Abstract Dyes and dyestuffs find use in a wide range ofindustries but are of primary importance to textile manu-facturing. Wastewater from the textile industry can con-tain a variety of polluting substances including dyes. In-creasingly, environmental legislation is being imposed tocontrol the release of dyes, in particular azo-based com-pounds, into the environment. The ability of microorgan-isms to decolourise and metabolise dyes has long beenknown, and the use of bioremediation based technologiesfor treating textile wastewater has attracted interest.Within this review, we investigate the mechanisms bywhich diverse categories of microorganisms, such as thewhite-rot fungi and anaerobic bacterial consortia, bringabout the degradation of dyestuffs.

Introduction

Dyes and dyestuffs are widely used within the food,pharmaceutical, cosmetic, textile and leather industries.Over 100,000 commercially available dyes exist andmore than 7×105 tonnes of dyestuff are produced annu-ally (Meyer 1981; Zollinger 1987). The human healthimpact of dyes used in the food industry, especially azo dyes and their degradation products, has caused con-cern for a number of years, with legislation controllingtheir use being developed in a variety of countries (Hildenbrand et al. 1999). Increasingly, the environmen-tal and subsequent health effects of dyes released in textile industry wastewater are becoming subject to scientific scrutiny. Wastewater from the textile industryis a complex mixture of many polluting substances rang-

ing from organochlorine-based pesticides to heavy metals associated with dyes and the dyeing process(Correia et al. 1994).

During textile processing, inefficiencies in dyeing re-sult in large amounts of the dyestuff being directly lost tothe wastewater, which ultimately finds its way into theenvironment. The amount of dye lost is dependent uponthe class of dye application used, varying from only 2%loss when using basic dyes to a 50% loss when certainreactive dyes are used (for a comprehensive review ofthis area, the types of dyes found in textile effluent andcolour discharge consent limits see O’Neill et al. 1999).Due to increasingly stringent environmental legislation,the textile industry in the UK and elsewhere is seeking todevelop effective wastewater remediation technologies,especially those that allow colour removal that is largelyunaffected by conventional treatment systems (O’Neill et al. 2000). Despite the existence of a variety of chemi-cal and physical treatment processes, bioremediation oftextile effluent is still seen as an attractive solution dueto its reputation as a low-cost, environmentally friendly,and publicly acceptable treatment technology (Banat et al. 1996).

A number of biological processes such as biosorptionhave been proposed as having potential application in removal of dyes from textile wastewater (Bustard et al.1998); however, this review will focus upon the decol-ourisation and degradation of textile dyes by both mixedand axenic cultures of bacteria and fungi.

Bacterial decolourisation and degradation of textile dyes

Actinomycetes

Actinomycetes, particularly Streptomyces species, areknown to produce extracellular peroxidases that have arole in the biodegradation of lignin. These prokaryoticperoxidases are involved in the initial oxidation of ligninto produce various water-soluble polymeric compounds.

G. McMullan (✉ ) · C. Meehan · A. Conneely · N. KirbyI.M. Banat · R. MarchantSchool of Environmental Studies, University of Ulster, Coleraine,County Londonderry, BT52 1SA, UKe-mail: g.mcmullan@ulst.ac.ukTel.: +44-28-70324755, Fax: +44-28-70324911

T. Robinson · P. Nigam · W.F. SmythSchool of Biomedical Sciences, University of Ulster, Coleraine,County Londonderry, BT52 1SA, UK

Appl Microbiol Biotechnol (2001) 56:81–87DOI 10.1007/s002530000587

M I N I - R E V I E W

G. McMullan · C. Meehan · A. Conneely · N. KirbyT. Robinson · P. Nigam · I. M. Banat · R. MarchantW. F. Smyth

Microbial decolourisation and degradation of textile dyes

Received: 12 October 2000 / Received revision: 22 November 2000 / Accepted: 24 November 2000 / Published online: 19 May 2001© Springer-Verlag 2001

Actinomycetes have also been shown to catalyse hydroxylation, oxidation, and dealkylation reactionsagainst various xenobiotic compounds (Goszczynski et al. 1994).

