A Marine Bivalve (Entodesma navicula) Extract Suppresses the Replication of Influenza Viruses and Flaviviruses in Association with Reduced MLKL Phosphorylation

Emerging influenza viruses and mosquito-borne flaviviruses continue to challenge animal and public health, highlighting the need for antivirals that remain effective after infection. This study evaluated the in vitro activity of a 70% ethanolic whole-specimen extract of the marine bivalve Entodesma navicula (ENE) against influenza A/Puerto Rico/8/1934 (H1N1; A/PR8), A/Wisconsin/15/2009 (H3N2), influenza B/Florida/78/2015, Zika virus, and dengue virus serotype 2. Antiviral activity was assessed using time-of-addition analysis, viability-based cytopathic-effect assays, reverse-transcription quantitative PCR, infectious virus titration, immunofluorescence, Western blotting, and flow cytometry. In the A/PR8 model, pre- and co-treatment produced no consistent inhibition. By contrast, post-infection ENE suppressed all tested viruses. For A/PR8, Zika virus, and dengue virus, this activity was supported by reduced viral genome abundance, infectious progeny, and viral antigen expression. In A/PR8-infected A549 cells, ENE reduced phosphorylation of receptor-interacting serine/threonine-protein kinase 3 (RIPK3) and mixed lineage kinase domain-like pseudokinase (MLKL). ENE also reduced the proportion of cells positive for phosphorylated MLKL within Zika- and dengue-virus-infected Vero E6 populations. These findings identify E. navicula as an underexplored source of post-infection antiviral bioactivity associated with attenuated necroptosis-related signaling. Isolation of active constituents and causal validation in primary cells and animal models are needed.