[Antimicrobial Properties of Eremoxylarin A Produced by Ascomycete of Sordariomycetes in Submerged Culture].

The fungal strain INA 01108 producing antibiotic substances with broad spectrum of antibacterial activity was isolated from the natural environment. By the morphological characteristics and DNA analysis it was shown to belong to Ascomycetes of Sordariomycetes. In submerged culture the strain produced at least four antibiotics. The major component of them was identified as eremophilane-type sesquiterpene eremoxylarin A. Eremoxylarin A is effective in vitro against grampositive bacteria, including methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin group glycopeptide antibiotics resistant Leuconostoc mesenteroides VKPM B-4177. The efficacy and toxicity of eremoxylarin A was determined on a murine staphylococcal sepsis model. The dose of 6.25 mg/kg provided 100% recovery and survival of the animals, while the dose of 3.12 mg/kg was close to the ED50. The chemical structure of eremoxylarin A allows to modify the antibiotic and such studies may be relevant to design a less toxic derivative without loss of the valuable antimicrobial properties.

[Endomembrane system of fungi: traditional and modern conceptions].

The review is devoted to the endomembrane system in Fungi, which is mostly presented in cell by membrane organelles, taking part in exo- and endocytosis. Main tenets of the modern model of exo- and endocytosis are covered by the example of the apical cells of filamentous fungi. Particular attention is given to studies of endomembrane system, which were carried out by electron microscopy in the last century--the era in science preceding the prevalence of methods of fluorescence microscopy, immune and molecular genetic tagging. Endomembrane organelles, which are described in classical studies, but have not been identified or differentiated from other organelles modern specialists are under consideration. Among these organelles are lomasomes, plasmalemmasomes, membranosomes and myelin-like bodies. Possible reasons for the "loss" of given structures in today's science and its place in modern proposed model of endomembrane system are discussed. In addition, some specific questions are mentioned, namely: the presence of mushroom mycelium endobionts, morphologically similar to endomembrane organelles, and the increase of number of endomembrane structures in bracket fungi, limited in nitrogen nutrition.

[Report of Quambalaria cyanescens associated with birch].

Long-term microbiological investigation of the pollen of silver birch (Betula pendula) in the Mos- cow, and Moscow region areas revealed that: almost one-third of the analyzed samples, contained the fungus identified by morphological, cultural, and molecular genetic techniques as Quambalaria cyanescens (de Hoog & G. A. de Vries) Z.W. de Beer, Begerow & R. Bauer. This species was previously known mostly as a syrmbiont of tropical plants of the generaEucalyptus and Cortyminbia and has not been isolated in Russia. We revealed a close association between Quambalaria cyanescens and silver birch. The micromycete was regulaly detected in pollen samples, as well as on the.inside and outside of the aments, on the surface of leaves and branches. It was never isolated from other plant species in the investigated area. The data on the morphological and cultural characteristics of the fungus, its cell ultrastructure, and occurrence are presented, as well as the phylogenetic analysis of the isolated strains.

[Effect of sodium chloride concentration in the medium on the composition of the membrane lipids and carbohydrates in the cytosol of the fungus Fusarium sp].

The fungus Fusarium sp. isolated from saline soil was identified by the ITS1-5.8S-ITS2 and the D1/D2 domains of LSU RNA as a member of the Fusarium incarnatum-equiseti species group. Its growth patterns on media with different NaCl concentrations indicated its adaptation as halotolerance. The mechanisms of halotolerance included accumulation of arabitol a five-atom noncyclic polyol, a decreased sterols/phospholipids ratio, elevated level of phosphatidic acids in the phospholipids, and increased unsaturation of phospholipids, which was especially pronounced in the idiophase. The mechanisms of halotolerance of the mycelial fungus Fusarium sp. are discussed in comparison with yeasts and yeastlike fungi.

[Longevity control in fungi and other organisms. The conception of scales].

