Large-scale nutrient and carbon dynamics along the river-estuary-ocean continuum.

Nutrient and carbon dynamics within the river-estuary-coastal water systems are key processes in understanding the flux of matter from the terrestrial environment to the ocean. Here, we analysed those dynamics by following a sampling approach based on the travel time of water and an advanced calculation of nutrient fluxes in the tidal part. We started with a nearly Lagrangian sampling of the river (River Elbe, Germany; 580 km within 8 days). After a subsequent investigation of the estuary, we followed the plume of the river by raster sampling the German Bight (North Sea) using three ships simultaneously. In the river, we detected intensive longitudinal growth of phytoplankton connected with high oxygen saturation and pH values and an undersaturation of CO2, whereas concentrations of dissolved nutrients declined. In the estuary, the Elbe shifted from an autotrophic to a heterotrophic system: Phytoplankton died off upstream of the salinity gradient, causing minima in oxygen saturation and pH, supersaturation of CO2, and a release of nutrients. In the shelf region, phytoplankton and nutrient concentrations were low, oxygen was close to saturation, and pH was within a typical marine range. Over all sections, oxygen saturation was positively related to pH and negatively to pCO2. Corresponding to the significant particulated nutrient flux via phytoplankton, flux rates of dissolved nutrients from river into estuary were low and determined by depleted concentrations. In contrast, fluxes from the estuary to the coastal waters were higher and the pattern was determined by tidal current. Overall, the approach is appropriate to better understand land-ocean fluxes, particularly to illuminate the importance of these fluxes under different seasonal and hydrological conditions, including flood and drought events.

Methylomonas albis sp. nov. and Methylomonas fluvii sp. nov.: Two cold-adapted methanotrophs from the river Elbe and emended description of the species Methylovulum psychrotolerans.

Three strains of methanotrophic bacteria (EbAT, EbBT and Eb1) were isolated from the River Elbe, Germany. These Gram-negative, rod-shaped or coccoid cells contain intracytoplasmic membranes perpendicular to the cell surface. Colonies and liquid cultures appeared bright-pink. The major cellular fatty acids were 12:0 and 14:0, in addition in Eb1 the FA 16:1ω5t was also dominant. Methane and methanol were utilized as sole carbon sources by EbBT and Eb1, while EbAT could not use methanol. All strains oxidize methane using the particulate methane monooxygenase. Both strains contain an additional soluble methane monooxygenase. The strains grew optimally at 15-25 °C and at pH 6 and 8. Based on 16S rRNA gene analysis recovered from the full genome, the phylogenetic position of EbAT is robustly outside any species clade with its closest relatives being Methylomonas sp. MK1 (98.24%) and Methylomonas sp. 11b (98.11%). Its closest type strain is Methylomonas methanica NCIMB11130 (97.91%). The 16S rRNA genes of EbBT are highly similar to Methylomonas methanica strains with Methylomonas methanica R-45371 as the closest relative (99.87% sequence identity). However, average nucleotide identity (ANI) and digital DNA-DNA-hybridization (dDDH) values reveal it as distinct species. The DNA G + C contents were 51.07 mol% and 51.5 mol% for EbAT and EbBT, and 50.7 mol% for Eb1, respectively. Strains EbAT and EbBT are representing two novel species within the genus Methylomonas. For strain EbAT we propose the name Methylomonas albis sp. nov (LMG 29958, JCM 32282) and for EbBT, we propose the name Methylomonas fluvii sp. nov (LMG 29959, JCM 32283). Eco-physiological descriptions for both strains are provided. Strain Eb1 (LMG 30323, JCM 32281) is a member of the species Methylovulum psychrotolerans. This genus is so far only represented by two isolates but Eb1 is the first isolate from a temperate environment; so, an emended description of the species is given.

Methane release from sediment seeps to the atmosphere is counteracted by highly active Methylococcaceae in the water column of deep oligotrophic Lake Constance.

Methane emissions from freshwater environments contribute substantially to global warming but are under strong control of aerobic methane-oxidizing bacteria. Recently discovered methane seeps (pockmarks) in freshwater lake sediments have the potential to bypass this control by their strong outgassing activity. Whether this is counteracted by pelagic methanotrophs is not well understood yet. We used a (3)H-CH4-radiotracer technique and pmoA-based molecular approaches to assess the activity, abundance and community structure of pelagic methanotrophs above active pockmarks in deep oligotrophic Lake Constance. Above profundal pockmarks, methane oxidation rates (up to 458 nmol CH4 l(-1) d(-1)) exceeded those of the surrounding water column by two orders of magnitude and coincided with maximum methanotroph abundances of 0.6% of the microbial community. Phylogenetic analysis indicated a dominance of members of the Methylococcaceae in the water column of both, pockmark and reference sites, with most of the retrieved sequences being associated with a water-column specific clade. Communities at pockmark and reference locations also differed in parts, which was likely caused by entrainment of sediment-hosted methanotrophs at pockmark sites. Our results show that the release of seep-derived methane to the atmosphere is counteracted by a distinct methanotrophic community with a pronounced activity throughout bottom waters.

