Grassland degradation amplifies the negative effect of nitrogen enrichment on soil microbial community stability.

Although nitrogen (N) enrichment is known to threaten the temporal stability of aboveground net primary productivity, it remains unclear how it alters that of belowground microbial abundance and whether its impact can be regulated by grassland degradation. Using data from N enrichment experiments at temperate grasslands with no, moderate, severe, and extreme degradation degrees, we quantified the temporal stability of soil microbial abundance (hereafter 'microbial community stability') using the ratio of the mean quantitative PCR to its standard deviation over 4 years. Both bacterial and fungal community stability sharply decreased when N input exceeded 30 g N m-2 year-1 in non-degraded grasslands, whereas a reduction in this threshold occurred in degraded grasslands. Microbial species diversity, species asynchrony, and species associations jointly altered microbial community stability. Interestingly, the linkages between plant and microbial community stability were strengthened in degraded grasslands, suggesting that plants and soil microbes might depend on each other to keep stable communities in harsh environments. Our findings highlighted the importance of grassland degradation in regulating the responses of microbial community stability to N enrichment and provided experimental evidence for understanding the relationships between plant and microbial community stability.

Soil fertility shifts the relative importance of saprotrophic and mycorrhizal fungi for maintaining ecosystem stability.

Soil microbial communities are essential for regulating the dynamics of plant productivity. However, how soil microbes mediate temporal stability of plant productivity at large scales across various soil fertility conditions remains unclear. Here, we combined a regional survey of 51 sites in the temperate grasslands of northern China with a global grassland survey of 120 sites to assess the potential roles of soil microbial diversity in regulating ecosystem stability. The temporal stability of plant productivity was quantified as the ratio of the mean normalized difference vegetation index to its standard deviation. Soil fungal diversity, but not bacterial diversity, was positively associated with ecosystem stability, and particular fungal functional groups determined ecosystem stability under contrasting conditions of soil fertility. The richness of soil fungal saprobes was positively correlated with ecosystem stability under high-fertility conditions, while a positive relationship was observed with the richness of mycorrhizal fungi under low-fertility conditions. These relationships were maintained after accounting for plant diversity and environmental factors. Our findings highlight the essential role of fungal diversity in maintaining stable grassland productivity, and suggest that future studies incorporating fungal functional groups into biodiversity-stability relationships will advance our understanding of their linkages under different fertility conditions.

Response of Soil Respiration and Its Components to Warming and Dominant Species Removal along an Elevation Gradient in Alpine Meadow of the Qinghai-Tibetan Plateau.

The Qinghai-Tibetan Plateau is experiencing unprecedented temperature rises and changes in plant community composition owing to global warming. Few studies focused on the combined effects of warming and changes in species composition on soil respiration (Rs). We conducted a 4-year experiment (2015-2018) to examine the influences of warming and dominant plant species removal on Rs and its autotrophic (Ra) and heterotrophic (Rh) components along an elevation gradient (3200, 3700, and 4000 m) for alpine meadow of the Qinghai-Tibetan Plateau. Results showed that warming positively affected Rs, and the stimulation of Rs gradually diminished at 3200 m but remained stable at 3700 and 4000 m as warming progressed. Warming did not influence Ra at all sites. Dominant species removal produced hysteretic behavior that decreased Ra (29%) at 3700 m but increased Ra (55%) at 4000 m in 2018. No significant effect of dominant species removal on Rh was observed. Significant interactive effects of warming and dominant species removal were detected only on Ra at 3700 and 4000 m. Accordingly, under future warming, soil organic matter decomposition at higher elevation will enhance positive feedback to atmospheric CO2 concentration more than that at lower elevation, thus accelerating soil organic carbon loss.

Critical transition of multifunctional stability induced by nitrogen enrichment in grasslands differing in degradation severity.

Nitrogen (N) enrichment poses a severe threat to ecosystem multifunctionality. Given increasing variability of ecosystem functioning and uncertainty under global change, a pressing question is how N enrichment affects temporal stability of multiple functions (i.e., 'multifunctional stability'). Whether the responses of multifunctional stability to N enrichment change with external disturbance, such as grasslands with different degradation statuses, remains unclear. We conducted multi-level N enrichment experiments at four grassland sites with no, moderate, severe, and extreme degradation statuses in Inner Mongolia, China. We measured temporal stability of five functions, comprising aboveground net primary productivity, soil total carbon (C) and N storage, and soil microbial biomass C and N storage, to explore how multifunctional stability responded to N enrichment. The temporal stability of most individual functions and multifunctional stability decreased sharply when N input exceeded 20 g N m-2 y-1 in the non-, moderately, and severely degraded grasslands, whereas the threshold declined to 10 g N m-2 y-1 in the extremely degraded grassland. The relative importance of plant and soil microbes in regulating multifunctional stability varied along the degradation gradient. In particular, plant species asynchrony and species richness showed strong positive relationships with multifunctional stability in the non- and moderately degraded grasslands, whereas soil microbial diversity, especially bacterial diversity, was positively associated with multifunctional stability in the severely and extremely degraded grasslands. Overall, our findings identified a critical threshold for N-induced multifunctional stability and called for context-specific biodiversity conservation strategies to buffer the negative effect of N enrichment on grassland ecosystem stability.

