Differential thermal analysis (DTA) was used to measure deep supercooling in flower buds of Prunus dulcis Mill., P. armeniaca L., P. davidiana (Carr.) Franch, P. persica (L.) Batsch, three sweet cherry (P. avium L.) selections, and `Bing' cherries (P. avium L.) during Winter 1990-91 and 1991-92. Low temperatures in Dec. 1990 killed many flower buds. After the freeze, dead flower primordia continued to produce low-temperature exotherms (LTEs) at temperatures near those of living primordia for >2 weeks. In Feb. 1992, cherry buds that had been killed by cooling to -33C again produced LTEs when refrozen the next day. As buds swelled, the median LTE (LTE50) of dead buds increased relative to that of living buds, and the number of dead buds that produced LTEs decreased. LTE artifacts from dead flower priimordia must be recognized when DTA is used to estimate LTE50 of field-collected samples.
Sorkel A. Kadir and Ed L. Proebsting
Michael Wisniewski and Glen Davis
The pit membrane of xylem parenchyma of peach plays an important role in deep supercooling. Enzyme hydrolysis of xylem tissue indicated that the pit membrane is rich in pectin. The objective of the present study was to determine if removal of calcium from the cell wall would effect deep supercooling by loosening the cell wall. Current year shoots of `Loring' peach were infiltrated with oxalic acid, EGTA, or sodium phosphate buffer for 24-48 hours and then prepared for either ultrastructural analysis or differential thermal analysis. The use of 5-50 mM oxalic acid resulted in a distinct reduction in the size of the low-temperature exotherm (LTE) with increasing concentration. Oxalic acid also produced a loosening and swelling of the pit membrane. The use of EGTA (100 mM) or NaP04 (150 mM) produced only a slight shift in the LTE to warmer temperatures when compared to fresh tissues. Heat treatments (30-100°C) also resulted in a gradual shift of the LTE to warmer temperatures. The data indicate that cross-linking of pectins may play a role in defining the pore structure of the pit membrane and that this area of the cell wall plays an integral role in deep supercooling of peach wood.
Fanyi Shen, Rongfu Gao, Wenji Liu, Wenjie Zhang, and Qi Zhao
It is known that the redistribution of water and the formation of dispersed water units appears to be a prerequisite for deep supercooling. A concentration of the cell solute results from the migration of water during extracelullar freezing and lowers the temperature of homogeneous nucleation, but we are convinced that nucleation of ice within cells may be initiated by a heterogeneous mechanism, except we consider a small spherical cave, the water can freeze on the wall of this cave. We are also convinced that the solid walls of the capillary exert an external potential on the water molecules, causing the shift of the triple point of the confined fluids. Based on Fletcher's work for spherical particle, we have gotten the formula of critical free energy in the process of heterogeneous nucleation of water in a small spherical cave. This presentation introduces the theoretical background and counts the drop of temperature in heterogeneous nucleation. Then, putting two actions (depression of triple point and process of heterogeneous nucleation) together, we have calculated the freezing point. Sometimes it is lower than –38 °C. Some phenomena can be explained by using this theory: 1) Water is at the tension status, which means that it wets plant tissue, so the triple point (melting point) of tissue water can be lowered. 2) The redistribution of water, formation of dispersed water units, and dry region preventing ice from propagating, all allow heterogeneous nucleation, then the two actions can be synthesized and the water would lead to deep supercooling. If the barriers were destroyed, heterogeneous nucleation and deep supercooling would certainly be lost. 3) This theory is only suited to rigid wall of small cave, so we understand why cell wall rigidity has been shown to affect freezing characteristics. Project 39870234 supported by National Nature Science Foundation.
