Anaerobic Hydrocarbon Biodegradation Research Articles

Microbial activities in hydrocarbon-laden wastewaters: Impact on diesel fuel stability and the biocorrosion of carbon steel

Anaerobic hydrocarbon biodegradation not only diminishes fuel quality, but also exacerbates the biocorrosion of the metallic infrastructure. While successional events in marine microbial ecosystems impacted by petroleum are well documented, far less is known about the response of communities chronically exposed to hydrocarbons. Shipboard oily wastewater was used to assess the biotransformation of different diesel fuels and their propensity to impact carbon steel corrosion. When amended with sulfate and an F76 military diesel fuel, the sulfate removal rate in the assay mixtures was elevated (26.8μM/d) relative to incubations receiving a hydroprocessed biofuel (16.1μM/d) or a fuel-unamended control (17.8μM/d). Microbial community analysis revealed the predominance of Anaerolineae and Deltaproteobacteria in F76-amended incubations, in contrast to the Beta- and Gammaproteobacteria in the original wastewater. The dominant Smithella-like sequences suggested the potential for syntrophic hydrocarbon metabolism. The general corrosion rate was relatively low (0.83 − 1.29±0.12mpy) and independent of the particular fuel, but pitting corrosion was more pronounced in F76-amended incubations. Desulfovibrionaceae constituted 50–77% of the sessile organisms on carbon steel coupons. Thus, chronically exposed microflora in oily wastewater were differentially acclimated to the syntrophic metabolism of traditional hydrocarbons but tended to resist isoalkane-laden biofuels.

Anaerobic hydrocarbon biodegradation and biocorrosion of carbon steel in marine environments: The impact of different ultra low sulfur diesels and bioaugmentation

The anaerobic biodegradation of ultra low sulfur diesels (ULSDs) by marine microbial communities was examined, along with the relationship of this metabolism to carbon steel biocorrosion. Fuel analysis revealed that the ULSDs differed based on the ratio of low to high molecular weight (LMW/HMW) n-alkanes. Desulfoglaeba alkanexedens, a sulfate-reducing bacterium capable of degrading LMW n-alkanes, was used as a positive control and to explore the impact of bioaugmentation. Metagenomic analysis was conducted to determine the genetic potential for anaerobic biodegradation of a range of hydrocarbons. Initial sulfate reduction rates were faster in incubations amended with ULSDs containing a higher ratio of LMW/HMW n-alkanes, but total sulfate loss was similar for all fuels. Sulfate removal in bioaugmented incubations exceeded stoichiometric expectations calculated for the mineralization of all paraffins. In combination with metagenomic analysis, these data confirmed that other microorganisms utilized hydrocarbons beyond the range exhibited by D. alkanexedens. A positive correlation between coupon weight loss and sulfate loss was observed, and pitting corrosion was more extensive in non-sterile incubations compared to sterile controls. This study demonstrates that while ULSDs vary in their susceptibility to early stage biodegradation, compositional differences were less important for overall fuel deterioration and steel biocorrosion patterns. Further, bioaugmentation stimulated other hydrocarbonclastic marine microorganisms.

Sequential biodegradation of complex naphtha hydrocarbons under methanogenic conditions in two different oil sands tailings

