[Molecular genetic determinants of intraspecific polymorphism of the phytopathogenic fungus Cryphonectria parasitica].

The review summarizes the current evidence on the phytopathogenic fungus Cryphonectria parasitica, which is a classic object for studying hypovirulence. Phenotypic manifestations of hypovirulence and themolecular mechanisms of action of the mycovirus Cryphonectria hypovirus (CHV) infecting the fungus are described in detail. Genetic determinants of vegetative incompatibility in C. parasitica (a phenomenon increasing polymorphism of the fungus and preventing CHV expansion) are considered. The data on C. parasitica polymorphism are correlated with the data on the distribution of different CHV species in the European, American, and Asian populations of the fungus.

[New producers of biologically active compounds--fungal strains of the genus Penicillium isolated from permafrost].

Screening of producers of secondary metabolites was carried out among 25 fungal strains of Penicillium genus isolated from permafrost in Arctic and Antarctic regions and Kamchatka. Nearly 50% of the investigated strains synthesize biologically active substances of alkaloid nature: ergot alkaloids, diketopiperazines, and quinoline derivatives. A large group of the identified metabolites belongs to mycotoxins. A strain of Penicillium waksmanii was found producing epoxiagroclavine-I and quinocitrinins. The main physiological and biochemical characteristics of this producer were investigated.

Secondary metabolites of the genus Penicillium from undisturbed and anthropogenically altered Antarctic habitats.

From undisturbed Antarctic habitats (permafrost sediments 30-150 thousand years of age, water of Radok Lake) and superficial deposits contaminated with petroleum products, we isolated 14 and 9 strains of Penicillium fungi, respectively. Comparison of the fungal complexes showed them to differ by species composition; only two species-P. palitans and P. solitum-were in the species lists of both groups. The identified secondary metabolites in the investigated strains belonged to diketopiperazine (group of roquefortines, rugulosuvin B), benzodiazepine (anacin, cyclopenins), quinoline alkaloids (viridicatins), clavine ergot alkaloids (α-cyclopiazonic acid, festuclavine, fumigaclavines), polycyclic indole alkaloids (communesin B, chaetoglobosin A), amino acid derivatives (N-acetyltryptamine, chrysogins, penicillin G), polyketides (citreoviridin A, mycophenolic acid), and terpenes (andrastins, phomenone). Strains isolated from anthropogenically altered habitats produced a more complete and characteristic profile of exometabolites, as compared with strains isolated from undisturbed habitats. It is only from contaminated soils there were isolated fungi that produced more structurally diverse secondary metabolites pertaining to polycyclic indole alkaloids and terpenoids. The fungi isolated from contaminated samples can be used in biodegradation of oil spills and bioremediation of the environment, and also as producers of promising biologically active compounds.

Microbial populations in Antarctic permafrost: biodiversity, state, age, and implication for astrobiology.

Antarctic permafrost soils have not received as much geocryological and biological study as has been devoted to the ice sheet, though the permafrost is more stable and older and inhabited by more microbes. This makes these soils potentially more informative and a more significant microbial repository than ice sheets. Due to the stability of the subsurface physicochemical regime, Antarctic permafrost is not an extreme environment but a balanced natural one. Up to 10(4) viable cells/g, whose age presumably corresponds to the longevity of the permanently frozen state of the sediments, have been isolated from Antarctic permafrost. Along with the microbes, metabolic by-products are preserved. This presumed natural cryopreservation makes it possible to observe what may be the oldest microbial communities on Earth. Here, we describe the Antarctic permafrost habitat and biodiversity and provide a model for martian ecosystems.

[Synthesis of alpha-cyclopiazonic acid by fungi of the genus Aspergillus].

The presence of alpha-cyclopiazonic acid has been studied among metabolites of Aspergillus fungi. The study was performed with 138 cultures of 13 species obtained from the All-Russia Collection of Microorganisms and the collection of our institute. alpha-Cyclopiazonic acid was most frequently encountered among the metabolites of the section Flavi (the ability to synthesize alpha-cyclopiazonic acid was expressed in 61% of the strains of A. flavus, 83% of the strains of A. oryzae, and all strains of A. tamarii). This expression index for A. versicolor was less than 5%. We showed for the first time that alpha-cyclopiazonic acid is produced by A. fumigatus and A. phoenicis (expression in 30% of the strains of either species).

[Secondary Metabolites of Fungi of the Usti Section, Genus Aspergillus and Their Application in Chemosystematics].

Secondary metabolites of 22 fungal strains (genus Aspergillus, section Usti) isolated at diverse geographic regions, including the Arctic permafrost deposits, were studied. The studied strains were found to synthesize a variety of biologically active compounds, structurally identified as drimane sesqueterpenoids, isoquinoline alkaloids (TMC-120 A−C, derivative 1), meroterpenoids (austalides О and J), and anthraquinone pigments (averufin, versicolorin C). Desferritriacetylfusigen production by A. calidoustus isolates is reported for the first time. The individual spectra of secondary metabolites were used for reidentification of 17 strains, of which 15 were identified as A. calidoustus and two, as A. pseudodeflectus.

