Basidiomycete Pleurotus Ostreatus Research Articles

The Nitrogen Content in the Fruiting Body and Mycelium of Pleurotus Ostreatus and Its Utilization as a Medium Component in Thraustochytrid Fermentation.

Following the idea of a circular bioeconomy, the use of side streams as substitutes for cultivation media (components) in bioprocesses would mean an enormous economic and ecological advantage. Costly compounds in conventional media for the production of the triterpene squalene in thraustochytrids are the main carbon source and complex nitrogen sources. Among other side streams examined, extracts from the spent mycelium of the basidiomycete Pleurotus ostreatus were best-suited to acting as alternative nitrogen sources in cultivation media for thraustochytrids. The total nitrogen (3.76 ± 0.01 and 4.24 ± 0.04%, respectively) and protein (16.47 ± 0.06 and 18.57 ± 0.18%, respectively) contents of the fruiting body and mycelium were determined. The fungal cells were hydrolyzed and extracted to generate accessible nitrogen sources. Under preferred conditions, the extracts from the fruiting body and mycelium contained 73.63 ± 1.19 and 89.93 ± 7.54 mM of free amino groups, respectively. Cultivations of Schizochytrium sp. S31 on a medium using a mycelium extract as a complex nitrogen source showed decelerated growth but a similar squalene yield (123.79 ± 14.11 mg/L after 216 h) compared to a conventional medium (111.29 ± 19.96 mg/L, although improvable by additional complex nitrogen source).

Discovery of Novel Terpenoids from the Basidiomycete Pleurotus ostreatus through Genome Mining and Coculture Optimization.

In our previous work, postredienes A-C, three unusual linear sesterterpenes with high antifungal activities, were isolated from Pleurotus ostreatus SY10 when cocultured with Trametes robiniophila SY636. However, their titers were low, and exploration of newly biosynthesized trace analogues is required. Herein, genome mining analysis predicted that 17 gene clusters are involved in terpenoid biosynthesis in P. ostreatus. Thus, coculture conditions for strains SY10 and SY636 were optimized using a single-factor test and Box-Behnken design. As a result, the titers of postredienes A-C were increased by over 2.5-fold, reaching 1.28 to 8.40 mg/L. Moreover, five new terpenoids, named postredienes D-H (1-5), were successfully isolated. Compound 1 exhibited activities against the human pathogenic fungi Candida albicans and Cryptococcus neoformans comparable to those of amphotericin B. Compound 2 represents a novel sesterterpene with a five-membered ring at C-7. The absolute configurations of 1-5 were elucidated by making the methoxyphenylacetic acid esters and acetonide derivatives, combined with ECD and NMR calculation. Two potential gene clusters and relevant biosynthetic pathways for 1-5 were subsequently proposed based on real-time reverse transcription-quantitative PCR (RT-qPCR) analysis. The current study provides new insights into the research of terpenoid biosynthesis genes in P. ostreatus and other basidiomycetes.

Effect of Different Inducer Sources on Cellulase Enzyme Production by White-Rot Basidiomycetes Pleurotus ostreatus and Phanerochaete chrysosporium under Submerged Fermentation

Cellulase enzymes attract a lot of research due to their industrial application. Diverse cellulase-producing organisms and substances that induce cellulase are highly sought after. This study aimed to evaluate the effect of different inducer sources on cellulase production by white rot fungi P. ostreatus CGMCC 3.7292 and P. chrysosporium CGMCC 3.7212 under submerged fermentation employing a completely randomized experimental design. The different inducer sources tested were nitrogen (yeast, potassium nitrate, sodium nitrate, ammonium sulphate, aqueous ammonia and urea), carbon (malt extract, glucose, fructose, carboxymethylcellulose, starch and xylose) and agro-biomass (stevia straw, wheat straw, oat straw, alfalfa straw, corn cobs and corn stover). These inducer sources strongly impacted enzyme activities by P. ostreatus CGMCC 3.7292 and P. chrysosporium CGMCC 3.7212. The suitable nitrogen and carbon inducer sources for cellulase activity by P. ostreatus and P. chrysosporium were yeast (1.354 U/mL and 1.154 U/mL) and carboxymethylcellulose (0.976 U/mL and 0.776 U/mL) while the suitable agro-biomass were wheat straw (6.880 U/mL) and corn stover (6.525 U/mL), respectively. The least inducer sources in terms of nitrogen, carbon and agro-biomass for cellulase activity by P. ostreatus and P. chrysosporium were urea (0.213 U/mL and 0.081 U/mL), glucose (0.042 U/mL and 0.035), xylose (0.042 U/mL and 0.035 U/mL) and stevia straw (1.555 U/mL and 0.960 U/mL). In submerged fermentation, the cellulase enzyme activity of P. ostreatus in response to various inducer sources was relatively higher than P. chrysosporium.

