Nitrogen and Water Regulate the Soil Microbial Carbon Cycle in Wheat Fields Primarily via the Pentose Phosphate Pathway

To clarify how nitrogen (N) and water regulate the microbe mediated carbon (C) cycle in farmland, a 3-year experiment was conducted in a wheat–maize rotation at Jiaozhou Station, North China. Twelve treatments combined four drip irrigation regimes (T1: no irrigation; T2: 40 mm irrigation at flowering; T3: 40 mm irrigation at the grain filling stage; T4: both, 40 mm each) and three N levels (N0: 0 kgN·hm−2; N1: 92 kgN·hm−2; N2: 184 kgN·hm−2). In this study, we measured wheat yield and biomass, soil organic carbon and nitrogen content, soil respiration, soil microbial community, and C-metabolic genes. The results showed that wheat yield increased with N, peaking at 8949.81 kg·hm−2 in the N2T3 treatment, while irrigation had no significant independent effect on yield but interacted with nitrogen fertilization: under identical nitrogen levels (N1, N2), yields in the T1 and T2 treatments were significantly lower than those in the T3/T4 treatments. The soil organic carbon content in N2 was significantly higher; the soil C/N ratio was highest in N2, and T3 resulted in a significantly higher C/N ratio than T1 under the same N level; total soil respiration in N0 was significant lower, and T4 had higher respiration than T2 under the same N level. N addition increased Actinobacteriota, Chloroflexi, Thaumarchaeota, and Ascomycota, while decreaing Proteobacteria and Acidobacteriota. No reduction in fungal phylum was observed with nitrogen addition. N application significantly upregulated key enzymes in the pentose phosphate pathway (e.g., transketolase K00615, transaldolase K00616), while irrigation increased phosphoserine aminotransferase (K00831) abundance and decreased methylmalonyl-CoA mutase (K01848) abundance. N2T3 maintains high SOC content while achieving maximum yield, promoting soil fertility retention. Compared to T4, N2T3 also enhances water use efficiency. The N2T3 treatment (high N and grain filling stage irrigation) achieved the optimal balance between high wheat yield and SOC sequestration.