Pinghai Ding* and Leslie H. Fuchigami
Differential exothermal characteristics in relations to bud water content and dormant periods were dynamically investigated within the buds of apple, pear, peach, plum, Grape, persimmon, and black walnut from late autumn to early spring. Differential thermal analysis (DTA) indicated that bud cold hardiness and two exotherms, the high temperature exotherm (HTE) and low temperature exotherm (LTE), were different among species and dormant periods. According to whether buds have deep supercooling during the dormant winter period the species tested can be divided into two groups. The first group, without supercooling, includes the buds of apple and pear, in which LTE was undetectable. The second group, with supercooling, includes the buds of peach, plum, grape, persimmon, and black walnut, in which LTE was detectable. The second group can be further divided into peach and plum subgroup, and grape, persimmon, and black walnut subgroup. Both HTE and LTE can be detected in the buds of peach and plum subgroup, in which bud cold hardiness can be further divided into three different stages; whereas in the buds of grape, persimmon and black walnut subgroup only LTE can be detected, in which bud cold hardiness can be further divided into five stages according to the detection dynamics of HTE and LTE. Bud differential exothermal characteristics and deep supercooling dynamics are closely related to bud water content and cold hardiness stages. No detection of LTE in the buds of apple and pear and no detection of HTE in the buds of grape, persimmon and black walnut were both closely associated with bud water content.
Cindy L. Flinn and Edward N. Ashworth
Thermal analysis of Forsythia × intermedia `Spectabilis' flower buds had previously detected the occurrence of low temperature exotherms (LTE) during freezing. The LTE apparently resulted from the freezing of supercooled water and corresponded to the death of the florets. The genus Forsythia encompasses a wide array of species and interspecific crosses ranging in flower bud hardiness and floret size. The ability of buds to supercool, the relationship between the LTE and flower bud hardiness, and the extent to which floret size affects both were studied in flower buds of the following Forsythia species: F. × intermedia `Spectabilis', F. × intermedia `Lynwood', F. `Meadowlark', F. suspensa var. fortunei, F. `Arnold Dwarf, F. europaea, F. giraldiana, F. × intermedia `Arnold Giant', F. japonica var. saxatilis, F. mandshurica, F. ovata, and F. viridissima. Flower buds used for thermal analysis were also used in subsequent size determinations. Hardiness evaluations were conducted using controlled freezing tests, and the sampling interval defined using the temperature range of the LTEs. Initial evaluation indicated a high degree of correlation (α>.50) between mean LTEs and mean killing temperatures. The Forsythia genus, with its broad range of bud hardiness and size provides an excellent system in which to study the mechanisms of supercooling. Thermal analysis of cultivars which exhibit LTEs can accurately assess bud hardiness with minimal plant material.
Rajeev Arora and Michael Wisniewski
The seasonal pattern of dehydrin accumulation was characterized during cold acclimation and deacclimation in the xylem tissues of genetically related (sibling) deciduous and evergreen peach (Prunus persica L.). Immunological studies indicate that a 60-kD polypeptide in peach xylem tissues is a dehydrin protein. Comparison of its accumulation pattern with seasonal fluctuations in cold hardiness indicate that dehydrin accumulated to high levels during the peak of cold acclimation. However, its accumulation was only weakly associated with cold hardiness during early stages of cold acclimation and during deacclimation. Our results indicate that factors related to supercooling rather than dehydrin accumulation may be primarily responsible for determining levels of cold hardiness during transition periods.
