Nitrogen Fixation and Microbial Communities Associated with Decomposing Seagrass Leaves in Temperate Coastal Waters

Seagrass meadows play pivotal roles in coastal biochemical cycles, with nitrogen fixation being a well-established process associated with living seagrass. Here, we tested the hypothesis that nitrogen fixation is also associated with seagrass debris in Danish coastal waters. We conducted a 52-day in situ experiment to investigate nitrogen fixation (proxied by acetylene reduction) and dynamics of the microbial community (16S rRNA gene amplicon sequencing) and the nitrogen fixing community (nifH DNA/RNA amplicon sequencing) associated with decomposing Zostera marina leaves. The leaves harboured distinct microbial communities, including distinct nitrogen fixers, relative to the surrounding seawater and sediment throughout the experiment. Nitrogen fixation rates were measurable on most days, but highest on days 3 (dark, 334.8 nmol N g^-1 dw h^-1) and 15 (light, 194.6 nmol N g^-1 dw h^-1). Nitrogen fixation rates were not correlated with the concentration of inorganic nutrients in the surrounding seawater or with carbon:nitrogen ratios in the leaves. The composition of nitrogen fixers shifted from cyanobacterial Sphaerospermopsis to heterotrophic genera like Desulfopila over the decomposition period. On the days with highest fixation, nifH RNA gene transcripts were mainly accounted for by cyanobacteria, in particular by Sphaerospermopsis and an unknown taxon (order Nostocales), alongside Proteobacteria. Our study shows that seagrass debris in temperate coastal waters harbours substantial nitrogen fixation carried out by cyanobacteria and heterotrophic bacteria that are distinct relative to the surrounding seawater and sediments. This suggests that seagrass debris constitutes a selective environment where degradation is affected by the import of nitrogen via nitrogen fixation.