Influence of chlorobenzoic acids on the growth and degradation potentials of PCB-degrading microorganisms

The biodegradation of polychlorinated biphenyls (PCBs) by diverse bacteria including those utilized in this study is often incomplete, a concomitant accumulation of chlorobenzoic acids (CBAs) are released as dead-end products. The build-up of these metabolites in the growth medium may result in feed-back inhibition and impede PCB biotransformation. In this investigation using GC-ECD and HPLC analyses, we confirmed that CBAs inhibit growth and PCB biodegradation potentials of five tropical bacteria namely, Pseudomonas aeruginosa SA-1, Enterobacter sp. SA-2, Ralstonia sp. SA-3, Ralstonia sp. SA-5 and Pseudomonas sp. SA-6. Among the four CBAs (2-CBA, 3-CBA, 4-CBA acids and 2,3-diCBA), 3-CBA was the strongest inhibitor followed by 4-CBA. Furthermore, we found that 3-CBA heavily inhibited growth of SA-3 and SA-6 on monochlorobiphenyls by 82–90% while elimination rate was inhibited by 71–88%. In the case of 2,3-diCBA, inhibition was generally less than 60%. However, effects of both acids were stronger in SA-3 than SA-6. We also found that 3-CBA and 2,3-diCBA completely inhibited carbon-chloride cleavage of 2-CB and 3-CB since cultivation in the absence of the acids resulted in recovery of 23–50% chloride in the culture fluids of organisms. These findings may therefore, have practical and ecological significance and are useful for improving the efficiency and the stability of some biological treatment processes.     

Degradation of chlorobiphenyls catalyzed by the bph-encoded biphenyl-2,3-dioxygenase and biphenyl-2,3-dihydrodiol-2,3-dehydrogenase of Pseudomonas sp. LB400

Abstract In order to characterize the metabolites produced in vivo by biphenyl-2,3-dioxygenase and biphenyl-2,3-dihydrodiol-2,3-dehydrogenase, the first two enzymes of the (polychloro)biphenyl catabolic pathway encoded by the bph locus of Pseudomonas sp. LB400, recombinant E. coli strains expressing the respective genes were constructed. Biphenyl-2,3-dioxygenase attack on 2,2'- or 2,4'-dichlorobiphenyl was shown to give rise to virtually quantitative ortho -dechlorination of these congeners by hydroxylation at the chlorinated carbon 2 and its unsubstituted neighbour. Elimination of hydrochloric acid directly leads to 2,3-dihydroxy-chlorobiphenyls and obviates the need for biphenyl-2,3-dihydrodiol-2,3-dehydrogenase for the catabolism of such congeners.     

Biodegradation of chlorinated alkanes and their commercial mixtures by Pseudomonas sp. strain 273

The biodegradation of chlorinated alkanes was studied under oxic conditions with the objective of identifying favorable and unfavorable intramolecular chlorination sequences with respect to the enzymes studied. Several dehalogenating bacterial strains were screened for their ability to degrade middle-chain polychlorinated alkanes as well as a commercial mixture. Of the organisms tested, the most promising was Pseudomonas sp. strain 273, which possesses an oxygenolytic dehalogenase. The effects of carbon chain length (C₆–C₁₆), halogen position, and overall chlorine content (14–61% w/w) were examined using both commercially available compounds and molecules synthesized in our laboratory. The effects of co-substrates, solvents, and inducing agents were also studied. The results with pure chlorinated alkanes showed that the relative positions of the chlorine atoms strongly influenced the total amount of dehalogenation achieved. The greatest dehalogenation yields were associated with terminally chlorinated alkanes. The α- and α,ω-chlorinated compounds yielded similar results. Vicinal chlorination had the most dramatic impact on degradation. When present on both ends or at the center of the molecule, no dehalogenation was detected. Although partial dehalogenation of 1,2-dichlorodecane was observed, it was likely due to a combination of β-oxidation and an abiotic mechanism. Cereclor S52 was appreciably dehalogenated in shake flasks only when 1,10-dichlorodecane was present as a co-substrate and after increasing the oil surface area through mechanical emulsification, demonstrating the importance of abiotic factors in degrading commercial polychlorinated alkane mixtures.     

