The electrical impedance spectroscopy of Scots pine (Pinus sylvestris L.) shoots in relation to cold acclimation

Electrical impedance spectroscopy (EIS) was applied to stems of Scots pine (Pinus sylvestris L.) in a provenance field trial during frost hardening to find an EIS parameter for assessing frost hardiness (FH) without a controlled freezing test. The FH of stems and needles assessed by controlled freezing tests was compared with the equivalent circuit EIS parameters of a distributed model of stems (not exposed to controlled freezing treatment) and with dry matter (DM) content of stems. Significant differences in the equivalent circuit parameters, FH and DM content were found between provenances. The relaxation time (tau(1)), describing the peak of the high frequency arc of the impedance spectrum, and the intracellular resistance (r(i)) of stems increased with increasing FH. According to the linear regression, the coefficient of determination (R(2)) between the FH of stems and needles with tau(1) of the stem was 0.87 and 0.89, and with r(i) of the stem 0.74 and 0.85, respectively. The relation between FH and tau(1) changed with the degree of hardiness. The highest coefficient of determination was 0.95 in September when the FH of needles, ranging from -10 degrees C to -25 degrees C, was predicted with an accuracy of +/-2.0 degrees C. The resistance parameter r(2), describing the width of the low frequency arc of the impedance spectrum, decreased prior to and during the initial hardening: significant differences were found between provenances. This indicates that r(2) was not related to frost hardening per se. It is concluded that it is possible to distinguish the hardening patterns of different provenances by tau(1) in the rapid phase of hardening without controlled freezing tests.     

水杨酸对大叶黄杨茎抗寒性和电阻抗图谱参数的影响

在抗寒锻炼前,对当年生大叶黄杨(Euonymus japonicus)扦插苗喷施不同浓度水杨酸,用电阻抗图谱(EIS)法和电导(EL)法估测茎的抗寒性,以探明抗寒锻炼期间水杨酸对大叶黄杨抗寒性和电阻抗图谱参数的影响,找到适合不经冷冻处理估测大叶黄杨茎抗寒性和经冷冻处理后测定其抗寒性的EIS参数。结果表明,水杨酸处理能够提高大叶黄杨茎的抗寒性,最适浓度为5.0mmol.L-1;不经冷冻处理茎的EIS参数电阻率r和r1、胞外电阻率re、胞内电阻率ri、弛豫时间τ、弛豫时间分布系数ψ与EL法测定的抗寒性有较高的相关性(r=0.70~0.87),说明不经冷冻处理样本用以上参数估测大叶黄杨茎抗寒性是可行的,r1为最佳参数;冷冻处理后茎的re、τ、ri求得的抗寒性与EL法测定的抗寒性有较高的相关性(r=0.85~0.94),说明冷冻处理后re、τ、ri可以作为测定大叶黄杨茎抗寒性的参数,re为最佳参数。     

The relation between growth cessation and frost hardening in Scots pines of different origins

The cessation of shoot elongation, diameter growth and needle elongation were compared with the initiation of frost hardening of the stems and needles in an 8-year-old provenance trial of Scots pine (Pinus sylvestris L.) established in central Finland. The saplings were of six different origins ranging from Estonia to northern Finland, forming a latitudinal gradient of ca. 10°N. The frost hardiness of the stems of current-year shoots was assessed by electrical impedance analysis and that of current-year needles by electrolyte leakage and visual scoring of damage. Artificial freezing tests were used in the assessments. The pattern of growth cessation (shoot and needle elongation, diameter growth) tended to follow the latitude of origin, i.e. growth ceased in the northernmost provenance first and in the southernmost one last. Both stems and needles of the northern provenances hardened earlier than the southern ones, but the differences in hardiness disappeared as hardening progressed. Growth cessation and initial hardening to -15°C were clearly correlated at the provenance level, indicating that growth must cease prior to hardening, and that earlier cessation of growth predicts earlier frost hardening of stems and needles. No differences in frost hardiness of stems were found at the provenance level at the end of the growing period in August. At that time, the frost hardiness of needles of the northernmost provenance was higher than that of other origins. Within the provenance, the stems were less hardy than the needles.     

