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PubMed2006

Bioremediation of soil contaminated with pentachlorophenol (PCP) using humic acids bound on zeolite.

We determined the toxicity of various chlorophenols, especially pentachlorophenol (PCP), on five bacterial strains and studied PCP biodegradation in soils amended with an organomineral complex (OMC) prepared from humic acids (organic part) bound on zeolite (inorganic part). Both components of OMC have excellent sorption properties and are of natural origin and therefore suitable to be used in the environment. Toxicity of chlorophenols depends not only on the number of chlorine atoms but also on their position on aromatic ring, and is thus regiospecific. Biodegradation of PCP was studied in three real completely characterized soil samples, Chernozem, Fluvisol, and Regosol, with and without the addition of OMC. The soils were sterilized and bioaugmented with the bacterial isolate Comamonas testosteroni CCM 7530. The immobilization effect of OMC in relation to PCP depends on the concentration of humic acids (HAs), the PCP concentration, and the content of organic carbon in soil. The microbial activity and the simulated action of acid rains led to the gradual release and biodegradation of the reversibly bound PCP without no initial toxic effect on indigenous or bioaugmented microorganisms. OMC appeared to be a good trap for PCP with potential applications in remediation technology because it reduces the potential toxicity of PCP to microbial community by lowering its bioavailability and thus facilitates its biodegradation.

Adsorption↗
PubMed2002

The role of cell bioaugmentation and gene bioaugmentation in the remediation of co-contaminated soils.

Soils co-contaminated with metals and organics present special problems for remediation. Metal contamination can delay or inhibit microbial degradation of organic pollutants such that for effective in situ biodegradation, bioaugmentation is necessary. We monitored the degradation of 2,4-dichlorophenoxyacetic acid (2,4-D) or 3-chlorobenzoate (3-CB) in two different soils with and without cadmium (Cd) contamination. Additionally, we evaluated the ability of bioaugmentation to enhance organic degradation in these co-contaminated soils. Finally, we determined whether enhanced degradation was due to survival of the introduced organism (cell bioaugmentation) or plasmid transfer to indigenous microbial populations (gene bioaugmentation). In Brazito soil, dual inoculation with a Cd-resistant bacterium plus a known 2,4-D-degrading bacterium, Ralstonia eutropha JMP134, enhanced 2,4-D degradation. Escherichia coli D11, which lacks chromosomal genes necessary for complete 2,4-D mineralization, was used for gene bioaugmentation in Madera soil. Significant gene transfer of the plasmid to the indigenous populations was observed, and the rate of 2,4-D degradation was enhanced relative to that of controls. Cell bioaugmentation was further demonstrated when (Comamonas testosteroni was used to enhance biodegradation of 3-CB in Madera soil. In this case no transfer of plasmid pBRC60 to indigenous soil recipients was observed. For the Madera soil, nonbioaugmented samples ultimately showed complete 2,4-D degradation. In contrast, nonbioaugmented Brazito soils showed incomplete 2,4-D degradation. These studies are unique in showing that both cell bioaugmentation and gene bioaugmentation can be effective in enhancing organic degradation in co-contaminated soils. Ultimately, the bioaugmentation strategy may depend on the degree of contamination and the time frame available for remediation.

2,4-Dichlorophenoxyacetic Acid↗
PubMed2006

Characterization of the terephthalate degradation genes of Comamonas sp. strain E6.

