Spatiotemporal and temperature-dependent disconnect between ammonia oxidation and dark DIC fixation in deep oligotrophic Lake Constance

Deep oligotrophic lakes hold over 80% of global lake water. In their hypolimnion, ammonia oxidation (the first step of nitrification) and non-photosynthetic fixation of dissolved inorganic carbon (DIC) are key processes, presumably linked by large populations of ammonia-oxidizing archaea (AOA). We used stable isotope-based activity measurements to follow both processes below the thermocline and in the central hypolimnion in deep oligotrophic Lake Constance. Throughout seasons, they varied substantially below the thermocline peaking at 139.0 NH4+ nmol l−1 d−1 oxidized and 14.6 nmol DIC l−1 d−1 fixed. At the center of the hypolimnion, they were rather stable averaging 7.5 nmol NH4+ l−1 d−1 and 1.3 nmol DIC l−1 d−1, respectively. However, both processes did not correlate in their spatiotemporal and temperature-related dynamics. Temperature manipulations (5–20°C) confirmed this disconnect with ammonia oxidation peaking at 10°C while dark DIC fixation increased exponentially with temperature. DIC fixation of single AOA cells centered at 2.17 × 10−18 mol C cell−1 d−1, explaining only 11% of overall DIC fixation. Metatranscriptomic analyses supported this, revealing that most DIC-fixation pathway transcripts originated from RubisCO-encoding cryptophytes, cyanobacteria, and Alpha- and Betaproteobacteria, rather than AOA or other nitrifiers. These non-nitrifier groups likely activated the Calvin cycle to maintain redox balance in the dark. Our findings provide a new perspective on nitrification-driven chemolithoautotrophy in oligotrophic lake hypolimnia, with freshwater AOA contributing a minor part to dark DIC fixation, likely explaining decoupled dynamics of ammonia oxidation and dark DIC fixation.