Stable isotope tracing in the Ianthella basta sponge holobiont

Ammonia-oxidizing archaea (AOA) frequently form symbiotic associations with marine sponges. While free-living AOA are generally considered metabolically constrained chemolithoautotrophs, sponge-associated AOA encode for a branched-chain amino acid (BCAA) transporter, suggesting mixotrophic potential. Here, we test the assimilation of 13C- and 15N-labeled BCAA and the 13C-labeled dissolved inorganic carbon fixation in the sponge holobiont Ianthella basta. We demonstrate that BCAA degradation fuels ammonia oxidation and quantify BCAA uptake at the single-cell level by combining stable isotope probing, catalyzed reporter deposition fluorescence in situ hybridization (CARD-FISH), and nanoscale secondary ion mass spectrometry (NanoSIMS). Our results reveal that the sponge-associated AOA, Candidatus Nitrosospongia ianthellae, assimilates BCAA as an additional carbon and nitrogen source, while being able to fix dissolved inorganic carbon. Our study reveals a novel nutritional interaction in sponge holobionts and challenges the perception of constrained metabolic capacities of AOA. Sequencing was performed at the Joint Microbiome Facility of the Medical University of Vienna and the University of Vienna under the project IDs JMF-2212-27, JMF-2303-04, and JMF-2407-25.