Microbial biodegradation of 4-chlorobiphenyl, a model compound of chlorinated biphenyls.

The biodegradation products of 4-chlorobiphenyl were analyzed in an Achromobacter sp. strain and a Bacillus brevis strain. Both strains generated the same metabolites, with 4-chlorobenzoic acid as the major metabolic product. Our results corroborate previous observations whereby most bacterial strains degrade the chlorobiphenyls via a major pathway which proceeds by an hydroxylation in position 2,3 and a meta-1,2 fission. However, we also detected several metabolites whose structure suggests the existence of other routes for the degradation of chlorinated biphenyls.     

Microbial ecology of chlorinated solvent biodegradation

This study focused on the microbial ecology of tetrachloroethene (PCE) degradation to trichloroethene, cis‐1,2‐dichloroethene and vinyl chloride to evaluate the relationship between the microbial community and the potential accumulation or degradation of these toxic metabolites. Multiple soil microcosms supplied with different organic substrates were artificially contaminated with PCE. A thymidine analogue, bromodeoxyuridine (BrdU), was added to the microcosms and incorporated into the DNA of actively replicating cells. We compared the total and active bacterial communities during the 50‐day incubations by using phylogenic microarrays and 454 pyrosequencing to identify microorganisms and functional genes associated with PCE degradation to ethene. By use of this integrative approach, both the key community members and the ecological functions concomitant with complete PCE degradation could be determined, including the presence and activity of microbial community members responsible for producing hydrogen and acetate, which are critical for Dehalococcoides‐mediated PCE degradation. In addition, by correlation of chemical data and phylogenic microarray data, we identified several bacteria that could potentially oxidize hydrogen. These results demonstrate that PCE degradation is dependent on some microbial community members for production of appropriate metabolites, while other members of the community compete for hydrogen in soil at low redox potentials.     

Total biodegradation of 4-chlorobiphenyl (4 CB) by a two-membered bacterial culture

Summary Several bacterial strains that can oxidize mono- and dichlorinated biphenyls with one unsubstituted ring have already been described. The major route for this biodegradation leads ultimately to the corresponding chlorobenzoic acid, but several other minor chlorinated metabolites that might possibly be of concern for the environment have also been described previously. Since none of the bacterial strains that are able to oxidize these chlorinated biphenyls in pure culture are known to degrade chlorobenzoic acid, the oxidation of these substrates by axenic cultures always generates chlorobenzoates plus several other metabolites. In the present study, we have estimated the biodegradation of 4-chlorobiphenyl (4CB) by a two-membered bacterial culture containing one strain able to grow on 4CB and to transform it into 4-chlorobenzoate (4CBA) and one strain able to degrade 4CBA. The results were encouraging, since it was shown that the degradation of 4CB was more rapid and complete with the double bacterial culture.     

Interactions between rhamnolipid biosurfactants and toxic chlorinated phenols enhance biodegradation of a model hydrocarbon-rich effluent

Surfactant-mediated treatment increases hydrocarbon solubilization and potentially facilitates biodegradation, unless toxic co-contaminants inhibiting microbial activity are present in the hydrocarbon mixture. We assessed the effect of rhamnolipids on the performance of a bacterial consortium degrading diesel fuel employed as a model hydrocarbon-rich effluent, co-contaminated with toxic phenol, 4-chlorophenol (4-CP) or 2,4-dichlorophenol (2,4-DCP). This approach led to the unexpected finding that rhamnolipids reduced toxicity of 4-CP and 2,4-DCP to the hydrocarbon-degrading cells. The facts that rhamnolipids decreased diesel fuel - water partition coefficient (K_FW) of 4-CP and 2,4-DCP and modified aggregate size distribution profiles of the dispersed diesel fuel - chlorinated phenols solutions, suggest the existence of specific interactions between rhamnolipids and the co-contaminants. Due to the polar nature of 4-CP and 2,4-DCP, possible explanations involve adsorption of 4-CP and 2,4-DCP on the surface of biosurfactant aggregates. This property of rhamnolipids is of interest to those using biosurfactants for microbial treatment of hydrocarbon-rich wastewaters co-contaminated with toxic compounds.     

