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.     

Survival and naphthalene-degrading activity of Rhodococcus sp. strain 1BN in soil microcosms

AIMS: The survival and activity of Rhodococcus sp. strain 1BN, inoculated into naphthalene-contaminated sandy-loam soil microcosms, were studied using classical and molecular methods. METHODS AND RESULTS: The naphthalene-degrading activity of 1BN in microcosms was examined through viable counts, CO2 production and naphthalene consumption, while its survival after inoculation was monitored by detecting the contemporary presence of alkane and naphthalene degradative genes and by analysing the 16S rDNA specific restriction profile. The inoculation of 1BN did not significantly enhance naphthalene degradation in the naphthalene-contaminated native soil, where 1BN maintained its catabolic activity also when in the presence of indigenous microflora. Instead the rate of naphthalene degradation by the inoculated 1BN was greater in sterile naphthalene-contaminated soil. The level of 1BN was only slightly higher after inoculation regardless of whether indigenous naphthalene-degrading bacteria were present or not and 1BN remained viable even when the substrate was depleted. CONCLUSIONS: This study documents the colonization and growth of 1BN in a non-sterile, naphthalene-added, sandy-loam soil having an active indigenous naphthalene-degrading population. SIGNIFICANCE AND IMPACT OF THE STUDY: An active and well-established naphthalene-degrading bacterial population in the native soil did not hamper the survival of the introduced 1BN that, through its activity, enhanced the mineralization rate of naphthalene.     

The microbial degradation of azimsulfuron and its effect on the soil bacterial community

AIMS: Azimsulfuron is a recently introduced sulfonylurea herbicide useful in controlling weeds in paddy fields. To date very little information is available on the biodegradation of this pesticide and on its effect on the soil microbial community. The aim of this work was to study its biodegradation both in slurry soil microcosms and in batch tests with mixed and pure cultures. METHODS AND RESULTS: Azimsulfuron was applied to forest bulk soil in order to study its effect on the structure of the bacterial soil community, as detectable by denaturant gradient gel electrophoresis (DGGE) analyses. Biodegradation and abiotic processes were investigated by HPLC analyses. In addition, a microbial consortium was selected, that was able to use azimsulfuron as the sole energy and carbon source. One of the metabolites produced by the consortium was isolated and identified through LC-MS analyses. Cultivable bacteria of the consortium were isolated and identified by 16S rDNA sequencing (1400 bp). CONCLUSIONS: Azimsulfuron treatment seems to have the ability to cause changes in the bacterial community structure that are detectable by DGGE analyses. It is easily biodegraded both in microcosms and in batch tests, with the formation of an intermediate that was identified as 2-methyl-4-(2-methyl-2H-tetrazol-5-yl)-2H-pyrazole-3-sulfonamide. SIGNIFICANCE AND IMPACT OF THE STUDY: The study increases the knowledge on the biodegradation of azimsulfuron and its effects on the soil microbiota.     

Responses of the anaerobic bacterial community to addition of organic C in chromium(VI)- and iron(III)-amended microcosms

Chromium (VI) is toxic to microorganisms and can inhibit the biodegradation of organic pollutants in contaminated soils. We used microcosms amended with either glucose or protein (to drive bacterial community change) and Fe(III) (to stimulate iron-reducing bacteria) to study the effect of various concentrations of Cr(VI) on anaerobic bacterial communities. Microcosms were destructively sampled based on microbial activity (measured as evolution of CO2) and analyzed for the following: (i) dominant bacterial community by PCR-denaturing gradient gel electrophoresis (DGGE) of the 16S rRNA gene; (ii) culturable Cr-resistant bacteria; and (iii) enrichment of iron-reducing bacteria of the Geobacteraceae family by real-time PCR. The addition of organic C stimulated the activities of anaerobic communities. Cr(VI) amendment resulted in lower rates of CO2 production in glucose microcosms and a slow mineralization phase in protein-amended microcosms. Glucose and protein amendments selected for different bacterial communities. This selection was modified by the addition of Cr(VI), since some DGGE bands were intensified and new bands appeared in Cr(VI)-amended microcosms. A second dose of Cr(VI), added after the onset of activity, had a strong inhibitory effect when higher levels of Cr were added, indicating that the developing Cr-resistant communities had a relatively low tolerance threshold. Most of the isolated Cr-resistant bacteria were closely related to previously studied Cr-resistant anaerobes, such as Pantoea, Pseudomonas, and Enterobacter species. Geobacteraceae were not enriched during the incubation. The studied Cr(VI)-contaminated soil contained a viable anaerobic bacterial community; however, Cr(VI) altered its composition, which could affect the soil biodegradation potential.     

