Tolypocladium Cylindrosporum Research Articles

Tetramic acid-motif natural products from a marine fungus Tolypocladium cylindrosporum FB06 and their anti-Parkinson activities.

The online version contains supplementary material available at 10.1007/s42995-023-00198-7.

Activity Against Musca domestica of Hypocrealean Fungi Isolated from Culicids in Central Brazil and Formulated in Vermiculite.

Musca domestica L. is a cosmopolitan nuisance of high sanitary importance. Entomopathogenic fungi are innovative and attractive tools for integrated control of the housefly to overcome insufficient levels of control caused by increasing resistance of this pest against chemical insecticides. High virulence of a fungal strain is a prerequisite to develop a mycoinsecticide, and the present study investigated the potential of hypocrealean fungi from the genera Beauveria, Clonostachys, Cordyceps, Akanthomyces, Metarhizium, and Tolypocladium, isolated from mosquitoes in Central Brazil against M. domestica. The highest mortalities (larvae, pupae, and adults) were caused by Metarhizium humberi IP 478 (98%) and IP 421 (90%), Metarhizium anisopliae IP 432 (85%), Beauveria bassiana IP 433 (82%), and Tolypocladium cylindrosporum IP 425 (68%) after a 23-day exposure of initially pre-pupating third instar larvae to conidia mixed with vermiculite. Lethal concentrations to kill 90% of adults of IP 433 and IP 478 were 5 × 107 and 108 conidia g-1 substrate, respectively. Fifty percent of adults were killed within 4 to 5days of exposure initially as pupae close to emergence to substrate treated with conidia of IP 478 or IP 433 at 1.1 × 108 conidia g-1, respectively. The other fungal strains tested were less virulent. The results demonstrate high potentials for conidial preparations in vermiculite of IP 433 and IP 478 as candidates for the biological control of both pre-pupating larvae, pupae, and emerging adults of houseflies.

Identification, Characterization, and Evaluation of Nematophagous Fungal Species of Arthrobotrys and Tolypocladium for the Management of Meloidogyne incognita.

Root-knot nematodes belonging to the genus Meloidogyne are agriculturally important pests, and biocontrol strategies offer safer alternatives for their management. In the present study, two fungal species from Indian soils were identified as Arthrobotrys thaumasia and Tolypocladium cylindrosporum based on morphological characteristics and further confirmed using molecular markers. In vitro evaluation of A. thaumasia against M. incognita and Caenorhabditis elegans showed 82 and 73% parasitism, respectively, whereas T. cylindrosporum gave 65.2 and 57.7% parasitism, respectively. Similarly, culture filtrates of A. thaumasia caused 57.7 and 53.7% mortality of M. incognita and C. elegans, respectively, whereas T. cylindrosporum caused higher mortality of 87.3 and 64%, respectively. Besides, greenhouse evaluation of both fungi against M. incognita infecting tomato significantly reduced nematode disease burden reflecting parasitic success measured as the total number of galls, egg masses, eggs per egg mass, and derived nematode multiplication factor. Application of A. thaumasia and T. cylindrosporum reduced nematode multiplication factor by 80 and 95%, respectively, compared with control. General metabolite profiling of tested fungi using gas chromatography–mass spectrometry and ultra-performance liquid chromatography–quadrupole/time of flight mass spectrometry reported for the first time here showed presence of various volatile and non-volatile compounds with nematicidal activity, viz., trimethyl-heptadiene, methyl-hexadecanol, dodecadienal, decane, terpendole E, dodecane, acetamido-6-anthraquinone, and hexadecanol. Also, other compounds such as undecane, dibutyl-disulfide, octadecenal, paganin, talathermophilin, dactylarin, tolypyridone A, tolypyridone B, pyridoxatin, and destruxin were identified, reported in the literature to possess antibacterial, antifungal, and insecticidal properties. This is the first report of the occurrence of both fungi from India and pioneer demonstration of T. cylindrosporum for root-knot nematode management.

