Selection of specific endophytic bacterial genotypes by plants in response to soil contamination

Plant-bacterial combinations can increase contaminant degradation in the rhizosphere, but the role played by indigenous root-associated bacteria during plant growth in contaminated soils is unclear. The purpose of this study was to determine if plants had the ability to selectively enhance the prevalence of endophytes containing pollutant catabolic genes in unrelated environments contaminated with different pollutants. At petroleum hydrocarbon contaminated sites, two genes encoding hydrocarbon degradation, alkane monooxygenase (alkB) and naphthalene dioxygenase (ndoB), were two and four times more prevalent in bacteria extracted from the root interior (endophytic) than from the bulk soil and sediment, respectively. In field sites contaminated with nitroaromatics, two genes encoding nitrotoluene degradation, 2-nitrotoluene reductase (ntdAa) and nitrotoluene monooxygenase (ntnM), were 7 to 14 times more prevalent in endophytic bacteria. The addition of petroleum to sediment doubled the prevalence of ndoB-positive endophytes in Scirpus pungens, indicating that the numbers of endophytes containing catabolic genotypes were dependent on the presence and concentration of contaminants. Similarly, the numbers of alkB- or ndoB-positive endophytes in Festuca arundinacea were correlated with the concentration of creosote in the soil but not with the numbers of alkB- or ndoB-positive bacteria in the bulk soil. Our results indicate that the enrichment of catabolic genotypes in the root interior is both plant and contaminant dependent.     

Changes in Microbial Community Composition and Function during a Polyaromatic Hydrocarbon Phytoremediation Field Trial

The purpose of this study was to investigate the mechanism by which phytoremediation systems promote hydrocarbon degradation in soil. The composition and degradation capacity of the bulk soil microbial community during the phytoremediation of soil contaminated with aged hydrocarbons was assessed. In the bulk soil, the level of catabolic genes involved in hydrocarbon degradation (ndoB, alkB, and xylE) as well as the mineralization of hexadecane and phenanthrene was higher in planted treatment cells than in treatment cells with no plants. There was no detectable shift in the 16S ribosomal DNA (rDNA) composition of the bulk soil community between treatments, but there were plant-specific and -selective effects on specific catabolic gene prevalence. Tall Fescue (Festuca arundinacea) increased the prevalence of ndoB, alkB, and xylE as well as naphthalene mineralization in rhizosphere soil compared to that in bulk soil. In contrast, Rose Clover (Trifolium hirtum) decreased catabolic gene prevalence and naphthalene mineralization in rhizosphere soil. The results demonstrated that phytoremediation systems increase the catabolic potential of rhizosphere soil by altering the functional composition of the microbial community. This change in composition was not detectable by 16S rDNA but was linked to specific functional genotypes with relevance to petroleum hydrocarbon degradation.     

Successful phytoremediation of crude-oil contaminated soil at an oil exploration and production company by plants-bacterial synergism

Phytoremediation is a promising approach for the cleanup of soil contaminated with petroleum hydrocarbons. This study aimed to develop plant-bacterial synergism for the successful remediation of crude oil-contaminated soil. A consortia of three endophytic bacteria was augmented to two grasses, Leptochloa fusca and Brachiaria mutica, grown in oil-contaminated soil (46.8 g oil kg^?1 soil) in the vicinity of an oil exploration and production company. Endophytes augmentation improved plant growth, crude oil degradation, and soil health. Maximum oil degradation (80%) was achieved with B. mutica plants augmented with the endophytes and it was significantly (P < 0.05) higher than the use of plants or bacteria individually. Moreover, endophytes showed more persistence, the abundance and expression of alkB gene in the rhizosphere as well as in the endosphere of the tested plants than in unvegetated soil. A positive relationship (r = 0.70) observed between gene expression and crude oil reduction indicates that catabolic gene expression is important for hydrocarbon mineralization. This investigation showed that the use of endophytes with appropriate plant is an effective strategy for the cleanup of oil-contaminated soil under field conditions.     

