Hexahydroacridine derivatives as novel anti-Salmonella agents: In vitro, In vivo, and In silico studies

The rise of multidrug-resistant (MDR) Salmonella enterica presents a critical global health concern and highlights the need for new antibacterial agents. This study investigates the activity of the hexahydroacridine derivatives D01 and D04 using a comprehensive set of in vitro, in vivo, and in silico approaches. Both compounds exhibited strong bactericidal effects against MDR S. enterica, with D04 showing superior potency, achieving a ≥6 log₁₀ reduction in CFU/mL within 24 h at 2 × MIC. D04 also reduced biofilm formation by 95% at 2 × MIC. Synergistic interactions with ciprofloxacin and tetracycline were observed for both derivatives. Serial passage experiments demonstrated stable MIC values, indicating minimal resistance development. In vivo assays using zebrafish and Galleria mellonella confirmed enhanced survival and decreased bacterial burden following treatment. Histopathological evaluation of murine intestinal tissue showed preserved epithelial integrity in treated groups. qPCR analysis revealed downregulation of virulence- and biofilm-associated genes and upregulation of stress response genes. Molecular docking studies further supported strong binding of D01 and D04 to the S. enterica InvA protein. Collectively, these results identify D01 and D04 as promising antibacterial and anti-virulence candidates with low resistance potential and favorable host compatibility.