The utilised primer sequences.

Respiratory infections remain a leading cause of morbidity and mortality, necessitating new therapeutic strategies. This study evaluated the antiviral and antibacterial activities of harmine, harmaline, and harmalacidine hydrochloride against H1N1 influenza virus and Staphylococcus aureus, key respiratory pathogens. The in vitro antiviral activity of the tested compounds against the H1N1 virus was evaluated using a plaque assay. Harmalacidine hydrochloride demonstrated notable activity, with an IC50 of 68.2 ± 0.8 µg/mL, while harmine and harmaline showed no significant effects at non-cytotoxic concentrations. The potential antibacterial action of the tested compounds was initially investigated by agar well diffusion method, which revealed clear zones of inhibition around the wells. Subsequently, their minimum inhibitory concentrations (MICs) were recorded using the broth microdilution method. Harmalacidine hydrochloride exhibited the highest antibacterial action with MICs from 16 to 128 µg/mL. Based on these findings, further investigations were conducted to assess the effect of harmalacidine hydrochloride on membrane integrity and permeability, cellular morphology, and biofilm formation. A noticeable reduction (p p AgrA) of S. aureus, suggesting antibacterial activity through inhibition of quorum sensing-mediated virulence. It also showed high affinity for H1N1 neuraminidase and polymerase basic protein 2 (PB2), indicating potential antiviral activity. However, experimental enzyme assays and in vivo studies are required to confirm the proposed antiviral and antibacterial mechanisms.