Strains Of White Rot Fungi Research Articles

Kinetic characterization of purified laccase produced from Trametes versicolor IBL-04 in solid state bio-processing of corncobs

A locally isolated white rot fungal strain Trametes versicolor IBL-04 produced high laccase activities in solid state bio-processing of corn cobs. Addition of glucose and yeast extract (C: N ratio; 25:1) enhanced laccase synthesis. Addition of Tween-80 and CuSO4 enhanced laccase production to 1012 U/mL under optimized process conditions. Laccase was further purified to 2.89-fold (specific activity of 840 U/mg) by ammonium sulfate fractional precipitation, dialysis, and Sephadex G-100 gel filtration chromatography. The purified laccase had a relative molecular mass of 63 kDa as detected by sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE). Best enzyme activity was at pH 5 and 40oC. Using 2,2′-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) as substrate, the enzyme showed maximum activity (Vmax) of 780 U/mL with a corresponding Michaelis constant (Km) value of 73µM. Among the different activators/inhibitors, Cu2+, Mn2+, and Fe2+ stimulated laccase activity, whereas EDTA and cystein inhibited the enzyme. The higher Vmax and lower Km for T. versicolor IBL-04laccase as compared to most of the reported laccases suggests its potential for industrial applications.

Screening and evaluation of white rot fungi to decolourise synthetic dyes, with particular reference to Antrodiella albocinnamomea

The ability of 60 white-rot fungal strains (38 species) to decolourise six structurally different synthetic dyes, namely Congo red, Orange G, Alizarin red, Crystal violet, Neutral red and Methylene blue, which, respectively, belong to the azo, anthraquinone-based, triphenylmethane and heterocyclic dye groups, were investigated by agar plate tests. Effects of different culture conditions on the decolourisation of Alizarin red by the Chinese-specific species of white-rot fungus, Antrodiella albocinnamomea, were investigated using liquid cultures. The results show that most of the isolates had the capacity to decolourise all synthetic dyes. Antrodiella albocinnamomea was first reported as having a strong decolourizing capacity and is closely related to Cerrena unicolor in the phylogenetic tree. In liquid culture, the optimum pH was 5.5 and, compared to other carbon and nitrogen sources tested, glucose and malt extracts resulted in the best decolourisation performances. The high activity of laccase produce by A. albocinnamomea was mainly responsible for its high decolourisation efficiency.

Hyperproductivity of extracellular enzymes from indigenous white rot fungi (P. chrysosporium IBL-03) by utilizing agro-wastes

An indigenous locally isolated white rot fungal strain Phanerochaete chrysosporium IBL-03was investigated for the hyper-production of ligninolytic enzymes from different agro-industrial wastes including wheat straw, rice straw, banana stalks, corncobs, corn stover, and sugarcane bagasse as substrate material in still culture fermentation technique. Screening experiments were performed at 30oC from 1 to 10 days and maximum enzyme activities were recorded after the 5th day of incubation on banana stalk. P. chrysosporium IBL-03 produced highestactivities of lignin peroxidase (LiP) and manganese peroxidase (MnP) but no laccase activity was detected in any fermented culture media. Production of ligninolytic enzymes was substantially enhanced through the optimization process. When banana stalk at 66.6 % moisture level and pH 4.5 was inoculated with 5mL spore suspension of P. chrysosporium IBL-03 at 35oC in the presence of molasses (1%) as carbon source, ammonium sulfate (0.2%) as nitrogen supplement, (1%) Tween-80 (0.3 mL) as surfactant and mediators (MnSO4 and veratryl alcohol) enhanced the LiP and MnP production up to 1040 and 965 (U/mL), respectively.

