Multiplexed Nanosensors for Real-Time Monitoring of VOC-Mediated Signalling Response in Plants.

To enhance crop performance, intercropping strategies leverage volatile organic compound (VOC)-driven interactions with companion plants that constitutively emit VOCs. Despite the agricultural importance, the mechanisms and kinetics of VOC-mediated sensory transduction in receiver plants eavesdropping on neighbouring non-kin emitters remain largely unknown due to a lack of appropriate non-destructive analytical tools. In this work, we employ multiplexed salicylic acid (SA) and H2O2 nanosensors in Brassica rapa subsp. Chinensis (pak choy) plants to visualize, in real time, reactive oxygen species (ROS) and SA signal transduction following exposure to constitutively-released VOCs from neighbouring aromatic plants-namely sweet basil and spearmint. Unique emitter-specific temporal signatures of ROS and SA were observed in receiver pak choy: sweet basil VOCs induced concomitant generation of ROS and SA at 30 min, whereas spearmint VOCs triggered SA production at 30 min, followed by ROS accumulation. The temporal data enabled the formulation of a diffusion model that quantifies the VOC perception threshold that triggers the distinct early ROS and SA signalling. Transcriptomics analysis at 2 h revealed that both emitters evoke largely distinct changes in pak choy, likely stemming from variations in speed and sequence of the early signal transduction, leading to different phenotypic outcomes. Intercropping with sweet basil led to enhanced pak choy biomass, stress resilience and secondary metabolite accumulation, whereas spearmint as companions had a limited impact. Our study captures in real time, the VOC-induced rapid signalling in receiver plants and its ensuing effect on growth. These nanosensor-enabled findings represent an important advance in deciphering how emitter-specific volatile cues are integrated into plant responses, guiding rational selection of beneficial companion plants for improved yield and nutritional profiles in sustainable agriculture.