Effect of chlorobenzoates on the degradation of polychlorinated biphenyls (PCB) by Pseudomonas stutzeri

Chlorobenzoic acids (CBA) are frequently dead-end products of partial aerobic biodegradation of polychlorinated biphenyls (PCB). When CBA produced from PCB accumulate in the growth medium, they can inhibit the bacterial growth and consequently, slow down PCB biodegradation. In this study, the effects of seven mono- and dichlorinated CBA on growth of Pseudomonas stutzeri on different substrates and on the PCB degradation by this strain in a liquid mineral medium were tested. 3-CBA was the strongest growth inhibitor for P. stutzeri growing on glucose, benzoate and biphenyl. It was found to inhibit heavily the elimination of some di- and trichlorinated biphenyls. In contrast, its influence on the elimination of more chlorinated congeners was much less significant.The authors are with the Department of Biochemical Technology, Faculty of Chemical Technology, Slovak Technical University, 812 37 Bratislava, Slovakia.     

Analysis of changes in congener selectivity during PCB degradation by Burkholderia sp. strain TSN101 with increasing concentrations of PCB and characterization of the bphBCD genes and gene products

We isolated and characterized a gram-negative bacterium, Burkholderia sp. strain TSN101, that can degrade polychlorinated biphenyls (PCBs) at concentrations as high as 150 μg Kaneclor 300/ml, a PCB mixture equivalent to Aroclor 1242. Growing cells of strain TSN101 degraded most of the tri- and tetrachlorobiphenyls in medium containing 25 μg Kaneclor 300/ml. Using PCB concentrations of 50–150 μg of Kaneclor 300/ml, the congener selectivity pattern was different and the pattern of chlorine substitution strongly affected degradation of some congeners. At 25 μg Kaneclor 300/ml, strain TSN101 degraded di- and trichlorinated congeners with chlorine substitutions at both the ortho and the para positions. At higher concentrations of Kaneclor 300, di- and trichlorobiphenyls with ortho substituents in both phenyl rings were not degraded well. Trichlorobiphenyls with para and meta substitutents were degraded equally well at all concentrations studied. The ability of strain TSN101 to degrade ortho and para-substituted congeners was confirmed using a defined PCB mixture with chlorine substituents at 2′- and 4′-positions. A 5-kb DNA fragment containing the bphBCD genes was cloned and sequenced. Comparison of the deduced amino acid sequences of these genes with related proteins indicated 99 and 98% sequence similarity to the BphB and BphD of Comamonas testosteroni strain B-356, respectively. The bphC gene product showed 74% sequence similarity to the BphC of Burkholderia cepacia strain LB400 and exhibited a narrow substrate specificity with strong affinity for 2,3-dihydroxybiphenyl. A bphC-disrupted mutant of Burkholderia sp. strain TSN101, constructed by gene replacement, lost the ability to utilize biphenyl, thus supporting the role of the cloned bph gene in biphenyl metabolism. Received: 18 February 1997 / Accepted: 19 August 1997     

Metabolism of hydroxybiphenyl and chloro-hydroxybiphenyl by biphenyl/chlorobiphenyl degradingPseudomonas testosteroni,strain B-356