Destructive activity of hydrocarbon-oxidizing micromycetes extracted from the substrates of the coastal areas, the Barents and White Seas

We analyzed the hydrocarbon-oxidizing activity of 33 micromycetes strains isolated from water and soils of the White Sea and Barents Sea coastal territories. The microscopic fungi studied were represented by the following genera: Acremonium, Aspergillus, Meyerozyma, Oidiodendron, Paradendryphiella, Penicillium, Pseudoqymnoascus, Tolypocladium, Trichoderma, Sarocladium, Talaromyces, Umbelopsis. Hydrocarbon-oxidizing activity of fungi was carried out in a laboratory experiment for two time periods: 14 and 28 days. The residual concentration of oil hydrocarbons in the medium was determined by the infrared spectrometry method. We revealed that micromycetes had different oil-destructive activity. The decrease in oil content for the strains over 14 days ranged from 11 to 83 %. Tolypocladium inflatum st.1, T. inflatum st.2, Penicillium thomii, Meyerozyma guilliermondii, P. simplicissimum, P. camemberti, P. solitum and Trichoderma minutisporum strains had the greatest oil destructive activity, reducing the concentration in the medium by 57-83 %. These micromycetes were isolated from coastal substrates, the content of oil products in which did not exceed the MPC. We found that some individual strains of microscopic fungi had high oil-destructive activity for a longer period of time: Tolypocladium cylindrosporum reduced the content of oil hydrocarbons for 28 days by 95 %. No clear relationship between the fungal biomass and the degree of decomposition of oil products was found, but some tolerant species of micromycetes can actively decompose oil hydrocarbons without significantly increasing their biomass. The identified active strains of hydrocarbon-oxidizing micromycetes can be used to create biopreparations with the prospect of their further use for bioremediation of oil spills in the northern marine ecosystems.

Conidiation under illumination enhances conidial tolerance of insect-pathogenic fungi to environmental stresses

Light is an important signal for fungi in the environment and induces many genes with roles in stress and virulence responses. Conidia of the entomopathogenic fungi Aschersonia aleyrodis, Beauveria bassiana, Cordyceps fumosorosea, Lecanicillium aphanocladii, Metarhizium anisopliae, Metarhizium brunneum, Metarhizium robertsii, Simplicillium lanosoniveum, Tolypocladium cylindrosporum, and Tolypocladium inflatum were produced on potato dextrose agar (PDA) medium under continuous white light, on PDA medium in the dark, or under nutritional stress (= Czapek medium without sucrose = MM) in the dark. The conidial tolerance of these species produced under these different conditions were evaluated in relation to heat stress, oxidative stress (menadione), osmotic stress (KCl), UV radiation, and genotoxic stress caused by 4-nitroquinoline 1-oxide (4-NQO). Several fungal species demonstrated greater stress tolerance when conidia were produced under white light than in the dark; for instance white light induced higher tolerance of A. aleyrodis to KCl and 4-NQO; B. bassiana to KCl and 4-NQO; C. fumosorosea to UV radiation; M. anisopliae to heat and menadione; M. brunneum to menadione, KCl, UV radiation, and 4-NQO; M. robertsii to heat, menadione, KCl, and UV radiation; and T. cylindrosporum to menadione and KCl. However, conidia of L. aphanocladii, S. lanosoniveum, and T. inflatum produced under white light exhibited similar tolerance as conidia produced in the dark. When conidia were produced on MM, a much stronger stress tolerance was found for B. bassiana to menadione, KCl, UV radiation, and 4-NQO; C. fumosorosea to KCl and 4-NQO; Metarhizium species to heat, menadione, KCl, and UV radiation; T. cylindrosporum to menadione and UV radiation; and T. inflatum to heat and UV radiation. Again, conidia of L. aphanocladii and S. lanosoniveum produced on MM had similar tolerance to conidia produced on PDA medium in the dark. Therefore, white light is an important factor that induces higher stress tolerance in some insect-pathogenic fungi, but growth in nutritional stress always provides in conidia with stronger stress tolerance than conidia produced under white light.