Responses of the Anaerobic Bacterial Community to Addition of Organic C in Chromium(VI)- and Iron(III)-Amended Microcosms

Chromium (VI) is toxic to microorganisms and can inhibit the biodegradation of organic pollutants in contaminated soils. We used microcosms amended with either glucose or protein (to drive bacterial community change) and Fe(III) (to stimulate iron-reducing bacteria) to study the effect of various concentrations of Cr(VI) on anaerobic bacterial communities. Microcosms were destructively sampled based on microbial activity (measured as evolution of CO₂) and analyzed for the following: (i) dominant bacterial community by PCR-denaturing gradient gel electrophoresis (DGGE) of the 16S rRNA gene; (ii) culturable Cr-resistant bacteria; and (iii) enrichment of iron-reducing bacteria of the Geobacteraceae family by real-time PCR. The addition of organic C stimulated the activities of anaerobic communities. Cr(VI) amendment resulted in lower rates of CO₂ production in glucose microcosms and a slow mineralization phase in protein-amended microcosms. Glucose and protein amendments selected for different bacterial communities. This selection was modified by the addition of Cr(VI), since some DGGE bands were intensified and new bands appeared in Cr(VI)-amended microcosms. A second dose of Cr(VI), added after the onset of activity, had a strong inhibitory effect when higher levels of Cr were added, indicating that the developing Cr-resistant communities had a relatively low tolerance threshold. Most of the isolated Cr-resistant bacteria were closely related to previously studied Cr-resistant anaerobes, such as Pantoea, Pseudomonas, and Enterobacter species. Geobacteraceae were not enriched during the incubation. The studied Cr(VI)-contaminated soil contained a viable anaerobic bacterial community; however, Cr(VI) altered its composition, which could affect the soil biodegradation potential.     

Low temperature bioremediation of oil-contaminated soil using biostimulation and bioaugmentation with a Pseudomonas sp. from maritime Antarctica

AIMS: To identify native Antarctic bacteria capable of oil degradation at low temperatures. METHODS AND RESULTS: Oil contaminated and pristine soils from Signy Island (South Orkney Islands, Antarctica) were examined for bacteria capable of oil degradation at low temperatures. Of the 300 isolates cultured, Pseudomonas strain ST41 grew on the widest range of hydrocarbons at 4 degrees C. ST41 was used in microcosm studies of low temperature bioremediation of oil-contaminated soils. Microcosm experiments showed that at 4 degrees C the levels of oil degradation increased, relative to the controls, with (i) the addition of ST41 to the existing soil microbial population (bioaugmentation), (ii) the addition of nutrients (biostimulation) and to the greatest extent with (iii) a combination of both treatments (bioaugmentation and biostimulation). Addition of water to oil contaminated soil (hydration) also enhanced oil degradation, although less than the other treatments. Analysis of the dominant species in the microcosms after 12 weeks, using temporal temperature gradient gel electrophoresis, showed Pseudomonas species to be the dominant soil bacteria in both bioaugmented and biostimulated microcosms. CONCLUSIONS: Addition of water and nutrients may enhance oil degradation through the biostimulation of indigenous oil-degrading microbial populations within the soil. However, bioaugmentation with Antarctic bacteria capable of efficient low temperature hydrocarbon degradation may enhance the rate of bioremediation if applied soon after the spill. SIGNIFICANCE AND IMPACT OF THE STUDY: In the future, native soil bacteria could be of use in bioremediation technologies in Antarctica.     

Involvement of fungi and bacteria in enhanced and nonenhanced biodegradation of carbendazim and other benzimidazole compounds in soil

