Ensayo de citotoxicidad de extractos de Pleurotusostreatus en diferentes líneas celulares para aplicaciones inmunonutricionales

Introducción: Los bioderivados propuestos como candidatos a ingredientes alimentarios suelen requerir ciertas evaluaciones para las aplicaciones inmunonutricionales Los hongos comestibles-medicinales son un surtidor de compuestos con estas potencialidades. Entre ellos, las setas Pleurotus ostreatus contienen metabolitos bioactivos, con importantes usos en la industria alimenticia y en la práctica terapéutica de la industria médico-farmacéutica. Los ensayos de citotoxicidad in vitro constituyen métodos valiosos para evaluarproductos de origen natural, como los extractos fúngicos. Objetivo: Evaluar la citotoxicidad de dos extractos obtenidos de la seta Pleurotus ostreatus en diferentes líneas celulares. Método: Se obtuvieron extractos hidrosolubles a partir del micelio y de los cuerpos fructíferos de Pleurotus ostreatus en laboratorios del Centro de Estudios de Biotecnología Industrial de la Universidad de Oriente. Se evaluó la citotoxicidad de los bioproductos por el ensayo de reducción del colorante resazurina sobre tres líneas celulares en el Laboratorio de Microbiología, Parasitología e Higiene (LMPH) de la Universidad de Amberes, Bélgica. Se utilizaron células no adherentes THP-1 (pre-monocitos de leucemia humana), células adherentes Caco-2 (epitelio de adenocarcinoma de colon humano) y células adherentes RAW 264.7 (macrófagos murinos). Resultados: Los extractos de Pleurotus ostreatus no resultaron citotóxicos para ninguna de las líneas celulares estudiadas humanas o murina, ya que no ocasionaron daños sobre la viabilidad de las célulasepiteliales del sistema gastrointestinal, nisobrelas células del sistema inmune empleadas. Conclusiones: Este resultado demuestra que ambos bioderivados fúngicos pueden ser aplicados con seguridad en estudios inmunonutricionales.

Introduction: Bioderivatives proposed as candidates for food ingredients usually require certain evaluations for immunonutritional applications. Edible-medicinal mushrooms are a source of compounds with these potentials. Among them, Pleurotus ostreatus mushrooms contain bioactive metabolites, with important uses in the food industry and in the therapeutic practice of the medical-pharmaceutical industry. In vitro cytotoxicity assays are valuable methods to evaluate products of natural origin, such as fungal extracts. Objective: To evaluate the cytotoxicity of two extracts obtained from the Pleurotus ostreatus mushroom in different cell lines. Method: Water-soluble extracts were obtained from the mycelium and fruiting bodies of Pleurotus ostreatus in laboratories of the Center for Industrial Biotechnology Studies of the Universidad de Oriente. The cytotoxicity of the bioproducts was evaluated by the resazurin dye reduction assay on three cell lines at the Laboratory of Microbiology, Parasitology and Hygiene (LMPH) of the University of Antwerp, Belgium. Non-adherent THP-1 cells (human leukemia pre-monocytes), Caco-2 adherent cells (human colon adenocarcinoma epithelium) and RAW 264.7 adherent cells (murine macrophages) were used. Results: Pleurotus ostreatus extracts were not cytotoxic for any of the human or murine cell lines studied, since they did not cause damage to the viability of the epithelial cells of the gastrointestinal system, nor to the immune system cells used. Conclusions: This result demonstrates that both fungal bioderivatives can be safely applied in immunonutritional studies.

Introdução: Bioderivados propostos como candidatos a ingredientes alimentícios geralmente requerem determinadas avaliações para aplicações imunonutricionais. Pleurotus ostreatus contêm metabólitos bioativos, com importantes utilizações na indústria alimentícia e na prática terapêutica da indústria médico-farmacêutica. Ensaios de citotoxicidade in vitro são métodos valiosos para avaliar produtos de origem natural, como extratos de fungos. Objetivo: Avaliar a citotoxicidade de dois extratos obtidos do cogumelo Pleurotus ostreatus em diferentes linhagens celulares. Método: Extratos hidrossolúveis foram obtidos do micélio e dos corpos frutíferos de Pleurotus ostreatus nos laboratórios do Centro de Estudos de Biotecnologia Industrial da Universidade de Oriente. A citotoxicidade dos bioprodutos foi avaliada pelo ensaio de redução do corante resazurina em três linhagens celulares no Laboratório de Microbiologia, Parasitologia e Higiene (LMPH) da Universidade de Antuérpia, Bélgica. Foram utilizadas células THP-1 não aderentes (pré-monócitos de leucemia humana), células aderentes Caco-2 (epitélio de adenocarcinoma do cólon humano) e células aderentes RAW 264.7 (macrófagos murinos). Resultados: Os extratos de Pleurotus ostreatus não foram citotóxicos para nenhuma das linhagens celulares humanas ou murinas estudadas, pois não causaram danos à viabilidade das células epiteliais do sistema gastrointestinal, nem às células do sistema imunológico utilizadas. Conclusões: Este resultado demonstra que ambos os bioderivados fúngicos podem ser aplicados com segurança em estudos imunonutricionais.

In vitro ?-amylase and ?-glucosidase inhibitory potential of Pleurotus ostreatus cv. Florida extract

Pleurotus ostreatus cv. Florida is one of the widely used edible mushroom. The polysaccharides from this mushrooms have been studied for antidiabetic potential; however, no efforts have been made to explore the potential of this mushroom to influence carbohydrate metabolizing enzymes viz. ?-amylase and ?-glucosidase. The present work was undertaken to investigate the inhibitory potential of Pleurotus ostreatus cv. Florida on enzymes ?-amylase and ?-glucosidase. Several concentrations of extracts were used to study inhibition of enzymatic activity of ?-amylase and ?-glucosidase. A dose dependent inhibitory effect on enzymes was observed. The current study, for the first time, uncovered ?-amylase and ?-glucosidase inhibitory potential of Pleurotus ostreatus cv. Florida. The study could be helpful to isolate and characterize compounds responsible for it.

