Freeze-thaw-induced embolism is a key limiting factor for perennial plants in frost-exposed environments. Gas bubbles are formed during freezing and expand during thawing resulting in xylem embolism. However, when water freezes, its volume increases by 9%, generating local pressures, which can affect the formation of bubbles. To characterize local dynamic of pressure-tension and physical state of the sap during freeze-thaw cycles, we simultaneously used ultrasonic acoustic emissions analysis and synchrotron-based high resolution computed tomography on the diffuse-porous species Betula pendula. Visualization of individual air-filled vessels was performed to measure freeze-thaw induced embolism after successive freeze-thaw cycles down to -10°C or -20°C during the leafy and the leafless periods. We also measured the distribution of gas bubbles in frozen xylem of Betula pendula, and made additional continuous monitoring of embolism spreading during one freeze-thaw cycle using a dedicated cooling system that allowed X-ray scanning during freezing and thawing. Experiments confirmed that ultrasonic emissions occurred after the onset of ice formation, together with bubble formation, whereas the development of embolism took place after thawing in all cases. The pictures of frozen tissues indicated that the positive pressure induced by the volumetric increase of ice can provoke inward flow from the cell wall toward the lumen of the vessels. We found no evidence that wider vessels within a tissue were more prone to embolism although the occurrence of gas bubbles in larger conduits would make them prone to earlier embolism. These results highlight the need to monitor local pressure as well as ice and air distribution during xylem freezing to understand the mechanism leading to frost-induced embolism.
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