Summary A biphenyl (BP) and chlorobiphenyl (CBP) metabolizingPseudomonas testosteroni, strain B-356 was also capable of utilizing 2-, 3-, and 4-hydroxybiphenyl. Data presented here suggest that utilization of biphenyl and mono-subtituted biphenyls involves the enzymes of the same pathway. Chloro-hydroxybiphenyls were also metabolized by strain B-356. The unsubstituted ring is first hydroxylated in position 2 and 3 and then cleaved in ameta 1, and 2, position to ultimately generate the benzoic acid derivatives. Since strain B-356 was capable of utilizing benzoic acid and mono-hydroxybenzoic acids, the utilization of biphenyl, 2-, 3-, and 4-hydroxybiphenyl is complete at non-toxic concentrations of the substrates. Chlorobenzoic acids and chloro-hydroxybenzoic acids were not metabolized further by this strain. Studies usingPseudomonas putida, strain KT2440 carrying cloned BP/CBP genes from strain B-356 provided further evidence for the presence of a common pathway for the metabolism of the above compounds inP. testosteroni, strain B-356. Suggestions are made on significance of the broad substrate specificity of the enzymes of biphenyl/chlorobiphenyl pathway in regard to their possible origin and in relation to PCB mixture degradation.     

Extensive biodegradation of highly chlorinated biphenyl and Aroclor 1242 by Pseudomonas aeruginosa TMU56 isolated from contaminated soils

A bacterial strain, designated TMU56, was isolated from soil that had been contaminated with electrical transformer fluid (Askarel) for over 35 years. The isolate was identified as Pseudomonas aeruginosa using its 16S rDNA sequence. This strain was found to grow on monochlorobiphenyls (CBs), including 2-chlorobenzoic acid and 4-chlorobenzoic acid. It was also found to grow on 2,4-, 2,5-, 2,2′-, and 4,4′-diCB, as well as on a wide range of other xenobiotic compounds. This is the first reported representative of the genus Pseudomonas that is capable of growing on 2,4,4′-triCB, 2,2′,5,5′-tetraCB and 2,2′,4,4′,5,5′-hexaCB as sole carbon sources. Washed benzoate-grown cells were able to degrade 89% and 56% of 2,4-diCB and 2,2′,4,4′,5,5′-hexaCB, respectively. Gas chromatography analysis of individual congeners in Aroclor 1242 (200 ppm) following a 4-day incubation showed 73.3% degradation of PCBs without the need for biphenyl as an inducer. The strain exhibited no noticeable specificity for the percentage of congener transformation or degree of chlorination.     

Influence of chroline substitution pattern on the degradation of polychlorinated biphenyls by eight bacterial strains

We compared the metabolism of eight di- and trichlorobiphenyls by eight bacterial strains chosen to represent a broad range of degradative activity against polychlorinated biphenyls (PCBs). The PCB congeners used were 2,3-, 2,3′-, 2,4′-, 3,3′-, 2,3,3′-, 2,4,4′-, 2,5,3′-, and 3,4,2′-chlorobiphenyl. The bacterial strains used wereCorynebacterium sp. MB1,Alcaligenes strainsA. eutrophus H850 andA. faecalis Pi434, andPseudomonas strains LB400 and H1130,P. testosteroni H430 and H336, andP. cepacia H201. The results indicated that both the relative rates of primary degradation of PCBs and the choice of the ring attacked were dependent on the bacterial strain used. The bacterial strains exhibited considerable differences in their relative reactivity preferences for attack on mono- and dichlorophenyl groups and in the degree to which the attack was affected by the chlorine substitution pattern on the nonreacting ring. For MB1 the reactivity pattern was 3-≥4-≫2-chlorophenyl with no attack on 2,4- or 2,5-chlorophenyl groups. This strain was relatively insensitive to the chlorine substitution pattern on the nonreacting ring. Strains H1130, H430, H201, and Pi434 exhibited the same reactivity preferences as MB1, but for these strains (and for all others tested) the chlorination pattern on the nonreacting ring had a strong effect. For strain H336 the reactivity preference was 4-≥2->2,4-≥3-chlorophenyl, with no evidence of attack on 2,5-chlorophenyl rings. For strains H850 and LB400 the relative reactivity was 2->2,5->3-≫2,4->4-chlorophenyl. On this basis we propose that the eight bacterial strains represent four distinct classes of biphenyl/PCB-dioxygenase activity. The types of products formed were largely strain-independent and were determined primarily by the chlorine substitution pattern on the reacting ring. When the reacting ring was an unsubstituted phenyl or a 2-chlorophenyl group, the products were chlorobenzoic acids in high yields; for a 3-chlorophenyl ring, both chlorobenzoic acids and chloroacetophenones in moderate yields; and for a 4- or 2,4-chlorophenyl group, chlorobenzoic acids in low yields with an apparent accumulation ofmeta ring-fission product. Strains H850 and LB400 were able to degrade the 3-chlorobenzoic acid that they produced from the degradation of 2,3′-chlorobiphenyl. We conclude that despite differences among strains in the specificity of the initial dioxygenase, the specificities of the enzymes responsible for the subsequent degradation to chlorobenzoic acid and/or chloroacetophenone are quite similar for all strains.     