Characterization of Hydrocarbon-Degrading Microbial Populations in Contaminated and Pristine Alpine Soils

Biodegradation of petroleum hydrocarbons in cold environments, including Alpine soils, is a result of indigenous cold-adapted microorganisms able to degrade these contaminants. In the present study, the prevalence of seven genotypes involved in the degradation of n-alkanes (Pseudomonas putida GPo1 alkB; Acinetobacter spp. alkM; Rhodococcus spp. alkB1, and Rhodococcus spp. alkB2), aromatic hydrocarbons (P. putida xylE), and polycyclic aromatic hydrocarbons (P. putida ndoB and Mycobacterium sp. strain PYR-1 nidA) was determined in 12 oil-contaminated (428 to 30,644 mg of total petroleum hydrocarbons [TPH]/kg of soil) and 8 pristine Alpine soils from Tyrol (Austria) by PCR hybridization analyses of total soil community DNA, using oligonucleotide primers and DNA probes specific for each genotype. The soils investigated were also analyzed for various physical, chemical, and microbiological parameters, and statistical correlations between all parameters were determined. Genotypes containing genes from gram-negative bacteria (P. putida alkB, xylE, and ndoB and Acinetobacter alkM) were detected to a significantly higher percentage in the contaminated (50 to 75%) than in the pristine (0 to 12.5%) soils, indicating that these organisms had been enriched in soils following contamination. There was a highly significant positive correlation (P < 0.001) between the level of contamination and the number of genotypes containing genes from P. putida and Acinetobacter sp. but no significant correlation between the TPH content and the number of genotypes containing genes from gram-positive bacteria (Rhodococcus alkB1 and alkB2 and Mycobacterium nidA). These genotypes were detected at a high frequency in both contaminated (41.7 to 75%) and pristine (37.5 to 50%) soils, indicating that they are already present in substantial numbers before a contamination event. No correlation was found between the prevalence of hydrocarbon-degradative genotypes and biological activities (respiration, fluorescein diacetate hydrolysis, lipase activity) or numbers of culturable hydrocarbon-degrading soil microorganisms; there also was no correlation between the numbers of hydrocarbon degraders and the contamination level. The measured biological activities showed significant positive correlation with each other, with the organic matter content, and partially with the TPH content and a significant negative correlation with the soil dry-mass content (P < 0.05 to 0.001).     

Abundance and Diversity of <Emphasis Type="Italic">n</Emphasis>-Alkane-Degrading Bacteria in a Forest Soil Co-Contaminated with Hydrocarbons and Metals: A Molecular Study on <Emphasis Type="Italic">alkB</Emphasis> Homologous Genes

Unraveling functional genes related to biodegradation of organic compounds has profoundly improved our understanding of biological remediation processes, yet the ecology of such genes is only poorly understood. We used a culture-independent approach to assess the abundance and diversity of bacteria catalyzing the degradation of n-alkanes with a chain length between C₅ and C₁₆ at a forest site co-contaminated with mineral oil hydrocarbons and metals for nearly 60 years. The alkB gene coding for a rubredoxin-dependent alkane monooxygenase enzyme involved in the initial activation step of aerobic aliphatic hydrocarbon metabolism was used as biomarker. Within the area of study, four different zones were evaluated: one highly contaminated, two intermediately contaminated, and a noncontaminated zone. Contaminant concentrations, hydrocarbon profiles, and soil microbial respiration and biomass were studied. Abundance of n-alkane-degrading bacteria was quantified via real-time PCR of alkB, whereas genetic diversity was examined using molecular fingerprints (T-RFLP) and clone libraries. Along the contamination plume, hydrocarbon profiles and increased respiration rates suggested on-going natural attenuation at the site. Gene copy numbers of alkB were similar in contaminated and control areas. However, T-RFLP-based fingerprints suggested lower diversity and evenness of the n-alkane-degrading bacterial community in the highly contaminated zone compared to the other areas; both diversity and evenness were negatively correlated with metal and hydrocarbon concentrations. Phylogenetic analysis of alkB denoted a shift of the hydrocarbon-degrading bacterial community from Gram-positive bacteria in the control zone (most similar to Mycobacterium and Nocardia types) to Gram-negative genotypes in the contaminated zones (Acinetobacter and alkB sequences with little similarity to those of known bacteria). Our results underscore a qualitative rather than a quantitative response of hydrocarbon-degrading bacteria to the contamination at the molecular level.     

