Richard Seifert

Highly isotopically depleted isoprenoids: molecular markers for ancient methane venting

Abstract We propose that organic compounds found in a Miocene limestone from Marmorito (Northern Italy) are source markers for organic matter present in ancient methane vent systems (cold seeps). The limestone contains high concentrations of the tail-to-tail linked, acyclic C20 isoprenoid 2,6,11,15-tetramethylhexadecane (crocetane), a C25 homolog 2,6,10,15,19-pentamethylicosane (PME), and a distinctive glycerol ether lipid containing 3,7,11,15-tetramethylhexadecyl (phytanyl-) moieties. The chemical structures of these biomarkers indicate a common origin from archaea. Their extremely 13C-depleted isotope compositions (δ13C ≈ −108 to −115.6‰ PDB) suggest that the respective archaea have directly or indirectly introduced isotopically depleted, methane-derived carbon into their biomass. We postulate that a second major cluster of biomarkers showing heavier isotope values (δ13C ≈ −88‰) is derived from sulfate-reducing bacteria (SRB). The observed biomarkers sustain the idea that methanogenic bacteria, in a syntrophic community with SRB, are responsible for the anaerobic oxidation of methane in marine sediments. Marmorito may thus represent a conceivable ancient scenario for methane consumption performed by a defined, two-membered bacterial consortium: (1) archaea that perform reversed methanogenesis by oxidizing methane and producing CO2 and H2; and (2) SRB that consume the resulting H2. Furthermore, the respective organic molecules are, unlike other compounds, tightly bound to the crystalline carbonate phase. The Marmorito carbonates can thus be regarded as “cold seep microbialites” rather than mere “authigenic” carbonates.

Hydrogen is an energy source for hydrothermal vent symbioses

The discovery of deep-sea hydrothermal vents in 1977 revolutionized our understanding of the energy sources that fuel primary productivity on Earth. Hydrothermal vent ecosystems are dominated by animals that live in symbiosis with chemosynthetic bacteria. So far, only two energy sources have been shown to power chemosynthetic symbioses: reduced sulphur compounds and methane. Using metagenome sequencing, single-gene fluorescence in situ hybridization, immunohistochemistry, shipboard incubations and in situ mass spectrometry, we show here that the symbionts of the hydrothermal vent mussel Bathymodiolus from the Mid-Atlantic Ridge use hydrogen to power primary production. In addition, we show that the symbionts of Bathymodiolus mussels from Pacific vents have hupL, the key gene for hydrogen oxidation. Furthermore, the symbionts of other vent animals such as the tubeworm Riftia pachyptila and the shrimp Rimicaris exoculata also have hupL. We propose that the ability to use hydrogen as an energy source is widespread in hydrothermal vent symbioses, particularly at sites where hydrogen is abundant.

Methane distribution in European tidal estuaries

Methane concentrations have been measured along salinity profilesin nine tidal estuaries in Europe (Elbe, Ems, Thames, Rhine,Scheldt, Loire, Gironde, Douro and Sado). The Rhine, Scheldt andGironde estuaries have been studied seasonally. A number ofdifferent methodologies have been used and they yieldedconsistent results. Surface water concentrations ranged from0.002 to 3.6 μM, corresponding to saturation ratios of 0.7 to1580 with a median of 25. Methane concentrations in thefresh-water end-members varied from 0.01 to 1.4 μM. Methaneconcentrations in the marine end-members were close to saturationoffshore and on the order of 0.1 μM in estuarine plumes. Methaneversus salinity profiles in river-dominated, stratified estuaries(Rhine and Douro) appeared rather erratic whereas those in thewell mixed, long-residence time estuaries (Elbe, Ems, Thames,Scheldt, Loire, Gironde and Sado) revealed consistent trends. Inthese systems dissolved methane initially decreases withincreasing salinity, then increases to a maximum at intermediateto high salinities before decreasing again going offshore. Tidalflats and creeks were identified as a methane source to estuarinewaters. The global estuarine flux of methane to the atmospherehas been calculated by combining the median water-air methanegradient (68.2 nmol dm−3) with a global area weighted transfercoefficient and the global area of estuaries. Estuaries emit 1.1to 3.0 Tg CH4 yr−1, which is less than 9% of the global marinemethane emission.

Unexpected occurrence of hopanoids at gas seeps in the Black Sea

Abstract High concentrations of free C32 bis-homohopanoic acids (up to 433 μg/g dry wt) occur in microbial mats at methane seeps in anoxic Black Sea waters. These compounds show a strong preference for the ‘geological’ 17α(H),21β(H)- over the ‘biological’ 17β(H),21β(H)-configuration (αβ/ββ ratios up to 30.7) and indicate the potential formation of αβ-hopanoids in modern environments. Strong 13C-depletions (δ13C as low as −78.4‰ PDB) indicate an in situ generation of these hopanoids by biota involved in the anaerobic cycling of methane carbon. The inferred presence of hopanoids indigenous to a permanently anoxic marine environment is significant because these lipids are not known to occur in strictly anaerobic bacteria.

Subsurface microbial methanotrophic mats in the Black Sea.

ABSTRACT A nodule-shaped microbial mat was found subsurface in sediments of a gas seep in the anoxic Black Sea. This mat was dominated by ANME-1 archaea and consumed methane and sulfate simultaneously. We propose that such subsurface mats represent the initial stage of previously investigated microbial reefs.

