Weakening of the ‘enzymatic latch’ mechanism following long-term fertilization in a minerotrophic peatland

Highlights

• N addition regulated the stoichiometry of BDG, NAG, and PHO activities.

• P played a more important role in downregulating BDG, NAG, and PHO activities.

• The ‘enzymatic latch’ became less critical after the long-term fertilization.

• The interaction between N and P exerted an important control on northern peatlands.

Abstract

Despite the global importance of understanding the effects of nitrogen (N) and phosphorus (P) deposition on carbon (C) cycling in northern peatlands, the control of N and P interactions through the ‘enzymatic latch’ mechanism has been largely overlooked. A long-term fertilization experiment in a moderate-rich fen in northeast China was conducted to evaluate the underlying mechanisms of how continuous N and P addition regulate key extracellular enzymes and their interactions, and the subsequent influences on organic C storage in peatlands. The results demonstrated that the growth of Sphagnum moss and vascular plants were both reduced by N addition but enhanced by P addition. Phenolic concentrations were higher in P addition treatments, as were phenol oxidase activities. In general, N addition played a critical role in regulating the stoichiometry of β-D-glucosidase, N-acetyl-β-glucosaminidase and phosphatase, while P addition was more important in regulating their activities. The direct and indirect effects due to fertilization and shifts in vegetational composition, respectively, weakened the ‘enzymatic latch’ mechanism that controls the decomposition of organic matter after long-term fertilization. Our results indicate that P likely plays a more important role than N in controlling microbial extracellular enzymatic activities and organic matter decomposition in northern minerotrophic peatlands. Consequently, the interactions between N and P is likely of primary significance in regulating the biogeochemical cycling of peatlands.

Introduction

Northern peatlands are important carbon (C) sinks that store one-third of the global soil C (500 ± 100 Gt), but only comprise approximately 3% of the total land surface (Gorham, 1991, Yu et al., 2010). Moreover, more C is stored in Sphagnum moss and litter than is fixed by all terrestrial global plants every year (Clymo and Hayward, 1982, Page and Baird, 2016). In China, peatlands are primarily located in the northeast and southwest, comprise an area of ~10,441 km², and store ~1.39 Gt organic carbon (Ma, 2013). The harsh conditions associated with northern peatlands (i.e., low nutrients, cold temperatures, and acidic and waterlogged soils) greatly limit microbial activity and decrease organic matter decomposition rates (Moore and Basiliko, 2006, Rydin and Jeglum, 2013).

Nitrogen (N) and phosphorus (P) are critical nutrients in peatlands and have profound effects on plant production, litter decomposition, and biogeochemical cycles in these environments (Limpens et al., 2006, Walbridge and Navaratnnam, 2006, Rydin and Jeglum, 2013). N and P are also important nutrients for microorganisms and are involved in many microbial biochemical processes (Sinsabaugh and Moorhead, 1994). Atmospheric reactive N deposition has increased by 59% from 12.64 kg N ha⁻¹ yr⁻¹ in the 1960s to 20.07 kg N ha⁻¹ yr⁻¹ in the past decade, due to increased human activities and massive resource consumption (Lü and Tian, 2014). The northern peatlands also encounter high rates of N deposition, which affect plant productivity, organic matter decomposition, and weaken their C sequestration capacities (Bragazza et al., 2012, Larmola et al., 2013, Wang et al., 2015b). The natural and anthropogenic emission of P increased globally from 2.6 to 4.0 Tg P yr⁻¹ between 1850 and 2013 (Wang et al., 2017). P addition may relieve P limitation, while stimulating biological N fixation (Benner and Vitousek, 2007), although it may not alleviate physiological stresses on Sphagnum mosses due to excessive N addition (Fritz et al., 2012). It is well known that interactions between N and P play key roles in regulating ecological processes and functions in a wide range of ecosystems (e.g., Elser et al., 2000, Zechmeister-Boltenstern et al., 2015). However, few studies have investigated the effects of long-term N and P inputs on the ecological processes and functions in peatlands (Bubier et al., 2011, Song et al., 2011, Larmola et al., 2013, Keuskamp et al., 2015). Such studies would provide a better understanding of the response and feedback mechanisms of N and P inputs into northern peatlands. In addition, changes in plant biomass and vegetation structure due to long-term nutrient input should be an important consideration underlying ecological function in peatlands (Bobbink et al., 2010).