The ability of actinomycetes to decolourise and min-eralise textile dyes was initially investigated by threegroups. In 1989, Ball et al. screened 20 strains of actino-mycetes, representing a wide range of genera, for their ability to decolourise the polymeric dye Poly R(Ball et al. 1989). Only three of the 20 strains were ob-served to significantly decolourise the dye: Streptomycesbadius 252, Streptomyces sp. strain EC22, and Thermo-monospora fusca MT800. Subsequently, Zhou and Zimmermann (1993) embarked on an even larger screen-ing process in which the decolourising capabilities of159 actinomycetes were investigated. Of particular inter-est in this study was the investigators’ use of actual textile effluents in the screening process. Five separateeffluents each containing a single dye of known concen-tration were used. Each dye was structurally distinct,ranging from the azo compound Reactive Red 147 to thephthalocyanine Reactive Blue 116. The widespread ability of actinomycetes to bring about dye decolourisat-ion was demonstrated by positive results being obtainedfor 83 of the isolates. The finding that actinomycetes arecapable of the aerobic decolourisation and degradationof azo dyes was significant given the recalcitrance of thecompounds to degradation by other bacteria under suchconditions.

Finally, but most significantly, a group based at theUniversity of Idaho and led by Don Crawford initiatedan investigation into the ability of ligninolytic microbes,both white-rot fungi and Streptomycetes, to mineraliseand decolourise textile dyes. Initially, 14 streptomyceteswere investigated for their ability to decolourise twopolymeric dyes, Poly B-411 and Poly R-478, as well asthe azo dye Remazol Brilliant Blue R (RBBR) (Pasti andCrawford 1991). With two of the dyes, RBBR and PolyB-411, identical results were essentially obtained, with astrong correlation between the ability of the isolate todecolourise dyes and its ligninolytic capability. This ob-servation, coupled with their finding that enhanced dyedecolourisation could be achieved as a result of extracel-lular H2O2 production when strains were grown in thepresence of glucose, suggested the involvement of per-oxidases in the decolourisation process. Previously, ex-tracellular peroxidases had been shown to be producedby Streptomyces species and the enzymes were shown tohave substrate specificities similar to the Mn(II)-peroxi-dase of P. chrysosporium (Pasti and Crawford 1991). In-terestingly, with the third dye, Poly R-478, no correlationbetween decolourising activity and ligninolytic activity(as measured by oxidation of veratryl alcohol, utilisationof syringic or coumaric acid, and acid-precipitable poly-meric lignins) was observed. The enzymatic processesinvolved in the decolourisation of this dye remain unex-plained.

Further work was carried out by Crawford’s group toinvestigate the mechanism by which Streptomyces

chromofuscus A11 decolourised and mineralised azodyes. Initial work confirmed that decolourisation was re-lated to the ligninolytic capabilities of the isolate but thatthe bacterial enzymatic systems responsible for degrada-tion of azo dyes had a different specificity than those of the white-rot fungus P. chrysosporium (Paszczynski et al. 1992). Subsequently, the actual decolourisation and degradation pathway for two azo dyes by both S.chromofuscus A11 and P. chrysosporium was elucidatedin a series of elegant experiments (Goszczynski et al.1994). It was proposed that the peroxidases of both organisms converted the azo dye to a cation radical thatwas susceptible to the nucleophilic attack of water or hy-drogen peroxide. This resulted in the simultaneous splitof the azo linkage both symmetrically and asymmetrical-ly to produce intermediates which subsequently undergoseveral redox reactions before producing more stable in-termediates (Fig. 1).

Recently, Burke and Crawford (1998) partially puri-fied a novel extracellular peroxidase from S. viridos-porus T7A in an effort to identify the class of peroxi-dase involved in dye decolourisation by Streptomycesspecies. The peroxidase was found to have a substratespecificity similar to that of fungal Mn-peroxidases and was not inhibited by KCN, an inhibitor of heme-peroxidases (Magnuson 1996). In addition, N-terminalamino acid sequences of the peroxidase were found toshare significant homology with a fungal Mn-peroxi-dase and actinomycete cellulases. Further purificationof the peroxidase in question is required to finally con-firm its true biochemical nature. The molecular mecha-nism by which this peroxidase is induced has also recently been investigated; with the gene oxyR, whichencodes an oxygen stress regulatory protein, apparentlyhaving some regulatory role (Ramachandran et al.2000).

