Relationships among cold hardiness, root growth potential and bud dormancy in three conifers

Greenhouse-cultured, container-grown ponderosa pine (Pinus ponderosa var. scopulorum Engelm.), interior Douglas-fir (Pseudotsuga menziesii var. glauca (Beissn.) Franco) and Engelmann spruce (Picea engelmannii (Parry) Engelm.) were cold acclimated and deacclimated in growth chambers over 19 weeks. Stem cold hardiness, total new root length at 14 days and days to bud break were measured weekly. Relationships among cold hardiness, root growth potential (RGP) and bud dormancy suggest that cold hardiness, which can be measured quickly, could provide a useful basis for estimating the two other parameters. During cold acclimation, there was a lag period in which stem cold hardiness remained at -15 degrees C and RGP was at a minimum, in all three species. Douglas-fir and Engelmann spruce buds remained fully dormant during this lag period. Ponderosa pine buds had no chilling requirement for the loss of dormancy, and reached quiescence during the lag period. Immediately following the lag period, as stem cold hardiness progressed to -22 degrees C, RGP increased to a high plateau in all three species, and Douglas-fir and Engelmann spruce buds approached quiescence. Cold deacclimation and bud development began immediately on exposure to warm, long days, but RGP remained high until stem cold hardiness returned to approximately -15 degrees C. At bud break, cold hardiness and RGP were at the minimum.     

Seasonal variations in soluble sugars and starch within woody stems of Cornus sericea L

Carbohydrate composition changed seasonally in red osier dogwood (Cornus sericea L.) stem tissues. Starch concentration was highest in fall and decreased to a minimum in midwinter. Coincident with the breakdown of starch in fall, there was an increase in the concentrations of soluble sugars. Soluble sugars were present in highest concentrations in midwinter. Glucose, fructose, sucrose, and raffinose were the predominant soluble sugars present in both bark and wood tissues. In early spring, the soluble sugar concentration decreased and the concentration of starch increased. The seasonal interchange between sugars and starch did not simply reflect a general quantitative shift in the balance between sugars and starch because qualitative changes in soluble sugars were also noted. The most striking changes involved the trisaccharide raffinose. Raffinose was barely detectable in summer and early fall, but increased to one fifth and one third of the total soluble sugars in January samples of bark and wood tissues, respectively. The potential physiological role of raffinose in overwintering red osier dogwood tissue is discussed.     

The dependance of root growth potential on light level,photosynthetic rate,and root starch content in jack pine seedlings

Number of new roots (root growth potential or RGP), new root length, photosynthesis, total nonstructural carbohydrate content of needles and roots, terminal bud condition, and shoot elongation were measured on jack pine container seedlings for 4 weeks at weekly intervals under greenhouse conditions of 100%, 20%, and 10% sunlight to simulate competition-induced, lower light levels in the field. Both lower light levels significantly reduced photosynthetic rate, RGP, new root length, total nonstructural carbohydrate (especially starch) content of needles and roots, speed of terminal bud flush, and shoot growth. Both light level and photosynthetic rate were positively correlated with RGP and new root length, indicating that jack pine seedlings may use current photosynthate as an energy source to support new root growth. RGP and new root length were also both negatively correlated with root starch content suggesting that jack pine seedlings may also use stored carbohydrates as a potential carbon source for root initiation and initial root growth.     

Frost hardening of Pinus radiata seedlings: effects of temperature on relative growth rate,carbon balance and carbohydrate concentration

Pinus radiata (D. Don) seedlings were grown for 100 days at day/night temperatures of 10/1, 15/1, 20/1 and 25/1 degrees C, to determine whether temperatures above a threshold of 5 degrees C influence frost hardiness development. Relationships between hardening and relative growth rate, carbohydrate concentration and net carbon balance were also investigated. Seedlings hardened at a nearly constant rate in each treatment, although the rate of hardening was strongly temperature dependent. It increased as the temperature declined, but in a curvilinear fashion. Temperatures below 9.5 degrees C were effective in hardening the seedlings. During the daily temperature cycle, dehardening occurred at temperatures above the threshold, whereas hardening occurred at temperatures below the threshold. The net difference between the two processes determined the development of frost hardiness. The development of frost hardiness was negatively correlated with relative growth rate and positively correlated with the accumulation of starch and sugars. We conclude that frost hardening is a complex process that is causally linked to carbohydrate concentrations.     

