Anaerobic dechlorination of polychlorobiphenyls (Aroclor 1242) by pasteurized and ethanol-treated microorganisms from sediments.

A polychlorobiphenyl (PCB)-dechlorinating inoculum eluted from upper Hudson River sediments was treated with either heat or ethanol or both. The treated cultures retained the ability to dechlorinate PCBs (Aroclor 1242) under strictly anaerobic conditions. The dechlorination activity was maintained in serial cultures inoculated with transfers of 1% inoculum when the transferred inoculum was treated each time in the same manner. No methane production was detected in any treated culture, although dechlorination of PCBs in the untreated cultures was always accompanied by methane production. All treated cultures preferentially removed meta chlorines, yielding a dechlorination pattern characterized by accumulation of certain ortho- and para-subsituted congeners such as 2-4-chlorobiphenyl (2-4-CB), 2,4-2-CB, and 2,4-4-CB. In contrast, the untreated cultures showed more extensive dechlorination activities, which almost completely removed both meta and para chlorines from Aroclor 1242. These results suggest that microorganisms responsible for the dechlorination of PCBs in the upper Hudson River sediments can be grouped into two populations according to their responses to the heat and ethanol treatments. Microorganisms surviving the heat and ethanol treatments preferentially remove meta chlorines, while microorganisms lost from the enrichment mainly contribute to the para dechlorination activity. These results indicate that anaerobic sporeformers are at least one of the physiological groups responsible for the reductive dechlorination of PCBs. The selection of a dechlorinating population by such treatments may be an important step in isolation of PCB-dechlorinating microorganisms.     

Biotransformations of Aroclor 1242 in Hudson River test tube microcosms.

A microcosm system to physically model the fate of Aroclor 1242 in Hudson River sediment was developed. In the dark at 22 to 25 degrees C with no amendments (nutrients, organisms, or mixing) and with overlying water being the only source of oxygen, the microcosms developed visibly distinct aerobic and anaerobic compartments in 2 to 4 weeks. Extensive polychlorinated biphenyl (PCB) biodegradation was observed in 140 days. Autoclaved controls were unchanged throughout the experiments. In the surface sediments of these microcosms, the PCBs were biologically altered by both aerobic biodegrading and reductive dechlorinating microorganisms, decreasing the total concentration from 64.8 to 18.0 micromol/kg of sediment in 1140 days. This is the first laboratory demonstration of meta dechlorination plus aerobic biodegradation in stationary sediments. In contrast, the primary mechanism of microbiological attack on PCBs in aerobic subsurface sediments was reductive dechlorination. The concentration of PCBs remained constant at 64.8 micromol/kg of sediment, but the average number of chlorines per biphenyl decreased from 3.11 to 1.84 in 140 days. The selectivities of microorganisms in these sediments were characterized by meta and para dechlorination. Our results provide persuasive evidence that naturally occurring microorganisms in the Hudson River have the potential to attack the PCBs from Aroclor 1242 releases both aerobically and anaerobically at rapid rates. These unamended microcosms represent a unique method for determining the fate of released PCBs in river sediments.     

Evidence for para Dechlorination of Polychlorobiphenyls by Methanogenic Bacteria

When microorganisms eluted from upper Hudson River sediment were cultured without any substrate except polychlorobiphenyl (PCB)-free Hudson River sediment, methane formation was the terminal step of the anaerobic food chain. In sediments containing Aroclor 1242, addition of eubacterium-inhibiting antibiotics, which should have directly inhibited fermentative bacteria and thereby should have indirectly inhibited methanogens, resulted in no dechlorination activity or methane production. However, when substrates for methanogenic bacteria were provided along with the antibiotics (to free the methanogens from dependence on eubacteria), concomitant methane production and dechlorination of PCBs were observed. The dechlorination of Aroclor 1242 was from the para positions, a pattern distinctly different from, and more limited than, the pattern observed with untreated or pasteurized inocula. Both methane production and dechlorination in cultures amended with antibiotics plus methanogenic substrates were inhibited by 2-bromoethanesulfonic acid. These results suggest that the methanogenic bacteria are among the physiological groups capable of anaerobic dechlorination of PCBs, but that the dechlorination observed with methanogenic bacteria is less extensive than the dechlorination observed with more complex anaerobic consortia.     