Other aerobic bacteria

With the notable exception of the actinomyctetes, theisolation of bacteria capable of the aerobic decol-ourisation and mineralisation of dyes, especially sulfonated azo dyes, has proven difficult. A number ofreports exist suggesting the aerobic conversion of specific azo dyes, including one notable report on thegreening of instant chocolate puddings (Dykes et al.1994). Certain carboxylated analogues of sulfonatedazo compounds are also utilised aerobically as sole car-bon and energy source by preadapted bacteria. Indeed,oxygen-insensitive azoreductases, Orange I azoreduc-tase[NAD(P)H:1-(4′-sulfophenylazo)-4-naphthol oxi-doreductase] and Orange II azoreductase [NAD(P)H:1-(4′-sulfophenylazo)-2-naphthol oxidoreductase], werepurified and characterised from a Pseudomonas strainK24 (Zimmermann et al. 1982, 1984). Both these enzymes have now been classified as being the sameenzyme, known correctly as azobenzene reductase (EC 1.6.6.7).

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Despite this it has been argued that unequivocal evi-dence for aerobic bacterial mineralisation of these com-pounds is absent from the literature (Blhmel et al. 1998).Recently, Blhmel et al. (1998) reported the isolation ofan unidentified bacterial strain, S5 (Genbank accessionnumber AF019037), capable of utilising the model sul-phonated azo compound 4-carboxy-4′-sulfoazobenzene(CSAB) as sole carbon and energy source. Elucidation ofthe degradation pathway demonstrated that the azo link-age of CSAB undergoes an initial reductive cleavage toform 4-aminobenzoate and 4-aminobenzenesulfonate,which are subsequently metabolised by conventional aromatic catabolic pathways (Blhmel et al. 1998). At-tempts to characterise the enzyme responsible for theazo-bond cleavage in crude cell extracts have so farproven unsuccessful.

In addition to azo dyes, the ability of bacteria to aero-bically metabolise other dye classes has also attracted in-terest but yielded little success. Recently, however,Sarnaik and Kanekar (1999) described the aerobic min-eralisation of the triphenylmethane dye, methyl violet,by a strain of Pseudomonas mendocina MCM B-402.Methyl violet, which has some commercial applicationsin addition to its recognised use as a bacteriological andhistological stain, was used by the isolate as sole carbonand energy source. Preliminary studies identified thatP. mendocina degraded the dye via a number of uniden-tified metabolites to phenol, which then entered the β-ketoadipic acid pathway.

Anaerobic bacterial decolourisation and degradation of textile dyes

Under anaerobic conditions many bacteria have been re-ported to readily decolourise azo-dyes. Initially, the bac-teria bring about the reductive cleavage of the azo link-age, which results in dye decolourisation and the pro-duction of colourless aromatic amines. The potentialtoxicity, mutagenicity, and carcinogenicity of such com-pounds is well-documented and has been reviewed else-where (Chung et al. 1992). Whilst concerns have longbeen expressed over the threat that such compoundshave on human health, their environmental impact isnow also troublesome. Baughmann and Weber (1994)demonstrated that in anoxic sediment environments un-charged azo dyes readily undergo biologically (and pre-sumably microbial)-mediated reduction to the corre-sponding amines.

The initial step in bacterial azo dye metabolism un-der anaerobic conditions involves the reductive cleavageof the azo linkage. This process is catalysed by a varietyof soluble cytoplasmic enzymes with low-substratespecificity which are known trivially as “azoreduc-tases”. Under anoxic conditions, these enzymes facili-tate the transfer of electrons via soluble flavins to theazo dye, which is then reduced. The role that such cyto-plasmic enzymes have in vivo is, however, uncertain.Reports indicate that intestinal bacteria decolourise cer-tain azo dyes and their polymeric derivatives at roughly

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Fig. 1 Proposed pathway forperoxidase-catalysed degrada-tion of sulfonated azo dyes.The compounds in parentheseshave not been found, but theirexistence is rationalised as necessary intermediates for theobserved final products. Thecompounds represented bynumbers in brackets have beenfound in reaction mixtures.Substitution pattern a (as in I),R1=R2=O and B=O; substitu-tion pattern b (as in II), R1=H,R2=OCH, and B=NH. [2a]2,6-dimethyl-1,4-benzoquinone,[4a] 4-nitrosobenzenesulfonicacid, [6b] 2-methoxyhydroqui-none, [7b] 2-methoxy-4-ami-nophenol, [8a] sulfanilic acid,[8b] sulfanilamide, [9a] 4-hy-droxybenznesulfonic acid, [9b]4-hydroxybenzenesulfonamide,[10a] benzenesulfonic acid,[10b] benzenesulfonamide,[11a] azobenzene-4,4′-disul-fonic acid, [12] ammonia. (Reproduced from Goszczynskiet al. 1994)

