Modeling of heavy nitrate corrosion in anaerobe aquifer injection water biofilm: a case study in a flow rig.

Heavy carbon steel corrosion developed during nitrate mitigation of a flow rig connected to a water injection pipeline flowing anaerobe saline aquifer water. Genera-specific QPCR primers quantified 74% of the microbial biofilm community, and further 87% of the community of the nonamended parallel rig. The nonamended biofilm hosted 6.3 × 10(6) SRB cells/cm(2) and the S(35)-sulfate-reduction rate was 1.1 µmol SO4(2-)/cm(2)/day, being congruent with the estimated SRB biomass formation and the sulfate areal flux. Nitrate amendment caused an 18-fold smaller SRB population, but up to 44 times higher sulfate reduction rates. This H2S formation was insufficient to form the observed Fe3S4 layer. Additional H2S was provided by microbial disproportionation of sulfur, also explaining the increased accessibility of sulfate. The reduced nitrate specie nitrite inhibited the dominating H2-scavenging Desulfovibrio population, and sustained the formation of polysulfide and Fe3S4, herby also dissolved sulfur. This terminated the availability of acetate in the inner biofilm and caused cell starvation that initiated growth upon metallic electrons, probably by the sulfur-reducing Desulfuromonas population. On the basis of these observations we propose a model of heavy nitrate corrosion where three microbiological processes of nitrate reduction, disproportionation of sulfur, and metallic electron growth are nicely woven into each other.

Hydrocarbon degradation by Dietzia sp. A14101 isolated from an oil reservoir model column.

The hydrocarbon-degrading strain Dietzia sp. A14101 was isolated from an oil reservoir model column inoculated with oil-field bacteria. The column was continuously injected with nitrate (0.5 mM) from the start of water flooding, which lead to a gradual development of nitrate reduction in the column. Strain A14101 was able to utilize a range of aliphatic hydrocarbons as sole carbon and energy source during aerobic growth. Whole oil gas chromatography analysis of the crude oil phase from aerobic pure cultures showed that strain A14101 utilized the near complete range of aliphatic components and aromatic components toluene and xylene. Longer n-alkanes >/=C(17) were utilized simultaneously with the shorter C(10) and C(15). After 120 days aerobic incubation, the whole oil gas chromatography profile of the crude oil phase was similar to that of heavily biodegraded oils. Anaerobic degradation of hydrocarbons with nitrate was not observed. Nitrate reduction was, however, observed during anaerobic growth on propionate, which suggests that strain A14101 grows on fatty acids in the column rather than on hydrocarbons.

Microbial response to reinjection of produced water in an oil reservoir.

The microbial response to produced water reinjection (PWRI) in a North Sea oil field was investigated by a combination of cultivation and culture-independent molecular phylogenetic techniques. Special emphasise was put on the relationship between sulphate-reducing bacteria (SRB) and nitrate-reducing bacteria (NRB), and results were used to evaluate the possibility of nitrate treatment as a souring management tool during PWRI. Samples were collected by reversing the flow of the injection water, which provided samples from around the injection area. The backflowed samples were compared to produced water from the same platform and to backflowed samples from a biocide-treated seawater injector, which was the previous injection water treatment of the PWRI well. Results showed that reinjection of produced water promoted growth of thermophilic SRB. Thermophilic fatty acid oxidising NRB and potential nitrate-reducing sulphide-oxidising bacteria were also found. The finding of thermophilic NRB makes nitrate treatment during PWRI possible, although higher nitrate concentration will be necessary to compensate for the increased SRB activity.

Microbial analysis of backflowed injection water from a nitrate-treated North Sea oil reservoir.

Reservoir souring in offshore oil fields is caused by hydrogen sulphide (H(2)S) produced by sulphate-reducing bacteria (SRB), most often as a consequence of sea water injection. Biocide treatment is commonly used to inhibit SRB, but has now been replaced by nitrate treatment on several North Sea oil fields. At the Statfjord field, injection wells from one nitrate-treated reservoir and one biocide-treated reservoir were reversed (backflowed) and sampled for microbial analysis. The two reservoirs have similar properties and share the same pre-nitrate treatment history. A 16S rRNA gene-based community analysis (PCR-DGGE) combined with enrichment culture studies showed that, after 6 months of nitrate injection (0.25 mM NO(3) (-)), heterotrophic and chemolithotrophic nitrate-reducing bacteria (NRB) formed major populations in the nitrate-treated reservoir. The NRB community was able to utilize the same substrates as the SRB community. Compared to the biocide-treated reservoir, the microbial community in the nitrate-treated reservoir was more phylogenetically diverse and able to grow on a wider range of substrates. Enrichment culture studies showed that SRB were present in both reservoirs, but the nitrate-treated reservoir had the least diverse SRB community. Isolation and characterisation of one of the dominant populations observed during nitrate treatment (strain STF-07) showed that heterotrophic denitrifying bacteria affiliated to Terasakiella probably contributed significantly to the inhibition of SRB.

