Analysis of a 2,4,6-trichlorophenol-dehalogenating enrichment culture and isolation of the dehalogenating member Desulfitobacterium frappieri strain TCP-A

An anaerobic, 2,4,6-trichlorophenol ortho-dehalogenating mixed culture was enriched from sediment of the river Saale (Germany). Two isolated dechlorinating colonies (MK1 and MK2) consisted of rods of different lengths and thicknesses, indicating heterogeneity. Following subcultivation with thiosulfate as alternative electron acceptor and cocultivation with Clostridium celerecrescensT, the 2,4,6-trichlorophenol-dehalogenating bacterium Desulfitobacterium frappieri strain TCP-A was isolated and characterized regarding its taxonomic properties and the spectrum of chlorophenols that it dehalogenated. Four other bacterial strains were coenriched and identified as organisms with closest phylogenetic relatedness to the Clostridium type strains C. indolis, C. glycolicum, C. hydroxybenzoicum and C. sporosphaeroides (16S rDNA sequence identities of 99.5, 99.2, 94.4, and 93.5%, respectively). Amplified ribosomal DNA restriction analysis of the original dehalogenating cultures MK1 and MK2 (when not exposed to thiosulfate) confirmed the microbial heterogeneity and revealed the presence of two additional species related to the type strains of C. celerecrescens and Clostridium propionicum. Only one copy of the 16S rRNA genes of Desulfitobacterium frappieri in each of the clone libraries of MK1 and MK2 (containing 136 and 56 clones, respectively) was found by dot-blot hybridization, suggesting a relatively low number of the dehalogenating bacterium within the enrichment culture.     

Enzymatic dehalogenation of pentachlorophenol by extracts from Arthrobacter sp. strain ATCC 33790.

Arthrobacter sp. strain ATCC 33790 was grown with pentachlorophenol (PCP) as the sole source of carbon and energy. Crude extracts, which were prepared by disruption of the bacteria with a French pressure cell, showed no dehalogenating activity with PCP as the substrate. After sucrose density ultracentrifugation of the crude extract at 145,000 x g, various layers were found in the gradient. One yellow layer showed enzymatic conversion of PCP. One chloride ion was released per molecule of PCP. The product of the enzymatic conversion was tetrachlorohydroquinone. NADPH and oxygen were essential for this reaction. EDTA stimulated the enzymatic activity by 67%. The optimum pH for the enzyme activity was 7.5, and the temperature optimum was 25 degrees C. Enzymatic activity was also detected with 2,4,5-trichlorophenol, 2,3,4-trichlorophenol, 2,4,6-trichlorophenol, and 2,3,4,5-tetrachlorophenol as substrates, whereas 3,4,5-trichlorophenol, 2,4-dichlorophenol, 3,4-dichlorophenol, and 4-chlorophenol did not serve as substrates.     

Effect of pH on the anaerobic dechlorination of chlorophenols in a defined medium

Anaerobic dehalogenation of aromatic compounds is a well-documented phenomenon. However, the effects of operating parameters such as pH have received little attention despite their potential impact on treatment processes using dehalogenating organisms. In this work the effect of pH on the dehalogenation of 2,4,6-trichlorophenol (2,4,6-TCP) was studied using defined media containing one of several non-fermentable buffering agents (MOPS, TRICINE, BICINE, CHES), and no chloride ions. The dechlorination process was followed by monitoring the disappearance of 2,4,6-TCP, as well as the appearance of its dehalogenation products, i.e., 2,4-dichlorophenol (2,4-DCP), 4-chlorophenol (4-CP), and chloride ions. The results indicate that dechlorination occurs only if the pH is within the range 8.0–8.8. The newly formed 2,4-DCP was also dehalogenated in the process. However, even within this pH range dechlorination ceased when all 2,4,6-TCP and 2,4-DCP was converted to 4-CP. Stoichiometric amounts of all dehalogenation products (including chloride) could be recovered at any stage during the process. In addition, the biomass concentration was measured. After an initial lag phase, it appeared that the rate of dechlorination per unit biomass (proportional to the Cl^– concentration divided by the biomass concentration) went through a rapid increase and then remained constant throughout the process. This indicates that the dechlorinating organism(s) either make up the entire population or constitute a stable fraction of it. Correspondence to: P. M. Armenante     

