Simultaneous sulfate and nitrate reduction in coastal sediments.

The oscillating redox conditions that characterize coastal sandy sediments foster microbial communities capable of respiring oxygen and nitrate simultaneously, thereby increasing the potential for organic matter remineralization, nitrogen (N)-loss and emissions of the greenhouse gas nitrous oxide. It is unknown to what extent these conditions also lead to overlaps between dissimilatory nitrate and sulfate respiration. Here, we show that sulfate and nitrate respiration co-occur in the surface sediments of an intertidal sand flat. Furthermore, we found strong correlations between dissimilatory nitrite reduction to ammonium (DNRA) and sulfate reduction rates. Until now, the nitrogen and sulfur cycles were assumed to be mainly linked in marine sediments by the activity of nitrate-reducing sulfide oxidisers. However, transcriptomic analyses revealed that the functional marker gene for DNRA (nrfA) was more associated with microorganisms known to reduce sulfate rather than oxidise sulfide. Our results suggest that when nitrate is supplied to the sediment community upon tidal inundation, part of the sulfate reducing community may switch respiratory strategy to DNRA. Therefore increases in sulfate reduction rate in-situ may result in enhanced DNRA and reduced denitrification rates. Intriguingly, the shift from denitrification to DNRA did not influence the amount of N2O produced by the denitrifying community. Our results imply that microorganisms classically considered as sulfate reducers control the potential for DNRA within coastal sediments when redox conditions oscillate and therefore retain ammonium that would otherwise be removed by denitrification, exacerbating eutrophication.

Insights into the Fundamental Physiology of the Uncultured Fe-Oxidizing Bacterium Leptothrix ochracea.

Leptothrix ochracea is known for producing large volumes of iron oxyhydroxide sheaths that alter wetland biogeochemistry. For over a century, these delicate structures have fascinated microbiologists and geoscientists. Because L. ochracea still resists long-term in vitro culture, the debate regarding its metabolic classification dates back to 1885. We developed a novel culturing technique for L. ochracea using in situ natural waters and coupled this with single-cell genomics and nanoscale secondary-ion mass spectrophotometry (nanoSIMS) to probe L. ochracea's physiology. In microslide cultures L. ochracea doubled every 5.7 h and had an absolute growth requirement for ferrous iron, the genomic capacity for iron oxidation, and a branched electron transport chain with cytochromes putatively involved in lithotrophic iron oxidation. Additionally, its genome encoded several electron transport chain proteins, including a molybdopterin alternative complex III (ACIII), a cytochrome bd oxidase reductase, and several terminal oxidase genes. L. ochracea contained two key autotrophic proteins in the Calvin-Benson-Bassham cycle, a form II ribulose bisphosphate carboxylase, and a phosphoribulose kinase. L. ochracea also assimilated bicarbonate, although calculations suggest that bicarbonate assimilation is a small fraction of its total carbon assimilation. Finally, L. ochracea's fundamental physiology is a hybrid of those of the chemolithotrophic Gallionella-type iron-oxidizing bacteria and the sheathed, heterotrophic filamentous metal-oxidizing bacteria of the Leptothrix-Sphaerotilus genera. This allows L. ochracea to inhabit a unique niche within the neutrophilic iron seeps.IMPORTANCELeptothrix ochracea was one of three groups of organisms that Sergei Winogradsky used in the 1880s to develop his hypothesis on chemolithotrophy. L. ochracea continues to resist cultivation and appears to have an absolute requirement for organic-rich waters, suggesting that its true physiology remains unknown. Further, L. ochracea is an ecological engineer; a few L. ochracea cells can generate prodigious volumes of iron oxyhydroxides, changing the ecosystem's geochemistry and ecology. Therefore, to determine L. ochracea's basic physiology, we employed new single-cell techniques to demonstrate that L. ochracea oxidizes iron to generate energy and, despite having predicted genes for autotrophic growth, assimilates a fraction of the total CO2 that autotrophs do. Although not a true chemolithoautotroph, L. ochracea's physiological strategy allows it to be flexible and to extensively colonize iron-rich wetlands.

Substrate and electron donor limitation induce phenotypic heterogeneity in different metabolic activities in a green sulphur bacterium.

Populations of genetically identical cells can display marked variation in phenotypic traits; such variation is termed phenotypic heterogeneity. Here, we investigate the effect of substrate and electron donor limitation on phenotypic heterogeneity in N2 and CO2 fixation in the green sulphur bacterium Chlorobium phaeobacteroides. We grew populations in chemostats and batch cultures and used stable isotope labelling combined with nanometer-scale secondary ion mass spectrometry (NanoSIMS) to quantify phenotypic heterogeneity. Experiments in H2 S (i.e. electron donor) limited chemostats show that varying levels of NH4+ limitation induce heterogeneity in N2 fixation. Comparison of phenotypic heterogeneity between chemostats and batch (unlimited for H2 S) populations indicates that electron donor limitation drives heterogeneity in N2 and CO2 fixation. Our results demonstrate that phenotypic heterogeneity in a certain metabolic activity can be driven by different modes of limitation and that heterogeneity can emerge in different metabolic processes upon the same mode of limitation. In conclusion, our data suggest that limitation is a general driver of phenotypic heterogeneity in microbial populations.

Cell-specific nitrogen- and carbon-fixation of cyanobacteria in a temperate marine system (Baltic Sea).

