The effect of climate warming on soil organic matter decomposition has major implications for soil carbon sequestration and nutrient circulation (Fontaine et al., 2007; Liang et al., 2019; Meyer et al., 2018). Soil microorganisms, such as soil organic matter decomposers, synthesize various extracellular enzymes to depolymerize macromolecular soil organic matter into absorbable substrates to fulfill their demands (e.g., C, N, and P) for growth and metabolism (Sinsabaugh et al., 2008, Allison et al., 2011). Extracellular enzymes thus reflect microbial resource constraints (Cui et al., 2021; Sinsabaugh et al., 2008) and are widely used to assess resource limitations (Chen et al., 2018; Cui et al. 2019; Guan et al., 2020; Jing et al., 2020; Moorhead et al., 2016; Peng et al., 2016; Waring et al., 2014). Although warming has been shown to affect numerous ecological processes, its effects on microbial resource limitations remain unclear.
The soil extracellular enzyme stoichiometric theory combines ecological metabolism theory with the stoichiometric theory of ecology to evaluate the resource limitation of microbial metabolism through the ratio of various enzyme activities (Chen et al., 2018 a, b; Cui et al., 2021; Cui et al., 2018; Sinsabaugh et al., 2008, 2012; Yuan et al., 2019; Zhou et al., 2020). Thus, enzyme stoichiometry is regarded as an effective tool for assessing the environmental drivers and resource constraints of microbial metabolism; therefore, it has been widely used to explore biogeochemical cycles (Nottingham et al., 2015; DeForest and Moorhead, 2020). The enzymes commonly used in enzymatic stoichiometry are mainly involved in the microbial acquisition of C (β-1,4-glucosidase, BG), N (β-1,4-N-acetylglucosaminidase, NAG), and P (acid phosphatase, AP, Sinsabaugh et al., 2008). Moorhead et al. (2013) proposed a vector model to identify the relative resource limitation of microorganisms by using the vector length and angle calculated from the enzymatic C:P versus C:N acquisition ratio. Using this approach, Moorhead et al. (2016) showed that soil microbial communities are more limited by P in montane tropical forests than in lowland tropical forests. Zheng et al. (2020) reported that warming reduced microbial C limitation in the mineral soil layer and increased P limitation in the organic layer in an alpine shrubland ecosystem. Lie et al. (2019) found that warming mitigated P limitation and increased N consumption in tropical forests by increasing the soil P availability. However, these studies have mainly focused on the 0–20 cm soil depth topsoil, and the knowledge of the effects of warming on microbial resource limitation below the 20 cm soil depth subsoil remains limited.
Subsoil accounts for over 60% of the soil C stock in the top 1 m of soil (Jobbágy and Jackson, 2000). The subsoil and topsoil differ in many crucial physical and chemical properties. Subsoil tends to have a lower temperature (Zogg et al., 1997) and oxygen concentration (DeAngelis et al., 2010) and higher soil moisture (Serna-Chavezet al., 2013) and pH (Fierer and Jackson, 2006) than topsoil. These physical and chemical properties play relevant roles in the regulation of extracellular enzyme activity by modifying the diffusion and adsorption of available substrates (Deng et al., 2019; Feng et al., 2019). Thus, the resource limitation for soil microorganisms in subsoil can substantially differ from that in topsoil. For example, fresh C input is more limited in subsoil than topsoil such that the subsoil may have greater C limitation than topsoil. Several recent studies have shown that warming decreases topsoil soil moisture, deepening root growth; therefore, C input via root exudation increases and C limitation decreases in subsoil (Querejeta et al., 2021; Zheng et al., 2020). The relief of C limitation in subsoil has been suggested to increase soil organic matter (SOM) decay (Shahzad et al., 2018; Sullivan et al., 2020). However, the understanding of how warming affects C and nutrient limitations in subsoil relative to topsoil is incomplete because few studies have focused on subsoil. Considering the subsoil C storage, slight changes in subsoil carbon may have significant effects on the global carbon cycle. Therefore, clarifying the impact of warming on microbial resource limitations in the subsoil is necessary.
Microbial community composition, recognized as context-dependent (e.g., climate and soil properties; Fierer and Jackson, 2006), has been shown to be a major factor regulating enzyme production (McGuire and Treseder, 2010; Schnecker et al., 2014; Stone et al., 2014; Strickland et al., 2009, Li et al., 2019) and may have a reciprocal influence on soil microbial resource limitations. Gram-positive (GP) and gram-negative (GN) bacteria are oligotrophic and copiotrophic bacteria respectively (Fierer et al., 2007; Fanin et al., 2018). GP uses recalcitrant carbon (C) compounds, exhibiting a higher C limitation than GN. Comparatively, GN uses labile C compounds (Fanin et al., 2018; Naylor and Coleman-Derr, 2018), showing a lower C limitation than GP. In addition, warming has been shown to increase, decrease, or have no effect on the abundances of GP and GN (Feng and Simpson, 2009; Frey et al., 2008; Karhu et al., 2010; Rinnan et al., 2008; Rinnan et al., 2009; Vanhala et al., 2011). Moreover, the impact of warming on the soil microbial community composition is depth-dependent (Dove et al., 2021). Although many studies have focused on the effects of warming on the microbial community structure, the relationship between microbial communities and microbial resource limitation has rarely been examined. Studying the relationship between microbial community structure and resource limitations would advance the understanding of the effects of warming on C, N, and P cycles.
To illustrate the outlined issues outlined above, we investigated microbial resource limitation and explored its relationships with microbial communities and soil properties in the topsoil and subsoil in a field manipulation experiment after 2 years of warming treatment in a subtropical Chinese fir plantation. Based on the commonly reported differences in abiotic factors and microbial communities between topsoil and subsoil, we hypothesized that warming would have different effects on microbial resource limitation between topsoil and subsoil (H1). Because of the critical role of microbial enzymes in nutrient acquisition, we also hypothesized that the microbial community structure plays a primary role in regulating microbial resource restriction (H2).