Urea fertilization increased CO2 and CH4 emissions by enhancing C-cycling genes in semi-arid grasslands.

Global climate change is predicted to increase exogenous N input into terrestrial ecosystems, leading to significant changes in soil C-cycling. However, it remains largely unknown how these changes affect soil C-cycling, especially in semi-arid grasslands, which are one of the most vulnerable ecosystems. Here, based on a 3-year field study involving N additions (0, 25, 50, and 100 kg ha−1 yr−1 of urea) in a semi-arid grassland on the Loess Plateau, we investigated the impact of urea fertilization on plant characteristics, soil properties, CO 2 and CH 4 emissions, and microbial C cycling genes. The compositions of genes involved in C cycling, including C fixation, degradation, methanogenesis, and methane oxidation, were determined using metagenomics analysis. We found that N enrichment increased both above- and belowground biomasses and soil organic C content, but this positive effect was weakened when excessive N was input (N100). N enrichment also altered the C-cycling processes by modifying C-cycle-related genes, specifically stimulating the Calvin cycle C-fixation process, which led to an increase in the relative abundance of cbbS , prkB , and cbbL genes. However, it had no significant effect on the Reductive citrate cycle and 3-hydroxypropionate bi-cycle. N enrichment led to higher soil CO 2 and CH 4 emissions compared to treatments without added N. This increase showed significant correlations with C degradation genes (bglA , per , and lpo), methanogenesis genes (mch, ftr, and mcr), methane oxidation genes (pmoA , pmoB , and pmoC), and the abundance of microbial taxa harboring these genes. Microbial C-cycling genes were primarily influenced by N-induced changes in soil properties. Specifically, reduced soil pH largely explained the alterations in methane metabolism, while elevated available N levels were mainly responsible for the shift in C fixation and C degradation genes. Our results suggest that soil N enrichment enhances microbial C-cycling processes and soil CO 2 and CH 4 emissions in semi-arid ecosystems, which contributes to more accurate predictions of ecosystem C-cycling under future climate change. • Urea fertilization increased CO 2 and CH 4 emissions in semi-arid grasslands. • Urea fertilization enhanced microbial C-cycling by altering C-related genes. • Microbial C fixation and C degradation genes were mainly driven by available N. • Microbial methane metabolism genes were mainly driven by pH. [ABSTRACT FROM AUTHOR]