Portal de Periódicos da CAPES

Dehardening of mountain birch ( Betula pubescens ssp. czerepanovii ) ecotypes at elevated winter temperatures

2004; Wiley; Volume: 162; Issue: 2 Linguagem: Inglês

10.1111/j.1469-8137.2004.01042.x

1469-8137

Kari Taulavuori, Erja Taulavuori, O. Skre, Jarle Nilsen, Bernt Igeland, Kari Laine,

Plant Water Relations and Carbon Dynamics

New PhytologistVolume 162, Issue 2 p. 427-436 Free Access Dehardening of mountain birch (Betula pubescens ssp. czerepanovii) ecotypes at elevated winter temperatures Kari M. J. Taulavuori, Corresponding Author Kari M. J. Taulavuori Department of Biology, University of Oulu, PO Box 3000, FIN-90014, Oulu, Finland;Author for correspondence: Kari M. J. Taulavuori Tel: +358 8 553 1512 Fax: +358 8 553 1061 Email: [email protected]Search for more papers by this authorErja B. Taulavuori, Erja B. Taulavuori Department of Biology, University of Oulu, PO Box 3000, FIN-90014, Oulu, Finland;Search for more papers by this authorOddvar Skre, Oddvar Skre Norway Forest Research Institute, Bergen, Norway;Search for more papers by this authorJarle Nilsen, Jarle Nilsen Department of Biology, University of Tromsø, NorwaySearch for more papers by this authorBernt Igeland, Bernt Igeland Department of Biology, University of Tromsø, NorwaySearch for more papers by this authorKari M. Laine, Kari M. Laine Department of Biology, University of Oulu, PO Box 3000, FIN-90014, Oulu, Finland;Search for more papers by this author Kari M. J. Taulavuori, Corresponding Author Kari M. J. Taulavuori Department of Biology, University of Oulu, PO Box 3000, FIN-90014, Oulu, Finland;Author for correspondence: Kari M. J. Taulavuori Tel: +358 8 553 1512 Fax: +358 8 553 1061 Email: [email protected]Search for more papers by this authorErja B. Taulavuori, Erja B. Taulavuori Department of Biology, University of Oulu, PO Box 3000, FIN-90014, Oulu, Finland;Search for more papers by this authorOddvar Skre, Oddvar Skre Norway Forest Research Institute, Bergen, Norway;Search for more papers by this authorJarle Nilsen, Jarle Nilsen Department of Biology, University of Tromsø, NorwaySearch for more papers by this authorBernt Igeland, Bernt Igeland Department of Biology, University of Tromsø, NorwaySearch for more papers by this authorKari M. Laine, Kari M. Laine Department of Biology, University of Oulu, PO Box 3000, FIN-90014, Oulu, Finland;Search for more papers by this author First published: 11 March 2004 https://doi.org/10.1111/j.1469-8137.2004.01042.xCitations: 46AboutSectionsPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Summary • The aim was to elucidate the effects of elevated winter temperatures on the dehardening process of mountain birch (Betula pubescens ssp. czerepanovii) ecotypes and to evaluate their susceptibility to frost damage under warming climate conditions. • Ecotypes from 60 to 71° N latitudes and 20–750 m altitudes were grown in northern Norway (70° N) and subjected to simulation of the photoperiod in southern Norway (60° N) by artificial illumination from September onwards. In November, the seedlings were transported to the south (60° N) to overwinter at ambient or 4°C above ambient temperatures. Frost hardiness and lipid peroxidation were determined during January–April. • The higher winter temperature accelerated dehardening, and there were significant differences between the ecotypes. Among tree individuals of southern origin, the alpine ecotype exhibited the most rapid rate of dehardening, whereas the oceanic type showed the slowest rate. Lipid peroxidation supported the above findings. • Since temperature elevation was unequal for the ecotypes with respect to climatic change, the frost hardiness results were normalized to obtain an equal +4°C temperature rise. The risk of frost injury seemed to be lowest in the northernmost ecotypes under a temperature elevation of +4°C, obviously due to their adaptation to a wider temperature range. Introduction The current atmospheric CO2 concentration of c. 370 ppm is expected to double by the end of 2100. Consequently, the globally averaged surface temperature is projected to increase by 1.4–5.8°C. The