Microbial metabolic limitation regulated by ecological stoichiometry in croplands

Carbon (C), nitrogen (N), and phosphorus (P) stoichiometry (C:N:P) profoundly influence material cycling and energy flow in terrestrial ecosystems by regulating nutrient allocation in plant–microbe–soil systems. Despite strong coupling between soil resources and microbial metabolism, multidimensional stoichiometric characteristics and their interrelationships remain poorly quantified, and the long-term effects of fertilization across climatic gradients are still unclear. Here, based on six long-term (27–44 years) fertilization experiments spanning ~17° of latitude from north to south, we examined no fertilizer, mineral fertilizer, and mineral plus organic fertilizer treatments, covering soil stoichiometry, available resource stoichiometry, microbial stoichiometry, and enzyme stoichiometry. Integrating ecological enzyme stoichiometry theory with metagenomics, we explored how fertilization and climate jointly regulate microbial metabolism via stoichiometric changes. Results showed that strong linkages existed among different stoichiometric types, and that both fertilization and climate significantly altered stoichiometric characteristics of soil, resources, microorganisms, and enzymes, with notable site-specific patterns. Compared to the control (CK), fertilization led to a more intricate stoichiometric network. Fertilization enhanced microbial C and P limitations at most experimental sites, and increasing temperature and precipitation shifted microbial N limitation toward P limitation. Microbial C limitation was mainly regulated by available resources and enzyme stoichiometry, whereas N/P limitation was driven by available resources and microbial stoichiometry. Overall, our findings highlight the critical role of multiple stoichiometric dimensions in regulating microbial metabolism and provide a theoretical basis for understanding and managing C, N, and P cycling in trans-regional agroecosystems.