Methane emissions in oil sands tailings ponds are sustained by anaerobic biodegradation of unrecovered hydrocarbons. Naphtha (primarily C6–C10; n- iso- and cycloalkanes) is commonly used as a solvent during bitumen extraction process and its residue escapes to tailings ponds during tailings deposition. To investigate biodegradability of hydrocarbons in naphtha, mature fine tailings (MFT) collected from Albian and CNRL tailings ponds were amended with CNRL naphtha at ∼0.2 wt% (∼2000 mg L−1) and incubated under methanogenic conditions for ∼1600 d. Microbial communities in both MFTs started metabolizing naphtha after a lag phase of ∼100 d. Complete biodegradation/biotransformation of all n-alkanes (except partial biodegradation of n-octane in CNRL MFT) followed by major iso-alkanes (2-methylpentane, 3-methylhexane, 2- and 4-methylheptane, iso-nonanes and 2-methylnonane) and a few cycloalkanes (derivatives of cyclopentane and cyclohexane) was observed during the incubation. 16S rRNA gene pyrosequencing showed dominance of Peptococcaceae and Anaerolineaceae in Albian MFT and Anaerolineaceae and Syntrophaceae in CNRL MFT bacterial communities with co-domination of Methanosaetaceae and “Candidatus Methanoregula” in archaeal populations during active biodegradation of hydrocarbons. The findings extend the known range of hydrocarbons susceptible to methanogenic biodegradation in petroleum-impacted anaerobic environments and help refine existing kinetic model to predict greenhouse gas emissions from tailings ponds.

Synthesis and characterization of anaerobic degradation biomarkers of n-alkanes via hydroxylation/carboxylation pathways.

Metabolite profiling is a powerful method in research on anaerobic biodegradation of hydrocarbons. Hydroxylation and carboxylation are proposed pathways in anaerobic degradation but very little direct evidence is available about metabolites and signature biomarkers. 2-Acetylalkanoic acid is a potential signature metabolite because of its unique and specific structure among possible intermediates. A procedure for the synthesis of four homologues with various carbon chain lengths was proposed and the characteristics of 2-acetyl- alkanoic acid esters were investigated using four derivatization processes, namely methyl, ethyl, n-butyl and trimethylsilyl esterification. Four intermediate fragments observed were at m/z 73 + 14n, 87 + 14n, 102 + 14n (n = 1, 2 and 4 for methyl, ethyl and n-butyl ester, respectively) and [M - 42]+ for three of the derivatization methods. For silylation, characteristic ions were observed at m/z 73, 117, [M - 42](+) and [M - 55](+). These are basic and significant data for the future identification of potential intermediates of the hydroxylation and carboxylation pathways in hydrocarbon degradation.

Exploring the capacity for anaerobic biodegradation of polycyclic aromatic hydrocarbons and naphthenic acids by microbes from oil-sands-process-affected waters

Both polycyclic aromatic hydrocarbons (PAHs) and naphthenic acids (NAs) are natural components of fossil fuels, but they are also widespread toxic and environmentally persistent pollutants. They are the major cause of environmental toxicity in oil-sands-process waters (OSPW). This study aimed to investigate the anaerobic biodegradation of the PAHs pyrene and 2-methylnaphthalene, and the NAs adamantane-1-carboxylic acid and a “natural” NA mixture (i.e., acid-extractable NAs from OSPW) under sulfate-reducing and methanogenic conditions by a microbial community derived from an oil sands tailings pond. Using gas-chromatography mass spectrometry (GC–MS), the rate of biodegradation was measured in relation to changes in bacterial community composition. Only 2-methylnaphthalene was significantly degraded after 260 days, with significantly more degradation under sulfate-reducing (40%) than methanogenic conditions (25%). During 2-methylnaphthalene biodegradation, a major metabolite was produced and tentatively identified as 2-naphthoic acid. Denaturing gradient gel electrophoresis (DGGE) demonstrated an increase in intensity of bands during the anaerobic biodegradation of 2-methylnaphalene, which derived from species of the genera Fusibacter, Alkaliphilus, Desulfobacterium, Variovorax, Thaurea, and Hydrogenophaga. Despite the biodegradation of 2-methylnaphthalene, this study demonstrates that, under anaerobic conditions, NAs and high-molecular-weight PAHs are the predominant molecules likely to persist in OSPW. Therefore alternative remediation strategies are required.

Biostimulation of petroleum-hydrocarbon-contaminated marine sediment with co-substrate: involved metabolic process and microbial community.

