Long-term fertilizer application alters microbial resource limitations in soil aggregates via nutrient and microbial resource allocation

Aims: Ecological stoichiometry reflects the stability of resources in farmland ecosystems. Long-term fertilization modulates soil nutrient resources in different soil aggregate sizes. However, how fertilization regimes alter nutrient resource limitations in different soil aggregate sizes remains unclear. Methods: We measured the activity of extracellular enzymes (EEAs) involved in the acquisition of nutrients such as soil carbon (C), nitrogen (N), and phosphorus (P), in conjunction with soil extracellular enzyme stoichiometry (EES) to reflect microbial resource demands. Four different fertilizing strategies (no fertilization, chemical fertilizer, chemical fertilizer plus cow manure, chemical fertilizer plus rice straw) were evaluated. Results: Applying fertilizer changed the size distribution of soil aggregates. The results from the EES model and vector analysis indicated that the application of cow manure could effectively alleviate carbon and nitrogen resource limitations (p < 0.05), as evidenced by changes in soil C: N, C: P, and N: P ratios. Microbial carbon limitation in microaggregates was greater than that in other soil aggregate sizes, whereas microbial nitrogen limitation in macroaggregates was the greatest among all treatments. Using random forest and partial least squares path models, we found that fertilization and soil aggregate size indirectly affected carbon limitation by influencing microbial biomass. Conclusions: Fertilization and soil aggregates not only affected nitrogen limitation directly but also indirectly by influencing nutrient resources, especially, the C: N ratio. Above all, in agricultural production, we can alleviate the nutrient limitations in the soil by adjusting the ratio of nutrients in fertilizers, providing an in-depth understanding for designing fertilizer formulations and appropriate fertilizing strategies.