Identification of nitrogen-incorporating bacteria in petroleum-contaminated arctic soils by using [15N]DNA-based stable isotope probing and pyrosequencing

Arctic soils are increasingly susceptible to petroleum hydrocarbon contamination, as exploration and exploitation of the Arctic increase. Bioremediation in these soils is challenging due to logistical constraints and because soil temperatures only rise above 0°C for ∼2 months each year. Nitrogen is often added to contaminated soil in situ to stimulate the existing microbial community, but little is known about how the added nutrients are used by these microorganisms. Microbes vary widely in their ability to metabolize petroleum hydrocarbons, so the question becomes: which hydrocarbon-degrading microorganisms most effectively use this added nitrogen for growth? Using [¹⁵N]DNA-based stable isotope probing, we determined which taxonomic groups most readily incorporated nitrogen from the monoammonium phosphate added to contaminated and uncontaminated soil in Canadian Forces Station-Alert, Nunavut, Canada. Fractions from each sample were amplified with bacterial 16S rRNA and alkane monooxygenase B (alkB) gene-specific primers and then sequenced using lage-scale parallel-pyrosequencing. Sequence data was combined with 16S rRNA and alkB gene C quantitative PCR data to measure the presence of various phylogenetic groups in fractions at different buoyant densities. Several families of Proteobacteria and Actinobacteria that are directly involved in petroleum degradation incorporated the added nitrogen in contaminated soils, but it was the DNA of Sphingomonadaceae that was most enriched in ¹⁵N. Bacterial growth in uncontaminated soils was not stimulated by nutrient amendment. Our results suggest that nitrogen uptake efficiency differs between bacterial groups in contaminated soils. A better understanding of how groups of hydrocarbon-degraders contribute to the catabolism of petroleum will facilitate the design of more targeted bioremediation treatments.     

Modulation of microbial consortia enriched from different polluted environments during petroleum biodegradation

Environmental microbial communities are key players in the bioremediation of hydrocarbon pollutants. Here we assessed changes in bacterial abundance and diversity during the degradation of Tunisian Zarzatine oil by four indigenous bacterial consortia enriched from a petroleum station soil, a refinery reservoir soil, a harbor sediment and seawater. The four consortia were found to efficiently degrade up to 92.0% of total petroleum hydrocarbons after 2 months of incubation. Illumina 16S rRNA gene sequencing revealed that the consortia enriched from soil and sediments were dominated by species belonging to Pseudomonas and Acinetobacter genera, while in the seawater-derived consortia Dietzia, Fusobacterium and Mycoplana emerged as dominant genera. We identified a number of species whose relative abundances bloomed from small to high percentages: Dietzia daqingensis in the seawater microcosms, and three OTUs classified as Acinetobacter venetianus in all two soils and sediment derived microcosms. Functional analyses on degrading genes were conducted by comparing PCR results of the degrading genes alkB, ndoB, cat23, xylA and nidA1 with inferences obtained by PICRUSt analysis of 16S amplicon data: the two data sets were partly in agreement and suggest a relationship between the catabolic genes detected and the rate of biodegradation obtained. The work provides detailed insights about the modulation of bacterial communities involved in petroleum biodegradation and can provide useful information for in situ bioremediation of oil-related pollution.     

The Use of a Combination of alkB Primers to Better Characterize the Distribution of Alkane-Degrading Bacteria

The alkane monooxygenase AlkB, which is encoded by the alkB gene, is a key enzyme involved in bacterial alkane degradation. To study the alkB gene within bacterial communities, researchers need to be aware of the variations in alkB nucleotide sequences; a failure to consider the sequence variations results in the low representation of the diversity and richness of alkane-degrading bacteria. To minimize this shortcoming, the use of a combination of three alkB-targeting primers to enhance the detection of the alkB gene in previously isolated alkane-degrading bacteria was proposed. Using this approach, alkB-related PCR products were detected in 79% of the strains tested. Furthermore, the chosen set of primers was used to study alkB richness and diversity in different soils sampled in Carmópolis, Brazil and King George Island, Antarctica. The DNA extracted from the different soils was PCR amplified with each set of alkB-targeting primers, and clone libraries were constructed, sequenced and analyzed. A total of 255 alkB phylotypes were detected. Venn diagram analyses revealed that only low numbers of alkB phylotypes were shared among the different libraries derived from each primer pair. Therefore, the combination of three alkB-targeting primers enhanced the richness of alkB phylotypes detected in the different soils by 45% to 139%, when compared to the use of a single alkB-targeting primer. In addition, a dendrogram analysis and beta diversity comparison of the alkB composition showed that each of the sampling sites studied had a particular set of alkane-degrading bacteria. The use of a combination of alkB primers was an efficient strategy for enhancing the detection of the alkB gene in cultivable bacteria and for better characterizing the distribution of alkane-degrading bacteria in different soil environments.     

