Nitrification Couples Microbial CO2 Fixation to Warming and Drought Responses in Alpine Grassland Soils

Microbial CO2 fixation in alpine grassland soils is highly sensitive to warming and drought. Nitrogen inputs from grazing may stimulate autotrophic nitrifiers, including ammonia-oxidizing archaea (AOA), ammonia-oxidizing bacteria (AOB), and nitrite-oxidizing bacteria (NOB). These nitrifiers assimilate carbon through CO2 fixation, but how their activity is associated with microbial CO2 fixation under warming and drought remains unclear. Here, we investigated microbial CO2 fixation and nitrification under altered temperature and moisture using urea-amended soil microcosms with 13CO2 labeling, DNA-stable isotope probing (DNA-SIP), metagenomics, and quantitative PCR. Warming increased CO2 fixation rates to 1.48-2.58 times those at 15 °C under moist conditions, whereas drought reduced it by 59-91%. Nitrification rates were positively correlated with CO2 fixation, whereas the CO2 fixation offset only 1.3-12.1% of associated N2O emissions (measured as CO2 equivalents). DNA-SIP and metagenomics indicated that nitrifiers contributed to microbial CO2 fixation, with AOA showing more pronounced 13C-labeling under combined warming and drought. Co-occurrence network indicated that AOA occupied more highly connected positions than AOB and NOB. This study provides microbial evidence that warming and drought reshape the linkage between nitrification and microbial CO2 fixation in urea-amended alpine grassland soil microcosms, with implications for carbon-nitrogen cycling and greenhouse-gas feedbacks.