Dechlorination of Four Commercial Polychlorinated Biphenyl Mixtures (Aroclors) by Anaerobic Microorganisms from Sediments

The rate, extent, and pattern of dechlorination of four Aroclors by inocula prepared from two polychlorinated biphenyl (PCB)-contaminated sediments were compared. The four mixtures used, Aroclors 1242, 1248, 1254, and 1260, average approximately three, four, five, and six chlorines, respectively, per biphenyl molecule. All four Aroclors were dechlorinated with the loss of meta plus para chlorines ranging from 15 to 85%. Microorganisms from an Aroclor 1242-contaminated site in the upper Hudson River dechlorinated Aroclor 1242 to a greater extent than did microorganisms from Aroclor 1260-contaminated sediments from Silver Lake, Mass. The Silver Lake inoculum dechlorinated Aroclor 1260 more rapidly than the Hudson River inoculum did and showed a preferential removal of meta chlorines. For each inoculum the rate and extent of dechlorination tended to decrease as the degree of chlorination of the Aroclor increased, especially for Aroclor 1260. The maximal observed dechlorination rates were 0.3, 0.3, and 0.2 μg-atoms of Cl removed per g of sediment per week for Aroclors 1242, 1248, and 1254, respectively. The maximal observed dechlorination rates for Hudson River and Silver Lake organisms for Aroclor 1260 were 0.04 and 0.21 μg-atoms of Cl removed per g of sediment per week, respectively. The dechlorination patterns obtained suggested that the Hudson River microorganisms were more capable than the Silver Lake organisms of removing the last para chlorine. These results suggest that there are different PCB-dechlorinating microorganisms at different sites, with characteristic specificities for PCB dechlorination.     

Reductive debromination of the commercial polybrominated biphenyl mixture firemaster BP6 by anaerobic microorganisms from sediments.

Anaerobic microorganisms eluted from three sediments, one contaminated with polybrominated biphenyls (PBBs) and two contaminated with polychlorinated biphenyls, were compared for their ability to debrominate the commercial PBB mixture Firemaster. These microorganisms were incubated with reduced anaerobic mineral medium and noncontaminated sediment amended with Firemaster. Firemaster averages six bromines per biphenyl molecule; four of the bromines are substituted in the meta or para position. The inocula from all three sources were able to debrominate the meta and para positions. Microorganisms from the Pine River (St. Louis, Mich.) contaminated with Firemaster, the Hudson River (Hudson Falls, N.Y.) contaminated with Aroclor 1242, and Silver Lake (Pittsfield, Mass.) contaminated with Aroclor 1260 removed 32, 12, and 3% of the meta plus para bromines, respectively, after 32 weeks of incubation. This suggests that previous environmental exposure to PBBs enhances the debromination capability of the sediment microbial community through selection for different strains of microorganisms. The Pine River inoculum removed an average of 1.25 bromines per biphenyl molecule during a 32-week incubation period, resulting in a mixture potentially more accessible to aerobic degradation processes. No ortho bromine removal was observed. However, when Firemaster was incubated with Hudson River microorganisms that had been repeatedly transferred on a pyruvate medium amended with Aroclor 1242, 17% of the meta and para bromines were removed after 16 weeks of incubation and additional debromination products, including 2-bromobiphenyl and biphenyl, were detected. This suggests the possibility for ortho debromination, since all components of the Firemaster mixture have at least one ortho-substituted bromine.(ABSTRACT TRUNCATED AT 250 WORDS)     

Reductive debromination of the commercial polybrominated biphenyl mixture firemaster BP6 by anaerobic microorganisms from sediments.

