Increasing deposition of atmospheric nitrogen (N) due to accelerated human activities is a threat to various ecosystems. However, there is a lack of long-term experimental evidence demonstrating the seasonal dynamics of plant species turnover that ultimately determine species richness in natural ecosystems under N enrichment. Moreover, the frequency of N addition also may affect species turnover in a community, but it is rarely studied.

To assess the responses of a plant community to N addition, we manipulated the amounts (0-50 g N m^-2 year^-1) and frequency (2 vs. 12 times year^-1) of N addition in an Inner Mongolian typical steppe ecosystem in northern China for 12 consecutive years (2008-2020). We measured species richness and density of plant in the growing seasons (May-September) from 2018 to 2020, starting 10 years after the initial N addition treatment.

Both species gain and species loss decreased with increasing amounts of N addition, resulting in a lower plant species turnover rate and greater similarity in the community between two adjacent months throughout the growing season. Species loss and species gain increased modestly under high N addition frequency. Species gain was more important than species loss in determining species richness after a decade of N application. In addition, plant density increased at high N amounts, mainly driven by enhanced clonal growth of the dominant species, Leymus chinensis.

Synthesis. Together, these results suggest that high levels of N deposition may suppress species richness due to aggravated soil chemical properties and may favor growth of a limited number of N-tolerant species compared to systems that experience low levels of N deposition. To conserve biodiversity and to facilitate restoration of degraded grassland ecosystems exposed to long-term N deposition, amelioration of the acidified soils induced by N deposition may be an important strategy to use.