Frost tolerance and biochemical changes during hardening and dehardening in contrasting white clover populations

Successful winter survival of perennial plants, like white clover, is dependent on proper timing of both hardening and dehardening. The purpose of this study was to investigate the regulation of these processes in two cultivars (AberCrest and AberHerald) and two Norwegian ecotypes (Særheim collected at 58°46′N lat. and Bodø at 67°20′N lat.) of white clover (Trifolium repens L.). For hardening and dehardening, plants were exposed to controlled temperature conditions and frost hardiness of stolons was tested by programmed freezing at the rate of 3°C per hour. In addition, stolons were analysed for starch, soluble sugars and soluble amino acids. Cultivars AberCrest and AberHerald, selected for growth at low temperature and winter hardiness in the United Kingdom, were significantly less hardy than the Norwegian populations. After six weeks of hardening (2 weeks at 6°C and 4 weeks at 0.5°C), estimated LT₅₀ values were ?13.8, ?13.0, ?17.8 and ?20.3°C for AberCrest, AberHerald, Saerheim and Bodø, respectively. The rate of dehardening increased with increasing temperature. At low temperature (6°C), the northern ecotype from Bodø was more resistant to dehardening than AberHerald. However, at 18°C the absolute rate of dehardening (°C day^?1) was twice as high in Bodø as in AberHerald plants. Stolon elongation during dehardening was initiated at lower temperatures in AberHerald than in plants of the Bodø ecotype. The content of total soluble sugars, sucrose and the amino acids proline and arginine were significantly higher in hardy plants of Bodø than in those of AberHerald. Sucrose levels decreased during dehardening and correlations between sucrose content and LT₅₀ during this process were statistically highly significant for both Bodø and AberHerald. The least hardy populations of white clover were characterized by thick stolons, long internodes and large leaves.     

Changing Environmental Effects on Frost Hardiness of Scots Pine During Dehardening

The effects of raised temperature and extended photoperiod onthe dehardening of quiescent and winter-hardy Scots pine saplingswere examined in an open-top-chamber experiment. The saplingswere exposed during winter to natural, square-curve fluctuating(between 1 and 11 °C with a 14 d interval), and constant(6 °C) temperatures with a natural and an extended (17 h)photoperiod. Frost hardiness of needles was determined by controlledfreezing tests and visual damage scoring. The constant 6 °Ctemperature treatment caused a gradual dehardening of needleswhereas under fluctuating temperatures the level of frost hardinessfluctuated. Trees exposed to extended photoperiods were lesshardy than under natural photoperiods after the initiation ofshoot elongation, but before this there were no clear differencesin frost hardiness between different photoperiodic treatments.The results indicate that the frost hardening competence ofScots pine changes during quiescence. Climate change; frost hardiness; hardening competence; photoperiod; Pinus sylvestris, Scots pine; temperature     

Plant resistance to cold stress: Mechanisms and environmental signals triggering frost hardening and dehardening

This introductory overview shows that cold, in particular frost, stresses a plant in manifold ways and that the plant’s response, being injurious or adaptive, must be considered a syndrome rather than a single reaction. In the course of the year perennial plants of the temperate climate zones undergo frost hardening in autumn and dehardening in spring. Using Scots pine (Pinus sylvestris L.) as a model plant the environmental signals inducing frost hardening and dehardening, respectively, were investigated. Over 2 years the changes in frost resistance of Scots pine needles were recorded together with the annual courses of day-length and ambient temperature. Both act as environmental signals for frost hardening and dehardening. Climate chamber experiments showed that short day-length as a signal triggering frost hardening could be replaced by irradiation with far red light, while red light inhibited hardening. The involvement of phytochrome as a signal receptor could be corroborated by respective night-break experiments. More rapid frost hardening than by short day or far red treatment was achieved by applying a short period (6 h) of mild frost which did not exceed the plant’s cold resistance. Both types of signals were independently effective but the rates of frost hardening were not additive. The maximal rate of hardening was − 0.93°C per day and frost tolerance of < − 72°C was achieved. For dehardening, temperature was an even more effective signal than day-length.     

