Tobacco black shank is a kind of soil-borne disease caused by the Oomycete Phytophthora parasitica. This disease is one of the most destructive diseases to tobacco (Nicotiana tabacum L.) growth worldwide. At present, various measures have been taken to control this disease, but they still have different challenges and limitations. Studies have shown that β-aminobutyric acid (BABA), a nonprotein amino acid, can enhance disease resistance in plants against different varieties of pathogens. However, it is unclear whether BABA can induce plants to resist Phytophthora parasitica infection. Therefore, this study aims to explore the effect and related mechanism of BABA against tobacco black shank. Our results showed that 5 mmol.L-1 BABA had an obvious anti-inducing effect on the pathogenic fungus and could effectively inhibit the formation of dark spots in the stems. The results also showed that a large amount of callose deposition was observed in BABA-treated tobacco. Furthermore, the application of BABA induced the accumulation of H2O2 in tobacco and effectively regulated the homeostasis of reactive oxygen in tobacco plants, reducing the toxicity of H2O2 to plants while activating the defense system. In addition, BABA spray treatment could induce an increase in the concentrations of salicylic acid (SA) and jasmonic acid-isoleucine (JA-Ile) in tobacco, and the gene expression results confirmed that BABA upregulated the expression of SA-related genes (PR1, PR2 and PR5), JA-related genes (PDF1.2) and ET-related genes (EFE26 and ACC oxidase) in tobacco plants. Taken together, BABA could activate tobacco resistance to black shank disease by increasing H2O2 accumulation, callose deposition, plant hormone (SA and JA-Ile) production, and SA-, JA-, and ET- signaling pathways.