Monitoring of microbial diversity and activity during bioremediation of crude oil-contaminated soil with different treatments

The present study compared the microbial diversity and activity during the application of various bioremediation processes to crude oil-contaminated soil. Five different treatments, including natural attenuation (NA), biostimulation (BS), biosurfactant addition (BE), bioaugmentation (BA), and a combined treatment (CT) of biostimulation, biosurfactant addition, and bioaugmentation, were used to analyze the degradation rate and microbial communities. After 120 days, the level of remaining hydrocarbons after all the treatments was similar, however, the highest rate (k) of total petroleum hydrocarbon (TPH) degradatioN was observed with the CT treatment (P < 0.05). The total bacterial counts increased during the first 2 weeks with all the treatments, and then remained stable. The bacterial communities and alkane monooxygenase gene fragment, alkB, were compared by denaturing gradient gel electrophoresis (DGGE). The DGGE analyses of the BA and CT treatments, which included Nocardia sp. H17-1, revealed a simple dominant population structure, compared with the other treatments. The Shannon-Weaver diversity index (H') and Simpson dominance index (D), calculated from the DGGE profiles using 16S rDNA, showed considerable qualitative differences in the community structure before and after the bioremediation treatment as well as between treatment conditions.     

Evaluation of inoculum addition to stimulate in situ bioremediation of oily-sludge-contaminated soil

A full-scale study evaluating an inoculum addition to stimulate in situ bioremediation of oily-sludge-contaminated soil was conducted at an oil refinery where the indigenous population of hydrocarbon-degrading bacteria in the soil was very low (10(3) to 10(4) CFU/g of soil). A feasibility study was conducted prior to the full-scale bioremediation study. In this feasibility study, out of six treatments, the application of a bacterial consortium and nutrients resulted in maximum biodegradation of total petroleum hydrocarbon (TPH) in 120 days. Therefore, this treatment was selected for the full-scale study. In the full-scale study, plots A and B were treated with a bacterial consortium and nutrients, which resulted in 92.0 and 89.7% removal of TPH, respectively, in 1 year, compared to 14.0% removal of TPH in the control plot C. In plot A, the alkane fraction of TPH was reduced by 94.2%, the aromatic fraction of TPH was reduced by 91.9%, and NSO (nitrogen-, sulfur-, and oxygen-containing compound) and asphaltene fractions of TPH were reduced by 85.2% in 1 year. Similarly, in plot B the degradation of alkane, aromatic, and NSO plus asphaltene fractions of TPH was 95.1, 94.8, and 63.5%, respectively, in 345 days. However, in plot C, removal of alkane (17.3%), aromatic (12.9%), and NSO plus asphaltene (5.8%) fractions was much less. The population of introduced Acinetobacter baumannii strains in plots A and B was stable even after 1 year. Physical and chemical properties of the soil at the bioremediation site improved significantly in 1 year.     

Verification of Degradation of <Emphasis Type="Italic">n</Emphasis>-Alkanes in Diesel Oil by <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> Strain WatG in Soil Microcosms

Degradation of n-alkanes in diesel oil by Pseudomonas aeruginosa strain WatG (WatG) was verified in soil microcosms. The total petroleum hydrocarbon (TPH) degradation level in two bioaugmentation samples was 51% and 46% for 1 week in unsterilized and sterilized soil microcosms, respectively. The TPH degradation in the biostimulation was of control level (15%). The TPH degradation in aeration-limited samples was clearly reduced when compared with that in aeration-unlimited ones under both sterilized and unsterilized conditions. Addition of WatG into soil microcosms was accompanied by dirhamnolipid production only in the presence of diesel oil. These findings suggest that degradation of n-alkanes in diesel oil in soil microcosms would be facilitated by bioaugmentation of WatG, with production of dirhamnolipid, and also by participation of biostimulated indigenous soil bacteria.     

