Growth of Phanerochaete chrysosporium on diesel fuel hydrocarbons at neutral pH

Generally, the white-rot fungus Phanerochaete chrysosporium performs its biodegradative activities in liquid culture while growing on easily utilized carbon sources such as malt- or potato-extract. However, less is known about the potential of this organism to grow directly on environmental pollutants without regard to special conditions. Growth of P. chrysosporium on a middle fraction (MF) of diesel fuel at neutral pH in mineral medium under non-ligninolytic conditions was explored. After 14 d, the GC-analyzable n-alkanes of 1000 mg l(-1)MF were reduced to background, with most biodegradation occurring by day 7 when quantified relative to the biodegradation of the internal fuel biodegradation marker, pristane. Investigations with n-hexadecane and unmodified diesel fuel further confirmed these biodegradation results. Biomass production was monitored and indicated that fungal biomass was more than 10 times less than positive controls (potato dextrose broth, PDB) but that biomass increased relative to negative controls. When P. chrysosporium was incubated with diesel fuel and PDB, fuel biodegradation was delayed for at least 4d and inhibited overall through 14 d. Experiments with P. chrysosporium growing on n-hexadecane in the presence of 1 mM 1-aminobenzotriazole (ABT), an inhibitor of the cytochrome P-450 enzyme system, resulted in inhibition of biomass production relative to positive controls implicating the utilization of this enzyme system in n-alkane metabolism. Finally, when P. chrysosporium was incubated in a non-aqueous phase liquid (NAPL) mixture of polycyclic aromatic hydrocarbons (PAHs) and MF, n-alkanes and phenanthrene were degraded in 2 weeks while anthracene, chrysene and benzo[a]pyrene were not.     

白腐菌应用于堆肥处理含木质素废物的研究

白腐菌对木质素具有较强的降解能力。通过在含大量木质素的模拟垃圾堆肥中采取添加白腐菌菌剂与不添加该菌剂2组实验对比,在相同堆肥处理条件下,接种菌剂的堆肥中木质素总量由274988mg降至15438mg,降解率达4386%,远高于未接种菌剂的堆肥中木质素的降解率,表明白腐菌可有效用于含木质素废弃物的堆肥处理,并有望于加速堆肥腐熟,提高堆肥效率。     

Fungal biodegradation of naphthalene: microcosms studies

The present work is aimed to ascertain naphthalene biodegradation capability of P. chrysosporium and T. harzianum in soil microcosms. Considering the high naphthalene volatility, a suitable soil microcosm was set-up and used. Several degradation tests were conducted with different C/N ratio media for the two fungi in order to enquire the best range of working conditions. The kinetic studies were conducted at a maximal naphthalene concentration of 600 mg kg(-1). During experimental time course naphthalene concentration, CO2 evolution as well as phytotoxicity tests were performed as monitoring parameters. The results shown in the current paper, put in evidence that T. harzianum, differently than in liquid culture, is not able to biodegrade naphthalene directly in soil microcosm, while P. chrysosporium in the same conditions biodegrades the PAH till about 600 mg kg(-1). As concern the founded kinetics for P. chrysosporium, a saturation shape in presence of N-limited medium (high C/N ratio) was evaluated while a growing form more than linear in no-N limited medium (normal C/N ratio) was determined.     

Phanerochaete chrysosporium吸附载体的选择及染料降解研究

研究了游离细胞与载体吸附培养、不同载体材料对Phanerochaete chrysosporium进行连续染料脱色及产酶能力的影响。结果表明,P.chrysosporium可在载体上良好生长,甚至生长到载体内部。木屑、玉米芯、花生壳3种载体材料中,以木屑载体吸附培养物的持续脱色和产酶效果最佳,该培养物经三轮连续脱色后对染料RB5仍能达到最高95%的脱色率,并产生596 U/L锰依赖过氧化物酶（MnP）和1 326 U/L木质素过氧化物酶（LiP）,对染料的持续脱色和产酶能力明显优于游离细胞培养物。     

Degradation of the antifouling compound Irgarol 1051 by manganese peroxidase from the white rot fungus Phanerochaete chrysosporium