Effects of the dioxin-like PCB 126 on larval summer flounder (Paralichthys dentatus)

Little is known about the sensitivity of teleost post-embryonic developmental stages (larval and metamorphic) to dioxin-like compounds. Larval and metamorphosing summer flounder (Paralichthys dentatus) were exposed to the dioxin-like polychlorinated biphenyl congener PCB 126, to compare their sensitivity to other fish species early life stages, and to document effects on metamorphic development, including degree of eye migration and gastric maturation. Median lethal doses (LD 50 s) ranged between 30 and 220 ng/g wet mass, indicating that pre- and early-metamorphic stages of summer flounder are equally sensitive to the embryos of some of the most vulnerable fish species tested. Consistent with the presence of a functional aryl hydrocarbon receptor pathway, dose-dependent induction of cytochrome P-4501A (CYP1A) at four days post-exposure was observed in liver, stomach, intestine, and kidney of metamorphosing larvae. Stage-dependent differences in the epithelial distribution of CYP1A immunoreactivity were observed in the developing stomach of fish exposed to relatively high PCB 126 doses. A single sublethal dose (15 ng/g) delayed metamorphic progress (determined by the degree of eye migration), and resulted in abnormally high levels of cell proliferation and abnormal gastric gland morphology in late metamorphic stages. These results suggest that the post-embryonic larval and metamorphic stages of summer flounder, and potentially other fish species with complex life histories, are vulnerable to the effects of dioxin-like compounds, including lethality, developmental delay, and malformations.     

Purification, characterisation, and inhibition by monoterpenes of acetylcholinesterase from the waxmoth, Galleria mellonella (L.)

Acetylcholinesterase (AChE) was purified from the brain of the waxmoth, Galleria mellonella (L.) by affinity chromatography followed by anion exchange chromatography. It resolved as a single band by polyacrylamide gel electrophoresis (PAGE) both non-denaturing and SDS (silver stained), and as a single peak by high pressure liquid chromatography (HPLC), in an overall yield of 32% representing 283-fold purification. This was a true acetylcholinesterase, with no activity as a non-specific cholinesterase (butyrylcholinesterase). The molecular weight determined by PAGE in the absence and presence of sodium dodecyl sulphate (SDS) was ca. 240,000 Da and 60,000 Da respectively, indicating an arrangement of tetrameric subunits. 2-Heptanone, the honeybee alarm pheromone, reversibly and competitively inhibited the purified Galleria AChE with a K_i value of 1.34×10⁻³ M. Furthermore, five monoterpenes associated with plant defence and representing a range of functional groups, also were reversible competitive inhibitors of the purified AChE from Galleria, which is consistent with previous data for electric eel AChE (Ryan, M.F., Byrne, O., 1988. Plant-insect coevolution and inhibition of acetylcholinesterase. Journal of Chemical Ecology 14, 1965-1975).     