The effect of long-term nitrate treatment on SRB activity, corrosion rate and bacterial community composition in offshore water injection systems.

Biogenic production of hydrogen sulphide (H(2)S) is a problem for the oil industry as it leads to corrosion and reservoir souring. Continuous injection of a low nitrate concentration (0.25-0.33 mM) replaced glutaraldehyde as corrosion and souring control at the Veslefrikk and Gullfaks oil field (North Sea) in 1999. The response to nitrate treatment was a rapid reduction in number and activity of sulphate-reducing bacteria (SRB) in the water injection system biofilm at both fields. The present long-term study shows that SRB activity has remained low at < or =0.3 and < or =0.9 microg H(2)S/cm(2)/day at Veslefrikk and Gullfaks respectively, during the 7-8 years with continuous nitrate injection. At Veslefrikk, 16S rRNA gene based community analysis by PCR-DGGE showed that bacteria affiliated to nitrate-reducing sulphide-oxidizing Sulfurimonas (NR-SOB) formed major populations at the injection well head throughout the treatment period. Downstream of deaerator the presence of Sulfurimonas like bacteria was less pronounced, and were no longer observed 40 months into the treatment period. The biofilm community during nitrate treatment was highly diverse and relative stable for long periods of time. At the Gullfaks field, a reduction in corrosion of up to 40% was observed after switch to nitrate treatment. The present study show that nitrate injection may provide a stable long-term inhibition of SRB in sea water injection systems, and that corrosion may be significantly reduced when compared to traditional biocide treatment.

Desulfotignum toluenicum sp. nov., a novel toluene-degrading, sulphate-reducing bacterium isolated from an oil-reservoir model column.

A Gram-negative, sulphate-reducing bacterium (strain H3(T)) was isolated from an oil-reservoir model column. The new isolate was able to oxidize toluene coupled to hydrogen sulphide production. For growth, the optimum salt concentration was 1.5 % (w/v), the optimum pH was 7.2 and the optimum temperature was 34 degrees C. The cells were straight to slightly curved rods, 0.6-1.0 microm in diameter and 1.4-2.5 microm in length. The predominant fatty acids were C(16 : 0), C(16 : 1)omega7c and C(17 : 0) cyclo, and the cells also contained dimethylacetals. Cloning and sequencing of a 1505 bp long fragment of the 16S rRNA gene showed that strain H3(T) is a member of the Deltaproteobacteria and is related closely to Desulfotignum balticum DSM 7044(T). The G+C content of the DNA was 52.0 mol% and the DNA-DNA similarity to D. balticum DSM 7044(T) was 56.1 %. Based on differences in DNA sequence and the unique property of toluene degradation, it is proposed that strain H3(T) should be designated a member of a novel species within the genus Desulfotignum, for which the name Desulfotignum toluenicum sp. nov. is proposed. The type strain is H3(T) (=DSM 18732(T)=ATCC BAA-1460(T)).

Microbial community diversity in seafloor basalt from the Arctic spreading ridges.

Microbial communities inhabiting recent (< or =1 million years old; Ma) seafloor basalts from the Arctic spreading ridges were analyzed using traditional enrichment culturing methods in combination with culture-independent molecular phylogenetic techniques. Fragments of 16S rDNA were amplified from the basalt samples by polymerase chain reaction, and fingerprints of the bacterial and archaeal communities were generated using denaturing gradient gel electrophoresis. This analysis indicates a substantial degree of complexity in the samples studied, showing 20-40 dominating bands per profile for the bacterial assemblages. For the archaeal assemblages, a much lower number of bands (6-12) were detected. The phylogenetic affiliations of the predominant electrophoretic bands were inferred by performing a comparative 16S rRNA gene sequence analysis. Sequences obtained from basalts affiliated with eight main phylogenetic groups of Bacteria, but were limited to only one group of the Archaea. The most frequently retrieved bacterial sequences affiliated with the gamma-proteobacteria, alpha-proteobacteria, Chloroflexi, Firmicutes, and Actinobacteria. The archaeal sequences were restricted to the marine Group 1: Crenarchaeota. Our results indicate that the basalt harbors a distinctive microbial community, as the majority of the sequences differed from those retrieved from the surrounding seawater as well as from sequences previously reported from seawater and deep-sea sediments. Most of the sequences did not match precisely any sequences in the database, indicating that the indigenous Arctic ridge basalt microbial community is yet uncharacterized. Results from enrichment cultures showed that autolithotrophic methanogens and iron reducing bacteria were present in the seafloor basalts. We suggest that microbial catalyzed cycling of iron may be important in low-temperature alteration of ocean crust basalt. The phylogenetic and physiological diversity of the seafloor basalt microorganisms differed from those previously reported from deep-sea hydrothermal systems.