Edward F. Durner
Edward F. Durner
Flower bud hardiness of ethephon-treated (100 mg·liter-1 in October), dormant pruned (in December) `Redhaven' peach (Prunus persica L. Batsch.) trees was studied from December through March using exotherm analysis. In early December, buds not treated with ethephon were 0.5C hardier than ethephon-treated buds. From mid-December through March, ethephon-treated buds were 0.5 to 2.1C hardier than nontreated buds. When a main effect of pruning was detected, buds from pruned trees were 0.8 to 2.8C less hardy than buds from nonpruned trees. On several dates, a significant interaction on flower bud hardiness between ethephon treatment and pruning was detected. For trees not treated with ethephon, buds from pruned trees were 1.8 to 2.2C less hardy than those from nonpruned trees. Pruning did not affect hardiness of buds from ethephon-treated trees. Ethephon delayed bloom to the 75% fully open stage by 9 days. Pruning accelerated bloom to the 75% fully open stage by 3 days compared to nonpruned trees. Flower bud dehardening under controlled conditions was also studied. As field chilling accumulated, flower buds dehardened more rapidly and to a greater extent when exposed to heat. Pruning accelerated and intensified dehardening. Ethephon reduced the pruning effect. The percentage of buds supercooling from any ethephon or pruning treatment did not change as chilling accumulated. In trees not treated with ethepbon, fewer buds supercooled as heat accumulated, and pruning intensified this effect. In pruned, ethephon-treated trees, fewer buds supercooled after exposure to heat. The number of buds supercooling in nonpruned trees did not change with heat accumulation. Flower bud rehardening after controlled dehardening was also evaluated. After dehardening in early February, there was no difference in the bud hardiness of pruned or nonpruned trees. Buds from ethepbon-treated trees were hardier than those from nontreated trees. With reacclimation, buds from pruned trees were not as hardy as those from nonpruned trees. The percentage of buds supercooling from ethephon-treated trees did not change with deacclimation or reacclimation treatments. After deacclimation in late February, buds from pruned trees were 2.2C less hardy than those from nonpruned trees. After reacclimation, buds from pruned, ethephon-treated trees rehardened 2.6C while buds from all other treatments remained at deacclimated hardiness levels or continued to deharden. Ethephon-treated pistils were shorter than nontreated pistils. Pistils from pruned trees were longer than those from nonpruned trees. Deacclimated pistils were longer than nondeacclimated pistils. Differences in hardiness among ethephon and pruning treatments were observed, but there was no relationship between pistil moisture and hardiness.
Michael Wisniewski, Glen Davis, and Katherine Bowers
Our previous research has indicated that the pit membrane regulates deep supercooling of xylem parenchyma in woody plants. This area of the cell wall is composed of three layers that may be rich in pectins. Since pectins may define the porosity of the cell wall they may also regulate deep supercooling. The present study examined pectin distribution in ray cells using monoclonal antibodies, that recognize un-esterified (JIM5) and methyl-esterified (JIM7) epitopes of pectin, in conjuction with immunogold electron microscopy. Antibodies were obtained courtesy of J. Paul Knox, John Innes Inst., U.K. Dormant and non-dormant tissues of Prunus persica, Cornus florida and Salix babylonica were utilized. Labelling with JIM7 revealed that methyl-esterified pectins were abundant and evenly distributed within the primary cell wall and amorphous layer. Labelling with JIM5 revealed that un-esterified pectins were located specifically within the pit membrane, in the outer region of the primary cell wall. No differences were observed between species, however, preliminary data indicated that JIM5 labelling was greater in dormant than in non-dormant tissues.
Sorkel A. Kadir and Edward L. Proebsting
Flower buds of 20 Prunus species showed quite different strategies to cope with low temperatures. Buds of most species deep supercooled. The two hardiest species, both from the subgenus Padus (P. padus L. and P. virginiana L.), did not supercool and survived -33C with no bud kill. Prunus serotina J.F. Ehrh., also in Padus, did supercool. Prunus nigra Ait., P. americana Marsh, P. fruticosa Pall., and P. besseyi L.H. Bailey had a low minimum hardiness level (MHL), small buds, and a low water content. Exotherms were no longer detectable from the buds of these species after 2 days at -7C and some buds survived -33C. Prunus triloba Lindl. and P. japonica Thunb. were similar to that group, but no buds survived -33C. Prunus davidiana (Carriere) Franch., P. avium L., and P. domestica L. had a relatively high MHL but hardened rapidly when the buds were frozen. Prunus persica (L.) Batsch., P. subhirtella Miq., P. dulcis (Mill) D. A. Webb, and P. emarginata (Dougl. ex Hook) Walp. deep supercooled, had large flower buds and a high MHL, and were killed in the Dec. 1990 freeze. Prunus salicina Lindl., P. hortulana L.H. Bailey, P. armeniaca L., and P. tomentosa Thunb. were in an intermediate group with a moderately low MHL and a moderate rate of hardiness increase while frozen. Prunus dulcis and P. davidiana had a low chilling requirement and bloomed early, whereas P. virginiana, P. fruticosa, P. nigra, and P. domestica had high chilling requirements and bloomed late.