Labile carbon input alleviates nitrogen-induced community instability in a meadow steppe

Global nitrogen (N) deposition continues to threaten plant diversity and ecosystem stability despite a recent slowdown in its increasing rates. Labile carbon (C) may help reduce excess N by alleviating microbial C starvations, but their role in mitigating the harmful effects of N enrichment remains unclear.

In a meadow steppe in northern China, we conducted a 9-year (2014-2022) field experiment with six levels of historical N addition (0, 2, 5, 10, 20, and 50 g N m–2 yr–1, 2014-2019) and three levels of labile C (0, 200, and 2000 g C m-2 yr-1).

Three years after ceasing N treatments (2020-2022), aboveground net primary productivity (ANPP) remained high under N addition. However, species richness and community stability continued to decline with increasing N addition rates. Labile C addition reduced the dominance of certain plant species within the community while it enhanced species asynchrony and belowground net primary productivity (BNPP). Boosted regression tree models indicated that the high levels of labile C inputs improved community stability by enhancing BNPP, which increased the relative importance of BNPP to the community stability from 7.5% to 27.4% as labile C input rose.

Synthesis. Our results highlight how labile C inputs can counteract the negative impacts of N enrichment on community stability via enhancing plant-microbe competition and increasing belowground biomass allocation.

Methods
Aboveground biomass was collected annually in mid-August (2020 to 2022) by clipping all vascular plants within a 1 × 1 m quadrat from a 2 × 2 m subplot. Biomass was sorted by species, separated from litter and dead standing biomass, and then weighed after being oven-dried at 70 °C for at least 48 h to estimate ANPP. Species richness was calculated as the total number of species per quadrat.

Belowground net primary productivity (BNPP) was measured using the root ingrowth core method (Wang et al., 2019a; Yang et al., 2022). At the end of the growing season in mid-September 2019, a single core (7 cm in diameter and 50 cm deep) was vertically drilled into the soil in each subplot. The soil was sieved to 2 mm to remove roots, debris, and rocks. A polyester mesh bag (mesh size 2 mm) was placed over a PVC tube (7 cm in diameter and 50 cm length) and inserted into the holes. The PVC tube was then slowly pulled out while the hole was filled with root-free sieved soil, which was compressed to match the original soil density.

In mid-September from 2020 to 2022, mesh bags were carefully pulled out from the holes, and the soil was sieved (mesh size 2 mm) to collect roots from each year. The screened soil was then returned to the polyester mesh bag and reinserted into the original hole. All root samples were washed, dried at 70 °C for 48 h, and weighed. BNPP was calculated based on the average root biomass from the ingrowth core in each subplot over the years 2020 to 2022.

During the growth period from 2020 to 2022, the leaves of L. chinensis were collected from all subplots, and the total N concentration of leaf samples was determined using an element analyzer (vario El III, Elementar Analysensysteme GmbH, Hanau, Germany) after grinding with a ball mill.

Following the biomass harvests in mid-August each year from 2020 to 2022, soil samples were collected using a soil auger (3 cm in diameter). The five-point method was employed to collect soil samples from the 0-10 cm depth, which were then combined to obtain a single homogenized composite sample per subplot. These samples were passed through a 2-mm sieve to remove visible plant roots before analyzing soil parameters, including soil pH, microbial biomass C (MBC), microbial biomass nitrogen (MBN), soil ammonium (NH4+-N) and nitrate (NO3−-N) concentrations. MBC and MBN were extracted using a 0.5 M K2SO4 solution following the chloroform fumigation extraction method (Lovell et al., 1995), and analyzed using an elemental analyzer (Liqui TOC, Analysensysteme, Germany). Soil NH4+-N and NO3−-N concentrations were extracted with a 2 M KCl solution and quantified using a continuous flow injection analyzer (FIAStar, Foss Tecator, Sweden).