Young volcanism and related hydrothermal activity at 5°S on the slow‐spreading southern Mid‐Atlantic Ridge

The effect of volcanic activity on submarine hydrothermal systems has been well documented along fast- and intermediate-spreading centers but not from slow-spreading ridges. Indeed, volcanic eruptions are expected to be rare on slow-spreading axes. Here we report the presence of hydrothermal venting associated with extremely fresh lava flows at an elevated, apparently magmatically robust segment center on the slow-spreading southern Mid-Atlantic Ridge near 5°S. Three high-temperature vent fields have been recognized so far over a strike length of less than 2 km with two fields venting phase-separated, vapor-type fluids. Exit temperatures at one of the fields reach up to 407°C, at conditions of the critical point of seawater, the highest temperatures ever recorded from the seafloor. Fluid and vent field characteristics show a large variability between the vent fields, a variation that is not expected within such a limited area. We conclude from mineralogical investigations of hydrothermal precipitates that vent-fluid compositions have evolved recently from relatively oxidizing to more reducing conditions, a shift that could also be related to renewed magmatic activity in the area. Current high exit temperatures, reducing conditions, low silica contents, and high hydrogen contents in the fluids of two vent sites are consistent with a shallow magmatic source, probably related to a young volcanic eruption event nearby, in which basaltic magma is actively crystallizing. This is the first reported evidence for direct magmatic-hydrothermal interaction on a slow-spreading mid-ocean ridge.

Metabolites of xenobiotica and mineral oil constituents linked to macromolecular organic matter in polluted environments

Abstract The type of association between pollutants and humic substances of soils, sediments and river waters has been investigated. Metabolites, which can arise from the microbiological degradation of polyaromatic hydrocarbons (PAH) and polychlorinated biphenyls (PCB), were cleaved from the macromolecular matrix by selective chemical degradation techniques (OH − , BCl 3 , Rh/H 2 ). Hydrolysis reactions performed with Na 18 OH proved that some metabolites of pollutants form stable ester bonds by condensation processes with functional groups of humic substances, a phenomenon which has major implications for transport, toxicity and bioavailability of xenobiotica.

The use of 13C-labelled polycyclic aromatic hydrocarbons for the analysis of their transformation in soil.

The formation of non-extractable residues during biodegradation and humification processes in soils and sediments represents a major sink for organic contaminants. The mode of incorporation of polycyclic aromatic hydrocarbons (PAH) and their metabolites into macromolecular organic matter during microbial degradation was studied applying 13C-labelled compounds. Mineralisation rates were determined by measuring the 13CO2 production. An incorporation of 13C-PAH-fragments into humic material could be traced by isotopic analysis of the bulk organic matter. Furthermore, selective chemical degradation reactions were performed to analyse the precise chemical structure of covalently bound 13C-labelled PAH fragments in soil humic substances. Structural assignments by GC-MS combined with isotope measurements on the bulk organic carbon and at the molecular level (Isotope Ratio Monitoring-GC-MS) provided useful information on the fate of xenobiotics within the soil.

Organic pollutants associated with macromolecular soil organic matter: Mode of binding☆

A study of ether-linked moieties in macromolecular bound residues of polycyclic aromatic hydrocarbons (PAH) generated in bioremediation experiments was performed using high temperature hydrolysis degradation with subsequent analysis of the products by GC-MS. This hydrolysis reaction was specifically designed to cleave ether bonds including relatively stable diarylether structures. Among the reaction products, aromatic alcohols representing typical microbiologically derived metabolites of PAH were found in addition to natural compounds. Thus, parts of the bound residues appeared to be linked within the macromolecular material by ether bonds. Model experiments with an oxidoreductase enzyme and aromatic alcohols indicate the formation of these ether bonds to be an enzyme-catalysed process.

In Vitro Study of Lipid Biosynthesis in an Anaerobically Methane-Oxidizing Microbial Mat

ABSTRACT The anaerobic oxidation of methane (AOM) is a key process in the global methane cycle, and the majority of methane formed in marine sediments is oxidized in this way. Here we present results of an in vitro 13CH4 labeling study (δ13CH4, ∼5,400‰) in which microorganisms that perform AOM in a microbial mat from the Black Sea were used. During 316 days of incubation, the 13C uptake into the mat biomass increased steadily, and there were remarkable differences for individual bacterial and archaeal lipid compounds. The greatest shifts were observed for bacterial fatty acids (e.g., hexadec-11-enoic acid [16:1Δ11]; difference between the δ13C at the start and the end of the experiment [Δδ13Cstart-end], ∼160‰). In contrast, bacterial glycerol diethers exhibited only slight changes in δ13C (Δδ13Cstart-end, ∼10‰). Differences were also found for individual archaeal lipids. Relatively high uptake of methane-derived carbon was observed for archaeol (Δδ13Cstart-end, ∼25‰), a monounsaturated archaeol, and biphytanes, whereas for sn-2-hydroxyarchaeol there was considerably less change in the δ13C (Δδ13Cstart-end, ∼2‰). Moreover, an increase in the uptake of 13C for compounds with a higher number of double bonds within a suite of polyunsaturated 2,6,10,15,19-pentamethyleicosenes indicated that in methanotrophic archaea there is a biosynthetic pathway similar to that proposed for methanogenic archaea. The presence of group-specific biomarkers (for ANME-1 and ANME-2 associations) and the observation that there were differences in 13C uptake into specific lipid compounds confirmed that multiple phylogenetically distinct microorganisms participate to various extents in biomass formation linked to AOM. However, the greater 13C uptake into the lipids of the sulfate-reducing bacteria (SRB) than into the lipids of archaea supports the hypothesis that there is autotrophic growth of SRB on small methane-derived carbon compounds supplied by the methane oxidizers.