We isolated Comamonas sp. strain E6, which utilizes terephthalate (TPA) as the sole carbon and energy source via the protocatechuate (PCA) 4,5-cleavage pathway. Two almost identical TPA degradation gene clusters, tphRICIA2IA3IBIA1I and tphRIICIIA2IIA3IIBIIA1II, were isolated from this strain. Based on amino acid sequence similarity, the genes tphR, tphC, tphA2, tphA3, tphB, and tphA1 were predicted to code, respectively, for an IclR-type transcriptional regulator, a periplasmic TPA binding receptor, the large subunit of the oxygenase component of TPA 1,2-dioxygenase (TPADO), the small subunit of the oxygenase component of TPADO, a 1,2-dihydroxy-3,5-cyclohexadiene-1,4-dicarboxylate (DCD) dehydrogenase, and a reductase component of TPADO. The growth of E6 on TPA was not affected by disruption of either tphA2I or tphA2II singly; however, the tphA2I tphA2II double mutant no longer grew on TPA, suggesting that both TPADO genes are involved in TPA degradation. Introduction of a plasmid carrying tphRIICIIA2IIA3IIBIIA1II conferred the TPA utilization phenotype on Comamonas testosteroni IAM 1152, which is able to grow on PCA but not on TPA. Disruption of either tphRII or tphCII on this plasmid resulted in the loss of the growth of IAM 1152 on TPA, suggesting that these genes are essential to convert TPA to PCA in E6. The genes tphA1II, tphA2II, tphA3II, and tphBII were expressed in Escherichia coli, and the resultant cell extracts containing TphA1II, TphA2II, and TphA3II converted TPA in the presence of NADPH into a product which was transformed to PCA by TphBII. On the basis of these results, TPADO was strongly suggested to be a two-component dioxygenase which consists of the terminal oxygenase component (TphA2 and TphA3) and the reductase (TphA1), and tphB codes for the DCD dehydrogenase.

Bacterial Proteins↗
PubMed2006

Oxidative stress by biphenyl metabolites induces inhibition of bacterial cell separation.

Cell-cell separation of a polychlorinated biphenyl (PCB)-degrading bacterium Comamonas testosteroni TK102 was monitored by flow cytometry. When monohydroxy metabolites of biphenyl (BP) (2-hydroxybiphenyl and 3-hydroxybiphenyl) were added to the culture, cell-cell separation of strain TK102 was inhibited at stationary phase. This inhibition was reproduced on non-PCB degrading bacteria such as Pseudomonas putida PpY101 and Escherichia coli MV1184, but was not observed on Pseudomonas aeruginosa PAO1. An opportunistic pathogen, P. aeruginosa PAO1, produces exopolysaccharide, which is known to scavenge damaging chemicals such as reactive oxygen species (ROS). The higher level of ROS and lipid peroxidants were detected in the cells treated by monohydroxybiphenyls. Fat-soluble vitamin E, which is a lipid radical scavenger, maintained bacterial cell separation during monohydroxybiphenyls treatment. Our results demonstrated that intracellular oxidative stress played an important role in the inhibition of bacterial cell separation during BP metabolism. This study shows that metabolites of environmental pollutants, such as monohydroxylated BP, inhibit bacterial cell separation by oxidative stress.

Antioxidants↗
PubMed2002

Bioaugmenting bioreactors for the continuous removal of 3-chloroaniline by a slow release approach.

The survival and activity of microbial degradative inoculants in bioreactors is critical to obtain successful biodegradation of non- or slowly degradable pollutants. Achieving this in industrial wastewater reactors is technically challenging. We evaluated a strategy to obtain complete and stable bioaugmentation of activated sludge, which is used to treat a 3-chloroaniline (3-CA) contaminated wastewater in a lab-scale semi-continuous activated sludge system. A 3-CA metabolizing bacterium, Comamonas testosteroni strain I2, was mixed with molten agar and encapsulated in 4 mm diameter open-ended silicone tubes of 3 cm long. The tubes containing the immobilized bacteria represented about 1% of the volume of the mixed liquor. The bioaugmentation activity of a reactor containing the immobilized cells was compared with a reactor with suspended I2gfp cells. From day 25-30 after inoculation, the reactor with only suspended cells failed to completely degrade 3-CA because of a decrease in metabolic activity. In the reactors with immobilized cells, however, 3-CA continued to be removed. A mass balance indicated that ca. 10% of the degradation activity was due to the immobilized cells. Slow release of the growing embedded cells from the agar into the activated sludge medium, resulting in a higher number of active 3-CA-degrading I2 cells, was responsible for ca. 90% of the degradation. Our results demonstrate that this simple immobilization procedure was effective to maintain a 3-CA-degrading population within the activated sludge community.

