Degradation of anaerobic reductive dechlorinationproducts of Aroclor 1242 by four aerobic bacteria

We studied the aerobic degradation of eight PCB congeners which comprise from 70 to 85% of the anaerobic dechlorination products from Aroclor 1242, including2-, 4-, 2,4-, 2,6-, 2,2'-, 2,4'-, 2,2',4-, and2,4,4'-chlorobiphenyl (CB), and the biodegradation of their mixtures designed to simulate anaerobic dechlorination profiles M and C. StrainsComamonas testosteroni VP44 and Rhodococcus erythreus NY05 preferentially oxidizeda para-substituted ring, while Rhodococcus sp. RHA1, similar to well known strain Burkholderia sp. LB400, preferably attackedan ortho-chlorinated ring. Strains with ortho-directed attack extensively degraded2,4'- and 2,4,4'-CB into 4-chlorobenzoate, while bacteria with para-directed attack transformed these congeners mostly into potentially problematicmeta-cleavage products. The strains that preferentiallyoxidized an ortho-substituted ring readily degradedseven of the eight congeners supplied individually; only 2,6-CB was poorly degraded. Degradationof 2,2'- and 2,4,4'-CB was reduced when present in mixtures M and C. Higher efficiencies of degradation of the individual congeners and defined PCB mixtures M and C and greater production of chlorobenzoates were observed with bacteria that preferentially attackan ortho-substituted ring. PCB congeners 2,4'-, 2,2',4-, and 2,4,4'-CB canbe used to easily identify bacteria with ortho-directed attack whichare advantageous for use in the aerobic stage of the two-phase (anaerobic/aerobic)PCB bioremediation scheme.     

Dechlorination of polychlorinated biphenyl congeners by an anaerobic microbial consortium

  An anaerobic methanogenic microbial consortium, developed in a granular form, exhibited extensive dechlorination of defined polychlorinated biphenyl (PCB) congeners. A 2,3,4,5,6-pentachlorobiphenyl was dechlorinated to biphenyl via 2,3,4,6-tetrachlorobiphenyl, 2,4,6-trichlorobiphenyl, 2,4-dichlorobi-phenyl and 2-chlorobiphenyl (CB). Removal of chlorine atoms from all three positions of the biphenyl ring, i.e., ortho, meta and para, was observed during this reductive dechlorination process. Biphenyl was identified as one of the end-products of the reductive dechlorination by GC-MS. After 20 weeks, the concentrations of the dechlorination products 2,4,6-CB, 2,4-CB, 2-CB and biphenyl were 8.1, 41.2, 3.0 and 47.8 μM respectively, from an initial 105 μM 2,3,4,5,6-CB. The extent and pattern of the dechlorination were further confirmed by the dechlorination of lightly chlorinated congeners including 2-CB, 3-CB, 4-CB, 2,4-CB and 2,6-CB individually. This study indicates that the dechlorination of 2,3,4,5,6-CB to biphenyl is due to ortho, meta and para dechlorination by this anaerobic microbial consortium. Received: 30 April 1996 / Received revision: 26 July 1996 / Accepted: 5 August 1996     

Aerobic degradation of polychlorinated biphenyls by Alcaligenes sp. JB1: metabolites and enzymes

In contrast to the degradation of penta-and hexachlorobiphenyls in chemostat cultures, the metabolism of PCBs by Alcaligenes sp. JB1 was shown to be restricted to PCBs with up to four chlorine substituents in resting-cell assays. Among these, the PCB congeners containing ortho chlorine substituents on both phenyl rings were found to be least degraded. Monochloro-benzoates and dichlorobenzoates were detected as metabolites. Resting cell assays with chlorobenzoates showed that JB1 could metabolize all three monochlorobenzoates and dichlorobenzoates containing only meta and para chlorine substituents, but not dichlorobenzoates possessing an ortho chlorine substituent. In enzyme activity assays, meta cleaving 2,3-dihydroxybiphenyl 1,2-dioxygenase and catechol 2,3-dioxygenase activities were constitutive, whereas benzoate dioxygenase and ortho cleaving catechol 1,2-dioxygenase activities were induced by their substrates. No activity was found for pyrocatechase II, the enzyme that is specific for chlorocatechols. The data suggest that complete mineralization of PCBs with three or more chlorine substituents by Alcaligenes sp. JB1 is unlikely.Abbreviations PCB polychlorinated biphenyls - CBA chlorobenzoate - D di - Tr tri - Te tetra - Pe penta- - H hexa     

