Biopreparation of cotton fabric with enzymes produced by solid-state fermentation

Fungal enzyme preparations from Phanerochaete chrysosporium, Aspergillus oryzae, Aspergillus giganteus and Trichoderma virens, produced by solid-state fermentation (SSF) on cotton seed-coat fragment waste as substrate and enzyme inducer were investigated in biopreparation of cotton fabric. Cotton seed-coat fragment is rich in lignin, cellulose and hemicelluloses, therefore enzyme complexes produced by target fungi on such a substrate can be used effectively to degrade impurities in cotton fabrics during biopreparation. Activities of extracellular hydrolytic and ligninolytic enzymes were determined from the SSF extract materials. The potential of the hydrolytic and accompanying oxidative enzymes in the whole SSF cultures was exploited in degradation of seed-coat fragments and other coloring materials of greige cotton fabric. Enzyme assays indicated that many extracellular enzymes have been produced under these conditions including both hydrolytic and oxidative enzymes. A. oryzae NRRL 3485 produced significantly higher amounts of both hydrolytic and oxidative enzymes than other tested fungi. Best results in removal of seed-coat fragments from cotton fabric were obtained by P. chrysosporium NCAIM (=ATCC 34541), P. chrysosporium VKM F-1767 and A. oryzae NRRL 3485 SSF enzyme complexes.     

Hydrolytic enzymes and their proteinaceous inhibitors in regulation of plant–pathogen interactions

This review considers the main groups of hydrolytic enzymes associated with plant pathogens, as well as proteinaceous inhibitors of these enzymes, acting as the components of plant defense system. The role of hydrolases is described in the development of a pathological process in plant tissues. Significance of hydrolase inhibitors in the development of plant resistance to pathogens is analyzed. It is proposed that specific interactions in the “host plant–pathogen” system, involving hydrolytic enzymes and their proteinaceous inhibitors, depend on the nutritional specialization of fungi.     

Hydrolytic enzymes and ability of arbuscular mycorrhizal fungi to colonize roots

The production of hydrolytic enzymes from external mycelia associated with roots and colonized soybean roots (Glycine max L.) inoculated with different arbuscular-mycorrhizal (AM) fungi of the genus GLOMUS:, and the possible relationship between these activities and the capacity of the AM fungi to colonize plant roots was studied. There were differences in root colonization and plant growth between the GLOMUS: strains, and also between two isolates of G. mosseae. Hydrolytic activities in the root and external mycelia associated with roots differed in the AM fungi tested. Correlations were only found between the endoxyloglucanase activity of the external mycelia associated with roots of the AM fungi tested and the percentage root colonization or plant growth. However, hydrolytic activities of roots colonized by the different endophytes correlated with those of external mycelia. The hydrolytic activities were not qualitatively different because the endoxyloglucanase from AM colonized roots and the external mycelia did not show a high degree of polymorphism in the different species of fungus tested. The possible role of the hydrolytic activity of external hyphae of AM fungi was discussed as a factor affecting fungal ability to colonize the root and influence plant growth.     

Contribution of hydrolytic enzymes produced by saprophytic fungi to the decrease in plant toxicity caused by water-soluble substances in olive mill dry residue

We studied the influence of saprophytic fungi on the toxic effect that the water-soluble substances in dry residues from olive (ADOR) have on the growth of plants. All saprophytic fungi were able to decrease the phytotoxicity of ADOR, although the toxicity of this residue did not decrease in the same way. Penicillium chrysogenum was able to reduce the toxicity of ADOR when this residue was applied at the highest dose of 15%. Fusarium lateritum, F. graminearum and Mucor racemosus were able to reduce the toxicity of ADOR when this residue was applied at the intermediate doses. However, F. oxysporum decreased the phytotoxicity of ADOR only when the residue was applied at the lowest dose of 2.5%. All saprophytic fungi tested produce endoglucanase, endopolymetylgalacturonase and endoxiloglucanase when grown in the presence of ADOR. A close relationship was found between the decrease in the phytotoxicity of ADOR and the amount of hydrolytic enzymes produced by the saprophytic fungi. These results shows that hydrolytic enzymes can be important in the degradation of phytotoxic substances present in olive mill dry residue.     

Inhibitory Effect of an Antimicrobial Preparation from Lipids of Marine Fishes on Tissue and Microbial Enzymes

A new food preservative from marine fish lipids, was obtained possessing pronounced activity in relation to bacteria and microscopic fungi. The effects of this preparation on enzymes of microorganisms and muscle tissue of marine hydrobionts were studied. In vitro, the preparation irreversibly inhibited acid and alkaline proteases and proteolytic and lipolytic enzymes of microorganisms and reduced enzyme activity in fish muscle tissue. The inhibitory effect of this preparation on enzymes contributes to stabilization of hydrolytic processes in meat of hydrobionts and suppresses microorganism growth during storage.     