most pronounced warming is predicted to occur at high latitudes, especially in winters, with an anticipated increase of c. 4–7°C for the boreal zone (Bach, 1988; Maxwell, 1992; Murray, 1995; Houghton et al., 2001). In perennial plant species, frost hardiness is strongly controlled and determined by the prevailing ambient temperatures, in that warm winter temperatures reduce frost hardiness (e.g. Sakai & Larcher, 1987). The increasing effect of climatic warming on the incidence of the risk of frost damage has been debated during the past two decades. Theoretical approaches have emphasized the possible risk, especially in late spring (e.g. Cannell & Smith, 1986; Hänninen, 1991), and this finding has been supported by some experimental and field studies (Repo et al., 1996; Taulavuori et al., 1997a; Gansert et al., 1999). However, the risk that the climate change will cause premature dehardening has been thought to be small for native species in Scandinavia (Ögren, 2001). Mountain birch (Betula pubescens ssp. czerepanovii) is the most important tree-line species in Scandinavian forests (Skre, 1993). The mountain birch forest zone constitutes the northernmost boundary of boreal forest in the Scandinavian mountain chain, and the forest is thus exposed to severe climatic restraints (Tenow, 1996). There was a horizontal zone at 410–430 m a.s.l. in Abisko, northern Sweden (68° N), where the mountain birch did not leaf in the summer of 1991. It was hypothesized that the damage was caused by a cold backlash in mid-April after a very warm first half of the month. Similar hazards have also been recorded in 1962, 1963 and 1985 (Tenow, 1996). Therefore, in addition to theoretical and experimental documentation, there is also some natural evidence to support the assumption that springtime frost damage evolves after unseasonably warm spells. Adaptation to locally variable climatic and meteorological conditions has resulted in the evolution of genetic ecotypes. Ecotypes in cold regions evolve to derive maximum benefit from the short period that enables photosynthesis, growth and reproduction. On the other hand, correct timing of growth cessation and the length of the dormant state in the overwintering period are necessary for the survival of plants at high northern latitudes. The dormant state is broken by two phases: accumulation of air temperatures near +5°C in winter (i.e. the chilling requirement for completion of rest) and the consequent long-term exposure to temperatures above a given threshold required for growth (i.e. the high-temperature requirement for onset of growth) (Hänninen, 1995). The latter requirement for growth resumption is probably more important than the chilling requirement for dormancy release (Häkkinen et al., 1998; Pop et al., 2000). The lower threshold air temperature for zero growth in birch is 7–7.8°C (Kullman, 1993). The chilling requirement consists of two components, the temperature and the duration of chilling, and the latter requirement for northern deciduous trees varies considerably both between and within species (Heide, 1993; Myking & Heide, 1995). The chilling requirement of birch species decreases significantly along with increasing latitude of origin (Myking & Heide, 1995). In addition, high altitude (e.g. Murray et al., 1989) and continental climate (Leinonen, 1996b) are factors that minimize the chilling requirement of origin, although the relative importance of these aspects is unknown. Thus, at northern high latitudes, the milder the climate, the higher is the chilling requirement, with a longer winter-to-spring time period of low risk of frost damage under warming climate. The present study consisted of experimental warming of mountain birch ecotypes (Table 1). The aim was to elucidate the effects of elevated winter temperatures on the dehardening process of these ecotypes. Dormancy and frost hardiness are separate adaptive traits, and frost hardiness does not directly reflect the state of dormancy at any time of the year. However, the rate of dehardening or the capacity to maintain frost hardiness depends indirectly on the state of dormancy, since