[Mycelial fungi maintained in the Russia Collection of Microorganisms (VKM IBPM RAS)].

Information on application of diversity of mycelial fungi maintained in the Russia Collection of Microorganisms (VKM) at the Institute for Biochemistry and Physiology of Microorganisms, Russian Academy of Sciences to research and biotechnology as producers is presented, as well as data on additions to the basic collection.

[Production of mycophenolic acid by fungi of the genus Penicillium link].

Out of 36 strains of fungi of the genus Penicillium, some of which were isolated from ancient permafrost soils, 14 strains synthesized mycophenolic acid (MPA). Maximal (over 500 mg/l) accumulation of MPA in culture liquid was observed in P. brevicompactum strains (VKM F-457, VKM F-477, and VKM F-1150). This was the first study to detect MPA in representatives of the species P. rugulosum; in three strains of this species (VKM FW-665, VKM FW-717, and VKM FW-733), the level of MPA accumulation exceeded 300 mg/l. The time course of the synthesis of MPA by the P. rugulosum strain VKM FW-733 was studied. It was shown that the synthesis of this metabolite was dramatically intensified at the stationary growth phase (ten days).

[Isolation of Penicillium DIERCKX 1901--a producer of diketopiperazine alkaloids rokefortin and 3,12-dihydrokefortin--from permafrost]. / Vydelennye iz vechnoi merzloty griby Penicillium aurantiogriseum DIERCKX 1901--produtsenty diketopiperazinovykh alkaloidov rokefortina i 3/12-digidrorokefortina.

Secondary metabolites of the three strains of Penicillium aurantiogriseum, isolated from permafrost sediments, were identified. It was found that these fungi synthesized diketopiperazine alkaloids roquefortine and 3,12-dihydroroquefortine. The strain VKM FW-766 synthesized alkaloids in in the course of certain growth-related processes. When the strain was grown on a mineral medium, time courses of roquefortine and 3,12-dihydroroquefortine concentrations were characterized by biphasic curves.

Exo-metabolites of mycelial fungi isolated in production premises of cheese-making and meat-processing plants.

Data were obtained on the species composition of mycelial fungi isolated from the air of workrooms and production premises in cheese-making and meat-processing plants. The strains studied were shown to be capable of producing various low molecular weight compounds. Many of them are mycotoxins such as α-cyclopiazonic acid (CPA), mycophenolic acid (MPA), citrinin, cladosporin, roquefortine and ergot alkaloids. The profiles of the secondary metabolites were used to elucidate the species' names of the isolated strains.

[Fungal diversity in the Antarctic active layer].

Taxonomic diversity of fungi in the samples of the active layer of Antarctica was investigated using conventional microbiological techniques and metagenomic analysis of total DNA extracted from environmental samples. The list of Antarctic microscopic fungi was expanded, including detection of the species representing a portion of the fungal complex, which is nonculturable or sterile on conventional nutrient media.

[Lipid synthesis by Geomyces pannorum under the impact of stress factors].

Lipogenic activity and fatty acid composition of two strains of Geomyces pannorum were studied in the course of fungal growth. The strains were isolated from an Arctic cryopeg lens (VKM FW-2241) and from Central Russia (VKM F-3808). The adaptive reactions in both strains towards the temperature decreasing to 2 degrees C involved intensification of the fatty acid desaturation. The degree of lipid unsaturation increased mainly due to a higher amount of alpha-linolenic acid (alpha C18:3) especially in the case of strain VKM FW-2241. Elevated NaCl concentration in the medium enhanced the level of linoleic acid (C18:2) which apparently played a specific role in osmoprotection. Strain VKM FW-2241 was more tolerant to increased salinity than strain VKM F-3808. Almost complete inhibition of the growth of strains VKM F-3808 and VKM FW-2241 occurred at salinity of 10 and 20%, respectively; however, the viability of the strains was not affected. Under the combined effect of high salinity and hypothermia, the ratio between C18:2 and alpha-C18:3 acids was intermediate, indicating that these acids were involved in two adaptation mechanisms. The inhibition of fungal growth under stress was found to result in lipid overproduction. An increased pool of energy-rich lipids in fungi possibly contributes to their strategy of cell survival.

[The structure of micromycete complexes in permafrost and cryopegs of the Arctic].

A comparative study of the structure of micromycete complexes has been performed. The samples of micromycetes were taken by boring from unique habitats: cryopegs (lenses of non-freezing hypersaline water in ancient permafrost horizons) and permafrost Arctic sediments of different age enclosing these cryopegs. The possibility of characterizing the above habitats by the structure of specific complexes of microscopic fungi using qualitative and quantitative indices at extremely low numbers of these organisms was demonstrated.

[Microbial communities of ancient seeds derived from permanently frozen Pleistocene deposits].