equivalent rates (Brown 1981). As it is unlikely thatthese polymeric dye molecules, or highly charged sul-phonated azo dyes, can actually pass through the bacte-rial cell membrane, then the possibility of non-cytoplas-mic “azoreductase” capabilities exists (Keck et al.1997). This subject will be returned to below in the sec-tion “Current state of the art”. Whilst the anaerobic re-duction of azo dyes is relatively easy to achieve, com-plete mineralisation of the molecule is difficult. Donlonet al. (1997) reported the partial mineralisation of theazo dye Mordant Orange 1 by a methanogenic granularsludge in a continuous-upflow anaerobic sludge blanket.In this study, however, complete mineralisation of onlyone of the azo cleavage products, 5-aminosalicylic acid,was possible, with the other product (1,4-phenylenedi-amine) accumulating in the reactor.

In addition to azo dyes, the bacterial metabolism ofother dye molecules under anoxic conditions has alsobeen studied. Henderson et al. (1997) demonstrated thata range of axenic bacterial cultures which are commonlyfound in the human gastro-intestinal tract, as well asconsortia of microbes from human, mouse, and rat intes-tines, all readily reduced the triphenylmethane dye mala-chite green. The metabolite produced, leucomalachitegreen, is a suspected carcinogen and raises concerns overthe continued use of malachite green in aquaculture invarious regions worldwide.

Anaerobic-aerobic biodegradation of dyes

To overcome the problem of the relative recalcitrance ofazo dye breakdown products under anoxic conditions, anumber of groups have used a two-stage treatment pro-cess (Oxspring et al. 1996; O’Neill et al. 2000). In thefirst anaerobic stage, the azo dye is readily reduced tothe corresponding colourless aromatic amines, which arethen metabolised relatively easily under aerobic condi-tions. A detailed review on the anaerobic treatment oftextile effluents can be found elsewhere (Delee et al.1998). In 1997, Kudlich et al. described the completemineralisation of the sulphonated azo dye Mordant Yellow 3 by the bacterium Sphingomonas sp. BN6 co-immobilised with an uncharacterised 5-aminosalicylate-degrading isolate in alginate beads. Although the beadswere maintained under aerobic conditions, their centresmaintained anoxic conditions and it was here that cellsof Sphingomonas sp. BN6 reductively cleaved MordantYellow 3 to produce 5-aminosalicylate and 6-amino-naphthalene-2-sulfonate. At the aerobic surface of thebeads, 6A2NS was also converted to 5AS by Sphingo-monas sp. BN6 before being mineralised by cells of theisolate 5AS1. These observations suggest that it may bepossible to develop biofilm-based reactors for the com-plete mineralisation of industrial-dye-contaminatedwastewaters.

Fungal decolourisation and degradation of textile dyes

White-rot fungi

By far the most widely studied of dye-decolourising mi-croorganisms are the white-rot fungi. This group of or-ganisms is central to the global carbon cycle as a resultof their ability to mineralise the woody plant materiallignin, which has a complex polymeric structure. In addi-tion to their natural substrate, white-rot fungi have beenfound to be capable of mineralising a diverse range ofpersistent organic pollutants, which distinguishes themfrom biodegradative bacteria that tend to be rather sub-strate-specific (Reddy 1995). The ability of these fungito degrade such a range of organic compounds resultsfrom the relatively non-specific nature of their lignino-lytic enzymes, such as lignin peroxidase (LiP), manga-nese peroxidase (MnP) (EC 1.11.1.13), and laccase.These enzymes and their catalytic properties are re-viewed elsewhere (Hattaka 1994), but briefly, LiP cata-lyses the oxidation of non-phenolic aromatic compoundssuch as veratryl alcohol, whilst MnP oxidises Mn2+ toMn3+ which is able to oxidise many phenolic compounds(Glenn et al. 1986). Laccase (benzenediol:oxygen oxido-reductase, EC 1.10.3.2) is a copper-containing enzymethat catalyses the oxidation of phenolic substrates bycoupling it to the reduction of oxygen to water (Edens et al. 1999).