The review deals mainly with gerontological processes that occur on the cellular-colonial level of organization in fungi and cellular-tissular level in other organisms. Aging and anti-aging mechanisms operating on these levels of organization can be considered as common ones for all living things. Fungi, as an object with tissular-like organization of thallus, afford a broad spectrum of possibilities as to solving the tasks of general gerontological import. Three basic (chronological, replicative, and cell-suicidal) and several auxiliary mechanisms of aging are singled out, the classification is given of stochastic aging factors accumulating in cells. It is shown that in complex multi-cellular organisms, aging and anti-aging mechanisms operate on the level of interactions between tissues, though in the base of their actions lie the aforesaid conservative basic mechanisms. Preliminary generalized conception of aging--the conception of scales--is put forward that is founded on the model of balanced and non-balanced counteractions between stressful impacts and various mechanisms of aging and anti-aging with different extent of genetic preprogramming. The importance is reaffirmed of mycological gerontology contribution to broadening of inferences on aging nature.

[Morphogenesis and ultrastructure of basidiomycetes Agaricus and Pleurotus mitochondria].

Mitochondrial morphogenesis in 31 strains of 9 species of Agaricus--A. arvensis Schaeff., A. bisporus (Lange) Imbach, A. bitorquis (Quel.) Sacc., A. campestris L., A. excellens (F. H. Moller) F. H. Moller, A. macrocarpus (F. H. Möller) F. H. Möller, A. silvaticus Schaeff., A. silvicola (Vittad.) Peck, A. xanthodermus Genev--and 2 strains of Pleurotus--P. ostreatus (Jacg.) P. Kumm., P. pulmonarius (Fr.) Quel.--has many common features in mitochondria distribution under favorable growth conditions (type 1) and in reconstruction of chondriom (fission or fragmentation) under unfavorable growth conditions and aging (type 2). The first type of mitochondria distribution was observed in heterokaryotic mycelium of some Agaricus strains and Pleurotus grown in agar medium during 7-14 days, and also in submerged mycelium of some Agaricus strains and Pleurotus. The second type of mitochondria distribution was observed in homokaryotic Agaricus strains under condition of starvation, in aging mycelium (28 days of growth), and in submerged mycelium of most of Agaricus strains. The first type of chondriom consists of small granular mitochondria in the apical cells and long snake-like network in subapical cells, and restores almost completely the mitochondrial network in the aging mycelium cells. The second type of chondriom consists of small granular mitochondria in all cells of mycelium. The surface of chondriom type 2 mitochondrial membrane was usually closely associated with ribosomes and changed crists. Such mycelium cells in A. bisporus strain Bs94 were TUNEL positive. So, the types of mitochondria morphogenesis in the Agaricus and Pleurotus mycelium cells are similar at different time and growth conditions and depend on complex of physiological and biochemical process in the mycelium cells.

[Proteolytic enzymes of the fungi: extracellular proteases of xylotrophic basidiomycetes].

Proteolytic enzymes of the fungi attract attention of investigators due to many reasons among which their large diversity, wide substrate specificity, stability under extreme conditions (pH, temperature) are most important. Their functional significance, including various processes, from hydrolysis of macromolecular substrates under deficiency of nitrogen compounds to initiation and maintenance of pathogenesis, is also very interesting. The present review deals with classification and biochemical properies of extracellular fungal proteinases, their physiological role as well as fields of practical application. Much attention is given to pecularities of extracellular proteinases of xylotrophic basidiomycetes-- an exceedingly important group of the fungi from the point of functioning of biological communities, and their participation in biodestruction of plant waste which are also used as source of food and medicinal preparations. Special attention is focused on regulation of synthesis and secretion of extracellular proteinases of xylotrophic basidiomycetes.

[Micromorphological features of Pleurotus pulmonarius (Fr.) Quel. and P. ostreatus (Jacq.) P. Kumm. strains in pure and binary culture with yeasts].

Micromorphology of oyster mushrooms Pleurotus ostreatus (Jacq.) Quel. and P. pulmonarius (Fr.) Quel. was studied in pure and binary culture with yeasts (Cryptococcus laurentii 1629, Rhodotorula minuta 2790, Sporidiobolus salmonicolor (see text for symbol) 31A-11, Candida krusie 3452, Pichia holstii 3438). The cultures were cultivated on malt-agar and water agar. Various mycelial structures were described: strands, rings, thin searching mycelium, clamps, crystals, head-like offshoots, mycelial fragments, chlamydospores, and coralloid hyphae. Vegetative mycelia interact in different ways (forming anastomoses, strands, system of thin anucleate hyphae) within the same culture. Head-like offshoots of mycelial cells, previously regarded as spores of asexual reproduction, appeared to lack nuclei and to be filled with polyphosphates. Coralloid hyphae, which induce yeast cell lysis after direct contact, were detected only in binary culture with yeasts under condition of nitrogen deficit. The same way of feeding is typical for carnivorous mushrooms.

