Temperature differentially regulates estuarine microbial NO production along a salinity gradient.

Nitrous oxide (NO) is atmospheric trace gas that contributes to climate change and affects stratospheric and ground-level ozone concentrations. Ammonia oxidizers and denitrifiers contribute to NO emissions in estuarine waters. However, as an important climate factor, how temperature regulates microbial NO production in estuarine water remains unclear. Here, we have employed stable isotope labeling techniques to demonstrate that the NO production in estuarine waters exhibited differential thermal response patterns between nearshore and offshore regions. The optimal temperatures (T) for NO production rates (NOR) were higher at nearshore than offshore sites. N-labeled nitrite (NO) experiments revealed that at the nearshore sites dominated by ammonia-oxidizing bacteria (AOB), the thermal tolerance of N-NOR increases with increasing salinity, suggesting that NO production by AOB-driven nitrifier denitrification may be co-regulated by temperature and salinity. Metatranscriptomic and metagenomic analyses of enriched water samples revealed that the denitrification pathway of AOB is the primary source of NO, while clade II NO-reducers dominated NO consumption. Temperature regulated the expression patterns of nitrite reductase (nirK) and nitrous oxide reductase (nosZ) genes from different sources, thereby influencing NO emissions in the system. Our findings contribute to understanding the sources of NO in estuarine waters and their response to global warming.