Enhanced production of ligninolytic enzymes and decolorization of molasses distillery wastewater by fungi under solid state fermentation

Selected isolates of fungi were grown on wheat straw and corncob in the presence of different moistening agents such as water, molasses, potato dextrose broth and distillery effluent. All the fungal isolates responded differently with respect to growth and ligninolytic enzyme production. Fungal growth on different substrates was checked by calculating ergosterol content, which varied widely within a single species when grown on different substrates. The maximum laccase production was obtained for Aspergillus flavus TERI DB9 grown on wheat straw with molasses. For manganese peroxidase, highest production was in Aspergillus niger TERI DB20 grown on corncob with effluent. Among the two isolates positive for lignin peroxidase, the highest production was in Fusarium verticillioides ITCC 6140. This immobilized fungal biomass was then used for decolorization of effluent from a cane molasses based distillery. Maximum decolorization (86.33%) was achieved in Pleurotus ostreatus (Florida) Eger EM 1303 immobilized on corncob with molasses in a period of 28 days.     

The commercially cultivated edible oyster mushrooms Pleurotus sajor-caju and P. pulmonarius are two separate species, similar in morphology but reproductively isolated

Two closely related commercially cultivated oyster mushroom species, Pleurotus pulmonarius and P. sajor-caju have been differentiated by traditional mating experiments as well as analysis of the variable ITS and IGS sequences of the ribosomal gene cluster. Molecular analysis of the variable ITS and IGS regions has allowed neither reliable differentiation between the morphologically similar species P. pulmonarius and P. sajor-caju nor confirmation of species identity of the P. sajor-caju strains CS-32, H-1, and H-2. Analysis of the sexual (mating) compatibility between haploid tester strains of these two species in monokaryon-monokaryon mating experiments has demonstrated complete reproductive isolation between P. pulmonarius and P. sajor-caju, thereby confirming that these are separate species.     

Bioconversion of agricultural lignocellulosic wastes through the cultivation of the edible mushrooms Agrocybe aegerita,Volvariella volvacea and Pleurotus spp.

Ten selected wild and commercial strains of Pleurotus ostreatus,Pleurotus eryngii,Pleurotus pulmonarius, Agrocybe aegerita andVolvariella volvacea were cultivated on three agricultural wastes, i.e. wheat straw (WS), cotton waste (CW) and peanut shells (PS). All species demonstrated significantly higher colonization rates on WS and CW than on PS. WS supported faster growth of A. aegerita and Pleurotus spp., whereas V. volvacea performed better on CW. Comparison of growth rates on composted and non-composted WS and CW substrates revealed that in the latter case faster colonization was achieved, particularly for Pleurotus spp. However, one commercial strain of V. volvacea presented higher growth rates when the composted CW medium was used. Furthermore, earliness in the fructification of P. ostreatus, P. pulmonarius and V. volvacea strains was promoted in CW substrates, while WS favoured earliness of P. eryngii and A. aegerita. Similarly, high sporophore yields were obtained by P. ostreatus and P. pulmonarius on both wastes, whereas WS enhanced yield and basidioma size of P. eryngii and A. aegerita strains and CW production of V. volvacea. The substrates cellulose:lignin ratios were found to be positively correlated to mycelial growth rates and to mushroom yield of P. ostreatus and P. pulmonarius; in addition, positive correlation was also detected for carbon:nitrogen ratio and mushroom yield in P. eryngii and A. aegerita and between cellulose content and mushroom yield for V. volvacea strains.     

Effect of lignin-derived phenolic monomers on the growth of the edible mushrooms Lentinus edodes,Pleurotus sajor-caju and Volvariella volvacea

Pleurotus sajor-caju was generally more tolerant to lignin-related phenolic monomers and tannin derivatives than Lentinus edodes and the straw mushroom, Volvariella volvacea. Several phenols, at up to 5 mM, enhanced mycelial growth of P. sajor-caju. No clear pattern was evident when the effects of phenols and tannins on the growth of V. volvacea and L. edodes were compared, but the lower concentrations of 4-hydroxybenzaldehyde and vanillin which were tested were markedly more toxic to the straw mushroom. The distribution of phenolic monomers and tannin derivatives in the agricultural wastes used for mushroom cultivation may be an important growth determinant. However, the differences in the growth inhibition profiles of L. edodes, P. sajor-caju and V. volvacea suggest that, alone, the effect of these compounds on fungal growth is unlikely to account for the varying abilities of the three mushroom species to grow and fruit on a particular lignocellulosic substrate.     