Aniline Compounds↗
PubMed2003

Evaluation of effective diffusion coefficient and intrinsic kinetic parameters on azo dye biodegradation using PVA-immobilized cell beads.

An immobilized mixed culture (Aeromonas hydrophila, Comamonas testosteroni, and Acinetobacter baumannii) was prepared by entrapment into phosphorylated polyvinyl alcohol (PVA) gel beads. The unsteady-state diffusion mechanism in a gel bead was applied to estimate the effective diffusion coefficients (D(e)) and the partition coefficients (K(p)) of azo dye. In addition, a simple method was developed to determine the intrinsic kinetic parameters of immobilized cells from observed reaction rates and the intrinsic kinetic parameters were then verified by fitting the experimental data into the reaction-diffusion model in a batch reactor running at a well-stirred state. The calculated effectiveness factor (eta(cal)) approached unity at Thiele modulus (Phi) < 0.3 (i.e., d(p) < 0.475 mm). The experimental effectiveness factor (eta(exp)) was in the range of 0.71-0.45 for a corresponding sphere diameter (d(p)) of 1.91 +/- 0.16 to 4.43 +/- 0.07 mm at an initial dye concentration of 200 mg/L. The results show that intraparticle diffusion resistance has a significant effect on the azo dye biodegradation rate.

Acinetobacter baumannii↗
PubMed2002

Structure of bacterial 3beta/17beta-hydroxysteroid dehydrogenase at 1.2 A resolution: a model for multiple steroid recognition.

The enzyme 3beta/17beta-hydroxysteroid dehydrogenase (3beta/17beta-HSD) is a steroid-inducible component of the Gram-negative bacterium Comamonas testosteroni. It catalyzes the reversible reduction/dehydrogenation of the oxo/beta-hydroxy groups at positions 3 and 17 of steroid compounds, including hormones and isobile acids. Crystallographic analysis at 1.2 A resolution reveals the enzyme to have nearly identical subunits that form a tetramer with 222 symmetry. This is one of the largest oligomeric structures refined at this resolution. The subunit consists of a monomer with a single-domain structure built around a seven-stranded beta-sheet flanked by six alpha-helices. The active site contains a Ser-Tyr-Lys triad, typical for short-chain dehydrogenases/reductases (SDR). Despite their highly diverse substrate specificities, SDR members show a close to identical folding pattern architectures and a common catalytic mechanism. In contrast to other SDR apostructures determined, the substrate binding loop is well-defined. Analysis of structure-activity relationships of catalytic cleft residues, docking analysis of substrates and inhibitors, and accessible surface analysis explains how 3beta/17beta-HSD accommodates steroid substrates of different conformations.

17-Hydroxysteroid Dehydrogenases↗
PubMed2001

Identification and functional characterization of CbaR, a MarR-like modulator of the cbaABC-encoded chlorobenzoate catabolism pathway.

In Comamonas testosteroni BR60 (formerly Alcaligenes sp. strain BR60), catabolism of the pollutant 3-chlorobenzoate (3CBA) is initiated by enzymes encoded by cbaABC, an operon found on composite transposon Tn5271 of plasmid pBRC60. The cbaABC gene product CbaABC converts 3CBA to protocatechuate (PCA) and 5-Cl-PCA, which are then metabolized by the chromosomal PCA meta (extradiol) ring fission pathway. In this study, cbaA was found to possess a sigma(70) type promoter. O(2) uptake experiments with whole cells and expression studies with cbaA-lacZ constructs showed that cbaABC was induced by 3CBA. Benzoate, which is not a substrate of the 3CBA pathway, was a gratuitous inducer, and CbaR, a MarR family repressor coded for by a divergently transcribed gene upstream of cbaABC, could modulate induction mediated by benzoate. Purified CbaR bound specifically to two regions of the cbaA promoter (P(cbaA)); site I, a high-affinity site, is between the transcriptional start point (position +1) and the start codon of cbaA, while site II, a lower-affinity site, overlaps position +1. 3CBA at concentrations as low as 40 microM interfered with binding to P(cbaA). PCA also interfered with binding, while benzoate only weakly disrupted binding. Unexpectedly, benzoate with a hydroxyl or carboxyl at position 3 improved CbaR binding. Data are also presented that suggest that an unidentified regulator is encoded on the chromosome that induces cbaABC in response to benzoate and 3CBA.