The intestinal microbiota of rats under the influence of biotechnological strains of microorganisms from various taxonomic groups

Introduction. The microbiota of the gastrointestinal tract (GIT) plays an essential role in maintaining human health. Many factors, including industrial pollutions with biotechnological strains of microbes, can affect the normal balance of intestinal microbiota. The biotechnological industry nowadays produces a wide range of products for medical and veterinary use, agriculture, food, chemical industries, etc. To develop hygienic standards that regulate the possible adverse effect of biotechnological strains of microorganisms on workers’ health, the intestinal microflora of rats in the experiment can be studied. The data obtained were used as the basic concept in elaborating state sanitary standards for limitations of the concentrations of biotechnological strains of microorganisms in the ambient air of the working area and settlements’ atmosphere. Materials and methods. We have tested 52 strains of microorganisms applied in biotechnology as producers of a variety of biological substances. They included members of different taxonomic groups: gram-positive and gram-negative bacteria, actinomycetes, molds, and yeasts. The experiments were carried out on conventional male and female white rats (290-320 g, body weight). Each test and control group of animals included eight animals. The strains of microorganisms mentioned above were given to animals by inhalation of minimal effective doses of microbes in the concentrations of 103-108 CFU/m3 during one month. To demonstrate possible adverse effects to gut microflora, the routine bacteriological examination of animal feces was performed. To do this, after the priming, the 10-fold dilutions of animal feces in sterile saline were inoculated onto a set of general-purpose and selective culture media for Enterobacteriaceae members, staphylococci, enterococci, clostridia, bifidobacteria, lactobacilli, and fungi, with subsequent identification of the genus of the isolated microorganism. After that, the concentrations of microorganisms were calculated and measured in lg of CFU/g of feces. The Institutional Ethical Committee of Animal Care and Use of the Pirogov Russian National Research Medical University approved all procedures involving animals. The results of experiments were analyzed with a simple t-test using Statistica (v.6.0, Stat Soft, USA) and Microsoft Office Excel 2007. Results were considered statistically significant when p < 0.05. Results. The most notable changes in the composition of the intestinal microbiota were observed after inhaling of yeasts of genus Candida at the level of 103-104 CFU/m3 and in cases of exposure to molds (Aspergillus awamori, Penicillium funiculosum, and Tolypocladium cylindrosporum) in the concentration of 2•104 CFU/m3, and gram-negative bacteria of the genus Alcaligenes and genus Pseudomonas at 5•105 CFU/m3. We observed a dramatic decrease of Escherichia coli and the increase of gram-positive bacteria (staphylococci, enterococci). For some genera of biotechnological strains, a significant decline in the content of lactobacilli was also shown. On the other hand, Rhodococcus did not cause any disturbances even at high concentrations in the ambient air. Conclusion. The obtained data can be used to develop biosafety and hygienic standards for industrial microbes to help decrease or minimize the occupational risk of infection or undesirable allergic effect when working with biotechnological strains of microbes in the ambient air of residential areas.

Cylindromicin from Arctic-Derived Fungus Tolypocladium sp. SCSIO 40433.

The fungus strain SCSIO 40433 was isolated from an Arctic-derived glacier sediment sample and characterized as Tolypocladium cylindrosporum. A new compound, cylindromicin (1), and seven known secondary metabolites (2–8) were isolated from this strain. The chemical structures of these compounds were elucidated by comprehensive spectroscopic analyses. Cylindromicin (1) featured a 3,4-dihydro-2H-pyran skeleton. The absolute configuration of compound 1 was assigned via interpretation of key Nuclear Overhauser Effect Spectroscopy (NOESY) correlations and Electronic Circular Dichroism (ECD) calculation. Cylindromicin (1) exhibited significant tyrosinase inhibition activity. This study highlights Polar fungi as a potential resource for new bioactive natural products.

