Brown spot disease is one of the main foliar diseases endangering the safe production of tobacco leaves in the Changsha tobacco-growing area. Previous studies have shown that Alternaria tenuissima (A. tenuissima) and Alternaria alternata(A. alternata) are the pathogenic fungi of brown spot disease in the Changsha tobacco-growing area. To clarify the biological characteristics of the two pathogens, this study compared the effects of temperature, light, carbon sources, and nitrogen sources on mycelial growth and sporulation. Furthermore, the indoor toxicity of nine chemical agents against the pathogens was investigated, and highly effective agents were selected for field trials. The biological charac-teristics experiments revealed that the optimal growth temperatures for A. tenuissima and A. alternata were 25 °C and 30 °C, respectively. The optimal carbon and nitrogen sources for A. tenuissima were glucose and sodium nitrate, and dark culture promoted sporulation. For A. alternata, the optimal carbon and nitrogen sources were maltose and peptone, and a 12 h dark/12 h light cycle favored sporulation. Indoor toxicity tests indicated significant differences in the toxicity of the nine fungicides against the two Alternaria species. For A. tenuissima, fluazinam exhibited the strongest toxicity (EC?? =1.326 2 mg/L), while thifluzamide was the weakest (EC??=5 121.437 3 mg/L). For A. alternata, fluazinam showed the highest toxicity (EC?? =0.004 8 mg/L), and picoxystrobin was the least effective (EC?? =8 052.050 0 mg/L). Field trial results demonstrated that 40% dimetachlone achieved a control efficacy of 67.70%, followed by 500 g/L fluazinam (66.66%), 32.5% benzoyl-azoxystrobin (54.86%), and 50% boscalid(25.50%). Based on the indoor toxicity and field trial results, alternating applications of 40% dimetachlone, 500 g/L fluazinam, and 32.5% benzoyl-azoxystrobin are recom-mended for chemical control of brown spot disease in the Changsha tobacco-growing area.