Properties of peach flower buds which facilitate supercooling

Water in dormant peach (Prunus persica [L.] Batsch. var. `Harbrite') flower buds deep supercooled. Both supercooling and the freezing of water within the bud axis and primordium as distinct components depended on the viability of the bud axis tissue. The viability of the primordium was not critical. Supercooling was prevented by wounding buds with a dissecting needle, indicating that bud structural features were important. Bud morphological features appeared to prevent the propagation of ice through the vascular tissue and into the primordium. In dormant buds, procambial cells had not yet differentiated into xylem vessel elements. Xylem continuity between the bud primordium and adjacent tissues did not appear to be established until buds had deacclimated. It was concluded that structural, morphological, and physiological features of the bud facilitated supercooling.     

The relationship between vascular differentiation and the distribution of ice within Forsythia flower buds

The relationship between vascular development and the distribution of ice within overwintering forsythia flower buds was examined. Previous experiments demonstrated that ice formed in dormant buds within the bud scales, sepals, and in the peduncle and lower portions of the developing flower. The current study demonstrated that these tissues contained columns of primary xylem forming a continuous network with the subtending stem tissues. The vascular traces within the developing petals, anther filaments and pistil were not fully differentiated. Xylem vessels were not present and only procambial cells were observed. Large ice crystals did not accumulate in these tissues. When vascular development resumed in the spring, coincident changes in the distribution of ice within buds were noted. Observations were consistent with the hypothesis that ice propagates into buds via the vascular system, and that the segregation of ice within bud tissues reflects the distribution of xylem.     

The Formation and Distribution of Ice within Forsythia Flower Buds

Differential thermal analysis detected two freezing events when dormant forsythia (Forsythia viridissima Lindl.) flower buds were cooled. The first occurred just below 0°C, and was coincident with the freezing of adjacent woody tissues. The second exotherm appeared as a spike between −10 and −25°C and was correlated with the lethal low temperature. Although this pattern of freezing was similar to that observed in other woody species, differences were noted. Both direct observations of frozen buds and examination of buds freeze-fixed at −5°C demonstrated that ice formed within the developing flowers at temperatures above the second exotherm and lethal temperature. Ice crystals had formed within the peduncle and in the lower portions of the developing flower. Ice also formed within the scales. In forsythia buds, the developing floral organ did not freeze as a unit as noted in other species. Instead the low temperature exotherm appeared to correspond to the lethal freezing of supercooled water within the anthers and portions of the pistil.     

黄花杓兰的花芽发育

对黄花杓兰（Cypripedium flavum P.F.Hunt et Summerh.）成年植株做了一个生长季的研究,提出了一年芽、二年芽和多年休眠芽的概念。指出由芽形成到植株开花需两年时间,其具体发育路线是：第一年6－7月份,根状茎顶端二年芽基部外侧有两个新的小芽产生,即“一年芽”,至9－10月份发育出7－9片幼叶,然后随气温下降停止生长;第2年4月份复苏,即为“二年芽”,二年芽在本生长季内发育成混合芽,但一般情况下只有一个充分发育,另一个未能充分发育并且一般将来也不再有发育的机会,被称为“多年休眠芽”;第3年5月份充分发育的二年芽长出地面,形成植株,迅速开花、结果,至9月底植株枯萎。本文还讨论了黄花杓兰发育过程与环境的关系。     

Cold hardiness in various organs and tissues of Rhododendron species and the supercooling ability of flower buds as the most susceptible organ

The freezing resistance of various organs and tissues was determined in 24 Rhododendron species (mainly Subgenus Tsutsutsi) having different ecological distributions. The order of hardiness for organ or tissue is as follows: leaf bud > wood ≧ bark > flower bud, and the flower bud is characterized as the most cold-susceptible organ. The relationship of killing temperature (KT) to northern distribution was the most significant in leaf buds compared to other organs and tissues. KTs of leaf buds for the most hardy species were ?45 °C (or below) and those for the most tender species were about ?23 °C, while KTs of flower buds were about ?28 °C for the former and ?16 °C for the latter. Although KTs of flower buds native to southwestern Japan were well correlated with the exothermic temperature distribution (ETD) of florets, those in the more northern species were generally lower than ETDs. The supercooling ability of flower buds appears to be sufficient to avoid the freezing stress since the extreme minimum temperature (EMT) at the northern limit of natural distribution for each tree species examined was not lower than the KT and ETD of the flower buds.     