Genotoxic effects of PCB 52 and PCB 77 on cultured human peripheral lymphocytes

Polychlorinated biphenyls (PCBs) are known to be carcinogenic, but the mechanisms of this action are uncertain. Most, but not all, studies have concluded that PCBs are not directly mutagenic, and that much if not all of the carcinogenic activity resides in the fraction of the PCB mixture that contains congeners with dioxin-like activity. The present study was designed to determine genotoxic effects of an ortho-substituted, non-coplanar congener, 2,2′,5,5′-tetrachlorobiphenyl (PCB 52), and a non-ortho-substituted coplanar congener with dioxin-like activity, 3,3′,4,4′-tetrachlorobiphenyl (PCB 77) on cultured human peripheral lymphocytes. DNA damage was assessed by use of the comet assay (alkaline single-cell gel electrophoresis). After cell cultures were prepared, test groups were treated with different concentrations of PCB 52 (0.2 and 1 μM) and PCB 77 (1 and 10 μM) for 1 h at 37 °C in a humidified carbon dioxide incubator, and compared to a DMSO vehicle control group. The cells were visually classified into four categories on the basis of extent of migration such as undamaged (UD), low damage (LD), moderate damage (MD) and high damage (HD). The highest concentration of PCBs 52 and 77 significantly increased DNA breakage in human lymphocytes (p < 0.001). Our results indicate that both the non-coplanar PCB 52 and coplanar PCB 77 cause DNA damage, and that the ortho-substituted congener was significantly more potent than the dioxin-like coplanar congener.     

Degradation of PCB congeners by bacterial strains

Biological in situ methods are options for the remediation of contaminated sites. An approach to quantify biodegradation by soil bacteria was developed, combining experiment with mathematical modelling. We performed in vitro assays to investigate the potential and kinetics of the wild-type degrader, Burkholderia sp. strain LB400 (expressing bph) and the genetically modified Pseudomonas fluorescens strains F113pcb and F113L::1180 (expressing bph under different promoters) to metabolise individual congeners of polychlorinated biphenyls (PCBs). Kinetics of metabolism was analysed using the Monod model. Results revealed similar patterns of degradable PCB congeners for LB400 and F113L::1180. The degree of PCB degradation was comparable for LB400 and F113L::1180 but was much lower for F113rifpcb. In additional mesocosm experiments with PCB-contaminated soil, the F113 derivatives demonstrated a good survival ability in willow (Salix sp.) rhizosphere. Strain F113L::1180 in combination with willow plants is expected to degrade a large spectrum of PCB congeners in soil. The data from the experiments were used to calculate the time scale of the degradation process in a PCB-contaminated soil. The uncertainty of the model predictions due to the uncertainties of experimental removal velocities and bacterial cell density in soil was quantified.     

Degradation of polychlorinated biphenyl (PCB) by a consortium obtained from a contaminated soil composed of Brevibacterium,Pandoraea and Ochrobactrum

An indigenous polychlorinated biphenyl (PCB)-degrading bacterial consortium was obtained from soils contaminated by transformer oil with a high content of PCBs. The PCB degrader strains were isolated and identified as Brevibacterium antarcticum, Pandoraea pnomenusa, and Ochrobactrum intermedium by 16S rRNA gene sequence phylogenetic analysis. The PCB-degrading ability of the consortium and of individual strains was determined by using GC/MS. The PCB-degrading capacities of the consortium were evaluated for three concentrations of transfomer oil ranging from 55 to 152 μM supplemented with 0.001% biphenyl and 0.1% of Tween 80 surfactant. PCB biodegradation by the consortium was favored in the presence of both additives and the greatest extent of biodegradation (67.5%) was obtained at a PCB concentration of 55 μM. Each bacterial species exhibited a particular pattern of degradation relating to specific PCB congeners. Isolated strains showed a moderate degradation capability towards tetra-, hepta-, and octa-chlorobiphenyls; although no effect on penta-, hexa-, and nona-chlorobiphenyls was observed. Recently, PCB degradation capacity was recognized in a Pandorea member; however, this is the first study that describes the ability of Brevibacterium and Ochrobactrum species to degrade PCBs.     