Characterization of a novel manganese peroxidase purified from solid state culture of Trametes versicolor IBL-04

A novel manganese peroxidase (MnP) produced by an indigenous white rot fungal strain Trametes versicolor IBL-04 in solid state medium of corncobs was purified and characterized. The fungus produced 964U/mL MnP in the presence of additional carbon (glucose) and nitrogen (yeast extract) supplements added at a C:N ratio of 25:1, 1mM Tween-80 (1mL), 1mM MnSO4 (1mL), and 1mM CuSO4 (1mL). The MnP was purified by ammonium sulfate fractionation (65% saturation) and dialysis, followed by Sephadex G-100 gel filtration chromatography. Purification procedures resulted in 2.4-fold purification with an overall yield and specific activity of 3.4% and 660 U/mg, respectively. The purified MnP was monomeric of molecular weight of 43 kDa, showing a single band on sodium dodecyl sulfate poly acrylamide gel electrophoresis (SDS-PAGE). The enzyme was optimally active at pH 5 and 50oC and was stable for 1 h over a broad range of pH (4-7) and temperature (40-65oC). Kinetic constants KM and Vmax of purified MnP were 70 µM and 540 U/mL for MnSO4 substrate. The effect of possible activators and inhibitors of enzyme were also investigated, and it was observed that EDTA, Cystein, and Ag+ caused MnP inhibition and inactivation to different extents, whereas MnP was activated by 4 and 3 mM of Cu2+ and Fe2+, respectively. High thermo-stability, low KM and high Vmax features of this novel MnP isolated from culture filtrate of T. versicolor IBL-04 suggests its suitability for various industrial and biotechnological applications.

Investigation of the potential of microbial stripping of dyed cotton fabric using white rot fungi

This paper reports the results of a novel approach of investigating the potential of biochemical stripping of cotton fabric dyed with reactive black B dye using five indigenous strains of white rot fungi (WRF) in Kirk’s basal salts medium. The color stripping was measured in terms of color strength (K/S value) before and after treatment. Among the five strains of WRF, Ganoderma lucidum IBL-05 showed the best stripping (20.11%) in 15 days of incubation under continuous shaking conditions (120 rpm) at pH 4.5 and 35 °C. The biochemical stripping process using G. lucidum IBL-05 was developed by optimizing various parameters like temperature, pH, additional carbon and nitrogen sources, low molecular weight mediators and metal ions. Supplementation with additional carbon and nitrogen sources enhanced the stripping to 67.73% under optimum conditions of pH 4 and temperature 35 °C. Wheat bran (1 g/100 mL) and peptone (0.2 g/ 100 mL) were found to be the best carbon and nitrogen sources, respectively. The addition of MnSO4 (1 mM) as a mediator of ligninolytic enzymes and ZnSO4 (1 mM) as a metal ion activator, further enhanced the stripping to 90.59% under optimum conditions generating a transparent effluent at the end of 15 days.

Black Liquor Decolorization by Selected White-Rot Fungi

Five different strains of white-rot fungi have been tested for their ability to decolorize black liquor on plates and on solid-state fermentation using vermiculite as the solid inert support. Since the high salt concentration inhibited the growth of all fungi, the black liquor was dialyzed against distilled water prior to use. A preliminary step on plates was carried out to qualitatively determine the capacity of the fungal strains for black liquor decolorization. Out of the five fungi studied, Phanerochaete sordida, Pycnoporus sanguineus, and Trametes elegans exhibited the more conspicuous decolorization halos in malt extract medium, while the decolorization by all the strains was not evident when a defined culture medium was used. Cultures on solid-state fermentation using vermiculite as solid support were also tested, the liquid phase was malt extract or glucose-based medium and supplemented with different black liquor concentrations. Decolorization of black liquor was largely affected by the fungal strain, the concentration of black liquor, and the carbon source. The percentage of color removal ranged from 6.14% to 91.86% depending on the fungal strain and culture conditions. Maximal decolorization was observed in malt extract cultures after 60 cultivation days. Interestingly, decolorization in malt extract medium increased with increasing black liquor concentration. The highest decolorization value was achieved by Steccherinum sp. which reduced up to 91.86% the color of the black liquor in malt extract medium; this percentage is equivalent to 5.2 g L(-1) of decolorized black liquor, the highest value reported to date. Traditional technologies used for the treatment of black liquor are not always effective and may not to be an environmentally friendly process. Vermiculite-white-rot fungi systems are presented in this work as a promising efficient alternative for the treatment of black liquor.