Composition and Structure of the Community of Mycelial Fungi in the Bottom Sediments of the White Sea

Fifty samples of bottom sediments were taken in 2016–2017 in the Kandalaksha Bay of the White Sea from the depth of 1.5–15.0 m, and 1419 colonies of mycelial fungi were obtained. Based on morphological and cultural features, a total of 136 morphotypes were classified, and 81 of these were identified down to the species level. Thirteen species were new to the White Sea. The most common species were Tolypocladium cylindrosporum, Penicillium chrysogenum, Tolypocladium inflatum, Penicillium glabrum, and the anamorph of Pseudogymnoascus pannorum. The dominance of Ascomycota was a common characteristic of the mycobiota due to the anamorphic species, and the class Sordariomycetes was the most diverse and numerous group. Assessment of species richness using a cumulative curve and the calculation of the expected total number of species adjusted using the Chao2 method evidenced that approximately 81% of the species diversity was found within the study. The ordination of samples by the nMDS method with the ANOSIM test showed the high importance of combining the samples into groups based on the year of sampling and type of sediment as well as the year of sampling and type of ecotope. Therefore, the type of sediment associated with the type of coast and the presence of fresh runoff were the most important factors for the formation of mycobiota. Moreover, the communities of mycelial fungi change from year to year in the studied bottom sediments.

Complete mitogenome of the entomopathogenic fungus Tolypocladium cylindrosporum

In this study, the complete mitogenome of an entomopathogenic fungus Tolypocladium cylindrosporum is assembled and annotated. This circular mitogenome is 34,698 bp in length and consists of two rRNA genes (rnl and rns), 26 tRNA genes, 14 standard protein-coding genes of the oxidative phosphorylation system, and four intergenic free-standing ORFs. A total of six introns (all group I) were identified in atp9, cob, cox1, and rnl, and they may encode ribosomal protein S3, LAGLIDADG or GIY-YIG endonucleases. Phylogenetic analysis based on concatenated mitochondrial protein sequences confirms T. cylindrosporum in Ophiocordycipitaceae.

Determinants of Coinfection in the Mycoviruses.

Coinfections of mycoviruses are generally common. The coinfecting mycoviruses are not necessarily the result of horizontal virus transmission among homologous fungal hosts compatible for anastomosis, but involve mycoviruses from phylogenetically diverse sources (Herrero and Zabalgogeazcoa, 2011; Osaki et al., 2016; Ran et al., 2016; Hao et al., 2018). An experimental study with different combinations of transmission scenarios among four partitiviruses showed a significant positive influence of one virus (Heterobasidion partitivirus13-an1) on its distantly related coinfecting partner Heterobasidion partitivirus 15-pa1, but no support of coinfection between two related viruses (Heterobasidion partitiviruses 11-au1 and 11-pa1) (Kashif et al., 2019). A more stable coinfection between distantly related species than conspecific strains was also reported among mycoviruses infecting the fungus, Heterobasidion parviporum (Vainio et al., 2015). Studies covering large geographical areas indicate that the mycoviruses in coinfections belong to the local fungal community (Ran et al., 2016; Arjona-Lopez et al., 2018). Arjona-Lopez et al. (2018) found a non-overlapping set of coinfecting mycoviruses in isolates of Rosellinia necatrix from Japan and the Mediterranean that match with mycoviruses from the respective local fungal pools. In general, transmission of mycoviruses occurs through anastomosis of vegetatively compatible strains of the same species, but phylogenetic evidence implies occasional transmission across vegetatively incompatible strains. The transmissions across vegetatively incompatible fungal hosts are poorly studied except in a few cases (Liu et al., 2003, 2016; Yaegashi et al., 2013a). In addition to transmission across heterologous fungi, interactions among mycoviruses play direct roles in coinfection. The interactions among coinfecting mycoviruses are diverse, ranging from synergistic to neutral to antagonistic. In some cases, mycovirus coinfection induces genome rearrangement of one of the coinfecting partners, likely through recombination (Sun and Suzuki, 2008). A more comprehensive review on the interactions among coinfecting mycoviruses is found in Hillman et al. (2018). The diversity of coinfecting mycoviruses and the diverse nature of interactions may imply that coinfections occur freely without any constraints. However, in examples from in vitro experiments the frequency of coinfection varies with different fungal systems and in many cases is lower than what would be expected from random incidences. For example, in 43 isolates of the ascomycete Tolypocladium cylindrosporum, coinfections of mycoviruses were reported in only in about 5% of the population (Herrero and Zabalgogeazcoa, 2011). In contrast, in almost 200 isolates of Ustilaginoidea virens examined multiple dsRNA elements were found in large samples, and the coinfection frequency of two common mycoviruses: Ustilaginoidea virens RNA virus 1 (Totiviridae) and Ustilaginoidea virens RNA virus 4 (unclassified) was close to 30% (Jiang et al., 2015). In our own study of about 200 North American isolates of Pseudogymnoascus destructans a fungus causing a deadly disease in bats, the coinfection incidence of a partitivirus (Thapa et al., 2016) and an unclassified mycovirus is close to 25%, but only in a geographically restricted area (unpublished). Independent segregation of coinfecting mycoviruses in the population has been described only in a few cases where the interaction is likely neutral, while in other cases one of the partners influences the presence of other. In the coinfection of two unrelated RNA viruses, Yado-nushi virus 1 (YnV1) and Yado-kari virus 1 (YkV1) in Rosellinia necatrix, YkV1 is dependent on YnV1. YkV1 does not segregate independently in the population (Zhang et al., 2016). Thus, the coinfection scenarios are likely influenced by the interactions between coinfecting viruses or at the level of mycovirus transmission, particularly during the transmission across vegetatively incompatible fungi or from other factors. Such constraints led us to investigate the determinants of mycovirus coinfections. Here, we discuss some of the determinants for mycovirus coinfection based on the literature available and provide our opinions on coinfection biology.

