Purpose <p>To determine the optimal agricultural management practice for improving soil health, this study examined the effects of straw incorporation on dissolved organic carbon (DOC) fractions and microbial community structures in paddy soil.</p> Materials and methods <p>The experiment was conducted over six rice growing seasons (6 year). Three treatments were established: annual on-site rice straw incorporation at 8 t ha^− 1 yr^− 1 (FR, full-rate straw incorporation), straw incorporation at 4 t ha^− 1 yr^− 1 (HR, half-rate straw incorporation), and the control (CK, no rice straw addition). Measured soil parameters included total organic carbon, total nitrogen, DOC, metagenomic profiling and DOC fluorescent components.</p> Results <p>The results showed that both HR and FR significantly increased the maximum fluorescence intensity (Fmax), with the C4 component particularly enhanced by 0.88 units in HR and 1.47 units in FR, and the C5 component increased by 0.73 units in FR relative to the CK (<i>P</i> &lt; 0.05). Straw incorporation promoted an elevation in the fluorescent components of DOC, especially the stable C4 and C5 substances, which exhibit stronger carbon sequestration potential. Additionally, straw incorporation facilitated the proliferation of specific nitrogen-fixing bacteria and lactic acid bacteria, such as <i>Leuconostoc</i> and <i>Lactobacillus brevis</i>. These probiotics participate in soil nutrient cycling, improve animal gut health, enhance immune systems, and even inhibit certain infectious and parasitic diseases. The results also revealed that environmental factors, including soil pH and organic matter content, significantly influenced the structure and functional dynamics of viral communities.</p> Conclusions <p>These findings demonstrate that full-rate straw incorporation enhances soil fertility and microbial community composition, modulates soil viruses’ diversity, and facilitates the development of a sustainable soil ecosystem.</p>