Long-term mono-cropping and excessive chemical fertilizer application reduce water-stable soil aggregate stability and soil N and C stocks in tobacco production. This study tested the hypothesis that rotation and fertilizer type affected the proportion of water-stable aggregates and aggregate-associated soil organic carbon (SOC) and total soil nitrogen (TSN) concentrations. Two planting systems (tobacco mono-cropping and tobacco-maize rotation) with five fertilizer treatments (0 and 75 kg N ha −1 , 450 kg oil cake + 75 kg N ha −1 , 15,000 kg farmyard manure + 60 kg N ha −1 , and 3000 kg straw + 75 kg N ha −1) were established in 2007. After 10 years, rotation and fertilizer type significantly affected soil aggregates and associated SOC and TSN at different soil depths. Rotation significantly increased mean weight diameter (MWD) and geometric mean diameter (GMD); GMD increased with increasing soil depth. Rotation and organic fertilizer effectively increased the proportion of large macro-aggregates and MWD and GMD, while significantly reducing the proportion of micro-aggregates. The effect of fertilization type and rotation on the SOC content of large macro-aggregates was more obvious than that on TSN content, but there was no significant effect on SOC and TSN contents of aggregates < 250 μm. Rotation and organic fertilizer significantly increased SOC stock and total nitrogen of soil macro-aggregates (> 250 μm). However, this trend weakened with increasing soil depth and decreasing aggregate size. Rotation and organic fertilizers used in tobacco production stabilized the proportion of macro-aggregates in surface soil and maintained SOC and TSN better. Keywords Flue-cured tobacco. Rotation. Organic fertilizer. Water-stable aggregate. Yunnan Abbreviations SOC Soil organic carbon TSN Total soil nitrogen BD Bulk density MWD Mean weight diameter GMD Geometric mean diameter SOM Soil organic matter SOCS Soil organic carbon stock TSNS Total soil nitrogen stock * Congming Zou