Cold-water coral reefs host diverse microbial communities that play key roles in nutrient cycling and deep-sea productivity. Among them, methylotrophic bacteria metabolize and cycle one-carbon (C1) compounds such as methane, methanol, methylated nitrogen and sulphur, linking food webs and having a central role in the global cycling of climate-active trace gases. To date, few methylotrophic bacteria have been cultured from cold-water coral reefs. In this study, we enriched methylotrophs from seawater, sediment and sponge samples collected in cold-water coral reefs off mid-Norway. Cultivation at 6-10 °C with ~20% methane in air using liquid and solid marine mineral salt media, combined with general and methanotroph-specific 16S rRNA gene screening, yielded 47 bacterial phylotypes spanning 22 genera from 19 families. Methanotrophs from the genera Methylococcus, Methylosinus and a pmoA-verified Methylocystis were enriched from seawater and sediment. Other phylotypes recovered included Methylophaga, Pelagibacter, Halomonas, Zhongshania and Rhodococcus. DNA stable isotope probing of a Plakortis sp. sponge traced methane-derived carbon into 16S rRNA gene sequences affiliated with sponge-associated Gammaproteobacteria and Methylocystis (Alphaproteobacteria). Supported by searching the genera's genomes, these findings reveal a broad range of bacteria potentially active in C1 cycling within deep, cold-water reef ecosystems.
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