Degradation of Polycyclic Aromatic Hydrocarbons by Co-Culture of Pleurotus ostreatus Florida and Azospirillum brasilense

Bacterial-fungal interactions are important in the functioning of natural ecosystems. We examined possible synergistic or antagonistic effects during the degradation of polycyclic aromatic hydrocarbons (PAHs) by a fungal–bacterial co-culture. Bacteria and fungi were grown in a liquid nutrient medium supplemented with PAH substrates. The degradation of PAHs and the identification of metabolites were checked by HPLC. Enzyme activities were spectrophotometrically measured with test substrates. Compared to monocultures, the co-culture yielded higher mycelium dry weights and higher numbers of bacterial colony-forming units (CFUs). Both organisms and their co-culture transformed three- and four-ring PAHs into the corresponding quinones. The degradation of PAHs was accompanied by the production of fungal extracellular laccase and versatile peroxidase, whose activities were higher in the co-culture than they were in the monocultures. The presence of exogenous indole-3-acetic acid (IAA) boosted PAH degradation and enzyme production. The xylotrophic basidiomycete Pleurotus ostreatus Florida and the plant-growth-promoting rhizobacterium Azospirillum brasilense exerted a positive mutual effect, including increases in mycelium dry weight, number of CFUs, degradation of PAHs, and production of fungal extracellular enzymes. IAA may be a factor in the interactions of P. ostreatus Florida with A. brasilense.

Functional Roles of LaeA-like Genes in Fungal Growth, Cellulase Activity, and Secondary Metabolism in Pleurotus ostreatus.

The global regulator LaeA plays crucial roles in morphological development and secondary metabolite biosynthesis in filamentous fungi. However, the functions of LaeA in basidiomycetes are less reported. The basidiomycete Pleurotus ostreatus is a well-known fungus used both in medicine and as food that produces polysaccharides and cellulolytic enzymes. In this study, we characterized three LaeA homologs (PoLaeA1, PoLaeA2, and PoLaeA3) in P. ostreatus. PoLaeA1 showed different expression patterns than PoLaeA2 and PoLaeA3 during different developmental stages. Silencing PoLaeA1 decreased the intracellular polysaccharide (IPS) content by approximately 28–30% and reduced intracellular ROS levels compared with those of the WT strain. However, silencing PoLaeA2 and PoLaeA3 decreased cellulase activity by 31–34% and 35–40%, respectively, and reduced the cytosolic Ca2+ content, compared with those of the WT strain. Further analysis showed that PoLaeA1 regulated IPS biosynthesis through intracellular ROS levels, whereas PoLaeA2 and PoLaeA3 regulated cellulase activity through intracellular Ca2+ signaling. Our results provide new insights into the regulation of polysaccharide biosynthesis and cellulase production in filamentous fungi.