Catabolic potential of multiple PCB transformation systems in Rhodococcus sp. strain RHA1

Summary Polychlorinated biphenyl (PCB) transformation activity of a strong PCB degrader, Rhodococcus sp. strain RHA1, was examined in different concentrations of PCBs. A extremely strong PCB transformation activity was observed on 30 g PCB/ml. At 50 and 100 g/ml, transformation activities were diminished. In the case of bphA insertion mutant, RDA1, transformation activity in the presence of ethylbezene was poor even at 30 g/ml. This indicated that the bphA dependent system would play a major role in PCB transformation by RHA1. Greater transformation activity of RHA1 was observed in the presence of ethylbenzene than in the presence of biphenyl.     

Dechlorination of Fenclor 54 and of a synthetic mixture of polychlorinated biphenyls by anaerobic microorganisms

Microorganisms obtained from a contaminated experimental soil were found to reductively dechlorinate the polychlorobiphenyls (PCBs) of ex-commercial Fenclor 54 and of a synthetic mixture of single congeners, under laboratory anaerobic conditions. The dechlorination rate and extent tended to increase as the chlorination degree of F 54 congeners increased. Several penta-chlorinated congeners temporarily accumulated during the final period of incubation. Dechlorination occurred primarily from the meta and para positions while ortho-sustituted congeners accumulated in the medium during incubation. The dechlorination pattern observed with these unacclimated microorganisms in both PCB mixtures could be only partially compared to patterns reported in the literature. The low product yield deriving from reductive dechlorination of PCBs, i.e. di-and tri-chlorinated biphenyls, and the slow rate of PCB biotransformation can be attributed to a lower dehalogenation capability of artificially contaminated soil microorganisms and, perhaps, also to the inadequacy of the adopted anaerobic medium. Correspondence to: F. Fava     

Fungal bioconversion of toxic polychlorinated biphenyls by white-rot fungus, Phlebia brevispora

Toxic coplanar polychlorinated biphenyls (Co-PCBs) were used as substrates for a degradation experiment with white-rot fungus, Phlebia brevispora TMIC33929, which is capable of degrading polychlorinated dibenzo-p-dioxins. Eleven PCB congener mixtures (7 mono-ortho- and 4 non-ortho-PCBs) were added to the cultures of P. brevispora and monitored by high resolution gas chromatography and mass spectrometry (HRGC/HRMS). Five PCB congeners, 3,3′,4,4′-tetrachlorobiphenyl, 2,3,3′,4,4′-pentachlorobiphenyl, 2,3′,4,4′,5-pentachlorobiphenyl, 3,3′,4,4′,5-pentachlorobiphenyl, and 2,3′,4,4′,5,5′-hexachlorobiphenyl were degraded by P. brevispora. To investigate the fungal metabolism of PCB, each Co-PCB was treated separately by P. brevispora and the metabolites were analyzed by gas chromatography and mass spectrometry (GC/MS) and identified on the basis of the GC/MS comparison with the authentic compound. Meta-methoxylated metabolite was detected from the culture containing each compound. Additionally, para-dechlorinated and -methoxylated metabolite was also detected from the culture with 2,3,3′,4,4′-pentachlorobiphenyl, 2,3′,4,4′,5-pentachlorobiphenyl, and 2,3′,4,4′,5,5′-hexachlorobiphenyl, which are mono-ortho-PCBs. In this paper, we identified the congener specific degradation of coplanar PCBs by P. brevispora, and clearly proved for the first time by identifying the metabolites that the white-rot fungus, P. brevispora, transformed recalcitrant coplanar PCBs.     

Microbial Reductive Dechlorination of Aroclor 1260 in Baltimore Harbor Sediment Microcosms Is Catalyzed by Three Phylotypes within the Phylum Chloroflexi

The specific dechlorination pathways for Aroclor 1260 were determined in Baltimore Harbor sediment microcosms developed with the 11 most predominant congeners from this commercial mixture and their resulting dechlorination intermediates. Most of the polychlorinated biphenyl (PCB) congeners were dechlorinated in the meta position, and the major products were tetrachlorobiphenyls with unflanked chlorines. Using PCR primers specific for the 16S rRNA genes of known PCB-dehalogenating bacteria, we detected three phylotypes within the microbial community that had the capability to dechlorinate PCB congeners present in Aroclor 1260 and identified their selective activities. Phylotype DEH10, which has a high level of sequence identity to Dehalococcoides spp., removed the double-flanked chlorine in 234-substituted congeners and exhibited a preference for para-flanked meta-chlorines when no double-flanked chlorines were available. Phylotype SF1 had similarity to the o-17/DF-1 group of PCB-dechlorinating bacteria. Phylotype SF1 dechlorinated all of the 2345-substituted congeners, mostly in the double-flanked meta position and 2356-, 236-, and 235-substituted congeners in the ortho-flanked meta position, with a few exceptions. A phylotype with 100% sequence identity to PCB-dechlorinating bacterium o-17 was responsible for an ortho and a double-flanked meta dechlorination reaction. Most of the dechlorination pathways supported the growth of all three phylotypes based on competitive PCR enumeration assays, which indicates that PCB-impacted environments have the potential to sustain populations of these PCB-dechlorinating microorganisms. The results demonstrate that the variation in dechlorination patterns of congener mixtures typically observed at different PCB impacted sites can potentially be mediated by the synergistic activities of relatively few dechlorinating species.     