Breakdown of low-level total petroleum hydrocarbons (TPH) in contaminated soil using grasses and willows

A phytoremediation study targeting low-level total petroleum hydrocarbons (TPH) was conducted using cool- and warm-season grasses and willows (Salix species) grown in pots filled with contaminated sandy soil from the New Haven Rail Yard, CT. Efficiencies of the TPH degradation were assessed in a 90-day experiment using 20–8.7–16.6 N-P-K water-soluble fertilizer and fertilizer with molasses amendments to enhance phytoremediation. Plant biomass, TPH concentrations, and indigenous microbes quantified with colony-forming units (CFU), were assessed at the end of the study. Switchgrass grown with soil amendments produced the highest aboveground biomass. Bacterial CFU's were in orders of magnitude significantly higher in willows with soil amendments compared to vegetated treatments with no amendments. The greatest reduction in TPH occurred in all vegetated treatments with fertilizer (66–75%) and fertilizer/molasses (65–74%), followed sequentially by vegetated treatments without amendments, unvegetated treatments with amendments, and unvegetated treatments with no amendment. Phytoremediation of low-level TPH contamination was most efficient where fertilization was in combination with plant species. The same level of remediation was achievable through the addition of grasses and/or willow combinations without amendment, or by fertilization of sandy soil.     

Bacterial community dynamics and polycyclic aromatic hydrocarbon degradation during bioremediation of heavily creosote-contaminated soil

Bacterial community dynamics and biodegradation processes were examined in a highly creosote-contaminated soil undergoing a range of laboratory-based bioremediation treatments. The dynamics of the eubacterial community, the number of heterotrophs and polycyclic aromatic hydrocarbon (PAH) degraders, and the total petroleum hydrocarbon (TPH) and PAH concentrations were monitored during the bioremediation process. TPH and PAHs were significantly degraded in all treatments (72 to 79% and 83 to 87%, respectively), and the biodegradation values were higher when nutrients were not added, especially for benzo(a)anthracene and chrysene. The moisture content and aeration were determined to be the key factors associated with PAH bioremediation. Neither biosurfactant addition, bioaugmentation, nor ferric octate addition led to differences in PAH or TPH biodegradation compared to biodegradation with nutrient treatment. All treatments resulted in a high first-order degradation rate during the first 45 days, which was markedly reduced after 90 days. A sharp increase in the size of the heterotrophic and PAH-degrading microbial populations was observed, which coincided with the highest rates of TPH and PAH biodegradation. At the end of the incubation period, PAH degraders were more prevalent in samples to which nutrients had not been added. Denaturing gradient gel electrophoresis analysis and principal-component analysis confirmed that there was a remarkable shift in the composition of the bacterial community due to both the biodegradation process and the addition of nutrients. At early stages of biodegradation, the alpha-Proteobacteria group (genera Sphingomonas and Azospirillum) was the dominant group in all treatments. At later stages, the gamma-Proteobacteria group (genus Xanthomonas), the alpha-Proteobacteria group (genus Sphingomonas), and the Cytophaga-Flexibacter-Bacteroides group (Bacteroidetes) were the dominant groups in the nonnutrient treatment, while the gamma-Proteobacteria group (genus Xathomonas), the beta-Proteobacteria group (genera Alcaligenes and Achromobacter), and the alpha-Proteobacteria group (genus Sphingomonas) were the dominant groups in the nutrient treatment. This study shows that specific bacterial phylotypes are associated both with different phases of PAH degradation and with nutrient addition in a preadapted PAH-contaminated soil. Our findings also suggest that there are complex interactions between bacterial species and medium conditions that influence the biodegradation capacity of the microbial communities involved in bioremediation processes.     