Widespread Ability of Ligninolytic Fungi to Degrade Hazardous Organic Pollutants as the Basis for the Self-Purification Ability of Natural Ecosystems and for Mycoremediation Technologies

The ability of sixteen wood- and soil-inhabiting basidiomycete strains and four ascomycete strains to degrade the most hazardous, widespread, and persistent pollutants (polycyclic aromatic hydrocarbons, oxyethylated nonylphenol, alkylphenol, anthraquinone-type synthetic dyes, and oil) was found. The disappearance of the pollutants, their main metabolites, and some adaptive properties (activities of ligninolytic enzymes, the production of emulsifying compounds and exopolysaccharides) were evaluated. The toxicity of polycyclic aromatic hydrocarbons decreased during degradation. New data were obtained regarding (1) the dependence of the completeness of polycyclic aromatic hydrocarbon degradation on the composition of the ligninolytic enzyme complex; (2) the degradation of neonol AF9-12 by higher fungi (different accessibilities of the oxyethyl chain and the aromatic ring of the molecules to different fungal genera); and (3) the production of an emulsifying agent in response to the presence in the cultivation medium of hydrophobic pollutants as the common property of wood- and soil-inhabiting basidiomycetes and ascomycetes. Promise for use in mycoremediation was shown in the wood-inhabiting basidiomycetes Pleurotus ostreatus f. Florida, Schizophyllum commune, Trametes versicolor MUT 3403, and Trametes versicolor DSM11372; the litter-decomposing basidiomycete Stropharia rugosoannulata; and the ascomycete Cladosporium herbarum. These fungi degrade a wide range of pollutants without accumulation of toxic metabolites and produce ligninolytic enzymes and emulsifying compounds.

Production of stable emulsions using β‐glucans extracted from Pleurotus ostreatus to encapsulate oxidizable compounds

AbstractThe basidiomycete Pleurotus ostreatus is a sustainable food source known to be rich in β‐glucans, functional compounds in mushrooms recognized for their numerous health, nutraceutical, and physicochemical properties. The aim of the present study was to extract β‐glucans from P. ostreatus powder and to study the feasibility of using the extract to prepare oil‐in‐water emulsions formulated for the encapsulation of lipophilic antioxidants by spray drying, for example, with an oil droplet size about 2 μm, a dry matter content of 40% w/w, and with a physical stability of at least 2 hrs. Hot‐water soluble β‐glucans fraction was extracted from P. ostreatus powder following a method adapted from literature. Emulsions were prepared by rotor‐stator homogenization using maltodextrin as wall material, β‐glucans extract as emulsifier, and commercial sunflower oil as a model for lipophilic active compounds. The emulsion stability was estimated from the evolution over time of the oil droplet size distribution measured by laser light diffraction and experimental data were investigated using one‐way ANOVA. Extracts containing up to 27% w/w water‐soluble β‐glucans were produced. Physically stable mono‐modal emulsions were obtained when maltodextrin/β‐glucans weight ratio was lower than about 500 to avoid depletion and β‐glucans/oil weight ratio was more than 0.014 to stabilize oil droplets. The usage of emulsifiers made‐up by polymeric constituents with inherent bioactivity, such as mushroom β‐glucans, could be a sustainable and healthy alternative to common emulsifiers.Practical applicationsA great deal of interest has been expressed in a unique dietary fiber, named β‐glucans, from the basidiomycete Pleurotus ostreatus, a mushroom which can grow efficiently on various clean by‐products of food processing, thus representing a sustainable food source. β‐Glucans' functionality is associated with their many healthy and physicochemical properties, for instance emulsifying properties. The usage of emulsifiers made‐up by polymeric constituents with inherent bioactivity, such as mushroom β‐glucans, could be a sustainable and healthy alternative to common emulsifiers.

Energy analysis of sugarcane bagasse after enzymatic catalysis process

Nowadays, countries produce approximately 1700 million tons of sugarcane per year. However, many countries now find it difficult to produce sufficient energy, generating a relevant socioeconomic impact. The objective of this work is to study the potential energy of sugarcane bagasse after decomposing fungus process. This fungus, the basidiomycete Pleurotus ostreatus, acts in the bioconversion of lignocellulosic residues, through the decomposition of the solid substrate in a process of enzymatic catalysis. The activation energy, Ea, varied from 12 to 145 kJ/mol, and the calorific value and the fixed carbon obtained were 21 MJ/kg and 82.4%, respectively, values considered acceptable for a solid biofuel. In the results of the enzymatic analysis performed through the laccase secretion and manganese peroxidase (MnP), measured by the Fourier transform infrared spectroscopy technique (FT-IR), it is possible to identify when there was degradation of the lignin due to the methoxy group (CH3) elongation. Scanning electron microscope (SEM) analysis showed partially fragmented structures, which occurred due to catalysis and pyrolysis processes, respectively.