Bacterial Proteins↗
PubMed2003

Bioaugmentation as a tool to protect the structure and function of an activated-sludge microbial community against a 3-chloroaniline shock load.

Bioaugmentation of bioreactors focuses on the removal of xenobiotics, with little attention typically paid to the recovery of disrupted reactor functions such as ammonium-nitrogen removal. Chloroanilines are widely used in industry as a precursor to a variety of products and are occasionally released into wastewater streams. This work evaluated the effects on activated-sludge reactor functions of a 3-chloroaniline (3-CA) pulse and bioaugmentation by inoculation with the 3-CA-degrading strain Comamonas testosteroni I2 gfp. Changes in functions such as nitrification, carbon removal, and sludge compaction were studied in relation to the sludge community structure, in particular the nitrifying populations. Denaturing gradient gel electrophoresis (DGGE), real-time PCR, and fluorescent in situ hybridization (FISH) were used to characterize and enumerate the ammonia-oxidizing microbial community immediately after a 3-CA shock load. Two days after the 3-CA shock, ammonium accumulated, and the nitrification activity did not recover over a 12-day period in the nonbioaugmented reactors. In contrast, nitrification in the bioaugmented reactor started to recover on day 4. The DGGE patterns and the FISH and real-time PCR data showed that the ammonia-oxidizing microbial community of the bioaugmented reactor recovered in structure, activity, and abundance, while the number of ribosomes of the ammonia oxidizers in the nonbioaugmented reactor decreased drastically and the community composition changed and did not recover. The settleability of the activated sludge was negatively influenced by the 3-CA addition, with the sludge volume index increasing by a factor of 2.3. Two days after the 3-CA shock in the nonbioaugmented reactor, chemical oxygen demand (COD) removal efficiency decreased by 36% but recovered fully by day 4. In contrast, in the bioaugmented reactor, no decrease of the COD removal efficiency was observed. This study demonstrates that bioaugmentation of wastewater reactors to accelerate the degradation of toxic chlorinated organics such as 3-CA protected the nitrifying bacterial community, thereby allowing faster recovery from toxic shocks.

Ammonia↗
PubMed2005

Medium composition affects the degree and pattern of cadmium inhibition of naphthalene biodegradation.

Metals have been reported to inhibit organic pollutant biodegradation; however, widely varying degrees and patterns of inhibition have been reported. To investigate the roles of medium composition and metal bioavailability on these different degrees and patterns of inhibition, we assessed the impact of cadmium on naphthalene biodegradation by a newly isolated strain of Comamonas testosteroni in three chemically-defined minimal salts media (MSM): Tris-buffered MSM, PIPES-buffered MSM, and Bushnell-Haas medium. Cadmium (total concentrations of 100 and 500 microM) inhibited biodegradation in each medium. Degrees of inhibition were different in each medium. Cadmium was most inhibitory in PIPES-buffered MSM and least inhibitory in Bushnell-Haas. For example, in Bushnell-Haas medium, 100 microM cadmium reduced the cell yield more than 4-fold compared to controls not containing cadmium. The same concentration of cadmium completely inhibited growth in PIPES-buffered MSM. No difference in inhibition was observed in any medium when cadmium was added 24 h before inoculation rather than when added within one minute of inoculation. Two patterns of inhibition were observed. Inhibition occurred in a dose dependent pattern in Tris- and PIPES-buffered MSM and in a non-dose dependent pattern in Bushnell-Haas. Specifically, in Bushnell-Haas, 100 microM total cadmium extended the lag phase by 23+/-8.66 h, whereas 500 microM did not extend the lag phase. Soluble, ionic cadmium (Cd2+) concentrations were measured and modeled in each medium to assess cadmium bioavailability. In media containing 500 microM total cadmium, bioavailability was highest in Tris- and PIPES-buffered MSM and lowest in Bushnell-Haas. In Bushnell-Haas, cadmium bioavailability was initially higher in the 500 microM treatments (196+/-21.2 microM) than in the 100 microM treatments (78.2+/-2.04 microM); however, after 12 h, bioavailability was higher in the 100 microM treatments (56.4+/-24.8 micro) than the 500 microM treatments (13.3+/-1.2 microM). These data suggest that the type of medium determines the degrees and patterns by which metals inhibit biodegradation and emphasize the importance of coupling metal toxicity and bioavailability data.