Irgarol 1051 (2-methylthio-4-tert-butylamino-6-cyclopropylamino-s-triazine), a derivative of s-triazine herbicide, is an antifouling compound used as an alternative to organotins. The compound is highly persistent and is known to be biodegraded only by the white rot fungus, Phanerochaete chrysosporium. We used partially purified manganese peroxidase (MnP) prepared from P. chrysosporium to evaluate its capacity to degrade Irgarol 1051. MnP degraded Irgarol 1051 to two major products, one identified as M1 (identical to GS26575, 2-methylthio-4-tert-butylamino-6-amino-s-triazine) and the other not identified but with same mass spectrum as M1 and a different ultraviolet spectrum. This report clearly demonstrates that this ligninolytic enzyme is involved in the degradation of Irgarol 1051.     

Use of fungi for the bioconversion of rice straw into cellulase enzyme

Cellulase production was carried out by solid state bioconversion (SSB) method using rice straw, a lignocellulosic material and agricultural waste, as the substrate of three Trichoderma spp. and Phanerochaete chrysosporium in lab-scale experiments. The results were compared to select the best fungi among them for the production of cellulase. Phanerochaete chrysosporium was found to be the best among these species of fungi, which produced the highest cellulase enzyme of 1.43 IU/mL of filter paper activity (FPase) and 2.40 IU/mL of carboxymethylcellulose activity (CMCase). The "glucosamine" and "reducing sugar" parameters were observed to evaluate the growth and substrate utilization in the experiments. In the case of Phanerochaete Chrysosporium, the highest glucosamine concentration was 1.60 g/L and a high concentration of the release of reducing sugar was measured as 2.58 g/L obtained on the 4th day of fermentation. The pH values were also recorded. The range of the pH was about 5.15 to 5.56 in the case of Phanerochaete Chrysosporium.     

紫外诱变黄孢原毛平革菌染料脱色研究

对黄孢原毛平革菌孢子悬浮液进行了紫外诱变,并将筛选出的优势菌ZWYB1与未经诱变的菌作对比实验,结果表明:诱变筛选得到的ZWYB1不仅可缩短对染料的脱色时间,提高对杂环类染料蕃红花红及高浓度染料400 mg/L的孔雀石绿的脱色效果,而且诱变后的菌种具有一定的脱色能力稳定性.     

紫外诱变黄孢原毛平革菌染料脱色研究

对黄孢原毛平革菌孢子悬浮液进行了紫外诱变，并将筛选出的优势菌ZWYB1与未经诱变的菌作对比实验，结果表明：诱变筛选得到的ZWYB1不仅可缩短对染料的脱色时间，提高对杂环类染料蕃红花红及高浓度染料400mg／L的孔雀石绿的脱色效果，而且诱变后的菌种具有一定的脱色能力稳定性。     

Degradation of veratrylglycerol-β-(pentachlorophenyl)ether,a model compound for lignin bound pentachlorophenol residues

The degradation of veratrylglycerol-β-(pentachlorophenyl)ether, a model compound for lignin derived “Bound residues” by Phanerochaete chrysosporium, Corynebacterium equi and photosensitized riboflavin was studied. Whereas C. equi was not able to degrade the model compound, the other two systems decomposed the model compound to pentachlorophenol, 3,4-dimethoxybenzylalcohol and 3,4-dimethoxybenzaldehyde.     

Oxidation of triphenylarsine to triphenylarsineoxide by Trichoderma harzianum and other fungi

Arsenic resistant strains of bacteria and fungi were isolated from soil contaminated by chemical warfare agents. Until now, no metabolic products of microbial attack against the phenyl residues of the model substrate triphenylarsine (TP) were found if it was incubated together with these strains in liquid culture assays. However, one of the isolated fungi, Trichoderma harzianum As 11, was found to oxidize TP to triphenylarsineoxide (TPO). The yeast Trichosporon mucoides SBUG 801 and the white-rot fungus Phanerochaete chrysosporium were also able to oxidize the As(III) in TP. In addition, P. chrysosporium transformed phenylarsineoxide (PAO) to phenylarsonic acid (PAA) under O2-atmosphere. By means of a respirometer system, the oxidation of TP by T. harzianum As 11 was confirmed by a significantly higher consumption of oxygen in the presence of these compounds. HPLC analysis of the oxidation products TPO and PAA in the medium of the assays provided evidence for the transfer reaction of As(III) to As(V) in organic bonds. The oxidation products TPO and PAA are more hydrophilic than TP and PAO. Therefore, it was concluded that particular fungi contribute to the mobilization of arsenic in soil contaminated by chemical warfare agents.     

