- Top of page
- Materials and Methods
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|
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 1. 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).
Download figure to PowerPoint
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
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.
- Top of page
- Materials and Methods
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 increased risk of spring frost injury (Myking & Heide, 1995). It has to be emphasized that the chilling requirement was not measured during the present study. However, there is evidence that two basic mechanisms control frost hardiness: long-term control through the annual ontogenetic cycle (Fuchigami et al., 1982; Leinonen, 1996a) and short-term control through direct environmental stimuli (e.g. Sakai & Larcher, 1987). Because of the long-term control of frost hardiness, it is understandable that the present results are in line with the hypotheses based on the chilling requirement.
A superficial evaluation of the ecotype response to dehardening (Order 4) under the investigated conditions closely follows the pattern presented as Hypothesis 1. The only exception is the ecotype FJ, which was not so susceptible to dehardening as was expected (Orders 1 and 2). However, the IC ecotype maintained its frost hardiness most effectively, as was expected. In turn, the most alpine ecotype NB had the highest rate of dehardening, in accordance with Hypothesis 1. The highest dehardening rate in the NB ecotype is in contradiction with Hypothesis 3. However, this does not exclude that daylength could affect the dehardening. Indeed, the dehardening order (Order 4) of the other ecotypes was as expected according to Hypothesis 3. The result is thus consistent with the stabilized opinion that rising temperature is the dominating factor for dehardening (e.g. Fuchigami et al., 1982), although it suggests that the photoperiod may also slightly modify the process (e.g. Linkosalo et al., 2000).
Because of the ontogenetic control of frost hardiness, the autumn photoperiod may have also affected the obtained result, since the northernmost ecotypes in the photoperiod of Bergen probably first entered into dormancy as a consequence of their longer critical daylength for the induction of dormancy (Cannell, 1990; Myking & Heide, 1995). Consequently, accumulation of chilling temperature and subsequent fulfilment of the chilling requirement started earlier in the northern ecotypes. This explains the significant interaction between the hardening temperature and ecotype in 2001: the northernmost ecotypes (NHa and FJ) dehardened most rapidly in the treatment with low hardening temperature (+9°C) and elevated winter temperature. In other words, the photoperiod of Bergen triggered soon the development of dormancy in the northernmost ecotypes, which started to accumulate chilling temperature. Once the chilling requirement had been met, the effect of elevated winter temperature became obvious. The ecotype NB made an exception from this pattern since it dehardened most effectively, probably as a consequence of the alpine climate it had adapted to, in accordance with Hypothesis 1.
The result reflects the response of each ecotype to given experimental manipulation, but does not provide information for evaluating the consequences of global climate warming. However, long-term temperature data from areas of the studied ecotypes are of great help in this evaluation. The temperature values shown in Table 1 illustrate well the wintertime climatic conditions to which the ecotypes have been adapted. Although the data concern only the period between 1931 and 1960, and do not extend to the present, it is nevertheless valid for two reasons. First, the decades 1930–1940 were warm in Scandinavia, followed by cooler periods until the 1980s, and since the 1990s warming had started again (Kullman, 2001). Cooler and warmer periods undoubtedly equalize the temperature differences of the whole period from 1930 to 2000, and provide a good, though approximate basis for considering environmental adaptation. Secondly, the ecotypes result from genetic adaptation, where 40 year (i.e. the lack of data from the 1960s to the 2000 s) is a short period, especially because of the longevity of trees. When these temperature parameters are compared to temperatures recorded during the experiment (Fig. 3), it is obvious that the ecotypes experienced the elevated temperatures unequally (Table 3). The ecotypes IC and NMe, in the elevated temperature conditions of Bergen, overwintered at temperatures of 4–7°C above the temperature where they have been adapted. The temperature elevation was exactly the same as predicted by the climatic change scenarios for the boreal zone in the near future (Bach, 1988; Maxwell, 1992; Murray, 1995; Houghton et al., 2001). The ambient temperature in Bergen as such provided a temperature elevation within the same range for the ecotypes NB and NHa, while the elevated temperature in Bergen provided conditions warmer than could be expected within the near future. However, the ecotype FJ overwintered at an unrealistic temperature elevation irrespective of the temperature treatment in Bergen.