硝酸盐还原条件下Fe0/厌氧微生物联合体系降解2,4,6-三氯酚

本文采用间歇试验,对硝酸盐还原条件下Fe0/厌氧微生物联合体系降解2,4,6-三氯酚(2,4,6-TCP)进行了研究。考察了不同硝酸盐浓度下,体系中pH、硝酸盐浓度以及硝酸盐还原活性的变化情况。结果表明:当2,4,6-TCP初始浓度为20mg/L时,硝酸盐对Fe0/厌氧微生物联合体系降解2,4,6-三氯酚具有明显的抑制作用;且随着硝酸盐浓度的升高,2,4,6-TCP的去除率降低,硝酸盐还原活性升高;体系先发生硝酸盐还原再进行2,4,6-TCP还原脱氯。     

Degradation of Chlorophenols by Alcaligenes eutrophus JMP134(pJP4) in Bleached Kraft Mill Effluent

The ability of Alcaligenes eutrophus JMP134(pJP4) to degrade 2,4-dichlorophenoxyacetic acid, 2,4,6-trichlorophenol, and other chlorophenols in a bleached kraft mill effluent was studied. The efficiency of degradation and the survival of strain JMP134 and indigenous microorganisms in short-term batch or long-term semicontinuous incubations performed in microcosms were assessed. After 6 days of incubation, 2,4-dichlorophenoxyacetate (400 ppm) or 2,4,6-trichlorophenol (40 to 100 ppm) were extensively degraded (70 to 100%). In short-term batch incubations, indigenous microorganisms were unable to degrade such of compounds. Degradation of 2,4,6-trichlorophenol by strain JMP134 was significantly lower at 200 to 400 ppm of compound. This strain was also able to degrade 2,4-dichlorophenoxyacetate, 2,4,6-trichlorophenol, 4-chlorophenol, and 2,4,5-trichlorophenol when bleached Kraft mill effluent was amended with mixtures of these compounds. On the other hand, the chlorophenol concentration and the indigenous microorganisms inhibited the growth and survival of the strain in short-term incubations. In long-term (>1-month) incubations, strain JMP134 was unable to maintain a large, stable population, although extensive 2,4,6-trichlorophenol degradation was still observed. The latter is probably due to acclimation of the indigenous microorganisms to degrade 2,4,6-trichlorophenol. Acclimation was observed only in long-term, semicontinuous microcosms.     

Environmental significance of O-demethylation of chloroanisoles by soil bacterial isolates as a mechanism that improves the overall biodegradation of chlorophenols

The biodegradation rate of chlorophenols in the environment seems to be limited by a competitive mechanism of O-methylation which produces chloroanisoles with a high potential of being bioconcentrated in living organisms. In this work we report for the first time the isolation of three soil bacterial strains able to efficiently degrade 2,4,6-trichloroanisole (2,4,6-TCA). These strains were identified as Xanthomonas retroflexus INBB4, Pseudomonas putida INBP1 and Acinetobacter radioresistens INBS1. In these isolates 2,4,6-TCA was efficiently metabolized in a minimal medium containing methanol and 2,4,6-TCA as the only carbon sources, with a concomitant release of 3 mol of chloride ion from 1 mol of 2,4,6-TCA, indicating complete dehalogenation of 2,4,6-TCA. 2,4,6-trichlorophenol (2,4,6-TCP) was identified as a degradative intermediate, indicating that 2,4,6-TCA underwent O-demethylation as the first step in the biodegradation process. 2,4,6-TCP was further transformed into 2,6-dichloro-para-hydroquinone (2,6-DCHQ) and subsequently mineralized. The degradation of chloroanisoles could improve the overall biodegradation of chlorophenols in the environment, because those chlorophenols previously biomethylated might also be later biodegraded. Xanthomonas retroflexus INBB4 has two O-demethylation systems: one is an oxygenase-type demethylase, and the other is a tetrahydrofolate (THF)-dependent O-demethylase. On the contrary O-demethylation of 2,4,6-TCA in P. putida INBP1 is just catalysed by an oxygenase-type NADH/NADPH-dependent O-demethylase, whereas in A. radioresistens INBS1 a THF-dependent O-demethylase activity was detected.     

Purification and characterization of 2,4,6-trichlorophenol-4-monooxygenase, a dehalogenating enzyme from Azotobacter sp. strain GP1.