Evaluación in vivo de la pirogenicidad de bioproductos fúngicos con potencial prebiótico / In vivo pyrogenicity evaluation of fungal bioproducts with prebiotic potential / Avaliação in vivo da pirogenicidade de bioprodutos fúngicos com potencial prebiótico

RESUMEN Introducción: Los productos naturales con actividad farmacológica requieren de evaluaciones preclínicas que justifiquen su empleo sobre una base científica. El ensayo de pirógenos es una prueba dentro de la Farmacología de Seguridad que se realiza para determinar la presencia de endotoxinas y constituye un método valioso, para demostrar la seguridad de bioderivados con potencial prebiótico en el campo de la inmunonutrición. Objetivo: Evaluar la pirogenicidad de bioproductos fúngicos de Pleurotus ostreatus (extractos acuosos del micelio y cuerpos fructíferos) y un biopreparado de levadura Kluyveromyces marxianus, empleando el ensayo de pirógenos en conejos Nueva Zelanda. Método: Se ensayaron concentraciones de 1,0 y 10,0 mg/mL de cada muestra por vía endovenosa en dosis de 0,5 y 5,0 mg/kg de peso. El diseño experimental cumplió las buenas prácticas de laboratorio según lo establecido por el International Council for Laboratory Animals Science y se realizó de acuerdo a los procedimientos normalizados de trabajo del Centro de Toxicología y Biomedicina, Santiago de Cuba. Resultados: Los extractos de Pleurotus ostreatus y el biopreparado de levadura (0,5 mg/kg) no mostraron signos de pirogenicidad. En los resultados del biopreparado (5,0 mg/kg), los valores de temperatura caen en un rango de incertidumbre, según la Farmacopea de Estados Unidos (USP) y se sugirió repetir el estudio. Conclusiones: Los extractos de Pleurotus ostreatus y el biopreparado de Kluyveromyces marxianus (0,5 mg/kg) no indujeron un aumento de temperatura significativo en los animales, lo cual sugiere que en estos bioproductos no existen niveles de endotoxinas que puedan provocar pirogenicidad.

ABSTRACT Introduction: Natural products with pharmacological activity require preclinical evaluations to justify their uses scientifically. The pyrogen assay is a safety pharmacology test performed to determine the presence of endotoxins and it is a valuable method to demonstrate the bio-derivative products safety and their prebiotic potential in the field of immunonutrition. Objective: To evaluate the pyrogenicity of fungal bioproducts from Pleurotus ostreatus (aqueous extracts from mycelium and fruiting bodies) and a biopreparation from Kluyveromyces marxianus yeast, using a pyrogen assay in New Zealand rabbits. Method: Concentrations of 1.0 and 10.0 mg/mL of each sample were tested intravenously at doses of 0.5 and 5.0 mg/kg body weight. The experimental design complied with good laboratory practices as established by the International Council for Laboratory Animal Science and was carried out according to the standard work procedures of the Centro de Toxicología y Biomedicina, Santiago de Cuba. Results: Pleurotus ostreatus extracts and the yeast biopreparation (0.5 mg/kg) showed no signs of pyrogenicity. In the biopreparation results (5.0 mg/kg), temperature values fall in the uncertainty range according to the United States Pharmacopoeia (USP), and therefore it was suggested to repeat the study. Conclusions: Pleurotus ostreatus extracts and Kluyveromyces marxianus biopreparation (0.5 mg/kg) did not induce a significant temperature increase in the animals, thereby suggesting that there are no endotoxin levels in such bioproducts that could cause pyrogenicity.

RESUMO Introdução: Produtos naturais com atividade farmacológica requerem avaliações pré-clínicas que justifiquem seu uso em bases científicas. O ensaio de pirogênio é um teste dentro da Farmacologia de Segurança que é realizado para determinar a presença de endotoxinas e é um método valioso para demonstrar a segurança de bioderivados com potencial prebiótico no campo da imunonutrição. Objetivo: Avaliar a pirogenicidade de bioprodutos fúngicos de Pleurotus ostreatus (extratos aquosos do micélio e corpos de frutificação) e de uma biopreparação da levedura Kluyveromyces marxianus, utilizando o ensaio pirogênico de coelho da Nova Zelândia. Método: Concentrações de 1,0 e 10,0 mg/mL de cada amostra foram testadas por via intravenosa nas doses de 0,5 e 5,0 mg/kg de peso. O desenho experimental obedeceu às boas práticas laboratoriais estabelecidas pelo Conselho Internacional para a Ciência dos Animais de Laboratório e foi realizado de acordo com os procedimentos de trabalho padrão do Centro de Toxicologia e Biomedicina, Santiago de Cuba. Resultados: Os extratos de Pleurotus ostreatus e a biopreparação de leveduras (0,5 mg/kg) não apresentaram sinais de pirogenicidade. Nos resultados da biopreparação (5,0 mg/kg), os valores de temperatura estão dentro de uma faixa de incerteza, segundo a Farmacopeia dos Estados Unidos (USP) e foi sugerido repetir o estudo. Conclusões: Os extratos de Pleurotus ostreatus e a biopreparação de Kluyveromyces marxianus (0,5 mg/kg) não induziram um aumento significativo da temperatura nos animais, o que sugere que não há níveis de endotoxinas nesses bioprodutos que possam causar pirogenicidade.

Evaluación in silico del efecto de compuestos fenólicos de Pleurotus ostreatus sobre la enzima 5-lipoxigenasa (5- LOX) / In silico evaluation of the effect of Pleurotus ostreatus phenolic compounds on the enzyme 5-lipoxygenase (5-LOX)

Introducción: Los hongos comestibles, en particular Pleurotus ostreatus, representan una importante fuente de metabolitos bioactivos con propiedades inmunomoduladoras, antioxidantes y antiinflamatorias. Trabajos recientes han demostrado que extractos y compuestos purificados a partir de esta seta, entre ellos, la fracción rica en fenoles, inhiben el factor nuclear kappa B(NF-κB), la cicloxigenasa (COX) y modulan cascadas de señalización relacionadas con el balance redox. De acuerdo con estos antecedentes, dichos compuestos podrían actuar, además, como inhibidores de la enzima 5- lipoxigenasa (5-LOX). Objetivo: Evaluar el efecto in silico de trece compuestos fenólicos presentes en la especie Pleurotus ostreatus sobre la enzima 5-LOX, al utilizar como compuesto de referencia la mangiferina. Métodos: El acoplamiento se llevó a cabo a través del programa AutoDock 4.2 (http://autodock.scripps.edu) y la estructura de 5 LOX se obtuvo con la base de datos de proteínas, PDB (www.wwpdb.org). Se estimaron la energía libre (ΔG), la constante de disociación (Ki) y la eficiencia de ligando (LE). Se obtuvieron los parámetros de similitud a un fármaco y los relacionados con la absorción, distribución, metabolismo, excreción y toxicidad (ADME/T) de los mejores modelos de acoplamiento. Resultados: Los mejores indicadores de ΔG y Ki, correspondieron a los ácidos homogentísico, clorogénico y gentísico, con valores de ΔG (-11,81; -12,28 y -11,67 kcal/moL) y Ki (2,19 10-9; 9,99 10-10, 2,79 10-9 M), respectivamente. La eficiencia de ligando alcanzó valores adecuados para estos tres compuestos fenólicos. El modelo de acoplamiento del ácido homogentísico mostró los mejores resultados en cuanto a la similitud a un fármaco y pruebas ADME/T. Conclusiones: El estudio in silico reveló las potencialidades de la fracción rica en fenoles de P. ostreatus, y en particular, del ácido homogentísico como inhibidor de la enzima 5 -LOX, y justifica el desarrollo de ensayos confirmativos in vitro/ in vivo que corroboren sus efectos antioxidantes y antinflamatorios(AU)