Bioremediation of soil polluted with fuels by sequential multiple injection of native microorganisms: Field-scale processes in Poland

Research was conducted to estimate impact of the multiple bioaugmentation on the treatment of soil contaminated by fuels - diesel oil and aircraft fuel. The bacteria used to inoculate the remediation plots were isolated from the polluted soil and proliferated in field conditions. The amount of biomass applied to the polluted soil was set to ensure the total number of bacteria in soil 10⁷-10⁸ cfu/g d.w. The multiple inoculation of soil with indigenous bacteria active in diesel oil and engine oil (plot A) degradation increased bioremediation effectiveness by 50% in comparison to the non-inoculated control soil and by 30% in comparison to the soil that was inoculated only once. The multiple inoculation of soil with indigenous microorganisms was then applied in bioremediation of the soil polluted with double high concentration of diesel oil (soil B) and in bioremediation of the soil polluted with aircraft fuel (soil C). The process efficiency was 80% and 98% removal of TPH for soil B and C, respectively.     

Kinetic Modeling and Error Analysis for Decontamination of Different Petroleum Hydrocarbon Components in Biostimulation of Oily Soil Microcosm

A microcosm study was constructed to investigate the effect of complex co-substrate (corn steep liquor, CSL) addition on indigenous bacterial community, rate and extent of petroleum hydrocarbons (PH) degradation in an oily soil with total petroleum hydrocarbons (TPH) content of 63353 mg kg^?1. TPH degradation was found to be characterized by a rapid phase of degradation during the first three weeks where 76% removal of TPH occurred, followed by a slower degradation phase, where further 7% of the initial TPH was removed by the end of incubation period, 35 d. Branched alkanes are more resistant to microbial degradation than n-alkanes. Furthermore, the unresolved complex mixtures (UCM) of hydrocarbons are less degradable than n- and iso-alkanes. Pristane (Pr) was the most recalcitrant aliphatic compound studied in this work. These results in addition to the extensive bacterial growth observed (from 10⁷ to 10¹⁰ CFU g^?1 soil) give strong support that the addition of CSL resulted in increased degradation rates. The indigenous bacteria grew exponentially during the incubation period of 35 d with a growth rate of 0.26 d^?1. Kinetic modeling was performed to estimate the rates of biodegradation of each hydrocarbon type component in the studied system. Five different error functions were used in this study to evaluate the fitness of the model equation to the obtained experimental data. This showed that the degradation of ∑nC₂₀-nC_24, ∑nC₃₅-nC₄₂ and nC₁₈ can be better represented by a second order model, whereas the TPH, total resolvable peaks (TRP), nC₁₇, UCM, ∑nC₁₀-nC₁₄, ∑nC₁₅-nC_19, ∑nC₂₅-nC₂₉, ∑nC₃₀-nC_34, ∑nC_n, and ∑isoC_n and isoprenoids Pr and phytane (Ph) were similarly following the first order model.     

Monitoring of petroleum hydrocarbon degradative potential of indigenous microorganisms in ozonated soil

This study was performed to investigate the petroleum hydrocarbon (PH) degradative potential of indigenous microorganisms in ozonated soil to better develop combined pre-ozonation/bioremediation technology. Diesel-contaminated soils were ozonated for 0–900min. PH and microbial concentrations in the soils decreased with increased ozonation time. The greatest reduction of total PH (TPH, 47.6%) and aromatics (11.3%) was observed in 900-min ozonated soil. The number of total viable heterotrophic bacteria decreased by three orders of magnitude in the soil. Ozonated soils were incubated for 9weeks for bioremediation. The number of microorganisms in the soils increased during the incubation period, as monitored by culture- and nonculture-based methods. The soils showed additional PH-removal during incubation, supporting the presence of PH-degraders in the soils. The highest removal (25.4%) of TPH was observed during the incubation of 180-min ozonated soil during the incubation while a negligible removal was shown in 900-min ozonated soil. This negligible removal could be explained by the existence of relatively few or undetected PH-degraders in 900-min ozonated soil. After a 9-week incubation of the ozonated soils, 180-min ozonated soil showed the lowest TPH concentration, suggesting that appropriate ozonation and indigenous microorganisms survived ozonation could enhance remediation of PH-contaminated soil. Microbial community composition in 9-week incubated soils revealed a slight difference between 900-min ozonated and unozonated soils, as analyzed by whole cell hybridization. Taken together, this study provided insight into indigenous microbial potential to degrade PH in ozonated soils.     