Microscopic Fungi of White Sea Sediments

The micromycete complex population size and taxonomic structure is determined for sediments (bottom soils) of the White Sea; species actively functioning in the littoral zone and at depths of 10–30 m are identified. The bottom-soil fungi population is pretty low (hundreds to several thousand colony-forming units (CFUs) per 1 g), while the species diversity is quite rich. In total, 70 species are identified; 90% of those are Ascomycota anamorphs (Capnodiales, Eurotiales, Hypocreales, Pleosporales, Saccharomycetales, and Incertae sedis), Zygomycota (Mucoromycota) (Mucorales and Umbelopsidales orders) constituted (8%), and the share of Basidiomycota (Agaricales order) is 2%. The actively functioning bottom-soil fungi are identified on the basis of their ability to develop on organic substrates (starch, cellulose, chitin, and pieces of laminaria thallus) and grow on seawater media at low temperatures and varying oxygen levels, i.e., under conditions similar to those observed in the studied ecotope. This fungi group includes some 20 species: Paradendryphiella salina, Acremonium tubakii, A. potronii, Pseudeurotium hygrophilum, Pseudogymnoascus pannorum, Emericellopsis sphaerospora, Oidiodendron periconioides, Parengyodontium album, Lecanicillium muscarium, and representatives of genera Tolypocladium and Sarocladium. These species are typical for marine and cold habitats; some of them are well-known chitinolytics and associates of insects and algae, and many of them are able to grow in anaerobic conditions. For some of the species (Aspergillus brasiliensis, A. sydowii, Cladosporium cladosporioides, Emericellopsis sphaerospora, Oidiodendron periconioides, Pseudeurotium hygrophilum, Tolypocladium cylindrosporum, T. tundrense, Umbelopsis vinacea, Penicillium spp., and Talaromyces spp.), this ability was discovered for the first time. Further studies are required to obtain detailed ecophysiological descriptions of marine isolates of species actively functioning in the bottom soils.

