The pinewood nematode (PWN, Bursaphelenchus xylophilus) is a devastating forest phytoparasite responsible for pine wilt disease, that poses a catastrophic ecological and economic threat to conifer forests worldwide. The limitations of current management strategies have been underscored by the recent 2025 PWN outbreak in France, emphasizing that traditional containment protocols must be improved. While conventional chemical control methods are effective, their environmental footprint, persistent toxicity, and risks to human health and non-target organisms highlight an urgent global demand for safer, sustainable alternatives. Phytochemicals - specifically those derived from bioactive essential oils - have emerged as highly promising biopesticides. Although some plant volatiles are already authorized as plant protection products within the European Union (e.g., geraniol, thymol), their specific application against PWN remains largely unexplored. In this study, a comprehensive in vitro screening of over 200 volatiles, essential oils, and essential oil fractions was conducted to evaluate their nematicidal potential against the PWN. Notably, complete PWN mortality was recorded for many of the tested compounds at a concentration of 1 mg/mL. Furthermore, over 40 of these candidates showed superior efficacy compared to the standard reference commercial nematicide, emamectin benzoate. The most potent candidates - predominantly oxygenated derivatives of medium-chain fatty acids - exhibited exceptionally rapid action achieving an up to 40-fold faster activity over the conventional treatment. To ensure these compounds align with European conservation goals, ecotoxicological data was paired with computational predictive modelling. These safety assessments demonstrated favourable environmental profiles, indicating significantly lower toxicity risks to non-target species compared to synthetic nematicides. By successfully bridging experimental screening with predictive computational frameworks, this work proposes a robust pipeline for developing high-potency, eco-friendly bionematicides. These findings offer innovative, sustainable tools for integrated pest management strategies, crucial for preserving vulnerable forest ecosystems.

Acknowledgements: To FCT–MCTES for structural funding for GreenIt/LS4Future (UID/04551/2025), CE3C/CHANGE (UID/00329/2025), MED/CHANGE (UID/05183/2025 and LA/P/0121/2020) and CQE/IMS (UID/00100/2025; UID/PRR/00100/2025 and LA/P/0056/2020).