Introduction: Edible mushrooms, Pleurotus ostreatus in particular, are an important source of bioactive metabolites with immunomodulatory, antioxidant and anti-inflammatory properties. Recent studies have shown that extracts and compounds purified from this mushroom, among them the phenol-rich fraction, inhibit nuclear factor kappa B (NF-κB), cyclooxygenase (COX), and modulate signaling cascades related to redox balance. According to these antecedents, such compounds could also act as inhibitors of the enzyme 5-lipoxygenase (5-LOX). Objective: Evaluate the in silico effect of 13 phenolic compounds present in the species Pleurotus ostreatus on the enzyme 5-LOX using mangiferin as reference compound. Methods: Docking was carried out with the software AutoDock 4.2 (http://autodock.scripps.edu) and the 5-LOX structure was obtained with the protein database PDB (www.wwpdb.org). Estimation was performed of free energy (ΔG), dissociation constant (Kd) and ligand efficiency (LE). Drug-likeness parameters were obtained, as well as those related to absorption, distribution, metabolism, excretion and toxicity (ADMET) of the best docking models. Results: The best ΔG and Kd indicators were homogentisic, chlorogenic and gentisic acids, with ΔG and Kd values of -11.81, -12.28, -11.67 kcal/mol, and 2.19 10-9, 9.99 10-10, 2.79 10-9 M, respectively. Ligand efficiency achieved adequate values for these three phenolic compounds. The docking model for homogentisic acid showed the best results in terms of drug likeness and ADMET tests. Conclusions: The in silico study revealed the potential of the phenol-rich fraction of P. ostreatus, homogentisic acid in particular, as an enzyme 5-LOX inhibitor, and justifies the development of confirmatory in vitro / in vivo assays to corroborate its antioxidant and anti-inflammatory effects(AU)

Micetismos: Parte 4: Síndromes tempranos con síntomas complejos / Mushroom poisonings. Part 4: early-onset syndromes with complex symptoms / Intoxicações por cogumelos. Parte 4: Síndromes precoces com sintomas complexos

En esta Parte 4 de la serie de cuatro artículos sobre micetismos se analizan los síndromes que se caracterizan por presentar un período de latencia muy corto, con la aparición de síntomas complejos en menos de 6 horas después de la ingestión de los macromicetos. Se discuten los siguientes micetismos: 1) Toxíndrome muscarínico o colinérgico periférico por especies de Inocybe y Clitocybe. 2) Toxíndrome inmunohemolítico o hemolítico por Paxillus. 3) Toxíndrome neumónico alérgico por Lycoperdon perlatum y por Pholiota nameko. 4) Toxíndrome panterínico o neurotóxico glutaminérgico por compuestos isoxazólicos o síndrome pantherina/muscaria. 5) Toxíndrome coprínico o cardiovascular. 6) Toxíndrome neurotóxico alucinogénico por psilocibina y derivados indólicos. 7) Toxíndrome psicotrópico por estirilpironas y gimnopilinas de Gymnopilus spectabilis o G. junonius. 8) Toxíndrome agudo de rabdomiólisis por Russula subnigricans. 9) Toxíndrome cianogénico por Marasmius oreades. 10) Toxíndrome inmunosupresor por tricotecenos macrocíclicos de Podostroma cornu-damae. 11) Toxíndrome hemolítico debido a ostreolisina de Pleurotus ostreatus y especies relacionadas. Se analizan los síntomas, las toxinas involucradas, los mecanismos de acción, cuando se conocen, y las especies causantes de los micetismos.

This Part 4 of the series of four articles on mushroom poisonings refers to early-onset syndromes, which are characterized by a very short latency period, and the appearance of complex symptoms in less than 6 hours after mushroom ingestion. The following mycetisms are discussed, (1) Peripheral cholinergic, or muscarinic syndrome due to Inocybe and Clitocybe species. (2) Immunohaemolytic or haemolytic syndrome by Paxillus. (3) Allergic pneumonic syndrome due to Lycoperdon perlatum, and Pholiota nameko. (4) Glutaminergic neurotoxic, or pantherinic syndrome by isoxazole compounds or pantherina/muscaria syndrome. (5) Coprinic or cardiovascular syndrome. (6) Hallucinogenic neurotoxic syndrome due to psilocybin and indole derivatives. (7) Psychotropic syndrome by styrylpirones and gymnopilins of Gymnopilus spectabilis or G. junonius. (8) Rhabdomyolysis acute syndrome due to Russula subnigricans. (9) Cyanogenic syndrome by Marasmius oreades. (10) Immunosuppressive syndrome by macrocyclic trichothecenes of Podostroma cornu-damae. (11) Haemolytic syndrome due to ostreolisine of Pleurotus ostreatus, and related species. The symptoms, toxins involved, mechanisms of action, when known, and the species of mushrooms responsible for the mycetisms are analyzed.

Nesta parte 4 da série de quatro artigos sobre intoxicação por cogumelos são analisadas síndromes que se caracterizam por apresentar um período de latência muito breve, com aparecimento de sintomas complexos em menos de 6 horas após a ingestão dos macromicetos. As seguintes intoxicações com cogumelos são discutidas: (1) Toxíndrome muscarínico ou colinérgico periférico por espécies de Inocybe e Clitocybe. (2) Toxíndrome imuno-hemolítica ou hemolítica por Paxillus. (3) Toxíndrome pneumônica alérgica por Lycoperdon perlatum e por Pholiota nameko. (4) Toxíndrome panterínica ou neurotóxica glutaminérgica por compostos isoxazólicos ou síndrome pantherina/muscaria. (5) Toxíndrome coprínica ou cardiovascular (6) Toxíndrome neurotóxico-alucinogênica por psilocibina e derivados indólicos. (7) Toxíndrome psicotrópica por estirilpironas e gimnopilinas de Gymnopilus spectabilis ou G. junonius. (8) Toxíndrome aguda de rabdomiólise por Russula subnigricans. (9) Toxíndrome cianogênica por Marasmius oreades. (10) Toxíndrome imunossupressora por tricotecenos macrocíclicos de Podostroma cornu-damae. (11) Síndrome hemolítica por ostreolisina de Pleurotus ostreatus e espécies relacionadas. São analisados os sintomas, as toxinas envolvidas, os mecanismos de ação, quando conhecidos, e as espécies de cogumelos responsáveis pelas intoxicações.