Methane turnover and methanotrophic communities in arctic aquatic ecosystems of the Lena Delta, Northeast Siberia.

Large amounts of organic carbon are stored in Arctic permafrost environments, and microbial activity can potentially mineralize this carbon into methane, a potent greenhouse gas. In this study, we assessed the methane budget, the bacterial methane oxidation (MOX) and the underlying environmental controls of arctic lake systems, which represent substantial sources of methane. Five lake systems located on Samoylov Island (Lena Delta, Siberia) and the connected river sites were analyzed using radiotracers to estimate the MOX rates, and molecular biology methods to characterize the abundance and the community composition of methane-oxidizing bacteria (MOB). In contrast to the river, the lake systems had high variation in the methane concentrations, the abundance and composition of the MOB communities, and consequently, the MOX rates. The highest methane concentrations and the highest MOX rates were detected in the lake outlets and in a lake complex in a flood plain area. Though, in all aquatic systems, we detected both, Type I and II MOB, in lake systems, we observed a higher diversity including MOB, typical of the soil environments. The inoculation of soil MOB into the aquatic systems, resulting from permafrost thawing, might be an additional factor controlling the MOB community composition and potentially methanotrophic capacity.

Methylosoma difficile gen. nov., sp. nov., a novel methanotroph enriched by gradient cultivation from littoral sediment of Lake Constance.

A novel methanotroph, strain LC 2(T), was isolated from the littoral sediment of Lake Constance by enrichment in opposing gradients of methane and oxygen, followed by traditional isolation methods. Strain LC 2(T) grows on methane or methanol as its sole carbon and energy source. It is a Gram-negative, non-motile, pale-pink-coloured methanotroph showing typical intracytoplasmic membranes arranged in stacks. Cells are coccoid, elliptical or rod-shaped and occur often in pairs. Strain LC 2(T) grows at low oxygen concentrations and in counter-gradients of methane and oxygen. It can grow on medium free of bound nitrogen, possesses the nifH gene and fixes atmospheric nitrogen at low oxygen pressure. It grows at neutral pH and at temperatures between 10 and 30 degrees C. Phylogenetically, it is most closely related to the genus Methylobacter, with the type strains of Methylobacter tundripaludum and Methylobacter psychrophilus showing 94 and 93.4 % 16S rRNA gene sequence similarity, respectively. Furthermore, the pmoA gene sequence of strain LC 2(T) is most closely related to pmoA gene sequences of Methylobacter strains (92 % similar to Methylobacter sp. LW 12 by deduced amino acid sequence identity). The DNA G+C content is 49.9 mol% and the major cellular fatty acid is 16 : 1omega7c (60 %). Strain LC 2(T) (=JCM 14076(T)=DSM 18750(T)) is described as the type strain of a novel species within a new genus, Methylosoma difficile gen. nov., sp. nov.

Cultivation of methanotrophic bacteria in opposing gradients of methane and oxygen.

In sediments, methane-oxidizing bacteria live in opposing gradients of methane and oxygen. In such a gradient system, the fluxes of methane and oxygen are controlled by diffusion and consumption rates, and the rate-limiting substrate is maintained at a minimum concentration at the layer of consumption. Opposing gradients of methane and oxygen were mimicked in a specific cultivation set-up in which growth of methanotrophic bacteria occurred as a sharp band at either c. 5 or 20 mm below the air-exposed end. Two new strains of methanotrophic bacteria were isolated with this system. One isolate, strain LC 1, belonged to the Methylomonas genus (type I methantroph) and contained soluble methane mono-oxygenase. Another isolate, strain LC 2, was related to the Methylobacter group (type I methantroph), as determined by 16S rRNA gene and pmoA sequence similarities. However, the partial pmoA sequence was only 86% related to cultured Methylobacter species. This strain accumulated significant amounts of formaldehyde in conventional cultivation with methane and oxygen, which may explain why it is preferentially enriched in a gradient cultivation system.

Preferential cultivation of type II methanotrophic bacteria from littoral sediments (Lake Constance).

Most widely used medium for cultivation of methanotrophic bacteria from various environments is that proposed in 1970 by Whittenbury. In order to adapt and optimize medium for culturing of methanotrophs from freshwater sediment, media with varying concentrations of substrates, phosphate, nitrate, and other mineral salts were used to enumerate methanotrophs by the most probable number method. High concentrations (>1 mM) of magnesium and sulfate, and high concentrations of nitrate (>500 microM) significantly reduced the number of cultured methanotrophs, whereas phosphate in the range of 15-1500 microM had no influence. Also oxygen and carbon dioxide influenced the culturing efficiency, with an optimal mixing ratio of 17% O(2) and 3% CO(2); the mixing ratio of methane (6-32%) had no effect. A clone library of pmoA genes amplified by PCR from DNA extracted from sediment revealed the presence of both type I and type II methanotrophs. Nonetheless, the cultivation of methanotrophs, also with the improved medium, clearly favored growth of type II methanotrophs of the Methylosinus/Methylocystis group. Although significantly more methanotrophs could be cultured with the modified medium, their diversity did not mirror the diversity of methanotrophs in the sediment sample detected by molecular biology method.