Substrate selectivity pattern of Comamonas testosteroni strain B-356 towards dichlorobiphenyls

In this work, we have investigated the substrate selectivity pattern of strain B-356 resting cell suspensions and cell lysates towards selected chlorobiphenyl congeners. The strain showed a preference for the double meta-substituted congener 3,3′-dichlorobiphenyl over the double ortho-substituted congener 2,2′-dichlorobiphenyl and the double para-substituted congener 4,4′-dichlorobiphenyl. The results are discussed with reference to the substrate selectivity pattern reported for Pseudomonas sp. strain LB400.     

Establishment of a Polychlorinated Biphenyl-Dechlorinating Microbial Consortium, Specific for Doubly Flanked Chlorines, in a Defined, Sediment-Free Medium

Estuarine sediment from Charleston Harbor, South Carolina, was used as inoculum for the development of an anaerobic enrichment culture that specifically dechlorinates doubly flanked chlorines (i.e., chlorines bound to carbon that are flanked on both sides by other chlorine-carbon bonds) of polychlorinated biphenyls (PCBs). Dechlorination was restricted to the para chlorine in cultures enriched with 10 mM fumarate, 50 ppm (173 μM) 2,3,4,5-tetrachlorobiphenyl, and no sediment. Initially the rate of dechlorination decreased upon the removal of sediment from the medium. However, the dechlorinating activity was sustainable, and following sequential transfer in a defined, sediment-free estuarine medium, the activity increased to levels near that observed with sediment. The culture was nonmethanogenic, and molybdate, ampicillin, chloramphenicol, neomycin, and streptomycin inhibited dechlorination activity; bromoethanesulfonate and vancomycin did not. Addition of 17 PCB congeners indicated that the culture specifically removes double flanked chlorines, preferably in the para position, and does not attack ortho chlorines. This is the first microbial consortium shown to para or meta dechlorinate a PCB congener in a defined sediment-free medium. It is the second PCB-dechlorinating enrichment culture to be sustained in the absence of sediment, but its dechlorinating capabilities are entirely different from those of the other sediment-free PCB-dechlorinating culture, an ortho-dechlorinating consortium, and do not match any previously published Aroclor-dechlorinating patterns.     

Brominated Biphenyls Prime Extensive Microbial Reductive Dehalogenation of Aroclor 1260 in Housatonic River Sediment

The upper Housatonic River and Woods Pond (Lenox, Mass.), a shallow impoundment on the river, are contaminated with polychlorinated biphenyls (PCBs), the residue of partially dechlorinated Aroclor 1260. Certain PCB congeners have the ability to activate or “prime” anaerobic microorganisms in Woods Pond sediment to reductively dehalogenate the Aroclor 1260 residue. We proposed that brominated biphenyls might have the same effect and tested the priming activities of 14 mono-, di-, and tribrominated biphenyls (350 μM) in anaerobic microcosms of sediment from Woods Pond. All of the brominated biphenyls were completely dehalogenated to biphenyl, and 13 of them primed PCB dechlorination. Measured in terms of chlorine removal and decrease in the proportion of hexa- through nonachlorobiphenyls, the microbial PCB dechlorination primed by several brominated biphenyls was nearly twice as effective as that primed by chlorinated biphenyls. Congeners containing a meta bromine primed Dechlorination Process N (flanked meta dechlorination), and congeners containing an unflanked para bromine primed Dechlorination Process P (flanked para dechlorination). Two ortho-substituted congeners, 2-bromobiphenyl and 2,6-dibromobiphenyl (2-BB and 26-BB), also primed Process N dechlorination. The most effective primers were 26-BB, 245-BB, 25-3-BB, and 25-4-BB. The microbial dechlorination primed by 26-BB converted ~75% of the hexa- through nonachlorobiphenyls to tri- and tetrachlorobiphenyls in 100 days and removed ~75% of the PCBs that are most persistent in humans. These results represent a major step toward identifying an effective method for accelerating PCB dechlorination in situ. The challenge now is to identify naturally occurring compounds that are safe and effective primers.     