Anaerobic microorganisms eluted from three sediments, one contaminated with polybrominated biphenyls (PBBs) and two contaminated with polychlorinated biphenyls, were compared for their ability to debrominate the commercial PBB mixture Firemaster. These microorganisms were incubated with reduced anaerobic mineral medium and noncontaminated sediment amended with Firemaster. Firemaster averages six bromines per biphenyl molecule; four of the bromines are substituted in the meta or para position. The inocula from all three sources were able to debrominate the meta and para positions. Microorganisms from the Pine River (St. Louis, Mich.) contaminated with Firemaster, the Hudson River (Hudson Falls, N.Y.) contaminated with Aroclor 1242, and Silver Lake (Pittsfield, Mass.) contaminated with Aroclor 1260 removed 32, 12, and 3% of the meta plus para bromines, respectively, after 32 weeks of incubation. This suggests that previous environmental exposure to PBBs enhances the debromination capability of the sediment microbial community through selection for different strains of microorganisms. The Pine River inoculum removed an average of 1.25 bromines per biphenyl molecule during a 32-week incubation period, resulting in a mixture potentially more accessible to aerobic degradation processes. No ortho bromine removal was observed. However, when Firemaster was incubated with Hudson River microorganisms that had been repeatedly transferred on a pyruvate medium amended with Aroclor 1242, 17% of the meta and para bromines were removed after 16 weeks of incubation and additional debromination products, including 2-bromobiphenyl and biphenyl, were detected. This suggests the possibility for ortho debromination, since all components of the Firemaster mixture have at least one ortho-substituted bromine.(ABSTRACT TRUNCATED AT 250 WORDS)     

Anaerobic ortho Dechlorination of Polychlorinated Biphenyls by Estuarine Sediments from Baltimore Harbor

Reductive dechlorination of the ortho moiety of polychlorinated biphenyls (PCBs) as well as of meta and para moieties is shown to occur in anaerobic enrichments of Baltimore Harbor sediments. These estuarine sediments ortho dechlorinated 2,3,5,6-chlorinated biphenyl (CB), 2,3,5-CB, and 2,3,6-CB in freshwater or estuarine media within a relatively short period of 25 to 44 days. ortho dechlorination developed within 77 days in marine medium. High levels of ortho dechlorination (>90%) occurred when harbor sediments were supplied with only 2,3,5-CB. Incubation with 2,3,4,5,6-CB or 2,3,4,5-CB resulted in the formation of the ortho dechlorination product 3,5-CB; however, para dechlorination of these congeners always preceded ortho chlorine removal. ortho dechlorination of PCBs is an exceedingly rare event that has not been reported previously for marine or estuarine conditions. The activity was reproducible and could be sustained through sequential transfers. In contrast, freshwater sediments incubated under the same conditions exhibited only meta and para dechlorinations. The results indicate that unique anaerobic dechlorinating activity is catalyzed by microorganisms in the estuarine sediments from Baltimore Harbor.     

Effect of Incubation Temperature on the Route of Microbial Reductive Dechlorination of 2,3,4,6-Tetrachlorobiphenyl in Polychlorinated Biphenyl (PCB)-Contaminated and PCB-Free Freshwater Sediments

We studied the influence of temperature (4 to 66(deg)C) on the microbial dechlorination of 2,3,4,6-tetrachlorobiphenyl (2,3,4,6-CB) incubated for 1 year in anaerobic sediments from Woods Pond in Lenox, Mass., and Sandy Creek Nature Center Pond (SCNC) in Athens, Ga. Seven discrete dechlorination reactions were observed, four of which occurred in both sediments. These were 2,3,4,6-CB (symbl) 2,4,6-CB, 2,3,4,6-CB (symbl) 2,3,6-CB, 2,4,6-CB (symbl) 2,6-CB, and 2,3,6-CB (symbl) 2,6-CB. Three additional reactions occurred only in Woods Pond sediment. These were 2,4,6-CB (symbl) 2,4-CB, 2,4-CB (symbl) 2-CB, and 2,4-CB (symbl) 4-CB. The dechlorination reactions exhibited at least four different temperature dependencies in SCNC sediment and at least six in Woods Pond sediment. We attribute the discrete dechlorination reactions to different polychlorinated biphenyl (PCB)-dechlorinating microorganisms with distinct specificities. Temperature influenced the timing and the relative predominance of parallel pathways of dechlorination, i.e., meta versus para dechlorination of 2,3,4,6-CB and ortho versus para dechlorination of 2,4,6-CB and 2,4-CB. meta dechlorination of 2,3,4,6-CB to 2,4,6-CB dominated at all tested temperatures except at 18 and 34(deg)C, where para dechlorination to 2,3,6-CB dominated in some replicates. The dechlorination of 2,4,6-CB was restricted to (symbl)15 to 30(deg)C in both sediments. Temperature affected the lag time preceding the dechlorination of 2,4,6-CB in both sediments and affected the preferred route of its dechlorination in Woods Pond sediment. para dechlorination dominated at 20(deg)C, and ortho dechlorination dominated at 15(deg)C, but at 18 and 22 to 30(deg)C the relative dominance of ortho versus para dechlorination of 2,4,6-CB varied. These data indicate that field temperatures play a significant role in controlling the nature and the extent of the PCB dechlorination that occurs at a given site.     