Novel salt and alkali tolerant neotropical basidiomycetes for dye decolorisation in simulated textile effluent

The initial alkaline pH and the concentration of sodium chloride in the synthetic liquid medium were a key factor in the capability of twenty-five white-rot fungi strains to decolorise the dye Reactive Blue 19. Six strains decolorised 90% of the dye at pH 8.0, and only Peniophora cinerea decolorized 90% of the dye at pH 9.0. Fourteen strains were capable of decolorising the dye in saline medium (sodium chloride 10 g l−1). P. ostreatus, P. cinerea and T. villosa were able to decolorize the dye both in medium with initial pH 8.0 or in saline medium. These three strains were selected and evaluated for simulated alkali-saline textile effluent decolorisation in different conditions: time of cultivation for effluent addition (0, 5, 7 and 9 days), initial pH (4.5 and 8.0) and agitation (0 and 120 rpm). P. ostreatus and P. cinerea decolorised the alkali-saline textile effluent by 93.0 and 25.4%, when the medium’s initial pH was 8.0 or 4.5, respectively, and the effluent was added in the 7th day of growth. T. villosa decolorized 40% when the effluent was added on the 9th day of cultivation at pH 4.5. Agitation increased the effluent decolorisation by T. villosa, but inhibition was observed for P. cinerea and P. ostreatus. The results showed that each fungus presented a specific behavior in relation to the best culture conditions for decolorisation of alkali-saline effluent containing reactive dyes. The strains of P. ostreatus, P. cinerea and T. villosa were considered as promising alternative for the biodegradation of this effluent, employing the strategy of effluent addition after a certain period of fungal growth.

Laccase production and simultaneous decolorization of synthetic dyes in unique inexpensive medium by new isolates of white rot fungus

Morphological and biochemical analysis of the newly isolated white rot fungal (WRF-1) strain has ability to secrete laccase in the economical medium consisted of synthetic dyes, groundnut shell (GNS) and cyanobacterial biomass (algal bloom) under submerged shaking condition at pH 5.0 and 30 °C ± 2 °C temperature. WRF-1 strain was found to decolorize synthetic dyes efficiently at pH 5.0 and 30 °C ± 2 °C temperature. The laccase activity of strain was purified to homogeneity by chromatography with yield up to 70%. The molecular mass of laccase was found to be 70 kDa by SDS-PAGE and isoelectric point was 4.8. Biotransformation of the dyes was followed spectrophotometrically and dyes were found to decolorize completely after 6 days of fermentation. LC-MS studies were used to decipher the degradation profile of synthetic dyes by WRF-1. Indigo carmine gets degraded to isatin sulfonic acid and 4-amino-3-methylbenzenesulphonic acid whereas methyl orange degraded metabolites were identified as p- N, N′-dimethylamine phenyldiazine and p-hydroxybenzene sulfonic acid. Thus the study would give a road map for the production and application of laccase enzyme on a larger scale using low cost substrate.

Degradation of selected pharmaceutical and personal care products (PPCPs) by white-rot fungi

Today, more than 3,000 pharmaceutical and personal care products (PPCPs) are used and released into the environment at low doses but they are barely degraded in wastewater treatment plants. One of the potential alternatives to effectively degrade PPCPs is based on the use of white-rot fungi (WRF) and involves the oxidative action of extracellular fungal enzymes. The aim of this work is to study the potential ability of three WRF strains, an anamorph species of Bjerkandera sp. R1, Bjerkandera adusta and Phanerochaete chrysosporium, to degrade PPCPs belonging to different therapeutic groups: anti-depressants (citalopram and fluoxetine), antibiotics (sulfamethoxazole), anti-inflammatory drugs (diclofenac, ibuprofen and naproxen), anti-epileptics (carbamazepine), tranquilizers (diazepam) and fragrances (celestolide, galaxolide and tonalide). The results reported complete degradation of all the PPCPs except for fluoxetine and diazepam, which were partially removed in percentages from 23 to 57%. In the case of fragrances, these compounds were neither detected in the fungal cultures nor in the abiotic controls, indicating the possibility of volatilization during the experiment.