The effect of metal-containing biocomposites of fungal origin on potato plants in vitro

The effect of metal (II)-containing composites based on extracellular metabolites of basidiomycetes Pleurotus ostreatus, Ganoderma lucidum, Grifola umbellata and Laetiporus sulphureus on the viability and response of potato plants in vitro has been investigated. The Lukyanovsky variety of potato, which is susceptible to ring rot, caused by the bacterium Clavibacter sepedonicus, was studied. The parameters investigated included biofilm formation by Clavibacter sepedonicus, various morphometric parameters of plants and the phytotoxicity of substances of fungal origin. The greatest anti-biofilm-forming effect was observed in metal-containing biocomposites based on G. lucidum; Fe- and Co-containing biopreparations inhibited the formation of Clavibacter sepedonicus biofilms by 40–50%. The plant height was adversely affected by composites, in the absence of metal (II), derived from L. sulphureus and P. ostreatus, as well as by a Co-containing composite derived from P. ostreatus. The decrease in plant growth, in comparison with the control, can be associated with the pronounced antibiotic properties of these basidiomycetes and cobalt. The remaining biocomposites studied did not have an adverse effect on the growth of potatoes in vitro. A number of morphometric parameters (length of internodes, number of leaves) remained virtually unchanged when exposed to biocomposites of fungal origin. In contrast to the vegetative part of plants, the biomass and length of the roots increased by 10–20% under the influence of biocomposites. Copper-containing composites derived from G. lucidum had no phytotoxic effect on plants and enhanced potato resistance to Clavibacter sepedonicus. The beneficial properties of biocomosites may be judged by the degree of stimulation of the physiological processes underlying the formation of the underground part of the plants, which is a prerequisite for increasing yields. The biocomposites are environmentally friendly because of their natural origin and being effective at very low doses. The results obtained using metal-containing biocomposites derived from G. lucidum and Gr. umbellata demonstrate the safety and possible improvement in health of potato plants by using biocomposites derived from cultures of higher fungi.

Glucose counteracts wood-dependent induction of lignocellulolytic enzyme secretion in monokaryon and dikaryon submerged cultures of the white-rot basidiomycete Pleurotus ostreatus

The secretome complexity and lignocellulose degrading capacity of Pleurotus ostreatus monokaryons mkPC9 and mkPC15 and mated dikaryon dkN001 were studied in submerged liquid cultures containing wood, glucose, and wood plus glucose as carbon sources. The study revealed that this white-rot basidiomycete attacks all the components of the plant cell wall. P. ostreatus secretes a variety of glycoside hydrolases, carbohydrate esterases, and polysaccharide lyases, especially when wood is the only carbon source. The presence of wood increased the secretome complexity, whereas glucose diminished the secretion of enzymes involved in cellulose, hemicellulose and pectin degradation. In contrast, the presence of glucose did not influence the secretion of redox enzymes or proteases, which shows the specificity of glucose on the secretion of cellulolytic enzymes. The comparison of the secretomes of monokaryons and dikaryons reveals that secretome complexity is unrelated to the nuclear composition of the strain.