Effects of carbon source and additives on biomass, exopolysaccharide production and morphology of Pleurotus ostreatus in submerged cultivation

Aims@#To investigate the influence of carbon sources and additives/surfactants on the mycelium growth and exopolysaccharides (EPS) production, including the morphology during submerged cultivation of Pleurotus ostreatus in the minimal-medium as the base medium. @*Methodology and results@#Pleurotus ostreatus was cultivated in different types of carbon sources to investigate the effects of carbon sources to mycelium growth and changes of mycelium morphology which directly affects the synthesis of EPS. In addition, additives or surfactants can increase the bioavailability of less soluble substrates in the cultured medium for the mycelium growth and indirectly affects the EPS production. In this study, the cultivation of P. ostreatus in the minimal-medium by using glucose as the carbon source with the addition of lecithin at 1% (w/v) gave the highest EPS production 4.53 ± 0.30 g/L, an increase of about 89.53% when compared to the cultivation without the addition of lecithin. Addition of lecithin changes morphology of the pellets outer layer and under microscope showing a dense hyphal network surrounding the pellets with the sizes of micro pellets almost 0.5-1.5 mm which contributed to the increase of EPS production after 14 days cultivation at 26 °C @*Conclusion, significance and impact of study@#The choice of the carbon source should not only be for high productivity rate of mycelium growth and EPS production, but a cheaper alternative source should also be considered. In conclusion, high mycelium biomass and EPS production was achieved either by changes of the morphology through the type of carbon source and addition of additives such as lecithin.

Development of a Novel Spawn (Block Spawn) of an Edible Mushroom, Pleurotus ostreatus, in Liquid Culture and its Cultivation Evaluation

Mushroom cultivation has gained increased attention in recent years. Currently, only four types of spawn, including sawdust spawn, grain spawn, liquid spawn, and stick spawn, are commonly available for mushroom cultivation. This limited spawn diversity has led to difficulty in selecting suitable inoculum materials in some cultivation. In this study, three small blocks of lignocellulosic agro-wastes and one block of a synthetic matrix were prepared as support for growing Pleurotus ostreatus in liquid medium. Mycelium-adsorbed blocks were then evaluated for their potential as block spawn for fructification. Our results indicated that the edible fungus was adsorbed and abundantly grew internally and externally on loofah sponge and synthetic polyurethane foam (PUF) supports and also has the ability to attach and grow on the surface of sugarcane bagasse and corncob supports. The mycelia of P. ostreatus adhered on corncob exhibited the highest metabolic activity, while those on the PUF showed the least activity. Mycelial extension rates of block spawns made of agro-waste materials were comparable to that of sawdust spawn, but the block spawn of PUF showed a significantly lower rate. No significant differences in cropping time and yield were observed among cultivations between experimental block spawns and sawdust spawns. Moreover, the corncob block spawn maintained its fruiting potential during an examined period of 6-month storage. The developed block spawn could be practically applied in mushroom cultivation.

Degradação do repelente DEET pelas lacases do Pleurotus ostreatus / Degradation of repellent DEET by Pleurotus ostreatus laccases

Objetivo: Avaliar a capacidade de remediação do DEET em meio líquido, pelo fungo de decomposição branca Pleurotus ostreatus usando como indutor enzimático os resíduos sólidos do cacau e realizar bioensaios de toxicidade com as amostras pós-tratamento, para aplicações em tratamentos de águas. Método: Foi realizada a produção enzimática com resíduos do Cacau. A biorremediação com o caldo enzimático foi realizada em erlenmeyers de 250mL, contendo a solução do composto, tampão acetato de sódio pH 5 e o caldo batata, incubados à 28°C, com rotação de 120 rpm, por 48 horas. Já com o fungo ativo, o mesmo foi incubado a 28 ºC e teve em seu meio a adição do composto. As amostras foram quantificados em Cromatografia líquida de alta performance (CLAE). O teste de adsorção foi feito com o fungo autoclavado e analisado após 14 dias. Resultado: O composto se apresentou possivelmente tóxico e a remediação mostrou uma tendência linear de degradação com o fungo de 39%. Conclusão: Pleurotus ostreatus é um candidato promissor para o tratamento de contimanações geradas por DEET.

Objective: We evaluated the remediation capacity of DEET in liquid medium by the white decomposition fungus Pleurotus ostreatus using the solid residues of cocoa as an enzymatic inducer and performed toxicity bioassays with the post-treatment samples, for water treatment applications. Method: Enzymatic production with cocoa residues was performed. Bioremediation with the enzyme broth was performed in a 250mL erlenmeyer flasks, containing the solution of the compound, sodium acetate buffer pH 5 and the potato broth, incubated at 28 °C, with rotation of 120 rpm, for 48 hours. With the active fungus, the same was incubated at 28 ºC and had in its culture medium the addition of the compound. The samples were quantified in high performance liquid chromatography (HPLC). The adsorption test was performed with the autoclaved fungus and analyzed after 14 days. Results: The compound was possibly toxic and the remediation showed a linear tendency of degradation of 39% with the fungus. Conclusion: Pleurotus ostreatus is a promising candidate for the treatment of contaminants generated by DEET.

Influence of Mycoremediation and Selected Surfactants on Growth Performance of Fluted Pumpkin In Crude Oil Impacted Soil and the Nephrotoxic Effects of Leaves Aqueous Extract on Wistar Rats