CaCl2对金叶女贞茎抗寒性和电阻抗图谱参数的影响

为探明CaCl₂处理对金叶女贞(Ligustrum vicaryi)茎抗寒性和电阻抗图谱参数的影响,找出抗寒性和电阻抗图谱参数的关系,2006年9月15日开始对金叶女贞当年生扦插苗进行CaCl₂处理,对照为清水;分别用电阻抗图谱法和电导法估测其抗寒性,并进行比较分析。结果表明：CaCl₂处理能在抗寒锻炼初期使金叶女贞茎的抗寒性提高3.5℃,金叶女贞茎的抗寒性与未冷冻处理茎的弛豫时间τ、冷冻处理后茎的胞外电阻率r_e、含水量之间均存在较高的相关性。     

测定植物抗寒性的电阻抗图谱法

电阻抗图谱(electrical impedance spectroscopy,EIS)分析作为测定植物抗寒性的一种方法,在农业、林业和园艺领域的应用正在不断扩大。该文从EIS的原理入手,讨论了影响电阻抗特性的生理和物理因子;介绍了测定EIS适用的模型;阐述了用EIS测定抗寒性的方法。在EIS分析中,胞外电阻率(re)是确定抗寒性最适用的一个参数,弛豫时间(τ1)是精确度最高的参数。     

电导法和电阻抗图谱法测定脱锻炼期间欧美杨×大青杨F_1子代抗寒性

利用电导（EL）法和电阻抗图谱（EIS）法对脱锻炼期间欧美杨‘107杨’（Populus×canadensis Moench cv.‘Neva’）×大青杨（P.ussuriensis Kom.）F1子代进行抗寒性测定,试图通过比较两种方法测定抗寒性结果的相关性,以确定EIS法快速测定‘107杨’×大青杨F1子代抗寒性的最佳参数。以8个杂种子代和母本‘107杨’3年生为试材,进行了电阻抗图谱法和电导法分析,结果表明：脱锻炼期间,各子代抗寒性均逐渐降低,依次为NK36〉NK41〉NK61〉NK57〉NK42〉NK60〉NK58〉NK56,均高于‘107杨’,EIS参数弛豫时间[EIS（τ）法]和EL法的抗寒性测定结果间存在明显的线性相关（R2=0.965）,但EIS法测定的抗寒性低于EL法,且未经冷冻处理样本的胞外电阻率（re）和弛豫时间（τ）与经冷冻处理EL测定的抗寒性有较高的相关性,R2分别为0.817和0.847。     

Intracellular resistance correlates with initial stage of frost hardening in willow (Salix viminalis)

Distributed model parameters of shoots of five clones of willow ( Salix viminalis ) were examined with electrical impedance analysis at the end of the growing season and with cold acclimation. The parameters were compared with regard to frost hardiness, linoleic (18:2) and linolenic (18:3) fatty acids, unsaturation/saturation ratio of fatty acids and dry weight content. The intracellular resistance (r_i) correlated best with changes in frost hardiness. The value of r_i rose from 1–2 Ω m in non-hardened to about 12 Ω m in hardened samples. In the initial stages of frost hardening, a linear relationship was found between r_i and frost hardiness and levels of 18:2 fatty acid, and an inverse relationship between r_i and levels of 18:3 fatty acid. The unsaturation/saturation ratio of fatty acids rose fairly rapidly in the initial stage of hardening. The dry weight content increased stepwise during the experimental period, and less steadily than r_i. In addition, equivalent circuit parameters changed in the prehardening phase, and were probably connected with cell differentiation and lignification. Frost hardiness by the visual method and by extracellular resistance, determined after controlled freezing tests, correlated well in the initial stages of hardening until about − 10°C but deviated upon further hardening.     

Effect of photoperiod and temperature on the development of frost hardiness in three Alnus species

The influence of short day and low temperature on cold acclimation of A. crispa (Ait.) Pursh, A. glutinosa (L.) Gaertn. and A. rubra Bong, was investigated. Two clones of each species originating from in vitro propagation were exposed to three daylength/temperature treatments. Periodically plantlets were exposed to controlled freezing temperature in order to evaluate their level of frost hardiness.
Short day (SD) and cold temperature (CT) and long day (LD) and cold temperature (CT) were the most effective treatments for the development of frost hardiness in shoots and roots of the three species tested. Short day (SD) and warm temperature (WT) induced a significant increase in hardiness in shoots of all three species. However, this treatment did not trigger root hardening. A. crispa was found to be the hardiest species followed by A. glutinosa and A. rubra . Intraspecific variation was observed between the two A. glutinosa clones. A glutinosa clone AG8, a Russian provenance, showed a greater freezing resistance than A. glutinosa clone AG2, a German provenance.     

The consequences of freezing temperatures followed by high irradiance on in vivo chlorophyll fluorescence and growth in Picea abies

Picea abies (L.) Karst. plants, propagated by cuttings, were subjected to one night of freezing temperatures (-5°C), high irradiance (1 200 or 1 800 μmol m⁻² s⁻¹), or freezing temperatures followed by high irradiance. The treatments were applied at bud burst, at time of shoot elongation, and when the shoots had ceased to elongate. The maximum quantum yield of photosynthesis, F_v/F_m, dry weight of branches and needles, and length and survival of shoots were measured. F_v/F_m and growth decreased after a night of freezing temperatures followed by high irradiance, at the time of bud burst and shoot elongation. High irradiance alone influenced F_v/F_m, but not growth. Freezing temperatures affected F_v/F_m, and growth at the time of shoot elongation. F₀ increased after a night of freezing temperatures and decreased with age of the current-year needles. It was concluded that the use of short-term measurements of chlorophyll fluorescence induction to predict changes in growth after a night of frost and subsequent high light was not a reliable method.     