Microbial Reductive Dechlorination of Aroclor 1260 in Anaerobic Slurries of Estuarine Sediments

Reductive dechlorination of Aroclor 1260 was investigated in anaerobic slurries of estuarine sediments from Baltimore Harbor (Baltimore, Md.). The sediment slurries were amended with 800 ppm Aroclor 1260 with and without the addition of 350 μM 2,3,4,5-tetrachlorobiphenyl (2,3,4,5-CB) or 2,3,5,6-tetrachlorobiphenyl (2,3,5,6-CB) and incubated in triplicate at 30°C under methanogenic conditions in an artificial estuarine medium. After 6 months, extensive meta dechlorination and moderate ortho dechlorination of Aroclor 1260 occurred in all incubated cultures except for sterilized controls. Overall, total chlorines per biphenyl decreased by up to 34%. meta chlorines per biphenyl decreased by 65, 55, and 45% and ortho chlorines declined by 18, 12, and 9%, respectively, when 2,3,4,5-CB, 2,3,5,6-CB, or no additional congener was supplied. This is the first confirmed report of microbial ortho dechlorination of a commercial polychlorinated biphenyl mixture. In addition, compared with incubated cultures supplied with Aroclor 1260 alone, the dechlorination of Aroclor 1260 plus 2,3,4,5-CB or 2,3,5,6-CB occurred with shorter lag times (31 to 60 days versus 90 days) and was more extensive, indicating that the addition of a single congener stimulated the dechlorination of Aroclor 1260.     

The reductive dechlorination of 2,3,4,5-tetrachlorobiphenyl in three different sediment cultures: evidence for the involvement of phylogenetically similar Dehalococcoides-like bacterial populations

Anaerobic cultures capable of reductively dechlorinating 2,3,4,5-tetrachlorobiphenyl (CB) were enriched from three different sediments, one estuarine, one marine and one riverine. Two different electron donors were used in enrichments with the estuarine sediment (elemental iron or a mixture of fatty acids). The removal of doubly flanked meta and para chlorines to form 2,3,5-CB and 2,4,5-CB was observed in all cultures. Bacterial community analysis of PCR-amplified 16S rRNA gene fragments revealed different communities in these cultures, with the exception of one common population that showed a high phylogentic relatedness to Dehalococcoides species. No Dehalococcoides-like populations were ever detected in control cultures to which no PCBs were added. In addition, the dynamics of this Dehalococcoides-like population were strongly correlated with dechlorination. Subcultures of the estuarine sediment culture demonstrated that the Dehalococcoides-like population disappeared when dechlorination was inhibited with 2-bromoethanesulfonate or when 2,3,4,5-CB had been consumed. These results provide evidence that Dehalococcoides-like populations were involved in the removal of doubly flanked chlorines from 2,3,4,5-CB. Furthermore, the successful enrichment of these populations from geographically distant and geochemically distinct environments indicates the widespread presence of these PCB-dechlorinating, Dehalococcoides-like organisms.     

The Dehalococcoides population in sediment-free mixed cultures metabolically dechlorinates the commercial polychlorinated biphenyl mixture aroclor 1260

