Contrasting Effects of Grass-Derived Endophytic Fungal VOCs on Early Growth of Spring Barley and Red Clover: From Stimulation to Suppression

Endophytic fungi can influence plant development through diverse molecular mechanisms; however, their volatile organic compound VOC-mediated effects on agriculturally relevant crops remain insufficiently characterized. In this study, we examined the effects of VOCs produced by six grass-root-associated endophytic fungi-<i>Cadophora fastigiata</i>, <i>Cordyceps fumosorosea</i>, <i>Chaetomium funicola</i>, <i>Epicoccum nigrum</i>, <i>Microdochium bolleyi</i>, and <i>Plectosphaerella cucumerina</i>-on early growth of spring barley (<i>Hordeum vulgare</i>) and red clover (<i>Trifolium pratense</i>). In plate-in-plate VOC exposure assays, we assessed root system traits, root hair formation, and biomass accumulation. Responses to fungal VOCs were fungal species-specific but similar across barley and red clover. VOCs emitted by <i>C. fastigiata</i> and <i>P. cucumerina</i> were consistently associated with increased root growth, root hair proliferation, and seedling biomass, whereas VOCs from <i>M. bolleyi</i> and <i>C. funicola</i> resulted in neutral or growth-suppressing effects. A complementary seed inoculation experiment was conducted with barley, which showed fungal species-dependent contrasting effects consistent with the observations of VOCs treatment. Gas chromatography-mass spectrometry (GC-MS) analysis revealed that <i>C. fastigiata</i>, the isolate associated with the strongest growth-promoting responses, emitted a diverse VOC profile dominated by sesquiterpenes, with 22 compounds identified. Together, these results demonstrate that VOCs emitted by grass-root-associated endophytic fungi exert reproducible, species-specific effects on early plant development occurring in phylogenetically distant species. The findings highlight the value of VOC-based assays for comparative functional screening of fungal isolates, providing a foundation for future studies that aim to link individual VOCs to plant growth responses.