Degradation of polychlorinated biphenyls (PCBs) by four bacterial isolates obtained from the PCB-contaminated soil and PCB-contaminated sediment

We investigated the PCB-degrading abilities of four bacterial strains isolated from long-term PCB-contaminated soil (Alcaligenes xylosoxidans and Pseudomonas stutzeri) and sediments (Ochrobactrum anthropi and Pseudomonas veronii) that were co-metabolically grown on glucose plus biphenyl which is an inducer of the PCB catabolic pathway. The aim of study was to determine the respective contribution of biomass increase and expression of degrading enzymes on the PCB degrading abilities of each isolate. Growth on 5 g l⁻¹ glucose alone resulted in the highest stimulation of the growth of bacterial strains, whereas grown on 10 mg l⁻¹, 100 mg l⁻¹, 1 g l⁻¹, or 5 g l⁻¹ biphenyl did not effected the bacterial growth. None of the strains used in this study was able to grow on PCBs as the sole carbon source. Cells grown on glucose exhibited enhanced degradation ability due to an increased biomass. Addition of biphenyl at concentrations of 1 or 5 g l⁻¹ did not increase total PCB degradation, but stimulated the degradation of highly chlorinated congeners for some of the strains. The degradation of di- and tri-chlorobiphenyls was significantly lower for cells grown on 5 g l⁻¹ biphenyl independently on glucose addition. The highest degradation of the PCBs was obtained for A. xylosoxidans grown in the presence of glucose. Thus A. xylosoxidans appears to be the most promising among the four bacterial isolates for the purpose of bioremediation.     

Effect of lignocellulose degradation products on microbial biomass and lipid production by the oleaginous yeast Cryptococcus curvatus

This work investigated effects of lignocellulose degradation products on cell biomass and lipid production by Cryptococcus curvatus. Furfural was found to have the strongest inhibitory effect. For the three phenolic compounds tested, vanillin was the most toxic, while PHB and syringaldehyde showed comparable inhibitions in the concentration range of 0–1.0 g/L. Generally little significant differences on the relative cell biomass and lipid contents at the same concentrations of tested compounds were observed between glucose and xylose as a sole carbon source. At 1.0 g/L of furfural, the cell biomass and lipid content decreased by 78.4% and 61.0% for glucose as well as 72.0% and 59.3% for xylose, respectively. C. curvatus ceased to grow at concentrations of PHB over 1.0 g/L or vanillin over 1.5 g/L. The strain could survive in the presence of syringaldehyde up to 2.0 g/L for glucose or 1.5 g/L for xylose. The compounds’ negative impact was reduced by an increase in inoculum size and a 10% (v/v) seed was detected to be optimal for cell biomass and lipid production. The results demonstrated C. curvatus could effectively utilize most of the dominant monosaccharides and cellobiose existing in lignocellulosic biomass hydrolysate in the presence of toxic compounds.     

Biocatalytic synthesis of monocyclic arene-dihydrodiols and -diols by Escherichia coli cells expressing hybrid toluene/biphenyl dioxygenase and dihydrodiol dehydrogenase genes

The hybrid toluene/biphenyl dioxygenase, which is encoded by the todC1 gene of Pseudomonas putida F1 and the bphA2A3A4 genes of Pseudomonas pseudoalcaligenes KF707, has substrate ranges wider than toluene dioxygenase endoced by the todC1C2BA genes of P. putida F1. We carried out growing cell reactions by Escherichia coli expressing the todC1-bphA2A3A4 genes for the comprehensive production of monocyclic arene-dihydrodiols. As a result, we successfully biotranformed acetophenone-related compounds (acetophenone, propiophenone, and butyrophenone) to the corresponding cis-dihydrodiols. Furthermore, we performed the bioconversion experiments by E. coli cells expressing the bphB (dihydrodiol dehydrogenase) gene in addition to todC1-bphA2A3A4 to produce a series of monocyclic arene-diols. Consequently, toluene, benzene, stylene, p-xylene, acetophenone, propiophenone, butyrophenone, and trifluoroacetophenone were converted to the corresponding vicinal diols. The antioxidative activity of these generated diol compounds was markedly higher than that of the substrate used.     

