Nicolas Tsesmetzis

Complementary Microorganisms in Highly Corrosive Biofilms from an Offshore Oil Production Facility

ABSTRACT Offshore oil production facilities are frequently victims of internal piping corrosion, potentially leading to human and environmental risks and significant economic losses. Microbially influenced corrosion (MIC) is believed to be an important factor in this major problem for the petroleum industry. However, knowledge of the microbial communities and metabolic processes leading to corrosion is still limited. Therefore, the microbial communities from three anaerobic biofilms recovered from the inside of a steel pipe exhibiting high corrosion rates, iron oxide deposits, and substantial amounts of sulfur, which are characteristic of MIC, were analyzed in detail. Bacterial and archaeal community structures were investigated by automated ribosomal intergenic spacer analysis, multigenic (16S rRNA and functional genes) high-throughput Illumina MiSeq sequencing, and quantitative PCR analysis. The microbial community analysis indicated that bacteria, particularly Desulfovibrio species, dominated the biofilm microbial communities. However, other bacteria, such as Pelobacter, Pseudomonas, and Geotoga, as well as various methanogenic archaea, previously detected in oil facilities were also detected. The microbial taxa and functional genes identified suggested that the biofilm communities harbored the potential for a number of different but complementary metabolic processes and that MIC in oil facilities likely involves a range of microbial metabolisms such as sulfate, iron, and elemental sulfur reduction. Furthermore, extreme corrosion leading to leakage and exposure of the biofilms to the external environment modify the microbial community structure by promoting the growth of aerobic hydrocarbon-degrading organisms.

Microbial redox processes in deep subsurface environments and the potential application of (per)chlorate in oil reservoirs

The ability of microorganisms to thrive under oxygen-free conditions in subsurface environments relies on the enzymatic reduction of oxidized elements, such as sulfate, ferric iron, or CO2, coupled to the oxidation of inorganic or organic compounds. A broad phylogenetic and functional diversity of microorganisms from subsurface environments has been described using isolation-based and advanced molecular ecological techniques. The physiological groups reviewed here comprise iron-, manganese-, and nitrate-reducing microorganisms. In the context of recent findings also the potential of chlorate and perchlorate [jointly termed (per)chlorate] reduction in oil reservoirs will be discussed. Special attention is given to elevated temperatures that are predominant in the deep subsurface. Microbial reduction of (per)chlorate is a thermodynamically favorable redox process, also at high temperature. However, knowledge about (per)chlorate reduction at elevated temperatures is still scarce and restricted to members of the Firmicutes and the archaeon Archaeoglobus fulgidus. By analyzing the diversity and phylogenetic distribution of functional genes in (meta)genome databases and combining this knowledge with extrapolations to earlier-made physiological observations we speculate on the potential of (per)chlorate reduction in the subsurface and more precisely oil fields. In addition, the application of (per)chlorate for bioremediation, souring control, and microbial enhanced oil recovery are addressed.

IMG/VR: A database of cultured and uncultured DNA viruses and retroviruses

Viruses represent the most abundant life forms on the planet. Recent experimental and computational improvements have led to a dramatic increase in the number of viral genome sequences identified primarily from metagenomic samples. As a result of the expanding catalog of metagenomic viral sequences, there exists a need for a comprehensive computational platform integrating all these sequences with associated metadata and analytical tools. Here we present IMG/VR (https://img.jgi.doe.gov/vr/), the largest publicly available database of 3908 isolate reference DNA viruses with 264 413 computationally identified viral contigs from >6000 ecologically diverse metagenomic samples. Approximately half of the viral contigs are grouped into genetically distinct quasi-species clusters. Microbial hosts are predicted for 20 000 viral sequences, revealing nine microbial phyla previously unreported to be infected by viruses. Viral sequences can be queried using a variety of associated metadata, including habitat type and geographic location of the samples, or taxonomic classification according to hallmark viral genes. IMG/VR has a user-friendly interface that allows users to interrogate all integrated data and interact by comparing with external sequences, thus serving as an essential resource in the viral genomics community.