The Xenon Test Chamber Q-SUN® for testing realistic tolerances of fungi exposed to simulated full spectrum solar radiation

The low survival of insect-pathogenic fungi when used for insect control in agriculture is mainly due to the deleterious effects of ultraviolet radiation and heat from solar irradiation. In this study, conidia of 15 species of entomopathogenic fungi were exposed to simulated full-spectrum solar radiation emitted by a Xenon Test Chamber Q-SUN XE-3-HC 340S (Q-LAB® Corporation, Westlake, OH, USA), which very closely simulates full-spectrum solar radiation. A dendrogram obtained from cluster analyses, based on lethal time 50 % and 90 % calculated by Probit analyses, separated the fungi into three clusters: cluster 3 contains species with highest tolerance to simulated full-spectrum solar radiation, included Metarhizium acridum, Cladosporium herbarum, and Trichothecium roseum with LT50 > 200 min irradiation. Cluster 2 contains eight species with moderate UV tolerance: Aschersonia aleyrodis, Isaria fumosorosea, Mariannaea pruinosa, Metarhizium anisopliae, Metarhizium brunneum, Metarhizium robertsii, Simplicillium lanosoniveum, and Torrubiella homopterorum with LT50 between 120 and 150 min irradiation. The four species in cluster 1 had the lowest UV tolerance: Lecanicillium aphanocladii, Beauveria bassiana, Tolypocladium cylindrosporum, and Tolypocladium inflatum with LT50 < 120 min irradiation. The QSUN Xenon Test Chamber XE3 is often used by the pharmaceutical and automotive industry to test light stability and weathering, respectively, but it was never used to evaluate fungal tolerance to full-spectrum solar radiation before. We conclude that the equipment provided an excellent tool for testing realistic tolerances of fungi to full-spectrum solar radiation of microbial agents for insect biological control in agriculture.

Production of fumonisins by endophytic strains of Tolypocladium cylindrosporum and its relation to fungal virus infection.

Fumonisins were first discovered in Fusarium verticillioides, a fungus associated to disease and asymptomatic infections in maize. Afterwards, other fungal taxa have been found to produce fumonisins. The entomopathogenic ascomycete Tolypocladium cylindrosporum has been isolated from soil and also as an endophyte from leaves of grasses. The objectives of this work were to determine the in vitro production of fumonisin B (FB) mycotoxins and the immunosuppressive compound cyclosporine A (CyA) in several strains of T. cylindrosporum, and to examine the effect of fungal virus infection and temperature in FB production. FB1 was detected in 30% of the strains, ranging from 0.16 to 5.52μgcm-2 in solid media, and FB2 was detected in 78% of the strains, ranging from 0.764 to 40.92μgcm-2. CyA was not detected in any strain. The mean FB2 concentration of the endophytic strain Tc37W was three times greater (p < 0.05) than that of any other strain. Up to 34% more of FB2 was detected in strains infected by the virus TcV3 than in the corresponding virus-free versions. The effect of temperature on FB2 content was interactively significantly dependent on fungal strain and growth medium; in the YES medium, the FB2 of virus-infected strains Tc37-1V and Tc37W increased by 67 and 16%, respectively, at 26°C as compared to 20°C. The FB concentration in some fungal strains was similar to that in fungi associated to food and feed intoxications.

Characterization of Tolypocladium cylindrosporum (Hypocreales: Ophiocordycipitaceae) and Its Impact Against Aedes aegypti and Aedes albopictus Eggs at Low Temperature.

We examined the growth characteristics of Tolypocladium cylindrosporum IBT 41712 and its potential to infect eggs of Aedes aegypti and Ae. albopictus at a low temperature (15°C). When grown on Sabouraud dextrose agar supplemented with yeast extract, the IBT 41712 formed white colonies turning to a slightly darker, off-white color when mature. The mycelia bore swollen conidiophores producing smooth-walled, oblong to cylindrical conidia with varying sizes, ranging from 1.5 to 3.5 μm long. To determine the optimum temperature for the fungus, we cultured the fungus at eight temperatures (4°C, 12°C, 15°C, 21°C, 28°C, 33°C, 37°C, and 40°C) and measured the diametric growth. The optimum temperature for growth was 28°C since it had the highest diametric growth rate (2.1 ± 0.05 mm/day) and the fastest sporulation period (within 8-10 days of incubation). There was no fungal growth at the 3 highest temperatures (33°C, 37°C, and 40°C) but plates incubated at 33°C, when shifted to optimal temperature (28°C), showed visible growth indicating that following incubation at 33°C, the fungus remained viable. The IBT 41712 successfully infected mosquito eggs at 15°C. Fungal treatment induced egg hatch on moist seed-germination paper and this effect was more pronounced in Ae. aegypti compared to Ae. albopictus. When treated eggs were immersed in dH2O 21 days posttreatment, larval hatch of both Ae. aegypti (control = 91%, 1 × 107 conidia/ml, fungal treatment = 0%) and Ae. albopictus (control = 85%, fungal treatment = 28%) was significantly lower in fungal treatment compared to the controls. The ability of the strain to grow in a wide temperature range, and effectively infect mosquito eggs and induce egg hatch at a low temperature warrants further investigation for its potential as a mosquito control agent targeting eggs that overwinter or undergo long diapause.