石油污染环境中固氮和寡氮营养细菌的分离鉴定及其特性

[背景]石油污染治理中的生物修复因无二次污染、处理成本低等优点受到人们的广泛关注,但由于石油烃向环境中大量输入,导致环境中氮源的相对不足成为制约生物修复效率的关键因素之一,因此筛选能够适应寡氮环境的微生物具有重要的生态意义.[目的]从辽河油田油藏水中筛选在不添加氮源培养基中生长的微生物,为石油污染环境生物修复提供候选菌...     

Biodegradation of Aliphatic and Aromatic Hydrocarbons by Nocardia sp. H17-1

We investigated the biodegradation of hydrocarbon components by Nocardia sp. H17-1 and the catabolic genes involved in the degradation pathways of both aliphatic and aromatic hydrocarbons. After 6 days of incubation, the aliphatic and aromatic fractions separated from Arabian light oil were degraded 99.0 ± 0.1% and 23.8 ± 0.8%, respectively. Detection of the catabolic genes involved in the hydrocarbon degradation indicated that H17-1 possessed the alkB genes for n-alkane biodegradation and catA gene for catechol 1,2-dioxygenase. However, H17-1 had neither the C23O gene for the degradation of aromatic hydrocarbons nor the catechol 2,3-dioxygenase activity. The investigation of the genes involved in the biodegradation of hydrocarbons supported the low degradation activity of H17-1 on the aromatic fractions.     

Microbial community analysis at crude oil‐contaminated soils targeting the 16S ribosomal RNA,xylM, C23O,and bcr genes

Aims: The analyses targeting multiple functional genes were performed on the samples of crude oil‐contaminated soil, to investigate community structures of organisms involved in monoaromatic hydrocarbon degradation. Methods and Results: Environmental samples were obtained from two sites that were contaminated with different components of crude oil. The analysis on 16S rRNA gene revealed that bacterial community structures were clearly different between the two sites. The cloning analyses were performed by using primers specific for the catabolic genes involved in the aerobic or anaerobic degradation of monoaromatic hydrocarbons, i.e. xylene monooxygenase (xylM), catechol 2,3‐dioxygenase (C23O), and benzoyl‐CoA reductase (bcr) genes. From the result of xylM gene, it was suggested that there are lineages specific to the respective sites, reflecting the differences of sampling sites. In the analysis of the C23O gene, the results obtained with two primer sets were distinct from each other. A comparison of these suggested that catabolic types of major bacteria carrying this gene were different between the two sites. As for the bcr gene, no amplicon was obtained from one sample. Phylogenetic analysis revealed that the sequences obtained from the other sample were distinct from the known sequences. Conclusions: The differences between the two sites were demonstrated in the analyses of all tested genes. As for aerobic cleavage of the aromatic ring, it was also suggested that analysis using two primer sets provide more detailed information about microbial communities in the contaminated site. Significance and Impact of the Study: The present study demonstrated that analysis targeting multiple functional genes as molecular markers is practical to examine microbial community in crude oil‐contaminated environments.     

Changes in microbial community composition and function during a polyaromatic hydrocarbon phytoremediation field trial

The purpose of this study was to investigate the mechanism by which phytoremediation systems promote hydrocarbon degradation in soil. The composition and degradation capacity of the bulk soil microbial community during the phytoremediation of soil contaminated with aged hydrocarbons was assessed. In the bulk soil, the level of catabolic genes involved in hydrocarbon degradation (ndoB, alkB, and xylE) as well as the mineralization of hexadecane and phenanthrene was higher in planted treatment cells than in treatment cells with no plants. There was no detectable shift in the 16S ribosomal DNA (rDNA) composition of the bulk soil community between treatments, but there were plant-specific and -selective effects on specific catabolic gene prevalence. Tall Fescue (Festuca arundinacea) increased the prevalence of ndoB, alkB, and xylE as well as naphthalene mineralization in rhizosphere soil compared to that in bulk soil. In contrast, Rose Clover (Trifolium hirtum) decreased catabolic gene prevalence and naphthalene mineralization in rhizosphere soil. The results demonstrated that phytoremediation systems increase the catabolic potential of rhizosphere soil by altering the functional composition of the microbial community. This change in composition was not detectable by 16S rDNA but was linked to specific functional genotypes with relevance to petroleum hydrocarbon degradation.     