The relationship between chemical structure and the enhancement of microbial degradation of three benzimidazole compounds in soil was determined. Preapplication of methyl benzimidazole-2-ylcarbamate (carbendazim or MBC), 2-aminobenzimidazole (2AB), and benzimidazole enhanced their degradation upon repeated application (self-enhanced degradation). MBC and 2AB cross-enhanced the degradation of each of these two compounds, whereas benzimidazole did not enhance the degradation of MBC. Thiabendazole (TBZ) did not enhance its own degradation or cross-enhance the degradation of MBC. No increase in the number of MBC-degrading fungi or in the capacity of soilborne fungi to degrade MBC was detected in soil exhibiting enhanced MBC degradation (MBC-history). A sharp increase in esterolytic activity in the microsomal fraction of Alternaria alternata capable of degrading MBC in culture was induced by the presence of MBC in the growth medium. 2AB was the main metabolite of MBC that accumulated in A. alternata cultures and in cell-free preparations. MBC was degraded much faster by mixed bacterial cultures that originated from MBC-history soil than in cultures from MBC-nonhistory soil. Fluctuations in the MBC degrading capacity of mixed bacterial cultures occurred during repeated subculturing of the mixed culture. Inoculation of nonhistory soil with mixed bacterial cultures resulted in enhanced MBC degradation, whereas inoculation with A. alternata did not enhance MBC degradation. It is suggested that while fungi contribute to MBC dissipation in soil, bacteria have a greater role in enhanced biodegradation of MBC in soil.     

Horizontal transfer of catabolic plasmids in the process of naphthalene biodegradation in model soil systems

The process of naphthalene degradation by indigenous, introduced, and transconjugant strains was studied in laboratory soil microcosms. Conjugation transfer of catabolic plasmids was demonstrated in naphthalene-contaminated soil. Both indigenous microorganisms and an introduced laboratory strain BS394 (pNF142::TnMod-OTc) served as donors of these plasmids. The indigenous bacterial degraders of naphthalene isolated from soil were identified as Pseudomonas putida and Pseudomonas fluorescens. The frequency of plasmid transfer in soil was 10(-5)-10(-4) per donor cell. The activity of the key enzymes of naphthalene biodegradation in indigenous and transconjugant strains was studied. Transconjugant strains harboring indigenous catabolic plasmids possessed high salicylate hydroxylase and low catechol-2,3-dioxygenase activities, in contrast to indigenous degraders, which had a high level of catechol-2,3-dioxygenase activity and a low level of salicylate hydroxylase. Naphthalene degradation in batch culture in liquid mineral medium was shown to accelerate due to cooperation of the indigenous naphthalene degrader P. fluorescens AP1 and the transconjugant strain P. putida KT2442 harboring the indigenous catabolic plasmid pAP35. The role of conjugative transfer of naphthalene biodegradation plasmids in acceleration of naphthalene degradation was demonstrated in laboratory soil microcosms.     

Effects of randomly methylated-β-cyclodextrins (RAMEB) on the bioavailability and aerobic biodegradation of polychlorinated biphenyls in three pristine soils spiked with a transformer oil

The low bioavailability of polychlorinated biphenyls (PCBs) in soils often results in their slow and partial aerobic biodegradation. The process can be enhanced by supplementing soils with cyclodextrins. However, pure cyclodextrins are expensive and we have therefore explored the use of a less costly technical grade mixture of randomly methylated-beta-cyclodextrins (RAMEB). RAMEB was tested at 0, 1, 3 and 5% (w/w) in the aerobic bioremediation and detoxification of a loamy-, a humic- and a sandy-soil, each artificially contaminated with a PCB-containing transformer oil (added PCBs: about 450 or 700 mg/kg), inoculated with an exogenous aerobic PCB-biodegrading bacterial co-culture and treated in slurry- and solid-phase laboratory conditions. Significant depletions of the spiked PCBs were observed in all microcosms of the three soils after 90 days of treatment; however, interesting yields of PCB dechlorination and detectable decreases of the original soil ecotoxicity were observed in the slurry-phase microcosms. RAMEB generally enhanced PCB-metabolism with effects which were dependent on the concentration at which it was applied, the physical-chemical nature of the amended soil, and the soil treatment conditions employed. RAMEB, which was slowly metabolized by soil microorganisms, enhanced the presence of PCBs and PCB-cometabolizing bacteria in the soil-water phase, suggesting that RAMEB enhances aerobic biodegradation of PCBs by increasing pollutant bioavailability in soil microcosms.     