Differences in frost hardiness of two Norway spruce morphotypes growing at Mt. Brocken, Germany

Norway spruce (Picea abies (L.) Karst.) exhibits strong ecotypic variation along altitudinal gradients in morphological traits, e.g. slenderness of crowns or arrangement of second-order branches. We were interested whether montane and lowland morphotypes differ in a key trait for the survival in cold environments, i.e. frost hardiness, and asked: (i) are montane morphotypes more resistant to frost damage and (ii) do they have a lower risk of frost damage by late frosts in spring than lowland morphotypes?We used the electrolyte leakage-method to measure frost hardiness on a monthly basis from October 2006 to May 2007 in stands of the montane and lowland morphotypes at Mt. Brocken in the Harz Mountains, Germany.LT₅₀ (i.e. the temperature that results in 50% of maximum electrolyte leakage) was assessed by freezing treatments in a frost chamber and was significantly influenced by morphotype, month and minimum ambient temperatures. LT₅₀ was significantly lower in the montane than in the lowland morphotype, with −107 °C and −49 °C, respectively. However, the interactions between morphotype with minimum ambient temperature or month were not significant. Thus, as frost hardiness of the two morphotypes responded to temperature in the same way, both morphotypes can be supposed to be exposed to the same risk of frost damage during hardening in autumn and dehardening in spring.     

Genetic and environmental contributions to the winter hardiness of conifers

Summary

Both genetic and environmental components are involved in the processes of winter hardening and dehardening which permit needles of conifers like Norway spruce to survive very low temperatures (< - 30°C) over winter and to recover fully in time for the following and subsequent seasons. One of the major environmental effects of increasing concentrations of atmospheric ozone (O₃) in recent summers has been to affect detrimentally the ability of conifer needles to harden properly and at the correct rate the following autumn. Part of the mechanism by which this occurs in the cytoplasm of needle cells has been traced to detrimental effects on both the Δ¹² fatty acid desaturase and the unusual Δ⁵ desaturase which appears to be part of the low temperature survival mechanism of conifers. The genetic component of winter hardening also involves needle lipids. Studies of lipids in the needles of Norway spruce trees of different provenances growing for many years in the UK have shown that they change during winter hardening as though they were still on trees in the original sites throughout Europe from which the seeds of these trees were initially collected.     

Photosynthetic downregulation in leaves of the Japanese white birch grown under elevated CO2 concentration does not change their temperature‐dependent susceptibility to photoinhibition

To determine the effects of elevated CO₂ concentration ([CO₂]) on the temperature‐dependent photosynthetic properties, we measured gas exchange and chlorophyll fluorescence at various leaf temperatures (15, 20, 25, 30, 35 and 40°C) in 1‐year‐old seedlings of the Japanese white birch (Betula platyphylla var. japonica), grown in a phytotron under natural daylight at two [CO₂] levels (ambient: 400 µmol mol^?1 and elevated: 800 µmol mol^?1) and limited N availability (90 mg N plant^?1). Plants grown under elevated [CO₂] exhibited photosynthetic downregulation, indicated by a decrease in the carboxylation capacity of Rubisco. At temperatures above 30°C, the net photosynthetic rates of elevated‐CO₂‐grown plants exceeded those grown under ambient [CO₂] when compared at their growth [CO₂]. Electron transport rates were significantly lower in elevated‐CO₂‐grown plants than ambient‐CO₂‐grown ones at temperatures below 25°C. However, no significant difference was observed in the fraction of excess light energy [(1 ? q_P)× F_v′/F_m′] between CO₂ treatments across the temperature range. The quantum yield of regulated non‐photochemical energy loss was significantly higher in elevated‐CO₂‐grown plants than ambient, when compared at their respective growth [CO₂] below 25°C. These results suggest that elevated‐CO₂‐induced downregulation might not exacerbate the temperature‐dependent susceptibility to photoinhibition, because reduced energy consumption by electron transport was compensated for by increased thermal energy dissipation at low temperatures.     

