The anti-respiratory syncytial virus activity of biochemicals from Pyrola incarnata.

Pyrola incarnata, a medicinal herb and functional food with diverse bioactive compounds, was systematically screened for in vitro anti-RSV activities of its constituents, combined with antiviral assays and computational analyses. Among ten compounds, pyrolin, gallic acid, and quercetin as main phytochemicals showed the most potent efficacy, with IC50 values of 0.56 ± 0.08 μM, 10.60 ± 1.08 μM, and 12.63 ± 1.00 μM, respectively, outperforming positive control ribavirin. Indirect immunofluorescence and high-content analysis indicated these three compounds target the early RSV replication cycle. Gallic acid showed inhibited RSV infection during virus pretreatment and adsorption, with inhibition rates of 72.79% and 48.25% (P < 0.01), respectively. Furthermore, gallic acid suppressed early post-infection replication, and mechanistically it disrupted RSV-F protein mediated membrane fusion, reducing cell-cell fusion to 22.52%, consistent with docking results. Moreover, exploratory network pharmacology suggests that gallic acid might exert anti-RSV effects by modulating targets such as MAPK, AKT1, and AKT2. In additional, it was discovered for the first time that pyrolin as the one of the component indicators from P. incarnata, markedly inhibited RSV infection during virus pretreatment and adsorption, with inhibition rates of 94.50% and 43.50% (P < 0.01) respectively. While quercetin also exhibited direct virucidal activity. In conclusion, P. incarnata acts as promising natural inhibitors targeting the early stages of RSV replication. These findings establish a scientific basis for future in vivo validation and the development of new drugs against respiratory syncytial virus.