Degradation of Mixed Aromatic Pollutants by Trametes Versicolor Strain 1

ABSTRACTOne of the most distributed groups of industrial pollutants consists of aromatic hydrocarbons. Because of the complex nature of such wastes the obtaining of new strains able to degrade different toxic aromatic compounds in mixtures is a challenge to many scientists exploring the area of environmental microbiology.In this study the strain of white rot fungus Trametes versicolor was cultivated in a medium comprising a mixture of phenol (0.2 g/l), resorcinol (0.1 g/l), p-cresol (0.1 g/l), and o-nitrophenol (0.14 g/l). Phenol, resorcinol and p-cresol were degraded in 24 hours and then a slow o-nitrophenol degradation was observed. There was about 50% decrease of o-nitrophenol concentration in 144 hours.The degradation of the tested concentrations of phenol, resorcinol and p-cresol in a mixture showed similar characteristics as in the experiments with each of them used as a single carbon substrate. The data showed that the presence of o-nitrophenol does not influence the biodegradation rate of other compounds included in the mixture. The capability of the investigated strain demonstrated its potential for future application in bioremediation technologies oriented to cleaning and preservation of industrially polluted water.

Growth inhibition of Microcystis aeruginosa by white-rot fungus Lopharia spadicea

Harmful cyanobacterial blooms cause water deterioration and threaten human health. It is necessary to remove harmful cyanobacteria with useful methods. A bio-treatment may be one of the best ways to do this. A strain of specific white-rot fungus, Lopharia spadicea, with algicidal ability was isolated. Its algicidal ability on algae under various conditions was determined using three main influence factors: initial chlorophyll-a content, initial pH, and algal cell mixture. The result showed that the chlorophyll-a content of Microcystis aeruginosa FACHB-912, Oocystis borgei FACHB-1108, and Microcystis flos-aquae FACHB-1028 decreased from 798+/-13, 756+/-40, and 773+/-24 microg/L to 0 within 39 h. L. spadicea could also remove more than 95% chlorophyll-a when initial chlorophyll-a content increased from 397+/-13 to 2,132+/-4 microg/L. Moreover, the strain has great removal ability under a broad initial pH range of 5.5 to 9.5. The chlorophyll-a content of the three algal strain mixtures decreased from about 672+/-23 microg/L to 0 within 45 h. After superoxide dismutase (SOD) and malondialdehyde (MAD) were assessed in a co-culture of L. spadicea, it was observed that an increase in MAD content was correlated with the decrease in chlorophyll-a content of M. aeruginosa FACHB-912. This result suggested that the algae was not only greatly inhibited but also severely damaged by the fungus.

English

The production of manganese peroxidase (MnP) by white rot fungus strain L-25 was carried out using potato-processing wastewater and the effects of amino acids in the potato-processing wastewater was investigated. The MnP was efficiently produced from the wastewater by the addition of glucose and the maximum MnP activity linearly increased with an increase in the glucose concentration. The initial pH affected the cell growth and also the production rate of the MnP. The maximum activity and the production rate of the MnP using the potato-processing wastewater-based medium were higher (ca. 2.5- fold) than that of the basal medium. Moreover, amino acids in the wastewater had significant effects on the MnP production. L-Glutamic acid, L-aspartic acid and L-serine induced the MnP secretion, on the other hand, L-phenylalanine, L-tyrosine, L-leucine and L-lysine inhibited it. The addition of L-leucine and L-lysine caused growth inhibition, while, L-phenylalanine and L-tyrosine blocked the MnP biosynthetic pathway. Ammonium ion released from the L-phenylalanine by the L-phenylalanine ammonia-lyase participated in the repression of the MnP biosynthetic pathway of the strain L-25.