Microbial reductive dechlorination of commercial polychlorinated biphenyl (PCB) mixtures (e.g., Aroclors) in aquatic sediments is crucial to achieve detoxification. Despite extensive efforts over nearly two decades, the microorganisms responsible for Aroclor dechlorination remained elusive. Here we demonstrate that anaerobic bacteria of the Dehalococcoides group derived from sediment of the Housatonic River (Lenox, MA) simultaneously dechlorinate 64 PCB congeners carrying four to nine chlorines in Aroclor 1260 in the sediment-free JN cultures. Quantitative real-time PCR showed that the Dehalococcoides cell titer in JN cultures amended with acetate and hydrogen increased from 7.07 x 10(6) +/- 0.42 x 10(6) to 1.67 x 10(8) +/- 0.04 x 10(8) cells/ml, concomitant with a 64.2% decrease of the PCBs with six or more chlorines in Aroclor 1260. No Dehalococcoides growth occurred in parallel cultures without PCBs. Aroclor 1260 dechlorination supported the growth of 9.25 x 10(8) +/- 0.04 x 10(8) Dehalococcoides cells per mumol of chlorine removed. 16S rRNA gene-targeted PCR analysis of known dechlorinators (i.e., Desulfitobacterium, Dehalobacter, Desulfuromonas, Sulfurospirillum, Anaeromyxobacter, Geobacter, and o-17/DF-1-type Chloroflexi organisms) ruled out any involvement of these bacterial groups in the dechlorination. Our results suggest that the Dehalococcoides population present in the JN cultures also catalyzes in situ dechlorination of Aroclor 1260 in the Housatonic River. The identification of Dehalococcoides organisms as catalysts of extensive Aroclor 1260 dechlorination and our ability to propagate the JN cultures under defined conditions offer opportunities to study the organisms' ecophysiology, elucidate nutritional requirements, identify reductive dehalogenase genes involved in PCB dechlorination, and design molecular tools required for bioremediation applications.     

Establishment of polychlorinated biphenyl-degrading enrichment culture with predominantly meta dechlorination.

Enrichment of polychlorinated biphenyl (PCB)-dechlorinating microorganisms from PCB-contaminated sediments from the Upper Hudson River, N.Y., was attempted. The enrichment strategy was to use pyruvate as the electron donor and dechlorination of Aroclor 1242 as the electron acceptor. The enrichment medium also contained non-PCB-contaminated Hudson River sediments, which were required for the PCB-dechlorinating activity. An enrichment culture (that had stable PCBT-dechlorinating activity over nine serial transfers during 1 year) was established under these conditions; however, the rate of dechlorination did not increase after the second serial transfer. Dechlorination occurred primarily from the meta positions of the biphenyl molecule. Hydrogen could be substituted for pyruvate as the electron donor with equal activity, but when acetate was used as the electron donor a delay in dechlorination was observed. Sulfate and bromethane sulfonate inhibited dechlorination activity. The pyruvate-Aroclor 1242 enrichment also dechlorinated Aroclors 1248, 1254, and 1260; the extent of chlorine removed was the greatest for Aroclor 1254. For comparison, nonautoclaved non-PCB-contaminated Hudson River sediments used in the assay also dechlorinated Aroclors, but only after 12 to 16 weeks of incubation. This suggests that PCB-dechlorinating organisms were also present in these sediments but in numbers lower than those in the enrichment culture.     

Establishment of polychlorinated biphenyl-degrading enrichment culture with predominantly meta dechlorination.

Enrichment of polychlorinated biphenyl (PCB)-dechlorinating microorganisms from PCB-contaminated sediments from the Upper Hudson River, N.Y., was attempted. The enrichment strategy was to use pyruvate as the electron donor and dechlorination of Aroclor 1242 as the electron acceptor. The enrichment medium also contained non-PCB-contaminated Hudson River sediments, which were required for the PCB-dechlorinating activity. An enrichment culture (that had stable PCBT-dechlorinating activity over nine serial transfers during 1 year) was established under these conditions; however, the rate of dechlorination did not increase after the second serial transfer. Dechlorination occurred primarily from the meta positions of the biphenyl molecule. Hydrogen could be substituted for pyruvate as the electron donor with equal activity, but when acetate was used as the electron donor a delay in dechlorination was observed. Sulfate and bromethane sulfonate inhibited dechlorination activity. The pyruvate-Aroclor 1242 enrichment also dechlorinated Aroclors 1248, 1254, and 1260; the extent of chlorine removed was the greatest for Aroclor 1254. For comparison, nonautoclaved non-PCB-contaminated Hudson River sediments used in the assay also dechlorinated Aroclors, but only after 12 to 16 weeks of incubation. This suggests that PCB-dechlorinating organisms were also present in these sediments but in numbers lower than those in the enrichment culture.     