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.     

The electrical impedance spectroscopy of Scots pine needles during cold acclimation

The electrical impedance spectroscopy (EIS) was applied to current-year needles of Scots pine ( Pinus sylvestris L.) in an 8-year provenance field trial in central Finland during frost hardening. The EIS analysis of the needles using a Model-A equivalent circuit indicated a sequence of events in the needles during their cold acclimation. Some of the EIS-parameters referred to maturation phenomena occurring during the pre-hardening phase at the end of the growing season, and some parameters displayed a clear coincidence with the frost hardening itself. Significant differences between provenances were found in several of the Model-A parameters. Extracellular resistance (r_e) and β -coefficient decreased in all provenances in the pre-hardening phase in August and until mid-September. In the same phase, both the intracellular resistance (r_i) and the cell membrane time constant (τ_m) first increased and then decreased. According to τ_m, r_e and β there was a clear gradation between provenances in the pre-hardening phase. From the end of September significant differences were found in the intracellular resistance between provenances, corresponding with the differences in their hardening pattern. The dry weight (DW) content of needles increased during the study period but no clear differences were found between the provenances.     

Effects of previous defoliation on pine looper larval performance

1 During outbreaks of the pine looper, Bupalus piniarius, its host, Pinus sylvestris, is severely defoliated. The larvae of this geometrid normally feed on mature needles. However, because trees are totally defoliated during outbreaks, the next generation is forced to feed on current-year needles. 2 Bupalus piniarius larvae feeding on previously defoliated trees may show lower performance either because of induced resistance or because larvae have to feed on needles not normally fed upon (current instead of mature). 3 These hypotheses were tested in an experiment where larvae were reared on (i) shoots naturally defoliated the previous year, and thus, bearing only current-year needles, (ii) non-defoliated shoots where larvae had access only to current-year needles, and (iii) control shoots with access to both current and mature needles. 4 There was no support for the induction hypothesis. Survival was lower on naturally defoliated shoots than on control shoots (81.3 vs. 90.9%), but survival was lower also on non-defoliated shoots where larvae had access only to current-year needles (78.8%). Data on larval feeding distribution showed a strong preference for mature needles. 5 Needle nitrogen concentration of current-year needles was 38% higher on defoliated trees than on non-defoliated trees. 6 It is concluded that defoliation affected larval performance primarily through the removal of the preferred type of needles and not because of an induced resistance. Effects of increased concentrations of allelochemicals in defoliated shoots, if present, were probably cancelled out by increased nitrogen concentrations.     

Effect of defoliation on tracheid production and the level of indole-3-acetic acid in Abies balsamea shoots

To simulate feeding by the spruce budworm ( Choristoneura fumiferana Clem.), the apical current-year shoots on 1-year-old branches in the uppermost whorl of 6-year-old balsam fir [ Abies balsamea (L.) Mill.] trees were either removed completely by debudding before the start of the growing season or defoliated 0, 50, 90 or 100% shortly after budbreak. Debudded branches were treated at the apical end with 0, 0.1 or 1.0 mg of indole-3-acetic acid (IAA) (g lanolin)⁻¹. Ninety % of the 1-year-old needles were also removed from some of the experimental branches. After ca 4 weeks of growth, the radial width of new xylem and the level of IAA were determined in the 1-year-old internode. The IAA content was measured by radioimmunoassay.
The removal or defoliation of current-year shoots inhibited tracheid production and decreased the IAA level. Exogenous IAA stimulated tracheid production and increased the IAA level in debudded branches. Current-year shoot defoliation also inhibited current-year shoot elongation. The inhibitory effect of current-year needle removal on all parameters generally increased with increasing intensity of defoliation. The removal of 1-year-old needles did not affect the IAA level or current-year shoot elongation, nor did it influence tracheid production in branches with current-year shoots. However, removal of 1-year-old needles inhibited tracheid production in debudded branches supplied with exogenous IAA. The results indicate that (1) IAA is involved in the control of tracheid production in the 1-year-old internode, (2) IAA is supplied primarily by current-year shoots, and (3) defoliation by the spruce budworm inhibits tracheid production partly by decreasing the supply of IAA.     