The decolourisation of dyes by white-rot fungi wasfirst reported by Glenn and Gold (1983), who developeda method to measure ligninolytic activity of Phaner-ochaete chrysosporium based upon the decolourisationof a number of sulfonated polymeric dyes. Subsequently,the decolourisation of dyes has been used by others torapidly assess the biodegradative capabilities of diversewhite-rot fungi (Field et al. 1993). In 1990, the first re-port appeared in the literature of sulfonated azo dyes be-ing decolourised, again by P. chrysosporium (Cripps et al. 1990), and a degradation pathway for this isolatewas elucidated (Goszczynski et al. 1994; Figure 1).Whilst P. chrysosporium remains the most widely stud-ied of white-rot fungi, Trametes (Coriolus) versicolor,Bjerkandera adusta, Pleurotus and Phlebia species, aswell as a variety of other isolates are increasingly beingstudied (Conneely et al. 1999; Heinfling et al. 1998; Kirby et al. 2000; Pointing et al. 2000; Swamy and Ramsay 1999).

Unfortunately, due to the inherent complexity both ofthe dye molecules themselves and the enzymatic mecha-nisms involved, the degradative pathways utilised bywhite-rot fungi other than P. chrysosporium remain un-studied, despite the exploitation of powerful analyticaltechniques (Smyth et al. 1999). Recently, our own groupbegan attempts to elucidate the degradation pathway ofcopper-phthalocyanine dyes by P. chrysosporium usingsuch analytical tools as HPLC, atomic absorption spec-trometry, and polarography (Conneely et al. 1999). Ini-tial data indicated that the dye structure was readily de-

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graded, resulting in decolourisation; however, both freecopper and organo-copper breakdown products werefound in culture supernatants and further work is neededto elucidate their identity.

Whilst it is clear that enzymes such as LiP, MnP, andlaccase play a significant role in dye metabolism bywhite-rot fungi, care must be taken not to exclude thepossibility of the existence of other degradative mecha-nisms. In many early studies, dyes were only added toculture media when conditions for the production of lig-ninolytic enzymes prevailed, thus excluding the potentialdiscovery of other, non-ligninolytic, processes, (Crippset al. 1990). For example, Pasti and Crawford (1991)have suggested the existence of alternative systems orenzymes in dye decolourisation by white-rot fungi. Theplasma membrane redox system of white-rot fungi hasbeen proposed as a potential mechanism for dye decol-ourisation, and it is interesting to draw parallels betweenthis and the redox-mediated mechanisms utilised by bac-teria under anaerobic conditions. Supportive evidencewas obtained by Stahl and Aust (1993), who found thatsuch a mechanism was responsible for the degradation of2,4,6-trinitrotoluene in P. chrysosporium. Kirby (1999),however, found little evidence to suggest that such amechanism was involved in decolourisation of RemazolBlack B by strains of P. chrysosporium, T. versicolor, orPhlebia tremellosa.

Vyas and Molitoris (1995) discovered that Pleurotusostreatus produced an unusual enzyme during the solid-state fermentation of wheat straw that was capable ofRemazol Brilliant Blue R (RBBR) decolourisation. Thisactivity was distinct from the MnP, laccase, and manga-nese-independent peroxidase activities also produced bythe isolate, and from LiP and veratryl alcohol oxidaseactivities, which were not detectable in P. ostreatus butare well-known in other white-rot fungi. The activitywas named RBBR oxygenase as it possessed a catalyticmetal centre (possiblely haeme) and was inhibited by a number of known oxygenase inhibitors (Vyas and Molitoris 1995). The work of Vyas and Molitoris is alsoof note as it documents a phenomenon seen by many researchers of white-rot fungal dye decolourisation: the sequential change of blue dyes to colourless through a rainbow of intermediates (Kirby 1999). Vyas and Molitoris (1995) proposed that such observations indi-cate that decolourisation is not a single-step reaction andthat these intermediate steps can be missed if enzyme activities are high.

Non-white-rot fungi

Whilst degradation pathways utilised by these fungi are not described in the literature, it is expected that they would be similar to those reported to be involved in the metabolism of other aromatic hydrocarbons (Wunderwald et al. 1997).