Microbial reductive dehalogenation of polychlorinated biphenyls

Under anaerobic conditions, microbial reductive dechlorination of polychlorinated biphenyls (PCBs) occurs in soils and aquatic sediments. In contrast to dechlorination of supplemented single congeners for which frequently ortho dechlorination has been observed, reductive dechlorination mainly attacks meta and/or para chlorines of PCB mixtures in contaminated sediments, although in a few instances ortho dechlorination of PCBs has been observed. Different microorganisms appear to be responsible for different dechlorination activities and the occurrence of various dehalogenation routes. No axenic cultures of an anaerobic microorganism have been obtained so far. Most probable number determinations indicate that the addition of PCB congeners, as potential electron acceptors, stimulates the growth of PCB-dechlorinating microorganisms. A few PCB-dechlorinating enrichment cultures have been obtained and partially characterized. Temperature, pH, availability of naturally occurring or of supplemented carbon sources, and the presence or absence of H(2) or other electron donors and competing electron acceptors influence the dechlorination rate, extent and route of PCB dechlorination. We conclude from the sum of the experimental data that these factors influence apparently the composition of the active microbial community and thus the routes, the rates and the extent of the dehalogenation. The observed effects are due to the specificity of the dehalogenating bacteria which become active as well as changing interactions between the dehalogenating and non-dehalogenating bacteria. Important interactions include the induced changes in the formation and utilization of H(2) by non-dechlorinating and dechlorinating bacteria, competition for substrates and other electron donors and acceptors, and changes in the formation of acidic fermentation products by heterotrophic and autotrophic acidogenic bacteria leading to changes in the pH of the sediments.     

Influence of Incubation Temperature on the Microbial Reductive Dechlorination of 2,3,4,6-Tetrachlorobiphenyl in Two Freshwater Sediments

We studied the impact of incubation temperatures on the dechlorination of 2,3,4,6-tetrachlorobiphenyl (2346-CB) in two sediments from different climates: polychlorinated biphenyl (PCB)-free sediment from Sandy Creek Nature Center Pond (SCNC) in Athens, Ga., and PCB-contaminated sediment from Woods Pond (WP) in Lenox, Mass. Sediment slurries were incubated anaerobically with 350 (mu)M 2346-CB for 1 year at temperatures ranging from 4 to 66(deg)C. Most of the 2346-CB was dechlorinated between 12 and 34(deg)C in both sediments and, unexpectedly, between 50 and 60(deg)C in WP sediment. This is the first report of PCB dechlorination at thermobiotic temperatures. The data reveal profound differences in dechlorination rate, extent, and products as a function of sediment and temperature. The highest observed rate of dechlorination of 2346-CB to trichlorobiphenyls occurred at 30(deg)C in both sediments, but the rate was higher for WP than for SCNC sediment (46 versus 16 (mu)mol liter(sup-1) day(sup-1)). For SCNC sediment the rate of dechlorination dropped sharply below 30(deg)C, but for WP sediments it was near optimal from 20 to 34(deg)C and then dropped sharply below 20(deg)C. In WP sediment most of the meta chlorines were removed between 8 and 34(deg)C and between 50 and 60(deg)C. para dechlorination was restricted from 18 to 34(deg)C and was optimal at 20(deg)C. ortho dechlorination occurred between 8 and 30(deg)C, with optima around 15 and 27(deg)C, but the extent was highly variable. In SCNC sediment complete meta dechlorination occurred from 12 to 34(deg)C and para dechlorination occurred from 18 to 30(deg)C; both were optimal at 30(deg)C. No ortho dechlorination was observed. Dechlorination products were 246-CB, 236-CB, and 26-CB (both sediments) and 24-CB, 2-CB, and 4-CB (WP sediment). The data suggest that in SCNC sediment similar factors controlled meta and para PCB dechlorination over a broad temperature range (18 to 30(deg)C) but that in WP sediment there were multiple temperature-dependent changes in the factors controlling ortho, meta, and para dechlorination. We attribute the differences observed in the two sediments to differences in their PCB-dechlorinating communities.     

