Publications by authors named "Casey R J Hubert"

28 Publications

  • Page 1 of 1

Genomic insights into diverse bacterial taxa that degrade extracellular DNA in marine sediments.

Nat Microbiol 2021 Jul 14;6(7):885-898. Epub 2021 Jun 14.

Division of Microbial Ecology, Centre for Microbiology and Environmental Systems Science, University of Vienna, Vienna, Austria.

Extracellular DNA is a major macromolecule in global element cycles, and is a particularly crucial phosphorus, nitrogen and carbon source for microorganisms in the seafloor. Nevertheless, the identities, ecophysiology and genetic features of DNA-foraging microorganisms in marine sediments are largely unknown. Here, we combined microcosm experiments, DNA stable isotope probing (SIP), single-cell SIP using nano-scale secondary isotope mass spectrometry (NanoSIMS) and genome-centric metagenomics to study microbial catabolism of DNA and its subcomponents in marine sediments. C-DNA added to sediment microcosms was largely degraded within 10 d and mineralized to CO. SIP probing of DNA revealed diverse 'Candidatus Izemoplasma', Lutibacter, Shewanella and Fusibacteraceae incorporated DNA-derived C-carbon. NanoSIMS confirmed incorporation of C into individual bacterial cells of Fusibacteraceae sorted from microcosms. Genomes of the C-labelled taxa all encoded enzymatic repertoires for catabolism of DNA or subcomponents of DNA. Comparative genomics indicated that diverse 'Candidatus Izemoplasmatales' (former Tenericutes) are exceptional because they encode multiple (up to five) predicted extracellular nucleases and are probably specialized DNA-degraders. Analyses of additional sediment metagenomes revealed extracellular nuclease genes are prevalent among Bacteroidota at diverse sites. Together, our results reveal the identities and functional properties of microorganisms that may contribute to the key ecosystem function of degrading and recycling DNA in the seabed.
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http://dx.doi.org/10.1038/s41564-021-00917-9DOI Listing
July 2021

Photooxidation and biodegradation potential of a light crude oil in first-year sea ice.

Mar Pollut Bull 2021 Apr 15;165:112154. Epub 2021 Mar 15.

University of Manitoba, Winnipeg, MB, Canada. Electronic address:

Disappearing sea ice in the Arctic region results in a pressing need to develop oil spill mitigation techniques suitable for ice-covered waters. The uncertainty around the nature of an oil spill in the Arctic arises from the ice-covered waters and sub-zero temperatures, and how they may influence natural attenuation efficiency. The Sea-ice Environmental Research Facility was used to create a simulated Arctic marine setting. This paper focuses on the potential for biodegradation of the bulk crude oil content (encapsulated in the upper regions of the ice), to provide insight regarding the possible fate of crude oil in an Arctic marine setting. Cheaper and faster methods of chemical composition analysis were applied to the samples to assess for weathering and transformation effects. Results suggest that brine volume in ice may not be sufficient at low temperatures to encompass biodegradation and that seawater is more suitable for biodegradation.
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http://dx.doi.org/10.1016/j.marpolbul.2021.112154DOI Listing
April 2021

Marine sediments harbor diverse archaea and bacteria with the potential for anaerobic hydrocarbon degradation via fumarate addition.

FEMS Microbiol Ecol 2021 04;97(5)

School of Marine Sciences, Sun Yat-Sen University, 2 Daxue Road, Xiangzhou District, Zhuhai 519082, China.

Marine sediments can contain large amounts of alkanes and methylated aromatic hydrocarbons that are introduced by natural processes or anthropogenic activities. These compounds can be biodegraded by anaerobic microorganisms via enzymatic addition of fumarate. However, the identity and ecological roles of a significant fraction of hydrocarbon degraders containing fumarate-adding enzymes (FAE) in various marine sediments remains unknown. By combining phylogenetic reconstructions, protein homolog modelling, and functional profiling of publicly available metagenomes and genomes, 61 draft bacterial and archaeal genomes encoding anaerobic hydrocarbon degradation via fumarate addition were obtained. Besides Desulfobacterota (previously known as Deltaproteobacteria) that are well-known to catalyze these reactions, Chloroflexi are dominant FAE-encoding bacteria in hydrocarbon-impacted sediments, potentially coupling sulfate reduction or fermentation to anaerobic hydrocarbon degradation. Among Archaea, besides Archaeoglobi previously shown to have this capability, genomes of Heimdallarchaeota, Lokiarchaeota, Thorarchaeota and Thermoplasmata also suggest fermentative hydrocarbon degradation using archaea-type FAE. These bacterial and archaeal hydrocarbon degraders occur in a wide range of marine sediments, including high abundances of FAE-encoding Asgard archaea associated with natural seeps and subseafloor ecosystems. Our results expand the knowledge of diverse archaeal and bacterial lineages engaged in anaerobic degradation of alkanes and methylated aromatic hydrocarbons.
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http://dx.doi.org/10.1093/femsec/fiab045DOI Listing
April 2021

Deep sea sediments associated with cold seeps are a subsurface reservoir of viral diversity.

ISME J 2021 Mar 1. Epub 2021 Mar 1.

School of Marine Sciences, Sun Yat-Sen University, Zhuhai, China.

In marine ecosystems, viruses exert control on the composition and metabolism of microbial communities, influencing overall biogeochemical cycling. Deep sea sediments associated with cold seeps are known to host taxonomically diverse microbial communities, but little is known about viruses infecting these microorganisms. Here, we probed metagenomes from seven geographically diverse cold seeps across global oceans to assess viral diversity, virus-host interaction, and virus-encoded auxiliary metabolic genes (AMGs). Gene-sharing network comparisons with viruses inhabiting other ecosystems reveal that cold seep sediments harbour considerable unexplored viral diversity. Most cold seep viruses display high degrees of endemism with seep fluid flux being one of the main drivers of viral community composition. In silico predictions linked 14.2% of the viruses to microbial host populations with many belonging to poorly understood candidate bacterial and archaeal phyla. Lysis was predicted to be a predominant viral lifestyle based on lineage-specific virus/host abundance ratios. Metabolic predictions of prokaryotic host genomes and viral AMGs suggest that viruses influence microbial hydrocarbon biodegradation at cold seeps, as well as other carbon, sulfur and nitrogen cycling via virus-induced mortality and/or metabolic augmentation. Overall, these findings reveal the global diversity and biogeography of cold seep viruses and indicate how viruses may manipulate seep microbial ecology and biogeochemistry.
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http://dx.doi.org/10.1038/s41396-021-00932-yDOI Listing
March 2021

Characterization of marine microbial communities around an Arctic seabed hydrocarbon seep at Scott Inlet, Baffin Bay.