Repeated application of carvone-induced bacteria to enhance biodegradation of polychlorinated biphenyls in soil

Carvone, the principal component of spearmint oil, induces biodegradation of polychlorinated biphenyls (PCB) by Arthrobacter sp. strain B1B. This study investigated the effectiveness of the repeated application of carvone-induced bacteria for bioremediation of Aroclor-1242-contaminated soil. Control treatments compared a single inoculation of carvone-induced cells, repeated applications of noninduced cells, and repeated applications of cell-free carvone/fructose medium. The results showed that repeated application of carvone-induced bacteria was the most effective treatment for mineralizing PCB, resulting in 27 ± 6% degradation of Aroclor 1242 after 9 weeks; whereas a single application of cells resulted in no significant degradation. Addition of cell-free, carvone/fructose medium resulted in 10% degradation of PCB, which suggests that this treatment stimulated biodegradation of PCB by the indigenous microflora. The di- and trichlorobiphenyls were the most readily degraded congeners. More highly chlorinated congeners, which had been previously shown to be degraded in liquid culture, were not substantially degraded in soil, indicating that low bioavailability may have limited their degradation. With the development of new technology, which permits automated in situ fermentation and delivery of degrader microorganisms, the repeated application of carvone-induced bacteria may facilitate bioremediation of PCB-contaminated soils. Received: 7 January 1998 / Received revision: 18 June 1998 / Accepted: 27 June 1998     

Isolation and characterization of a novel polychlorinated biphenyl-degrading bacterium, Paenibacillus sp. KBC101

The biphenyl-utilizing bacterial strain KBC101 has been newly isolated from soil. Biphenyl-grown cells of KBC101 efficiently degraded di- to nonachlorobiphenyls. The isolate was identified as Paenibacillus sp. with respect to its 16S rDNA sequence and fatty acid profiles, as well as various biological and physiological characteristics. In the case of highly chlorinated biphenyl (polychlorinated biphenyl; PCB) congeners, the degradation activities of this strain were superior to those of the previously reported strong PCB degrader, Rhodococcus sp. RHA1. Recalcitrant coplanar PCBs, such as 3,4,3,4-CB, were also efficiently degraded by strain KBC101 cells. This is the first report of a representative of the genus Paenibacillus capable of degrading PCBs. In addition to growth on biphenyl, strain KBC101 could grow on dibenzofuran, xanthene, benzophenone, anthrone, phenanthrene, naphthalene, fluorene, fluoranthene, and chrysene as sole sources of carbon and energy. Paenibacillus sp. strain KBC101 presented heterogeneous degradation profiles toward various aromatic compounds.     

Anaerobic Dehalogenation of Hydroxylated Polychlorinated Biphenyls by Desulfitobacterium dehalogenans

Ten years after reports on the existence of anaerobic dehalogenation of polychlorinated biphenyls (PCBs) in sediment slurries, we report here on the rapid reductive dehalogenation of para-hydroxylated PCBs (HO-PCBs), the excreted main metabolites of PCB in mammals, which can exhibit estrogenic and antiestrogenic activities in humans. The anaerobic bacterium Desulfitobacterium dehalogenans completely dehalogenates all flanking chlorines (chlorines in ortho position to the para-hydroxyl group) from congeners such as 3,3′,5,5′-tetrachloro-4,4′-dihydroxybiphenyl.     

Bioremediation of polychlorinated biphenyl-contaminated soil using carvone and surfactant-grown bacteria

Partial bioremediation of polychlorinated biphenyl (PCB)-contaminated soil was achieved by repeated applications of PCB-degrading bacteria and a surfactant applied 34 times over an 18-week period. Two bacterial species, Arthrobacter sp. strain B1B and Ralstonia eutrophus H850, were induced for PCB degradation by carvone and salicylic acid, respectively, and were complementary for the removal of different PCB congeners. A variety of application strategies was examined utilizing a surfactant, sorbitan trioleate, which served both as a carbon substrate for the inoculum and as a detergent for the mobilization of PCBs. In soil containing 100 μg Aroclor 1242 g⁻¹ soil, bioaugmentation resulted in 55–59% PCB removal after 34 applications. However, most PCB removal occurred within the first 9 weeks. In contrast, repeated addition of surfactant and carvone to non-inoculated soil resulted in 30–36% PCB removal by the indigenous soil bacteria. The results suggest that bioaugmentation with surfactant-grown, carvone-induced, PCB-degrading bacteria may provide an effective treatment for partial decontamination of PCB-contaminated soils. Received: 9 March 2000 / Received revision: 27 June 2000 / Accepted: 16 July 2000     