Detailed investigations have been carried out on onesuch isolate, a strain of Geotrichum candidum Dec1 iso-

lated from soil and capable of decolourising a number ofanthraquinone dyes (Kim et al. 1995). The broad sub-strate specificity exhibited by this isolate led Kim andShoda (1999a) to propose the existence of an extracellu-lar peroxidase-type enzyme. Subsequently, a glycosylat-ed haeme-based peroxidase (DyP) was purified from G.candidum which had properties distinct from LiP, MnP,horseradish peroxidase, and other peroxidases previouslyreported (Kim and Shoda 1999a). The potential impor-tance of this isolate for dye decolourisation rests with itsrobustness in comparison to P. chrysosporium and otherwhite-rot fungi. DyP is produced constitutively and, un-like the activity of P.chrysosporium ligninolytic en-zymes, is unaffected by the shear forces of shake flasksor stirred tank reactors (Kim and Shoda 1998, 1999b).Such properties should allow a number of G. candidum-Dec1-based bioreactors to be developed for the treatmentof diverse organic pollutants including dyestuffs. Recent-ly the gene (dyp) encoding DyP was cloned into the het-erologous expression host Aspergillus oryzae to allowgreater protein production in a safe host (Sugano et al.2000). Molecular analysis of the protein confirmed thatDyP is distinct from other peroxidases in the plant peroxidase superfamily, having a novel haeme-bindingregion.

Current state of the art

The role of bacterial cytoplasmic and extracellular “azo-reductases” in azo dye reduction in vivo has recentlybeen clarified by investigators at the University ofStuttgart, Germany (Keck et al. 1997; Kudlich et al.1997; Russ et al. 2000). Their work focused upon thegram-negative isolate Sphingomonas sp. BN6, which hasrecently, through molecular analysis, been shown to re-present a novel species named Sphingomonas xenophagaafter its ability to “eat foreign compounds” (Stolz et al.2000). It was demonstrated that an extracellular mecha-nism of dye reduction existed in addition to the nonspe-cific cytoplasmic enzymes which functioned as “azore-ductases” by transferring electrons via reduced flavingroups to the dye molecule and thus bringing about apurely chemical reduction. Keck et al. (1997) demon-strated that certain quinone-based compounds generatedduring metabolism of specific substrates acted as media-tors shuttling redox equivalents to azo dye moleculesfrom the bacterial membrane (Keck et al. 1997; Kudlichet al. 1997) (Fig. 2). To investigate if cytoplasmic en-zymes played any role in dye decolourisation in vivo, aflavin reductase [NAD(P)H:flavin oxidoreductase] wascloned and overexpressed in both E. coli and S. xeno-phaga (Russ et al. 2000). In cell extracts the strains withoverexpressed flavin reductase demonstrated elevatedazo reductase activity; however, in whole cell studiesthese strains showed little improvement in their dye-decolourising capabilities. These findings demonstratethat the cytoplasmic azoreductases previously describedin the literature are likely to be a laboratory artefact and

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play no significant role in dye decolourisation in vivo.Finally, Russ et al. (2000) also provided evidence that re-ports of aerobic azo reductases could be explained by theisolates in which such a phenomenon was described hav-ing elevated flavin reductase activities.

Whilst many groups have demonstrated the ability ofwhite-rot fungi to decolourise and degrade textile dyes,the development of potential remediation solutions forthe textile industry based upon these microorganisms hasbeen slow. A range of reactor systems aimed at optimi-sing production of ligninolytic enzymes has been devel-oped, including stirred tank reactors (Linko 1988),packed-bed reactors (Feijoo et al. 1995), and rotatingdisk reactors (Kirk et al. 1986). Few of these studieshave focused upon the ability of such reactors to remedi-ate pollutants, with the exception of Allemann et al.(1995), Zhang et al. (1999), and Kapdan et al. (2000),who investigated pentachlorophenol, Orange II azo dye,and Everzol Turquoise Blue G dye degradation, respec-tively. Our own group utilised the rotating-tube bioreac-tor system described by Allemann et al. (1995) to inves-tigate the remediation of actual textile effluent by P.chrysosporium (Kirby 1999). This reactor configurationproved to be robust enough to continuously decolourisethree different mixed azo dye effluents by greater than90% over the 38-day operating period (Kirby 1999).Whilst such findings are encouraging, many problemsstill face researchers in this area due to the lack of reli-able, robust, and economic white-rot-fungi-based reac-tors. In addition, the volumes of effluent generated bymany of the larger dyeing plants worldwide needs to beconsidered, and it may be that extraction and concentra-tion of dyes and other pollutants is required before biore-mediation is feasible (Nigam et al. 2000).

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