Sci Total Environ 2021 Mar 7;762:143961. Epub 2020 Dec 7.

Geomicrobiology Group, Department of Biological Sciences, University of Calgary, 2500 University Dr NW, Calgary, Alberta T2N 1N4, Canada.

Seabed hydrocarbon seeps present natural laboratories for investigating responses of marine ecosystems to petroleum input. A hydrocarbon seep near Scott Inlet, Baffin Bay, was visited for in situ observations and sampling in the summer of 2018. Video evidence of an active hydrocarbon seep was confirmed by methane and hydrocarbon analysis of the overlying water column, which is 260 m at this site. Elevated methane concentrations in bottom water above and down current from the seep decreased to background seawater levels in the mid-water column >150 m above the seafloor. Seafloor microbial mats morphologically resembling sulfide-oxidizing bacteria surrounded areas of bubble ebullition. Calcareous tube worms, brittle stars, shrimp, sponges, sea stars, sea anemones, sea urchins, small fish and soft corals were observed near the seep, with soft corals showing evidence for hydrocarbon incorporation. Sediment microbial communities included putative methane-oxidizing Methyloprofundus, sulfate-reducing Desulfobulbaceae and sulfide-oxidizing Sulfurovum. A metabolic gene diagnostic for aerobic methanotrophs (pmoA) was detected in the sediment and bottom water above the seep epicentre and up to 5 km away. Both 16S rRNA gene and pmoA amplicon sequencing revealed that pelagic microbial communities oriented along the geologic basement rise associated with methane seepage (running SW to NE) differed from communities in off-axis water up to 5 km away. Relative abundances of aerobic methanotrophs and putative hydrocarbon-degrading bacteria were elevated in the bottom water down current from the seep. Detection of bacterial clades typically associated with hydrocarbon and methane oxidation highlights the importance of Arctic marine microbial communities in mitigating hydrocarbon emissions from natural geologic sources.
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http://dx.doi.org/10.1016/j.scitotenv.2020.143961DOI Listing
March 2021

Sensitive quantification of dipicolinic acid from bacterial endospores in soils and sediments.

Environ Microbiol 2021 Mar 21;23(3):1397-1406. Epub 2020 Dec 21.

Department of Biological Sciences, University of Calgary, Calgary, T2N 1N4, Canada.

Endospore-forming bacteria make up an important and numerically significant component of microbial communities in a range of settings including soils, industry, hospitals and marine sediments extending into the deep subsurface. Bacterial endospores are non-reproductive structures that protect DNA and improve cell survival during periods unfavourable for bacterial growth. An important determinant of endospores withstanding extreme environmental conditions is 2,6-pyridine dicarboxylic acid (i.e. dipicolinic acid, or DPA), which contributes heat resistance. This study presents an improved HPLC-fluorescence method for DPA quantification using a single 10-min run with pre-column Tb chelation. Relative to existing DPA quantification methods, specific improvements pertain to sensitivity, detection limit and range, as well as the development of new free DPA and spore-specific DPA proxies. The method distinguishes DPA from intact and recently germinated spores, enabling responses to germinants in natural samples or experiments to be assessed in a new way. DPA-based endospore quantification depends on accurate spore-specific DPA contents, in particular, thermophilic spores are shown to have a higher DPA content, meaning that marine sediments with plentiful thermophilic spores may require spore number estimates to be revisited. This method has a wide range of potential applications for more accurately quantifying bacterial endospores in diverse environmental samples.
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http://dx.doi.org/10.1111/1462-2920.15343DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8048543PMC
March 2021

Thermogenic hydrocarbon biodegradation by diverse depth-stratified microbial populations at a Scotian Basin cold seep.

Nat Commun 2020 11 17;11(1):5825. Epub 2020 Nov 17.

Department of Biological Sciences, University of Calgary, Calgary, AB, T2N 1N4, Canada.

At marine cold seeps, gaseous and liquid hydrocarbons migrate from deep subsurface origins to the sediment-water interface. Cold seep sediments are known to host taxonomically diverse microorganisms, but little is known about their metabolic potential and depth distribution in relation to hydrocarbon and electron acceptor availability. Here we combined geophysical, geochemical, metagenomic and metabolomic measurements to profile microbial activities at a newly discovered cold seep in the deep sea. Metagenomic profiling revealed compositional and functional differentiation between near-surface sediments and deeper subsurface layers. In both sulfate-rich and sulfate-depleted depths, various archaeal and bacterial community members are actively oxidizing thermogenic hydrocarbons anaerobically. Depth distributions of hydrocarbon-oxidizing archaea revealed that they are not necessarily associated with sulfate reduction, which is especially surprising for anaerobic ethane and butane oxidizers. Overall, these findings link subseafloor microbiomes to various biochemical mechanisms for the anaerobic degradation of deeply-sourced thermogenic hydrocarbons.
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http://dx.doi.org/10.1038/s41467-020-19648-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7673041PMC
November 2020

Hydrocarbon seepage in the deep seabed links subsurface and seafloor biospheres.

Proc Natl Acad Sci U S A 2020 05 30;117(20):11029-11037. Epub 2020 Apr 30.