Extracellular enzymes and the pathogenesis of nematophagous fungi

Nematophagous fungi are an important group of soil microorganisms that can suppress the populations of plant-parasitic nematodes. The pathogenic mechanisms of nematophagous fungi are diverse: They can be parasitical–mechanical through producing specialized capturing devices, or toxin-dependent. During infections, a variety of virulence factors may be involved against nematodes by nematophagous fungi. In this review, we present up-to-date information on the modes of infection by nematophagous fungi. The roles of extracellular hydrolytic enzymes and other virulence factors involved in infection against nematodes were summarized. The biochemical properties and peptide sequences of a special group of enzymes, the serine proteases, were compared, and their implications in infections were discussed. We also discussed the impact of emerging new techniques on our understanding of this unique group of fungi.     

Inhibitory effect of antimicrobial preparation from lipids of marine fishes on tissue and microbial enzymes

We obtained a new food preservative from marine fish lipids possessing pronounced activity in relation to bacteria and microscopic fungi. The effects of this preparation on enzymes of microorganisms and muscle tissue of marine hydrobionts were studied. In vitro the preparation irreversibly inhibited acid and alkaline proteases and proteolytic and lipolytic enzymes of microorganisms and reduced enzyme activity in fish muscle tissue. The inhibitory effect of this preparation on enzymes contributes to stabilization of hydrolytic processes in meat of hydrobionts and suppresses microorganism growth in storage.     

Activity of native hydrolytic enzymes and their association with the cell wall of three ectomycorrhizal fungi

The ecological and biogeochemical relevance of hydrolytic enzymes associated with the fungal cell wall has been poorly studied in ectomycorrhizal (ECM) fungi. We used a modified sequential extraction procedure to investigate the activity of various hydrolytic enzymes (β-glucosidase, acid-phosphatase, leucine-aminopeptidase, chitinase, xylanase and glucuronidase) and their association with the cell wall of three ECM fungi (Rhizopogon roseolus, Paxillus involutus and Piloderma croceum). Fungi were grown on C-rich solid medium under three different P concentrations (3.7, 0.37 and 0.037 mM). The sequential extraction procedure classifies enzymes as: (a) cytosolic, (b) loosely bound, (c) hydrophobically bound, (d) ionically bound and (e) covalently bound. Results showed that for the same fungus absolute enzymatic activity was affected by P concentration, whilst enzymatic compartmentalization among the cytosol and the cell wall fractions was not. The association of enzymes with the cell wall was fungus- and enzyme-specific. Our data indicate also that enzymes best known for being either extracellular or cytosolic or both, do act in muro as well. The ecological implications of cell wall-bound enzymes and the potential applications and limitations of sequential extractions are further discussed.     

Extracellular peptidases of insect-associated fungi and their possible use in biological control programs and as pathogenicity markers

This review deals with characteristics of peptidases of fungi whose life cycles are associated with insects to varying degrees. The review examines the characteristic features of the extracellular peptidases of entomopathogenic fungi, the dependence of the specificity of these peptidases on the ecological characteristics of the fungi, and the role of peptidases in the development of the pathogenesis. Data on the properties and physiological role of hydrolytic enzymes of symbiotic fungi in “fungal gardens” are also considered in detail. For the development of representations about mechanisms of control over populations of insect pests, special attention is given to analysis of possibilities of genetic engineering for the creation of entomopathogens with enhanced virulence. Clarification of the role of fungi and their secreted enzymes and careful environmental studies are still required to explain their significance in the composition of the biota and to ensure widespread adoption of these organisms as effective biological control agents. The systematization and comparative analysis of the existing data on extracellular peptidases of insect-associated fungi will help in the planning of further work and the search for markers of pathogenesis and symbiosis.     