Application of a Bayesian nonparametric model to derive toxicity estimates based on the response of Antarctic microbial communities to fuel‐contaminated soil

Ecotoxicology is primarily concerned with predicting the effects of toxic substances on the biological components of the ecosystem. In remote, high latitude environments such as Antarctica, where field work is logistically difficult and expensive, and where access to adequate numbers of soil invertebrates is limited and response times of biota are slow, appropriate modeling tools using microbial community responses can be valuable as an alternative to traditional single‐species toxicity tests. In this study, we apply a Bayesian nonparametric model to a soil microbial data set acquired across a hydrocarbon contamination gradient at the site of a fuel spill in Antarctica. We model community change in terms of OTUs (operational taxonomic units) in response to a range of total petroleum hydrocarbon (TPH) concentrations. The Shannon diversity of the microbial community, clustering of OTUs into groups with similar behavior with respect to TPH, and effective concentration values at level x, which represent the TPH concentration that causes x% change in the community, are presented. This model is broadly applicable to other complex data sets with similar data structure and inferential requirements on the response of communities to environmental parameters and stressors.     

Cultivation-dependent and -independent approaches for determining bacterial diversity in heavy-metal-contaminated soil

In recent years, culture-independent methods have been used in preference to traditional isolation techniques for microbial community analysis. However, it is questionable whether uncultured organisms from a given sample are important for determining the impact of anthropogenic stress on indigenous communities. To investigate this, soil samples were taken from a site with patchy metal contamination, and the bacterial community structure was assessed with a variety of approaches. There were small differences in microscopic epifluorescence bacterial counts. Denaturing gradient gel electrophoresis (DGGE) profiles of 16S rRNA gene fragments (16S-DGGE) amplified directly from soil samples were highly similar. A clone library generated from the most contaminated sample revealed a diverse bacterial community, which showed similarities to pristine soil communities from other studies. However, the proportion of bacteria from the soil samples that were culturable on standard plate-counting media varied between 0.08 and 2.2%, and these values correlated negatively with metal concentrations. The culturable communities from each sample were compared by 16S-DGGE of plate washes and by fatty acid profiling of individual isolates. Each approach indicated that there were considerable differences between the compositions of the culturable communities from each sample. DGGE bands from both culture-based and culture-independent approaches were sequenced and compared. These data indicated that metal contamination did not have a significant effect on the total genetic diversity present but affected physiological status, so that the number of bacteria capable of responding to laboratory culture and their taxonomic distribution were altered. Thus, it appears that plate counts may be a more appropriate method for determining the effect of heavy metals on soil bacteria than culture-independent approaches.     

Comparative study of rhizobacterial community structure of plant species in oil-contaminated soil

In this study, the identity and distribution of plants and the structure of their associated rhizobacterial communities were examined in an oil-contaminated site. The number of plant species that formed a community or were scattered was 24. The species living in soil highly contaminated with total petroleum hydrocarbon (TPH) (9,000-4,5000 mg/g-soil) were Cynodon dactylon, Persicaria lapathifolia, and Calystegia soldanella (a halophytic species). Among the 24 plant species, the following have been known to be effective for oil removal: C. dactylon, Digitaria sanguinalis, and Cyperus orthostachyus. Denaturing gradient gel electrophoresis (DGGE) profile analysis showed that the following pairs of plant species had highly similar (above 70%) rhizobacterial community structures: Artemisia princeps and Hemistepta lyrata; C. dactylon and P. lapathifolia; Carex kobomugi and Cardamine flexuosa; and Equisetum arvense and D. sanguinalis. The major groups of rhizobacteria were Betaproteobacteria, Gamma-proteobacteria, Chloroflexi, Actinobacteria, and unknown. Based on DGGE analysis, P. lapathifolia, found for the first time in this study growing in the presence of high TPH, may be a good species for phytoremediation of oil-contaminated soils and in particular, C. soldanella may be useful for soils with high TPH and salt concentrations. Overall, this study suggests that the plant roots, regardless of plant species, may have a similar influence on the bacterial community structure in oil-contaminated soil.     