Department of Biological Sciences, University of Calgary, Calgary, AB T2N 1N4, Canada;

Marine cold seeps transmit fluids between the subseafloor and seafloor biospheres through upward migration of hydrocarbons that originate in deep sediment layers. It remains unclear how geofluids influence the composition of the seabed microbiome and if they transport deep subsurface life up to the surface. Here we analyzed 172 marine surficial sediments from the deep-water Eastern Gulf of Mexico to assess whether hydrocarbon fluid migration is a mechanism for upward microbial dispersal. While 132 of these sediments contained migrated liquid hydrocarbons, evidence of continuous advective transport of thermogenic alkane gases was observed in 11 sediments. Gas seeps harbored distinct microbial communities featuring bacteria and archaea that are well-known inhabitants of deep biosphere sediments. Specifically, 25 distinct sequence variants within the uncultivated bacterial phyla and and the archaeal order occurred in significantly greater relative sequence abundance along with well-known seep-colonizing members of the bacterial genus , in the gas-positive sediments. Metabolic predictions guided by metagenome-assembled genomes suggested these organisms are anaerobic heterotrophs capable of nonrespiratory breakdown of organic matter, likely enabling them to inhabit energy-limited deep subseafloor ecosystems. These results point to petroleum geofluids as a vector for the advection-assisted upward dispersal of deep biosphere microbes from subsurface to surface environments, shaping the microbiome of cold seep sediments and providing a general mechanism for the maintenance of microbial diversity in the deep sea.
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http://dx.doi.org/10.1073/pnas.2002289117DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7245137PMC
May 2020

Anaerobic microbial communities and their potential for bioenergy production in heavily biodegraded petroleum reservoirs.

Environ Microbiol 2020 08 7;22(8):3049-3065. Epub 2020 May 7.

School of Natural and Environmental Sciences, Newcastle University, Newcastle Upon Tyne, NE1 7RU, UK.

Most of the oil in low temperature, non-uplifted reservoirs is biodegraded due to millions of years of microbial activity, including via methanogenesis from crude oil. To evaluate stimulating additional methanogenesis in already heavily biodegraded oil reservoirs, oil sands samples were amended with nutrients and electron acceptors, but oil sands bitumen was the only organic substrate. Methane production was monitored for over 3000 days. Methanogenesis was observed in duplicate microcosms that were unamended, amended with sulfate or that were initially oxic, however methanogenesis was not observed in nitrate-amended controls. The highest rate of methane production was 0.15 μmol CH g oil d , orders of magnitude lower than other reports of methanogenesis from lighter crude oils. Methanogenic Archaea and several potential syntrophic bacterial partners were detected following the incubations. GC-MS and FTICR-MS revealed no significant bitumen alteration for any specific compound or compound class, suggesting that the very slow methanogenesis observed was coupled to bitumen biodegradation in an unspecific manner. After 3000 days, methanogenic communities were amended with benzoate resulting in methanogenesis rates that were 110-fold greater. This suggests that oil-to-methane conversion is limited by the recalcitrant nature of oil sands bitumen, not the microbial communities resident in heavy oil reservoirs.
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http://dx.doi.org/10.1111/1462-2920.14995DOI Listing
August 2020

Natural attenuation of spilled crude oil by cold-adapted soil bacterial communities at a decommissioned High Arctic oil well site.

Sci Total Environ 2020 Jun 29;722:137258. Epub 2020 Feb 29.

Geomicrobiology Group, Department of Biological Sciences, University of Calgary, EEEL Building, 2500 University Drive NW, Calgary, Alberta T2N 1N4, Canada. Electronic address:

Romulus C-42 is a decommissioned oil well on Ellesmere Island in the Canadian High Arctic, and is the northernmost well to have produced oil and gas anywhere in the world. The remote site has been untouched since a crude oil spill in 1972, offering a rare opportunity to examine natural attenuation in Arctic soils >40 years after a pollution event. Bacterial community composition in crude oil contaminated soils was significantly different from adjacent background soils. Two members of the genus Rhodanobacter (Alphaproteobacteria) were found consistently in contaminated soils, whereas two members of the genus Sphingomonas (Gammaproteobacteria) appeared opposite to each other, one consistently within the oil contaminated soil and the other consistently in non-oil contaminated soils. GC of soil hydrocarbon extracts revealed moderate levels of biodegradation relative to the original oil produced in 1972. Despite conditions permissive for bacterial activity (>0 °C) being limited to only a few months each year, natural attenuation by cold adapted soil microbial communities has taken place since the oil spill over 40 years ago. Rhodanobacter and Sphingomonas lineages are associated with contaminated and baseline conditions in this extreme environment, revealing the utility of bacterial diversity measurements for assessing long-term responses of Arctic soils to pollution. ORIGINALITY-SIGNIFICANCE STATEMENT: Romulus C-42, the northernmost onshore drilling well in the world, was decommissioned following a small crude oil spill in 1972. Soil bacterial diversity profiles obtained >40 years later revealed significant differences in oil contaminated soils relative to adjacent non-oil contaminated background soils, consistent with evidence for moderate biodegradation of spilled crude oil having taken place since 1972. The results indicate that microbial diversity profiling is an effective tool for assessing natural attenuation in remote High Arctic soils with a history of oil pollution.
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http://dx.doi.org/10.1016/j.scitotenv.2020.137258DOI Listing
June 2020

Comparison of sulfide-oxidizing Sulfurimonas strains reveals a new mode of thiosulfate formation in subsurface environments.

Environ Microbiol 2020 05 2;22(5):1784-1800. Epub 2020 Jan 2.

School of Natural and Environmental Sciences, Newcastle University, Devonshire Building (3rd floor), Newcastle upon Tyne, NE1 7RU, UK.