Decolourization of Municipal Effluent and Sludge by Pleurotus sajor-caju and Pleurotus ostreatus

Summary The Americana Municipal Treatment Station, S?o Paulo, Brazil, manages 400 l of effluent s⁻¹, from domestic and textile origin, which produces an average of 20 t of sludge per day. The decolourization of the effluent and sludge by three strains of Pleurotus (Pleurotus sajor-caju F2, F6 and Pleurotus ostreatus) was evaluated. The strains of P. sajor-caju F2 and F6 were able to decolourize the sludge, while P. ostreatus was less efficient. Detoxification was appraised with three bioassays comprising the cnidarian Hydra attenuata, the alga Selenastrum capricornutum and lettuce seeds. After exposure to fungi, effluent toxicity decreased but not that of its sludge. Strain P. sajor-caju F6 presented signs of toxicity shown by electron microscopy in the presence of the effluent. The three strains produced high amounts of manganese-peroxidase (Mn–P) and laccase in the presence of the sludge. Although P. ostreatus produced large amount of Mn–P and laccase enzymes, these enzymes did not result in decolourization of the sludge, suggesting that other factors are likely to be involved. Carbon content decreased only in the treatment with P. ostreatus.     

Bioconversion of sugar cane crop residues with white-rot fungiPleurotus sp.

Four mushroom strains ofPleurotus spp. were cultivated on sugar cane crop residues for 30 days at 26°C. Biochemical changes affected the substrate as a result of fungal growth, in terms of nitrogen, lignin, cellulose and hemicellulose contents. All strains showed a strong ligninolytic activity together with variable cellulolytic and xylanolytic action.Pleurotus sajor-caju attacked lignin and cellulose at the same rate, showing a degradation of 47% and 55%, respectively. A better balance was shown by theP. ostreatus-P. pulmonarius hybrid, which exhibited the poorest cellulolytic action (39%) and the highest ligninolytic activity (67%). The average composition of mushroom fruit bodies, in terms of nitrogen, carbohydrates, fats and amino acid profiles, was determined. Crude protein and total carbohydrate varied from 23% to 33% and 36% to 68% of dry matter, respectively. Fat ranged from 3.3% to 4.7% and amino acid content from 12.2% to 22.2%. Slight evidence for a nitrogen fixing capability was encountered in the substrate to fruit body balance.     

Mineral content of the fruiting bodies of Pleurotus sajor-caju (FR.) SINGER cultivated on different kinds of biomass

Pleurotus sajor-caju (FR. ) SINGER was cultivated on different organic wastes, namely sericulture waste, Populus deltoides MARSH , and Eupatorium adenophorum SPRENG . Paddy straw was taken as the control and all the data were compared with it. The mineral contents of the fruiting bodies of Pleurotus sajor-caju and the substrates on which the mushroom was grown were analyzed. Among the eight minerals determined (calcium, phosphorus, potassium, magnesium, sodium, iron, manganese and zinc), the potassium content was highest followed by phosphorus, magnesium and sodium. Analysis of the mineral contents of the substrates before cultivation had also been carried out. The mineral contents of the fruiting bodies of Pleurotus sajor-caju were found to be different on different substrates. It was also observed that the mineral contents of the fruiting bodies of Pleurotus sajor-caju increase when cultivated on substrates with higher mineral contents. The maximum mineral contents per 100 g of the substrates before cultivation were Ca – 347 mg; P – 151 mg; K – 1,805 mg; Na – 127 mg; Mg – 227 mg; Fe – 53 mg; Mn – 10 mg and zn – 3.1 mg. The mineral contents of the fruiting bodies of Pleurotus sajor-caju per 100 g ranged as follows: Ca – 25.1 mg to 35.3 mg; P – 448 mg to 602 mg; K – 2,146 mg to 2350 mg; Na – 139 mg to 229 mg; Mg – 153 mg to 224 mg; Fe – 9.74 mg to 20.75 mg; Mn – 2.5 mg to 4.0 mg and Zn – 2.2 mg to 3.1 mg.     