Effects of diurnal temperature variation on microbial community and petroleum hydrocarbon biodegradation in contaminated soils from a sub‐Arctic site

Contaminated soils are subject to diurnal and seasonal temperature variations during on‐site ex‐situ bioremediation processes. We assessed how diurnal temperature variations similar to that in summer at the site from which petroleum hydrocarbon‐contaminated soil was collected affect the soil microbial community and the extent of biodegradation of petroleum hydrocarbons compared with constant temperature regimes. Microbial community analyses for 16S rRNA and alkB genes by pyrosequencing indicated that the microbial community for soils incubated under diurnal temperature variation from 5°C to 15°C (VART5‐15) evolved similarly to that for soils incubated at constant temperature of 15°C (CST15). In contrast, under a constant temperature of 5°C (CST5), the community evolved significantly different. The extent of biodegradation of C10–C16 hydrocarbons in the VART5‐15 systems was 48%, comparable with the 41% biodegradation in CST15 systems, but significantly higher than CST5 systems at 11%. The enrichment of Gammaproteobacteria was observed in the alkB gene‐harbouring communities in VART5‐15 and CST15 but not in CST5 systems. However, the Actinobacteria was abundant at all temperature regimes. The results suggest that changes in microbial community composition as a result of diurnal temperature variations can significantly influence petroleum hydrocarbon bioremediation performance in cold regions.     

Morphological,biochemical and molecular identification of petroleum hydrocarbons biodegradation bacteria isolated from oil polluted soil in Dhahran,Saud Arabia

Accumulation of petroleum hydrocarbon residual considered a major environmental problem in the kingdom of Saudi Arabia cause of intensive efforts for oil detecting. Until now, In situ biodegradation considered the most effective method for petroleum hydrocarbon residual biodegradation. The aim of this study is isolation and identification biodegradable capability bacteria from contaminated sites in Khurais oil field, Dhahran, Saud Arabia via Different morphological and biochemical and molecular methods. Furthermore, degradation level in contaminated liquid medium and soil were evaluated. Three bacterial strains were selected from petroleum-contaminated soils of Khurais oil field depending on their capacity to grow in the existence of hydrocarbon components and identified according to morphological, biochemical. Interestingly, 16S rDNA sequencing fingerprinting results confirmed our bacterial identification as Bacillus subtilis, Pseudomonas aeruginosa and Bacillus cereu. Phyllogenetic tree was constructed and genetic similarity was calculated according to alignments results. Biodegradation patterns for different three isolates were reflected varied degradation ability for three isolates regarding incubation time. Different features were studied for three biodegrading bacterial strains and identified as Bacillus subtilis, Pseudomonas aeruginosa and Bacillus cereus. Remarkable degradation rate % patterns for hydrocarbons residual were recorded for all three isolates with varied.     

Assessment of in-situ bioremediation at a refinery waste-contaminated site and an aviation gasoline contaminated site

A combination of geochemical, microbiological and isotopic methods were used to evaluate in-situ bioremediation of petroleum hydrocarbons at one site contaminated with refinery waste and a second site contaminated with aviation gasoline at Alameda Point, California. At each site, geochemical and microbiological characteristics from four locations in the contaminated zone were compared to those from two uncontaminated background locations. At both sites, the geochemical indicators of in-situbiodegradation includeddepleted soil gas and groundwater oxygen, elevated groundwater alkalinity, and elevated soil gas carbon dioxide and methane in the contaminated zone relative to the background. Radiocarbon content of methane and carbon dioxide measured in soil gas at both sites indicated that they were derived from hydrocarbon contaminant degradation. Direct microscopy of soil core samples using cell wall stains and activity stains, revealed elevated microbial numbers and enhanced microbial activities in contaminated areas relative to background areas, corroborating geochemical findings. While microbial plate counts and microcosm studies using soil core samples provided laboratory evidence for the presence of some microbial activity and contaminant degradation abilities, they did not correlate well with either contaminant location, geochemical, isotopic, or direct microscopy data.     

