Developmental stage-dependent and wound-triggered formation dynamics of volatiles in Pleurotus cornucopiae

Volatile organic compounds (VOCs) emitted by fungi play crucial roles in ecological interactions, including spore dispersal, defense, and communication with other organisms. However, developmental regulation and biosynthetic relationships among individual fungal VOCs remain poorly understood. In this study, we investigated growth-stage-dependent and wound-induced VOC emissions in the basidiomycete Pleurotus cornucopiae by combining nondestructive headspace sampling, GC-MS analysis, controlled wounding and feeding experiments, and real-time monitoring using proton-transfer-reaction time-of-flight mass spectrometry (PTR-ToF-MS). Developmental profiling revealed distinct emission patterns of fatty-acid-derived oxylipins and sesquiterpenes, with sesquiterpenes accumulating predominantly during the late fruiting body stages associated with spore release, whereas eight-carbon (C8) VOCs were transiently enhanced during the early reproductive stages. Mechanical wounding triggered an immediate burst of C8 VOCs including 1-octen-3-ol, 1-octen-3-one, and 3-octanone. PTR-ToF-MS analysis resolved their production kinetics at second-scale resolution, demonstrating the nearly simultaneous formation of 1-octen-3-ol and 1-octen-3-one, followed by delayed accumulation of 3-octanone. Cell-free enzyme assays indicated that enzymatic cleavage of linoleic acid 10(S)-hydroperoxide predominantly yielded 1-octen-3-ol. Feeding experiments and tissue disruption analyses further suggested that 3-octanone formation requires metabolic cooperation between damaged and adjacent intact tissues. Taken together, these results indicated that P. cornucopiae controls VOC production in a growth-and damage-dependent manner. We hope that our findings will facilitate future investigations on the ecological and physiological roles of dynamic VOC regulation during mushroom fruiting body development and in response to wounding.