Introduction
Nitrous oxide (N2O), a non-carbon dioxide (CO2) greenhouse gas, has a global warming potential nearly 300-fold greater than that of CO2 over a 100-year lifespan (Dijkstra et al., 2013). The accumulation of N2O in the atmosphere will deplete stratospheric ozone and contribute to global warming (Ravishankara et al., 2009). The main sources of atmospheric N2O are closely associated with soil nitrogen (N) cycling (i.e., nitrification and denitrification) of terrestrial ecosystems, which contribute to ~56–70% of global N2O emissions (Butterbach-Bahl et al., 2013). Grasslands host one of the most widely distributed vegetation types on earth, and grassland ecosystems are the main component of terrestrial ecosystems (Scurlock et al., 2002). On the Qinghai-Tibetan Plateau (QTP), alpine grassland ecosystems (e.g., alpine meadows and alpine steppes) are huge nitrogen (N) reservoirs because of sluggish microbial decomposition (Yang et al., 2018; Zhang et al., 2020). However, the substantial labile N stored in alpine soils, which is a large source of N2O, is often neglected (Mao et al., 2020). Global change, particularly atmospheric N deposition and changing precipitation regimes, has considerable consequences for storage and patterns of N in alpine ecosystems (Fu et al., 2017; Lin et al., 2016). Given that alpine grasslands may possess the capacity for N2O release and are sensitive to global change (Xiao et al., 2020), understanding how alpine soil N2O emissions respond to N deposition and precipitation changes is crucial for predicting future atmospheric N2O concentrations.
The main regulatory factors for plant communities and soil ecological processes in grasslands are N and water. Field simulations of the impact of atmospheric N deposition on N2O emissions are not scarce, especially in the alpine grasslands of the QTP. However, reports of the effects of N addition in these ecosystems are inconsistent. N addition has been shown to significantly increase soil N2O emissions, because N input elevates the concentration of inorganic N and the abundance of functional microbes in the soil (Geng et al., 2019; Peng et al., 2018; Wu et al., 2020; Yan et al., 2018). In addition, a greater labile carbon (C) supply (e.g., litter decomposition or root exudation) under N enrichment provides substrate C for heterotrophic denitrifiers, thereby stimulating N2O emissions (Brown et al., 2012; Dijkstra et al., 2013). However, Zhu et al. (2015) showed that N input did not affect N2O emissions. A possible interpretation of this finding is that low temperature and inadequate soil moisture limit the activities of microorganisms associated with N cycling in cold conditions (Banerjee et al., 2016; Curtis et al., 2006; Schaufler et al., 2010). Despite this work on grasslands, the response of N2O emissions to long-term N deposition on the QTP remains understudied.
Soil N2O emissions are also susceptible to hydrologic variations (Knapp et al., 2002). Generally, changes in soil water content influence N mineralization and organic matter degradation, which then affect the provision of N and C reactants for N cycling processes. On a global scale, elevated precipitation in grassland ecosystems accelerates N2O emissions while decreased precipitation mitigates N2O emissions. These processes are predominantly regulated by shifts in soil water availability (Li et al., 2020). By contrast, Liu et al. (2014) showed that short-term water increment did not affect N2O emissions from semiarid steppes. Even increased precipitation decreased N2O emission in arid grasslands (Cai et al., 2016). This finding may be attributable to soil leaching and run-off events caused by the increased rainfall, which intensified the loss of inorganic N in soil and thereby limited soil N cycling (Cregger et al., 2014). Little is known about how long-term precipitation changes impact N2O emissions on the QTP. Both N and water affect soil biogeochemical cycles. N deposition and variation in precipitation usually occur simultaneously; thus, their effects are interdependent (Harpole et al., 2007). The combined effect of N deposition and altered precipitation on N2O emissions is also unknown. N-cycling microbiomes play a crucial role in regulating soil N dynamics and global climate stabilization. On the QTP it is also unclear how pivotal N-cycling functional microorganisms respond to global change and which microbes better explain N2O emissions.
Due to multifactorial climate change and intensive interventions targeting anthropogenic activities, the environmental conditions of the QTP have undergone dramatic changes in the past few decades (Gong et al., 2017). The amount, frequency, and intensity of precipitation increased from 1975 to 2014 (Ge et al., 2017). The QTP is also confronting pronounced N deposition, with an average of ~8 kg N ha−1 year−1 (Lü et al., 2007). The alpine steppes, the largest grassland ecosystem on the QTP, are extremely sensitive to global change (Ding et al., 2016; Wang et al., 2011). Therefore, understanding the effects of N enhancement and altered precipitation on N2O emissions in the alpine steppes is essential. This study consists of altered precipitation and N addition manipulation experiments that were conducted in an alpine steppe on the QTP in 2013. We monitored the N2O flux during the 2020 growing season (May to October) based on in-situ experiments. To identify the key abiotic and biotic factors regulating N2O emissions, we measured N2O flux on six consecutive days in mid-August (during peak plant growth). Soils were also collected to measure abiotic parameters and functional microbes, including nitrifiers (ammonia-oxidizing bacteria: AOB; ammonia-oxidizing archaea: AOA) and denitrifiers (nirS -,nirK -, and nosZ gene-containing microorganisms). The objectives of the study were to (1) assess whether N2O emissions were altered by long-term N addition, precipitation changes, and their interaction; and (2) identify the mechanisms that regulated N2O emissions under N addition and altered precipitation patterns.