Microbial communities from the surface of ancient seeds of higher plants and embedding frozen material dated to the late Pleistocene (formed about 30 thousand years ago) were studied by various methods: scanning electron microscopy, epifluorescence microscopy, and inoculation of nutrient media, followed by identification of isolated cultures. Both prokaryotic and eukaryotic microorganisms were found on the surface of ancient seeds. The total quantity of bacterial cells determined by direct counting and dilution plating (CFU) for the samples of ancient seeds exceeded the value in the embedding frozen material by one to two orders of magnitude. This pattern was not maintained for mycelial fungi; their quantity in the embedding material was also rather high. A significant difference was revealed between the microbial communities of ancient seeds and embedding frozen material. These findings suggest that ancient plant seeds are a particular ecological niche for microorganisms existing in permafrost and require individual detailed study.

[Halo- and psychrotolerant Geomyces fungi from arctic cryopegs and marine deposits].

Comparative characterization of Geomyces isolates was performed. The isolates were obtained from Arctic cryopegs and the surrounding ancient marine deposits, from nonsaline permafrost soils, and from temperate environments. Microbiological (cultural and morphological) and molecular criteria were used to confirm the identification of the isolates as Geomyces pannorum. The isolates from cryopegs and surrounding marine deposits were shown to differ from those obtained from nonsaline soils and temperate environments in their ability to grow at negative temperatures (-2 degrees C) under increased salt concentration (10%). The results are discussed in relation to the possible inheritance of the adaptive characteristics acquired in specific environments.

Thiol-dependent serine proteinase from Paecilomyces lilacinus: purification and catalytic properties.

An extracellular thiol-dependent serine proteinase was isolated from culture medium filtrate of the microscopic fungus Paecilomyces lilacinus with a yield of 33%. The enzyme is inactivated by specific inhibitors of serine proteinases, phenylmethylsulfonyl fluoride, as well as by chloromercuribenzoate and mercury acetate, but is resistant to chelating agents. The proteinase has broad specificity, hydrolyzes proteins and p-nitroanilides of N-acylated tripeptides, exhibiting maximal activity in hydrolysis of substrates containing long hydrophobic and aromatic residues (norleucine, leucine, phenylalanine) as well as arginine at the P1 position. The enzyme has a molecular weight of 33 kD. The enzyme is most active at pH 10.0-11.5; it is thermostable and is characterized by broad optimum temperature range (30-60 degrees C), displaying about 25% of maximal activity at 0 degrees C. The N-terminal sequence of the enzyme (Gly-Ala-Thr-Thr-Gln-Gly-Ala-Thr-Gly/Ile-Xxx-Gly) has no distinct homology with known primary structures of serine proteinases from fungi and bacilli. Based on its physicochemical and enzymatic properties, the serine proteinase from P. lilacinus can be classified as a thiol-dependent subtilisin-like enzyme.

[The fungus Penicillium variabile sopp 1912 isolated from permafrost deposits as a producer of rugulovasines].

It has been established that relict fungi Penicillium variabile Sopp can synthesize clavine alkaloids, rugulovasines A and B, which are revealed in this species for the first time. Submerged cultivation of the strain-producer revealed several microcycles of conidia formation. The synthesis of alkaloids was also of a cyclic character. The synchronism beween the cycles of rugulovasine biosynthesis and conidia formation was revealed. Zinc ions stimulated fungal growth but had a negative effect on the biosynthesis of rugulovasines.

[Germination of phytopathogenic fungi conidia]. / Zakonomernosti prorastaniia konidii fitopatogennykh gribov.

The autoregulation of conidium germination in phytopathogenic micromycetes of the genera Fusarium, Botrytis, and Bipolaris was studied. It was shown that Trichoderma longibrachiatum was less competitive than Fusarium oxysporum after their simultaneous inoculation but inhibited the phytopathogen growth in the case of earlier introduction. In the latter case, no autoinhibition of the germination of F. oxysporum conidia occurred; moreover, cooperative effect was observed, i.e., the number of germinated F. oxysporum conidia increased with an increase in their density.

[The quantity and structure of the root-associated microbial complexes of two greenhouse rose cultivars]. / Chislennost' i struktura mikrobnykh kompleksov kornevykh sistem teplichnykh roz.

The study of the root-associated microbial complexes of affected and healthy rose plants of two cultivars (Grand gala and Royal velvet) grown in a greenhouse showed that the biomass of eukaryotic microorganisms in the rhizoplane and rhizosphere of healthy rose plants and in the surrounding soil was considerably lower than in the same loci of affected plants. In contrast, the biomass of root-associated prokaryotic microorganisms was higher in the case of healthy than in the case of affected rose plants. The root-associated bacterial complexes of both affected and healthy rose plants were dominated by the genera Arthrobacter, Rhodococcus, and Myxobacterium and did not contain phytopathogenic bacteria. The root-associated fungal complex of healthy roses was dominated by fungi of the genus Trichoderma, whereas that of the affected rose plants was dominated by the species Aureobasidium microstictum. The affected cane cuttings and cankers occurring on affected canes were found to contain Coniothyrium fuckelii (the causal fungus of rose stem canker) and sclerotia of Botrytis cinerea (the causal fungus of gray rot). The micromycete complex of healthy rose plants was not so diverse as was the micromycete complex of affected rose plants.