oksana Tomilova ◽

Olga Polenogova ◽

Ulyana Rotskaya ◽

...

Keyword(s):

Fungal Infection ◽

Bacterial Communities ◽

Galleria Mellonella ◽

Fungal Growth ◽

16S Rrna Gene Sequencing ◽

Illumina Miseq ◽

Rrna Gene ◽

Cultivable Bacteria ◽

Stress Related Genes ◽

Rrna Gene Sequencing

Various insect bacterial associates are involved in pathogeneses caused by entomopathogenic fungi. The outcome of infection (fungal growth or decomposition) may depend on environmental factors such as temperature. The aim of this study was to analyze the bacterial communities and immune response of Galleria mellonella larvae injected with Cordyceps militaris and incubated at 15 °C and 25 °C. We examined changes in the bacterial CFUs, bacterial communities (Illumina MiSeq 16S rRNA gene sequencing) and expression of immune, apoptosis, ROS and stress-related genes (qPCR) in larval tissues in response to fungal infection at the mentioned temperatures. Increased survival of larvae after C. militaris injection was observed at 25 °C, although more frequent episodes of spontaneous bacteriosis were observed at this temperature compared to 15 °C. We revealed an increase in the abundance of enterococci and enterobacteria in the midgut and hemolymph in response to infection at 25 °C, which was not observed at 15 °C. Antifungal peptide genes showed the highest expression at 25 °C, while antibacterial peptides and inhibitor of apoptosis genes were strongly expressed at 15 °C. Cultivable bacteria significantly suppressed the growth of C. militaris. We suggest that fungi such as C. militaris may need low temperatures to avoid competition with host bacterial associates.

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Entomopathogenic fungal (Hyphomycetes) collection: assessment of conidial viability

Pesquisa Agropecuária Brasileira ◽

10.1590/s0100-204x1999000800023 ◽

1999 ◽

Vol 34 (8) ◽

pp. 1497-1503 ◽

Cited By ~ 3

Author(s):

Marcos Rodrigues de Faria ◽

Irene Martins ◽

Raquel Mello ◽

Myrian Silvana Tigano

Keyword(s):

Liquid Nitrogen ◽

Beauveria Bassiana ◽

Entomopathogenic Fungi ◽

Culture Collection ◽

Nomuraea Rileyi ◽

Paecilomyces Farinosus ◽

Viability Loss ◽

Conidial Viability

Twenty four strains of the entomopathogenic fungi (Hyphomycetes) Beauveria bassiana, Metarrhizium anisopliae, Nomuraea rileyi, Paecilomyces farinosus, P. fumosoroseus and P. lilacinus, maintained in the culture collection of Embrapa-Centro Nacional de Pesquisa de Recursos Genéticos e Biotecnologia (Cenargen) and preserved by lyophilization and in liquid nitrogen, had their conidial viability assessed. Germination rates of 16- to 84-month-old cultures stored in liquid nitrogen decreased, on average, less than 13.3%. For 29- to 49-month-old cultures preserved by lyophilization, the viability loss ranged, on average, from 28.6 to 94.5%. The results demonstrated the efficiency of the tested methods, especially liquid nitrogen, in preserving the viability of entomopathogenic fungi.

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