Concentration of fungal ligninolytic enzymes by ultrafiltration and their use in distillery effluent decolorization

Extracellular ligninolytic enzymes secreted by seven different fungi grown under solid-state fermentation using agro-residue as substrate were extracted through successive extractions. In general, most of the enzymes were recovered during first and second extractions. These extractants were then subjected to ultrafiltration using a 10 kDa membrane for further concentration. The permeates collected after every filtration and the final retentate were analyzed for protein and enzymes. In all the isolates, enzyme concentration was maximum in first retentate, which reduced significantly in second, third, and fourth retentate. Maximum per unit laccase (14.44 U g⁻¹) and MnP production (142.2 U g⁻¹) was observed in Fusarium verticillioides TERIDB16 while maximum LiP production (137.42 U g⁻¹) was in Alternaria gaisen TERIDB6. The retentate was further used for checking its decolorization efficiency of undiluted distillery effluent. The maximum decolorization (37%) was obtained using the enzyme extract of Pleurotus florida EM1303.     

Protein enrichment and digestibility of soft rush (<Emphasis Type="Italic">Juncus effusus</Emphasis>) and rice residues using edible mushrooms <Emphasis Type="Italic">Pleurotus ostreatus</Emphasis> and <Emphasis Type="Italic">Pleurotus sajor-caju</Emphasis>

Pleurotus species are found to be among the most efficient lignocellulolytic types of white-rot fungi. Rice is the main grain cultivated in the extreme south of Brazil. Defatted rice bran and straw are by-products of low aggregate value. Soft rush (Juncus effusus) is a common native plant also very abundant in the region. In the present work, we evaluated changes in substrate composition after growth of two white-rot fungal species: Pleurotus ostreatus and Pleurotus sajor-caju, aiming to increase protein content and digestibility from substrates through solid fermentations and obtain edible mushrooms of high aggregate value. For that, defatted rice bran, defatted rice straw and soft rush were utilized as substrate. The influence of the variables thermal treatment temperature of substrate, substrate moisture and concentration were evaluated on the protein content, digestibility and biological efficiency. The highest protein enrichment of rice bran in P. sajor-caju-fermented medium was due the fact that there was no fructification in these media, while for the P. ostreatus-fermented medium, part of the synthesized protein was converted into mushrooms. The highest protein enrichments were verified in medium with 80% moisture and 25% soft rush (47.1% using P. ostreatus and 49.0% using P. sajor-caju). A higher digestible protein increase was obtained for both species in media with 70% moisture and 25% soft rush.     

Biodegradation of gossypol by the white oyster mushroom,Pleurotus florida,during culturing on rice straw growth substrate,supplemented with cottonseed powder

The capacity of the white oyster mushroom, Pleurotus florida to biodegrade gossypol was studied, when grown on rice straw supplemented with cottonseed powder. The mushroom fruiting bodies did not contain any residues of gossypol at concentrations of cottonseed powder 0.15–0.60% nitrogen contents of rice straw at the end of mycelial ramification. However, the cottonseed supplementation (at 0.30% N level itself) caused a doubling in the mushroom yield and its protein content, per unit weight straw substrate. The mushroom mycelium when grown on synthetic medium in liquid cultures was able to biodegrade gossypol. A pre-incubation period of 5 days before the addition of gossypol into the culture medium, an inoculum load 10 mg and an incubation period of 10 days at 25 °C caused the biodegradation of 100 g gossypol. Increased concentrations of gossypol required increased duration and increased inoculum levels to effect biodegradation. However, the effect was more pronounced with an increase in inoculum density. The fungal monoculture when grown in rice straw (powder) (5%) + glucose (1%) liquid culture medium, showed an increase in hexosamine content and laccase activity that produced an increased degradation of gossypol over an incubation period from 5 to 25 days. Enzymic extracts of the mycelial monoculture raised on the chopped rice straw substrate when incubated with 100 g of gossypol demonstrated its biodegradability; the increase in enzyme concentration showed enhanced gossypol degradation. This study adds to the world list of organic compounds that Pleurotus is able to biodegrade, and explains the cause of non-yellowing of the white oyster mushroom (P. florida) fruiting bodies, during culture on rice straw with supplementation of cottonseed powder for enhancing the mushroom yields.     