Microbial cometabolism of sucralose,a chlorinated disaccharide,in environmental samples

During the rapid mineralization in soil of sucralose (4-chloro-4-deoxy-,D-galactopyranosyl-1,6-dichloro-1,6-dideoxy-,D-fructofuranoside), a metabolic product was formed that appears to be the corresponding unsaturated aldehyde. During the slow and incomplete mineralization of sucralose in lake water, which was not increased by the addition of nitrogen and phosphorus, the same compound was produced. That product was further metabolized by microorganisms in lake water and soil. Mineralization was also slow in sewage under aerobic conditions, but organic products were not detected. Little or no CO₂ was formed from the disaccharide in flooded soil or anaerobic sewage. Bacteria in culture did not use sucralose as a carbon source but did convert it to the presumed unsaturated aldehyde, 1,6-dichloro-1,6-dideoxy-D-fructose and possibly the uronic acid of sucralose. Sucralose carbon was not incorporated into cells of two sucralose-metabolizing bacteria or the microbial biomass of sewage or lake water. The chlorinated disaccharide was slowly metabolized by a galactose oxidase preparation. It is concluded that the chlorinated sugar is acted on microbiologically by cometabolism.     

Plasmid-mediated mineralization of 4-chlorobiphenyl.

Strains of Alcaligenes and Acinetobacter spp. were isolated from a mixed culture already proven to be proficient at complete mineralization of monohalogenated biphenyls. These strains were shown to harbor a 35 X 10(6)-dalton plasmid mediating a complete pathway for 4-chlorobiphenyl (4CB) oxidation. Subsequent plasmid curing of these bacteria resulted in the abolishment of the 4CB mineralization phenotype and loss of even early 4CB metabolism by Acinetobacter spp. Reestablishment of the Alcaligenes plasmid, denoted pSS50, in the cured Acinetobacter spp. via filter surface mating resulted in the restoration of 4CB mineralization abilities. 4CB mineralization, however, proved to be an unstable characteristic in some subcultured strains. Such loss was not found to coincide with any detectable alteration in plasmid size. Cultures capable of complete mineralization, as well as those limited to partial metabolism of 4CB, produced 4-chlorobenzoate as a metabolite. Demonstration of mineralization of a purified 14C-labeled chlorobenzoate showed it to be a true intermediate in 4CB mineralization. Unlike the mineralization capability, the ability to produce a metabolite has proven to be stable on subculture. These results indicate the occurrence of a novel plasmid, or evolved catabolic plasmid, that mediates the complete mineralization of 4CB.     

Novel biotransformations of 4-chlorobiphenyl by a Pseudomonas sp

A bacterium, tentatively identified as a representative of the genus Pseudomonas (strain MB86), was isolated from soil contaminated by wood-preservation chemicals by using 4-chlorobenzoate as an enrichment substrate. The pseudomonad was able to grow on 4-chlorobenzoic acid and 4-chlorobiphenyl as sole carbon and energy sources. Spent culture medium from 4-chlorobiphenyl-grown cells contained 4-chlorobenzoic acid, 4'-chloroacetophenone, 2-hydroxy,2-[4'-chlorophenyl] ethane, and 2-oxo,2-[4'-chlorophenyl] ethanol as metabolites. 4'-Chloroacetophenone was produced in large amounts, possibly as a dead-end metabolite.     

Microbial degradation of chlorinated phenols

The chlorinated phenols comprise a large group of toxic, man-made chemicals that are serious environmental pollutants. Microorganisms can degrade many, but not all, of the chlorinated phenols, often using chlorophenol-specific catabolic enzymes. Novel technologies are evolving for using specific microorganisms to clean contaminated soils and waters of chlorophenols.     

Biodegradation of chlorinated biphenyls and benzoic acids by a Pseudomonas strain

Summary A Pseudomonas strain able to grown on biphenyl and 2- and 4-chlorobiphenyl has been isolated from soil. Benzoate-grown cultures of this strain were able to cometabolize other chlorobiphenyls to the corresponding chlorobenzoates. In contrast to most of the chlorobiphenyl-degrading strains described previously in the literature, which are reported to form chlorobenzoates as end metabolites from chlorobiphenyls, this strain is also able to further cometabolize chlorobenzoates to form ring-cleaved compounds.     

Novel biotransformations of 4-chlorobiphenyl by a Pseudomonas sp.

A bacterium, tentatively identified as a representative of the genus Pseudomonas (strain MB86), was isolated from soil contaminated by wood-preservation chemicals by using 4-chlorobenzoate as an enrichment substrate. The pseudomonad was able to grow on 4-chlorobenzoic acid and 4-chlorobiphenyl as sole carbon and energy sources. Spent culture medium from 4-chlorobiphenyl-grown cells contained 4-chlorobenzoic acid, 4'-chloroacetophenone, 2-hydroxy,2-[4'-chlorophenyl] ethane, and 2-oxo,2-[4'-chlorophenyl] ethanol as metabolites. 4'-Chloroacetophenone was produced in large amounts, possibly as a dead-end metabolite.     