Immunogold localization of pectins and glycoproteins in tissues of peach with reference to deep supercooling

Living xylem tissues and floral buds of several species of woody plants survive exposure to freezing temperatures by deep supercooling. A barrier to water loss and the growth of ice crystals into cells is considered necessary for deep supercooling to occur. Pectins, as a constituent of the cell wall, have been implicated in the formation of this barrier. The present study examined the distribution of pectin in xylem and floral bud tissues of peach (Prunus persica). Two monoclonal antibodies (JIM5 and JIM7) that recognize homogalacturonic sequences with varying degrees of esterification were utilized in conjunction with immunogold electron microscopy. Results indicate that highly esterified epitopes of pectin, recognized by JIM7, were the predominant types of pectin in peach and were uniformly distributed throughout the pit membrane and primary cell walls of xylem and floral bud tissues. In contrast, un-esterified epitopes of pectin, recognized by JIM5, were confined to the outer surface of the pit membrane in xylem tissues. In floral buds, these epitopes were localized in middle lamellae, along the outer margin of the cell wall lining empty intercellular spaces, and within filled intercellular spaces. JIM5 labeling was more pronounced in December samples than in July/August samples. Additionally, epitopes of an arabinogalactan protein, recognized by JIM14, were confined to the amorphous layer of the pit membrane. The role of pectins in freezing response is discussed in the context of present theory and it is suggested that pectins may influence both water movement and intrusive growth of ice crystals at freezing temperatures.     

Frost Injury as a Possible Inciting Factor in Bud and Shoot Necroses of Fraxinus excelsior L.

Large numbers of European ash have died in Poland in all age classes during the last ten years. The characteristic symptom occurring on shoots of planted and self‐sown seedlings was bark necroses starting from the shoot apex, necrotic buds, or leaf and twig scars. The results showed that in the bud tissue of cold acclimated European ash extracellular and intracellular ice formation occurred at approximately ?9 and ?32°C, respectively. In deacclimated plants in spring water supercooling is limited by the heterogenous ice nucleation temperature and consequently the cold tolerance is ?9 to ?4°C for bud tissues and ?13 to ?9°C for shoots. Isolations of fungi were performed from dead buds and from necroses occurring on the main stem. Alternaria alternata, Fusarium lateritium and Phomopsis scobina were among the fungi occurring in both these organs at frequencies of more than 7%. Cylindrocarpon heteronemum, Diplodia mutila and Tubercularia vulgaris from necroses were only isolated in frequencies; 3.3, 1.2 and 5.4%, respectively. It seems likely that freezing injury is the inciting factor, which combined with fungal colonization manifests itself as fatal damage to European ash buds and shoots.     

Winter bud content according to position in 3-year-old branching systems of 'Granny Smith' apple

An investigation was made of the number of preformed organs in winter buds of 3-year-old reiterated complexes of the 'Granny Smith' cultivar. Winter bud content was studied with respect to bud position: terminal buds were compared on both long shoots and spurs according to branching order and shoot age, while axillary buds were compared between three zones (distal, median and proximal) along 1-year-old annual shoots in order 1. The percentage of winter buds that differentiated into inflorescences was determined and the flowers in each bud were counted for each bud category. The other organ categories considered were scales and leaf primordia. The results confirmed that a certain number of organs must be initiated before floral differentiation occurred. The minimum limit was estimated at about 15 organs on average, including scales. Total number of lateral organs formed was shown to vary with both bud position and meristem age, increasing from newly formed meristems to 1- and 2-year-old meristems on different shoot types. These differences in bud organogenesis depending on bud position, were consistent with the morphogenetic gradients observed in apple tree architecture. Axillary buds did not contain more than 15 organs on average and this low organogenetic activity of the meristems was related to a low number of flowers per bud. In contrast, the other bud categories contained more than 15 differentiated organs on average and a trade-off was observed between leaf and flower primordia. The ratio between the number of leaf and flower primordia per bud varied with shoot type. When the terminal buds on long shoots and spurs were compared, those on long shoots showed more flowers and a higher ratio of leaf to flower primordia.     

Supercooling Ability, Water Content and Hardiness of Rhododendron Flower Buds during Cold Acclimation and Deacclimation

The relationship between supercooling ability and water contentand killing temperature of flower buds during cold acclimationand deacclimation were studied using R. kiusianum and R. x akebono.The occurrence of multiple floret exotherms and their shiftto a narrow range at lower subzero temperatures, as well asthe marked decrease of florets water content, were observedas the symptoms of cold acclimation occuring in flower budsfrom fall to winter, and vice versa in spring buds during deacclimation.In R. kiusianum, the fully acclimated period was from Novemberto March and two months longer than that of R. x akebono. Thesupercooling ability of the former was about –25°Cand about –20°C in the latter. Although the watermigration within bud tissues during the freezing process wasdetermined in the acclimated and deacclimated buds for R. xakebono, no significant water changes could be observed, evenin the acclimated buds. Thus, it is conceivable that deep supercoolingin florets may result not necessarily from water migration fromflorets and bud axes to scales in response to freezing, butfrom low water content in situ of cold-acclimated or artificiallydehydrated flower buds. (Received July 29, 1981; Accepted October 12, 1981)     