Production of plant protoplasts by enzymes from selected wood-destroying fungi

The goal of this work was to select higher fungi for the production of enzymes able to produce plant protoplasts. The hydrolytic capacity of various species of wood-detsroying fungi was examined to select those with the maximal hydrolytic capacity. The selection criteria included attack on cellulose, and oligo-and polysaccharides by each fungus species. Lenzites elegans, gonoderma applanatum and Pycnoporus sanguineus produced thr richest enzymatic mixtures, which were evaluated for their protoplasting ability. L. elegans has the best enzymatic mixture able to [roduced plant protoplasts with the shortest time for maceration of plant tissues, followed by G. applanatum and P. sanguineus. The potential of these fungi and the simplicity of enzyme proudtcion suggest the possibility of commercial applications.Correepondence to: A. R. Sampietro     

Phospholipase A2 and phospholipase B activities in fungi

As saprophytes or disease causing microorganisms, fungi acquire nutrients from dead organic material or living host organisms. Lipids as structural components of cell membranes and storage compartments play an important role as energy-rich food source. In recent years, it also has become clear that lipids have a wide range of bioactive properties including signal transduction and cell to cell communication. Thus, it is not surprising that fungi possess a broad range of hydrolytic enzymes that attack neutral lipids and phospholipids. Especially during infection of a mammalian host, phospholipase A(2) (PLA(2)) enzymes released by fungi could play important roles not only for nutrient acquisition and tissue invasion, but for intricate modulation of the host's immune response. Sequencing of fungal genomes has revealed a wide range of genes encoding PLA(2) activities in fungi. We are just beginning to become aware of the significance these enzymes could have for the fungal cells and their interaction with the host.     

Rio Tinto as a niche for acidophilus enzymes of industrial relevance

Lignocellulosic residues are amongst the most abundant waste products on Earth. Therefore, there is an increasing interest in the utilization of these residues for bioethanol production and for biorefineries to produce compounds of industrial interest. Enzymes that breakdown cellulose and hemicellulose into oligomers and monosaccharides are required in these processes and cellulolytic enzymes with optimum activity at a low pH area are desirable for industrial processes. Here, we explore the fungal biodiversity of Rıo Tinto, the largest acidic ecosystem on Earth, as far as the secretion of cellulolytic enzymes is concerned. Using colorimetric and industrial substrates, we show that a high proportion of the fungi present in this extremophilic environment secrete a wide range of enzymes that are able to hydrolyze cellulose and hemicellulose at acidic pH (4.5–5). Shotgun proteomic analysis of the secretomes of some of these fungi has identified different cellulases and hemicellulolytic enzymes as well as a number of auxiliary enzymes. Supplementation of pre-industrial cocktails from Myceliophtora with Rio Tinto secretomes increased the amount of monosaccharides released from corn stover or sugar cane straw. We conclude that the Rio Tinto fungi display a good variety of hydrolytic enzymes with high industrial potential.     

Hydrolytic activities of anaerobic fungi from wild blue bull (Boselaphus tragocamelus)

The anaerobic fungi play an active role in the plant fibre degradation by producing a wide array of potential hydrolytic enzymes in the rumen. In present study, 12 anaerobic fungal strains were isolated from the faecal samples of wild blue bull, and identified as species of Piromyces, Anaeromyces, Orpinomyces and Neocallimastix based on their morphological characteristics. Isolate WNG-12 (Piromyces sp.), showed maximum filter paper cellulase (23 mIU ml(-1)) and xylanase (127 mIU ml(-1)) activity, while WNG-5 (Piromyces sp.) showed maximum carboxymethyl cellulase activity (231 mIU ml(-1)). Based on the results obtained, it can be stated that Piromyces sp. WNG-12 may be a promising isolate in utilizing fibre rich diets in the rumen as evidenced by the production of hydrolytic enzymes in vitro.     

Alternatives to Trichoderma reesei in biofuel production

Mutant strains of Trichoderma reesei are considered indisputable champions in cellulase production among biomass-degrading fungi. So, it is not surprising that most R&D projects on bioethanol production from lignocellulosics have been based on using T. reesei cellulases. The present review focuses on whether any serious alternatives to T. reesei enzymes in cellulose hydrolysis exist. Although not widely accepted, more and more data have been accumulated that demonstrate that fungi belonging to the genera Penicillium, Acremonium and Chrysosporium might represent such alternatives because they are competitive to T. reesei on some important parameters, such as protein production level, cellulase hydrolytic performance per unit of activity or milligram of protein.     

Biosynthesis of hydrolytic enzymes during cocultivation of macro- and micromycetes

Effects of co-cultivation of higher Basidiomycetes and Phycomycetes on the biosynthesis of cellulases, amylases, and proteases was studied. Four optimal pairs of fungal cultures were selected. Of these, three pairs belonged to higher fungi, and one pair was constituted by fungi of distinct ecological groups, a macromycete and a micromycete. The activities of amylase and protease were 1.5 to 2 times higher, and the activity of cellulase was lower during the growth of higher fungi associations. The mixed association of the macromycete Schizophyllum commune and the micromycete Mucor sp. was the most active producer of hydrolytic enzymes. During the growth of this mixed association, a fourfold and 1.5-fold increases were observed in the activity of endoglucanase and protease, respectively, paralleled by stimulation of amylase formation.     