[DNA quantification in nuclei of cultivated mushroom with DAPI staining]. / Kolichestvennoe opredeleniie DNK v iadrakh shampin'ona dvusporovogo pri okrashivanii reaktivom DAPI.

Agaricus bisporus (Lange) Imbach is actively cultivated amphithallic basidiomycete, in which various strains are primary homothallic, heterothallic or secondary homothallic. Countings of relative nuclear DNA content by means of DAPI stain and its comparison in different strains can help to understand the mushroom's life cycle features. The authors for the first time observed change of nuclear phases in basidia of A. bisporus strains with different types of life cycle and revealed that DNA content in diploid nuclei is about 1.3 times higher than in haploid ones. The method is highly sensitive and can be used for quantitative measurings of nuclear DNA even in objects with nuclei of about 1 mkm in diameter.

[Ultrastructure of the cell wall in vegetative mycelia of Agaricus bisporus (Lange) Imbach]. / Osobennosti ul'trastruktury kletochnoi stenki vegetativnogo mitseliia Agaricus bisporus (Lange) Imbach.

A comparative description of the cell wall ultrastructure was made for vegetative mycelia in Agaricus bisporus, other species of Agaricus, and Pleurotus ostreatus. Apart from well known ultrastructural features of cell walls of the examined strains, inclusions of different kinds were observed. The cell wall thickness varied in different Agaricus strains. Maximum differences between the examined strains were 1.4 times. The cell wall thickness in P. ostreatus strains was by 3 to 4 times less than that in Agaricus strains. Using electron micrographs, a brief description of mycelia and A. bisporus basidia ultrastructure was composed.

[Antimicrobial activity of Laetiporus sulphureus strains grown in submerged culture]. / Antimikrobnaia aktivnost' shtammov Laetiporus sulphureus v usloviiakh glubinnogo kul'tivirovaniia.

Cultural conditions for growth and fruit body formation were elaborated to four strains of Laetiporus sulphureus isolated from nature. All strains demonstrated antimicrobial activity against a wide spectrum of gram-positive and gram-negative bacteria during agar and submerged cultivation including methicillin-resistant strain of Staphylococcus aureus (MRSA) and glycopeptide-resistant strain of Leuconostoc mesenteroides. Antifungal activity was not found. The level of antimicrobial activity during submerged cultivation reached maximum after seven days of growth on specific medium with soybean meal and corn liquid; the next four weeks its increasing was not so manifested. Antimicrobial activity correlated with orange pigment secretion and cultural liquid acidification to pH 2.0-2.8 that indicates on acid nature of synthesized products.

[Selection of the optimal conditions for the procurement and regeneration of protoplasts of the industrial strain of Streptomyces rimosus, the producer of oxytetracycline, and the effect of the protoplasting process on the antibiotic activity]. / Podbor optimal'nykh uslovii polucheniia i regeneratsii protoplastov promyshlennogo shtamma Streptomyces rimosus produtsent oksitetratsiklina i vliianie protoplastirovaniia na antibioticheskuiu aktivnost'.

Optimal conditions for protoplasting of the Streptomyces rimosus industrial strain No. 1 producing oxytetracycline were developed. Observation of the early stages of the protoplast regeneration in microchambers showed that there were two regeneration types: normal and anomalous. The latter was likely defined by the glycine effect on cell wall synthesis. It was accompanied by the stage in which the protoplasts had the form of multiplying protoplast-like cells. The protoplasting of the S. rimosus culture producing oxytetracycline resulted in an increase in the variability of an antibiotic producing property and the frequency of low active variants.

[Cytologic control of protoplast formation and regeneration in Streptomyces erythraeus, strain BTCC-2]. / Tsitologicheskii kontrol' polucheniia i regeneratsii protoplastov Streptomyces erythraeus shtamm BTCC-2.

Experiments on protoplast formation and regeneration in S. erythraeus, strain BTCC-2 (Saccharopolyspora erythrae) were performed under microscopic control at all the stages. It was shown that the highest protoplast titer was provided by the mycelium grown in one step in the absence of glycine. For characterizing the protoplasts formed by the mycelium grown under different conditions, their regeneration capacity was estimated by microscopic examination of the protoplasts after 15-20-hour growth in microchambers and evaluation of the regeneration efficiency 7-10 hours later. Of interest was the fact of spontaneous development of colonies consisting of the protoplast-like cells (L-cells) in 15-20 hours. Such colonies were formed only by the protoplasts grown from the mycelium incubated in one step in the absence of glycine or in the presence of 0.1 per cent of glycine. Such conditions provided also the maximum efficiency of the protoplast regeneration. The long-term storage of protoplasts led to a decrease in their viability.