This study investigated the mycoremediation effects of Pleurotus ostreatus and selected surfactants (Triton x-100 and meshed Costus aferstem)on the growth performance of fluted pumpkin (Telfairia occidentalis) in crude oil impacted soil and their effects on the electrolytes, urea and creatinine levels of Wistar rats fed with aqueous leaf extract of fluted pumpkin (Telfairia occidentalis) cultivated on the amended soil. Crude oil highly impacted soil excavated from an oil spill site at Obeche community in Ogba/Egbema/Ndoni Local Government Area of Rivers State, Nigeria was used. Remediation was induced using white rot fungus (Pleurotus ostreatus), phyto-emulsified surfactant (Costus aferstem) and chemical surfactant (Triton x-100). Seven experimental cells (20cm diameter x 20cm high polypropylene bags) labelled A-G, each containing 2kg of polluted soil were used. The application of amendments to the crude oil impacted soil affected the growth of fluted pumpkin. Cell D (PSS + Triton x-100) was observed to have the highest number of leaves (15.00 ± 0.70), highest vine length (45.00 ± 0.00), highest fresh weight of leaves (16.50 ± 0.70) when compared with other cells. The control cell, cell A (CISS) and cell F (CISS + Pleurotus ostreatus+ Triton x-100) also were observed to have a positive effect on the growth performance of fluted pumpkin. When the aqueous leaf extract of fluted pumpkin was administered to the Wistar rats, potassium level was observed to be decreased in groups C to G with cell D having the lowest value (3.99 ± 0.00) when compared with group A (29.39±34.44). The results also indicated that groups C, D, F, and G were decreased when compared with group A for sodium, only group E was decreased when compared with group A for Chlorine, all the groups for Calcium where significantly (p?0.05) decreased when compared with group A. There was a significant (p?0.05) difference when group A is compared with group G for Bicarbonate. Histopathological evaluation of the kidney of Wistar rats revealed the presence of congested renal vessels and haemorhage.

Yield, Nutritional Content, and Antioxidant Activity of Pleurotus ostreatus on Corncobs Supplemented with Herb Residues

Improper disposal of herb residues in China has caused severe problems to the surrounding environment and human safety. Three herb residues, i.e., compound Kushen injection residues (CKI) and part one and part two of Qizhitongluo Capsule residues (QC1 and QC2, respectively), were used for the cultivation of Pleurotus ostreatus. The effect of the supplementation of corncobs (CC) with different herb residues on yield, nutritional composition, and antioxidant activity of P. ostreatus was investigated. Compared to the control, the higher mycelial growth rate was observed on substrates CC +30% CKI and CC +30% QC1, while the higher yield was obtained from substrates CC +30% QC2 and CC +30% CKI. Moreover, chemical analysis of fruit bodies revealed that the addition of herb residues to CC significantly increased proteins, amino acids, ashes, minerals (Na and Ca), and total phenolic contents but significantly reduced carbohydrates and IC50 values of DPPH radicals. In addition, no heavy metals (Pb, Cd, and As) were detected in the fruiting bodies harvested from different substrate combinations. These results demonstrated that mixtures of CC with herb residues might be utilized as a novel, practical, and easily available substrate for the cultivation of P. ostreatus, which is beneficial for the effective management of herb residues.

A polysaccharide fraction extracted from Pleurotus ostreatus mycelial biomass inhibit Sarcoma 180 tumor

ABSTRACT Fungi of Pleurotus genus have attracted a great interest due to their medicinal properties such as anti-inflammatory, antimicrobial and antitumor. These properties are attributed mainly to polysaccharides synthesized by Pleurotus. This work aimed to study the mycelial growth of P. ostreatus in submerged culture, evaluating the influence of the initial concentration of substrate (20 and 40 g/L of glucose) and the pH (4 and 6) on kinetic parameters of production of biomass. The effectiveness of different doses (10, 30 and 50 mg/kg) of a mycelium polysaccharide fraction extracted from P. ostreatus in reducing Sarcoma 180 development in mice was also verified. In the range of this study, maximum concentration of mycelial biomass (about 12.8 g/L) was obtained using 40.0 g/L of glucose, at pH 4.0. The total biomass productivity (Px) was not significantly affected by substrate concentration and pH, reaching values of 0.034 g/L.h. Sarcoma 180 tumor weight was reduced in 74.1, 75.5 and 53.7% when 10, 30 and 50 mg/kg were administered, respectively. These results show the high antitumor potential of intracellular polysaccharide fraction of mycelial biomass of P. ostreatus, particularly at lower doses of 10 and 30 mg/kg.

Partial removal and detoxification of Malachite Green and Crystal Violet from laboratory artificially contaminated water by Pleurotus ostreatus / Eliminación parcial y desintoxicación de Verde Malaquita y Cristal Violeta del laboratorio con agua contaminada artificialmente por Pleurotus ostreatus / Extração e desintoxicação de Verde Malaquita e Cristal Violeta de água contaminada artificialmente em laboratório por Pleurotus ostreatus

Abstract The triphenylmethane Malachite Green (MG) and Crystal Violet (CV) dyes are cationic dyes and mix with domestic wastewater when dumped; increasing, among others, the chemical and biological oxygen demand and can cause acute toxicity at different trophic levels. Promoting the removal (decolorization) of MG and CV, and laccase activity (54.8 ± 8.9 and 30.6 ± 2.9 UL-1 respectively) by using P. ostreatus viable biomass needed parameters such as pH (4.5 and 6.0), temperature (25 to 30 °C), stirring speed (120 rpm), percentage of inoculum (2% v/v), and dye concentration (20 and 10 mg L-1). In adsorption studies, it was showed that an acidic pH favors the adsorption of both dyes and the model of pseudo-second order describes best the phenomenon of adsorption. Finally, the germination index (GI), using Lactuca sativa seeds for the initial dyes solutions, was < 50%; demonstrating its high phytotoxic effect. When dye solutions were treated with viable biomass, the GI increased, leaving open the possibility to perform future research to determine if the aqueous solutions, post-treated with P. ostreatus, could be used in treatments that generate less toxic water which could be used in processes that do not require potable water.

Resumen Los colorantes trifenilmetánicos Verde Malaquita (MG) y Crystal Violeta (CV) son catiónicos y al ser vertidos se mezclan con aguas residuales domésticas, incrementando, entre otros, la demanda química y biológica de oxígeno; pudiendo causar toxicidad aguda en diferentes niveles tróficos. En este estudio se encontró que los parámetros pH (4,5 y 6,0), temperatura (25 y 30 °C), velocidad de agitación (120 r.p.m.), porcentaje de inóculo (2 % v/v) y concentración de colorante (20 y 10 mgL-1), presentaron un efecto significativo (p < 0.05) para favorecer la remoción (decoloración) de MG y CV, así como la actividad lacasa (54,76 ± 8,91 y 30,59 ± 2,89 UL-1 respectivamente) al utilizar biomasa viable de P. ostreatus. En los estudios de adsorción se evidenció que pH ácidos favorecen la adsorción de ambos colorantes y que el modelo de Pseudo-segundo orden describe mejor el fenómeno de quimisorción. Finalmente los índices de germinación (IG) empleando semillas de Lactuca sativa, para los colorantes iniciales fueron < 50 %; demostrando su efecto fitotóxico elevado. Cuando las soluciones de colorantes fueron tratadas con biomasa viable, el IG aumentó, dejando abierta la puerta para la realización de investigaciones futuras con la intensión de determinar si las soluciones acuosas, postratadas con P ostreatus, pueden ser utilizadas en tratamientos que generen aguas menos tóxicas y que estas puedan ser empleadas en otros procesos que no requieran agua potable.

