X-ray phase contrast imaging of Vitis spp. buds reveals freezing pattern and correlation between volume and cold hardiness

Authors: KOVALESKI, ALISSON - Cornell University; Londo, Jason; FINKELSTEIN, KENNETH - Cornell University

Submitted to: Scientific Reports
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/15/2019
Publication Date: 10/18/2019
Citation: Kovaleski, A.P., Londo, J.P., Finkelstein, K. 2019. X-ray phase contrast imaging of Vitis spp. buds reveals freezing pattern and correlation between volume and cold hardiness. Scientific Reports. 9:14949. https://doi.org/10.1038/s41598-019-51415-2.
DOI: https://doi.org/10.1038/s41598-019-51415-2

Interpretive Summary: Grapevines, like all woody perennial crops, produce buds during the growing season which go dormant during times of stress. To survive winter temperatures, these buds must adjust their cold hardiness as temperatures fluctuate from day to day. The mechanism that grapevine's use to survive winter temperatures is called supercooling, a process that involves suppressing the freezing point of water to temperatures as low as -40°. What isn't known is what happens when supercooling fails. Specifically, where in the bud does the first ice crystal form, and how does ice move through the bud. In this study, we used X-ray phase contrast imaging to watch the formation of ice inside dormant grapevine buds. High powered x-ray beams were passed from the Cornell High Energy Synchrotron Source (CHESS) facility through the buds while the buds were gradually frozen. We discovered that the lethal freezing event occurs at the center of the bud and takes several minutes to propagate through the bud structure. Additionally we measured how the bud volume changes as dormant buds begin losing cold hardiness as they approach budbreak. Our results demonstrated that the supercooling mechanism remains intact as the bud loses cold hardiness. However, as the bud volume increases due to tissue expansion, the supercooling mechanism is lost. This result suggests that the dormant bud cannot resume growth while supercooling remains intact, and that bud expansion signals the end of cold hardiness in grapevine.

Technical Abstract: Grapevine (Vitis spp.) buds must survive winter temperatures in order to resume growth when suitable conditions return in spring. They do so by developing cold hardiness through deep supercooling, but the mechanistic process of supercooling in buds remains largely unknown. Here we use synchrotron X-ray phase contrast imaging to study cold hardiness-related characteristics of V. amurensis, V. riparia, and V. vinifera buds: time-resolved 2D imaging was used to visualize freezing; and microtomography was used to evaluate morphological changes during deacclimation. Bud cold hardiness was determined (low temperature exotherms; LTEs) using needle thermocouples during 2D imaging as buds were cooled with a N2 gas cryostream. Resolution in 2D imaging did not allow for ice crystal identification, but freezing was assessed by movement of tissues coinciding with LTE values. Freezing was observed to propagate from the center of the bud toward the outer bud scales. The freezing events observed lasted several minutes. Additionally, loss of supercooling ability appears to be correlated with increases in bud tissue volume during the process of deacclimation, but major increases in volume occur after most of the supercooling ability is lost, suggesting growth resumption processes are limited by deacclimation state.