Soya lecithin effects on the aerobic biodegradation of polychlorinated biphenyls in an artificially contaminated soil

The effects of the phytogenic surfactant soya lecithin (SL) on the aerobic biodegradation of polychlorinated biphenyls (PCBs) spiked into a synthetic soil were studied. Soil was spiked with both biphenyl (4 g/kg) and Fenclor 42 (1,000 mg/kg) and treated in aerobic batch slurry-phase microcosms (17.5% w/v). Microcosms were prepared either with or without the exogenous aerobic PCB-dechlorinating bacterial co-culture ECO3 (inoculum:10(8) CFU/mL). In some inoculated microcosms, SL was added at 15 or 30 g/kg. Indigenous bacteria having the capability of metabolizing biphenyl and 2-chlorobenzoic acid were found to develop in the microcosms during the experiment, and were responsible for the significant PCB biodegradation and dechlorination observed in the uninoculated controls. The addition of ECO3 bacteria resulted in only a slight PCB biodegradation increase. In the presence of SL, a higher availability of biphenyl- and chlorobenzoic acid-degrading bacteria and higher PCB biodegradation and dechlorination yields were observed; the effects increased proportionally with the concentration of the applied SL. A significant decrease of soil ecotoxicity was also revealed in SL-supplemented microcosms. At both concentrations, SL was found to be a good carbon source for both the indigenous and ECO3 bacteria, as well as a product capable of enhancing the PCB bioavailability in the microcosms.     

Comparison of bioremediation strategies for soil impacted with petrochemical oily sludge

Different bioremediation techniques (natural attenuation, biostimulation and bioaugmentation) in contaminated soils with two oily sludge concentrations (1.5% and 6.0%) in open and closed microcosms systems were assessed during 90 days. The results showed that the highest biodegradation rates were obtained in contaminated soils with 6% in closed microcosms. Addition of microbial consortium and nutrients in different concentrations demonstrated higher biodegradation rate of total petroleum hydrocarbons (TPH) than those of the natural attenuation treatment. Soils treated in closed microcosms showed highest removal rate (84.1 ± 0.9%) when contaminated at 6% and bacterial consortium and nutrients in low amounts were added. In open microcosms, the soil contaminated at 6% using biostimulation with the highest amounts of nutrients (C:N:P of 100:10:1) presented the highest degradation rate (78.7 ± 1.3%). These results demonstrate that the application of microbial consortium and nutrients favored biodegradation of TPH present in oily sludge, indicating their potential applications for treatment of the soils impacted with this important hazardous waste.     

Reduction of Perchlorate and Nitrate by Microbial Communities in Vadose Soil

Perchlorate contamination is a concern because of the increasing frequency of its detection in soils and groundwater and its presumed inhibitory effect on human thyroid hormone production. Although significant perchlorate contamination occurs in the vadose (unsaturated) zone, little is known about perchlorate biodegradation potential by indigenous microorganisms in these soils. We measured the effects of electron donor (acetate and hydrogen) and nitrate addition on perchlorate reduction rates and microbial community composition in microcosm incubations of vadose soil. Acetate and hydrogen addition enhanced perchlorate reduction, and a longer lag period was observed for hydrogen (41 days) than for acetate (14 days). Initially, nitrate suppressed perchlorate reduction, but once perchlorate started to be degraded, the process was stimulated by nitrate. Changes in the bacterial community composition were observed in microcosms enriched with perchlorate and either acetate or hydrogen. Denaturing gradient gel electrophoresis analysis and partial sequencing of 16S rRNA genes recovered from these microcosms indicated that formerly reported perchlorate-reducing bacteria were present in the soil and that microbial community compositions were different between acetate- and hydrogen-amended microcosms. These results indicate that there is potential for perchlorate bioremediation by native microbial communities in vadose soil.     

Reduction of perchlorate and nitrate by microbial communities in vadose soil

Perchlorate contamination is a concern because of the increasing frequency of its detection in soils and groundwater and its presumed inhibitory effect on human thyroid hormone production. Although significant perchlorate contamination occurs in the vadose (unsaturated) zone, little is known about perchlorate biodegradation potential by indigenous microorganisms in these soils. We measured the effects of electron donor (acetate and hydrogen) and nitrate addition on perchlorate reduction rates and microbial community composition in microcosm incubations of vadose soil. Acetate and hydrogen addition enhanced perchlorate reduction, and a longer lag period was observed for hydrogen (41 days) than for acetate (14 days). Initially, nitrate suppressed perchlorate reduction, but once perchlorate started to be degraded, the process was stimulated by nitrate. Changes in the bacterial community composition were observed in microcosms enriched with perchlorate and either acetate or hydrogen. Denaturing gradient gel electrophoresis analysis and partial sequencing of 16S rRNA genes recovered from these microcosms indicated that formerly reported perchlorate-reducing bacteria were present in the soil and that microbial community compositions were different between acetate- and hydrogen-amended microcosms. These results indicate that there is potential for perchlorate bioremediation by native microbial communities in vadose soil.     

