Litter amendment rather than phosphorus can dramatically change inorganic nitrogen pools in a degraded grassland soil by affecting nitrogen-cycling microbes

Phosphorus fertilisation and increasing litter input are widely employed to restore the degraded grasslands. Despite the key roles of nitrogen-cycling microbes in determining the soil nitrogen dynamics and development of grassland degradation, little is known about their responses to these restoration efforts. Here, a microcosm experiment, with soils collected from a degraded Tibetan alpine meadow, was conducted to investigate the responses of nitrogen-cycling microbes to litter and phosphorus amendments, and their links with the changes in soil properties. Copies of the corresponding nitrogen-cycling genes (nifH, amoA, narG, nirK, and nirS genes) and their mRNAs were determined using real-time PCR. The results showed that the litter amendment significantly stimulated the transcription of nifH and nirS genes, but reduced the copies of amoA gene and bacterial amoA mRNA. It also significantly increased soil dissolved organic carbon, available phosphorus, ammonium nitrogen, and microbial biomass concentrations, but decreased soil inorganic and nitrate nitrogen concentrations. The phosphorus amendment exerted little effects on soil properties and nitrogen-cycling microbes, while the litter phosphorus interactions significantly offset the individual negative effects of the litter and phosphorus amendments on the denitrifier abundance. The soil nitrate and inorganic nitrogen concentrations were positively correlated with the amoA genes and bacterial amoA mRNA copies, but negatively correlated with nirS mRNA copies. These results indicate that litter addition may decrease soil nitrate and inorganic nitrogen concentrations by suppressing nitrifiers and stimulating nirS gene expression, highlighting the vital roles of nitrogen-cycling microbes in determining the soil nitrogen dynamics during the restoration of degraded grasslands.