Influence of diet on synthesis and utilisation of lipids for reproduction by the tsetse fly Glossina morsitans

Fatty acid composition of lipids from adult Glossina morsitans was unaffected by an in vitro fed diet of cow blood which induces production of under-sized offspring compared to that of flies fed on a superior diet of pig blood. The commonest fatty acids were C_16:0, C_16:1 and C_18:1 and only small differences in proportions were detected between virgin and pregnant females. Rate of lipid accumulation by males and females was the same and was unaffected by diet, but males achieved a maximum of 2.5 mg on day 9 while both virgin and fertilised females reached a maximum of 5,0 mg on day 14 of adult life. Lipid content of pregnant flies then fell to 3.0 mg on the day of larviposition and accumulation began again. A cow blood diet reduced the extent to which the lipids were utilised for larval growth and this was reflected in an altered secretory activity cycle in the female uterine gland. However, no effect on the growth of adult fat body was detectable in such flies. Mating and fertilisation, which influence reproductive events through activity of the endrocine system do not seem to affect the acquisition of lipid reserves by female Glossina. However, they clearly exert considerable influence over distribution of such reserves between fat body and uterine gland, which distribution is also affected by diet.     

Efficiency of defined strains and of soil consortia in the biodegradation of polycyclic aromatic hydrocarbon (PAH) mixtures

The microbiological characteristics of the bacterialdegradation of mixtures of five polycyclic aromatichydrocarbons (PAH), phenanthrene, fluorene,anthracene, fluoranthene and pyrene, wereinvestigated. Three pure bacterial strains using oneor several of these PAH as carbon sources wereselected. The interactions between PAH during thedegradation of PAH pairs by each of these strains werestudied and their effects on the kinetics and thebalance of degradation were characterised. Competitionbetween PAH and degradation by cometabolism werefrequently observed. Mixed cultures of two or threestrains, although possessing the global capacity tomineralise the set of five PAH, achieved limiteddegradation of the mixture. In contrast, a consortiumfrom a PAH-contaminated soil readily mineralised thefive-PAH mixture. The results suggested that soilconsortia possessed a wider variety of strains capableto compensate for the competitive inhibition betweenPAH as well as specialised strains that mineralisedpotentially inhibitory PAH metabolites produced bycometabolism.     

Influence of citric acid amendments on the availability of weathered PCBs to plant and earthworm species

A series of small and large pot trials were conducted to assess the phytoextraction potential of several plant species for weathered polychlorinated biphenyls (PCBs) in soil (105 microg/g Arochlor 1268). In addition, the effect of citric acid on PCB bioavailability to both plants and earthworms was assessed. Under small pot conditions (one plant, 400 g soil), three cucurbits (Cucurbita pepo ssp pepo [zucchini] and ssp ovifera [nonzucchini summer squash], Cucumis sativus, cucumber) accumulated up to 270 microg PCB/g in the roots and 14 microg/g in the stems, resulting in 0.10% contaminant removal from soil. Periodic 1 mM subsurface amendments of citric acid increased the stem and leaf PCB concentration by 330 and 600%, respectively, and resulted in up to a 65% increase in the total amount of contaminant removed from soil. Although citric acid at 10 mM more than doubled the amount of PCB desorbed in abiotic batch slurries, contaminant accumulation by two earthworm species (Eisenia foetida and Lumbricus terrestris) was unaffected by citric acid at 1 and 10 mM and ranged from 11-15 microg/g. Two large pot trials were conducted in which cucurbits (C. pepo ssp pepo and ssp ovifera, C. sativus) and white lupin (Lupinus albus) were grown in 70 kg of PCB-contaminated soil White lupin was the poorest accumulator of PCBs, with approximately 20 microg/g in the roots and 1 microg/g in the stems. Both C. pepo ssp ovifera (summer squash) and C. sativus (cucumber) accumulated approximately 65-100 microg/g in the roots and 6-10 microg/g in the stems. C. pepo ssp pepo (zucchini) accumulated significantly greater levels of PCB than all other species, with 430 microg/g in the roots and 22 microg/g in the stems. The mechanism by which C. pepo spp pepo extracts and translocates weathered PCBs is unknown, but confirms earlier findings on the phytoextraction of other weathered persistent organic pollutants such as chlordane, p,p'-DDE, and polycyclic aromatic hydrocarbons.     