Genome sequence of the lignocellulose degrading fungus Phanerochaete chrysosporium strain RP78

White rot fungi efficiently degrade lignin, a complex aromatic polymer in wood that is among the most abundant natural materials on earth. These fungi use extracellular oxidative enzymes that are also able to transform related aromatic compounds found in explosive contaminants, pesticides and toxic waste. We have sequenced the 30-million base-pair genome of Phanerochaete chrysosporium strain RP78 using a whole genome shotgun approach. The P. chrysosporium genome reveals an impressive array of genes encoding secreted oxidases, peroxidases and hydrolytic enzymes that cooperate in wood decay. Analysis of the genome data will enhance our understanding of lignocellulose degradation, a pivotal process in the global carbon cycle, and provide a framework for further development of bioprocesses for biomass utilization, organopollutant degradation and fiber bleaching. This genome provides a high quality draft sequence of a basidiomycete, a major fungal phylum that includes important plant and animal pathogens.     

Biosynthesis of Hydrolytic Enzymes during Cocultivation of Macro- and Micromycetes

Effects of cocultivation of higher basidiomycetes and zygomycetes on biosynthesis of cellulases, amylases, and proteases were studied. Four optimal pairs of cultures of fungi were selected. Of these, three pairs belonged to higher fungi and one pair was constituted by fungi of distinct ecological groups, a macromycete and a micromycete. The activities of amylase and protease were 1.5–2 times higher and the activity of cellulase was lower during the growth of associations of higher fungi. The mixed association of the macromycete Schizophyllum commune and the micromycete Mucor sp. was the most active producer of hydrolytic enzymes. During the growth of this mixed association, increases of 4 and 1.5 times were observed in the activity of endoglucanase and protease, respectively, paralleled by stimulation of amylase formation.     

Endophytic fungi: expanding the arsenal of industrial enzyme producers

Endophytic fungi, mostly belonging to the Ascomycota, are found in the intercellular spaces of the aerial plant parts, particularly in leaf sheaths, sometimes even within the bark and root system without inducing any visual symptoms of their presence. These fungi appear to have a capacity to produce a wide range of enzymes and secondary metabolites exhibiting a variety of biological activities. However, they have been only barely exploited as sources of enzymes of industrial interest. This review emphasizes the suitability and possible advantages of including the endophytic fungi in the screening of new enzyme producing organisms as well as in studies aiming to optimize the production of enzymes through well-known culture processes. Apparently endophytic fungi possess the two types of extracellular enzymatic systems necessary to degrade the vegetal biomass: (1) the hydrolytic system responsible for polysaccharide degradation consisting mainly in xylanases and cellulases; and (2) the unique oxidative ligninolytic system, which degrades lignin and opens phenyl rings, comprises mainly laccases, ligninases and peroxidases. The obvious ability of endophytic fungi to degrade the complex structure of lignocellulose makes them useful in the exploration of the lignocellulosic biomass for the production of fuel ethanol and other value-added commodity chemicals. In addition to this, endophytic fungi may become new sources of industrially useful enzymes such as lipases, amylases and proteases.     

Enzymatic systems involved in decomposition reflects the ecology and taxonomy of saprotrophic fungi

One hundred and eleven strains of Basidiomycota, 39 strains of Ascomycota and 2 strains of Mucoromycotina belonging to wood decomposers that cause white-rot (WR) or brown-rot (BR), other wood associated saprotrophs (WA), litter decomposing cord-forming Basidiomycota (LDF), and saprotrophic microfungi (SA), were screened for the production of hydrolytic enzymes and laccase. The presence of enzyme-encoding genes was also analysed in the published genomes of saprotrophic fungi. Several genes, including those for acidic phosphatase, β-glucosidase and N-acetylglucosaminidase, were common in the genomes with enzyme activity widely displayed by fungi, while other enzymes, such as certain hemicellulases or laccase, were produced less frequently. Enzyme production by saprotrophic fungi was shaped by the combination of their ecophysiology and taxonomy. Basidiomycota exhibited higher activities of all enzymes, except alkaline phosphatase, α-glucosidase, N-acetylglucosaminidase, α-mannosidase and α-fucosidase, than Ascomycota. The SA and BR fungi showed distinct enzyme production patterns, while the enzyme production by WR, LDF and WA was similar. Differences among species were typically reflected in the level of enzyme activity rather than in the absence of enzymes. Enzyme screening results showed that in several cases, fungi exhibited enzyme activity without the presence of the corresponding gene and vice versa. This indicates that the use of genome-derived information for the prediction of potential enzyme production has substantial limitations and cannot replace functional screening of fungal cultures.