frost hardiness depends on the ontogenetic cycle of a plant (e.g. Fuchigami et al., 1982) in addition to environmental fluctuations (Repo et al., 1990). There is evidence that dehardening of northern woody species starts gradually much before bud break, although frost hardiness is eventually lost in parallel with the resumption of growth (see Repo, 1992; Taulavuori et al., 1997a,b). The buds maintain a high degree of frost hardiness until opening, while the cambial activity of stem tissue arises due to warm spells in the spring, making the plant vulnerable to frost damage (e.g. Zalasky, 1976). In the present study the frost hardiness in stem tissue (e.g. cambium) was determined and followed in late winter and spring. Because of their shared dependency on ontogenetic development, we decided to test if the rate of dehardening is linked to their chilling requirement, and which of the following hypotheses best fits the observed pattern of the dehardening rate. Table 1. Seed origins representing the mountain birch (Betula pubescens ssp. czerepanovii) ecotypes investigated in the experiment Seed origin Latitude Altitude Dist. to ocean Tmean* Tminabs** NB (Blefjell, N) 60° N 750 m 100 km −4.3°C −12.6°C IC (Hafnaskogur, IC) 63° N 50 m < 5 km ±0.0°C −9.3°C NMe (Melbu, N) 68° N 20 m < 5 km +0.6°C −9.7°C FJ (Kevo/FIN) 69° N 250 m 200 km −11.9°C −33.3°C NHa (Hammerfest/N) 71° N 50 m < 5 km −4.2°C −17.2°C * Mean winter (Dec–Mar) temperature based on 30-year data (1931–1960). ** Absolute minimum temperature in Mar based on 30-year data (1931–1960). Hypothesis 1 Dehardening of the mountain birch ecotypes follows the pattern derived from their chilling requirement. This assumes that the chilling requirement of ecotypes decreases towards: (i) northern high latitudes; (ii) high altitudes; and (iii) continental sites (Fig. 1). Figure 1Open in figure viewerPowerPoint Three-dimensional plot to illustrate the basis for Hypothesis 1. E denotes the hypothetical ecotype with the highest chilling requirement and a low risk of spring frost damage. E′ stands for the opposite ecotype with the lowest chilling requirement and a consequent high risk of spring frost damage. This is based on the following assumptions: chilling requirement of ecotypes decreases towards (1) northern high latitudes (2) high altitudes and (3) continental sites. The NB, FJ, NHa, NMe and IC are the investigated ecotypes (see Table 1 for their spatial parameters). According to Hypothesis 1, birch ecotypes of different climatic origin are to be arranged from low to high chilling requirement (and low to high capacity to maintain frost hardiness under warming winter climate, respectively) given as: Order 1 (Hypothesis 1): NB; FJ < NHa; Nme < IC Hypothesis 2 The assumptions given in the context of Hypothesis 1 may be generalized as: The chilling requirement of ecotypes decreases towards harsh winter climate. Thus, if ‘harshness’ of winter is a reflection of local temperature conditions, the order derived from Hypothesis 1 can be modified as: Order 2 (Hypothesis 2): FJ < NHa; NB < Nme; IC According to this hypothesis, the ecotypes are arranged from low to high temperature climate (see Table 1), which reflects the capacity to maintain frost hardiness on a similar basis as in Hypothesis 1. In addition, there is evidence that light climate plays a role in the start of ontogenetic development from dormant to active growth phase (Linkosalo et al., 2000). Therefore, we performed the present experiment in a common garden-like system: the seedlings experienced the same photoperiod over frost hardening to dehardening phases irrespective of their ecotype. Once light has an effect on the ontogenetic development during dehardening, the ambient daylength in Bergen (60° N) should favour the dormancy release of the ecotypes from higher latitudes. The hypothetical order to maintain frost hardiness is thus: Order 3 (Hypothesis 3): NHa; FJ; Nme < IC < NB In order to follow the dehardenining process we determined the frost hardiness (LT50) of stems of the mountain birch seedlings. In addition, we analysed lipid peroxidation (malondialdehyde) to investigate