We analysed N2 - and carbon (C) fixation in individual cells of Baltic Sea cyanobacteria by combining stable isotope incubations with secondary ion mass spectrometry (SIMS). Specific growth rates based on N2 - and C-fixation were higher for cells of Dolichospermum spp. than for Aphanizomenon sp. and Nodularia spumigena. The cyanobacterial biomass, however, was dominated by Aphanizomenon sp., which contributed most to total N2 -fixation in surface waters of the Northern Baltic Proper. N2 -fixation by Pseudanabaena sp. and colonial picocyanobacteria was not detectable. N2 -fixation by Aphanizomenon sp., Dolichospermum spp. and N. spumigena populations summed up to total N2 -fixation, thus these genera appeared as sole diazotrophs within the Baltic Sea's euphotic zone, while their mean contribution to total C-fixation was 21%. Intriguingly, cell-specific N2 -fixation was eightfold higher at a coastal station compared to an offshore station, revealing coastal zones as habitats with substantial N2 -fixation. At the coastal station, the cell-specific C- to N2 -fixation ratio was below the cellular C:N ratio, i.e. N2 was assimilated in excess to C-fixation, whereas the C- to N2 -fixation ratio exceeded the C:N ratio in offshore sampled diazotrophs. Our findings highlight SIMS as a powerful tool not only for qualitative but also for quantitative N2 -fixation assays in aquatic environments.

Viral infection of Phaeocystis globosa impedes release of chitinous star-like structures: quantification using single cell approaches.

Phaeocystis globosa is an ecologically important bloom-forming phytoplankton, which sequesters substantial amounts of inorganic carbon and can form carbon-enriched chitinous star-like structures. Viruses infecting P. globosa (PgVs) play a significant regulatory role in population dynamics of the host species. However, the extent to which viruses alter host physiology and its carbon assimilation on single cell level is still largely unknown. This study demonstrates for the first time the impact of viral infection on carbon assimilation and cell morphology of individual axenic P. globosa cells using two single cell techniques: high resolution nanometre-scale Secondary-Ion Mass Spectrometry (nanoSIMS) approach and atomic force microscopy (AFM). Up until viral lysis (19 h post infection), the bulk carbon assimilation by infected P. globosa cultures was identical to the assimilation by the non-infected cultures (33 µmol C l(-1)). However, single cell analysis showed that viral infection of P. globosa impedes the release of star-like structures. Non-infected cells transfer up to 44.5 µmol C l(-1) (36%) of cellular biomass in the form of star-like structures, suggesting a vital role in the survival of P. globosa cells. We hypothesize that impediment of star-like structures in infected P. globosa cells may inactivate viral infectivity by forming flocculants after cell lysis. Moreover, we show that substantial amounts of newly produced viruses (≈ 68%) were attached to P. globosa cells prior to cell lysis. Further, we speculate that infected cells become more susceptible for grazing which provides potential reasons for the sudden disappearance of PgVs in the environment. The scenarios of enhanced grazing is at odds to the current perspective that viral infections facilitates microbial mediated processes by diverting host material away from the higher trophic levels.

Impacts of atmospheric anthropogenic nitrogen on the open ocean.

Increasing quantities of atmospheric anthropogenic fixed nitrogen entering the open ocean could account for up to about a third of the ocean's external (nonrecycled) nitrogen supply and up to approximately 3% of the annual new marine biological production, approximately 0.3 petagram of carbon per year. This input could account for the production of up to approximately 1.6 teragrams of nitrous oxide (N2O) per year. Although approximately 10% of the ocean's drawdown of atmospheric anthropogenic carbon dioxide may result from this atmospheric nitrogen fertilization, leading to a decrease in radiative forcing, up to about two-thirds of this amount may be offset by the increase in N2O emissions. The effects of increasing atmospheric nitrogen deposition are expected to continue to grow in the future.

Global impact and application of the anaerobic ammonium-oxidizing (anammox) bacteria.

In the anaerobic ammonium oxidation (anammox) process, ammonia is oxidized with nitrite as primary electron acceptor under strictly anoxic conditions. The reaction is catalysed by a specialized group of planctomycete-like bacteria. These anammox bacteria use a complex reaction mechanism involving hydrazine as an intermediate. The reactions are assumed to be carried out in a unique prokaryotic organelle, the anammoxosome. This organelle is surrounded by ladderane lipids, which make the organelle nearly impermeable to hydrazine and protons. The localization of the major anammox protein, hydrazine oxidoreductase, was determined via immunogold labelling to be inside the anammoxosome. The anammox bacteria have been detected in many marine and freshwater ecosystems and were estimated to contribute up to 50% of oceanic nitrogen loss. Furthermore, the anammox process is currently implemented in water treatment for the low-cost removal of ammonia from high-strength waste streams. Recent findings suggested that the anammox bacteria may also use organic acids to convert nitrate and nitrite into dinitrogen gas when ammonia is in short supply.

Massive expansion of marine archaea during a mid-Cretaceous oceanic anoxic event.

Biogeochemical and stable carbon isotopic analysis of black-shale sequences deposited during an Albian oceanic anoxic event (approximately 112 million years ago) indicate that up to 80 weight percent of sedimentary organic carbon is derived from marine, nonthermophilic archaea. The carbon-13 content of archaeal molecular fossils indicates that these archaea were living chemoautotrophically. Their massive expansion may have been a response to the strong stratification of the ocean during this anoxic event. Indeed, the sedimentary record of archaeal membrane lipids suggests that this anoxic event marks a time in Earth history at which certain hyperthermophilic archaea adapted to low-temperature environments.