This study investigated the effect of acetate and methanol as co-substrates on anaerobic biodegradation of total petroleum hydrocarbons (TPHs, C10-C40) in marine sediment. The findings evidenced that the degradation of TPH can be enhanced by adding acetate or methanol. The addition of acetate was generally more favorable than the addition of methanol for the TPH degradation. Both sulfate reduction and methanogenesis occurred in the acetate-treated sediment. However, the depletion of SO4 (2-) inhibited sulfate reduction over the incubation period. Only methanogenesis was prevalent in the methanol-treated sediment within the whole incubation period. The degradation of TPH fractions with higher carbon number ranges (C31-C40) was speculated to be more favored under sulfate-reducing condition, while TPH fractions with lower carbon number ranges (C10-C20) were preferentially degraded under methanogenic condition. The 16S rRNA clone library-based analysis revealed that the addition of different co-substrates led to distinct structures of the microbial community. Clones related to sulfate-reducing Desulfobacterales were the most abundant in the sediment dosed with acetate. Clones related to Clostridiales predominated in the sediment dosed with methanol. Acetoclastic methanogens were found to be the predominant archaeal species in the sediment dosed with acetate, while both acetoclastic methanogens and hydrogenotrophic methanogens accounted for large proportions in the sediment dosed with methanol. The results obtained in this study will contribute to more comprehensive knowledge on the role of acetate and methanol as co-substrates in biostimulation of petroleum-hydrocarbon-contaminated marine sediment.

Arsenic Cycling in Hydrocarbon Plumes: Secondary Effects of Natural Attenuation.

Monitored natural attenuation is widely applied as a remediation strategy at hydrocarbon spill sites. Natural attenuation relies on biodegradation of hydrocarbons coupled with reduction of electron acceptors, including solid phase ferric iron (Fe(III)). Because arsenic (As) adsorbs to Fe-hydroxides, a potential secondary effect of natural attenuation of hydrocarbons coupled with Fe(III) reduction is a release of naturally occurring As to groundwater. At a crude-oil-contaminated aquifer near Bemidji, Minnesota, anaerobic biodegradation of hydrocarbons coupled to Fe(III) reduction has been well documented. We collected groundwater samples at the site annually from 2009 to 2013 to examine if As is released to groundwater and, if so, to document relationships between As and Fe inside and outside of the dissolved hydrocarbon plume. Arsenic concentrations in groundwater in the plume reached 230 µg/L, whereas groundwater outside the plume contained less than 5 µg/L As. Combined with previous data from the Bemidji site, our results suggest that (1) naturally occurring As is associated with Fe-hydroxides present in the glacially derived aquifer sediments; (2) introduction of hydrocarbons results in reduction of Fe-hydroxides, releasing As and Fe to groundwater; (3) at the leading edge of the plume, As and Fe are removed from groundwater and retained on sediments; and (4) downgradient from the plume, patterns of As and Fe in groundwater are similar to background. We develop a conceptual model of secondary As release due to natural attenuation of hydrocarbons that can be applied to other sites where an influx of biodegradable organic carbon promotes Fe(III) reduction.

Draft Genome Sequence of Uncultivated Desulfosporosinus sp. Strain Tol-M, Obtained by Stable Isotope Probing Using [13C6]Toluene.

A draft Desulfosporosinus genome was assembled from the metagenome of a methanogenic [13C6]toluene-degrading community. The Desulfosporosinus sp. strain Tol-M genome is distinguished from that of previously published Desulfosporosinus strain by containing bss, bbs, and bam genes encoding enzymes for anaerobic biodegradation of monoaromatic hydrocarbons and lacking dsrAB genes for dissimilatory sulfate reduction.