Effects of growth stage and hardening conditions on the association between frost resistance and the expression of the cold-induced protein COR14b in barley

Sowing date, being determinant for growth stage, may play a decisive role in optimising freezing resistance of winter annual plants. In cereal species, in spite of the abundant literature analysing the factors responsible for the acquisition of frost resistance through the cold hardening process, the involvement of the growth stage per se, has been seldom considered, especially at the earlier vegetative phases. In this work the contribution of growth stage in determining resistance to freezing temperature has been analysed in field and growth chamber experiments using winter and spring barley cultivars exposed to different hardening conditions. Field damage was assessed twice during winter on plants sown at three different dates. In the growth chamber experiments several acclimation treatments at 11/7 and/or 3/1 °C (day/night) were simulated. In both field and laboratory experiments the development of cold acclimation was monitored by means of a COR14b specific antibody, since in previous studies the expression of COR14b was found genetically linked to frost resistance. The lowest resistance, found in the youngest plants and in spring cultivars, however, was not always associated with the lowest level of COR14b accumulation. COR14b accumulation correlated with frost resistance at the earlier field sampling date and in plants grown at 11/7 °C. In a following phase of the hardening process (second sampling in field and 4 weeks at 3/1 °C in growth chamber) the accumulation of COR14b was independent of plant stage and genotype, showing no association with freezing resistance. Results suggest that growth stage is crucial for the achievement of maximal resistance in barley, but not for COR14b expression.     

On the mechanisms of frost injury and frost hardening of spruce chloroplasts

Hill reaction and noncyclic photophosphorylation of isolated class C chloroplasts of spruce (Picea abies (L.) Karst.), as well as ¹⁴CO₂ fixation by whole needles at constant laboratory conditions proceeded at high rates during spring and early summer, declined during late summer and autumn by about 60%, remained at this level during winter, and recovered quickly in early spring. During summer, the whole needles proved to be frost labile, since after exposure to-20°C and careful thawing, fast chlorophyll degradation occurred. In addition, only photosynthetically inactive chloroplasts could be isolated from those precooled needles. On the contrary, during winter the photochemical activities of plastids from freshly harvested needles did not differ from those of artificially frozen-thawed needles. When isolated spruce chloroplasts were exposed to the same subfreezing temperatures as the whole needles, no influence of freezing on the photochemical activities was observed, irrespective of whether the plastids were isolated from frost sensitive or frost hardened needles. It is concluded that frost damage to spruce chloroplasts is due to an attack of membrane toxic compounds or lytic enzymes which were liberated upon freezing from more labile compartments. Frost hardening of the chloroplasts, as determined by the stability of chlorophyll after exposure of the needles to low temperatures, as well as by the isolation of photosynthetically active chloroplasts from such precooled needles, appeared to depend at least on 2 processes: (i) an alteration of the composition of the photosynthetically active membranes and (ii) and additional stabilization of these membranes by protecting substances. The first process was indicated by a large increase (decrease) of the capability of isolated chloroplasts for PMS-mediated photophosphorylation which accompanied natural or artificial frost hardening (dehardening). Production of cryoprotecting compounds was suggested by a significant higher stability against NaCl observed with class C chloroplasts isolated from frost hardened needles as compared to that of plastids from frost labile material. The decrease of the capability for both, the ferricyanide dependent photoreactions of the plastids and the CO₂ fixation by whole needles, which was observed during the frost hardening phase, cannot be due to freezing injuries; it rather appears to be a consequence of the frost hardening process.     

Periodicity of cambial activity in Abies balsamea. I. Effects of temperature and photoperiod on cambial dormancy and frost hardiness

