Unraveling the mechanism of action of a biostimulant: A successful integration of ecophysiologycal investigation and high-throughput phenotyping

Biostimulant (bs) products are a promising technological solution in agriculture, providing a key strategy to sustain crop quality and yield under climate challenges. To maximize product benefits and expand their application scope, beyond the progress made in formulation and characterization, it is essential to develop a structured strategy to study their mechanisms of action, ensuring robustness to account for both bs diversity and crop-specific responses. In this study, we used combination of ecophysiological analyses and High-Throughput Phenotyping to unveil the performance and mode of action of YaraAmplix OPTIVI (Yara International S.A.), a commercially registered bs under Regulation (EU) 2019/1009, to mitigate the negative effects of water stress effects in grapevine. Our solid approach indicated that the bs induces structural and functional modifications in leaf and stem tissues, promoting a water-saving strategy under well-watered conditions, while enhancing transpiration and photosynthetic efficiency during water stress. This dual response sustained carbon assimilation and accelerated recovery upon rehydration. The effect was driven by a drastic reduction in water and osmotic potential, triggering an anisohydric response under moderate stress while preserving stomatal control and protecting the photosynthetic apparatus under severe stress. Moreover, starch quantification in shoots revealed that enhanced photosynthetic efficiency led to greater reserve accumulation, improving stress resilience. High-temporal-resolution weighing lysimeters detected transpirative changes just days after the first application, while multispectral 3D imaging analysis identified leaf angle and Green Leaf Index as the most responsive parameters to water stress, effectively capturing early responses and indicating improved stress tolerance. Future field research should further assess the ability of bs to enhance plant resilience and performance in applied agricultural contexts while also investigating its potential effects on fruit quality traits and ripening kinetics. DLP was funded by Project PE9 GRINS, SPOKE 6 – WP1 activity - CUP D13C22002160001. CL and AF activities were in the frame of “POC Plant Flow Solutions”, NODES Spoke 2, Green technologies and sustainable industries – CUP D17G22000150001.