title: High ice nucleation activity located in blueberry stem bark is linked to primary freeze initiation and adaptive freezing behaviour of the bark.
abstract: Controlled ice nucleation is an important mechanism in cold-hardy plant tissues for avoiding excessive supercooling of the protoplasm, for inducing extracellular freezing and/or for accommodating ice crystals in specific tissues. To understand its nature, it is necessary to characterize the ice nucleation activity (INA), defined as the ability of a tissue to induce heterogeneous ice nucleation. Few studies have addressed the precise localization of INA in wintering plant tissues in respect of its function. For this purpose, we recently revised a test tube INA assay and examined INA in various tissues of over 600 species. Extremely high levels of INA (-1 to -4 °C) in two wintering blueberry cultivars of contrasting freezing tolerance were found. Their INA was much greater than in other cold-hardy species and was found to be evenly distributed along the stems of the current year's growth. Concentrations of active ice nuclei in the stem were estimated from quantitative analyses. Stem INA was localized mainly in the bark while the xylem and pith had much lower INA. Bark INA was located mostly in the cell wall fraction (cell walls and intercellular structural components). Intracellular fractions had much less INA. Some cultivar differences were identified. The results corresponded closely with the intrinsic freezing behaviour (extracellular freezing) of the bark, icicle accumulation in the bark and initial ice nucleation in the stem under dry surface conditions. Stem INA was resistant to various antimicrobial treatments. These properties and specific localization imply that high INA in blueberry stems is of intrinsic origin and contributes to the spontaneous initiation of freezing in extracellular spaces of the bark by acting as a subfreezing temperature sensor.
authors.affiliation: Division of Plant Sciences, National Institute of Agrobiological Sciences, Kannondai 2-1-2, Tsukuba, Ibaraki 305-8602, Japan.
authors.forename: Tadashi
authors.initials: T
authors.lastname: Kishimoto
authors.affiliation: Division of Plant Sciences, National Institute of Agrobiological Sciences, Kannondai 2-1-2, Tsukuba, Ibaraki 305-8602, Japan Present address: International Patent Organism Depository, National Institute of Technology and Evaluation, Kisarazu, Chiba 292-0818, Japan.
authors.forename: Hideyuki
authors.initials: H
authors.lastname: Yamazaki
authors.affiliation: Division of Plant Sciences, National Institute of Agrobiological Sciences, Kannondai 2-1-2, Tsukuba, Ibaraki 305-8602, Japan.
authors.forename: Atsushi
authors.initials: A
authors.lastname: Saruwatari
authors.affiliation: Division of Plant Sciences, National Institute of Agrobiological Sciences, Kannondai 2-1-2, Tsukuba, Ibaraki 305-8602, Japan Applied Biological Science, Graduate School of Science and Technology, Tokyo University of Science, Yamazaki 2641, Noda, Chiba 278-8510, Japan.
authors.forename: Hiroki
authors.initials: H
authors.lastname: Murakawa
authors.affiliation: Faculty of Life and Environmental Sciences, University of Tsukuba, Tennodai 1-1-1, Tsukuba 305-8572, Japan.
authors.forename: Yoshihiko
authors.initials: Y
authors.lastname: Sekozawa
authors.affiliation: Applied Biological Science, Graduate School of Science and Technology, Tokyo University of Science, Yamazaki 2641, Noda, Chiba 278-8510, Japan.
authors.forename: Kazuyuki
authors.initials: K
authors.lastname: Kuchitsu
authors.affiliation: Nanoscale Organisation and Dynamics Group, University of Western Sydney, Penrith, NSW 2751, Australia.
authors.forename: William S
authors.initials: WS
authors.lastname: Price
authors.affiliation: Division of Plant Sciences, National Institute of Agrobiological Sciences, Kannondai 2-1-2, Tsukuba, Ibaraki 305-8602, Japan isikawam@affrc.go.jp.
authors.forename: Masaya
authors.initials: M
authors.lastname: Ishikawa
keywords: Blueberry; Vaccinium ashei; Vaccinium corymbosum; cold hardiness; extracellular freezing; freezing tolerance; ice nucleation; infra-red thermography
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