The effect of salinity, redox mediators and temperature on anaerobic biodegradation of petroleum hydrocarbons in microbial fuel cells

Microbial fuel cells (MFCs) need to be robust if they are to be applied in the field for bioremediation. This study investigated the effect of temperature (20–50°C), salinity (0.5–2.5% (w/v) as sodium chloride), the use of redox mediators (riboflavin and anthraquinone-2-sulphonate, AQS) and prolonged fed-batch operation (60 days) on biodegradation of a petroleum hydrocarbon mix (i.e. phenanthrene and benzene) in MFCs. The performance criteria were degradation efficiency, % COD removal and electrochemical performance. Good electrochemical and degradation performance were maintained up to a salinity of 1.5% (w/v) but deteriorated by 35-fold and 4-fold respectively as salinity was raised to 2.5%w/v. Degradation rates and maximum power density were both improved by approximately 2-fold at 40°C compared to MFC performance at 30°C but decreased sharply by 4-fold when operating temperature was raised to 50°C. The optimum reactor performance obtained at 40°C was 1.15mW/m2 maximum power density, 89.1% COD removal and a degradation efficiency of 97.10%; at moderately saline (1% w/v) conditions the maximum power density was 1.06mW/m2, 79.1% COD removal and 91.6% degradation efficiency. This work suggests the possible application of MFC technology in the effective treatment of petroleum hydrocarbons contaminated site and refinery effluents.

Synthesis of anaerobic degradation biomarkers alkyl-, aryl- and cycloalkylsuccinic acids and their mass spectral characteristics.

Anaerobic biodegradation of petroleum hydrocarbons has been reported to proceed predominantly via fumarate addition to yield substituted succinate metabolites. These metabolites, commonly regarded as signature biomarkers, are specific indicators of anaero- bic hydrocarbon degradation by microbial activity. To the best of our knowledge, mass spectrometry information for 2-(1-methylalkylj succinic acids, 2-arylsuccinic acids, 2-cycloalkylsuccinic acids and/or their derivatives is still incomplete, especially for the analysis of environmental samples. Here, a novel approach is proposed for the successful synthesis of five hydrocarbon-derived succinic acids. The characteristic fragments of 2-[1-methylalkyllsuccinic acid diesters were investigated by four derivatization processes (methyl, ethyl, n-butyl and trimethylsilyl esterification], some of which are not available in official Libraries. Under electron ionization mass spec- trometry conditions, informative fragments of various molecular masses have been obtained. Results confirmed characteristic differ- ences among the derivatization processes of the chemically synthesized compounds. In the case of 2-[cyclo)alkylsuccinate esters, four intermediate fragments were observed at m/z 114 + 14n, 118 + 28n, [M - [17 + 14n1]]+ and [M - (59 + 14n)]+ (n = 1, 2 and 4 for methyl, ethyl and n-butyl ester]. However, for silylation the abundant fragment ions are at m/z 262, 217, 172, 147, 73 and [M - 15]+. These data provide information for the identification of hydrocarbon-derived succinic acids as anaerobic biodegradation intermediates in hydrocarbons- rich environments.

Anaerobic hydrocarbon and fatty acid metabolism by syntrophic bacteria and their impact on carbon steel corrosion.

The microbial metabolism of hydrocarbons is increasingly associated with the corrosion of carbon steel in sulfate-rich marine waters. However, how such transformations influence metal biocorrosion in the absence of an electron acceptor is not fully recognized. We grew a marine alkane-utilizing, sulfate-reducing bacterium, Desulfoglaeba alkanexedens, with either sulfate or Methanospirillum hungatei as electron acceptors, and tested the ability of the cultures to catalyze metal corrosion. Axenically, D. alkanexedens had a higher instantaneous corrosion rate and produced more pits in carbon steel coupons than when the same organism was grown in syntrophic co-culture with the methanogen. Since anaerobic hydrocarbon biodegradation pathways converge on fatty acid intermediates, the corrosive ability of a known fatty acid-oxidizing syntrophic bacterium, Syntrophus aciditrophicus was compared when grown in pure culture or in co-culture with a H2-utilizing sulfate-reducing bacterium (Desulfovibrio sp., strain G11) or a methanogen (M. hungatei). The instantaneous corrosion rates in the cultures were not substantially different, but the syntrophic, sulfate-reducing co-culture produced more pits in coupons than other combinations of microorganisms. Lactate-grown cultures of strain G11 had higher instantaneous corrosion rates and coupon pitting compared to the same organism cultured with hydrogen as an electron donor. Thus, if sulfate is available as an electron acceptor, the same microbial assemblages produce sulfide and low molecular weight organic acids that exacerbated biocorrosion. Despite these trends, a surprisingly high degree of variation was encountered with the corrosion assessments. Differences in biomass, initial substrate concentration, rates of microbial activity or the degree of end product formation did not account for the variations. We are forced to ascribe such differences to the metallurgical properties of the coupons.