Mycelial growth and solid-state fermentation of lignocellulosic waste by white-rot fungus Phanerochaete chrysosporium under lead stress

Lignocellulosic biomass is an abundant renewable resource difficult to degrade. Its bioconversion plays important roles in carbon cycles in nature, which may be influenced by heavy metals in environment. Mycelial growth and the degradation of lignocellulosic waste by lignin-degrading fungus Phanerochaete chrysosporium under lead stress were studied. It was shown that P. chrysosporium could grow in liquid media with 400 mg L?1 Pb(II), and mycelial dry weight was reduced by 54% compared to the control. Yellow mycelia in irregular short-strip shape formed in Pb-containing media, whereas the control showed ivory-white regular mycelial pellets. Two possible responses to Pb stress were: dense hyphae, and secretion from mycelia to resist Pb. During solid-state fermentation of straw, fungal colonization capability under Pb stress was positively correlated with the removal efficiency of soluble-exchangeable Pb when its content was higher than 8.2 mg kg?1 dry mass. Carboxymethyl cellulase activity and cellulose degradation were inhibited at different Pb concentrations, whereas low Pb concentrations increased xylanase and ligninolytic enzyme activities and the hemicellulose and lignin degradation. Cluster analyses indicated that Pb had similar effects on the different microbial indexes related to lignin and hemicellulose degradation. The present findings will advance the understandings of lignocellulose degradation by fungi under Pb pollution, which could provide useful references for developing metal-polluted waste biotreatment technology.     

白腐真菌F1对染料脱色特性的研究

本研究从我国生物资源中筛选出一株白腐真菌F1,确定了其生长条件 ,实现了扩大培养 ,并研究了F1与黄孢原毛平革菌对 4种难降解染料的脱色效果。试验结果表明 ,F1对中性深黄GRL、酸性媒介漂蓝B和刚果红的脱色效率都超过 90 % ;白腐真菌能在短时间内将有毒染料酸性媒介漂蓝B降低到较低浓度 ;白腐真菌对染料脱色的同时 ,自身能够繁殖生长。     

Biodegradation of aromatic compounds by white rot and ectomycorrhizal fungal species and the accumulation of chlorinated benzoic acid in ectomycorrhizal pine seedlings

The capability of different white rot (WR, Heterobasidion annosum, Phanerochaete chrysosporium, Trametes versicolor) and ectomycorrhizal (ECM, Paxillus involutus, Suillus bovinus) fungal species to degrade different aromatic compounds and the absorption of 3-chlorobenzoic acid (3-CBA) by ECM pine seedlings was examined. The effect of aromatic compounds on the fungal biomass development varied considerably and depended on (a) the compound, (b) the external concentration, and (c) the fungal species. The highest effect on the fungal biomass development was observed for 3-CBA. Generally the tolerance of WR fungi against aromatic compounds was higher than that of the biotrophic fungal species. The capability of different fungi to degrade aromatic substances varied between the species but not generally between biotrophic and saprotrophic fungi. The highest degradation capability for aromatic compounds was detected for T. versicolor and H. annosum, whereas for Phanerochaete chrysosporium and the ECM fungi lower degradation rates were found. However, Paxillus involutus and S. bovinus showed comparable degradation rates at low concentrations of benzoic acid and 4-hydroxybenzoic acid. In contrast to liquid cultures, where no biodegradation of 3-CBA by S. bovinus was observed, mycorrhizal pines inoculated with S. bovinus showed a low capability to remove 3-CBA from soil substrates. Additional X-ray microanalytical investigations showed, that 3-CBA supplied to mycorrhizal plants was accumulated in the root cell cytoplasm and is translocated across the endodermis to the shoot of mycorrhizal pine seedlings.     