Sulfur-oxidizing Sulfurimonas spp. are widespread in sediments, hydrothermal vent fields, aquifers and subsurface environments such as oil reservoirs where they play an important role in the sulfur cycle. We determined the genome sequence of the oil field isolate Sulfurimonas sp. strain CVO and compared its gene expression during nitrate-dependent sulfide oxidation to the coastal sediment isolate Sulfurimonas denitrificans. Formation of elemental sulfur (S ) and high expression of sulfide quinone oxidoreductase (SQR) genes indicates that sulfide oxidation in both strains is mediated by SQR. Subsequent oxidation of S was achieved by the sulfur oxidation enzyme complex (SOX). In the coastal S. denitrificans, the genes are arranged and expressed as two clusters: soxXY Z AB and soxCDY Z H, and sulfate was the sole metabolic end product. By contrast, the oil field strain CVO has only the soxCDY Z H cluster and not soxXY Z AB. Despite the absence of the soxXY Z AB cluster, strain CVO oxidized S to thiosulfate and sulfate, demonstrating that soxCDY Z H genes alone are sufficient for S oxidation in Sulfurimonas spp. and that thiosulfate is an additional metabolic end product. Screening of publicly available metagenomes revealed that Sulfurimonas spp. with only the soxCDY Z H cluster are widespread suggesting this mechanism of thiosulfate formation is environmentally significant.
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http://dx.doi.org/10.1111/1462-2920.14894DOI Listing
May 2020

Sediment cooling triggers germination and sulfate reduction by heat-resistant thermophilic spore-forming bacteria.

Environ Microbiol 2020 01 2;22(1):456-465. Epub 2019 Dec 2.

School of Natural and Environmental Sciences, Newcastle University, Newcastle upon Tyne, UK.

Thermophilic endospores are widespread in cold marine sediments where the temperature is too low to support growth and activity of thermophiles in situ. These endospores are likely expelled from warm subsurface environments and subsequently dispersed by ocean currents. The endospore upper temperature limit for survival is 140°C, which can be tolerated in repeated short exposures, potentially enabling transit through hot crustal fluids. Longer-term thermal tolerance of endospores, and how long they could persist in an environment hotter than their maximum growth temperature, is less understood. To test whether thermophilic endospores can survive prolonged exposure to high temperatures, sediments were incubated at 80-90°C for 6, 12 or 463 days. Sediments were then cooled by 10-40°C, mimicking the cooling in subsurface oil reservoirs subjected to seawater injection. Cooling the sediments induced sulfate reduction, coinciding with an enrichment of endospore-forming Clostridia. Different Desulfofundulus, Desulfohalotomaculum, Desulfallas, Desulfotomaculum and Desulfofarcimen demonstrated different thermal tolerances, with some Desulfofundulus strains surviving for >1 year at 80°C. In an oil reservoir context, heat-resistant endospore-forming sulfate-reducing bacteria have a survival advantage if they are introduced to, or are resident in, an oil reservoir normally too hot for germination and growth, explaining observations of reservoir souring following cold seawater injection.
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http://dx.doi.org/10.1111/1462-2920.14866DOI Listing
January 2020

Freezing Tolerance of Thermophilic Bacterial Endospores in Marine Sediments.

Front Microbiol 2019 3;10:945. Epub 2019 May 3.

Geomicrobiology Group, Department of Biological Sciences, University of Calgary, Calgary, AB, Canada.

Dormant endospores of anaerobic, thermophilic bacteria found in cold marine sediments offer a useful model for studying microbial biogeography, dispersal, and survival. The dormant endospore phenotype confers resistance to unfavorable environmental conditions, allowing dispersal to be isolated and studied independently of other factors such as environmental selection. To study the resilience of thermospores to conditions relevant for survival in extreme cold conditions, their viability following different freezing treatments was tested. Marine sediment was frozen at either -80°C or -20°C for 10 days prior to pasteurization and incubation at +50°C for 21 days to assess thermospore viability. Sulfate reduction commenced at +50°C following both freezing pretreatments indicating persistence of thermophilic endospores of sulfate-reducing bacteria. The onset of sulfate reduction at +50°C was delayed in -80°C pretreated microcosms, which exhibited more variability between triplicates, compared to -20°C pretreated microcosms and parallel controls that were not frozen in advance. Microbial communities were evaluated by 16S rRNA gene amplicon sequencing, revealing an increase in the relative sequence abundance of thermophilic endospore-forming in all microcosms. Different freezing pretreatments (-80°C and -20°C) did not appreciably influence the shift in overall bacterial community composition that occurred during the +50°C incubations. Communities that had been frozen prior to +50°C incubation showed an increase in the relative sequence abundance of operational taxonomic units (OTUs) affiliated with the class , relative to unfrozen controls. These results show that freezing impacts but does not obliterate thermospore populations and their ability to germinate and grow under appropriate conditions. Indeed the majority of the thermospore OTUs detected in this study (21 of 22) could be observed following one or both freezing treatments. These results are important for assessing thermospore viability in frozen samples and following cold exposure such as the very low temperatures that would be encountered during panspermia.
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http://dx.doi.org/10.3389/fmicb.2019.00945DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6509201PMC
May 2019

Metabolic potential of uncultured bacteria and archaea associated with petroleum seepage in deep-sea sediments.

Nat Commun 2019 04 18;10(1):1816. Epub 2019 Apr 18.

Department of Biological Sciences, University of Calgary, Calgary, AB, T2N 1N4, Canada.

The lack of microbial genomes and isolates from the deep seabed means that very little is known about the ecology of this vast habitat. Here, we investigate energy and carbon acquisition strategies of microbial communities from three deep seabed petroleum seeps (3 km water depth) in the Eastern Gulf of Mexico. Shotgun metagenomic analysis reveals that each sediment harbors diverse communities of chemoheterotrophs and chemolithotrophs. We recovered 82 metagenome-assembled genomes affiliated with 21 different archaeal and bacterial phyla. Multiple genomes encode enzymes for anaerobic oxidation of aliphatic and aromatic compounds, including those of candidate phyla Aerophobetes, Aminicenantes, TA06 and Bathyarchaeota. Microbial interactions are predicted to be driven by acetate and molecular hydrogen. These findings are supported by sediment geochemistry, metabolomics, and thermodynamic modelling. Overall, we infer that deep-sea sediments experiencing thermogenic hydrocarbon inputs harbor phylogenetically and functionally diverse communities potentially sustained through anaerobic hydrocarbon, acetate and hydrogen metabolism.
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http://dx.doi.org/10.1038/s41467-019-09747-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6472368PMC
April 2019

Historical Factors Associated With Past Environments Influence the Biogeography of Thermophilic Endospores in Arctic Marine Sediments.

