Background: Anaerobic soil disinfestation (ASD) has been proven to be an effective and environmentally friendly method for controlling soil-borne plant diseases. Mechanisms of ASD-mediated pathogen suppression are not fully elucidated but appear to depend on the carbon (C) sources used and involve a combination of abiotic and biotic factors. This study sought to assess the impacts of ASD with different C sources on soil chemical properties, microbial activity, and antagonistic compounds, and identify the major factor(s) driving suppression of tomato bacterial wilt caused by Ralstonia solanacearum. Methods: A pot experiment was conducted containing five treatments, i.e., untreated control (CK), anaerobic treatment without C source (ASD-CK), anaerobic treatment with rice bran (ASD-R), wheat bran (ASD-W) and peanut bran (ASD-P). Results: All the ASD treatments significantly reduced disease incidence by 83–100% and simultaneously promoted tomato growth, while anaerobic treatments with C sources achieved a better effect than anaerobic treatment alone. This could be associated with the improved soil chemical (lowered Eh, NO3−, SO42− and elevated pH) and biological (elevated dehydrogenase and urease activities) properties and elevated production of antagonistic compounds (Fe2+, Mn2+, citric acid, succinic acid, and ammonia) by anaerobic treatments with C sources. Redundancy analysis further indicated that the elevated ammonia (11.1%, P = 0.002), Mn2+ (5.3%, P = 0.002), citric acid (1.8%, P = 0.046) and urease activity (1.0%, P = 0.036) were the major factors driving disease suppression, which all achieved the highest value in ASD-P. Conclusions: In summary, the incorporation of organic materials that improve antagonistic compounds (especially ammonia) production could induce higher inhibition effect against tomato bacterial wilt during the ASD process. [ABSTRACT FROM AUTHOR]