Application of Pleurotus ostreatus β-glucans for oil–in–water emulsions encapsulation in powder

In this study, sunflower oil containing α-tocopherol (model for oxidizable lipophilic compounds) was encapsulated using a wall material made of maltodextrin and two different extracts rich in β-glucans and proteins, obtained from the basidiomycete Pleurotus ostreatus. A mixture of maltodextrin and acacia gum was used as control wall material. The aim was to assess P. ostreatus extracts as a sustainable and healthy alternative to common emulsifiers. After the evaluation of the emulsion stability, four powders were produced by spray drying and then analysed for the particle morphology and oil droplets distribution within the solid matrix (by scanning electron microscopy), the emulsion microstructure after their dissolution in water (by laser light diffraction) and the encapsulation efficiency (by solvent extraction of surface oil). In addition, powders were analysed in terms of oil protection against oxidation by spectroscopic determination of conjugated dienes and HPLC evaluation of vitamin E. Results showed that P. ostreatus extracts provided suitable emulsifying properties both in the liquid and in the spray dried emulsions. Moreover, it allowed excellent protection of vitamin E and polyunsaturated fatty acids against oxidation. Hence, P. ostreatus extracts can be exploited as innovative emulsifier to provide physical protection to functional, oxygen sensitive lipophilic ingredients by microencapsulation.

Antiradical and antidiabetic activity of Pleurotus ostreatus extracts

Mushrooms have been an important part of the diet for centuries. There are more than 2,000 different species of mushrooms found in the nature, but only 25 species are generally used in the human diet, and even fewer are grown commercially. One of the more popular mushrooms for cultivation and culinary use are mushrooms of the genus Pleurotus. The objectives of this study were: (i) preparation of isolates from the basidiomycetes Pleurotus ostreatus by sequential extraction with cold and hot water, extraction with hot water and extraction with hot alkali solution for determination of antiradical activity and (ii) preparation of water and 70% ethanol extracts for determination of antidiabetic activity. The antiradical activity of the extracts was determined using spectrophotometric method based on inhibition of free radical 2,2- diphenyl-1-picrylhydrazyl. Antidiabetic activity was determined using α- amylase inhibition assay with DNSA reagent and spectrophotometric determination. As a result of the work carried out, it was found that the preparations obtained by extraction exhibit antiradical activity. Regarding antidiabetic activity, all tested extracts showed considerable antidiabetic activity. Moreover, water extracts were inhibiting α-amylase significantly more than ethanol extracts at higher concentrations of dry extract (50 and 100 mg/mL), whereas at lower concentrations (1 and 10 mg/mL) of ethanol extracts were more efficient than water ones.

Fortification of dairy product Narine with polysaccharides from oyster mushroom

Mushrooms are rich in biologically active substances. Basidiomycete oyster mushroom or Pleurotus ostreatus is not only widely used in food, but also have radioprotection, preventive effect on recurrent respiratory tract infections, hypocholesterolic action, antitumor effect, strong immunostimulatory activity and other beneficial medical effects. It is considered that β-glucans play significant role in the biological activities of the oyster mushroom. The aim of this research was to study the effect of the addition of various β-glucan containing preparations obtained from the submerged biomass of P. ostreatus on the process of milk fermentation and properties of the finished product. Preparations of β-glucans obtained from submerged cultured P. ostreatus biomass were obtained using different extraction protocols. The content of β-glucans in preparations was determined using Assay Kit (Megazyme). Preparations were added to milk in different concentrations before the introduction of starter culture. A starter containing Lactobacillus acidophilus n.v. Er 317/402 strain «Narine» was used for fermentation. The titratable acidity was controlled during the fermentation. Physicochemical, structural-mechanical properties (water holding capacity) of the obtained samples were studied. The sensory evaluation of the products was evaluated. Results show that the addition of preparations increase the glycolytic activity. Structural-mechanical properties of yogurt samples fortified by β-glucan-containing preparations depend on the preparation and its concentration. Sensory evaluation showed that experimental samples differed not only from the control sample but also from each other.

Pleurotus ostreatus as a Biocatalyst to Obtain Bicyclic Hydroxylactones with Three or Four Methyl Groups

The purpose of the study was to explore microbial transformations in cultures of basidiomycetes Pleurotus ostreatus, two bicyclic unsaturated lactones occurring in the form of two diastereoisomers. Some of these strains were able to transform unsaturated lactones into four known and three new derivatives. The structures of all lactones were established on the basis of spectroscopic data. Both substrates and products caused a complete inhibition of growth of A. alternata and F. linii strains.