Front Microbiol 2019 28;10:245. Epub 2019 Feb 28.

School of Civil Engineering and Geosciences, Newcastle University, Newcastle upon Tyne, United Kingdom.

Selection by the local, contemporary environment plays a prominent role in shaping the biogeography of microbes. However, the importance of historical factors in microbial biogeography is more debatable. Historical factors include past ecological and evolutionary circumstances that may have influenced present-day microbial diversity, such as dispersal and past environmental conditions. Diverse thermophilic sulfate-reducing are present as dormant endospores in marine sediments worldwide where temperatures are too low to support their growth. Therefore, they are dispersed to here from elsewhere, presumably a hot, anoxic habitat. While dispersal through ocean currents must influence their distribution in cold marine sediments, it is not clear whether even earlier historical factors, related to the source habitat where these organisms were once active, also have an effect. We investigated whether these historical factors may have influenced the diversity and distribution of thermophilic endospores by comparing their diversity in 10 Arctic fjord surface sediments. Although community composition varied spatially, clear biogeographic patterns were only evident at a high level of taxonomic resolution (>97% sequence similarity of the 16S rRNA gene) achieved with oligotyping. In particular, the diversity and distribution of oligotypes differed for the two most prominent OTUs (defined using a standard 97% similarity cutoff). One OTU was dominated by a single ubiquitous oligotype, while the other OTU consisted of ten more spatially localized oligotypes that decreased in compositional similarity with geographic distance. These patterns are consistent with differences in historical factors that occurred when and where the taxa were once active, prior to sporulation. Further, the influence of history on biogeographic patterns was only revealed by analyzing microdiversity within OTUs, suggesting that populations within standard OTU-level groupings do not necessarily share a common ecological and evolutionary history.
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http://dx.doi.org/10.3389/fmicb.2019.00245DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6403435PMC
February 2019

Metabolites of an Oil Field Sulfide-Oxidizing, Nitrate-Reducing sp. Cause Severe Corrosion.

Appl Environ Microbiol 2019 02 23;85(3). Epub 2019 Jan 23.

School of Natural and Environmental Sciences, Newcastle University, Newcastle upon Tyne, United Kingdom.

Oil reservoir souring and associated material integrity challenges are of great concern to the petroleum industry. The bioengineering strategy of nitrate injection has proven successful for controlling souring in some cases, but recent reports indicate increased corrosion in nitrate-treated produced water reinjection facilities. Sulfide-oxidizing, nitrate-reducing bacteria (soNRB) have been suggested to be the cause of such corrosion. Using the model soNRB sp. strain CVO obtained from an oil field, we conducted a detailed analysis of soNRB-induced corrosion at initial nitrate-to-sulfide (N/S) ratios relevant to oil field operations. The activity of strain CVO caused severe corrosion rates of up to 0.27 millimeters per year (mm y) and up to 60-μm-deep pitting within only 9 days. The highest corrosion during the growth of strain CVO was associated with the production of zero-valent sulfur during sulfide oxidation and the accumulation of nitrite, when initial N/S ratios were high. Abiotic corrosion tests with individual metabolites confirmed biogenic zero-valent sulfur and nitrite as the main causes of corrosion under the experimental conditions. Mackinawite (FeS) deposited on carbon steel surfaces accelerated abiotic reduction of both sulfur and nitrite, exacerbating corrosion. Based on these results, a conceptual model for nitrate-mediated corrosion by soNRB is proposed. Ambiguous reports of corrosion problems associated with the injection of nitrate for souring control necessitate a deeper understanding of this frequently applied bioengineering strategy. Sulfide-oxidizing, nitrate-reducing bacteria have been proposed as key culprits, despite the underlying microbial corrosion mechanisms remaining insufficiently understood. This study provides a comprehensive characterization of how individual metabolic intermediates of the microbial nitrogen and sulfur cycles can impact the integrity of carbon steel infrastructure. The results help explain the dramatic increases seen at times in corrosion rates observed during nitrate injection in field and laboratory trials and point to strategies for reducing adverse integrity-related side effects of nitrate-based souring mitigation.
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http://dx.doi.org/10.1128/AEM.01891-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6344618PMC
February 2019

Thermophilic endospores associated with migrated thermogenic hydrocarbons in deep Gulf of Mexico marine sediments.

ISME J 2018 08 29;12(8):1895-1906. Epub 2018 Mar 29.

Department of Biological Sciences, University of Calgary, Calgary, AB, Canada.

Dormant endospores of thermophilic bacteria (thermospores) can be detected in cold marine sediments following high-temperature incubation. Thermospores in the cold seabed may be explained by a dispersal history originating in deep biosphere oil reservoir habitats where upward migration of petroleum fluids at hydrocarbon seeps transports viable cells into the overlying ocean. We assessed this deep-to-shallow dispersal hypothesis through geochemical and microbiological analyses of 111 marine sediments from the deep water Eastern Gulf of Mexico. GC-MS and fluorescence confirmed the unambiguous presence of thermogenic hydrocarbons in 71 of these locations, indicating seepage from deeply sourced petroleum in the subsurface. Heating each sediment to 50 °C followed by 16S rRNA gene sequencing revealed several thermospores with a cosmopolitan distribution throughout the study area, as well as thermospores that were more geographically restricted. Among the thermospores having a more limited distribution, 12 OTUs from eight different lineages were repeatedly detected in sediments containing thermogenic hydrocarbons. A subset of these were significantly correlated with hydrocarbons (p < 0.05) and most closely related to Clostridiales previously detected in oil reservoirs from around the world. This provides evidence of bacteria in the ocean being dispersed out of oil reservoirs, and suggests that specific thermospores may be used as model organisms for studying warm-to-cold transmigration in the deep sea.
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http://dx.doi.org/10.1038/s41396-018-0108-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6052102PMC
August 2018

Distribution of thermophilic endospores in a temperate estuary indicate that dispersal history structures sediment microbial communities.