PCB congener selective biodegradation by the white rot fungus Pleurotus ostreatus in contaminated soil

Six strains of white rot fungi were tested for their biodegradation ability of low chlorinated polychlorinated biphenyl (PCB) commercial mixture (Delor 103) in real soil system. Phanerochaete chrysosporium and Trametes versicolor did not show any ability to degrade PCBs in soil. On the contrary, four strains of Pleurotus ostreatus were able to remove about 40% of Delor 103 in two months. All P. ostreatus strains decomposed PCBs selectively with the preference for congeners with chlorine atoms in ortho > meta > para position. Degradation efficiency decreased with increasing number of chlorination.     

Degradation of CL-20 by white-rot fungi

In previous studies, we found that the emerging energetic chemical, CL-20 (C6H6N12O12, 2,4,6,8,10,12-hexanitro-2,4,6,8,10,12-hexaazaisowurtzitane), can be degraded following its initial denitration using both aerobic and anaerobic bacteria. The C and N mass balances were not determined due to the absence of labeled starting compounds. The present study describes the degradation of the emerging contaminant by Phanerochaete chrysosporium using ring-labeled [15N]-CL-20 and [14C]-CL-20. Ligninolytic cultures degraded CL-20 with the release of nitrous oxide (N2O) in amounts corresponding to 45% of the nitrogen content of CL-20. When ring-labeled [15N]-CL-20 was used, both 14N14NO and 15N14NO were observed, likely produced from -NO2 and N-NO2, respectively. The incubation of uniformly labeled [14C]-CL-20 with fungi led to the production of 14CO2 (> 80%). Another ligninolytic fungus, Irpex lacteus, was also able to degrade CL-20, but as for P. chrysosporium, no early intermediates were observed. When CL-20 was incubated with manganese peroxidase (MnP), we detected an intermediate with a [M-H]- mass ion at 345 Da (or 351 and 349 Da when using ring-labeled and nitro-labeled [15N]-CL-20, respectively) matching a molecular formula of C6H6N10O8. The intermediate was thus tentatively identified as a doubly denitrated CL-20 product. The concomitant release of nitrite ions (NO2-) with CL-20 degradation by MnP also supported the occurrence of an initial denitration prior to cleavage and decomposition.     

Benzo[a]pyrene removal from soil by Phanerochaete chrysosporium grown on sugarcane bagasse and pine sawdust

The capacity of Phanerochaete chrysosporium grown on soil with added sugarcane baggase (BP) and pine sawdust (PS) to remove benzo(a)pyrene (BaP) was studied. A half factorial two-level experiment 2(4-1) was designed to determine the effect of: type of lignocellulosic material (BP and PS) for fungus growth, age of fungus (5 and 10d), amount of lignocellulosic material (10% and 15% w/w) and soil moisture content (water holding capacity of 45% and 56% w/w). Inoculum obtained at different ages showed that the capacity of P. chrysosporium to remove BaP depends on the lignocellulosic used and on inoculum age. Abiotic BaP removal was affected significantly (p<0.05) by inoculum age, type of lignocellulosic added and soil moisture content. The removal of BaP by lignocellulosic material was more effective by young inocula (71.97 mg BaP kg(-1) dry soil), with high percentage of added lignocellulosic (71.57 mg BaP kg(-1) dry soil) and at low soil moisture content (73.07 mg BaP kg(-1) dry soil). When fungus was grown on BP, maximum BaP removal rate was obtained at 5d of incubation (10.85 mg BaP d(-1)l(-1) and 50.12 mg BaP kg(-1) dry soil), while in PS maximum BaP removal was obtained at 10d of incubation (12.06 mg BaP d(-1)l(-1) and 39.94 mg BaP kg(-1) dry soil).     