The relationship between from hardiness and growth potential, and their dependence on temperature and photoperiod, was investigated in the one-year-old cambium of balsam fir [Abies balsamea (L.) Mill.]. Six-year-old trees were exposed for 9 weeks to either the natural environment or one of 4 controlled environments in the fall (18 September-18 November), spring (12 April–14 June) and summer (19 July – 19 September). The 4 controlled environments were (1) WS, warm temperature (24/20°C in day/night) + short day (8 h). (2) WL. warm temperature (24/20°C) + long day (8 h + 1 h night break), (3) CS. cold temperature (9/5°C) + short day (8 h) and (4) CL, cold temperature (9/5°C) + long day (8 h + 1 h night break). At the beginning and end of each exposure, cambial activity was measured by recording the number of xylem, cambium and phloem cells, frost hardiness was estimated from the cambium's ability to survive freezing to –40°C, and cambial growth potential was deduced from the duration of the cell cycle and the production of xylem, cambium and phloem cells in cuttings cultured for 4 weeks with exogenous indole-3-acetic acid (IAA) under environmental conditions favourable for cambial activity. In the natural environment, frost hardening began in September and was completed in November, while dehardening occurred when the cambium reactivated. CL, CS, and to a lesser extent WS, promoted hardening in the summer and fall, but did not prevent dehardening in the spring. The cambial growth potential in the natural environment declined from a maximum in April to a low level in June, reached a minimum in September, then increased to a high level in November. This potential was promoted by CL and CS on all dates by WL in the summer and fall. The ratio of xylem to phloem induced by IAA treatment was greatest in June and least in September in cuttings from trees exposed to the natural environment, and was increased by CL and CS in the fall. The cambium in intact branches of trees protected from chilling during the fall and winter resumed cell cycling after less than 9 weeks of dormancy, but produced mostly or only phloem in the subsequent growing period. It is concluded that the frost hardiness of the cambium, the IAA-induced cycling of cambial cells, and IAA-induced xylem to phloem ratio vary independently with season, temperature and photoperiod, and that the periodicity of these processes is regulated endogenously.     

Divergent mechanisms of frost-hardiness in two populations of the gall fly, Eurosta solidaginsis

Two populations of the gall fly Eurosta solidaginsis utilize different strategies to endure seasonal exposure to temperatures below freezing. Both populations are freezing tolerant. In north temperate populations, supercooling points rise from ?10.2°C to ?6.2°C following exposures to temperatures below freezing. This level is maintained throughout winter and ensures frequent and prolonged periods of tissue freezing. South temperate populations depress the supercooling point to ?14.2°C during autumn and early winter, and this depression precludes extracellular ice formation during periods of supra-optimal temperature fluctuations. During mid-winter, supercooling points rise to the same level as in northern groups.Both populations accumulate three principal cryoprotective agents following first frost exposures (glycerol, sorbitol and trehalose). Cryoprotectants levels do not peak in northern populations until 4–6 weeks after first frost. In southern populations the accumulation profile is characterized by a high initial rate of synthesis, a protective overshoot and pronounced seasonal fluctuations. The relative survival advantages of each strategy are discussed.     

Frost dehardening and rehardening of floral buds of deciduous azaleas are influenced by genotypic biogeography

Temperate-zone woody perennials may resist cold dehardening and reharden effectively after unseasonably warm winter conditions to avoid frost damage. Few controlled experiments have examined dehardening kinetics or the impact of dehardening on rehardening capacity after cold temperatures return. We used nine genotypes of deciduous azalea from eight known provenances to study the influence of biogeographical origin on floral bud dehardening and rehardening after controlled dehardening. Buds cold acclimated in the field were placed in warm conditions to stimulate dehardening. Visual assays were conducted periodically over 11 days of dehardening to evaluate survival of immature florets at subfreezing temperatures. A rehardening regime was applied to three genotypes after 1, 3, 5, and 8 days of dehardening. Dehydrin abundance after dehardening and rehardening was estimated for selected genotypes. Floral buds from warmer-climate azaleas Rhododendron canescens, Rhododendron prunifolium, and Rhododendron viscosum variety serrulatum exhibited lower mid-winter hardiness than did the colder-climate azaleas Rhododendron calendulaceum, Rhododendron canadense, Rhododendron prinophyllum, and Rhododendron viscosum variety montanum. The dehardening rates of the “low dehardening-resistant” R. canadense, R. canescens, and R. viscosum var. serrulatum were at least twice the rates of “high dehardening resistant” Rhododendron arborescens, Rhododendron atlanticum, R. calendulaceum, R. prinophyllum, R. prunifolium, and R. viscosum var. montanum throughout the time-course. Genotypes originating in colder and warmer climates did not always exhibit high and low dehardening-resistance, respectively. Dehardening was associated with declining levels of dehydrins in R. prinophyllum and the two R. viscosum varieties. All tested genotypes rehardened in response to cold even after 8 days of dehardening. The high-altitude variety of R. viscosum had substantially larger rehardening-capacity than the low-altitude variety. Rehardening was associated with increasing levels of dehydrins in both R. viscosum varieties. Mid-winter hardiness ≥26.0 °C, dehardening rates ≤1.0 °C day⁻¹, a capacity to reharden, and the ability to accumulate dehydrins could all be important winter survival strategies for genotypes originating in colder climates.     

