Abstract: To acquire entomopathogenic fungi with high pathogenicity against the tobacco beetle (Lasioderma serricorne) larvae, cadavers of naturally infected L. serricorne and the tobacco moth (Ephestia elutella) were collected from tobacco storage facilities. Entomopathogenic fungi were isolated and purified via the tissue separation method, and their taxonomic positions were clarified based on morphological characteristics and homology analysis of the rDNA-ITS region sequences. The growth rate and sporulation yield of the isolated strains were determined using the cross-measurement method and hemocytometer counting method, respectively. Finally, the bioactivity of each isolated strain against L. serricorne larvae was assayed in vitro by the dipping method, and their biocontrol potential was evaluated. Results indicated that 15 strains (Bb001-Bb015) of Beauveria bassiana were isolated and identified. When cultured on SDAY medium at 26 ℃ and 80% relative humidity for 10 days, the growth rates of these strains ranged from 2.59 to 4.59 mm·d–1, and the sporulation yields varied from 1.54×107 to 5.75×107 spores·cm–2. Bioactivity assay results showed that at a concentration of 1×108 spores·mL–1, the corrected mortalities of L. serricorne larvae reached 91.38% and 84.48% at 10 days post-inoculation. Gradient dose tests revealed that larval mortality increased with the elevation of spore concentration, and the survival time of L. serricorne larvae inoculated with Bb001 was shorter than that of those inoculated with Bb013 at all tested concentrations. The median lethal concentration (LC50) of Bb001 was 4.52×10⁴ spores·mL–1, which was lower than that of Bb013 (1.02×105 spores·mL–1). In conclusion, B. bassiana strains Bb001 and Bb013 exhibit high pathogenicity against L. serricorne larvae, with Bb001 demonstrating superior efficacy. These strains can serve as potential fungal resources for the biological control of L. serricorne larvae.