Toxic effects and signal regulation of Eucalyptus robusta volatiles on Vicia faba cells

To understand the cytological mechanism of allelopathy of Eucalyptus robusta, the toxic effects of volatiles from E. robusta were studied by microscopic, cytochemical and qRT-PCR techniques, taking volatile oil from E. robusta and its main components α-pinene and eucalyptol as donors, and using root cells and leaf guard cells of Vicia faba as targets. The results were as follows: (1) The growth of radicle of V. faba were exhibited and showed a time-concentration dependent effects under the treatments of the volatiles of Eucalyptus robusta. The allelopathic effects were volatile oil, α-pinene and eucalyptol in descending order. (2) When Vicia faba roots were exposed to Eucalyptus robusta volatiles, the activity of root border cells decreased, the micronucleus rate of cells in the meristematic zone increased, mitotic index decreased, and the cell cycle of most cells was arrested in the prophase of division. (3) Under the action of the volatiles of E. robusta, the activity of NADPH oxidase increased, reactive oxygen species (ROS) burst in leaf guard cells of Vicia faba, microfilament polymerization, and stomatal aperture decreased. At the same time, the leaf epidermis strip of V. faba was treated with Eucalyptus robusta volatiles, the nuclear distortion rate of leaf guard cells increased. Moreover, the treatment of E. robusta volatiles led to the decrease of guard cell activity and caspase-dependent apoptosis in Vicia faba. However, the guard cell survival rates increased when the leaf epidermis strips of V. faba were exposed to volatiles from Eucalyptus robusta combined with different concentrations of Ca2+channel blocker (LaCl3), ROS scavenger ascorbic acid (AsA), and nitrate reductase inhibitors (NaN3), which indicated that the volatiles of E. robusta changed the signal regulation of Ca2+, ROS and NO. These results suggested that the cytotoxicity and genotoxicity of the volatiles of E. robusta altered the signal transduction pathway of the receptor cells, induced the genetic aberration of the root tip cells, then led to the dysfunction of protective function and stomatal movement of the receptor root border cells, which affected the root growth and photosynthesis of the receptor, and ultimately hindered the growth of receptor. The results provide a theoretical basis for scientific planting and management of E. robusta planting area.