Novel Alkane Hydroxylase Gene (alkB) Diversity in Sediments Associated with Hydrocarbon Seeps in the Timor Sea,Australia

Hydrocarbon seeps provide inputs of petroleum hydrocarbons to widespread areas of the Timor Sea. Alkanes constitute the largest proportion of chemical components found in crude oils, and therefore genes involved in the biodegradation of these compounds may act as bioindicators for this ecosystem''s response to seepage. To assess alkane biodegradation potential, the diversity and distribution of alkane hydroxylase (alkB) genes in sediments of the Timor Sea were studied. Deduced AlkB protein sequences derived from clone libraries identified sequences only distantly related to previously identified AlkB sequences, suggesting that the Timor Sea maybe a rich reservoir for novel alkane hydroxylase enzymes. Most sequences clustered with AlkB sequences previously identified from marine Gammaproteobacteria though protein sequence identities averaged only 73% (with a range of 60% to 94% sequence identities). AlkB sequence diversity was lower in deep water (>400 m) samples off the continental slope than in shallow water (<100 m) samples on the continental shelf but not significantly different in response to levels of alkanes. Real-time PCR assays targeting Timor Sea alkB genes were designed and used to quantify alkB gene targets. No correlation was found between gene copy numbers and levels of hydrocarbons measured in sediments using sensitive gas chromatography-mass spectrometry techniques, probably due to the very low levels of hydrocarbons found in most sediment samples. Interestingly, however, copy numbers of alkB genes increased substantially in sediments exposed directly to active seepage even though only low or undetectable concentrations of hydrocarbons were measured in these sediments in complementary geochemical analyses due to efficient biodegradation.Alkanes are saturated hydrocarbons that are widespread in marine environments due to a variety of anthropogenic and natural sources. They constitute the major fraction of hydrocarbon components found in crude oils and refined petroleum and are also produced by various marine organisms (e.g., zooplankton) as cellular components (2, 44). Alkanes are considered as pollutants, with short-chained alkanes acting as solvents toward cellular membranes and other lipid components (34) while longer-chained alkanes may contribute to the formation of oil films and slicks that may limit nutrient and oxygen exchange (21). Importantly, alkanes also serve as important carbon and energy sources for some microorganisms. In marine environments, alkanes succumb to various removal and dispersal processes such as dissolution, photochemical oxidation, evaporation, adsorption, and sedimentation. However, the greatest removal pathway for alkanes in marine sediments is via biodegradation by bacteria (13). This mechanism also mediates the transfer of oil-derived carbon to higher trophic levels (28, 37), and therefore these bacteria have an important role in carbon cycling in environments subject to long-term inputs of hydrocarbons such as marine seep-associated ecosystems. Alkane biodegradation is mediated by a diverse range of marine bacteria using various electron acceptors although degradation generally proceeds at greater rates under aerobic conditions than under anaerobic conditions, where the process is relatively slow (8, 26).In the presence of oxygen, well-characterized alkane oxidation pathways are initiated by an activation step whereby oxygen is introduced to the alkane substrate before further catabolic steps can proceed. A number of oxygen-dependent alkane hydroxylase enzyme systems have been discovered that catalyze this initial step including the soluble di-iron methane monooxygenases and the membrane-bound copper-containing methane monooxygenases, both of which act upon short-chain alkanes (i.e., C₁ up to C₈). Integral membrane non-heme iron alkane hydroxylases (the alk system) that are related to the well-characterized AlkB of Pseudomonas putida GPo1 (also known as Pseudomonas oleovorans TF4-1 I) act upon longer-chain alkanes (i.e., C₅ to C₁₆) (40). Other systems exist that include alkane-hydroxylating cytochrome P450 enzymes in addition to other enzyme systems that are known to exist based purely on chemical analyses of metabolites formed during alkane degradation experiments (22, 25, 29); however, knowledge pertaining to the enzymes and genes involved as well as their importance in the environment is limited. Only recently have genes involved in the degradation of long-chain alkanes (e.g., C₃₂ and C₃₆) been identified in Acinetobacter sp. strain DSM 17874 (39) though there is no information about the presence or importance of such enzymes in the environment.Although various chemical and microbiological aspects of petroleum oil and alkane biodegradation in marine systems have been relatively well studied, there is a general lack of knowledge concerning the diversity or abundance of the functional genes involved. The biochemical and molecular aspects of alkB genes and the enzymes they encode have been relatively well studied, and this has enabled the development of molecular tools for the study of alkB genes in the environment (19). Elevated levels of hydrocarbons or the introduction of hydrocarbons to environments has been shown to increase gene copy numbers, indicating the potential use of alkB genes as bioindicators of oil pollution and/or biodegradation (16, 33, 36, 43). However, to date only one study has used culture-independent molecular methods to examine the diversity of alkB genes in a marine environment (20), and no studies have examined hydrocarbon-degrading genes where natural hydrocarbon seepage occurs.In this study, the diversity and relative abundance of alkB genes were examined in sediments of the Timor Sea, a region where natural seeps are sources of widespread petroleum hydrocarbons. It was hypothesized that (i) novel alkB genes may exist in this unique tropical marine environment, (ii) that variations in gene diversity would be found in sediments with different hydrocarbon levels, and (iii) that the abundance of certain alkB gene types may reflect the levels of measured hydrocarbons in sediments, and therefore this assay could be used as a complementary tool for monitoring petroleum inputs into sediments of the Timor Sea.     