The enzyme which catalyzes the dehalogenation of 2,4,6-trichlorophenol (TCP) was purified to apparent homogeneity from an extract of TCP-induced cells of Azotobacter sp. strain GP1. The initial step of TCP degradation in this bacterium is inducible by TCP; no activity was found in succinate-grown cells or in phenol-induced cells. NADH, flavin adenine dinucleotide, and O2 are required as cofactors. As reaction products, 2,6-dichlorohydroquinone and Cl- ions were identified. Studies of the stoichiometry revealed the consumption of 2 mol of NADH plus 1 mol of O2 per mol of TCP and the formation of 1 mol of Cl- ions. No evidence for membrane association or for a multicomponent system was obtained. Molecular masses of 240 kDa for the native enzyme and 60 kDa for the subunit were determined, indicating a homotetrameric structure. Cross-linking studies with dimethylsuberimidate were consistent with this finding. TCP was the best substrate for 2,4,6-trichlorophenol-4-monooxygenase (TCP-4-monooxygenase). The majority of other chlorophenols converted by the enzyme bear a chloro substituent in the 4-position. 2,6-Dichlorophenol, also accepted as a substrate, was hydroxylated in the 4-position to 2,6-dichlorohydroquinone in a nondehalogenating reaction. NADH and O2 were consumed by the pure enzyme also in the absence of TCP with simultaneous production of H2O2. The NH2-terminal amino acid sequence of TCP-4-monooxygenase from Azotobacter sp. strain GP1 revealed complete identity with the nucleotide-derived sequence from the analogous enzyme from Pseudomonas pickettii and a high degree of homology with two nondehalogenating monooxygenases. The similarity in enzyme properties and the possible evolutionary relatedness of dehalogenating and nondehalogenating monooxygenases are discussed.     

Reductive dechlorination of chlorophenols by a pentachlorophenol- acclimated methanogenic consortium.

Anaerobic digester sludge fed 5,300 mg of acetate per liter, 3.4 microM pentachlorophenol, and nutrients for 10 days biotransformed pentachlorophenol by sequential ortho dechlorinations to produce 2,3,4,5-tetrachlorophenol and 3,4,5-trichlorophenol. Upon acclimation to 3.4 microM pentachlorophenol for 6 months, the methanogenic consortium removed chlorines from the ortho, meta, and para positions of pentachlorophenol and its reductive dechlorination products. Pentachlorophenol was degraded to produce 2,3,4,5-tetrachlorophenol, 2,3,4,6-tetrachlorophenol, and 2,3,5,6-tetrachlorophenol. Dechlorination of 2,3,4,5-tetrachlorophenol produced 3,4,5-trichlorophenol, which was subsequently degraded to produce 3,4-dichlorophenol and 3,5-dichlorophenol. 2,3,4,6-Tetrachlorophenol was dechlorinated at the ortho and meta positions to produce 2,4,6-trichlorophenol and 2,4,5-trichlorophenol. 2,3,5,6-Tetrachlorophenol yielded 2,3,5-trichlorophenol, followed by production of 3,5-dichlorophenol. 2,4,6-Trichlorophenol was degraded to form 2,4-dichlorophenol, and 2,4,5-trichlorophenol was dechlorinated at two positions to form 2,4-dichlorophenol and 3,4-dichlorophenol. Of the three dichlorophenols produced (2,4-dichlorophenol, 3,4-dichlorophenol, and 3,5-dichlorophenol), only 2,4-dichlorophenol was degraded significantly within 3 weeks, to produce 4-chlorophenol.     

Utilization of d-carnitine by Pseudomonas sp. AK 1

Abstract The degradation of chlorophenols by Alcaligenes eutrophus JMP134 (pJP4) was studied. The strain grew on 2,4,6-trichlorophenol or 2,4,6-tribromophenol as the sole carbon and energy source. Complete degradation of 2,4,6-trichlorophenol was confirmed by chloride release and gas chromatography analysis of supernatants from growth cultures. The 2,3,5-, 2,3,4-, 2,3,6-and 2,4,5-isomers of trichlorophenol did not support growth. However, up to 40% of 2,4,5-trichlorophenol was mineralized during growth of A. eutrophus on chemostats fed with either phenol (0.4 mM) or 2,4,6-trichlorophenol (0.4 mM) plus 2,4,5-trichlorophenol (0.1 mM). Growth on 2,4,6-trihalophenols was also observed in A. Eutrophus JMP222, the strain lacking pJP4, suggesting that this new degradative ability reported for A. eutrophus is not related to pJP4 encoded catabolic functions.     