Differential regulation of Pleurotus ostreatus dye peroxidases gene expression in response to dyes and potential application of recombinant Pleos-DyP1 in decolorization.

Dye-decolorizing peroxidase (DyP) from the white rot basidiomycete Pleurotus ostreatus is a heme peroxidase able to oxidize diverse substrates, including recalcitrant phenols and dyes. This study analyzed the effect of chemical dyes on P. ostreatus growth, DyP activity and the expression of four Pleos-dyp genes during the time-course of Pleurotus ostreatus cultures containing either Acetyl Yellow G (AYG), Remazol Brilliant Blue R (RBBR) or Acid Blue 129 (AB129) dyes. Additionally, Pleos DyP1 was heterologously expressed in the filamentous fungus Trichoderma atroviride in order to explore the potential of a secreted recombinant enzyme for decolorizing different dyes in cultures and plate assays. The addition of dyes had an induction effect on the enzymatic activity, with the fermentations undertaken using RBBR and AYG dyes presenting the highest total DyP activity. DyP gene expression profiles displayed up/down regulation during the culture of three Pleos-dyp genes (Pleos-dyp1, Pleos-dyp2 and Pleos-dyp4), while Pleos-dyp3 transcript was not detected under any of the culture conditions studied. A 14-fold relative induction level (log2) increase for Pleos-dyp2 and Pleos-dyp4 in AB129 and AYG, respectively, was also found. The presence of AB129 resulted in the highest Pleos-dyp1 gene induction and repression level, corresponding to 11.83 and -14.6-fold relative expression and repression levels, respectively. The lowest expression level of all genes was observed in RBBR, a response which is associated with the growth phase. The filamentous fungus Trichoderma atroviride was successfully transformed for the heterologous expression of Pleos-dyp1. The modified strains (TaDyP) were able to decolorize mono-azo, di-azo, anthraquinone and anthracenedione dyes with extracellular DyP1 activity found in the culture supernatant. After 96 h of culture, the recombinant TaDyP strains were able to degrade (decolorize) 77 and 34% of 0.05mM AB129 and 0.25mM AYG, respectively.

Transposon-associated epigenetic silencing during Pleurotus ostreatus life cycle.

Transposable elements constitute an important fraction of eukaryotic genomes. Given their mutagenic potential, host-genomes have evolved epigenetic defense mechanisms to limit their expansion. In fungi, epigenetic modifications have been widely studied in ascomycetes, although we lack a global picture of the epigenetic landscape in basidiomycetes. In this study, we analysed the genome-wide epigenetic and transcriptional patterns of the white-rot basidiomycete Pleurotus ostreatus throughout its life cycle. Our results performed by using high-throughput sequencing analyses revealed that strain-specific DNA methylation profiles are primarily involved in the repression of transposon activity and suggest that 21 nt small RNAs play a key role in transposon silencing. Furthermore, we provide evidence that transposon-associated DNA methylation, but not sRNA production, is directly involved in the silencing of genes surrounded by transposons. Remarkably, we found that nucleus-specific methylation levels varied in dikaryotic strains sharing identical genetic complement but different subculture conditions. Finally, we identified key genes activated in the fruiting process through the comparative analysis of transcriptomes. This study provides an integrated picture of epigenetic defense mechanisms leading to the transcriptional silencing of transposons and surrounding genes in basidiomycetes. Moreover, our findings suggest that transcriptional but not methylation reprogramming triggers fruitbody development in P. ostreatus.

Somatic transposition and meiotically driven elimination of an active helitron family in Pleurotus ostreatus.

Helitrons constitute a superfamily of DNA transposons that were discovered in silico and are widespread in most eukaryotic genomes. They are postulated to mobilize through a “rolling-circle” mechanism, but the experimental evidence of their transposition has been described only recently. Here, we present the inheritance patterns of HELPO1 and HELPO2 helitron families in meiotically derived progeny of the basidiomycete Pleurotus ostreatus. We found distorted segregation patterns of HELPO2 helitrons that led to a strong under-representation of these elements in the progeny. Further investigation of HELPO2 flanking sites showed that gene conversion may contribute to the elimination of such repetitive elements in meiosis, favouring the presence of HELPO2 vacant loci. In addition, the analysis of HELPO2 content in a reconstructed pedigree of subclones maintained under different culture conditions revealed an event of helitron somatic transposition. Additional analyses of genome and transcriptome data indicated that P. ostreatus carries active RNAi machinery that could be involved in the control of transposable element proliferation. Our results provide the first evidence of helitron mobilization in the fungal kingdom and highlight the interaction between genome defence mechanisms and invasive DNA.