Biodegradation, Environmental↗
PubMed2003

Origin of the different pH activity profile in two homologous ketosteroid isomerases.

Two homologous Delta5-3-ketosteroid isomerases from Comamonas testosteroni (TI-WT) and Pseudomonas putida biotype B (PI-WT) exhibit different pH activity profiles. TI-WT loses activity below pH 5.0 due to the protonation of the conserved catalytic base, Asp-38, while PI-WT does not. Based on the structural analysis of PI-WT, the critical catalytic base, Asp-38, was found to form a hydrogen bond with the indole ring NH of Trp-116, which is homologously replaced with Phe-116 in TI-WT. To investigate the role of Trp-116, we prepared the F116W mutant of TI-WT (TI-F116W) and the W116F mutant of PI-WT (PI-W116F) and compared kinetic parameters of those mutants at different pH levels. PI-W116F exhibited significantly decreased catalytic activity at acidic pH like TI-WT, whereas TI-F116W maintained catalytic activity at acidic pH like PI-WT and increased the kcat/Km value by 2.5- to 4.7-fold compared with TI-WT at pH 3.8. The crystal structure of TI-F116W clearly showed that the indole ring NH of Trp-116 could form a hydrogen bond with the carboxyl oxygen of Asp-38 like that of PI-WT. The present results demonstrate that the activities of both PI-WT and TI-F116W at low pH were maintained by a tryptophan, which was able not only to lower the pKa value of the catalytic base but also to increase the substrate affinity. This is one example of the strategy nature can adopt to evolve the diversity of the catalytic function in the enzymes. Our results provide insight into deciphering the molecular evolution of the enzyme and creating novel enzymes by protein engineering.

Aspartic Acid↗
PubMed1993

Use of a simplified cell blot technique and 16S rRNA-directed probes for identification of common environmental isolates.

A simple technique in which rRNA-targeted oligodeoxynucleotide probes are used to identify bacteria immobilized on membranes is described. By using colony lifts, bacteria are directly transferred from plates to untreated nitrocellulose membranes. Alternatively, cells resuspended from colonies can be applied to membranes by using a vacuum manifold under high-salt conditions. Blotted cells are baked and hybridized under stringent conditions by using standard protocols. Treatment of blotted cells with sodium dodecyl sulfate, urea, formaldehyde, or protease had no apparent effect on hybridization signals. Hybridization to rRNA from cells that had been stored refrigerated for 6 days was readily detected; however, fivefold more cells (approximately 10(7) cells) were required to obtain hybridization signals comparable to those generated by cells not subjected to storage. The sequences of oligonucleotide probes specific for Pseudomonas cepacia, Comamonas testosteroni, and Acinetobacter calcoaceticus and a group probe identifying Pseudomonas aeruginosa, Pseudomonas mendocina, Pseudomonas fluorescens, Comamonas acidovorans, and "Flavobacterium" lutescens are presented. In conjunction with the colony lift hybridization procedure, bacteria isolated from river water were identified by using these probes.

Bacteria↗
PubMed1996

Biodegradation of polychlorinated biphenyls by rhizobia: a novel finding.

Metabolism of simple aromatic compounds in rhizobial strains has been a subject of study for a few decades, due either to the significance of nutritional diversity in the inoculum survival during agricultural applications or to the importance of plant phenolics in the microbe-plant cross-talk and signal-transduction. Here, we report the capability of rhizobial strains to catabolize polychlorinated biphenyls (PCBs). In order to identify the genes in these strains that mediate the catabolism of PCBs we used the bphABC genes from Comamonas testosteroni strain B-356. Our results showed that genomic DNAs from all four rhizobial strains studied hybridized strongly with the Comamonas-derived probe, indicating the presence of a similar genetic system. This is a novel and interesting finding indicating for the first time, perhaps, of a role of rhizobia in recycling of aromatic compounds in nature and, certainly, opening a new avenue to be explored in the field of bioremediation.