Recycling spent larval food of Corcyra cephalonica Stainton for preparing spawn and sporophore of Pleurotus sajor-caju (Fr.) Singer

Large scale production of the rice moth, Corcyra cephalonica Stainton in pearl millet grain medium leads to a huge accumulation of spent larval medium in commercial insectaries. We attempted bioconversion of spent larval medium of C. cephalonica (CLM) for cultivation of the mushroom Pleurotus sajor-caju (Fr.) Singer, to increase the usage of these residues. Maximum efficiency limits of CLM for spawn run, sporophore cropping and as bed substrate were assessed with varying combinations of sorghum and rice straw. Sorghum grains and rice straw were the best substrates for spawn run and sporophore yield respectively. Having been crushed, macerated, heated and sterilized, CLM could also become a suitable substrate along with sorghum or rice straw. Sorghum and CLM at 16.7% + 83.3% and 33.3% + 66.7% combinations were very effective in supporting mycelial growth and quicker colonization of fungus, and mother spawn yield. The spawn that was obtained from these combinations yielded higher sporophore as well. The fungus did not rapidly colonize on other combinations (50% + 50%, 66.7% + 33.3% and 83.3% + 16.7%), and was completely unable to grow on CLM 100%. Combination of rice straw and CLM at 75% + 25% and 50% + 50% as bed substrate contributed higher sporophore yield. Analysis of the substrates indicated variation in their chemical and mineral composition, but they were good sources of N, P and Ca. The prospects of exploring CLM for the mushroom cultivation are discussed.     

Growth of higher fungi on wheat straw and their impact on the digestibility of the substrate

Summary The influence of the growth of three higher fungi on the composition of wheat straw was investigated. Pleurotus pulmonarius, P. sajor-caju and Lentinus edodes grew very well on lignocellulosic substrates, breaking down a considerable amount of lignin. The initial lignin concentration of straw was halved after 12 weeks of fungal growth, doubling the enzymic digestibility. Together with lignin, the higher fungi consumed half of the amount of hemicellulose (i.e. 15%), leaving cellulose fairly intact, which should remain as an energy source for ruminants.     

Solid state fermentation of Saccharum munja residues into food through Pleurotus cultivation

The technical feasibility of using Saccharum munja as a substrate for the cultivation of the oyster mushroom, Pleurotus sajor-caju, is evaluated. The biological efficiency of mushroom production is compared on different substances-S. munja, S. munja plus paddy straw, and paddy straw. Although the efficiency is low on S. munja, the ready availability of this weed large areas holds a favourable option for its use in mushroom cultivation. The crude proteins, fats, carbohydrates, and energy values are also lower though the differences are not great. The degradation of three major components-cellulose, hemicellulose, and lignin has been observed, which proves P. sajor-caju to be a lignocellulolytic fungus. The nitrogen and mineral analysis of post-mushroom production S. munja (spent substrate) compares favorably with the dry cow-feed ration showing an enhanced protein content in the spent substrate.     