stress response in cell membranes, which are the primary site of freezing injury (Steponkus, 1990). Lipid peroxidation is a widely used stress indicator of plasma membranes (e.g. Taulavuori et al., 2001), which undergo both biochemical and biophysical changes during cold acclimation. The purpose of lipid peroxidation analysis in the present investigation was to provide additional information about the physiological state of stress attributed to cold hardiness. The final aim of this work was to evaluate the risk of the studied mountain birch ecotypes to frost damage under warming climate conditions. Materials and Methods Experimental design Seeds from 6 to 8 trees per population from five different mountain birch (B. pubescens Ehrh. ssp. czerepanovii (Orlova)) ecotypes (Table 1, Fig. 1) were sown into the glasshouse of the University of Tromsø during 2 yrs, 2000 and 2001. In 2000 the seeds were stratified for 20 d and sown on 30 June, and in 2001 they were stratified for 28 d and sown 1 month later (6 August). The material in 2000 was kept at 18°C and in the ambient light conditions with a 24 h photoperiod until September. The material in 2001 was also kept at 18°C. The 24 h photoperiod was supplemented with an artificial lamp system (Osram 58 W/30; 200 µmol m−2 s−1) because of the late season for growth. On the 7 and 18 of September (2000 and 2001, respectively), the seedlings were potted in 10 cm containers with fertilized peat (Skre, 1993) and watered regularly once a week with SUBERBA nutrient solution equivalent to 10 g N m−2 yr−1 and placed at two autumnal temperatures (+9°C and +15°C) for hardening in the ambient daylight of Bergen (60° N). On 1 and 22 November (in 2000 and 2001, respectively), the seedlings were transferred for overwintering to the Norwegian Forest Research Institute in Bergen. They were subdivided between two glasshouse compartments with different temperatures: either ambient (unheated) or approx. 4°C above ambient (heated) with an ambient day length of 6 h. The daylight intensity in the glasshouse was reduced by c. 20–50% relative to the outside. The daily minimum temperatures during the overwintering period are shown in Fig. 2. Figure 2Open in figure viewerPowerPoint Daily minimum temperatures in Bergen during the overwintering periods used in the experiments in (a) 2000–2001 and (b) 2001–2002. Day 1 denotes November 1. The lower curve stands for ambient and the upper curve for elevated temperatures. To sum up, the experimental design consists of dehardening determinations during two springs. During both springs (2001 and 2002), a sample set containing 40 seedlings was transported three times to the University of Oulu (Finland) for further analysis. The sampling dates were: 18 January, 1 February and 22 March in 2001, and 24 January, 21 February and 4 April in 2002. Frost hardiness The frost hardiness test was similar to that described by Gansert et al. (1999) for another mountain birch species (Betula ermanii), with the exception of propanol infiltration, which was substituted by a standard shaking procedure in Millipore water. Main shoots were taken from below the top crown and exposed to the following temperatures in the following order: −10, −20, −40, −70 and −196°C (= LN) (Month 1 and 2), and −2, −10, −20, −40 and −196°C (Month 3). Stem pieces of 10–15 mm were moistened, wrapped in aluminium foil and frozen to the test temperature. The freeze-thaw procedure, the consequent electrolyte leakage and the calculation of LT50 were done as described earlier for Pinus sylvestris (Taulavuori et al., 1997b, 2000), Vaccinium myrtillus (Taulavuori et al., 1997, 1997a) and V. vitis-idaea (Taulavuori et al., 2001). The special adaptation of the method (preliminary results, unpublished data) to the stem material of birch species required: (1) dissection of buds after the freeze-thaw procedure to obtain a homogeneous organ (stem); (2) the remaining internode was cut into 3 segments of approx. 