The effect of chlorination degree and substitution pattern on the interactions of polychlorinated biphenyls with model bacterial membranes

Polychlorinated biphenyls (PCB) are persistent organic pollutants that due to their chemical resistivity and inflammability found multiple applications. In spite of the global ban for PCB production, due to their long half-lives periods, PCB accumulate in the soils, so effective bioremediation of the polluted lands is of crucial importance. Some of the 209 PCB congeners exhibit increased toxicity to soil bacteria and their presence impoverish the soil decomposer community and slows down the degradation of environmental pollutants in the soils. The exact mechanism of PCB antimicrobial activity is unknown, but it is strictly related with the membrane activity of PCB. Therefore, to shed light on these interactions we applied Langmuir monolayers formed by selected phospholipids as model bacterial membranes. In our studies we tested 5 PCB congeners differing in the degree of chlorination and the distribution of the chlorine substituents around the biphenyl frame. Special attention was paid to tetra-substituted PCB because of their increased presence in the environment and disubstituted PCB being their degradation products. To characterize the model membranes as Langmuir monolayers, we used surface pressure measurements, Brewster angle microscopy and Grazing Incidence X-ray Diffraction. It turned out that among the tetra-substituted PCB the ortho-substituted non-dioxin like compound was much more membrane destructive than the flat dioxin-like congener. On the contrary, among the di-substituted PCB the flat para-substituted 2,2′-dichlorobiphenyl turned out to exhibit high membrane activity.     

A Novel Transformation of Polychlorinated Biphenyls by Rhodococcus sp. Strain RHA1

We have characterized a biphenyl degrader, Rhodococcus sp. strain RHA1. Biphenyl-grown cells of strain RHA1 efficiently transformed 45 components in the 62 major peaks of a polychlorinated biphenyl (PCB) mixture of Kanechlors 200, 300, 400, and 500 within 3 days, which includes mono- to octachlorobiphenyls. Among the intermediate metabolites of PCB transformation, di- and trichlorobenzoic acids were identified. The gradual decrease of these chlorobenzoic acids during incubation indicated that these chlorobenzoic acids would also be degraded by this strain. The effect of the position of chlorine substitution was determined by using PCB mixtures that have chlorine substitutions mainly at either the ortho or the meta position. This strain transformed both types of congeners, and strong PCB transformation activity of RHA1 was indicated. RHA1 accumulated 4-chlorobenzoic acid temporally during the transformation of 4-chlorobiphenyl. The release of most chloride in the course of 2,2(prm1)-dichlorobiphenyl degradation was observed. These results suggested that RHA1 would break down at least some PCB congeners into smaller molecules to a considerable extent.     

Mineralization of low-chlorinated biphenyls by Burkholderia sp. strain LB400 and by a two-membered consortium upon directed interspecies transfer of chlorocatechol pathway genes

The biphenyl-mineralizing bacterium Burkholderia sp. strain LB400 also utilized 3-chloro-, 4-chloro-, 2,3′-dichloro- and 2,4′-dichlorobiphenyl for growth. By the attack of the initial enzyme a chlorine was eliminated dioxygenolytically from position 2 of one of the aromatic rings when hydrogens of both were substituted by chlorine. The strain mineralized 3-chloro- and 2,3′-dichlorobiphenyl via the central intermediate 3-chlorobenzoate through its chlorocatechol pathway enzymes, but excreted stoichiometric amounts of 4-chlorobenzoate from 4-chloro- and 2,4^′-dichlorobiphenyl. These two compounds were mineralized by a co-culture of strain LB400 and a derivative of the (methyl-) benzoate-degrading strain Pseudomonas putida mt-2 (TOL). The complete degradation was achieved upon transfer of a cluster of at least five genes, encoding the regulated chlorocatechol pathway operon, from strain LB400 to strain mt-2. This transfer was demonstrated by the polymerase chain reaction. Received: 15 April 1998 / Received revision: 12 June 1998 / Accepted: 19 June 1998