[Development of protoplasts of Streptomycetes on media favoring the regeneration and formation of L-forms]. / Razvitie protoplastov streptomitsetov na sredakh, sposobstvuiushchikh regeneratsii i obrazovaniiu L-form.

The protoplasts of three Streptomyces species and their regenerative ability were studied using light microscopy. When Streptomyces lividans and S. erythraeus protoplasts are cultivated on regeneration media, their regeneration is not synchronous during the first day; some protoplasts revert to yield the mycelial form and also L-forms of these cultures are produced. If the protoplasts are transferred to a medium inducing L-forms, they grow and multiply for a long time with the production of L-form colonies. This process is maintained if S. lividans L-form cells are passaged on the medium inducing L-forms, but the protoplasts revert to yield the mycelial form on the regeneration medium.

[Isolation and properties of the protoplasts of Streptomyces erythraeus]. / Poluchenie i svoistva protoplastov Streptomyces erythraeus.

The paper is concerned with development of conditions for cultivation of the erythromycin-producing organism and preparation and maintenance of its stable viable protoplasts. Optimal conditions for the culture growth and protoplasting were developed. Two-stage cultivation of the organism on media PB and S provided dense diffuse or diffuse local growth characterized by low differentiation and higher homogenicity. The incubation time at stage I was 66 hours and that at stage II was not more than 24 hours at respective temperatures. The culture was incubated on a shaker in the presence of glycine at the minimum concentration. The presence of glycine in the medium altered the culture cell walls which was evident from changing of staining by Gram from + to +/- . Treatment of such a culture with lysozyme for 30 minutes provided formation of up to 2.10(9) protoplasts per 1. ml. It is possible to maintain the protoplasts in frozen state at -20 degrees C in medium P for 1 month. Under such storage conditions the titer of the viable protoplasts as compared to the initial one decreased only 2-fold after the one-month storage.

[Cytological study of early stages of regeneration of protoplasts of fungi and Streptomyces]. / Tsitologicheskoe izuchenie rannikh étapov regeneratsii protoplastov gribov i streptomitsetov.

Early stages of Penicillium chrysogenum 51 and Streptomyces lividans 66 protoplast regeneration on solid media were studied microscopically under conditions of microcompartments. It was shown that at the early regeneration stages there were both rapid reversion into the mycelial form and a retarded one. In P. chrysogenum retarded regeneration resulted in formation of hypha-like structures or protoplast breaking into fragments of various sizes. Some of the fragments restored the cell walls and mycelial organization whereas the others lysed. As a result of the breaking and compartmentalization of the viable areas one protoplasts formed several centers of P. chrysogenum colony reversion. Retarded regeneration of protoplasts in S. lividans 66 resulted in their growth and multiplication in the protoplast-like L-form. On media with penicillin, glycine and horse serum there were isolated colonies of S. lividans L-forms subject to passages or reversion depending on the medium composition.

[Formation of A-factor-type regulators by representatives of the genus Micromonospora]. / Obrazovanie reguliatorov tipa A-faktora predstaviteliami roda Micromonospora.

It is known that substances similar in their chemical structure and biological action to A-factor, a regulator of Streptomyces griseus differentiation are widely observed in streptomycetes. The present paper is concerned with studies on capacity for producing A-factor-like regulators in 13 strains belonging to 11 species of genus Micromonospora. It was shown that 7 strains had an A-factor-deficient test mutant effect analogous to the effect of A-factor. Relation of A-factor-like substance production to biosynthesis of sisomicin and sporogenesis was investigated in M. fusca var. sisomicini, strain RIA 1572. A-Factor-like substances were isolated from submersion culture of strain 1572 on the 6th-8th day of the culture growth. By the time it corresponded to the highest number of the spores in the medium. The highest levels of sisomicin were detected on the 3rd day of the fermentation process. After that period its concentration markedly lowered. Exogenic addition of A-factor had no effect on sisomicin biosynthesis but resulted in increased numbers of the spores.