Reductive dechlorination of chlorophenols by a pentachlorophenol- acclimated methanogenic consortium.

Anaerobic digester sludge fed 5,300 mg of acetate per liter, 3.4 microM pentachlorophenol, and nutrients for 10 days biotransformed pentachlorophenol by sequential ortho dechlorinations to produce 2,3,4,5-tetrachlorophenol and 3,4,5-trichlorophenol. Upon acclimation to 3.4 microM pentachlorophenol for 6 months, the methanogenic consortium removed chlorines from the ortho, meta, and para positions of pentachlorophenol and its reductive dechlorination products. Pentachlorophenol was degraded to produce 2,3,4,5-tetrachlorophenol, 2,3,4,6-tetrachlorophenol, and 2,3,5,6-tetrachlorophenol. Dechlorination of 2,3,4,5-tetrachlorophenol produced 3,4,5-trichlorophenol, which was subsequently degraded to produce 3,4-dichlorophenol and 3,5-dichlorophenol. 2,3,4,6-Tetrachlorophenol was dechlorinated at the ortho and meta positions to produce 2,4,6-trichlorophenol and 2,4,5-trichlorophenol. 2,3,5,6-Tetrachlorophenol yielded 2,3,5-trichlorophenol, followed by production of 3,5-dichlorophenol. 2,4,6-Trichlorophenol was degraded to form 2,4-dichlorophenol, and 2,4,5-trichlorophenol was dechlorinated at two positions to form 2,4-dichlorophenol and 3,4-dichlorophenol. Of the three dichlorophenols produced (2,4-dichlorophenol, 3,4-dichlorophenol, and 3,5-dichlorophenol), only 2,4-dichlorophenol was degraded significantly within 3 weeks, to produce 4-chlorophenol.     

Removal of chlorophenols from synthetic solutions using Phanerochaete chrysosporium

The potential use of the fungus Phanerochaete chrysosporium to remove chlorophenols (phenol, o-chlorophenol, p-chlorophenol and 2,4,6-trichlorophenol) from aqueous solutions was evaluated. The kinetics of both adsorption and desorption of phenolic compounds was rapid for all adsorbates. The maximum adsorptions of phenol and chlorophenols onto the Phanerochaete chrysosporium were 1.23 mmol/g for phenol, 1.49 mmol/g for o-chlorophenol, 1.78 mmol/g for p-chlorophenol and 2.14 mmol/g for 2,4,6-trichlorophenol. The affinity order was as follows: 2,4,6-trichlorophenol > p-chlorophenol > o-chlorophenol > phenol. Phenol and chlorophenols binding with Phanerochaete chrysosporium were clearly pH dependent. The adsorption of phenol and chlorophenols increased as pH increased. Desorption of phenol or chlorophenols was achieved using methanol solution (30% (v/v)). Phanerochaete chrysosporium is suitable for reuse for more than ten cycles without noticeable loss of adsorption capacity.     

Oxidative 4-dechlorination of 2,4,6-trichlorophenol catalyzed by horseradish peroxidase

 The well-known and easily available horseradish peroxidase (HRP) catalyzes the H₂O₂-dependent oxidative 4-dechlorination of the pollutant 2,4,6-trichlorophenol, which is recalcitrant to many organisms except those producing ligninases. UV-visible spectroscopy and gas chromatography-mass spectrometry identified the oxidized reaction product as 2,6-dichloro-1,4-benzoquinone. NMR and IR spectroscopic data further supported the above characterization. Experimental evidence for the elimination of HCl from the substrate was acquired by detecting the decrease in pH of the reaction mixture, and by observing the presence of the β-chlorocyclopentadienone cation fragment in the mass spectrum of 2,6-dichloro-1,4-benzoquinone. Consequently, nucleophilic attack by water on the 2,4,6-trichlorocyclohexadienone cation was proposed to give the final product. Our results indicate an oxidative dechlorination pathway catalyzed by HRP for 2,4,6-trichlorophenol, similar to that by extracellular lignin peroxidases. The relative catalytic efficiency of HRP seems higher than that of lignin peroxidases. The HRP-H₂O₂ catalytic system could be utilized in the degradation of polychlorinated phenols for industrial and biotechnological purposes. Received: 20 November 1998 / Accepted: 29 January 1999     

Removal of chlorophenols from aquatic systems using the dried and dead fungus Pleurotus sajor caju