The rapid degradation of bisphenol A induced by the response of indigenous bacterial communities in sediment

In the present study, sediment was spiked with bisphenol A (BPA) solution to explore the interaction between indigenous bacterial communities and BPA biodegradation in sediment. Results showed that BPA could be adsorbed to the sediment and then biodegraded rapidly. Biodegradation efficiency of BPA in treatments with 10 and 50 mg/L BPA reached 64.3 and 61.8% on the first day, respectively. Quantitative polymerase chain reaction and denaturing gradient gel electrophoresis analysis indicated that BPA affected the densities, species, and diversities of bacteria significantly. The response of bacterial community to BPA favored BPA biodegradation by promoting the growth of BPA-reducing bacteria and inhibiting other competitors. According to the results of sequencing, Pseudomonas and Sphingomonas played vital roles in the degradation of BPA. They presented over 73% of the original bacterial community, and both of them were promoted by BPA comparing with controls. Laccase and polyphenol oxidase contributed to the degradation of BPA and metabolic intermediates, respectively. This paper illustrates the rapid biodegradation of BPA induced by the response of indigenous bacterial communities to the BPA stress, which will improve the understandings of BPA degradation in sediment.

     

Drought‐resistant fungi control soil organic matter decomposition and its response to temperature

Microbial‐mediated decomposition of soil organic matter (SOM) ultimately makes a considerable contribution to soil respiration, which is typically the main source of CO₂ arising from terrestrial ecosystems. Despite this central role in the decomposition of SOM, few studies have been conducted on how climate change may affect the soil microbial community and, furthermore, on how possible climate‐change induced alterations in the ecology of microbial communities may affect soil CO₂ emissions. Here we present the results of a seasonal study on soil microbial community structure, SOM decomposition and its temperature sensitivity in two representative Mediterranean ecosystems where precipitation/throughfall exclusion has taken place during the last 10 years. Bacterial and fungal diversity was estimated using the terminal restriction fragment length polymorphism technique. Our results show that fungal diversity was less sensitive to seasonal changes in moisture, temperature and plant activity than bacterial diversity. On the other hand, fungal communities showed the ability to dynamically adapt throughout the seasons. Fungi also coped better with the 10 years of precipitation/throughfall exclusion compared with bacteria. The high resistance of fungal diversity to changes with respect to bacteria may open the controversy as to whether future ‘drier conditions’ for Mediterranean regions might favor fungal dominated microbial communities. Finally, our results indicate that the fungal community exerted a strong influence over the temporal and spatial variability of SOM decomposition and its sensitivity to temperature. The results, therefore, highlight the important role of fungi in the decomposition of terrestrial SOM, especially under the harsh environmental conditions of Mediterranean ecosystems, for which models predict even drier conditions in the future.     

Composition of soil microbial communities enriched on a mixture of aromatic hydrocarbons

Soil contaminated with C5+, which contained benzene (45%, wt/wt), dicyclopentadiene (DCPD) plus cyclopentadiene (together 20%), toluene (6%), styrene (3%), xylenes (2%), naphthalene (2%), and smaller quantities of other compounds, served as the source for isolation of 55 genomically distinct bacteria (standards). Use of benzene as a substrate by these bacteria was most widespread (31 of 44 standards tested), followed by toluene (23 of 44), xylenes (14 of 44), styrene (10 of 44), and naphthalene (10 of 44). Master filters containing denatured genomic DNAs of all 55 standards were used to analyze the community compositions of C5+ enrichment cultures by reverse sample genome probing (RSGP). The communities enriched from three contaminated soils were similar to those enriched from three uncontaminated soils from the same site. The compositions of these communities were time dependent and showed a succession of Pseudomonas and Rhodococcus spp. before convergence on a composition dominated by Alcaligenes spp. The dominant community members detected by RSGP were capable of benzene degradation at all stages of succession. The enrichments effectively degraded all C5+ components except DCPD. Overall, degradation of individual C5+ hydrocarbons followed first-order kinetics, with the highest rates of removal for benzene.     

