DNA-stable isotope probing (DNA-SIP) identifies marine sponge-associated bacteria actively utilizing dissolved organic matter (DOM).

Sponges possess exceptionally diverse associated microbial communities and play a major role in (re)cycling of dissolved organic matter (DOM) in marine ecosystems. Linking sponge-associated community structure with DOM utilization is essential to understand host-microbe interactions in the uptake, processing, and exchange of resources. We coupled, for the first time, DNA-stable isotope probing (DNA-SIP) with 16S rRNA amplicon sequencing in a sponge holobiont to identify which symbiotic bacterial taxa are metabolically active in DOM uptake. Parallel incubation experiments with the sponge Plakortis angulospiculatus were amended with equimolar quantities of unlabelled (12 C) and labelled (13 C) DOM. Seven bacterial amplicon sequence variants (ASVs), belonging to the phyla PAUC34f, Proteobacteria, Poribacteria, Nitrospirae, and Chloroflexi, were identified as the first active consumers of DOM. Our results support the predictions that PAUC34f, Poribacteria, and Chloroflexi are capable of organic matter degradation through heterotrophic carbon metabolism, while Nitrospirae may have a potential mixotrophic metabolism. We present a new analytical application of DNA-SIP to detect substrate incorporation into a marine holobiont with a complex associated bacterial community and provide new experimental evidence that links the identity of diverse sponge-associated bacteria to the consumption of DOM.