Comparative studies of fungal degradation of single or mixed bioaccessible reactive azo dyes

A screening using several fungi (Phanerochaete chrysosporium, Pleurotus ostreatus, Trametes versicolor and Aureobasidium pullulans) was performed on the degradation of syringol derivatives of azo dyes possessing either carboxylic or sulphonic groups, under optimized conditions previously established by us. T. versicolor showed the best biodegradation performance and its potential was confirmed by the degradation of differently substituted fungal bioaccessible dyes. Enzymatic assays (lignin peroxidase, manganese peroxidase, laccase, proteases and glyoxal oxidase) and GC-MS analysis were performed upon the assay obtained using the most degraded dye. The identification of hydroxylated metabolites allowed us to propose a possible metabolic pathway. Biodegradation assays using mixtures of these bioaccessible dyes were performed to evaluate the possibility of a fungal wastewater treatment for textile industries.     

Chloroperoxidase-mediated transformation of highly halogenated monoaromatic compounds

Peroxidase transformations of widely distributed pollutants, tetra- and penta-chlorinated phenols and anilines, were studied using different peroxidases. Chloroperoxidase from Caldariomyces fumago was able to transform tetra- and penta-chlorinated phenols and anilines, while horseradish peroxidase, lignin peroxidase from Phanerochaete chrysosporium and versatile peroxidase from Bjerkandera adusta were able only to transform the halogenated phenols. Chloroperoxidase showed a specific activity on pentachlorophenol two orders of magnitude higher than lignin peroxidase and horseradish peroxidase, and one order of magnitude higher than versatile peroxidase. The main product from peroxidase oxidation in all cases was a polymeric and insoluble material. The insolubilization of halogenated phenols and anilines permits their removal, reduces their bioavailability, and thus reduces their environmental impact. The other minor products from the enzymatic transformation of highly chlorinated compounds were determined by mass spectrometry. Tetrachloroquinone, dimers and trimers of halogenated compounds were also identified. Chloroperoxidase was able to halogenate tetrachloroaniline to form pentachloroaniline.     

Loofa sponge immobilized fungal biosorbent: a robust system for cadmium and other dissolved metal removal from aqueous solution

The potential of loofa sponge discs to immobilize fungal biomass of Phanerochaete chrysosporium (a known biosorbent) was investigated as a low cost biosorbent for the removal of Cd(II) ions from aqueous solution. A comparison of the biosorption of Cd(II) by immobilized and free fungal biomass from 10 to 500 mg l(-1) aqueous solutions showed an increase in uptake of over 19% when the biomass is immobilized (maximum biosorption capacity of 89 and 74 mg Cd(II) g(-1) biomass for immobilized and free biomass respectively at a solution pH of 6). Equilibrium was established within 1h and biosorption was well defined by the Langmuir isotherm model. The immobilized biomass could be regenerated using 50 mM HCl, with up to 99% metal recovery and reused in ten biosorption-desorption cycles without significant loss of capacity. This study suggests that such an immobilized biosorbent system has the potential to be used in the industrial removal/recovery of cadmium and other pollutant metal ions from aqueous solution.     

Study of the degradation of dyes by MnP of Phanerochaete chrysosporium produced in a fixed-bed bioreactor

The production of ligninolytic enzymes by the fungus Phanerochaete chrysosporium in a fixed-bed tubular bioreactor, filled with cubes of nylon sponge, operating in semi-solid-state conditions, was studied. Maximum individual manganese-dependent peroxidase (MnP) and lignin peroxidase (LiP) activities of 1293 and 225 U/l were detected.The in vitro decolourisation of two structurally different dyes (Poly R-478, crystal violet) by the extracellular liquid obtained in the above-mentioned bioreactor was monitored in order to determine its degrading capability. The concentration of some compounds (sodium malonate, manganese sulphate) from the reaction mixture was optimised in order to maximise the decolourisation levels. A percentage of Poly R-478 decolourisation of 24% after 15 min of dye incubation was achieved.On the other hand, a methodology for a long treatment of these dyes based on the continuous addition of MnP enzyme and H(2)O(2) was developed. Moreover, this enzymatic treatment was compared with a photochemical decolourisation process. The former allowed to maintain the degradation rate almost constant for a long time, resulting in a decolourisation percentage of 70% and 30% for crystal violet and Poly R-478, respectively, after 2 h of treatment. As for the latter, it was not able to degrade Poly R-478, whereas crystal violet reached a degradation of 40% in 2 h.