Effect of light regimes on the utilisation of an exogenous carbon source by freshwater biofilm bacterial communities

This study reports the novel use of nucleic acid stable isotope probing (NA-SIP) to identify metabolically active ([¹³C]-acetate assimilating) bacteria in freshwater biofilms. Currently, a little is known of the factors affecting the structure and activity of these complex microbial biofilm communities, although it is likely that they are influenced by riparian vegetation through attenuation of light and alteration of allochthonous inputs of carbon. NA-SIP was used to investigate the effect of varying light regimes on [¹³C]-acetate assimilating bacteria within laboratory biofilm microcosms. Differences in clone libraries of 16S rRNA and rRNA genes from ¹³C-labelled and unlabelled nucleic acids indicated differential uptake of acetate and the rapid transfer of ¹³C to organisms at a higher trophic level. Biofilm communities incubated in the dark changed least over time and retained a significant fraction of phototrophic organisms. Incubation under elevated light caused the greatest change in bacterial community structure. Contrary to expectation, a complete loss of chlorophyll containing organisms occurred within this treatment, challenging current thinking that elevated light promotes communities dominated by photoautotrophs in nutrient enriched environments.     

The inoculation method affects colonization and performance of bacterial inoculant strains in the phytoremediation of soil contaminated with diesel oil

Plants in combination with microorganisms can remediate soils, which are contaminated with organic pollutants such as petroleum hydrocarbons. Inoculation of plants with degrading bacteria is one approach to improve remediation processes, but is often not successful due to the competition with resident microorganisms. It is therefore of high importance to address the persistence and colonization behavior of inoculant strains. The objective of this study was to determine whether the inoculation method (seed imbibement and soil inoculation) influences bacterial colonization, plant growth promotion and hydrocarbon degradation. Italian ryegrass was grown in non-sterilized soil polluted with diesel and inoculated with different alkane-degrading strains Pantoea sp. ITSI10, Pantoea sp. BTRH79 and Pseudomonas sp. MixRI75 individually as well as in combination. Inoculation generally had a beneficial effect on plant biomass production and hydrocarbon degradation, however, strains inoculated in soil performed better than applied by seed imbibement. Performance correlated with the colonization efficiency of the inoculated strains. The highest hydrocarbon degradation was observed in the treatment, in which all three strains were inoculated in combination into soil. Our study revealed that besides the degradation potential and competitive ability of inoculant strains the inoculation method plays an important role in determining the success of microbial inoculation.     

Influence of PCBs on the predator-prey relation between bacteria and protozoa in soil

Abstract This work deals with the impact of a possible accidental pollutant, pyralene (Prodelec, France; PCBs in trichlorobenzene), intoduced into the soil. Its influence on the predator-prey relation between bacteria and amoebae was studied by comparing the population dynamics of (i) an inoculated bacterial population ( A. lipoferum ) chosen as a biological tracer, (ii) the indigenous bacterial microflora, (iii) the infigenous amoebae. In the absence of pyralene the inoculated bacterial population decreased from 10⁷ to 10⁴ bacteria g⁻¹ soil (dw), grazed by the infigenous amoebae whose numbers increased 3-fold. In contrast, in presence of 2500 ppm of pyralene the introduced bacteria survived at a higher level (3·10⁶ bacteria g⁻¹ soil (dw)) while the number of amoebae diminished slightly. No predation occurred with PCB contamination. The indigenous bacterial microflora was not affected quantitatively by pyralene. In pure liquid culture with 500 ppm of pyralene added, bacterial growth was inhibited and an amoebal strain isolated from an inoculated uncontaminated soil was killed. We conclude that the active form of the amoebae were killed, and encystement was inhibited by pyralene in the soil. Hence the protozoa were unable to regulate the introduced A. lipoferum strain as they did in the absence of the pollutant.     

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

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