The Influence of Photoperiod and Temperature on the Development of Frost Hardiness in Seedlings of Pinus silvestris and Picea abies

The influence of short days and low temperature on the development of frost hardiness in seedlings of Scots pine (Pinus silvestris L.) and Norway spruce [Picea abies (L.) Karst.], grown for 6 months in glasshouses and climate chambers, was investigated. The degree of hardiness was estimated by freezing the shoots of the seedlings to predetermined temperatures. After 8 weeks in a glasshouse the viability of the seedlings was determined by establishing bud flushing. The most effective climate for the development of frost hardiness was short days (SD) and low temperature (2°C); the next most effective was SD and room temperature (20°C). However, long days (LD) and low temperature also had a marked effect on the development of hardiness. A combination of 3 weeks’treatment with SD and 20°C, and 3 weeks with SD and 2°C gave the same results as 6 weeks with SD and 2°C. The results clearly demonstrate the importance of the photoperiod prior to low temperature for the development of frost hardiness. In conclusion both short days and low temperature induce frost hardiness development. Probably this occurs by initiation of different processes in the two cases. The degree of frost hardiness development appears to depend on the sum of these different processes and on the timing between them.     

Dehydration strain avoidance and tolerance in plant cold hardiness

Cell strain during freezing is typically characterized by either a measure of cell volume contraction or a measure of the fraction of cell water frozen. If the measure of strain at the killing temperature increases during cold acclimation, it is concluded that hardiness is controlled by strain tolerance, and if the measure of strain does not increase during cold acclimation it is concluded that hardiness is controlled by strain avoidance. It is demonstrated that the measure of cell volume contraction is not mathematically equivalent to the measure of the fraction of water frozen. Thus the conclusion regarding the relative role of avoidance and tolerance in frost hardiness depends upon the choice of strain measurement. The parameters determining the two measures of strain indicate that if cell injury is related to membrane area contraction or expansion then the measure of cell volume contraction is likely to be the better measure of strain; however, if cell injury is related to solution effects, the measure of the fraction of water frozen is likely to be the better measure of strain. In addition, the temperature dependence of each strain measure is derived to aid in the visualization of the freezing process, and to accommodate calculation of the measures of strain from data obtained at above freezing temperatures.     

The effects of long-term elevation of air temperature and CO on the frost hardiness of Scots pine

The frost hardiness of 20 to 25-year-old Scots pine (Pinus sylvestris L.) saplings was followed for 2 years in an experiment that attempted to simulate the predicted climatic conditions of the future, i.e. increased atmospheric CO₂ concentration and/or elevated air temperature. Frost hardiness was determined by an electrolyte leakage method and visual damage scoring on needles. Elevated temperatures caused needles to harden later and deharden earlier than the controls. In the first year, elevated CO₂ enhanced hardening at elevated temperatures, but this effect disappeared the next year. Dehardening was hastened by elevating CO₂ in both springs. The frost hardiness was high (相似文献   

The effect of soil temperature on the bud phenology, chlorophyll fluorescence, carbohydrate content and cold hardiness of Norway spruce seedlings

The effects of soil temperature on the shoot phenology, carbohydrate dynamics, chlorophyll fluorescence and cold hardiness of 4-year-old Norway spruce seedlings ( Picea abies L. Karst.) were studied. The experiment was carried out under controlled conditions in the Joensuu dasotrons. Air conditions were similar but soil temperatures differed by treatments (9, 13, 18 and 21°C) during the second growing period in the dasotrons. The after-effects of the treatments were investigated during the third growing period following the treatments. Low soil temperature increased the starch content of needles and delayed the loss of starch at the end of the growing season. The photochemical efficiency ( F _v/ F _m) of the PSII of the current-year needles was reduced at the lowest soil temperature. The cold hardiness of needles correlated with the soluble sugar content. The differences in soil temperature had no effect on the timing of bud burst. No after-effects from the treatments were observed during the third growing period in the dasotrons.     

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.     

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     

Application of Impedance Spectroscopy for Selecting Frost Hardy Varieties of English Ryegrass

The frost hardening and frost damage of 12 varieties of Englishryegrass (Lolium perenne) was studied by electrical impedancespectroscopy. For the measurement of the impedance spectrum(80 Hz to 1 MHz) a 10 mm length sample was cut from the stemabove the growing point, but the growing point was included.The impedance spectra were analysed by an asymmetric distributedcircuit model. The impedance spectra were measured at two phasesof hardiness and after freezing, i.e. (a) before hardening,(b) after hardening in controlled conditions, and (c) aftercontrolled frost exposure at -16 °C after hardening. Twomodel parameters, i.e. intra- and extracellular resistance,increased with hardening. The intracellular resistance and theskewness factor before hardening, and the ratio between thosetwo parameters before and after hardening, strongly correlatedwith hardening of different varieties of English ryegrass. Theextracellular resistance and the relaxation time decreased asa result of the frost exposure at -16 °C. Cold acclimation; electrical impedance; English ryegrass; frost hardiness; impedance spectroscopy; Lolium perenne