Frequency and C:N:P stoichiometry of organic inputs determines intensity of net C balance in paddy soils

The frequency and nutrient composition of organic inputs jointly regulate soil organic carbon (SOC) dynamics, but their interactive effects on microbial carbon use efficiency (CUE), priming effects (PE), and net soil C balance remain poorly understood in flooded paddy systems. We performed a 40-day incubation experiment using a 13C-labeled simulated root-exudate mixture (glucose:oxalic acid:alanine, 65:30:5) under two input modes (single substrate input vs. continuous substrate input) and four C:N:P stoichiometries. Single substrate inputs generated an early pulse in labile-C mineralization that was 39–64 % greater than under continuous addition, and mineralization rates declined with increasing nutrient supply. Single-pulse addition triggered an early peak in the metabolic quotient (qCO2) and lower tracer-based CUE, whereas continuous addition maintained steadier microbial activity and higher CUE. The C:N:P stoichiometry of the added substrate strongly controlled C partitioning: stoichiometrically balanced inputs reduced CO2–C losses and increased 13C incorporation into microbial biomass and SOC pool. Pulse inputs typically induced negative PEs, whereas continuous inputs tended to cause positive PEs. Therefore, the net C balance was consistently greater following single substrate inputs than following continuous inputs; across nutrient treatments, single pulses produced substantially larger short-term C retention. Combining 13C tracing, enzyme assays and kinetic modelling, we demonstrate that under balanced nutrient inputs, microbes respire less of the added C and allocate more into biomass and necromass, which subsequently contributes to more stable SOC pool. This study provides mechanistic guidance for using C:N:P-balanced amendments to increase SOC retention in flooded cropping systems. © 2025 Elsevier Ltd.