Aroclors↗
PubMed2001

Genetic diversity among 3-chloroaniline- and aniline-degrading strains of the Comamonadaceae.

We examined the diversity of the plasmids and of the gene tdnQ, involved in the oxidative deamination of aniline, in five bacterial strains that are able to metabolize both aniline and 3-chloroaniline (3-CA). Three strains have been described and identified previously, i.e., Comamonas testosteroni I2 and Delftia acidovorans CA28 and BN3.1. Strains LME1 and B8c were isolated in this study from linuron-treated soil and from a wastewater treatment plant, respectively, and were both identified as D. acidovorans. Both Delftia and Comamonas belong to the family Comamonadaceae. All five strains possess a large plasmid of ca. 100 kb, but the plasmids from only four strains could be transferred to a recipient strain by selection on aniline or 3-CA as a sole source of carbon and/or nitrogen. Plasmid transfer experiments and Southern hybridization revealed that the plasmid of strain I2 was responsible for total aniline but not 3-CA degradation, while the plasmids of strains LME1 and B8c were responsible only for the oxidative deamination of aniline. Several transconjugant clones that had received the plasmid from strain CA28 showed different degradative capacities: all transconjugants could use aniline as a nitrogen source, while only some of the transconjugants could deaminate 3-CA. For all four plasmids, the IS1071 insertion sequence of Tn5271 was found to be located on a 1.4-kb restriction fragment, which also hybridized with the tdnQ probe. This result suggests the involvement of this insertion sequence element in the dissemination of aniline degradation genes in the environment. By use of specific primers for the tdnQ gene from Pseudomonas putida UCC22, the diversity of the PCR-amplified fragments in the five strains was examined by denaturing gradient gel electrophoresis (DGGE). With DGGE, three different clusters of the tdnQ fragment could be distinguished. Sequencing data showed that the tdnQ sequences of I2, LME1, B8c, and CA28 were very closely related, while the tdnQ sequences of BN3.1 and P. putida UCC22 were only about 83% identical to the other sequences. Northern hybridization revealed that the tdnQ gene is transcribed only in the presence of aniline and not when only 3-CA is present.

Aniline Compounds↗
PubMed2001

Diversity in kinetics of trichloroethylene-degrading activities exhibited by phenol-degrading bacteria.

Whole-cell kinetics of phenol- and trichloroethylene (TCE)-degrading activities expressed by 13 phenol-degrading bacteria were analyzed. The Ks (apparent affinity constant in Haldane's equation) values for TCE were unexpectedly diverse, ranging from 11 microM to over 800 microM. The Vmax/Ks values for phenol were three orders of magnitude higher than the values for TCE in all bacteria analyzed, suggesting that these bacteria preferentially degrade phenol rather than TCE. A positive correlation between Ks for phenol and Ks for TCE was found, i.e., bacteria exhibiting high Ks values for phenol showed high Ks values for TCE, and vice versa. A comparison of the Ks values allowed grouping of these bacteria into three types, i.e., low-, moderate- and high-Ks types. Pseudo-first-order degradation-rate constants for TCE at 3.8 microM were found to be adequate to rapidly discriminate among the three types of bacteria. When bacteria were grown on phenol at the initial concentration of 2 mM, Comamonas testosteroni strain R5, a representative of low-Ks bacteria, completely degraded TCE at 3.8 microM, while strain P-8, a representative of high-Ks bacteria, did not. A mixed culture of these two bacteria poorly degraded TCE under the same conditions, where P-8 outgrew R5. These results suggest that low-Ks bacteria should be selectively grown for effective bioremediation of TCE-contaminated groundwater.

Betaproteobacteria↗
PubMed2003

Post-translational modification of Rhodococcus R312 and Comamonas NI1 nitrile hydratases.