Environ Microbiol 2018 03 23;20(3):1134-1147. Epub 2018 Feb 23.

School of Natural and Environmental Sciences, Newcastle University, Newcastle upon Tyne, NE1 7RU, UK.

Endospores of thermophilic bacteria are found in cold and temperate sediments where they persist in a dormant state. As inactive endospores that cannot grow at the low ambient temperatures, they are akin to tracer particles in cold sediments, unaffected by factors normally governing microbial biogeography (e.g., selection, drift, mutation). This makes thermophilic endospores ideal model organisms for studying microbial biogeography since their spatial distribution can be directly related to their dispersal history. To assess dispersal histories of estuarine bacteria, thermophilic endospores were enriched from sediments along a freshwater-to-marine transect of the River Tyne in high temperature incubations (50°C). Dispersal histories for 75 different taxa indicated that the majority of estuarine endospores were of terrestrial origin; most closely related to bacteria from warm habitats associated with industrial activity. A subset of the taxa detected were marine derived, with close relatives from hot deep marine biosphere habitats. These patterns are consistent with the sources of sediment in the River Tyne being predominantly terrestrial in origin. The results point to microbial communities in estuarine and marine sediments being structured by bi-directional currents, terrestrial run-off and industrial effluent as vectors of passive dispersal and immigration.
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http://dx.doi.org/10.1111/1462-2920.14056DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6849807PMC
March 2018

Using Thermodynamics to Predict the Outcomes of Nitrate-Based Oil Reservoir Souring Control Interventions.

Front Microbiol 2017 19;8:2575. Epub 2017 Dec 19.

School of Civil Engineering and Geosciences, Newcastle University, Newcastle upon Tyne, United Kingdom.

Souring is the undesirable production of hydrogen sulfide (HS) in oil reservoirs by sulfate-reducing bacteria (SRB). Souring is a common problem during secondary oil recovery via water flooding, especially when seawater with its high sulfate concentration is introduced. Nitrate injection into these oil reservoirs can prevent and remediate souring by stimulating nitrate-reducing bacteria (NRB). Two conceptually different mechanisms for NRB-facilitated souring control have been proposed: nitrate-sulfate competition for electron donors (oil-derived organics or H) and nitrate driven sulfide oxidation. Thermodynamics can facilitate predictions about which nitrate-driven mechanism is most likely to occur in different scenarios. From a thermodynamic perspective the question "Which reaction yields more energy, nitrate driven oxidation of sulfide or nitrate driven oxidation of organic compounds?" can be rephrased as: "Is acetate driven sulfate reduction to sulfide exergonic or endergonic?" Our analysis indicates that under conditions encountered in oil fields, sulfate driven oxidation of acetate (or other SRB organic electron donors) is always more favorable than sulfide oxidation to sulfate. That predicts that organotrophic NRB that oxidize acetate would outcompete lithotrophic NRB that oxidize sulfide. However, sulfide oxidation to elemental sulfur is different. At low acetate HS oxidation is more favorable than acetate oxidation. Incomplete oxidation of sulfide to S is likely to occur when nitrate levels are low, and is favored by low temperatures; conditions that can be encountered at oil field above-ground facilities where intermediate sulfur compounds like S may cause corrosion. These findings have implications for reservoir management strategies and for assessing the success and progress of nitrate-based souring control strategies and the attendant risks of corrosion associated with souring and nitrate injection.
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http://dx.doi.org/10.3389/fmicb.2017.02575DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5742191PMC
December 2017

Microbial Communities in a High Arctic Polar Desert Landscape.

Front Microbiol 2016 31;7:419. Epub 2016 Mar 31.

School of Civil Engineering and Geosciences, Newcastle University Newcastle upon Tyne, UK.

The High Arctic is dominated by polar desert habitats whose microbial communities are poorly understood. In this study, we used next generation sequencing to describe the α- and β-diversity of microbial communities in polar desert soils from the Kongsfjorden region of Svalbard. Ten phyla dominated the soils and accounted for 95% of all sequences, with the Proteobacteria, Actinobacteria, and Chloroflexi being the major lineages. In contrast to previous investigations of Arctic soils, relative Acidobacterial abundances were found to be very low as were the Archaea throughout the Kongsfjorden polar desert landscape. Lower Acidobacterial abundances were attributed to characteristic circumneutral soil pHs in this region, which has resulted from the weathering of underlying carbonate bedrock. In addition, we compared previously measured geochemical conditions as possible controls on soil microbial communities. Phosphorus, pH, nitrogen, and calcium levels all significantly correlated with β-diversity, indicating landscape-scale lithological control of available nutrients, which in turn, significantly influenced soil community composition. In addition, soil phosphorus and pH significantly correlated with α-diversity, particularly with the Shannon diversity and Chao 1 richness indices.
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http://dx.doi.org/10.3389/fmicb.2016.00419DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4814466PMC
April 2016

Activity and community structures of sulfate-reducing microorganisms in polar, temperate and tropical marine sediments.

ISME J 2016 Apr 11;10(4):796-809. Epub 2015 Sep 11.

Department of Geological Sciences, Bolin Centre for Climate Research, Stockholm University, Stockholm, Sweden.

Temperature has a fundamental impact on the metabolic rates of microorganisms and strongly influences microbial ecology and biogeochemical cycling in the environment. In this study, we examined the catabolic temperature response of natural communities of sulfate-reducing microorganisms (SRM) in polar, temperate and tropical marine sediments. In short-term sediment incubation experiments with (35)S-sulfate, we demonstrated how the cardinal temperatures for sulfate reduction correlate with mean annual sediment temperatures, indicating specific thermal adaptations of the dominant SRM in each of the investigated ecosystems. The community structure of putative SRM in the sediments, as revealed by pyrosequencing of bacterial 16S rRNA gene amplicons and phylogenetic assignment to known SRM taxa, consistently correlated with in situ temperatures, but not with sediment organic carbon concentrations or C:N ratios of organic matter. Additionally, several species-level SRM phylotypes of the class Deltaproteobacteria tended to co-occur at sites with similar mean annual temperatures, regardless of geographic distance. The observed temperature adaptations of SRM imply that environmental temperature is a major controlling variable for physiological selection and ecological and evolutionary differentiation of microbial communities.
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http://dx.doi.org/10.1038/ismej.2015.157DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4796921PMC
April 2016

Survival of Desulfotomaculum spores from estuarine sediments after serial autoclaving and high-temperature exposure.