Assessment of Anaerobic Biodegradation of Aromatic Hydrocarbons: The Impact of Molecular Biology Approaches

One of the most cost effective methods of pollution remediation is through natural attenuation where the resident microorganisms are responsible for the breakdown of pollutants (Dou et al. 2008). Other forms of bioremediation – such as analogue enrichment, composting and bio-venting – also use the microbes already present in a contaminated site to enhance the remediation process. In order for these approaches to be successful, in an industrial setting, some form of monitoring needs to take place enabling conclusions to be drawn about the degradation processes occurring. In this review we look at some key molecular biology techniques that have the potential to act as a monitoring tool for industries dealing with contaminated land.

Bioconversion of coal: new insights from a core flooding study

A pressurized core flooding experiment was performed to better understand in situ coal bioconversion processes. The core flooding experiment was conducted using a biaxial core holder packed with subbituminous coal particles (250–150 μm grain size) obtained from the Highvale mine in Alberta, Canada. The coal pack was inoculated with a methanogenic microbial culture enriched from coal and was continuously flooded with mineral salt medium and an organic carbon/nitrogen nutrient supplement (tryptone). The changes in the physical properties of the coal pack during the core flooding suggested coal bioconversion to methane under the experimental conditions. Colonization and bioconversion of coal by microbes was evident from the change in core permeability and presence of metabolites and gas (CH4 and CO2) in the effluent. A total of 1.52 μmol of CH4 was produced per gram of coal during the 90 days experiment at 22 °C. Signature metabolites consistent with anaerobic biodegradation of hydrocarbons, e.g., carboxylic acids, were identified in effluent samples throughout incubation. The transient nature of metabolites in effluent samples supports fermentation of coal constituents and nutrient supplement to simple molecules such as acetic acid, which served as a substrate for methanogenesis during the bioconversion process. Accumulation of carboxylic acids such as succinic acid in the effluent also demonstrates that the coal bioconversion process may be used for extraction of other value-added products apart from CH4 generation. Importantly, results presented here suggest that coal bioconversion by biostimulation under reservoir conditions is a scalable technology with potential for energy generation and for overall reduction of greenhouse gas emissions.

Molecular characterization of phenanthrene-degrading methanogenic communities in leachate-contaminated aquifer sediment

The existence of polycyclic aromatic hydrocarbons in the various environments has aroused great environmental concerns due to their potential hazards to human health. The presence of polycyclic aromatic hydrocarbons in aquifer is particularly sensitive where groundwater is used as a source of potable water. Anaerobic biodegradation of polycyclic aromatic hydrocarbons is an attractive option for remediation of contaminated aquifer sediment. Bacterial and archaeal community structures of phenanthrene-degrading aquifer sediment under methanogenic condition were investigated using clone library analysis in combination with microcosm study. The bacterial members were all affiliated with γ-Proteobacteria. Phylum Euryarchaeota was the predominant archaeal group, represented by genera Methanosarcina, Methanobacterium and Thermogymnomonas. Both bacteria (genera Citrobacter and Pseudomonas) and archaea (genus Methanosarcina) might have links to phenanthrene degradation process. This work might provide some new insights into developing strategies for the isolation of the putative polycyclic aromatic hydrocarbons degraders under methanogenic condition and bioremediating polycyclic aromatic hydrocarbons in leachate-contaminated aquifer sediment.