Biodegradation of aged diesel in diverse soil matrixes: impact of environmental conditions and bioavailability on microbial remediation capacity

While bioremediation of total petroleum hydrocarbons (TPH) is in general a robust technique, heterogeneity in terms of contaminant and environmental characteristics can impact the extent of biodegradation. The current study investigates the implications of different soil matrix types (anthropogenic fill layer, peat, clay, and sand) and bioavailability on bioremediation of an aged diesel contamination from a heterogeneous site. In addition to an uncontaminated sample for each soil type, samples representing two levels of contamination (high and low) were also used; initial TPH concentrations varied between 1.6 and 26.6 g TPH/kg and bioavailability between 36 and 100 %. While significant biodegradation occurred during 100 days of incubation under biostimulating conditions (64.4–100 % remediation efficiency), low bioavailability restricted full biodegradation, yielding a residual TPH concentration. Respiration levels, as well as the abundance of alkB, encoding mono-oxygenases pivotal for hydrocarbon metabolism, were positively correlated with TPH degradation, demonstrating their usefulness as a proxy for hydrocarbon biodegradation. However, absolute respiration and alkB presence were dependent on soil matrix type, indicating the sensitivity of results to initial environmental conditions. Through investigating biodegradation potential across a heterogeneous site, this research illuminates the interplay between soil matrix type, bioavailability, and bioremediation and the implications of these parameters for the effectiveness of an in situ treatment.     

Functional gene diversity of soil microbial communities from five oil-contaminated fields in China

To compare microbial functional diversity in different oil-contaminated fields and to know the effects of oil contaminant and environmental factors, soil samples were taken from typical oil-contaminated fields located in five geographic regions of China. GeoChip, a high-throughput functional gene array, was used to evaluate the microbial functional genes involved in contaminant degradation and in other major biogeochemical/metabolic processes. Our results indicated that the overall microbial community structures were distinct in each oil-contaminated field, and samples were clustered by geographic locations. The organic contaminant degradation genes were most abundant in all samples and presented a similar pattern under oil contaminant stress among the five fields. In addition, alkane and aromatic hydrocarbon degradation genes such as monooxygenase and dioxygenase were detected in high abundance in the oil-contaminated fields. Canonical correspondence analysis indicated that the microbial functional patterns were highly correlated to the local environmental variables, such as oil contaminant concentration, nitrogen and phosphorus contents, salt and pH. Finally, a total of 59% of microbial community variation from GeoChip data can be explained by oil contamination, geographic location and soil geochemical parameters. This study provided insights into the in situ microbial functional structures in oil-contaminated fields and discerned the linkages between microbial communities and environmental variables, which is important to the application of bioremediation in oil-contaminated sites.