Effects of Long-Term Combination of Organic and Inorganic Fertilizers on Bacterial Community Structure, Ecological Network, and Key Species in Fluvo-Aquic Soil

Based on the long-term fertilization station in fluvo-aquic soil region, this study investigated how bacterial community structure, ecological network, and key species in response to fertilization.

Based on the long-term experiment started from 1990, soil was sampled after wheat maturity in 2023, and high-throughput sequencing and ecological network analysis were used to examine dynamics in soil organic carbon (SOC) and other nutrients, enzyme activity, bacterial community composition, ecological network and stochastic process under four treatments: no fertilization (CK), mineral nitrogen (N), phosphorus (P), and potassium (K) (NPK), NPK+straw (NPKS), and NPK+manure (NPKM).

The combined application of organic and inorganic fertilizers significantly increased SOC and other soil nutrients, as well as soil enzyme activity. Compared with CK, SOC content under NPKS and NPKM increased by 52.1% and 81.9%, respectively, and particulate carbon increased by 60.6% and 137.4%, respectively, and easily oxidized organic carbon increased by 45.3% and 63.4%, respectively; additionally, β-N-acetylglucosaminidase activity increased by 7.2% and 12.6%, respectively, while alkaline phosphatase activity increased by 166.4% and 216.2% (P<0.05), respectively. Notably, β-1,4-glucosidase activity was the highest under NPKS (63.82 μmol·g^-1·d^-1). In terms of bacterial diversity, α-diversity significantly decreased under NPKS compared with CK, with reductions of 5.4%, 5.2%, and 2.6% in the Ace, Chao1, and Shannon indices, respectively (P<0.05). Fertilization treatment alerted bacterial community structure, while NPKS and NPKM exhibited similar compositions. Compared with CK, NPKS significantly reduced the relative abundance of Chloroflexi, Gemmatimonadota, and Methylomirabilota, while NPKM significantly increased the relative abundance of Bacteroidota (P<0.05). Redundancy analysis identified ammonium nitrogen (NH₄⁺-N), nitrate nitrogen (NO₃^--N), and organic carbon (SOC) as the primary environmental factors shaping microbial community structure. Network analysis showed, compared with CK, NPKM increased the complexity, stability of the bacterial community network and the proportion of positive correlations between species. Furthermore, both NPKS and NPKM significantly enhanced the relative abundances of eight keystone taxa, including members of Actinobacteriota (order Microtrichales), Chloroflexi (order Thermomicrobiales), Bacteroidota (orders Chitinophagales and Cytophagales), Myxococcota (uncultured order bacteriap25), and Proteobacteria (order Burkholderiales) (P<0.05). These keystone taxa were closely associated with soil carbon, nitrogen, phosphorus, and other material cycling, as well as plant growth promotion. Partial least squares path modeling suggested that fertilization did not directly impact key species but exerted an indirect influence by significantly affecting soil pH, regulating community composition, and increasing soil nutrient availability.

Long-term combined application of organic and inorganic fertilizers enhanced soil nutrient content and extracellular enzyme activity, regulated microbial community composition and structure, affected bacterial network complexity and stability, and increased the relative abundance of key species involved in soil nutrient cycling and material transformation. These findings provided valuable insights into the interactions between soil properties and microbial communities under long-term fertilization, contributing to a deeper understanding of bacterial community dynamics and key species in agricultural ecosystems.