Enhanced Degradation of n-Hexadecane in Diesel Fuel Contaminated Soil by the Addition of Fermented Whey

The objective of this work has been to investigate the possibility of using fermented whey as an organic growth supplement in order to enhance the aerobic degradation of n-hexadecane in soil. Fermented whey was added at different dosages to nutrient amended soil microcosms contaminated with 5000 mg diesel fuel kg^?1 dry weight (dw). The target substance was ¹⁴C-labeled n-hexadecane, and the biodegradation was monitored by analysis of evolved ¹⁴CO₂. Biodegradation curves were fitted to a three-half-order kinetics model. Enhanced biodegradation was observed in sand at 7 and 22°C and in loamy sand at 22°C but the effect was most pronounced in the sand soil at 22°C. The addition of 6 or 60 ml fermented whey kg^{? 1} soil dw increased the degree of n-hexadecane biodegradation at the end of the experiment, 167 days, from 49% in the untreated sand to 60 or 67%, respectively. This increase in biodegradation was characterized by an increase in the amount of substrate biodegradation by first-order kinetics despite a decrease in the first order rate constant, k₁. The highest concentration of fermented whey, 60 ml kg^?1, gave rise to substrate competition, diauxie, which resulted in an extended lag phase.     

Simazine treatment history determines a significant herbicide degradation potential in soils that is not improved by bioaugmentation with Pseudomonas sp. ADP

AIMS: To study biological removal of the herbicide simazine in soils with different history of herbicide treatment and to test bioaugmentation with a simazine-degrading bacterial strain. METHODS AND RESULTS: Simazine removal was studied in microcosms prepared with soils that had been differentially exposed to this herbicide. Simazine removal was much higher in previously exposed soils than in unexposed ones. Terminal restriction fragment length polymorphism analysis and multivariate analysis showed that soils previously exposed to simazine contained bacterial communities that were significantly impacted by simazine but also had an increased resilience. The biodegradation potential was also related to the presence of high levels of the atz-like gene sequences involved in simazine degradation. Bioaugmentation with Pseudomonas sp. ADP resulted in an increased initial rate of simazine removal, but this strain scarcely survived. After 28 days, residual simazine removals were the same in bioaugmented and not bioaugmented microcosms. CONCLUSIONS: In soils with a history of simazine treatment bacterial communities were able to overcome subsequent impacts with the herbicide. The success of bioaugmentation was limited by the low survival of the introduced strain. SIGNIFICANCE AND IMPACT OF THE STUDY: Conclusions from this work provided insights on simazine biodegradation potential of soils and the convenience of bioaugmentation.     

Synergistic relationships in algal-bacterial microcosms for the treatment of aromatic pollutants

The potential of algal-bacterial microcosms was studied for the biodegradation of salicylate, phenol and phenanthrene. The isolation and characterization of aerobic bacterial strains capable of mineralizing each pollutant were first conducted. Ralstonia basilensis was isolated for salicylate degradation, Acinetobacter haemolyticus for phenol and Pseudomonas migulae and Sphingomonas yanoikuyae for phenanthrene. The green alga Chlorella sorokiniana was then cultivated in the presence of the pollutants at different concentrations, showing increasing inhibitory effects in the following order: salicylate < phenol < phenanthrene. The synergistic relationships in the algal-bacterial microcosms were clearly demonstrated, since for the three contaminants tested, a substantial removal (>85%) was recorded only in the systems inoculated with both algae and bacteria and incubated under continuous lighting. This study presents, to our knowledge, the first reported case of photosynthesis-enhanced biodegradation of toxic aromatic pollutants by algal-bacterial microcosms in a one-stage treatment.     