The effect of varying levels of sodium bicarbonate on polychlorinated biphenyl dechlorination in Hudson River sediment cultures

The addition of different concentrations of sodium bicarbonate had a profound effect on 2,3,4,5-chlorobiphenyl (2,3,4,5-CB) dechlorination in Hudson River sediment cultures. The most extensive dechlorination was observed in cultures to which 100 mg l(-1) bicarbonate was added. Cultures amended with 1000 mg l(-1) bicarbonate had the least extensive dechlorination, with 2,4-CB and 2,5-CB as predominant end-products. A significant loss of total chlorinated biphenyl mass was observed in cultures to which < or = 500 mg l(-1) bicarbonate was added, suggesting that degradation beyond chlorinated biphenyls occurred. The dynamics of acetate formation were different among the treatments, with high acetate concentrations detected throughout the 303-day experiment in cultures to which 1000 mg l(-1) bicarbonate had been added. Sodium bicarbonate addition also had a significant impact on bacterial community structure as detected by polymerase chain reaction-denaturant gradient gel electrophoresis (PCR-DGGE) of 16S rRNA gene fragments. Three putative polychlorinated biphenyl (PCB) dechlorinators were identified; one Dehalococcoides-like population was detected in all enrichment cultures, whereas two Dehalobacter-like populations were only detected in the enrichment cultures with the most extensive dechlorination. These results suggest that the availability of bicarbonate, and potentially sodium, may affect PCB dechlorination in Hudson River sediment and thus need to be taken into consideration when assessing the fate of PCBs or implementing bioremediation.     

Temperature determines the pattern of anaerobic microbial dechlorination of Aroclor 1260 primed by 2,3,4,6-tetrachlorobiphenyl in Woods Pond sediment.

Reductive dechlorination of the Aroclor 1260 residue in Woods Pond (Lenox, Mass.) sediment samples was investigated for a year at incubation temperatures from 4 to 66 degrees C. Sediment slurries were incubated anaerobically with and without 2,3,4,6-tetrachlorobiphenyl (2346-CB; 350 microM) as a primer for dechlorination of the Aroclor 1260 residue. Dechlorination of the Aroclor residue occurred only in live samples primed with 2346-CB and only at 8 to 34 degrees C and 50 to 60 degrees C. The extent and pattern of polychlorinated biphenyl (PCB) dechlorination were temperature dependent. At 8 to 34 degrees C, the dechlorination resulted in 28 to 65% decreases of the hexathrough nonachlorobiphenyls and corresponding increases in the tri- and tetrachlorobiphenyls. At 12 to 30 degrees C, 30 to 40% of the hexa- through nonachlorobiphenyls were dechlorinated in just 3 months. The optimal temperature for overall chlorine removal was 20 to 27 degrees C. We observed four different microbial dechlorination processes with different but partially overlapping temperature ranges, i.e., Process N (flanked meta dechlorination) at 8 to 30 degrees C, Process P (flanked para dechlorination) at 12 to 34 degrees C, Process LP (unflanked para dechlorination) at 18 to 30 degrees C, and Process T (a very restricted meta dechlorination of specific hepta- and octachlorobiphenyls) at 50 to 60 degrees C. These temperature ranges should aid in the development of strategies for the enrichment and isolation of the microorganisms responsible for each dechlorination process. The incubation temperature determined the relative dominance of the four PCB dechlorination processes and the extent and products of dechlorination. Hence, understanding the effects of temperature on PCB dechlorination at contaminated sites should assist in predicting the environmental fate of PCBs or planning bioremediation strategies at those sites.     

Two anaerobic polychlorinated biphenyl-dehalogenating enrichments that exhibit different para-dechlorination specificities.

