Long-term manure inputs differentially regulate microbial carbon and nitrogen resource allocation strategies to achieve co-benefits for crop yield and soil carbon sequestration in low fertility croplands

Enhancing both crop productivity and soil organic carbon (SOC) in low fertility croplands poses a critical challenge for sustainable agriculture. Manure input has been proposed as a strategy to achieve these dual goals, with soil microorganisms acting as the key drivers in carbon and nitrogen cycling. However, microbial activities in these soils are constrained by resource limitation and stoichiometric imbalances. The underlying mechanisms through which manure stimulates microbial physiological responses to mediate these co-benefits remain poorly understood. A 16-year field experiment integrating SOC fractionation, enzymatic stoichiometry, and stoichiometric modeling was conducted to quantify soil microbial resource limitations, extracellular enzyme activities, carbon and nitrogen use efficiencies (CUE and NUE), and their regulatory relationships with SOC fractions and crop yield under three treatments: chemical fertilizer (FP), replacing 50% chemical N with manure N (RMN), and completely replacing chemical N with manure N (M). The results indicated that all fertilization treatments increased crop yield, but only the RMN and M treatments enhanced SOC sequestration, with rates of 0.5±0.1 and 0.7±0.0 Mg ha⁻¹ yr⁻¹. Specifically, compared to the FP treatment, the accumulation of dissolved organic carbon (DOC), microbial necromass carbon (MNC), and mineral-associated organic carbon (MAOC) increased by 14.5% in RMN and 127.8% in M, 70.4% in RMN and 183.9% in M, 73.0% in RMN and 161.2% in M. Partial least squares structural equation modeling further indicated that microbial resource allocation strategies adapted to alleviated nitrogen limitation and increased carbon demand are the central mechanisms. These are manifested as reduced investment in C- and N‑acquiring enzymes, with decreased NUE accompanied by increased CUE. These strategies are associated with the manure‑induced expansion of soil total dissolved nitrogen (TDN) pool and DOC pool. These was associated with increased root nitrogen uptake and was associated with carbon flow towards microbial biomass and necromass, thereby further enhancing the formation of MAOC and total SOC. Notably, the high CUE leads to a trade‑off between particulate organic carbon (POC) retention and MAOC formation. These findings suggest that long‑term manure input is associated with differential regulation of microbial carbon‑nitrogen resource allocation strategies, alleviates microbe‑resource stoichiometric imbalance, and consequently enhances SOC sequestration and optimizes nitrogen supply, offering a framework for the sustainable management of low‑fertility or degraded soils to improve soil quality and productivity.