The microbial conversion of different agricultural residues and its biological efficiency

A number of agricultural residues such as Saccharum munja Roxb. (Sarkanda), Oryza sativa L. (Paddy straw, as the control), Vinna unguiculata (L.) Walp (Cowpeas), Abelmoschus esculentum (L.) Moench (Lady's finger), Zea mays L. (Maize) and Cyamopsis tetragonoloba (L.) Taub. (Guar) were used for the cultivation of Pleurotus sajor-caju (Fr.) Singer. The biological efficiency of the fruit bodies of Pleurotus sajor-caju from the above mentioned substrates were found to be 13.47, 11.20, 8.37, 8.31, 6.87, and 6.08 percent, respectively. S. munja and paddy straw were found to be the best substrates for the growth of P. sajor-caju followed by Z. mays, V. unguiculata, A. esculentum, and C. tetragonoloba.     

Recycling of spent shiitake substrate for production of the oyster mushroom,Pleurotus sajor-caju

Pleurotus sajor-caju was produced on a basal medium containing spent shiitake substrate plus 10% wheat bran and 10% millet. An analysis of the fibrous composition of the spent shiitake substrate revealed that 85% of the original hemicellulose, 44% of the original cellulose and 77% of the original lignin was not consumed during production of a full crop (78% biological efficiency) of shiitake (63-day harvest period). To produce P. sajor-caju, the spent shiitake substrate was ground, air dried, supplemented, pasteurized with live steam and spawned. Highest yields (79% biological efficiency) of P. sajor-caju were obtained by supplementing the spent shiitake basal medium with 12% soybean and 1% CaCO₃. Increases in percentage biological efficiency and mushroom size were positively correlated with increasing levels of CaCO₃ added to the basal medium.     

Biological Treatment of a Pulp and Paper Industry Effluent by Pleurotus spp.

In this work, the ability of Pleurotus spp.:P. sajor-caju; P. platypus and P. citrinopileatus to treat pulp and paper mill effluent on a laboratory and pilot scale were studied. On the laboratory scale treatment, P. sajor-caju decolorized the effluent by 66.7% on day 6 of incubation. Inorganic chloride liberated by P. sajor-caju was 230.9% (814.0 mg/dl) and the COD was reduced by 61.3% (1302.0 mg/dl) on day 10 of treatment. In the pilot scale treatment maximum decolorization was obtained by P. sajor-caju (60.1%) on day 6 of the incubation. Inorganic chloride content was increased by 524.0 mg/dl (113.0%) and the COD was reduced by 1442.0 mg/dl (57.2%) by P. sajor-caju on day 7 of incubation. These results revealed that the treatment of pulp and paper mill effluent by P. sajor-caju proved as better candidate for the purpose than P. platypus and P. citrinopileatus.     

Bioconversion and biotransformation of coir pith for economic production of Pleurotus florida: chemical and biochemical changes in coir pith during the mushroom growth and fructification

Coir pith represents ∼50% of the waste from the coir industries and was tested for its potential in serving as a growth substrate for the production of species of oyster mushroom, Pleurotus florida. Due to its high lignin (∼48%) content and amorphous powdery nature, coir pith supported poor mushroom mycelial growth and yields were considerably low (∼25% bioconversion efficiency). Pre-treating coir pith with hot water did not prove economical to produce the mushroom yields. Acid swelling and alkali delignification of coir pith though served to change the structure of coir pith; the mushroom yields were not improved. Amendment of coir pith with rice (Oryza sativa) straw and horse gram (Dolichos biflorus) plant residue tended to greatly modify the physical characteristics of the inoculated mushroom bed. Such a supplementation of coir pith growth substrate resulted in production of mushroom yields with 110–125% bioconversion efficiency. Implications of supplementing coir pith with rice straw/horse gram plant residue in terms of holocellulose:lignin ratio are discussed. Sensorially, the mushrooms so produced did not differ from that on rice straw, the economic growth substrate recommended for production of the mushroom yields on commercial scale. Changes in cellulose, hemicellulose and lignin contents of coir pith amended with rice straw were studied. Cellulase, hemicellulase and protease enzyme activities in the amended coir pith substrate showed a continuous increase from inoculation till the end of fructification, whereas laccase activity decreased during fructification, in consonance with decreased lignin degradation during fructification.     