Tetramic Acids and Pyridone Alkaloids from the Endolichenic Fungus Tolypocladium cylindrosporum

Three new tetramic acid derivatives, tolypocladenols A1, A2, and B (1-3), a new pyridone alkaloid, tolypyridone A (4), and a new coumarin derivative, 3,8-dihydroxy-4-(4-hydroxyphenyl)-6-methylcoumarin (5), together with four known compounds (6-9) were isolated from the endolichenic fungus Tolypocladium cylindrosporum, which inhabits the lichen Lethariella zahlbruckneri. Structures of these compounds were determined by comprehensive analysis of spectroscopic data and single-crystal X-ray diffraction determination. Bioassay of the isolated compounds found that pyridoxatin (7) was cytotoxic to human cancer cells by induction of G0/G1 cell cycle arrest and apoptosis.

Some properties and possible biological role of peptidase inhibitors from the entomopathogenic fungus Tolypocladium cylindrosporum.

The activities of secreted and mycelial inhibitors of proteolytic enzymes from fungi of the order Hypocreales have been investigated. Inhibitors of bromelain, papain, and trypsin of low molecular mass (about 1kDa) and a subtilisin proteinaceous inhibitor with molecular mass of 45kDa were revealed in the culture liquid of the fungus Tolypocladium cylindrosporum. The subtilisin inhibitor from T. cylindrosporum has antibiotic properties, significantly decreased the activity of purified bacterial enzymes, and prevented the growth of the bacterium Pseudomonas sp. Data suggesting the existence in fungi of the Hypocreales order of two pools of peptidase inhibitors have been obtained.

Susceptibility of Cabbage Maggot Larvae (Diptera: Anthomyiidae) to Hypocreales Entomopathogenic Fungi

The pathogenicity of six Metarhizium spp., four Beauveria bassiana (Balsamo) Vuillemin, and four Tolypocladium cylindrosporum Gams (Ascomycota: Hypocreales) fungal pathogens exposed to third-instar Delia radicum L. was evaluated in laboratory bioassays. The presence of intra- and intergeneric variations concerning the pathogenicity of the isolates was investigated. Results show that all Metarhizium spp. and T. cylindrosporum isolates caused a noteworthy mortality to the third instar and consequently reduced adult eclosion. The well-known standard, F52 strain (identified as Metarhizium brunneum), resulted in up to 79% reduction in D. radicum eclosion. The other Metarhizium isolates including UAMH 9197 (Metarhizium anisopliae) and UAMH 2801 (M. brunneum), as well as T. cylindrosporum DAOM 167325 and DAOM 183952, produced a mean eclosion reduction of >50%. While the pathogenicity of Metarhizium spp. and T. cylindrosporum is similar, the B. bassiana isolates are undoubtedly less pathogenic. Based on the results obtained with the selected isolates, no intrageneric differences relative to the pathogenicity of the isolates appeared to be present. Globally, this study deepened the knowledge about D. radicum susceptibility toward Hypocreales entomopathogenic fungi, chiefly T. cylindrosporum. The implications of this study regarding the development of a biological control agent are discussed.