3–5 mm to accelerate the rate of electrolyte leakage out of the tissue. Normalized hardiness at +4°C temperature elevation Because of the climatic adaptations, the ecotypes experienced unequal temperature elevations in Bergen. Therefore, frost hardiness was normalized at the +4°C temperature elevation corresponding to the mean winter temperatures of each ecotype, in order to allow evaluation and comparison of the susceptibility of the ecotypes to frost injuries under conditions of global climatic warming. The normalization was performed as follows (Fig. 3): Figure 3Open in figure viewerPowerPoint An example of normalized hardiness (NH) at +4°C. The numerals in the parentheses refer to the text. 1 The ‘Ambient’ and ‘Elevated’ treatments were calculated to obtain the temperature elevation experienced by a given ecotype. The calculated temperature elevation was based on the difference between the mean temperatures in winter adapted for seed origin (Table 1) and the mean temperatures experienced in Bergen. The mean temperatures in Bergen were averages of the weekly mean values from the same period (December–March) as the reference values in Table 1. The long-term data from vicinity of origins of all the ecotypes concern the period 1931–1960, and provide an adequate basis to evaluate environmental adaptation (genetic) because of the long life span of trees. The resulting mean temperatures in Bergen were +5.5 and +2.1°C for the elevated and ambient temperature treatments in the winter of 2000–2001 and +6.9 and +3.5°C in the following winter, respectively. 2 The determined LT50 values of the last sampling for each year (March and April in 2001 and 2002, respectively) were plotted against the true temperature elevation obtained by this method. 3 Normalized hardiness (NH) was obtained through extrapolation of the response line to intercept the true temperature elevation at +4°C. 4 Subsequent interpolation to the Y-axis. Lipid peroxidation Lipid peroxidation was analysed by the malondialdehyde (MDA) method (Hodges et al., 1999, Taulavuori et al., 2001), with the exception of minor modifications during homogenization. A 0.4 g sample was homogenized in liquid nitrogen with a mortar and pestle. The homogenized tissue powder was suspended in 5 ml of 0.1% TCA on ice, after addition of 0.6 g of PVPP (poly vinyl-poly pyrrolidone) in 4 ml of 0.1% TCA (trichloro acetic acid). Extraction was continued as described by Taulavuori et al. (2001), including the following steps: centrifugation, TCA/TBA (thiobarbituric acid) addition, heat/cool cycle and another round of centrifugation. The absorbance of supernatant was read at 440, 532 and 600 nm, and each sample had a reference without TBA. The MDA equivalents were calculated according to Hodges et al. (1999). Statistical analyses The statistical analyses were performed with the anova General Linear Model (SPSS software) stepwise as follows. The main effects of the factors: (1) year, (2) month, (3) seed origin (ecotype), (4) autumn temperature, and (5) winter temperature were tested first (Table 2). Since the temporal variation (yearly, monthly) in ambient temperature determines frost hardiness, and the temperature differed markedly between the years, further analysis of the data was performed using successive years as separate cases. Each month was analysed step by step. Since the autumn temperature (Table 2) showed no main effect on frost hardiness, the data based on the two autumn temperatures were pooled, thus resulting in a replicate number of four (n = 4). The data on lipid peroxidation were analysed similarly. Table 2. anova results from the 2-year data of the main effects on frost hardiness on mountain birch (Betula pubescens ssp. czerepanovii) ecotypes Source of variation SS d.f. F Corrected model (R2 = 0.88) 113817.32 119 6.95*** Intercept 217990.17 1 1583.19*** Y (year) 34767.46 1 252.50*** M (month) 12154.21 2 44.14*** E (ecotype) 8071.95 4 14.66*** AT (autumn temperature) 119.36 1 0.867 NS WT (winter temperature) 8009.47 1 58.17*** Year × M 662.43 2 2.41 NS × E 6940.14 4 12.60*** × AT 507.22 1 3.69 NS × WT 6881.23 1 49.98*** × M × E 2228.50 8 2.02 NS × M × AT 770.41 2 2.80 NS × M × WT 597.34 2 2.17 NS × AT × WT 125.98 1 0.92 NS × E × AT 2756.49 4 5.00** × E × WT 2065.76 4 3.75** × M × E × AT 1009.30 8 0.92 NS × M × E × WT 1634.85 8 1.48 NS × M × AT × WT 389.49 2 1.41 NS × E × AT × WT 1066.94 4 1.94 NS × M × E × AT × WT 2751.66 8 2.50* Month × E 2136.99 8 1.94 NS × AT 281.48 2 1.02 NS × WT 2197.90 2 7.98** × E × AT 1598.54 8 1.45 NS × E × WT 3689.69 8 3.35** × AT × WT 805.36 2 2.93 NS × E × AT × WT 2143.44 8 1.95 NS Ecotype × AT 1077.12 4 1.96 NS × WT 1360.43 4 2.47* × AT × WT 464.49 4 0.84 NS Autumn temperature × WT 517.67 1 3.77 NS Error 15696.74 114 Total 359337.13 234 Corrected total 129514.06 233 The F-values contain indication of significance as follows: *, P < 0.05; **, P < 0.01; ***, P < 0.01; NS, nonsignificant. Results Main effects on frost hardiness The anova table (Table 2) shows the highly significant effects of temporal variation (year, month) on frost hardiness. For example, the averaged frost hardiness based on the complete data of 2001 and 2002 shows LT50 = −44 and −19°C, respectively. The seedlings were thus twice as hardy during the first studied year compared to the second. This reflects the higher winter temperature of approx. 2°C in 2002 and the younger physiological age of the seedlings in that year. In addition, the last sampling in 2002 occurred c. 2 weeks later. Frost hardiness decreased throughout the experiments in both years, indicating the degree of dehardening as follows: LT50 =−49 (January 2001) and −29°C (January 2002), LT50 = −47 (February 2001) and −18.6°C (February 2002), and LT50 =−35 (March 2001) and −9°C (April 2002). The difference between the autumn temperatures (+9 and +15°C) had no effect on frost hardiness, although it interacted with ecotype and year partly due to the above reasons. As a consequence of interaction, the northernmost ecotypes (NHa and FJ) dehardened easier in 2001 when they first experienced the lower growing temperature before overwintering at elevated winter temperatures. However, winter temperature and seed origin (i.e. ecotype) showed a highly significant effect on frost hardiness. The approx. 4°C elevation in winter temperature from ambient levels in Bergen thus reduced the frost hardiness of mountain birch seedlings. The effects of the seed population were variable. The 2- year patterning of the ecotypes (from the most dehardened to hardened) was as shown in Order 4 (differences at P < 0.5). Order 4 NB ≤ NHa, FJ, NMe ≤ IC The Iceland ecotype (IC) thus maintained its frost hardiness most conservatively, while the NB ecotype from the nearby latitude (63° N), but from a much higher altitude, dehardened most rapidly. Spring 2001 The elevated winter temperature resulted in reduced frost hardiness at P < 0.01 in January (Fig. 4a), but there were no differences between the ecotypes. Two week later, at the beginning of February, the effect of elevated winter temperature remained significant (P < 0.01) and the effect of seed origin had become visible (P < 0.01) (Fig. 4b). At this stage, the ecotypes NB, FJ and NHa appeared to have dehardened most rapidly. At the end of March (week 26), the effect of winter temperature was most marked (P < 0.001). In addition, there was a significant seed origin–winter temperature interaction (P < 0.01) as the ecotypes IC, NMe and FJ displayed a high level of frost hardiness at ambient temperature (i.e. LT50 > −60°C), while the frost hardiness of each ecotype had gone up to −20°C or more at the elevated temperature (Fig. 4c). The effect of seed origin was also greatest at this time, showing that the ecotypes NB (i.e. the southernmost ecotype from high altitude) and NHa (i.e. the northernmost ecotype near the sea level) had dehardened most rapidly. Figure 4Open in figure viewerPowerPoint Frost hardiness (mean LT50, °C below zero ± se, n = 4) of the mountain birch (Betula pubescens ssp. czerepanovii) ecotypes during the spring of 2001: (a) January (b) February and (c) March. The different small letters indicate the statistical difference between the seed origins (ecotypes) at P < 0.05. The abbreviated seed origins are as in Table 1. Lipid peroxidation measured as malondialdehyde content was greatest in Jan (Fig. 5). It differed from the content assessed at the two latter dates at P < 0.05. Interestingly, though