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In greenhouse vegetable production, reductive soil disinfestation (RSD) effectively mitigates soil-borne diseases, yet its impact on the dynamics of soil organic carbon (SOC) has not been adequately examined. This study investigated the distribution of soil aggregates and the organic carbon retention mechanism following intensive RSD treatment. Greenhouse experiments, including control (CK), wheat straw (RSD), and wheat straw with chemical fertilizer (RSD + NP) treatments, indicated augmentation in the formation of macro-aggregates (>2 mm and 0.25–2 mm) under RSD, particularly in the RSD + NP treatment. Silty clay particles transform into macro- and micro-aggregates. Fourier infrared spectroscopy highlighted the augmentation of carbon-containing functional groups in SOC, with aliphatic carbon accumulating in macro-aggregates and aromatic carbon in silt clay. Laboratory culture experiments employing different C/N ratios (RSD1 with wheat straw, RSD2 with kiwifruit branches) underscored the beneficial impact of low C/N ratio organic matter on coarse and fine macro-aggregate content, mean weight diameter, geometric mean diameter, and reduced silt clay. Low C/N ratios enhance SOC retention in large aggregates, while high ratios stabilise micro-aggregate carbon. This research underscores the severe degradation in continuous greenhouse cropping systems and emphasises RSD’s dual benefits—disease prevention and improved SOC retention. Implementing RSD requires careful consideration of organic material choices, specifically their C/N ratios, as a pivotal factor influencing SOC dynamics.