Microbial degradation of chlorinated benzenes

Chlorinated benzenes are important industrial intermediates and solvents. Their widespread use has resulted in broad distribution of these compounds in the environment. Chlorobenzenes (CBs) are subject to both aerobic and anaerobic metabolism. Under aerobic conditions, CBs with four or less chlorine groups are susceptible to oxidation by aerobic bacteria, including bacteria (Burkholderia, Pseudomonas, etc.) that grow on such compounds as the sole source of carbon and energy. Sound evidence for the mineralization of CBs has been provided based on stoichiometric release of chloride or mineralization of (14)C-labeled CBs to (14)CO(2). The degradative attack of CBs by these strains is initiated with dioxygenases eventually yielding chlorocatechols as intermediates in a pathway leading to CO(2) and chloride. Higher CBs are readily reductively dehalogenated to lower chlorinated benzenes in anaerobic environments. Halorespiring bacteria from the genus Dehalococcoides are implicated in this conversion. Lower chlorinated benzenes are less readily converted, and mono-chlorinated benzene is recalcitrant to biotransformation under anaerobic conditions.     

Microbial degradation of chlorinated phenols

Chlorophenols have been introduced into the environment through their use as biocides and as by-products of chlorine bleaching in the pulp and paper industry. Chlorophenols are subject to both anaerobic and aerobic metabolism. Under anaerobic conditions, chlorinated phenols can undergo reductive dechlorination when suitable electron-donating substrates are available. Halorespiring bacteria are known which can use both low and highly chlorinated congeners of chlorophenol as electron acceptors to support growth. Many strains of halorespiring bacteria have the capacity to eliminate ortho-chlorines; however only bacteria from the species Desulfitobacterium hafniense (formerly frappieri) can eliminate para- and meta-chlorines in addition to ortho-chlorines. Once dechlorinated, the phenolic carbon skeletons are completely converted to methane and carbon dioxide by other anaerobic microorganisms in the environment. Under aerobic conditions, both lower and higher chlorinated phenols can serve as sole electron and carbon sources supporting growth. The best studied strains utilizing pentachlorophenol belong to the genera Mycobacterium and Sphingomonas. Two main strategies are used by aerobic bacteria for the degradation of chlorophenols. Lower chlorinated phenols for the most part are initially attacked by monooxygenases yielding chlorocatechols as the first intermediates. On the other hand, polychlorinated phenols are converted to chlorohydroquinones as the initial intermediates. Fungi and some bacteria are additionally known that cometabolize chlorinated phenols.     

Microbial transformation of chlorinated benzoates

Chlorinated benzoates enter the environment through their use as herbicides or as metabolites of other halogenated compounds. Ample evidence is available indicating biodegradation of chlorinated benzoates to CO₂ and chloride in the environment under aerobic as well as anaerobic conditions. Under aerobic conditions, lower chlorinated benzoates can serve as sole electron and carbon sources supporting growth of a large list of taxonomically diverse bacterial strains. These bacteria utilize a variety of pathways ranging from those involving an initial degradative attack by dioxygenases to those initiated by hydrolytic dehalogenases. In addition to monochlorinated benzoates, several bacterial strains have been isolated that can grow on dichloro-, and trichloro- isomers of chlorobenzoates. Some aerobic bacteria are capable of cometabolizing chlorinated benzoates with simple primary substrates such as benzoate. Under anaerobic conditions, chlorinated benzoates are subject to reductive dechlorination when suitable electron-donating substrates are available. Several halorespiring bacteria are known which can use chlorobenzoates as electron acceptors to support growth. For example, Desulfomonile tiedjei catalyzes the reductive dechlorination of 3-chlorobenzoate to benzoate. The benzoate skeleton is mineralized by other microorganisms in the anaerobic environment. Various dichloro- and trichlorobenzoates are also known to be dechlorinated in anaerobic sediments.     

Microbial biodegradation of polyaromatic hydrocarbons

Polycyclic aromatic hydrocarbons (PAHs) are widespread in various ecosystems and are pollutants of great concern due to their potential toxicity, mutagenicity and carcinogenicity. Because of their hydrophobic nature, most PAHs bind to particulates in soil and sediments, rendering them less available for biological uptake. Microbial degradation represents the major mechanism responsible for the ecological recovery of PAH-contaminated sites. The goal of this review is to provide an outline of the current knowledge of microbial PAH catabolism. In the past decade, the genetic regulation of the pathway involved in naphthalene degradation by different gram-negative and gram-positive bacteria was studied in great detail. Based on both genomic and proteomic data, a deeper understanding of some high-molecular-weight PAH degradation pathways in bacteria was provided. The ability of nonligninolytic and ligninolytic fungi to transform or metabolize PAH pollutants has received considerable attention, and the biochemical principles underlying the degradation of PAHs were examined. In addition, this review summarizes the information known about the biochemical processes that determine the fate of the individual components of PAH mixtures in polluted ecosystems. A deeper understanding of the microorganism-mediated mechanisms of catalysis of PAHs will facilitate the development of new methods to enhance the bioremediation of PAH-contaminated sites.     