no statistical difference emerged, lipid peroxidation tended to be less pronounced in populations with the highest degree of frost hardiness (IC ≤ FJ ≤ NB ≤ NMe ≤ NHa). Figure 5Open in figure viewerPowerPoint Lipid peroxidation (mean ± se, n = 4) of mountain birch (Betula pubescens ssp. czerepanovii) ecotypes during the spring of 2001: (a) January (b) February and (c) March. The abbreviated seed origins are as in Table 1. Spring 2002 Frost hardiness was unaffected by either the winter temperature or the seed origin in January (Fig. 6a). The elevated winter temperature appeared to have decreased frost hardiness only in April (week 15) (Fig. 6c). In addition, seed origin did not affect frost hardiness except marginally (P < 0.1) in January. As a whole, however, the southernmost population from the high altitude (NB) dehardened most rapidly, in accordance with the dehardening in 2001. The greatest difference between the years due to seed origin was seen in the response of the Iceland population: these seedlings were the most conservative in 2001 and nonconservative in 2002 in maintaining their high level of frost hardiness. Figure 6Open in figure viewerPowerPoint Frost hardiness (mean LT50, °C below zero ± se, n = 4) of the mountain birch (Betula pubescens ssp. czerepanovii) ecotypes during the spring of 2002: (a) January (b) February and (c) April. Abbreviated seed origins are as in Table 1. Lipid peroxidation content followed a similar pattern as in the spring of 2001. The only statistically significant response was found in response to sampling date (Fig. 7). As in 2001, lipid peroxidation decreased from January (P < 0.05). In addition, a slight recovery in lipid peroxidation, as in 2001, was also observed in February and April. Moreover, the rate of lipid peroxidation was lowest in the most frost-hardy populations (NHa ≤ NMe ≤ NB ≤ FJ ≤ IC), as in the previous year. Figure 7Open in figure viewerPowerPoint Lipid peroxidation (mean ± se, n = 4) of the mountain birch (Betula pubescens ssp. czerepanovii) ecotypes during the spring of 2002: (a) January (b) February and (c) April. The abbreviated seed origins are as in Table 1. Normalized hardiness Normalization of the frost hardiness data (Table 3) resulted in the following pattern of responses to a temperature elevation of +4°C. In the winter of 2000–2001, the frost hardiness order of the ecotypes (from the most dehardened to hardened) was as shown in Order 5. In the winter of 2001–2002, the order was almost the same (Order 6). The result indicates that the northernmost ecotypes were most tolerant and the southernmost ecotypes least tolerant against a +4°C elevation in winter temperatures compared to the temperature they were adapted to: Table 3. Temperature elevations experienced by the mountain birch (Betula pubescens ssp. czerepanovii) ecotypes in the experiment and the respective normalized hardiness corresponding to +4°C temperature elevations of a given ecotype Ecotype T elevation (±°C) Normalized hardiness (–°C) Elevated Ambient 2000–2001 NB 9.8 6.4 14 IC 5.5 2.1 46 NMe 4.9 1.5 32 FJ 17.4 14 < 100 NHa 9.7 6.3 55 2001–2002 NB 11.2 7.8 1 IC 6.9 3.5 12 NMe 7.5 4.1 18 FJ 18.8 15.4 43 NHa 11.3 7.7 13 Order 5 NB < Nme < IC < NHa < FJ Order 6 NB < IC; NHa < Nme < FJ Discussion The present study demonstrates that a 4°C rise in winter temperature may significantly accelerate the dehardening process of mountain birch in spring. In accordance, accelerated dehardening by rising winter temperature was also reported in Scots pine (Repo et al., 1996) and bilberry (Taulavuori et al., 1997a). Elevated winter temperatures may result in either premature or delayed bud break (e.g. Murray et al., 1989). The former is due to accumulation of day degrees after the fulfilled chilling requirement (Hänninen, 1995), while the latter is due to incomplete fulfilment of the chilling requirement at excessively high winter temperatures (Murray et al., 1989). Such a chilling deficit and subsequent delay in bud break is unlikely in Scandinavia, and the likely effects of climatic warming include earlier bud burst, a longer growing season and an in