Soil is the largest reservoir of terrestrial organic C in the biosphere, and more than half of soil C is stored in forest ecosystems (Schlesinger and Bernhardt, 2013). Tropical forest C accounts for 55% of the total forest C stock, 32% of which is stored in soil (Pan et al., 2011). Soil microorganisms regulate soil C sequestration and decomposition process and therefore have a strong impact on the terrestrial C cycle. Integration of more realistic and accurate key parameters, such as carbon use efficiency (hereafter, CUE), defined as the fraction of growth to total assimilation C (Geyer et al., 2016), could reduce the uncertainty in the long-term soil C response to climate change (Luo and Schuur, 2020; Tao et al., 2023). N is one of the essential nutrients for soil microbial growth and metabolism. Therefore, the association between microbial CUE and soil N availability has garnered increasing attention with augmented N deposition during the last decade (Feng et al., 2022; Liu et al., 2018b; Spohn et al., 2016a; Yuan et al., 2019).
The response of microbial CUE to N deposition varied across ecosystems. Microbial CUE could positively, negatively, or neutrally respond to an increase in N availability, limiting our prediction of soil C storage under future N deposition. For example, studies have reported that higher nitrogen availability could reduce microbial respiration (Moore et al., 2021; Xiao et al., 2020) and conversely increase its growth (Manzoni et al., 2012; Spohn et al., 2016b), and finally led to an increase in microbial CUE (Lee and Schmidt, 2014; Spohn et al., 2016b; Zhran et al., 2021). Nevertheless, a six-year N addition experiment revealed a decreased CUE in a temperate grassland, which was attributed to a decline in the fungi to bacteria ratio (F/B) (Riggs and Hobbie, 2016). Similarly, Luo et al. (2020) found a decrease in CUE with a four-year N addition in a Tibetan grassland, which was induced by an increased dominance of fast-growing bacteria. Widdig et al. (2020) and Ma et al. (2023) reported an unchanged CUE with N addition and attributed this to a similar climate and soil texture. Overall, the effect of N addition on microbial CUE was achieved by changing the availability of soil nutrients, altering microbial community, and influencing soil pH value. Microbial CUE increased with increasing N availability, especially in N-limited ecosystems, as they decreased the investment in secreting enzymes for nutrient mining and reduced respiring C (Chen and Yu, 2020; Manzoni et al., 2012). The decomposition and uptake rates of microbial communities vary based on their composition, resulting in different adaptation strategies to changes in the nutrient environment. Recent research has indicated a negative association between CUE and the ratio of F/B, suggesting that fungi might exhibit lower CUE compared to bacteria due to their increased investment in the secretion of extracellular enzymes (Soares and Rousk, 2019; Ullah et al., 2021). Similarly, Pold et al. (2020) proposed that CUE of the slow-growing bacterial population was inferior to that of the fast-growing bacterial population. Soil pH is an important factor affecting microbial CUE (Hu et al., 2022). On the one hand, lower pH values may cause environmental stress for microbes, resulting in lower CUE (Rousk et al., 2009). On the other hand, lower pH values could influence iron reduction (Hall et al., 2016; Lipson et al., 2010; Schulz et al., 2016) and its related mineral protection for organic matter and hence for C availability (Ye et al., 2018; Chen et al., 2020) and CUE. Furthermore, pH has a significant impact on the composition of microbial communities. Higher microbial diversity allows for the utilization of various carbon sources, thereby increasing CUE (Domeignoz-Horta et al., 2020).
Despite the fact that extensive research has been conducted to date, our knowledge regarding the microbial response to N deposition and its impact on CUE is still inadequate. Presently, research efforts have predominantly concentrated on examining the topsoil layer (less than 30 cm) of agricultural and grassland environments. However, there is inadequate data available regarding forest ecosystems, specifically the underlying soil layers. The subsoil which lies beneath the 30 cm depth, accumulates more than 50% of SOC and retains it for an extended duration (Chen et al., 2023; Jobbágy and Jackson, 2000; Rumpel and Kögel-Knabner, 2011). In addition, the response of microbial CUE to N addition changed with soil layers. Studies found microbial CUE decreased or did not change with N addition in topsoil (0 − 10 cm), while in deeper soils (10 − 30 cm), it increased with N availability, which was called ‘flexible CUE’ (Manzoni, 2017). However, a recent global-scale study unveiled a correlation between the increase in soil depth to 100 cm and a corresponding decrease in soil microbial biomass as well as bacterial diversity (He et al., 2023). Therefore, it is not clear whether microbial CUE in soil layers below 30 cm decreases with a decrease of C input and the increase of microbial death in an anaerobic environment, nor is the regulating mechanism clear.
To enhance the precision of predicting the impact of N deposition on the feedback of soil C stock in forest ecosystems, it is imperative to gain a comprehensive understanding of the mechanisms that underlie the diverse responses of microbial CUE to N deposition. An alternative method evaluates how microbes change their resource use in response to substrate stoichiometry using assays of extracellular enzymes that are based on community-level resource capture. Compared to the labeled approach, the stoichiometrically defined CUE encompasses the biochemical CUE within it (Schimel et al., 2022). To indicate the factors and mechanisms of microbial CUE response in the top and deep soil layer to N addition, we conducted a short-term N addition experiment with soils collected from two tropical forests in southern China, both topsoil (0 − 10 cm) and deep soil (60 − 80 cm). To investigate the overall reactions of microbial CUE in forest soil to N addition, we additionally gathered a comprehensive global dataset and employed a meta-analysis approach to systematically and quantitatively evaluate the findings. We aimed to assess (1) the impact of N addition on forest soil microbial CUE, (2) the variability in microbial CUE response to N addition across different soil layers, and (3) the potential similarities or differences in the underlying mechanisms of microbial CUE response to N addition between the top and deep soil layers. We hypothesized that (1) N addition would have no effects on forest topsoil microbial CUE because microbial could be influenced by more factors than on subsoil, and (2) SOC was the determining factor when considering deep soil as it was considerably less abundant in the deeper soil layer.