Degradation of fluorene and fluoranthene by the basidiomycete Pleurotus ostreatus

The dependence of the degree of fluorene and fluoranthene degradation by the fungus Pleurotus ostreatus D1 on the culture medium composition has been studied. Polycyclic aromatic hydrocarbons (PAHs) have been transformed in Kirk’s medium (under conditions of laccase production) with the formation of a quinone metabolite and 9-fluorenone upon the use of fluoranthene and fluorene as substrates, respectively. More complete degradation with the formation of an intermediate metabolite, phthalic acid that has undergone subsequent utilization, has occurred in basidiomycete-rich medium (under the production of both laccase and versatile peroxidase). The formation of phthalic acid as a metabolite of fluoranthene degradation by lignolytic fungi has been revealed for the first time. The data allow the supposition that both extracellular laccase and laccase on the mycelium surface can participate in the initial stages of PAH metabolism, while versatile peroxidase is necessary for the oxidation of the formed metabolites. A scheme of fluorene metabolism by Pleurotus ostreatus D1 is suggested.

Validation of Reference Genes for Transcriptional Analyses in Pleurotus ostreatus by Using Reverse Transcription-Quantitative PCR.

Recently, the lignin-degrading basidiomycete Pleurotus ostreatus has become a widely used model organism for fungal genomic and transcriptomic analyses. The increasing interest in this species has led to an increasing number of studies analyzing the transcriptional regulation of multigene families that encode extracellular enzymes. Reverse transcription (RT) followed by real-time PCR is the most suitable technique for analyzing the expression of gene sets under multiple culture conditions. In this work, we tested the suitability of 13 candidate genes for their use as reference genes in P. ostreatus time course cultures for enzyme production. We applied three different statistical algorithms and obtained a combination of stable reference genes for optimal normalization of RT-quantitative PCR assays. This reference index can be used for future transcriptomic analyses and validation of transcriptome sequencing or microarray data. Moreover, we analyzed the expression patterns of a laccase and a manganese peroxidase (lacc10 and mnp3, respectively) in lignocellulose and glucose-based media using submerged, semisolid, and solid-state fermentation. By testing different normalization strategies, we demonstrate that the use of nonvalidated reference genes as internal controls leads to biased results and misinterpretations of the biological responses underlying expression changes.

Decolourization of remazol brilliant blue R by enzymatic extract and submerged cultures of a newly isolated Pleurotus ostreatus MR3

A local white-rot fungus basidiomycete Pleurotus ostreatus MR3 was isolated from MacRitchie Reservoir Park, Singapore. Among all the ligninolytic activities, laccase was the only enzyme detected in the supernatant when the fungus was grown in liquid culture. This newly isolated white rot fungus was able to completely decolourise remazol brilliant blue R (RBBR) in-vivo on agar plates within five days and in the liquid culture (in the presence of inducers) within three days. The addition of inducers was able to enhance laccase production and therefore enhanced in-vivo RBBR decolourisation. Veratryl alcohol was shown to be the best inducer for laccase production with the maximum laccase activity reaching about 5.99 U/mL. Cu 2+ also had a positive effect on laccase production, the laccase activity being enhanced to 5.24 U/mL. In-vitro RBBR decolourisation using the laccase from P. ostreatus MR3 was much comparable to that using the commercial laccase from Trameters versicolor . Keywords: Dyes, remazol brilliant blue R, Pleurotus ostreatus MR3, decolourisation, inducers, laccase activity African Journal of Biotechnology Vol. 12(39), pp. 5778-5783