Coupled steel slag and biochar amendment correlated with higher methanotrophic abundance and lower CH₄ emission in subtropical paddies

Aerobic methanotrophs in paddies serve as methane (CH₄) filters and thereby reduce CH₄ emissions. Amending soil with waste products can mitigate CH₄ emissions in crops, but little is known about the impacts of amendments with steel slag and biochar on the populations and activities of aerobic methanotrophs in rice cropland. We used real-time quantitative PCR detecting system and high-throughput sequencing to determine the effects of slag and biochar amendments on CH₄ emission, abundance, and community structure of methanotrophs, and the relationships between soil properties and the abundance and community composition of methanotrophs during the rice growing season in both early and late paddies. Soil salinity and pH were significantly higher for an amendment with both slag and biochar than the control in both the early and late paddies, and pH was significantly higher for a slag amendment in the late paddy. Cumulative CH₄ emission was lower for the slag and slag + biochar amendments than the control in early paddy by-34.1%. Methanotrophic abundance was three- and sixfold higher for the slag + biochar amendment than the control in the early and late paddies (p < 0.05), respectively. The abundance of different groups of methanotrophs varied among the treatments. The relative abundance of Methylosarcina was higher for the slag amendment than the control, and the relative abundance of Methylomonas was lower for biochar, and slag + biochar amendments than the control. The relative abundance of Methylocystis was higher for the slag and slag + biochar amendments than the control in the early paddy, and the relative abundance of Methylocystis was higher for the slag, biochar, and slag + biochar amendments in the late paddy. Univariate and multivariate analyses indicated that the higher abundance of methanotrophic bacteria for the slag and slag + biochar amendments was correlated with soil pH, salinity, soil organic carbon, and C/N ratio, and the relative abundances of Methylocystis, Methylomonas, and Methylosarcina were associated with the effective mitigation of CH₄ emission in the paddies. A discriminant general analysis indicated that the total population of methanotrophs was larger for the slag + biochar amendment than the control, and that this effect was only weakly correlated with changes in the soil properties, demonstrating that this effect on the size and species composition of methanotrophic soil populations was mostly associated with a direct effect of the slag + biochar amendmen