Anthracene biodegradation under nitrate-reducing condition and associated microbial community changes

Anaerobic biodegradation of polycyclic aromatic hydrocarbons (PAHs) and degraders in the subsurface environment have aroused increasing attention. Molecular techniques are especially useful when isolates are hard to obtain. Nitrate-reducing microcosms inoculated with aquifer sediment were constructed to investigate anthracene biodegradation. The associated microbial community changes were characterized using terminal restriction fragment length polymorphism analysis (TRFLP) in combination with 16S rRNA gene clone library analysis. A nearly complete removal of anthracene was achieved after an eighty day incubation under the nitrate-reducing condition. The two molecular techniques revealed a significant shift of microbial community structure, coupled with anthracene biodegradation. Species of genera Paracoccus, Herbaspirillum, Azotobacter, and Rhodococcus were grouped into four major operational taxonomic units (OTUs) in the library that was constructed with the microcosm sample on day 80. The enrichment of these genera might have links to anthracene biodegradation under the nitrate-reducing condition. Microbial consortia likely played a part in anthracene degradation.

Changes in bacterial communities from anaerobic digesters during petroleum hydrocarbon degradation

Anaerobic biodegradation of petroleum hydrocarbons (PHC) to methane has been recognized to occur in oil reservoirs and contaminated surface sites alike. This process could be employed efficiently for the treatment of contaminated materials, including petrochemical wastes and PHC-contaminated soil, since no external electron acceptor is required. Moreover, the controlled production of methane in digestion plants, similarly to the anaerobic digestion (AD) of energy crops or organic residues, would enable for energy recovery from these wastes. At present, little is known about the bacterial communities involved in and responsible for hydrocarbon fermentation, the initial step in PHC conversion to methane. In the present study, the fate of two different methanogenic communities derived from the AD of wastewater (WWT) and of biowaste, mixed with PHC-contaminated soil (SWT), was monitored during incubation with PHC using denaturing gradient gel electrophoresis (DGGE) of 16S rDNA genes amplified with Bacteria-specific primers. During 11 months of incubation, slight but significant degradation of PHC occurred in both sludges and distinct bacterial communities were developing. In both sludges, Bacteroidetes were found. In addition, in WWT, the bacterial community was found to be dominated by Synergistetes and Proteobacteria, while Firmicutes and unidentified members were abundant in SWT. These results indicate that bacterial communities from anaerobic digesters can adapt to and degrade petroleum hydrocarbons. The decontamination of PHC-containing waste via fermentative treatment appears possible.

Dual Biomarkers of Anaerobic Hydrocarbon Degradation in Historically Contaminated Groundwater

This study reports that ongoing in situ anaerobic hydrocarbon biodegradation at a manufactured gas plant impacted site is occurring, 9 years after the initial investigation. Groundwater samples from the site monitoring wells (MW) were analyzed for biomarkers by GC-MS, end-point PCR, and quantitative PCR (qPCR). Metabolic biomarkers included specific intermediates of anaerobic naphthalene and/or 2-methylnaphthalene degradation: 2-naphthoic acid (2-NA); 5,6,7,8-tetrahydro-2-NA (TH-2-NA); hexahydro-2-NA (HH-2-NA); and carboxylated-2-methylnaphthalene (MNA). The analogues of gene bssA, encoding alpha subunit of enzyme benzylsuccinate synthase, were used as a genetic biomarker. Results indicate 1-2 orders of magnitude higher abundance of total bacteria in the impacted wells than in the unimpacted wells. End-point PCR analysis of bssA gene, with degenerate primers, indicated the presence of hydrocarbon degrading bacteria within the plume. In qPCR analysis, using primers based on toluene-degrading denitrifying or sulfate-reducing/methanogenic bacteria, bssA genes were detected only in MW-24, located downstream from the source. Metabolic biomarkers were detected in multiple wells. The highest abundance of 2-NA (6.7 μg/L), TH-2-NA (2.6 μg/L), HH-2-NA, and MNA was also detected in MW-24. The distribution of two independent biomarkers indicates that the site is enriched for anaerobic hydrocarbon biodegradation and provides strong evidence in support of natural attenuation.