Resumen Os corantes de tipo trifenilmetano Verde Malaquita (VM) e Cristal Violeta (CV) são corantes catiônicos e se misturam com águas residuais domésticas quando descartadas; aumentando, entre outros, as demandas químicas e biológicas de oxigênio, podendo causar toxicidade aguda em diferentes níveis tróficos. Promoveu-se a remoção (descoloração) de VM e CV, e atividade da lacase (54.8 ± 8.9 e 30.6 ± 2.9 UL-1 respectivamente) utilizando como parâmetros necessários para a biomassa viável de P. ostreatus como pH (4,5 e 6,0), temperatura (25 a 30 °C), velocidade de agitação (120 RPM), porcentagem de inócuo (2 % v/v), e concentração de corante (20 e 10 mg L-1). Em estudos de absorção, se demonstrou que um pH mais ácido favorece a absorção de ambos corantes e o modelo de pseudo-segunda ordem descreve melhor o fenômeno da absorção. Finalmente, o índice de germinação (IG), utilizando sementes de Lactuca sativa para as soluções iniciais dos corantes, foi < 50 %; demonstrando assim seu alto efeito fitotóxico. Quando as soluções de corante foram tratadas com a biomassa viável, o IG aumentou, deixando em aberto a possibilidade de realizar futuras investigações para determinar se as soluções aquosas, tratadas com P. ostreatus, poderiam ser utilizadas em tratamentos que gerem águas menos tóxicas, que poderia ser utilizada em processos que não requerem água potável.

Viral Effects of a dsRNA Mycovirus (PoV-ASI2792) on the Vegetative Growth of the Edible Mushroom Pleurotus ostreatus

A double-stranded RNA (dsRNA) mycovirus was detected in malformed fruiting bodies of Pleurotus ostreatus strain ASI2792, one of bottle cultivated commercial strains of the edible oyster mushroom. The partial RNA-dependent RNA polymerase (RdRp) gene of the P. ostreatus ASI2792 mycovirus (PoV-ASI2792) was cloned, and a cDNA sequences alignment revealed that the sequence was identical to the RdRp gene of a known PoSV found in the P. ostreatus strain. To investigate the symptoms of PoV-ASI2792 infection by comparing the isogenic virus-free P. ostreatus strains with a virus-infected strain, isogenic virus-cured P. ostreatus strains were obtained by the mycelial fragmentation method for virus curing. The absence of virus was verified with gel electrophoresis after dsRNA-specific virus purification and Northern blot analysis using a partial RdRp cDNA of PoV-ASI2792. The growth rate and mycelial dry weight of virus-infected P. ostreatus strain with PoV-ASI2792 mycovirus were compared to those of three virus-free isogenic strains on 10 different media. The virus-cured strains showed distinctly higher mycelial growth rates and dry weights on all kinds of experimental culture media, with at least a 2.2-fold higher mycelial growth rate on mushroom complete media (MCM) and Hamada media, and a 2.7-fold higher mycelial dry weight on MCM and yeastmalt-glucose agar media than those of the virus-infected strain. These results suggest that the infection of PoV mycovirus has a deleterious effect on the vegetative growth of P. ostreatus.

Maintenance culture medium and inoculum based on peach palm leaves for Pleurotus spp. production / Meio de manutenção e inóculo a partir de folhas de pupunheira para a produção de Pleurotus spp

This study evaluated the use of immersion water from peach palm leaves (PPLDA) as a component of the culture medium for the maintenance of Pleurotus spp. and the use of agricultural waste, peach palm leaves, as inoculum support for the fungi. The performance of the inoculum based on peach palm leaves (PPL) for the production of Pleurotus spp. fruiting bodies was compared with that using wheat grains (WG) as inoculum support. PPLDA culture medium (immersion water of peach palm leaves, dextrose, and agar) showed a lower radial velocity of mycelial growth for both fungi than that obtained with the culture medium WDA (wheat extract, dextrose and agar), commonly used as maintenance medium for Pleurotus spp. However, the type of inoculum support does not significantly influence the linear velocity of P. ostreatus mycelial growth, reaching 6.71 mm/day on wheat grains and 6.18 mm/day on peach palm leaves. Thus, when the inoculum based on peach palm leaves is utilized, the immersion water used for preparing this support can be used for preparing the PPLDA maintenance culture medium, diminishing the production costs of Pleurotus mushrooms. Data also showed that when Pleurotus sajor-caju was cultivated on peach palm leaves, using PPL as inoculum support, the fruiting bodies production parameters (Y = 47%, BE = 3% and Pr = 0.2 g/day) did not differ from that obtained using WG.(AU)

Este trabalho avaliou a utilização da água de imersão das folhas de pupunheira como componente do meio de cultivo para manutenção da cepa fúngica e a produção de inóculo de Pleurotus spp. utilizando como suporte folhas de pupunheira, um resíduo agrícola. Ainda, para verificar a capacidade do inóculo à base de folhas de pupunheira (FP) de produzir corpos frutíferos, foi realizado um experimento comparando FP com o inóculo à base de grãos de trigo (GT). Para tanto, realizou-se a comparação das velocidades de crescimento radial entre o meio de cultivo TDA (extrato de trigo, dextrose e ágar) utilizado para a manutenção de fungos do gênero Pleurotus e o meio FPDA (água de imersão de folhas de pupunheira, dextrose e ágar), e a comparação das velocidades de crescimento linear entre os grãos de trigo, utilizados como suporte para o inóculo de fungos do gênero Pleurotus e as folhas de pupunheira. O meio de cultivo TDA apresentou velocidade radial superior ao meio FPDA para as duas espécies, no entanto, o tipo de suporte não influenciou significativamente a velocidade de crescimento linear de P. ostreatus , que apresentou 6,71 mm/dia em grãos de trigo e 6,18 mm/dia em folhas de pupunheira. Assim, quando o inóculo à base de folhas de pupunheira é utilizado, a água de imersão deste suporte pode ser aproveitada para a confecção do meio de cultivo de manutenção (FPDA), diminuindo os custos produção. Observou-se que Pleurotus sajor-caju , quando cultivado em folhas de pupunheira utilizando inóculo FP, apresentou cerca de 47% de rendimento, 3% de eficiência biológica e 0,2 g/dia de produtividade, sem diferença significativa com o inóculo GT.(AU)