Horizontal transfer of catabolic plasmids in the process of naphthalene biodegradation in model soil systems

The process of naphthalene degradation by indigenous, introduced, and transconjugant strains was studied in laboratory soil microcosms. Conjugation transfer of catabolic plasmids was demonstrated in naphthalene-contaminated soil. Both indigenous microorganisms and an introduced laboratory strain BS394 (pNF142::TnMod-OTc) served as donors of these plasmids. The indigenous bacterial degraders of naphthalene isolated from soil were identified as Pseudomonas putida and Pseudomonas fluorescens. The frequency of plasmid transfer in soil was 10^?5–10^?4 per donor cell. The activity of the key enzymes of naphthalene biodegradation in indigenous and transconjugant strains was studied. Transconjugant strains harboring indigenous catabolic plasmids possessed high salicylate hydroxylase and low catechol-2,3-dioxygenase activities, in contrast to indigenous degraders, which had a high level of catechol-2,3-dioxygenase activity and a low level of salicylate hydroxylase. Naphthalene degradation in batch culture in liquid mineral medium was shown to accelerate due to cooperation of the indigenous naphthalene degrader P. fluorescens AP1 and the transconjugant strain P. putida KT2442 harboring the indigenous catabolic plasmid pAP35. The role of conjugative transfer of naphthalene biodegradation plasmids in acceleration of naphthalene degradation was demonstrated in laboratory soil microcosms.     

Anaerobic Benzene Biodegradation: A Microcosm Survey

Benzene-amended microcosms prepared with saturated soil or sediment from five hydrocarbon-contaminated sites and one pristine site were monitored for a year and a half to determine the rate of benzene biodegradation under a variety of electron-accepting conditions. Sustainable benzene degradation was observed under specific conditions in microcosms from four of the six sites. Significant differences were observed between sites with respect to lag times before the onset of degradation, rates of degradation, sustainability of the activity, and environmental conditions supporting degradation. Benzene degradation was observed under sulfate-reducing, nitrate-reducing, and iron(III)-reducing conditions, but not under methanogenic conditions. The presence of competing substrates such as toluene, xylenes, and ethylbenzene was found to inhibit anaerobic benzene degradation in microcosms where sulfate or possibly nitrate was the electron acceptor for benzene degradation, but not in microcosms from where iron(III) was the electron acceptor. The presence of organic matter, indicated by a high fraction organic carbon (f_oc), also appeared to inhibit the biodegradation of benzene; microcosms constructed with soils with the highest f_oc exhibited the longest lag times before the onset of benzene degradation. The initial extent of hydrocarbon contamination did not appear to correlate with anaerobic benzene-degrading activity.     

Bioremediation of polyaromatic hydrocarbon contaminated soils by native microflora and bioaugmentation with Sphingobium chlorophenolicum strain C3R: A feasibility study in solid- and slurry-phase microcosms

The aim of the research was to verify if a Sphingobium chlorophenolicum strain C3R was effective in the degradation of phenanthrene (Ph) in agricultural soil co-contaminated by metals and mixtures of PAHs. The presence of PAHs in mixtures produced interactive effects which could either increase or decrease the utilization rate of Ph by C3R and by the native bacterial microflora. Bioaugmentation significantly improved the biodegradation rate of Ph in the presence of both cadmium and arsenic and PAH mixtures. The augmented C3R strain persisted in inoculated microcosms as monitored by the DGGE analysis and outcompeted some indigenous bacteria. The potential role of the soil bacteria in PAH degradation could be envisaged. The results indicate the applicability of S. chlorophenolicum C3R toward in situ bioremediation of sites contaminated with phenanthrene alone or co-contaminated with low molecular weight PAHs and with cadmium and arsenate.     

Use of a Whole-Cell Biosensor and Flow Cytometry to Detect AHL Production by an Indigenous Soil Community During Decomposition of Litter

Quorum sensing, mediated by acylated homoserine lactones (AHLs), is well described for pure culture bacteria, but few studies report detection of AHL compounds in natural bacterial habitats. In this study, we detect AHL production during a degradation process in soil by use of whole-cell biosensor technology and flow cytometry analysis. An indigenous soil bacterium, belonging to the family of Enterobacteriaceae, was isolated and transformed with a low-copy plasmid harboring a gene encoding an unstable variant of the green fluorescent protein (gfpASV) fused to the AHL-regulated P_luxI promoter originating from Vibrio fischeri. This resulted in a whole-cell biosensor, responding to the presence of AHL compounds. The biosensor was introduced to compost soil microcosms amended with nettle leaves. After 3 days of incubation, cells were extracted and analyzed by flow cytometry. All microcosms contained induced biosensors. From these microcosms, AHL producers were isolated and further identified as species previously shown to produce AHLs. The results demonstrate that AHL compounds are produced during degradation of litter in soil, indicating the presence of AHL-mediated quorum sensing in this environment.