Heat treatment of wheat straw by immersion in hot water decreases mushroom yield in Pleurotus ostreatus

BackgroundThe oyster mushroom, Pleurotus ostreatus, is cultivated worldwide. It is one of the most appreciated mushrooms due to its high nutritional value. Immersion of the substrate in hot water is one of the most popular and worldwide treatment used for mushroom farmers. It is cheap and easy to implement.AimsTo compare the yields obtained during mushroom production of P. ostreatus using different pre-treatments (immersion in hot water, sterilization by steam and the use of fungicide) to determine if they influence mushroom crop.MethodsFour different treatments of substrate (wheat straw) were carried out: (i) immersion in hot water (IHW); (ii) steam sterilization; (iii) chemical; and (iv) untreated. The residual water from the IHW treatment was used to evaluate the mycelium growth and the production of P. ostreatus.ResultsCarbendazim treatment produced highest yields (BE: 106.93%) while IHW produced the lowest BE with 75.83%. Sugars, N, P, K and Ca were found in residual water of IHW treatment. The residual water increased the mycelium growth but did not increase yields.ConclusionsWe have proved that IHW treatment of substrate reduced yields at least 20% when compared with other straw treatments such as steam, chemical or untreated wheat straw. Nutrients like sugars, proteins and minerals were found in the residual water extract which is the resultant water where the immersion treatment is carried out. The loss of these nutrients would be the cause of yield decrease. Alternative methods to the use of IHW as treatment of the substrate should be considered to reduce economical loss.     

Mycelium growth kinetics and optimal temperature conditions for the cultivation of edible mushroom species on lignocellulosic substrates

The influence of environmental parameters on mycelial linear growth ofPleurotus ostreatus, P. eryngii, P. pulmonarius, Agrocybe aegerita, Lentinula edodes, Volvariella volvacea andAuricularia auricula-judae was determined in two different nutrient media in a wide range of temperature, forming the basis for the assessment of their temperature optimaV. volvacea grew faster at 35°C,P. eryngii at 25°C,P. ostreatus andP. pulmonarius at 30°C,A. aegerita at 25 or 30°C andA. auricula-judae at 20 or 25°C depending on the nutrient medium used andL. edodes at 20 or 30°C depending on the strain examined. The mycelium extension rates were evaluated on seven mushroom cultivation substrates: wheat straw, cotton gin-trash, peanut shells, poplar sawdust, oak sawdust, corn cobs and olive press-cake. The mycelium extension rates (linear growth and colonization rates) were determined by the ‘race-tube’ technique, and were found to be the highest on cotton gin-trash, peanut shells and poplar sawdust forPleurotus spp. andA. aegerita. Wheat straw, peanut shells and particularly cotton gin-trash supported fast growth ofV. volvacea, whereas wheat straw was the most suitable substrate forL. edodes andA. auricula-judae. Supplemented oak sawdust and olive press-cake were poor substrates for most species examined, white almost all strains performed adequately on corn cobs.     

Influence of formaldehyde-treated soybean and commercial nutrient supplementation on mushroom (Pleurotus sajor-caju) yield and in-vitro dry matter digestibility of spent substrate

Summary Pleurotus sajor-caju 537 was grown on chopped, pasteurized wheat straw non-supplemented and supplemented with formaldehyde-treated soybean, commercial delayed-release nutrient (SpawnMate II SE) or vegetable oil. Yield was 2.1-fold higher for substrate supplemented (12% dry wt) with low-volume formaldehyde-treated soybean as compared to non-supplemented substrate. Mushroom yield from substrate supplemented with commercial nutrient was 1.7-fold higher than yield from non-supplemented substrate. As the supplement level increased, the mushroom yield response increased. The yield ranged from 3.56 kg/m² for non-supplemented substrate to 7.36 kg/m² for substrate supplemented (12% dry wt) with formaldehyde-treated soybean. The type of supplement affected in vitro dry matter digestibility (IVDMD) of spent substrate; commercial supplement resulted in higher IVDMD compared to formaldehyde-treated substrate. An opportunity exists for commercial development of a nutrient(s) specifically designed for Pleurotus cultivation.