Production of lignocellulolytic enzymes from floriculture residues using Pleurotos ostreatus / Producción de enzimas lignocelulolíticas a partir de residuos de floricultura empleando Pleurotus ostreatus / Produçâo de enzimas lignoceluliticas a partir de residuos de floricultura utilizando Pleurotus ostreatus

Floriculture is a vital agro-industrial sector in the Colombian economy; the export of flowers positively impacts employment and the balance of trade. However, this industry could negatively impact the environment if its waste products are not handled properly. These flower residues, rich in lignin, hemicellulose and cellulose, could be a cost-effective raw material to produce enzymes. Here, we evaluate the production of lignocellulolytic enzymes by degradation of Chrysanthemum and Rosa residues using Pleurotus ostreatus, and manganese sulfate and copper sulfate as inductors. From the two residues, we obtained laccase, manganese peroxidase, endoglucanase, exoglucanase, and (3-glucosidase. The use of inductors, favored all enzyme activities except for (3-glucosidase. The enzymes that displayed the highest activity were laccase (4,693.4 U/L and 2,640 U/L from the residues of Chrysanthemum and Rosa, respectively) and (3-glucosidase (9,513 U/L and 6,811.9 U/L). The enzyme that showed the lowest activity was endoglucanase (11.5 U/L and 15.4 U/L). Under the conditions evaluated, the best substrate for enzyme production is Chrysanthemum wastes; the extracts obtained had higher enzymatic activity than the extracts from Rosa residues.

En Colombia la floricultura es un sector agro-industrial importante, con impactos positivos en el empleo y la balanza comercial. Sin embargo, tiene impacto negativo en el medio ambiente porque genera alto volumen de residuos. Estos residuos, ricos en lignina, hemicelulosa y celulosa, podrían ser una materia prima de bajo costo para la producción de enzimas. En este trabajo se estudió en la producción de enzimas lignocelulíticas por la degradación con Pleurotus ostreatus de residuos de Chrysanthemum y Rosa, usando como inductores sulfato de manganeso y de cobre. A partir de ambos residuos se obtuvieron lacasa, manganeso peroxidasa, endoglucanasa, exoglucanasa y p-glucosidasa. Los inductores favorecieron todas las actividades enzimáticas, excepto para p-glucosidasa. Las enzimas que tuvieron mayores actividades fueron lacasa (4,693.4 U/L y 2,640 U/L a partir del residuo de Chrysanthemum y Rosa, respectivamente) y p-glucosidasa (9,513 U/L y 6,811.9 U/L). La enzima que tuvo menor actividad fue endoglucanasa (11.5 U/L y 15.4 U/L). Bajo las condiciones evaluadas, el mejor residuo para producción de enzimas fue Chrysanthemum, porque los extractos tuvieron mayor actividad enzimática que los producidos a partir de Rosa.

Na Colombia, a floricultura è um importante setor da indùstria agrícola, com impactos positivos sobre o sector laboral na balança comercial. Além disto, tem impacto negativo sobre o meio ambiente, pois gera grandes volumes de resíduos. Estes resíduos, com altos conteùdos em lignina, hemicelulose e celulose, podem ser uma matèria-prima de baixo custo para a produçâo de enzimas. Neste traballio foi estudada a produçâo da enzimas lignoceluliticas pela degradaçâo com Pleurotus ostreatus de residuos de Chrysanthemum e Rosa, utilizando como indutores sulfatos de cobre e manganês. A partir destes resíduos foram obtidos lacase, manganês peroxidase, endoglucanase, exoglucanase e β-glicosidase. O uso de indutores favoreceu as atividades enzimáticas, exceto β-glicosidase. As enzimas que apresentaram atividades mais elevadas foram lacase (4,693.4 U/L e 2,640 U/L a partir do Chrysanthemum e Rosa, respetivamente) e β-glicosidase (9,513 U/L e 6,81.9 U/L). A enzima que apresentou menor atividade foi a endoglucanase (11.5 U/L e 15.4 U/L). Nas condiçôes testadas, o melhor resíduo para a produçâo da enzima foi Chrysanthemum, porque os extratos tinham uma atividade enzimática mais elevada que aqueles produzidos a partir de Rosa.

Optimization of submerged culture for biomass and polysaccaharide of Pleurotus ostreatus BPPTCC 6017 using response surface methodology

Aims: Polysaccharide of Pleurotus ostreatus is one of the fungal polysaccharide which has been widely studied, produced by extracting the fruiting body. An alternative method for producing polysaccharide of P. ostreatus directly from the mycelia instead of the fruiting body is through submerged culture. This study was aimed to determine the optimum submerged culture conditions for producing biomass and intracellular polysaccharide of the oyster mushroom. Methodology and results: P. ostreatus BPPTCC 6017 was collected from traditional mushroom farm in West Java, Indonesia. Submerged fermentation was conducted in 1000 mL medium (2 L flask). Four variables were tested: temperature, pH, agitation, and fermentation time, using central composite design of the response surface methodology. Mycelial biomass produced, was extracted to obtain water-soluble and alkali-soluble polysaccharide. Experimental data obtained were fitted to a second-order polynomial equation using multiple regression analysis and also analysed by appropriate statistical methods. The 3-D response surface plots derived from the mathematical models were applied to determine the optimum conditions: temperature 27.89 °C, initial pH medium 5.49, agitation 124.08 rpm, and fermentation time 11.44 days. The predicted results of the models were 33.75 g/L mycelia, 0.33 g/L water-soluble polysaccharide, and 0.64 g/L alkali-soluble polysaccharide. Those results were then verified on the optimum conditions, and produced 32.00±1.25 g/L mycelia, 0.29±0.01 g/L water-soluble polysaccharide and 0.60±0.02 g/L alkali-soluble polysaccharide, were close to the theoretical predictions. Conclusion, significance and impact study: The present study was a first effort to assess and obtain the optimum conditions for producing the biomass and polysaccharides of the strain P. ostreatus BPPTCC 6017 using submerged fermentation

Catalyzed Mediator-Based Decolorization of Five Synthetic Dyes by Pleurotus ostreatus ARC280 Laccase.