Succession in the petroleum reservoir microbiome through an oil field production lifecycle

Subsurface petroleum reservoirs are an important component of the deep biosphere where indigenous microorganisms live under extreme conditions and in isolation from the Earth’s surface for millions of years. However, unlike the bulk of the deep biosphere, the petroleum reservoir deep biosphere is subject to extreme anthropogenic perturbation, with the introduction of new electron acceptors, donors and exogenous microbes during oil exploration and production. Despite the fundamental and practical significance of this perturbation, there has never been a systematic evaluation of the ecological changes that occur over the production lifetime of an active offshore petroleum production system. Analysis of the entire Halfdan oil field in the North Sea (32 producing wells in production for 1–15 years) using quantitative PCR, multigenic sequencing, comparative metagenomic and genomic bins reconstruction revealed systematic shifts in microbial community composition and metabolic potential, as well as changing ecological strategies in response to anthropogenic perturbation of the oil field ecosystem, related to length of time in production. The microbial communities were initially dominated by slow growing anaerobes such as members of the Thermotogales and Clostridiales adapted to living on hydrocarbons and complex refractory organic matter. However, as seawater and nitrate injection (used for secondary oil production) delivered oxidants, the microbial community composition progressively changed to fast growing opportunists such as members of the Deferribacteres, Delta-, Epsilon- and Gammaproteobacteria, with energetically more favorable metabolism (for example, nitrate reduction, H2S, sulfide and sulfur oxidation). This perturbation has profound consequences for understanding the microbial ecology of the system and is of considerable practical importance as it promotes detrimental processes such as reservoir souring and metal corrosion. These findings provide a new conceptual framework for understanding the petroleum reservoir biosphere and have consequences for developing strategies to manage microbiological problems in the oil industry.

Perchlorate and chlorate reduction by the Crenarchaeon Aeropyrum pernix and two thermophilic Firmicutes

This study reports the ability of one hyperthermophilic and two thermophilic microorganisms to grow anaerobically by the reduction of chlorate and perchlorate. Physiological, genomic and proteome analyses suggest that the Crenarchaeon Aeropyrum pernix reduces perchlorate with a periplasmic enzyme related to nitrate reductases, but that it lacks a functional chlorite-disproportionating enzyme (Cld) to complete the pathway. Aeropyrum pernix, previously described as a strictly aerobic microorganism, seems to rely on the chemical reactivity of reduced sulfur compounds with chlorite, a mechanism previously reported for perchlorate-reducing Archaeoglobus fulgidus. The chemical oxidation of thiosulfate (in excessive amounts present in the medium) and the reduction of chlorite result in the release of sulfate and chloride, which are the products of a biotic-abiotic perchlorate reduction pathway in Ae. pernix. The apparent absence of Cld in two other perchlorate-reducing microorganisms, Carboxydothermus hydrogenoformans and Moorella glycerini strain NMP, and their dependence on sulfide for perchlorate reduction is consistent with the observations made on Ar. fulgidus. Our findings suggest that microbial perchlorate reduction at high temperature differs notably from the physiology of perchlorate- and chlorate-reducing mesophiles and that it is characterized by the lack of a chlorite dismutase and is enabled by a combination of biotic and abiotic reactions.

Microbial and isotopic evidence for methane cycling in hydrocarbon-containing groundwater from the Pennsylvania region

The Pennsylvania region hosts numerous oil and gas reservoirs and the presence of hydrocarbons in groundwater has been locally observed. However, these methane-containing freshwater ecosystems remain poorly explored despite their potential importance in the carbon cycle. Methane isotope analysis and analysis of low molecular weight hydrocarbon gases from 18 water wells indicated that active methane cycling may be occurring in methane-containing groundwater from the Pennsylvania region. Consistent with this observation, multigenic qPCR and gene sequencing (16S rRNA genes, mcrA, and pmoA genes) indicated abundant populations of methanogens, ANME-2d (average of 1.54 × 104 mcrA gene per milliliter of water) and bacteria associated with methane oxidation (NC10, aerobic methanotrophs, methylotrophs; average of 2.52 × 103 pmoA gene per milliliter of water). Methane cycling therefore likely represents an important process in these hydrocarbon-containing aquifers. The microbial taxa and functional genes identified and geochemical data suggested that (i) methane present is at least in part due to methanogens identified in situ; (ii) Potential for aerobic and anaerobic methane oxidation is important in groundwater with the presence of lineages associated with both anaerobic an aerobic methanotrophy; (iii) the dominant methane oxidation process (aerobic or anaerobic) can vary according to prevailing conditions (oxic or anoxic) in the aquifers; (iv) the methane cycle is closely associated with the nitrogen cycle in groundwater methane seeps with methane and/or methanol oxidation coupled to denitrification or nitrate and nitrite reduction.