Anaerobic biodegradation of petroleum hydrocarbons of a fuel oil contaminated aquifer

Anaerobic biodegradation of petroleum hydrocarbons of a fuel oil contaminated aquifer

Anaerobic biodegradation of polycyclic aromatic hydrocarbons with amendment of iron(III) in mangrove sediment slurry

Mangrove sediment, influenced by tidal cycles, switches between low-oxygen and non-oxygen conditions, and iron is abundant in it. Polycyclic aromatic hydrocarbon (PAH) contamination often occurs in mangrove wetlands. In the present paper, the effects of iron [Fe(III)] amendment on the biodegradation of four mixed PAHs, namely fluorene (Fl), phenanthrene (Phe), fluoranthene (Flua) and pyrene (Pyr), in mangrove sediment slurries, with and without the inoculation of the enriched PAH-degrading bacterial consortia, under low-oxygen (2 ± 0.3% O 2) and non-oxygen (0% O 2) conditions were investigated. Under both oxygen conditions and for all four PAHs, the highest PAHs biodegradation was observed in the groups with the inoculation of the enriched PAH-degrading consortia, while the groups without the inoculum and without Fe(III) amendment had the lowest biodegradation. However, the amendment of Fe(III) did not show any significant improvement on the biodegradation of all the four mixed PAHs.

Vertical distribution and anaerobic biodegradation of polycyclic aromatic hydrocarbons in mangrove sediments in Hong Kong, South China

The vertical distribution of polycyclic aromatic hydrocarbons (PAHs) at different sediment depths, namely 0–2 cm, 2–4 cm, 4–6 cm, 6–10 cm, 10–15 cm and 15–20 cm, in one of the most contaminated mangrove swamps, Ma Wan, Hong Kong was investigated. It was the first time to study the intrinsic potential of deep sediment to biodegrade PAHs under anaerobic conditions and the abundance of electron acceptors in sediment for anaerobic degradation. Results showed that the total PAHs concentrations (summation of 16 US EPA priority PAHs) increased with sediment depth. The lowest concentration (about 1300 ng g − 1 freeze-dried sediment) and the highest value (around 5000 ng g − 1 freeze-dried sediment) were found in the surface layer (0–2 cm) and deeper layer (10–15 cm), respectively. The percentage of high molecular weight (HMW) PAHs (4 to 6 rings) to total PAHs was more than 89% at all sediment depths. The ratio of phenanthrene to anthracene was less than 10 while fluoranthene to pyrene was around 1. Negative redox potentials (Eh) were recorded in all of the sediment samples, ranging from − 170 to − 200 mv, with a sharp decrease at a depth of 6 cm then declined slowly to 20 cm. The results suggested that HMW PAHs originated from diesel-powered fishing vessels and were mainly accumulated in deep anaerobic sediments. Among the electron acceptors commonly used by anaerobic bacteria, sulfate was the most dominant, followed by iron(III), nitrate and manganese(IV) was the least. Their concentrations also decreased with sediment depth. The population size of total anaerobic heterotrophic bacteria increased with sediment depth, reaching the peak number in the middle layer (4–6 cm). In contrast, the aerobic heterotrophic bacterial count decreased with sediment depth. It was the first time to apply a modified electron transport system (ETS) method to evaluate the bacterial activities in the fresh sediment under PAH stress. The vertical drop of the ETS activity suggested that the indigenous bacteria were still active in the anaerobic sediment layer contaminated with PAHs. The biodegradation experiment further proved that the sediment collected at a depth of 10–15 cm harbored anaerobic PAH-degrading bacterial strains (two Sphingomonas, one Microbacterium, one Rhodococcus and two unknown species) with some intrinsic potential to degrade mixed PAHs consisting of fluorene, phenanthrene, fluoranthene and pyrene under low oxygen (2% O 2) and non-oxygen (0% O 2) conditions. This is the first paper to report the anaerobic PAH-degrading bacteria isolated from subsurface mangrove sediment.