Seasonal acclimatization in metabolic rate of the fan-fingered gecko,Ptyodactylus hasselquistii (Reptilia: Gekkonidae)

The resting metabolic rate of the fan-fingered gecko Ptyodactylus hasselquistii of various body masses was determined in relation to ambient temperatures ranging from 20 to 35°C during winter and summer acclimatization. Oxygen consumption (ml g⁻¹ h⁻¹) decreased with increasing mass at each temperature. The intraspecific exponents of body mass in relation to metabolic rate ranged from 0.62 to 0.79. Winter-acclimatized geckos had significantly lower metabolic rates than summer-acclimatized geckos at different temperatures, especially at low temperature (20°C). The pattern of acclimatization exhibited by P. hasselquistii may conserve energy during inactivity in winter and make activity more easily achieved during active seasons.     

Effects of artificial frost hardening and winter stress on net photosynthesis, photosynthetic electron transport and RuBP carboxylase activity in seedlings of Pinus silvestris

Net photosynthesis of seedlings of Pinus silvestris has been measured and compared with the activities of photosynthetic electron transport and extracted RuBP carboxylase. The effects of prolonged frost hardening (photoperiod 8 h, + 3°C) followed by winter stress at subzero temperatures were analysed. There was a parallel effect of frost hardening and winter stress on the photosynthetic properties of both intact seedlings and isolated chloroplast thylakoids. The activity of extracted RuBP carboxylase was less affected by the treatments. In relation to earlier works we conclude that the decay of net photosynthesis in winter climate is determined by the electron transport properties of the chloroplast thylakoids, i.e. by the pool sizes of photosynthetically active plastoquinone. The results of this work justify the definition of two phases in the response of conifers towards autumn and winter climates: I. Frost hardening occurs at temperatures slightly above zero and it does not affect the efficiency of photosynthesis as defined by the quantum yield at rate limiting light absorption. II. Winter stress occurs at subzero temperatures and it is characterized by a suppression of the photosynthetic efficiency as a result of damage within the photosynthetic apparatus.     

Effect of frost nights and day and night temperature during dormancy induction on frost hardiness, tolerance to cold storage and bud burst in seedlings of Norway spruce

For trees, the ability to obtain and maintain sufficient levels of frost hardiness in late autumn, winter and spring is crucial. We report that temperatures during dormancy induction influence bud set, frost hardiness, tolerance to cold storage, timing of bud burst and spring frost hardiness in seedlings of Norway spruce (Picea abies (L.) Karst.). Bud set occurred later in 12°C than in 21°C, and later in cool nights (7°C) than in constant temperature. One weekly frost night (−2.5°C) improved frost hardiness. Cool nights reduced frost hardiness early, but improved hardiness later during cold acclimation. Buds and stems were slightly hardier in 21°C than in 12°C, while needles were clearly hardier in 12°C. Cold daytime temperature, cool nights and one weekly frost night improved cold storability (0.7°C). Seedlings receiving high daytime temperatures burst buds later, and were less injured by light frost some days after bud burst.     