Soil enzymes are the principal media by which soil microorganisms participate in soil biogeochemical processes including organic matter decomposition and nutrient cycling, and they are sensitive to soil nutrient characteristics (Kuijper et al., 2012). The ‘enzymatic latch’ is a well-known mechanism positing that the low phenol oxidase (POX) activity within peatlands leads to the build-up of phenolics ([PHEN]), which then inhibits the activity of hydrolytic enzymes, and thereby facilitates the accumulation of organic matter (Freeman et al., 2001, Freeman et al., 2004, Freeman et al., 2012). Phenolics inhibit decomposition by binding to the reactive sites of hydrolytic enzymes (Brouns et al., 2014). Importantly, POX partially oxidize phenolics into simple organic compounds (Duran et al., 2002, Freeman et al., 2004). Due to their importance in peatland nutrient cycling, POX activity and the concentration of soluble phenolics within peatlands have been investigated in recent years in the context of warming climates (Jassey et al., 2012) and long-term fertilization (Pinsonneault et al., 2016a). However, the effect of long-term fertilization on POX, and especially the interaction of N and P, is still unclear. For example, Bragazza et al. (2006) observed that POX activity increased nearly three-fold when the external N input was increased from 0.2 to 2 g N m⁻² yr⁻¹. However, Pinsonneault et al. (2016a) suggested that POX activity could be significantly reduced by long-term N additions. Nevertheless, most studies have not accounted for the role of P in regulating POX activity.

β-D-glucosidase (BDG), N-acetyl-β-glucosaminidase (NAG), and phosphatase (PHO) directly constrain the decomposition of organic C, and changes in their activities are reliable indicators of organic C decomposition in peatlands (Freeman et al., 1995, Shackle et al., 2000, Kuijper et al., 2012). BDG is an enzyme used to acquire C via the hydrolysis of cellulose to glucose, which is a labile C resource that supports microbial metabolisms (Kuijper et al., 2012, Dunn et al., 2014). Increases in external nutrient inputs result in the release of greater BDG to meet C demands (Sinsabaugh and Moorhead, 1994, Dunn et al., 2014). POX activity may be influenced by fertilization and can also affect the activity of BDG (Freeman et al., 2004). Evolutionary-economic principles (Allison et al., 2010) assert that microorganisms produce enzymes due to the balance between resource supply and demand. Consequently, NAG, the primary enzyme for N acquisition via chitin decomposition (Kang et al., 2005), may exhibit increased activity due to the addition of P. Similarly, N additions can also lead to increases in the activity of PHO, which releases phosphate by catalyzing the breakdown of phosphate monoesters (Dunn et al., 2014). Therefore, the stoichiometric trade-off between N and P (i.e., N and P interactions) may be the primary factor controlling the above-mentioned biochemical processes in peatlands.

In this study, the effects of N and P addition on the ‘enzymatic latch’ mechanism were evaluated in a 10-year N and P fertilization experiment in the Hani peatland in the Changbai Mountains (Bu et al., 2011). To the best of our knowledge, no investigation has evaluated the effects of N and P interactions on the regulation of soil enzymes involved in organic C cycling via the ‘enzymatic latch’ mechanism. We hypothesized that (1) sufficient nutrient supply would enhance POX activity, therefore decreasing [PHEN]; (2) the demand for P by microorganisms would increase in the long-term N fertilized plots due to the possible shift towards P limitation; (3) microorganisms would release more NAG to enhance the acquisition of N and stoichiometrically balance N and P as P deficiency subsided after P application; (4) hydrolase activities would increase owing to lower [PHEN], thereby accelerating soil organic matter decomposition.