Anthropogenic N deposition alters soil organic matter biochemistry and microbial communities on decaying fine roots

Fine root litter is a primary source of soil organic matter (SOM), which is a globally important pool of C that is responsive to climate change. We previously established that ~20 years of experimental nitrogen (N) deposition has slowed fine root decay and increased the storage of soil carbon (C; +18%) across a widespread northern hardwood forest ecosystem. However, the microbial mechanisms that have directly slowed fine root decay are unknown. Here, we show that experimental N deposition has decreased the relative abundance of Agaricales fungi (?31%) and increased that of partially ligninolytic Actinobacteria (+24%) on decaying fine roots. Moreover, experimental N deposition has increased the relative abundance of lignin‐derived compounds residing in SOM (+53%), and this biochemical response is significantly related to shifts in both fungal and bacterial community composition. Specifically, the accumulation of lignin‐derived compounds in SOM is negatively related to the relative abundance of ligninolytic Mycena and Kuehneromyces fungi, and positively related to Microbacteriaceae. Our findings suggest that by altering the composition of microbial communities on decaying fine roots such that their capacity for lignin degradation is reduced, experimental N deposition has slowed fine root litter decay, and increased the contribution of lignin‐derived compounds from fine roots to SOM. The microbial responses we observed may explain widespread findings that anthropogenic N deposition increases soil C storage in terrestrial ecosystems. More broadly, our findings directly link composition to function in soil microbial communities, and implicate compositional shifts in mediating biogeochemical processes of global significance.     

Bacterial succession in a petroleum land treatment unit

Bacterial community dynamics were investigated in a land treatment unit (LTU) established at a site contaminated with highly weathered petroleum hydrocarbons in the C(10) to C(32) range. The treatment plot, 3,000 cubic yards of soil, was supplemented with nutrients and monitored weekly for total petroleum hydrocarbons (TPH), soil water content, nutrient levels, and aerobic heterotrophic bacterial counts. Weekly soil samples were analyzed with 16S rRNA gene terminal restriction fragment (TRF) analysis to monitor bacterial community structure and dynamics during bioremediation. TPH degradation was rapid during the first 3 weeks and slowed for the remainder of the 24-week project. A sharp increase in plate counts was reported during the first 3 weeks, indicating an increase in biomass associated with petroleum degradation. Principal components analysis of TRF patterns revealed a series of sample clusters describing bacterial succession during the study. The largest shifts in bacterial community structure began as the TPH degradation rate slowed and the bacterial cell counts decreased. For the purpose of analyzing bacterial dynamics, phylotypes were generated by associating TRFs from three enzyme digests with 16S rRNA gene clones. Two phylotypes associated with Flavobacterium and Pseudomonas were dominant in TRF patterns from samples during rapid TPH degradation. After the TPH degradation rate slowed, four other phylotypes gained dominance in the community while Flavobacterium and Pseudomonas phylotypes decreased in abundance. These data suggest that specific phylotypes of bacteria were associated with the different phases of petroleum degradation in the LTU.     

Influence of soil pollution on the composition of a microbial community

The abundance dynamics and composition of indigenous soil microbial communities were studied in soils polluted with naphthalene, dioctyl phthalate, diesel fuel, and crude oil. DGGE analysis of the 16S rRNA genes amplified from the total soil DNA revealed that the bacterial community of uncontaminated soil was more diverse and included no dominant species. In the soil samples polluted with the crude oil, diesel fuel, or dioctyl phthalate, Pseudomonas became the dominant bacteria since the third day of the experiment. In the soil polluted with naphthalene, two genera of bacteria (Pseudomonas and Paenibacillus) were dominant in population on the third day of the experiment, while on the 21th day of the experiment Arthrobacter became dominant. During the experiment, the average number of indigenous bacterial degraders increased approximately by two orders of magnitude. While the key genes of naphthalene catabolism, nahAc and nahH, were not detected in the pristine soil, they were found in a significant amount on the third day after naphthalene addition. Three degrader strains harboring the plasmids of naphthalene biodegradation (IncP-9 group) were isolated on the third day from the soil polluted with naphthalene. Two of these plasmids, although isolated from various degraders, were shown to be identical.     