Cryo-scanning electron microscopic study on freezing behaviors of tissue cells in dormant buds of larch (Larix kaempferi)

The freezing behavior of dormant buds in larch, especially at the cellular level, was examined by a Cryo-SEM. The dormant buds exhibited typical extraorgan freezing. Extracellular ice crystals accumulated only in basal areas of scales and beneath crown tissues, areas in which only these living cells had thick walls unlike other tissue cells. By slow cooling (5 °C/day) of dormant buds to −50 °C, all living cells in bud tissues exhibited distinct shrinkage without intracellular ice formation detectable by Cryo-SEM. However, the recrystallization experiment of these slowly cooled tissue cells, which was done by further freezing of slowly cooled buds with LN and then rewarming to −20 °C, confirmed that some of the cells in the leaf primordia, shoot primordia and apical meristem, areas in which cells had thin walls and in which no extracellular ice accumulated, lost freezable water with slow cooling to −30 °C, indicating ability of these cells to adapt by extracellular freezing, whereas other cells in these tissues retained freezable water with slow cooling even to −50 °C, indicating adaptation of these cells by deep supercooling. On the other hand, all cells in crown tissues and in basal areas of scales, areas in which cells had thick walls and in which large masses of ice accumulated, had the ability to adapt by extracellular freezing. It is thought that the presence of two types of cells exhibiting different freezing adaptation abilities within a bud tissue is quite unique and may reflect sophisticated freezing adaptation mechanisms in dormant buds.     

The Exogenous Hormornal Control of the Development of Regenerated Flower Buds in Hyacinthus orientalis

The critical role of exogenous hormone on inducing the initiation of different floral organs in the regenerated flower bud and controlling their numbers was further evidenced. The initiation of the flower buds was first induced from the perianth explants of Hyacinthus orientalis L. cv. White pearl by a combination of 2 mg/L 6-BA and 0.1 mg/L 2,4-D, and then a continuous initiation of over 100 tepals (a flower bud of H. orientalis in situ has only 6 tepals) was successfully controlled by maintenance of such a hormone concentration. However, a change of hormonal concentration (2 mg/L 6-BA and 0-0.000 1 mg/L 2,4-D) caused cessation of continuous initiation of the tepals but gave rise to induction of stamen initiation. Keeping the changed hormone concentrations could successfully control the continuous initiation of over 20 stamens (a flower bud of H. orientalis in situ has only 6 stamens). The experiment showed that the number of identical floral organs in the regenerated flower buds can be controlled by certain defined concentrations of the exogenous hormones, and the amount of the induced identical floral organs has no effect on the differentiation sequence of the different floral organs in the regenerated flower bud. Based on a systematic research on controlling the differentiation of the floral organs from both the perianth explants and the regenerated flower buds by the exogenous hormones in H. orientalis over the past decade, the authors put forward here a new idea on the role of phytohormone in controlling the automatic and sequential differentiation of the different floral organs in flower development. The main points are as follows: 1. the development of flower bud in plant is a process in which all of the floral organs are automatically and sequentially differentiated from the flower meristem. 2. Experiments in vitro showed that the effect of exogenous hormones in controlling the initiation of different floral organs is strictly concentration dependent, i.e., one kind of the floral organ can continuously and repeatedly initiate from the flower meristem as long as it is maintained in that specific concentration of the exogenous hormone which is suitable for the initiation of that particular kind of floral organ. 3. It shows that the flower buds in situ must be automatically able to adjust the endogenous hormonal concentrations just after the completion of the differentiation of one whorl of floral organ to suit the differentiation of the next whorl. Thus, the phytohormone in different concentrations takes after many change-over switches of the organ differentiation and plays a connective and regulatory role between the differentiation of every two whorls of the floral organ. In other words, these change-over switches play the roles of inhibiting the expression of the genes which control the initiation of the floral organs in the first whorl, meanwhile, activating the expression of the genes which control the initiation of the floral organs in the second whorl during the successive initiation of the different floral organs from the flower bud. It results in the automatic and sequential initiation of the various floral organs from the floral meristem.      