Nitrile hydratases (NHases) are industrially significant iron- and cobalt-containing enzymes used in the large-scale synthesis of acrylamide. Previous reports have shown that the active site peptides of NHases are post-translationally modified by oxidation of cysteine residues, and that these modifications are essential for catalysis. We report mass spectrometric evidence of the oxidation states of the active site cysteines in the iron coordination spheres of two iron-containing nitrile hydratases, namely R312 NHase from Rhodococcus rhodochrous strain R312 and NI1 NHase from Comamonas testosteroni. At least one of these cysteines is oxidised to a sulfinic acid (SO(2)H) and there is also evidence suggesting an additional oxidation to a sulfenic acid (SOH). This is the first evidence for the presence of these oxidation states for full-length NHases and for Fe-NHases from different microorganisms. The presence of these covalent modifications was confirmed by performing mass spectrometry on the active site peptide of R312 NHase, under native, reduced and carboxymethylated conditions. We also show the nitrosylation of the iron by mass spectrometry, as well as the release of NO by photoirradiation.

Binding Sites↗
PubMed1999

Degradation of anaerobic reductive dechlorination products 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, including 2-, 4-, 2,4-, 2,6-, 2,2'-, 2,4'-, 2,2', 4-, and 2,4,4'-chlorobiphenyl (CB), and the biodegradation of their mixtures designed to simulate anaerobic dechlorination profiles M and C. Strains Comamonas testosteroni VP44 and Rhodococcus erythreus NY05 preferentially oxidized a para-substituted ring, while Rhodococcus sp. RHA1, similar to well known strain Burkholderia sp. LB400, preferably attacked an ortho-chlorinated ring. Strains with ortho-directed attack extensively degraded 2,4'- and 2,4,4'-CB into 4-chlorobenzoate, while bacteria with para-directed attack transformed these congeners mostly into potentially problematic meta-cleavage products. The strains that preferentially oxidized an ortho-substituted ring readily degraded seven of the eight congeners supplied individually; only 2,6-CB was poorly degraded. Degradation of 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 attack an ortho-substituted ring. PCB congeners 2,4'-, 2,2',4-, and 2,4,4'-CB can be used to easily identify bacteria with ortho-directed attack which are advantageous for use in the aerobic stage of the two-phase (anaerobic/aerobic) PCB bioremediation scheme.

Anaerobiosis↗
PubMed2005

Effects of pH amendment on metal working fluid wastewater biological treatment using a defined bacterial consortium.

The aim of this study was to determine whether pH amendment of a highly alkaline metal working fluid (MWF) wastewater would improve biological treatment in a bioreactor system following introduction of a bacterial inoculum (comprised of the following strains: Agrobacterium radiobacter, Comamonas testosteroni, Methylobacterium mesophilicum, Microbacterium esteraromaticum, and Microbacterium saperdae). The pH values tested were 6, 7, 8, and 9. Three replicate batch mode bioreactors inoculated with the bacterial inoculum (plus an abiotic control bioreactor) were operated for each of the four pH conditions. After 14 days, the final mean chemical oxygen demand (COD) reduction at pH 9 was 50 +/- 1.4%; at pH 8, 58 +/- 1.4%; pH 7, 65 +/- 1.0%; and pH 6, 75 +/- 2.7% of the initial COD (approximately 10,000 mg L(-1)), respectively. Interestingly, within 5 days, the pH in all inoculated bioreactors progressed toward pH 8. However, all abiotic control bioreactors remained at the pH at which they were amended. The fate of the inoculum, determined by denaturing gradient gel electrophoresis (DGGE) and by cluster analysis of the resulting DGGE profiles, revealed that the inocula survived throughout operation of all pH-amended bioreactors. Length-heterogeneity polymerase chain reaction (PCR) was used to track the population dynamics of individual strains. After 7 days of operation, M. esteraromaticum was the most abundant population in all bioreactors, regardless of pH. From our findings, it appears necessary to adjust the MWF wastewater from pH 9 to between 6 and 7, to achieve optimal biological treatment rates.

Biodegradation, Environmental↗