The normalized hardiness (Fig. 2, Table 3) provides a tool for relating the present results to climatic warming. The ecotype NB seems most susceptible to climatic warming by a +4°C temperature elevation, while the northernmost ecotypes NHa and FJ were relatively well buffered against it, especially the FJ exhibiting the ecotype from the coldest climate within this study (Table 2). This is obviously due to the adaptation of this ecotype to northern and relatively continental conditions at a relatively high altitude, which altogether provide a wide range of annual temperature changes. The significance of the +4°C elevation is thus relatively smaller for an ecotype adapted to a harsh rather than a mild climate. It should be emphasized, however, that climatic warming is most intensive in the northernmost areas (Bach, 1988; Maxwell, 1992; Murray, 1995; Houghton et al., 2001), i.e. equalizing somewhat the magnitude of its effects between the ecotypes.
During the first year of the experiment (i.e. 2000) the growth period was a month longer than it was the next year. Cool periods during the growing season, i.e. unfavourable growing conditions, may disturb development and ripening of the overwintering tissues of Japanese mountain birch (Betula ermanii) (Gansert, 2002). Consistently, the frost hardiness of each studied ecotype was poor (Fig. 6) in January 2002, obviously due to unfavourable growing conditions, although the winter was also approx. 1.5°C milder (Fig. 3). However, the latter cannot explain the low degree of frost hardiness in 2002, since frost hardiness in elevated temperature treatment (approx. +5.5°C) in 2001 was greater than in ambient conditions in 2002 (approx. +3.5°C), for example.
Seasonal changes in lipid peroxidation patterns were high during the winter and tended to decrease in the spring (Cakmak et al., 1995). Consistently, our data show the highest rate of lipid peroxidation in January in both years (Figs 5a and 7a). The purpose of the hardened state, however, is to stabilize and alleviate the increase in lipid peroxidation caused by freezing stress conditions (Wang et al., 1995; Yang et al., 2002). Therefore, a low degree of springtime lipid peroxidation indicates a high degree of frost hardiness in relation to temperature experienced, i.e. low stress. It is thus understandable that the ecotypes IC and FJ with maximum frost hardiness (Fig. 4c) also exhibited the lowest level of lipid peroxidation in 2001 (Fig. 5c). In addition, NHa, being one of the most dehardened ecotypes, showed the highest degree of lipid peroxidation. In accordance with high degree of lipid peroxidation in NHa in March 2001, freezing stress may peak in spring, which is able to induce a temporary increase in lipid peroxidation (Polle et al., 1996; Zhou & Leul, 1998; Benson et al., 1999). On the other hand, in 2002, the ecotype IC belonged to the rapidly dehardened population and showed the highest degree of lipid peroxidation. Lipid peroxidation analysis thus generally supports the dehardening findings based on the determinations of hardiness.
To sum up, an interpretation of the results based on statistical tests shows that the different mountain birch ecotypes dehardened in line with Hypothesis 1 and the presumed chilling requirement. The most alpine and southern ecotype (NB) exhibited the greatest, and the other southern ecotype (IC) from the mild climate the lowest, rate in the dehardening process. However, projection of the results on the difference between the adapted and experienced temperatures indicates that the risk of frost injury is lowest in the northernmost ecotypes under a temperature elevation of +4°C. This indicates adaptability of these ecotypes to a wider range of temperatures, including the absolute minima. Moreover, the most alpine ecotype (NB) was among the populations with the most probable risk of frost injury, regardless of which of the above perspectives was used to interpret the results. The results on all the ecotypes in 2002 demonstrate that a low level of frost hardiness and a respective high risk of frost damage may occur after an unfavourable growing season.