Effects of community-accessible biochar and compost on diesel-contaminated soil

Petroleum pollution is a global problem that requires effective and accessible remediation strategies that takes ecosystem functioning into serious consideration. Bioremediation can be an effective tool to address the challenge. In this study, we used a mesocosm experiment to evaluate the effects of locally sourced and community produced biochar and compost amendments on diesel-contaminated soil. At the end of the 90-day experiment, we quantified the effects of the amendments on total petroleum hydrocarbons (C9-C40) (TPH) and soil pH, organic matter, aggregate stability, soil respiration, extractable phosphorus, extractable potassium, and micronutrients (Mg, Fe, Mn, and Zn). We observed significantly higher TPH degradation in compost-amended soils than in controls and soils amended with biochar. We propose that the addition of compost improved TPH biodegradation by augmenting soil nutrient content and microbial activity. Our results suggest that community-accessible compost can improve TPH biodegradation, and that implementation is possible at the community level.     

Microbial communities in different soil types do not converge after diesel contamination

AIMS: To study the comparative effect of diesel addition and simulated bioremediation on the microbial community in three different soil types.METHODS AND RESULTS: Three different soils were amended with diesel and bioremediation treatment simulated by addition of nutrients. The progress of bioremediation, and the effect on the indigenous microbial communities, was monitored using microbiological techniques. These included basal respiration, sole carbon source utilization patterns using both a commercially-available substrate set and a set designed to highlight changes in hydrocarbon-utilizing bacteria, and phospholipid fatty acid (PLFA) profiling. The development of active hydrocarbon-degrading communities was indicated by the disappearance of diesel, increases in soil respiration and biomass, and large changes in the sole carbon source utilization patterns and PLFA profiles compared with control soils. However, comparison of the relative community structure of the three soils using PLFA profiling showed that there was no tendency for the community structure of the three different soil types to converge as a result of contamination. In fact, they became more dissimilar as a result. Changes in the sole carbon source utilization patterns using the commercially-available set of carbon sources indicated the same result as shown by PLFA profiling. The specially selected set of carbon sources yielded no additional information compared with the commercially-available set.CONCLUSIONS: Diesel contamination does not result in the development of similar community profiles in different soil types.SIGNIFICANCE AND IMPACT OF THE STUDY: The results suggest that different soils have different inherent microbial potential to degrade hydrocarbons, a finding that should be taken into account in impact and risk assessments. Following the development of the microbial community and its recovery is a useful and sensitive way of monitoring the impact and recovery of oil-contaminated soils.     

凉水保护区土壤产类漆酶-多铜氧化酶细菌群落结构

在凉水国家级自然保护区3种主要林型红松(Pinus koraiensis)、白桦(Betula platyphylla)及云杉(Picea dietrich)林采集林下土壤样品,以铜离子作为筛选剂处理后,结合平板分离法与基于16S rDNA V3区片段的变性梯度凝胶电泳(DenaturingGradient Gel Electrophoresis,DGGE)技术,调查了土壤样品中产类漆酶-多铜氧化酶(laccase-like multicopper oxidase,LMCO)细菌的群落结构。这是研究产类漆酶-多铜氧化酶细菌在环境中存在的种、属及分布的新尝试。平板分离获得10株细菌均为芽孢杆菌属(Bacillus sp.),其中梭状芽孢杆菌(Bacillus fusiformis)未见相关报道。通过DGGE图谱分析可知,产类漆酶-多铜氧化酶细菌在研究地不同林型土壤中的群落结构无明显差异,在红松林土壤中多样性最为丰富。DGGE条带测序结果表明,取样地土壤中产类漆酶细菌主要为罗尔斯顿菌属(Ralstonia sp.)、肠杆菌属(Enterobacter sp.)、芽孢杆菌属和一些未培养细菌。