In this study, the potential use of the fungus Pleurotus sajor caju to remove phenols (i.e., phenol, o-chlorophenol, p-chlorophenol and 2,4,6-trichlorophenol) from aqueous solutions was evaluated. Biosorption of phenol or chlorophenols reached equilibrium in 4 h. The maximum adsorptions of phenol and chlorophenols onto the Pleurotus sajor caju were 0.95 mmol/g for phenol, 1.24 mmol/g for o-chlorophenol, 1.47 mmol/g for p-chlorophenol and 1.89 mmol/g for 2,4,6-trichlorophenol. The affinity order was as follows: 2,4,6-trichlorophenol> p-chlorophenol> o-chlorophenol>phenol. Phenol and chlorophenols bindings onto Pleurotus sajor caju were clearly pH dependent. The adsorption of phenol and chlorophenols increased with increasing pH. Desorption was achieved using methanol solution (30%, v/v). Pleurotus sajor caju biomass is suitable for reuse for more than five cycles without noticeable loss of adsorption capacity.     

厌氧微生物菌群XH-1对2,4,6-三氯苯酚的降解特性

有机污染物2,4,6-三氯苯酚(2,4,6-TCP)普遍存在于地下水和河流底泥等厌氧环境中。为了探究厌氧微生物菌群XH-1对2,4,6-TCP的降解能力,本研究以2,4,6-TCP为底物,接种XH-1建立微宇宙培养体系,并以中间产物4-氯苯酚(4-CP)和苯酚为底物分别进行分段富集培养,利用高效液相色谱分析底物的降解转化,同时基于16S rRNA基因高通量测序分析微生物群落结构变化。结果表明: 2,4,6-TCP(122 μmol·L^-1)以0.15 μmol·d^-1的速率在80 d内被完全降解转化,降解中间产物分别为2,4-二氯苯酚(2,4-DCP)、4-氯苯酚和苯酚,所有中间产物最终在325 d被完全降解。高通量测序结果表明,脱卤杆菌和脱卤球菌可能驱动2,4,6-TCP还原脱氯,其中,脱卤球菌可能在4-CP的脱氯转化中发挥重要作用,并与丁酸互营菌和产甲烷菌联合作用彻底降解2,4,6-TCP。     

Enhanced dechlorination of 2,4,6-trichlorophenol by anaerobic microbial populations in the presence of ethanol

Summary A consortium of anaerobic microorganisms was grown on acetate, ethanol, glucose or methanol and dechlorinated 50 umol 2,4,6-trichlorophenol, through 2,4-dichlorophenol, to 4-chlorophenol. The highest rate of dechlorination of 2,4,6-trichlorophenol was observed when ethanol was used as a growth substrate.     

Biodegradation of phenol and chlorophenols with defined mixed culture in shake-flasks and a packed bed reactor

Pseudomonas testosteroni CPW301 degraded phenol and 4-chlorophenol simultaneously, but degradation rates of these compounds were affected by 4-chlorophenol. Phenol increased the cell concentration and therefore the degradation efficiency of 4-chlorophenol was improved. Pseudomonas solanacearum TCP114 could degrade only 2,4,6-trichlorophenol. A defined mixed culture of P. testosteroni CPW301 and P. solanacearum TCP114 could treat phenol, 4-chlorophenol, and 2,4,6-trichlorophenol completely and overcome the inhibition of substrates to other microorganisms. The degradation capacity of the packed bed reactor (PBR) was higher than that of the continuous stirred tank reactor, but the PBR was unsuitable for oxygen-sensitive microorganisms.     

Aerobic degradation of 2,4-dichlorophenoxyacetic acid and other chlorophenols by <Emphasis Type="Italic">Pseudomonas</Emphasis> strains indigenous to contaminated soil in South Africa: Growth kinetics and degradation pathway