ISME J 2015 Mar 17;9(4):922-33. Epub 2015 Mar 17.

School of Earth and Ocean Sciences, Cardiff University, Cardiff CF10 3AT, Wales, UK.

Bacterial spores are widespread in marine sediments, including those of thermophilic, sulphate-reducing bacteria, which have a high minimum growth temperature making it unlikely that they grow in situ. These Desulfotomaculum spp. are thought to be from hot environments and are distributed by ocean currents. Their cells and spores upper temperature limit for survival is unknown, as is whether they can survive repeated high-temperature exposure that might occur in hydrothermal systems. This was investigated by incubating estuarine sediments significantly above (40-80 °C) maximum in situ temperatures (∼ 23 °C), and with and without prior triple autoclaving. Sulphate reduction occurred at 40-60 °C and at 60 °C was unaffected by autoclaving. Desulfotomaculum sp. C1A60 was isolated and was most closely related to the thermophilic D. kuznetsovii(T) (∼ 96% 16S rRNA gene sequence identity). Cultures of Desulfotomaculum sp. C1A60, D. kuznetsovii(T)and D. geothermicum B2T survived triple autoclaving while other related Desulfotomaculum spp. did not, although they did survive pasteurisation. Desulfotomaculum sp. C1A60 and D. kuznetsovii cultures also survived more extreme autoclaving (C1A60, 130 °C for 15 min; D. kuznetsovii, 135 °C for 15 min, maximum of 154 °C reached) and high-temperature conditions in an oil bath (C1A60, 130° for 30 min, D. kuznetsovii 140 °C for 15 min). Desulfotomaculum sp. C1A60 with either spores or predominantly vegetative cells demonstrated that surviving triple autoclaving was due to spores. Spores also had very high culturability compared with vegetative cells (∼ 30 × higher). Combined extreme temperature survival and high culturability of some thermophilic Desulfotomaculum spp. make them very effective colonisers of hot environments, which is consistent with their presence in subsurface geothermal waters and petroleum reservoirs.
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http://dx.doi.org/10.1038/ismej.2014.190DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4817712PMC
March 2015

Endospores of thermophilic bacteria as tracers of microbial dispersal by ocean currents.

ISME J 2014 Jun 19;8(6):1153-65. Epub 2013 Dec 19.

1] Division of Microbial Ecology, Department of Microbiology and Ecosystem Science, Faculty of Life Sciences, University of Vienna, Vienna, Austria [2] Austrian Polar Research Institute, Vienna, Austria.

Microbial biogeography is influenced by the combined effects of passive dispersal and environmental selection, but the contribution of either factor can be difficult to discern. As thermophilic bacteria cannot grow in the cold seabed, their inactive spores are not subject to environmental selection. We therefore conducted a global experimental survey using thermophilic endospores that are passively deposited by sedimentation to the cold seafloor as tracers to study the effect of dispersal by ocean currents on the biogeography of marine microorganisms. Our analysis of 81 different marine sediments from around the world identified 146 species-level 16S rRNA phylotypes of endospore-forming, thermophilic Firmicutes. Phylotypes showed various patterns of spatial distribution in the world oceans and were dispersal-limited to different degrees. Co-occurrence of several phylotypes in locations separated by great distances (west of Svalbard, the Baltic Sea and the Gulf of California) demonstrated a widespread but not ubiquitous distribution. In contrast, Arctic regions with water masses that are relatively isolated from global ocean circulation (Baffin Bay and east of Svalbard) were characterized by low phylotype richness and different compositions of phylotypes. The observed distribution pattern of thermophilic endospores in marine sediments suggests that the impact of passive dispersal on marine microbial biogeography is controlled by the connectivity of local water masses to ocean circulation.
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http://dx.doi.org/10.1038/ismej.2013.225DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4030223PMC
June 2014

Improving PCR efficiency for accurate quantification of 16S rRNA genes.

J Microbiol Methods 2013 May 22;93(2):148-52. Epub 2013 Mar 22.

Petroleum Microbiology Research Group, University of Calgary, Department of Biological Sciences, Calgary, Alberta, T2N 1N4, Canada.

Quantitative real-time PCR is a valuable tool for microbial ecologists. To obtain accurate absolute quantification it is essential that PCR efficiency for pure standards is close to amplification efficiency for test samples. Counter to normal expectation that PCR efficiency might be lower in environmental DNA, due to the presence of PCR inhibitors, we report the counterintuitive observation that PCR efficiency of pure standards can be lower than for environmental DNA. This can lead to overestimation of gene abundances if not corrected. SYBR green-based qPCR assays of 16S rRNA genes targeting Bacteria, Syntrophus and Smithella spp., Marinobacter spp., Methanomicrobiales, Methanosarcinaceae, and Methanosaetaceae in samples from methanogenic crude oil biodegradation enrichments were tested. In five out of the six assays, PCR efficiency was lower with pure standards than with environmental DNA samples. We developed a solution to this problem based on amending pure clone standards with a background of non-target environmental 16S rRNA genes which significantly improved PCR efficiency of standards in the qPCR assays that exhibited this phenomenon. Overall this method of qPCR standard preparation achieved a more reliable and robust quantification of 16S rRNA genes. We believe this may be a potentially common issue in microbial ecology that often goes unreported, as intuitively one would not expect standards to have poorer PCR efficiency than samples.
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http://dx.doi.org/10.1016/j.mimet.2013.03.010DOI Listing
May 2013

Microbial diversity and anaerobic hydrocarbon degradation potential in an oil-contaminated mangrove sediment.

BMC Microbiol 2012 Aug 30;12:186. Epub 2012 Aug 30.