Hardening and Dehardening of Lolium perenne in Response to Fluctuating Temperatures

Hardening and dehardening responses of two contrasting varietiesof Lolium perenne, measured as LT50 estimates, were followedin fluctuating temperature environments. Unhardened seedlingswere exposed to hardening environments for 7, 14, and 21 d inall combinations of 2, 4, 6, 8 and 10 C with either high dayand low night temperatures or low day and high night temperatures.Seedlings hardened for 28 d at 2 C were exposed to dehardeningenvironments in all combinations of 4, 6, 8, 10, and 12 C withhigh day and low night temperatures. A low day, or night, temperature of 2 C in combination withany other temperature increased hardening compared with theconstant higher temperature. For Premo, a hardy variety, thisincrease was 3 C when night temperature was reduced from 10to 2 C in combination with a day temperature of 10 C. Similarly,a low night temperature reduced the dehardening response ofPremo to higher day temperatures. At 12 C this effect on LT60was greater than 2 C. Much smaller responses to daily periodsof low temperature were found for the less hardy variety, GrasslandsRuanui. During each 24-h period, exposures to 2 C of longer than 4h were required to achieve greater hardening than that achievedin continuous 10 C treatments. Hardiness was not improved furtherby exposures longer than 8 h. Responses to diurnal temperature fluctuations were discussedin relation to possible mechanisms and to changes in hardinessduring the winter under different weather systems. Lolium perenne, cold hardening, cold dehardening, diurnal temperature fluctuations, varieties     

Relationship between respiratory depletion of sugars and loss of cold hardiness in coniferous seedlings over-wintering at raised temperatures: indications of different sensitivities of spruce and pine

Long-term effects of elevated winter temperatures on cold hardiness were investigated for Norway spruce (Picea abies L. Karst.), lodgepole pine (Pinus contorta Dougl.) and Scots pine (Pinus sylvestris L.). Two-year-old seedlings with the same pre-history of growth and cold hardening in the field were maintained from early December to late March at two field sites in northern Sweden and in a cold room. The temperatures at these locations averaged –13·5, –8·9 and 5·5°C, respectively. Following treatments, carbohydrate contents and cold tolerances were assessed. Needle respiration was also analysed during the 5·5°C treatment. Cold tolerance of lodgepole pine and Scots pine was much reduced following the 5·5°C treatment. Cold tolerance was somewhat reduced in lodgepole pine following the –8·9 °C treatment, but was essentially maintained in spruce throughout all treatments. The cold tolerance of needles was strongly correlated with their soluble sugar contents. Spruce maintained cold hardiness by having larger reserves of sugars and lower rates of respiration which decreased more rapidly as sugars were depleted. Tolerance of lodgepole pine to frost desiccation was also much reduced following the 5·5°C treatment.     

Winter hardening and glutathione status in the bilberry (Vaccinium myrtillus) in response to trace gases (CO2, O3) and nitrogen fertilization

Bilberry plants (Vaccinium myrtillus L.) at a field site in northern Finland (65°N) were subjected to nitrogen fertilization [6.5 mmol m^?2 NH₄NO₃× Ca(OH)₂] at the beginning of 3 growing seasons in late May and to trace gas fumigation (CO₂ and O₃) for 5 months (May–September) in 1993–1995 in order to investigate frost resistance and glutathione concentrations during the winter hardening period, and to assess the correlation between these variables. Harvesting was performed twice in the autumn of both 1994 and 1995, and the two-year data for each harvest were pooled. The frost resistance of the bilberry stems increased by about 10°C during the hardening period between the two harvests. Nitrogen fertilization increased the frost resistance towards late autumn. The fumigation treatments had no marked effect on it. The combination of elevated CO₂ and nitrogen fertilization induced a decrease in frost resistance. Increases in total glutathione concentrations and the proportion of reduced glutathione (GSH) in the stems were evident during hardening. Nitrogen fertilization positively affected the total glutathione concentration and the proportion of GSH at the beginning of the hardening period but the effect disappeared during the hardening process. Trace gas fumigation as such had no marked effect on glutathione concentration. Increases in glutathione concentrations during hardening did not correlate with frost resistance, possibly due to different timing of the appearence of the response to fertilization treatment, i.e., glutathione responded in the beginning of hardening while frost resistance at the end. The lack of correlation with frost resistance, and especially the different responses to nitrogen fertilization, may reflect the indirect role of glutathione in the development of winter hardening, as a transport and storage form of reduced nitrogen and sulphur. In conclusion, winter hardening and glutathione status in the bilberry seems to be sensitive to nitrogen fertilization, and not affected by elevated CO₂ and O₃.     

