Deoxygenation effects on the interaction between microbial metabolisms and dissolved organic matter cycling in the seasonally anoxic Mariager Fjord (Denmark, North Sea)

Seawater stores as much carbon in the form of dissolved organic matter (DOM) as there is CO₂ in the atmosphere. Over a period of just 50 years (from 1960 to 2010) global oceanic oxygen reserves have been reduced by 2% and the anoxic waters have quadrupled, mainly due to anthropogenic global warming and eutrophication. Ocean deoxygenation leads to an expansion of oxygen minimum zones (OMZs), which contain higher concentrations of DOM (carbon and sulfur (DOS)) than the oxygenated ocean. Microbial metabolisms are directly responsible for the production, degradation and recycling of marine DOM but there is no consensus about their role in DOM accumulation in OMZs. Recent advances in analytical chemistry characterize the DOM at the molecular level in unprecedented detail, revealing new insights into its source and history by Fourier transform ion-cyclotron resonance mass spectrometry (FT-ICR-MS). Current progress in sequencing technology can predict specific functions contributing to the molecular activity of microbial communities in environmental samples by metatranscriptomics, or to specific substrate assimilation by quantitative DNA stable isotope probing (qSIP). In this study, we investigated the interaction between microbes and DOM in the water column of the Mariager Fjord (Denmark, North Sea). Due to nutrient loading from land combined with its topography, Mariager Fjord has anoxic bottom waters between spring and late fall, but it is typically flushed with oxygen-rich seawater from the Kattegat during winter. In spring 2023, we sampled waters at three depths (5, 15, 25 m) with an O₂ range from oxic-to-hypoxic conditions (99, 65 and 4 % O₂, respectively). Natural environmental samples were combined with incubations targeting (a) abiotic and biotic interactions in the presence or absence of oxygen; and (b) organosulfur cycling by active uncultivated microbes assimilating the ¹³C-DOS substrates methionine and taurine. Samples were analyzed for elemental organic and inorganic geochemistry, microbial diversity (16S rRNA sequencing), FT-ICR-MS, qSIP and metatranscriptomics. Our results showed clear changes on the microbial community composition and gene expression depending on the oxygen concentration. The surface oxic waters were dominated by Planctomycetes and Actinobacteria, while the hypoxic nitrite-enriched waters were dominated by Gammaproteobacteria and Bacteroidota. Expressed genes diversity was highest in the hypoxic waters, with reverse dissimilatory sulfate reduction and sulfur oxidation genes present in the metatranscriptomes, even though the waters were not sulfidic. Regarding organosulfur cycling, only bacteria assimilated ¹³C-DOS in the water column. Methionine was mainly utilized in oxic layers by Gammaproteobacteria, Alphaproteobacteria and Actinobacteria, while taurine was only assimilated in hypoxic waters mostly by Bacteroidota. Largest differences in DOM molecular composition between oxic-to-hypoxic samples were related to N- and S-containing compounds, although autochthonous terrigenous DOM input in the fjord dominated the DOM signature more drastically than oxygen variations. Overall, our study includes novel implementation of state-of-art methods to elucidate new links between the microbial biosphere with the chemical diversity of DOM in the context of a changing, deoxygenated ocean.