Laboratório de Ecologia Molecular Microbiana, Instituto de Microbiologia Paulo de Góes, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil.

Background: Mangrove forests are coastal wetlands that provide vital ecosystem services and serve as barriers against natural disasters like tsunamis, hurricanes and tropical storms. Mangroves harbour a large diversity of organisms, including microorganisms with important roles in nutrient cycling and availability. Due to tidal influence, mangroves are sites where crude oil from spills farther away can accumulate. The relationship between mangrove bacterial diversity and oil degradation in mangrove sediments remains poorly understood.

Results: Mangrove sediment was sampled from 0-5, 15-20 and 35-40 cm depth intervals from the Suruí River mangrove (Rio de Janeiro, Brazil), which has a history of oil contamination. DGGE fingerprinting for bamA, dsr and 16S rRNA encoding fragment genes, and qPCR analysis using dsr and 16S rRNA gene fragment revealed differences with sediment depth.

Conclusions: Analysis of bacterial 16S rRNA gene diversity revealed changes with depth. DGGE for bamA and dsr genes shows that the anaerobic hydrocarbon-degrading community profile also changed between 5 and 15 cm depth, and is similar in the two deeper sediments, indicating that below 15 cm the anaerobic hydrocarbon-degrading community appears to be well established and homogeneous in this mangrove sediment. qPCR analysis revealed differences with sediment depth, with general bacterial abundance in the top layer (0-5 cm) being greater than in both deeper sediment layers (15-20 and 35-40 cm), which were similar to each other.
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http://dx.doi.org/10.1186/1471-2180-12-186DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3579730PMC
August 2012

Dispersal of thermophilic Desulfotomaculum endospores into Baltic Sea sediments over thousands of years.

ISME J 2013 Jan 26;7(1):72-84. Epub 2012 Jul 26.

Center for Geomicrobiology, Department of Bioscience, Aarhus University, Aarhus, Denmark.

Patterns of microbial biogeography result from a combination of dispersal, speciation and extinction, yet individual contributions exerted by each of these mechanisms are difficult to isolate and distinguish. The influx of endospores of thermophilic microorganisms to cold marine sediments offers a natural model for investigating passive dispersal in the ocean. We investigated the activity, diversity and abundance of thermophilic endospore-forming sulfate-reducing bacteria (SRB) in Aarhus Bay by incubating pasteurized sediment between 28 and 85 °C, and by subsequent molecular diversity analyses of 16S rRNA and of the dissimilatory (bi)sulfite reductase (dsrAB) genes within the endospore-forming SRB genus Desulfotomaculum. The thermophilic Desulfotomaculum community in Aarhus Bay sediments consisted of at least 23 species-level 16S rRNA sequence phylotypes. In two cases, pairs of identical 16S rRNA and dsrAB sequences in Arctic surface sediment 3000 km away showed that the same phylotypes are present in both locations. Radiotracer-enhanced most probable number analysis revealed that the abundance of endospores of thermophilic SRB in Aarhus Bay sediment was ca. 10(4) per cm(3) at the surface and decreased exponentially to 10(0) per cm(3) at 6.5 m depth, corresponding to 4500 years of sediment age. Thus, a half-life of ca. 300 years was estimated for the thermophilic SRB endospores deposited in Aarhus Bay sediments. These endospores were similarly detected in the overlying water column, indicative of passive dispersal in water masses preceding sedimentation. The sources of these thermophiles remain enigmatic, but at least one source may be common to both Aarhus Bay and Arctic sediments.
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http://dx.doi.org/10.1038/ismej.2012.83DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3524260PMC
January 2013

Massive dominance of Epsilonproteobacteria in formation waters from a Canadian oil sands reservoir containing severely biodegraded oil.

Environ Microbiol 2012 Feb 8;14(2):387-404. Epub 2011 Aug 8.

School of Civil Engineering and Geosciences, Newcastle University, Newcastle upon Tyne, UK.

The subsurface microbiology of an Athabasca oil sands reservoir in western Canada containing severely biodegraded oil was investigated by combining 16S rRNA gene- and polar lipid-based analyses of reservoir formation water with geochemical analyses of the crude oil and formation water. Biomass was filtered from formation water, DNA was extracted using two different methods, and 16S rRNA gene fragments were amplified with several different primer pairs prior to cloning and sequencing or community fingerprinting by denaturing gradient gel electrophoresis (DGGE). Similar results were obtained irrespective of the DNA extraction method or primers used. Archaeal libraries were dominated by Methanomicrobiales (410 of 414 total sequences formed a dominant phylotype affiliated with a Methanoregula sp.), consistent with the proposed dominant role of CO(2) -reducing methanogens in crude oil biodegradation. In two bacterial 16S rRNA clone libraries generated with different primer pairs, > 99% and 100% of the sequences were affiliated with Epsilonproteobacteria (n = 382 and 72 total clones respectively). This massive dominance of Epsilonproteobacteria sequences was again obtained in a third library (99% of sequences; n = 96 clones) using a third universal bacterial primer pair (inosine-341f and 1492r). Sequencing of bands from DGGE profiles and intact polar lipid analyses were in accordance with the bacterial clone library results. Epsilonproteobacterial OTUs were affiliated with Sulfuricurvum, Arcobacter and Sulfurospirillum spp. detected in other oil field habitats. The dominant organism revealed by the bacterial libraries (87% of all sequences) is a close relative of Sulfuricurvum kujiense - an organism capable of oxidizing reduced sulfur compounds in crude oil. Geochemical analysis of organic extracts from bitumen at different reservoir depths down to the oil water transition zone of these oil sands indicated active biodegradation of dibenzothiophenes, and stable sulfur isotope ratios for elemental sulfur and sulfate in formation waters were indicative of anaerobic oxidation of sulfur compounds. Microbial desulfurization of crude oil may be an important metabolism for Epsilonproteobacteria indigenous to oil reservoirs with elevated sulfur content and may explain their prevalence in formation waters from highly biodegraded petroleum systems.
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http://dx.doi.org/10.1111/j.1462-2920.2011.02521.xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3490369PMC
February 2012