Microbial biodegradation of cellophase.

The microbial biodegradation of cellophane (U.C.B.--Division Sidac) was studied. Preliminary experiments with pure cultures of seven cellulolytic microorganisms (Aspergillus sp., Penicillium sp., Chaetomium crispatum, Ch. globosum, Sclerotium rolfsii and two actinomycetes) revealed that the substrate as such was very recalcitrant, probably due to the occurrence of insoluble coating agents. Therefore, mixed cultures of the above mentioned cellulolytic microorganisms were used as inoculum. The cellophane showed a slow microbial degradation which starts only after 37 days of incubation. This long lag-phase is due to the unaltered presence of the coating agents. However, when the coating agents are extracted with tetrahydrofuran, the biodegradation starts after 10 days, resulting in a biodegradation rate of 85% after 52 days of incubation and a protein content of 30%. The endproduct (30% protein, 60% soluble sugars, 10% residual substrate) will probably be useful as compost.     

Evaporation kinetics of polychorinated biphenyls during biodegradation experiments

Summary During microbial degradation of polychlorinated biphenyls (PCB) in liquid media two processes take place simultaneously: elimination of PCB and evaporation of PCB. The physical loss of PCB due to evaporation causes frequently false positive results in long-term biodegradation experiments. Therefore, if only the PCB concentration is to be measured, its determination in both liquid and gaseous phase is essential for a correct appraisal of biodegradation. The kinetics of PCB evaporation have been monitored and the evaporation rate constants for individual PCB congeners have been determined. The values of evaporation rate constants show a good corelation with the values of the 1-octanol/water partition coefficient.     

Effects of humic substances on the bioavailability and aerobic biodegradation of polychlorinated biphenyls in a model soil.

The very high hydrophobicity of polychlorinated biphenyls (PCBs) strongly reduces their bioavailability in aged contaminated soils, thus limiting their bioremediation. The biodegradability of PCBs in heavily contaminated soils can be significantly enhanced by soil treatment with surface-active agents. In this work, the effects of naturally occurring surfactants such as humic substances (HS) on the aerobic biodegradation of PCBs in a model soil were studied. The soil was amended with biphenyl (4 g/kg), Fenclor 42 (1,000 mg/kg), the aerobic PCB-biodegrading bacterial co-culture ECO3 (inoculum: 10(8)CFU/mL), and treated in aerobic batch slurry-phase conditions (17.5% w/v) with and without the addition of HS at the rates of 1.5 and 3.0% (w/w). Low PCBs biodegradation and dechlorination yields were observed in the HS-free microcosms, probably as a result of the rapid disappearance of inoculated bacteria. The presence of HS influenced significantly the activity of the specialized biomass and the biodegradation of PCBs in the microcosms. The microcosms that received HS at the 1.5% rate showed a higher persistence of the specialized bacteria and yields of PCB biodegradation and dechlorination about 150 and 100%, respectively, larger than those found for the HS-free microcosms. Lower stimulating effects were observed in the microcosms added with the HS at 3.0% rate. These effects were attributed to an increased solubilization of PCBs in the hydrophobic domains of the humic supramolecular associations and to a different accessibility of PCBs by the specialized bacteria at the different rates of HS addition. Although the slurry-phase treatment generally showed a decrease of the original soil ecotoxicity, the addition of the originally non-toxic HS decreased soil ecotoxicity for the Collembola animal biomarker and increased that towards the Lepidium sativum vegetal biomarker.     

Microbial degradation of chlorinated acetophenones.

A defined mixed culture, consisting of an Arthrobacter sp. and a Micrococcus sp. and able to grow with 4-chloroacetophenone as a sole source of carbon and energy, was isolated. 4-Chlorophenyl acetate, 4-chlorophenol, and 4-chlorocatechol were identified as metabolites through comparison of retention times and UV spectra with those of standard substances. The proposed pathway was further confirmed by investigation of enzymes. The roles of the two collaborating strains were studied by growth experiments and on the level of enzymes. If transient accumulation of 4-chlorophenol was avoided either by the use of phenol-absorbing substances or by careful supplement of 4-chloroacetophenone, the Arthrobacter sp. was able to grow as a pure culture with 4-chloroacetophenone as a sole source of carbon and energy. Several mono-, di-, and trichlorinated acetophenones were mineralized by the Arthrobacter sp.