Responsiveness of entomopathogenic fungi to menadione-induced oxidative stress

Entomopathogenic fungi are predisposed to ROS induced by heat and UV–A radiation when outside the insect host. When inside the host, they are subject to phagocytic cells that generate ROS to eliminate invading pathogens. The oxidative stress tolerance of the entomopathogenic fungi Aschersonia aleyrodis (ARSEF 430 and 10276), Aschersonia placenta (ARSEF 7637), Beauveria bassiana (ARSEF 252), Isaria fumosorosea (ARSEF 3889), Lecanicillium aphanocladii (ARSEF 6433), Metarhizium acridum (ARSEF 324), Metarhizium anisopliae (ARSEF 5749), Metarhizium brunneum (ARSEF 1187 and ARSEF 5626), Metarhizium robertsii (ARSEF 2575), Tolypocladium cylindrosporum (ARSEF 3392), Tolypocladium inflatum (ARSEF 4877), and Simplicillium lanosoniveum (ARSEF 6430 and ARSEF 6651) was studied based on conidial germination on a medium supplemented with menadione. Conidial germination was evaluated 24 h after inoculation on potato dextrose agar (PDA) (control) or PDA supplemented with menadione. The two Aschersonia species (ARSEF 430, 7637, and 10276) were the most susceptible fungi, followed by the two Tolypocladium species (ARSEF 3392 and 4877) and the M. acridum (ARSEF 324). Metarhizium brunneum (ARSEF 5626) and M. anisopliae (ARSEF 5749) were the most tolerant isolates with MIC 0.28 mM. All fungal isolates, except ARSEF 5626 and ARSEF 5749, were not able to germinate at 0.20 mM.

Conidial water affinity is an important characteristic for thermotolerance in entomopathogenic fungi

The interspecific thermotolerance of several species of entomopathogenic fungi was evaluated based on the conidial water affinity. The species were divided between hydrophilic and hydrophobic conidia. The species with hydrophobic conidia were Beauveria bassiana (ARSEF 252), Metarhizium brunneum (ARSEF 1187), Metarhizium robertsii (ARSEF 2575), Isaria fumosorosea (ARSEF 3889) and Metarhizium anisopliae s.l. (ARSEF 5749). The species with hydrophilic conidia were Tolypocladium cylindrosporum (ARSEF 3392), Tolypocladium inflatum (ARSEF 4877), Simplicillium lanosoniveum (ARSEF 6430), Lecanicillium aphanocladii (ARSEF 6433), S. lanosoniveum (ARSEF 6651), Aschersonia placenta (ARSEF 7637) and Aschersonia aleyrodis (ARSEF 10276). The conidial surface tension of each isolate was also studied. Conidial suspensions were exposed to 38, 41 or 45 °C. After exposure, the suspensions were inoculated on media and conidial germination was evaluated. Considerable differences in thermotolerance were found among the 12 entomopathogenic fungal species. Species with hydrophobic conidia were generally more thermotolerant than species with hydrophilic conidia. All isolates with hydrophobic conidia showed higher conidial surface tension than the isolates with hydrophilic conidia.

Molecular characterization of a novel victorivirus from the entomopathogenic fungus Beauveria bassiana

New Zealand isolates of the entomopathogenic fungus Beauveria were examined for the presence of dsRNAs and virus-like particles. Seven out of nine isolates contained one or more high-molecular-weight dsRNAs and all seven contained isometric virus particles ranging in size from 30 to 50 nm. B. bassiana isolate ICMP#6887 contained a single dsRNA band of ~6 kb and isometric virus-like particles of ~50 nm in diameter. Sequencing revealed that the virus from ICMP#6887 had a genome of 5,327 nt with two overlapping ORFs coding for a putative coat protein (CP) and an RNA-dependent RNA-polymerase (RdRp). The sequence showed a highest CP identity of 58.3 % to Tolypocladium cylindrosporum virus 1 (TcV1) and a highest RdRp identity of 48.8 % to Sphaeropsis sapinea RNA virus 1 (SsRV1). Since both TcV1 and SsRV1 belong to the genus Victorivirus, the new virus from B. bassiana ICMP#6887 was tentatively assigned the name Beauveria bassiana victorivirus 1 (BbVV1-6887).