Aim: The aim of the present study was to evaluate qualitatively the decolorization of five dyes by Pleurotus ostreatus (P. ostreatus) ARC280 using solid medium. The laccase produced by the fungus was used in terms of its concentration and thermal stability for enzymatic decolorization and also in combination with Hydroxybenzotriazole (HBT) as a redox mediator. Study Design: Qualitative evaluation of decolorization of dyes and determining the best conditions required for decolorization in the presence and absence of HBT. Place and Duration of Study: Department of Microbial Chemistry, Genetic Engineering and Biotechnology Division, National Research Centre (NRC), Cairo, Egypt, between January 2013 and February 2014. Methodology: P. ostreatus ARC280 fungal ability for dyes decolorization was qualitatively evaluated on solid medium containing (g/L): dye, 0.1; glucose, 10; agar, 30; 100 mL mineral solution and 100 mL wheat bran washing water obtained by boiling 50 g of wheat bran in 1000 mL of distilled water. The efficiency of decolorization was expressed in terms of decolorization percentage as follows: Decolorization (%) = 100 × Absorbance t0 - Absorbance tf Absorbance t0 Where Absorbance t0 is the absorbance at the optimumwavelength of the reaction mixture before incubation with the enzyme and Absorbance tf is the absorbance at the optimum wavelength after incubation time. Results: The enzyme was efficient in decolorizing Acid Blue C.I. 220 (100%), Dichlorophenol indophenol sodium salt D 5110 (92.6%) and Brilliant Green C.I. 42040 (78.6%) after 6 h of incubation at 30ºC. In the presence of HBT (1 mM), Lanasol Red 6G was greatly affected by HBT as a laccase mediator system with decolorization percentage of 53.85% instead of 10.90 in case of laccase alone, however the enzyme could not efficiently decolorize Foron Yellow Brown S 2RFLI dye even in presence of HBT. The decolorization efficiency of all dyes was decreased by increasing reaction temperature from 30 to 50ºC. The absorbance reduction at the maximum wavelength was recorded with all the tested dyes. Conclusion: The results obtained clearly confirmed the role of P. ostreatus ARC280 laccase and its mediated system in the decolorization of structurally different dyes.

Purification, Biochemical Characterization and Applications of Pleurotus ostreatus ARC280 Laccase.

Aims: To purify, characterize, and apply the laccase produced by submerged fermentation using an edible mushroom Pleurotus ostreatus ARC280. Study Design: Laccase purification and characterization were designed using the most recent approaches and statistical studies of triplicate results values. Place and Duration of Study: Department of Microbial Chemistry, Genetic Engineering and Biotechnology Division, National Research Centre, Dokki, Cairo, Egypt, between May 2011 and January 2013. Methodology: P. ostreatus ARC280 laccase was purified using ammonium sulfate precipitation (40-80%), followed by gel filtration using Sephadex G100 column chromatography. The resulted pure laccase was analyzed on SDS-PAGE (12%). Laccase activity parameters such as temperature, pH, stability, metal ions and kinetic constants were studied. Laccase was applied to reduce four tumor cell lines growth and as antibacterial and antifungal agent. Results: P. ostreatus ARC280 laccase was purified using ammonium sulphate followed by Sephadex G-100 chromatographic column by about 148 purification fold with Mr of 85kDa. Optimum P. ostreatus ARC280 purified laccase activity was recorded at 50ºC and at pH 6.0, 3.0, 4.5 for Syringaldazine (SGZ), 2,2'-azinobis-(3-ethylbenzthiazoline-6-sulfonic) acid (ABTS) and 2, 6-dimethoxyphenol (DMP) as substrates, respectively. The purified enzyme was more stable in alkaline pH range and retained about 37.42, 73.51, 85.65, 87.7, 88.49, 93.65, 92.86 and 100.0 % of the initial activity after 5hrs of incubation at pH 3.0, 4.0, 5.0, 6.0, 7.0, 8.0, 9.0 and 10.0, respectively. Hg2+ caused complete inhibition at all tested concentrations; however Mn2+ (2.5x10-3M) caused laccase activation by about 190 and 330% after 1 and 24 hrs, respectively. Km and Vmax were calculated and found to be 0.074, 2.857 and 0.476 μM and 1.563, 2.500 and 2.632 μmol min-1 for SGZ, DMP and ABTS, respectively. The purified enzyme has the ability to reduce four tested cell lines growth in vitro with percentage reduction of 16.8, 23.4, 15.2 and 23.4% for HePG2, HCT116, A549 and MCF7, respectively. On the other hand, the enzyme was found to have antibacterial and antifungal activities against Escherichia coli and Candida albicans respectively. Conclusion: This enzyme seems to be a prospective enzyme for further biotechnological exploitation such as anticancer and antimicrobial activity applications.

Efficiency of treatments for controlling Trichoderma spp during spawning in cultivation of lignicolous mushrooms

Trichoderma spp is the cause of the green mold disease in mushroom cultivation production. Many disinfection treatments are commonly applied to lignocellulose substrates to prevent contamination. Mushroom growers are usually worried about the contaminations that may occur after these treatments during handling or spawning. The aim of this paper is to estimate the growth of the green mold Trichoderma sp on lignocellulose substrates after different disinfection treatments to know which of them is more effective to avoid contamination during spawning phase. Three different treatments were assayed: sterilization (121 ºC), immersion in hot water (60 and 80 ºC), and immersion in alkalinized water. Wheat straw, wheat seeds and Eucalyptus or Populus sawdust were used separately as substrates. After the disinfection treatments, bagged substrates were sprayed with 3 mL of suspension of conidia of Trichoderma sp (10(5) conidia/mL) and then separately spawned with Pleurotus ostreatus or Gymnopilus pampeanus. The growth of Trichoderma sp was evaluated based on a qualitative scale. Trichoderma sp could not grow on non-sterilized substrates. Immersions in hot water treatments and immersion in alkalinized water were also unfavorable treatments for its growth. Co- cultivation with mushrooms favored Trichoderma sp growth. Mushroom cultivation disinfection treatments of lignocellulose substrates influence on the growth of Trichoderma sp when contaminations occur during spawning phase. The immersion in hot water at 60 ºC for 30 min or in alkalinized water for 36 h, are treatments which better reduced the contaminations with Trichoderma sp during spawning phase for the cultivation of lignicolous species.

Development of bioprocess for the production of laccase by Pleurotus ostreatus MTCC 1802 using evolutionary optimization technique.

For cost effective production of laccase enzyme (benzenediol: oxygen oxidoreductase) from P. ostreatus MTCC 1802 through solid sate fermentation, physico-chemical parameters such as temperature (20-35 ºC), incubation period (9-17 days) and substrate (Neem bark and wheat bran, in various ratios, w/w) were optimized first by one parameter at time approach and then obtained optimum conditions were considered as zero level in evolutionary optimization factorial design technique. At statistically optimized conditions yield of laccase was found 303.59+16.8) U/gds after 13 days of incubation at 25 ºC taking wheat bran and neem bark as substrate at a ratio of 3:2 (w/w). The results obtained could be a base line for industrial scale production of laccase.