Supercooling characteristics of isolated peach flower bud primordia

The amount of unfrozen water in dormant peach (Prunus persica [L.] Batsch, cv Redhaven) flower buds, isolated primordia, and bud axes was determined during freezing using pulse nuclear magnetic resonance methods. Differential thermal analysis studies were conducted on whole buds and isolated primordia in the presence of ice nucleation. The results showed that some of the water in isolated primordia remained supercooled in the presence of ice nucleation. Although most tissue water froze (57.5%) following ice nucleation at −2.5°C, a considerable amount of water was found to supercool. In the presence of ice nucleation, increased hydration of isolated primordia resulted in the elimination of the supercooling characteristic. The structural integrity of isolated primordia appeared to be essential for supercooling.     

The Role of Light in Leaf and Flower Bud Break of the Peach (Prunus persica)

The effect of light on peach leaf and flower bud break was examined. It was found that leafless dormant shoots were light-perceptive organs. Darkness, after light preconditioning during dormancy, reduced leaf bud opening; however, light was obligatory when the shoots were preconditioned in the dark. Relatively short exposures to light were sufficient to stimulate leaf bud break. Terminal buds were less inhibited by darkness than were laterals. Flower bud break was inhibited in light after dark preconditioning. The red region of the spectrum was found to be active; the phytochorome system seems to be involved in the light reactions, as the red light effect was reversible with subsequent far-red illumination. Supplementary light, producing long-day conditions, could partly compensate for insufficient chilling. A possible sequence of reactions in the plant is suggested.     

荷花玉兰休眠芽幼叶的形态和发育特征

对荷花玉兰休眠芽的形态和发育特征进行了解剖观察。结果表明:幼叶多直立,个别旋抱状;叶片沿中脉向近轴面,在同株和异株的芽间随机性向左或向右纵向对折;叶芽内的外1～3层和花芽内的幼叶常枯死;花芽最内一层幼叶柄与其托叶贴生,并且叶片多完全退化,个别发育出较小的正常叶片。芽内幼叶枯死,是适应性的生理退化而非病害或营养不良现象,在演化上可能与其托叶替代幼叶作为芽鳞进行保护作用有相关性。     

穗花牡荆花芽分化过程中形态和生理指标变化

以穗花牡荆为研究材料,通过探究其花芽分化进程和生理特性,为花期调控技术提供成花机理。采用物候期观察和石蜡切片相结合的方法并测定花芽分化过程中相关生理指标,研究花发育过程中的形态和生理变化。结果表明,穗花牡荆花芽分化为一年多次分化型,其进程可划分为七个时期：未分化期、总轴花序原基分化期、初级分轴花序原基分化期、次级分轴花序原基分化期、小花原基分化期、花器官分化前期和花器官分化后期。同一植株不同位置花芽及同一花序中不同单花分化的进程不同,第一季花期后各阶段的花芽分化形态常存在重叠。花芽分化过程中不同时期叶片和花芽的可溶性糖和可溶性蛋白质含量均有上升下降的变化,总体上叶片中营养物质含量高于花芽保证营养供应。花芽分化过程中,IAA、ABA、CTK和GA3整体水平上先升后降有利于花芽分化进行。研究认为,花芽中大量的可溶性糖和蛋白质积累及较高的碳氮比,有利于穗花牡荆花芽形态分化顺利完成。低水平的GA3/ABA和IAA/CTK有利于花序的形成,ABA/CTK和ABA/IAA比值升高促进小花原基和小花萼片原基的分化, GA3/CTK、GA3/ABA和GA3/IAA比值升高促进花瓣原基、雄雌蕊原基发育。     

The effect of LAB 173711 and ethephon on time of flowering and cold hardiness of peach flower buds

Peach flowers are often killed during bloom by spring frosts. LAB 173711, a compound with abscisic (ABA)-like activity, and ethephon delayed flowering in peach trees. In greenhouse experiments, LAB 173711, at concentrations of 10^?3–10^?2 M, was most effective in delaying bloom when applied after a 5°C cold storage period, rather than before the dormancy breaking treatment. In contrast, ethephon delayed bloom most effectively when applied before 5°C cold storage; ethephon caused flower bud abscission when treatments were made after the chilling requirement had been satisfied. In field experiments, ethephon delayed flowering by 6–7 days, which reduced bud injury after a spring frost during bloom. No flower bud injury was found on ethephon-treated trees after temperatures of ?4.3°C; whereas without ethephon 25% of the flower buds were frost damaged. LAB 173711 delayed the time to 50% bloom by 2–3 days. However, this was not long enough to avoid low-temperature injury to the flower buds.     

