Long-term manure application enhances carbon use efficiency in soil aggregates by regulating microbial communities in cropland

Abstract

Microbial carbon use efficiency (CUE), a key metric reflecting the allocation of carbon between microbial growth and respiration, plays a central role in predicting soil carbon (C) dynamics. Microbial CUE is influenced by soil aggregates that create nutrient heterogeneity, and long-term fertilization may alter these heterogeneous microsites. However, the effects of long-term fertilization on microbial CUE across different aggregates remain unclear, limiting mechanistic understanding of soil C cycling at the microscale. Here, a 34-year field experiment was conducted with five treatments including no fertilizer (CT), mineral nitrogen (N), mineral N, phosphorus and potassium (NPK), manure-only (M), and combined mineral fertilizer and manure (NPKM). Microbial CUE was measured in bulk soil and in >2 mm, 0.25–2 mm, and <0.25 mm aggregates using an ¹⁸O-H₂O labeling approach. Metagenomic sequencing, enzyme activity assays, and substrate analyses were integrated to examine how fertilization altered interactions among aggregates, substrate quality, and microbial communities affecting CUE. Results showed that, compared to CT, manure application (M and NPKM) significantly increased microbial CUE both in bulk soil and aggregates. While microbial activity and CUE differed among aggregates under CT and NPK, manure application homogenized CUE across aggregates. Further analysis revealed that manure application regulated enzyme activities and substrate quality, which affected key bacterial modules (M11 and M14) and consequently modulated microbial CUE. These findings underscore the role of manure in enhancing microbial CUE and suggest that accounting for manure-induced microscale homogenization can improve predictions of SOC dynamics in global C models.