Use of white rot fungi to improve the feed value of rice straw

Introduction In many countries straw is a main feed for cattle in the winter or in the dry season. However, the feed value of straw is too low to obtain high production levels. One of the main reason for the low feed value of straw is the high lignin level (for rice straw up to 5 %), preventing microbes to access the valuable carbohydrates and causing a low feed intake. In the past many attempts have been published to increase the feed value, using sodium hydroxide, urea and ammonia. Rodrigues et al. (2008) showed that enzymes of fungi are capable of increasing the digestibility of straw in rumen fluid. The aim of the present research was in investigate if growing fungi can be used to unseal the straw and to increase its feed value. Rice straw samples were incubated for different periods with two strains of white rot fungi before being fermented in rumen fluid, using the gas production technique.

Study on the influence of high salts content on fungal treatment of saline wastewaters

We investigate in this study an assay of aerobic treatment of wastewaters from the sea-foodprocessing industry with selected strains of white rot fungi. Because the effluent was highly charged with salts, the effect of high salts concentrations on growth and enzyme production for Trametes trogii and Phanerochaete chrysosporium was firstly studied. Results showed that the two selected strains tolerated high concentrations of NaCl up to 20 g l-1 for both mycelia growth and enzymes production. The production of laccase by T. trogii was inhibited by 50% for 30 g l-1 of added NaCl (512 mM). The enzyme production inhibition was less important for MnP and LiP produced by P. chrysosporium. On the other hand, the industrial wastewaters, not highly charged with organic matter (COD ¼ 3 g l-1), contained 25 g l-1 of salts and 0.3g l-1 of ammonia. But, the wastewaters were toxic according to microtoxicity and phytotoxicity tests since the percentage of bioluminescence inhibition of Vibrio fischeri (% BI) was 60% and the germination index of Lepidium sativum not exceeded 40%. The two strains grew on 80% of the effluent and the enzymes production was less than the enzymes production in the case of the control. This effluent concentration partially inhibited the COD removal efficiency but the strains were able to decrease the amount of organic matter contained up to 90%. Furthermore, the toxicity exhibited for the raw effluent was removed and the treated wastewaters by the two strains remained non toxic.

Degradation of phenanthrene by the endophytic fungus Ceratobasidum stevensii found in Bischofia polycarpa

Some strains of white rot fungi, non-lignolytic fungi and litter-decomposing basidiomycetes have been recognized as PAH degraders. The purpose of our research was to enlarge the scope of PAH-degrading fungi and explore the huge endophytic microorganism resource for bioremediation of PAHs. In this study, phenanthrene was used as a model PAHs compound. Nine strains of endophytic fungi isolated from four kinds of plant from Eupharbiaceae were screened for degradation of phenanthrene. The endophytic fungus Ceratobasidum stevensii (strain B6) isolated from Bischofia polycarpam showed high degradation efficiency and was selected for further studies. Into the fungal culture, 100 mg l(-1) phenanthrene was added, and after 10 days of incubation, about 89.51% of the phenanthrene was removed by strain B6. Extracellular ligninolytic enzyme activities of strain B6 were tested. The results showed that manganese peroxidase [MnP] was the predominant ligninolytic enzyme and that its production was greatly induced by the presence of phenanthrene. To confirm the involvement of MnP in phenanthrene degradation, promotion and inhibition studies on MnP in different concentration level of Mn(2+) and NaN(3) were performed. Additionally, fungal mycelium-free and resuspended experiments were carried out. The results showed no apparent correlation between MnP activity and phenanthrene degradation. The mycelium and fresh medium were the crucial factors affecting the degradation of phenanthrene. To date, this is the first report on PAH degradation by Ceratobasidum stevensii. This study suggests that endophytic fungi might be a novel and important resource for microorganisms that have PAH-degrading capabilities.