Preservation of ancestral Cretaceous microflora recovered from a hypersaline oil reservoir

Microbiology of a hypersaline oil reservoir located in Central Africa was investigated with molecular and culture methods applied to preserved core samples. Here we show that the community structure was partially acquired during sedimentation, as many prokaryotic 16S rRNA gene sequences retrieved from the extracted DNA are phylogenetically related to actual Archaea inhabiting surface evaporitic environments, similar to the Cretaceous sediment paleoenvironment. Results are discussed in term of microorganisms and/or DNA preservation in such hypersaline and Mg-rich solutions. High salt concentrations together with anaerobic conditions could have preserved microbial/molecular diversity originating from the ancient sediment basin wherein organic matter was deposited.

MIxS-HCR: A MIxS extension defining a minimal information standard for sequence data from environments pertaining to hydrocarbon resources

Here we introduce a MIxS extension to facilitate the recording and cataloguing of metadata from samples related to hydrocarbon resources. The proposed MIxS-HCR package incorporates the core features of the MIxS standard for marker gene (MIMARKS) and metagenomic (MIMS) sequences along with a hydrocarbon resources customized environmental package. Adoption of the MIxS-HCR standard will enable the comparison and better contextualization of investigations related to hydrocarbon rich environments. The insights from such standardized way of reporting could be highly beneficial for the successful development and optimization of hydrocarbon recovery processes and management of microbiological issues in petroleum production systems.

Comparative metagenomics of hydrocarbon and methane seeps of the Gulf of Mexico

Oil and gas percolate profusely through the sediments of the Gulf of Mexico, leading to numerous seeps at the seafloor, where complex microbial, and sometimes animal communities flourish. Sediments from three areas (two cold seeps with contrasting hydrocarbon composition and a site outside any area of active seepage) of the Gulf of Mexico were investigated and compared. Consistent with the existence of a seep microbiome, a distinct microbial community was observed in seep areas compared to sediment from outside areas of active seepage. The microbial community from sediments without any influence from hydrocarbon seepage was characterized by Planctomycetes and the metabolic potential was consistent with detrital marine snow degradation. By contrast, in seep samples with methane as the principal hydrocarbon, methane oxidation by abundant members of ANME-1 was likely the predominant process. Seep samples characterized by fluids containing both methane and complex hydrocarbons, were characterized by abundant Chloroflexi (Anaerolinaceae) and deltaproteobacterial lineages and exhibited potential for complex hydrocarbon degradation. These different metabolic capacities suggested that microorganisms in cold seeps can potentially rely on other processes beyond methane oxidation and that the hydrocarbon composition of the seep fluids may be a critical factor structuring the seafloor microbial community composition and function.

Damage to offshore production facilities by corrosive microbial biofilms

In offshore production facilities, large amounts of deaerated seawater are continuously injected to maintain pressure in oil reservoirs and equivalent volumes of fluids, composed of an oil/gas, and water mixture are produced. This process, brewing billions of liters of biphasic fluids particularly rich in microorganisms, goes through complex steel pipeline networks that are particularly prone to biofilm formation. Consequently, offshore facilities are frequently victims of severe microbiologically influenced corrosion. Understanding of microbiologically influenced corrosion is constantly growing. In the laboratory, the inventory of potentially corrosive microorganisms is increasing and microbial biochemical and bioelectrical processes are now recognized to be involved in corrosion. However, understanding of corrosive multispecies biofilms and the complex metabolic processes associated with corrosion remains a considerable challenge as simple laboratory biofilms comprising pure or defined mixed cultures poorly represent the complexity of in situ biofilms. Complementary, antagonistic, and parallel microbial pathways occur within the complex microbial and inorganic matrix of the biofilms which can lead to high corrosion rates. This mini-review explores models of microbiologically influenced corrosion and places them in the context of the multispecies biofilms observed in situ. Consequences of mitigation strategies on biofilm corrosiveness and dispersal are also discussed.