Three indigenous pseudomonads, Pseudomonas putida DLL-E4, Pseudomonas reactans and Pseudomonas fluorescens, were isolated from chlorophenol-contaminated soil samples collected from a sawmill located in Durban (South Africa). The obtained isolates were tested for their ability to degrade chlorophenolic compounds: 2,4-dichlorophenoxyacetic acid (2,4-D), 2,4-dichlorophenol (2,4-DCP) and 2,4,6-trichlorophenol (2,4,6-TCP) in batch cultures. The isolates were found to effectively degrade up to 99.5, 98.4 and 94.0% with a degradation rate in the range of 0.67–0.99 (2,4-D), 0.57–0.93 (2,4-DCP) and 0.30–0.39 (2,4,6-TCP) mgL^–1 day^–1 for 2,4-D; 2,4-DCP and 2,4,6-TCP, respectively. The degradation kinetics model revealed that these organisms could tolerate up to 600 mg/L of 2,4-DCP. Catechol 2,3-dioxygenase activity detected in the crude cell lysates of P. putida DLL-E4 and P. reactans was 21.9- and 37.6-fold higher than catechol 1,2-dioxygenase activity assayed, suggesting a meta-pathway for chlorophenol degradation by these organisms. This is also supported by the generally high expression of C23O gene (involved in meta-pathway) relative to tfdC gene (involved in ortho-pathway) expression. Results of this study will be helpful in the exploitation of these organisms and/or their enzymes in bioremediation strategies for chlorophenol-polluted environment.     

Isolation of Pseudomonas pickettii strains that degrade 2,4,6-trichlorophenol and their dechlorination of chlorophenols.

Three strains of Pseudomonas pickettii that can grow with 2,4,6-trichlorophenol (2,4,6-TCP) as the sole source of carbon and energy were isolated from different mixed cultures of soil bacterial populations that had been acclimatized to 2,4,6-TCP. These strains released 3 mol of chloride ion from 1 mol of 2,4,6-TCP during the complete degradation of the TCP. Of these strains, P. pickettii DTP0602 in high-cell-density suspension cultures dechlorinated various chlorophenols (CPs). Cells that were preincubated with 2,4,6-TCP converted isomers of 4-CP to the corresponding chloro-p-hydroquinones, but those preincubated with 4-CP converted CPs lacking a chlorine atom(s) at the o position to isomers of chlorocatechol. The ability of DTP0602 to dechlorinate 2,4,6-TCP was induced by 2,6-dichlorophenol, 2,3,6- and 2,4,6-TCP, and 2,3,4,6-tetrachlorophenol and was repressed in the presence of succinate or glucose.     

Isolation of Pseudomonas pickettii strains that degrade 2,4,6-trichlorophenol and their dechlorination of chlorophenols.

Three strains of Pseudomonas pickettii that can grow with 2,4,6-trichlorophenol (2,4,6-TCP) as the sole source of carbon and energy were isolated from different mixed cultures of soil bacterial populations that had been acclimatized to 2,4,6-TCP. These strains released 3 mol of chloride ion from 1 mol of 2,4,6-TCP during the complete degradation of the TCP. Of these strains, P. pickettii DTP0602 in high-cell-density suspension cultures dechlorinated various chlorophenols (CPs). Cells that were preincubated with 2,4,6-TCP converted isomers of 4-CP to the corresponding chloro-p-hydroquinones, but those preincubated with 4-CP converted CPs lacking a chlorine atom(s) at the o position to isomers of chlorocatechol. The ability of DTP0602 to dechlorinate 2,4,6-TCP was induced by 2,6-dichlorophenol, 2,3,6- and 2,4,6-TCP, and 2,3,4,6-tetrachlorophenol and was repressed in the presence of succinate or glucose.     

Role of Chrysonilia sitophila in the quality of cork stoppers for sealing wine bottles

The contribution of Chrysonilia sitophila in cork stopper manufacture was studied and a simulation of the industrial processing of cork stoppers was performed. Stoppers cut from slabs where mold development was inhibited were compared with others cut from slabs colonized by C. sitophila alone or with several molds, in terms of physical properties and chemical taints. C. sitophila does not produce 2,4,6-trichloroanisole, guaiacol, or 1-octene-3-ol on cork slabs incubated for 66 days. Since some chlorophenol-related compounds contaminate cork slabs during the production processes, metabolic tests were performed to investigate the capability of molds to produce 2,4,6-trichloroanisole by methylation of 2,4,6-trichlorophenol. Degradation of 2,4,6-trichlorophenol by C. sitophila resulted in a very high level of degradation without production of 2,4,6-trichloroanisole. C. sitophila restricted growth of other molds on maturing slabs for at least 30 days. These results show that C. sitophila can be exploited by industrial producers of cork stoppers since it is able to inhibit the development of other molds and it does not produce the compounds responsible for ‘cork-taint’, even in the presence of chlorophenols. Journal of Industrial Microbiology & Biotechnology (2000) 24, 256–261. Received 28 July 1999/ Accepted in revised form 05 January 2000