Two anaerobic polychlorinated biphenyl (PCB)-dechlorinating enrichments with distinct substrate specificities were obtained: a 2,3,4,6-tetrachlorobiphenyl (2346-CB) para-dechlorinating enrichment derived from Aroclor 1260-contaminated Woods Pond (Lenox, Mass.) sediment and a 2,4,6-trichlorobiphenyl (246-CB) unflanked para-dechlorinating enrichment derived from PCB-free Sandy Creek Nature Center (Athens, Ga.) sediment. The enrichments have been successfully transferred to autoclaved soil slurries over 20 times by using 300 to 350 microM 2346-CB or 246-CB. Both enrichments required soil for successful transfer of dechlorination activity. The 2346-CB enrichment para dehalogenated, in the absence or presence of 2346-CB, only 4 of 25 tested para halogen-containing congeners: 234-CB, 2345-CB, 2346-CB, and 2,4,6-tribromobiphenyl (246-BrB). In the presence of 246-CB, the 246-CB enrichment para dehalogenated 23 of the 25 tested congeners. However, only three congeners (34-CB, 2346-CB, and 246-BrB) were dehalogenated in the absence of 246-CB, indicating that these specific congeners initiate dehalogenation in this enrichment culture. The addition of the 2346-CB (para)-dechlorinating enrichment did not further stimulate the 2346-CB-primed dechlorination of the Aroclor 1260 residue in Woods Pond sediment samples. Compared to the addition of the primer 246-CB or the 246-CB unflanked para-dechlorinating enrichment alone, the addition of both 246-CB (300 microM) and the 246-CB enrichment stimulated the unflanked para dechlorination of the Aroclor 1260 residue in Woods Pond sediments. These results indicate that the two enrichments contain different PCB-dechlorinating organisms, each with high substrate specificities. Furthermore, bioaugmentation with the enrichment alone did not stimulate the desired dechlorination in PCB-contaminated Woods Pond sediment.     

Extensive degradation of Aroclors and environmentally transformed polychlorinated biphenyls by Alcaligenes eutrophus H850.

We have isolated and characterized a strain of Alcaligenes eurtrophus, designated H850, that rapidly degrades a broad and unusual spectrum of polychlorinated biphenyls (PCBs) including many tetra- and pentachlorobiphenyls and several hexachlorobiphenyls. This strain, which was isolated from PCB-containing dredge spoils by enrichment on biphenyl, grows well on biphenyl and 2-chlorobiphenyl but poorly on 3- and 4-chlorobiphenyl. Capillary gas-chromatographic analysis showed that biphenyl-grown resting cells of H850 degraded the components of 38 of the 41 largest peaks of Aroclor 1242 and 15 of the 44 largest peaks of Aroclor 1254, resulting in an overall reduction of PCBs by 81% for Aroclor 1242 (10 ppm) and 35% for Aroclor 1254 (10 ppm) in 2 days. Furthermore, H850 metabolized the predominantly ortho-substituted PCB congeners that resulted from the environmental transformation of the more highly chlorinated congeners of Aroclor 1242 by the upper Hudson River anaerobic meta-, para-dechlorination agent system C (J. F. Brown, R. E. Wagner, Jr., D. L. Bedard, M. J. Brennan, J. C. Carnahan, R. J. May, and J. J. Tofflemire, Northeast Environ. Sci. 3:167-179, 1984). The congener selectivity patterns indicate that a two-step process consisting of anaerobic dechlorination followed by oxidation by H850 can effectively degrade all of the congeners in Aroclor 1242 and possibly all those in Aroclor 1254.     

Extensive degradation of Aroclors and environmentally transformed polychlorinated biphenyls by Alcaligenes eutrophus H850

We have isolated and characterized a strain of Alcaligenes eurtrophus, designated H850, that rapidly degrades a broad and unusual spectrum of polychlorinated biphenyls (PCBs) including many tetra- and pentachlorobiphenyls and several hexachlorobiphenyls. This strain, which was isolated from PCB-containing dredge spoils by enrichment on biphenyl, grows well on biphenyl and 2-chlorobiphenyl but poorly on 3- and 4-chlorobiphenyl. Capillary gas-chromatographic analysis showed that biphenyl-grown resting cells of H850 degraded the components of 38 of the 41 largest peaks of Aroclor 1242 and 15 of the 44 largest peaks of Aroclor 1254, resulting in an overall reduction of PCBs by 81% for Aroclor 1242 (10 ppm) and 35% for Aroclor 1254 (10 ppm) in 2 days. Furthermore, H850 metabolized the predominantly ortho-substituted PCB congeners that resulted from the environmental transformation of the more highly chlorinated congeners of Aroclor 1242 by the upper Hudson River anaerobic meta-, para-dechlorination agent system C (J. F. Brown, R. E. Wagner, Jr., D. L. Bedard, M. J. Brennan, J. C. Carnahan, R. J. May, and J. J. Tofflemire, Northeast Environ. Sci. 3:167-179, 1984). The congener selectivity patterns indicate that a two-step process consisting of anaerobic dechlorination followed by oxidation by H850 can effectively degrade all of the congeners in Aroclor 1242 and possibly all those in Aroclor 1254.