Freezing behaviours in wintering Cornus florida flower bud tissues revisited using MRI

How plant tissues control their water behaviours (phase and movement) under subfreezing temperatures through adaptative strategies (freezing behaviours) is important for their survival. However, the fine details of freezing behaviours in complex organs and their regulation mechanisms are poorly understood, and non‐invasive visualization/analysis is required. The localization/density of unfrozen water in wintering Cornus florida flower buds at subfreezing temperatures was visualized with high‐resolution magnetic resonance imaging (MRI). This allowed tissue‐specific freezing behaviours to be determined. MRI images revealed that individual anthers and ovules remained stably supercooled to ?14 to ?21 °C or lower. The signal from other floral tissues decreased during cooling to ?7 °C, which likely indicates their extracellular freezing. Microscopic observation and differential thermal analyses revealed that the abrupt breakdown of supercooled individual ovules and anthers resulted in their all‐or‐nothing type of injuries. The distribution of ice nucleation activity in flower buds determined using a test tube‐based assay corroborated which tissues primarily froze. MRI is a powerful tool for non‐invasively visualizing unfrozen tissues. Freezing events and/or dehydration events can be located by digital comparison of MRI images acquired at different temperatures. Only anthers and ovules preferentially remaining unfrozen are a novel freezing behaviour in flower buds. Physicochemical and biological mechanisms/implications are discussed.     

Passion fruit flowers: Kunitz trypsin inhibitors and cystatin differentially accumulate in developing buds and floral tissues

In order to better understand the physiological functions of protease inhibitors (PIs) the PI activity in buds and flower organs of passion fruit (Passiflora edulis Sims) was investigated. Trypsin and papain inhibitory activities were analyzed in soluble protein extracts from buds at different developmental stages and floral tissues in anthesis. These analyses identified high levels of inhibitory activity against both types of enzymes at all bud stages. Intriguingly, the inhibitory activity against both proteases differed remarkably in some floral tissues. While all organs tested were very effective against trypsin, only sepal and petal tissues exhibited strong inhibitory activity against papain. The sexual reproductive tissues (ovary, stigma-style and stamen) showed either significantly lower activity against papain or practically none. Gelatin–SDS–PAGE assay established that various trypsin inhibitors (TIs) homogenously accumulated in developing buds, although some were differentially present in floral organs. The N-terminal sequence analysis of purified inhibitors from stamen demonstrated they had homology to the Kunitz family of serine PIs. Western-blot analysis established presence of a ∼60 kDa cystatin, whose levels progressively increased during bud development. A positive correlation between this protein and strong papain inhibitory activity was observed in buds and floral tissues, except for the stigma-style. Differences in temporal and spatial accumulation of both types of PIs in passion fruit flowers are thus discussed in light of their potential roles in defense and development.     

Influence of different temperatures on bud break and 1-aminocyclopropane-1-carboxylic acid content of young peach trees

Effects of different temperatures on bud break and 1-aminocyclopropane-1-carboxylic acid (ACC) content were determined by using potted two-year-old ‘Akatsuki’ peach trees. One group of trees were subjected to 1°C for four weeks and then transferred to a growth chamber at 24°C, while the other was kept at 24 °C throughout the experiment. After four-week temperature treatments floral and vegetative bud break were evaluated weekly and bud break percentage was calculated. Bud break was greater under 1 °C than 24 °C in both November and December. The time required to release buds from dormancy was shorter in December than November. In November ACC content in peach buds increased after one and two weeks, then decreased in the forth week under both treatments. However, in December ACC content after two and four weeks showed a similar trend under 1 °C and a reverse trend under 24 °C. It was higher under low temperature treatment. These data indicate that chilling requirements for bud break of peach seems to be associated with the promotion of ethylene biosynthesis caused by low temperature stress.     

Dormancy in peach (Prunus persica L.) flower buds

Morphological studies were carried out with peach flower buds collected monthly in 1989 and 1990, from two months before leaf fall (7 March) until two to three weeks before bloom (7/8 August). Chilled (2–4°C for 30 days) and unchilled buds were exposed to 20 to 25°C, 100% RH and continuous light. Gibberellin A₃ (3 ng or 30 ng) was applied to some of the non-chilled cuttings at three days intervals. Then, 12, 19, and 26 days after they were planted, the buds were sampled and processed for histological studies. Cultured flower buds (chilled or unchilled) had accelerated anther and gynoecium morphogenesis after 12 days under controlled conditions, compared to buds processed immediately after collection from the field. Chilling treatment augmented the bud culture effect, while Gibberellin A₃ applications to the excised buds retarded bud morphogenesis to a stage comparable to that of buds collected directly from the field. This, suggests that the comparatively high levels of Gibberellin A_1/3 we previously found in mid winter [15, 18] could be at least one of the factors that controls floral bud dormancy by retarding anther and gynoecium development.