Regulation of the loss of frost hardiness in Pinus radiata by photoperiod and temperature

Abstract. In controlled environments, the interactive effects of warm (16: 8°C, day: night) and cool (12: 4°C, day: night) temperatures and long (13.5 h) and short (10 h) photoperiods on the dehardening of seedlings of Pinus radiata D. Don were investigated. In another experiment, the effect of four photoperiods from 9 to 14 h was examined. In a third, dehardening at constant temperatures from 5 to 17°C was followed. There was no evidence for an interaction between photoperiod and temperature. Dehardening was temporarily delayed by photoperiods below about 10 h, but there was no other quantitative effect of photoperiod. At constant temperatures, the rate of dehardening was initially constant but declined as the minimum summer frost hardiness was reached. In the initial phase the rate of dehardening was a linear function of temperature, increasing from 0.05°C day⁻¹ at 8°C to 0.30 °C day⁻¹ at 17°C. Temperature controlled the loss of frost hardiness by regulating the rate of dehardening.     

Altitudinal and seasonal variation of frost resistance of Fagus crenata and Betula ermanii along the Pacific slope of Mt. Fuji, Japan

1 Frost resistance of Fagus crenata (Siebold's beech) and Betula ermanii (Japanese mountain birch) was investigated with respect to the species' altitudinal distribution on the Pacific slope of Mt. Fuji from 1996 to 1997. Flint's Index of Injury, which is based on electrolyte leakage from freeze-injured tissue, was used to assess frost hardiness of shoots produced in the previous growing season.
2 Fagus crenata is found on the lower slopes (700–1600 m a.s.l.). Mid- to late November hardening of shoots was enhanced, midwinter damage below −30 °C reduced and dehardening delayed nearer the upper limit. To here temperatures began to rise at least 3 weeks before dehardening began. Shade crown shoots were more susceptible to deep-freeze damage than light crown shoots. If the ultimate upper distribution limit was determined by frost hardiness, F. crenata would be expected to occur up to 1800 m altitude.
3 Betula ermanii is found between 1600 m and 2800 m, and intensive hardening occurred at all altitudes during the second half of October. Frost hardiness increased considerably with altitude up to the forest limit, where frost acclimation preceded the temperature decline by 2 weeks. Once maximum frost resistance had been attained freezing to −47 °C failed to cause tissue injury. Dehardening began slightly later at the tree line, but the time–course was the same at all altitudes. Main and lateral shoots did not differ in frost hardiness.
4 Comparison of monthly air temperature minima over the past 66 years with the course of frost resistance showed that F. crenata and B. ermanii found on the Pacific slope of Mt. Fuji were unlikely to suffer damage by frost.
5 The observed uppermost distribution limit for B. ermanii at 2800 m altitude on Mt. Fuji is considered both with our observations and with previous hypotheses.     

Effect of low and high temperatures on the photosynthetic performance of Lantana camara L. Leaves in darkness

Low and high temperatures are known as most important factors influencing plant performance and distribution. Plants of Lantana camara L. coming from two distinct geographical populations (Iberian Peninsula and Galápagos Islands) were cultivated in a common garden experiment, and their leaves were subjected to thermal treatments (from +20.0 to ?7.5°C during the winter and from +20.0 to +50.0°C during the summer) in a programmable water bath in darkness. Their photosynthetic performance and their recovery capacity after the thermal treatment were evaluated by measuring chlorophyll fluorescence, net photosynthesis rate, and leaf necrosis. In general, L. camara photosynthetic apparatus showed a wide range of temperature tolerance in darkness, showing optimal functioning of its photosystem II just after exposure to temperatures between ?2.5 and +35.0°C for the Iberian population and between +10.0 and +25.0°C for the Galápagos population. Just after exposure to low and high temperatures, gradual cold and heat-induced photoinhibition was recorded for both populations. After 24 h, leaves of L. camara demonstrated a great recovery capacity from ?2.5 to +42.5°C. However, leaves of the treatments from ?5.0°C down and +47.50°C up showed permanent damages to the photosynthetic apparatus and to the leaf tissues. Slight interpopulation differences were found only at extreme temperatures.