Fibrolytic enzymes produced in solid-state fermentation by two strains of white-rot fungi

Fibrolytic enzymes produced in solid-state fermentation by two strains of white-rot fungi

Detoxification of atrazine by selected white-rot fungal strain in optimized conditions

Detoxification of atrazine by selected white-rot fungal strain in optimized conditions

Effect of high ammonia concentrations on fungal treatment of Tunisian landfill leachates

Landfill leachates (LFL) collected from Djebel Chekir are highly loaded with COD ranging from 4.5 to 45 g l–1, ammonia ranging from 0.64 to 4.99 g l–1 and salts (14–32 mScm–1). They also contain high concentrations of phenols and hydrocarbons. Consequently, LFL exhibit high toxicity since they completely inhibit the bioluminescence of Vibrio fischeri and the germination of Lepidium sativum seeds. Assays of a biological treatment of LFL, in order to detoxify the effluent by selected strains of white rot fungi (Trametes trogii and Phanerochaete chrysosporium), were carried out. Since the effluent contained an important amount of ammonia which is problematic for microorganisms, we first studied the effect of different concentrations of ammonia on mycelia growth and enzymes secretion. Results obtained during 14 days of fungal culture showed that ammonium chloride was not problematic at concentrations up to 2 g l–1 added to the synthetic medium. However, at a concentration of 5 g l–1 added, it exerted inhibitor effects on mycelia growth and enzymes secretion. Experiments with real LFL using T. trogii and P. chrysosporium were carried out at 50% of diluted effluent. COD removal efficiencies for T. trogii and P. chrysosporium were 79% and 68%, respectively. This was accompanied by an important enzyme secretion for each strain and a reduction in phenols and hydrocarbons concentrations. Consequently, an important reduction in the toxicity of the 50% diluted LFL was achieved.

Evaluation of wild and commercial types of Pleurotus strains for their ability to decolorize cibacron black W-NN textile dye

Biological decolorisation of Cibacron Black W-NN, was comparatively studied using five commercial (Pleurotus djamor, P. ostreatus, P. citrinopileatus, P. eryngiiand P. sajor-caju) and three wild types of (Pleurotus ostreatus, P. ostreatus and P. eryngii) white rot fungi strains on agar plates. The initial dye concentrations in the medium were 20, 50, 100, 200 and 500 mg/L, respectively. All P. ostreatus strains and P. sajor-caju fully decolorized Cibacron Black W-NN without any problem. Other organisms were found to be negatively affected from increasing dye concentrations and were able to decolorize the dye used up to a certain concentration (50 mg/L for P. citrinopileatus; 100 mg/L for commercial P. eryngiiand 200 mg/L for wild P. eryngii). Key words: Agar plate screening, biological decolorisation, Pleurotus spp., textile dye

Vitality and genetic fidelity of white-rot fungi mycelia following different methods of preservation

Basidiomycetes present specific problems with regard to their preservation, because most of them do not form resistant propagules in culture but exist only as mycelium. Usually these fungi can only be preserved by serial transfer on agar (labour-intensive procedures that can increase the danger of variation or loss of physiological or morphological features), or cryopreserved in liquid nitrogen (expensive). Cryopreservation at −80 °C and lyophilisation could be good alternatives. In this work we set up and tested six protocols of cryopreservation at −80 °C, and 12 protocols of lyophilisation on 15 isolates of white-rot fungi (WRF) belonging to 10 species. The tested protocols were mainly characterized by the use of different growth media, protectants, time and number of perfusion with protectants and finally by the typology and origin of the samples to be cryopreserved (mycelium/agar plug, whole colony) or to lyophilise (mycelium/agar plug, mycelium fragment, whole colony). Cryopreservation and lyophilisation outcomes were checked, at morphological (macro- and microscopic features), physiological (growth rate and laccase, Mn-independent and Mn-dependent peroxidases activities) and genetic level (Amplified Fragment Length Polymorphisms analysis – AFLP). Vitality of all fungi was successfully preserved by all cryopreservation protocols at −80 °C, and by two lyophilisation methods. Our results showed that cryopreservation at −80 °C did not produce morphological changes in any isolate, while two isolates were affected by lyophilisation. None of the physiological features were lost, even though growth rate and enzyme activities were somehow influenced by all preservation methods. AFLP analysis showed that only the two isolates that varied in their morphology after lyophilisation produced a different DNA fingerprint pattern in comparison with that obtained before lyophilisation. These findings provide evidence that cryopreservation at −80 °C and lyophilisation are suitable alternatives to liquid nitrogen cryopreservation for preservation of some WRF strains.