Nitrogen deficiency in soil mediates multifunctionality responses to global climatic drivers.

Natural and anthropogenic processes that decrease the availability of nitrogen (N) frequently occur in soil. Losses of N may limit the multiple functions linked to carbon, N and phosphorous cycling of soil (soil multifunctionality, SMF). Microbial communities and SMF are intimately linked. However, the relationship between soil microbial communities and SMF in response to global changes under N deficiency has never been examined in natural ecosystems. Here, soil samples from nine temperate arid grassland sites were used to assess the importance of microbial communities as driver of SMF to climate change and N deficiency. SMF was significantly decreased by drought and drought-wetting cycles, independent of the availability of soil N. Interestingly, temperature changes (variable temperature and warming) significantly increased SMF in N-poor conditions. However, this was at the expense of decreased SMF resistance. Deterministic assembly-driven microbial α-diversity and particularly fungal α-diversity, but not ß-diversity, were generally found to play key roles in maintaining SMF in N-poor soil, irrespective of the climate. The results have two important implications. First, the absence of the stability offered by ß-diversity means N-poor ecosystems will be particularly sensitive to global climate changes. Second, fungi are more important than bacteria for maintaining SMF in N-poor soil under climate changes.

Nonlinear responses of total belowground carbon flux and its components to increased nitrogen availability in temperate forests.

With the increased interest in allocating more carbon (C) belowground for C sequestration, total belowground C flux (TBCF) and its dynamics have become an important research topic. However, it remains uncertain whether TBCF responds nonlinearly to increased nitrogen (N) availability and how its main components (root and ectomycorrhizal (ECM) fungi) contribute to TBCF. We established a four-year N addition experiment with control, low-N, medium-N, and high-N fertilization treatments in a N-limited temperate forest in northern China. We measured TBCF and its three main components including root respiration, ECM fungal respiration, and root production. The involved edaphic and plant factors were also measured. TBCF showed a nonlinear response to the increasing amounts of N addition, accelerated by 10.37% in low-N addition and restrained by 10.29% in high-N addition. Contrasting patterns of the contributions of root respiration and ECM fungal respiration to TBCF implies different strategies of investment in roots and ECM fungi under the different N-availability statuses. The ratio of production and respiration in roots under N addition was nearly 1:2, which indicated that when soil N availability increases, roots prefer to lose C by overflow respiration rather than fix C in new biomass. The low-N addition increased TBCF by directly increasing root respiration and indirectly increasing coarse root biomass. The medium-N addition positively affected TBCF by increasing root respiration but this positive effect was cancelled out by the significantly negative effect of the increased soil total N concentration. The decrease in soil pH was the most effective pathway to decrease TBCF in high-N addition. Because of a large-scale reforestation program for C sink management in recent years, our findings of the nonlinear response of TBCF to different N fertilization treatments could provide insight for predicting belowground C sequestration potential and its response to atmospheric N deposition.

Selection of suitable reference genes for qRT-PCR normalisation under different experimental conditions in Eucommia ulmoides Oliv.

Normalisation of data, by choosing the appropriate reference genes, is fundamental for obtaining reliable results in quantitative real-time PCR (qPCR). This study evaluated the expression stability of 11 candidate reference genes with different varieties, developmental periods, tissues, and abiotic stresses by using four statistical algorithms: geNorm, NormFinder, BestKeeper, and RefFinder. The results indicated that ubiquitin-conjugating enzyme S (UBC) and ubiquitin-conjugating enzyme E2 (UBC E2) could be used as reference genes for different E. ulmoides varieties and tissues, UBC and histone H4 (HIS4) for different developmental periods, beta-tubulin (TUB) and UBC for cold treatment, ubiquitin extension protein (UBA80) and HIS4 for drought treatment, and ubiquitin-60S ribosomal protein L40 (UBA52) and UBC E2 for salinity treatment. UBC and UBC E2 for the group "Natural growth" and "Total", UBA80 and UBC for the group "Abiotic stresses". To validate the suitability of the selected reference genes in this study, mevalonate kinase (MK), phenylalanine ammonia-lyase (PAL), and 4-coumarate-CoA ligase (4CL) gene expression patterns were analysed. When the most unstable reference genes were used for normalisation, the expression patterns had significant biases compared with the optimum reference gene combinations. These results will be beneficial for more accurate quantification of gene expression levels in E. ulmoides.