Assessment of seedling storability of Quercus robur and Pinus sylvestris

Pedunculate oak (Quercus robur L.) and Scots pine (Pinus sylvestris L.) seedlings were lifted on several occasions during autumn 1997 to determine the relationships between storability and frost hardiness. On each lifting date their physiological status was determined by assessment of shoot and root electrolyte leakage and frost hardiness, assessed as freeze-induced electrolyte leakage. Additional seedlings were simultaneously cold-stored for field planting and assessment of preplanting root growth potential in April 1998. First year field performance was determined the following winter. Storability and cold acclimation patterns differed between the two species. Both were negatively affected by early lifting, but oak was less sensitive with respect to survival, and pine attained tolerance to cold storage more rapidly and earlier with respect to growth increment. The correlations between shoot frost hardiness and performance suggest that freeze-induced shoot electrolyte leakage (SEL_diff?20) below a threshold of 5% is a good storability predictor for Scots pine in Denmark. A completely reliable criterion for pedunculate oak could not be established.     

Fall fertilization of Pinus resinosa seedlings: nutrient uptake, cold hardiness, and morphological development

? Fall fertilization may increase plant nutrient reserves, yet associated impacts on seedling cold hardiness are relatively unexplored.

? Bareroot red pine (Pinus resinosa Ait.) seedlings in north-central Minnesota, USA were fall fertilized at the end of the first growing season with ammonium nitrate (NH₄NO₃) at 0, 11, 22, 44, or 89 kg N ha^?1. Seedling morphology and cold hardiness [assessed by freeze induced electrolyte leakage (FIEL)] were evaluated six weeks after fertilization and following the second growing season.

? Seedling height and number of needle primordia increased with fertilizer rate for both sampling years. Seedlings fertilized with 44 and 89 kg N ha^?1 attained target height (15 cm) after the second growing season. Shoot and root N concentration increased after the first growing season in fall fertilized seedlings compared to controls. Fall fertilized seedlings had lower FIEL (i.e., increased cold hardiness) compared to controls when tested at ?40 °C after the first growing season, but no significant differences in FIEL of control and fertilized seedlings were observed after the second growing season.

? Results suggest that fall fertilization of red pine seedlings can help render desired target height in the nursery, while maintaining or increasing cold hardiness levels.

     

Cold hardiness of interspecific hybrids between Pinus strobus and P. wallichiana measured by post-freezing needle electrolyte leakage

Interspecific hybrids between eastern white pine (Pinus strobus L.) and Himalayan blue pine (P. wallichiana A. B. Jacks.) were developed in Ontario, Canada, to introduce blister rust (Cronartium ribicola Fisch.) resistance genes to P. strobus. There is concern that introducing blister rust resistance has resulted in reduced cold hardiness of the progeny compared with non-hybridized eastern white pine. To test the efficacy of backcrossing with P. strobus to improve cold hardiness, 1-year-old seedlings from hybrid crosses differing in P. strobus genome composition were artificially freeze-tested. In Experiment 1, unhardened seedlings were allowed to acclimate to progressively lower temperatures in a growth room, whereas in Experiment 2, seedlings were hardened outdoors under natural weather conditions in Sault Ste Marie, Ontario. Needle cold injury was determined by calculating relative electrical conductivity based on post-freezing electrolyte leakage. Results indicated that needle fascicles from unhardened seedlings of all genotypes in the greenhouse tolerated -5 degrees C for 3 hours with little or no injury. Cold hardiness increased in parallel with declining growth room minimum temperature over the 7-week period of hardening. Cold hardiness was improved for hybrid crosses with increased Pinus strobus genome composition in Experiment 2, but the results were less conclusive in Experiment 1.     

Relationship between temperature, respiratory loss of sugar and premature dehardening in dormant Scots pine seedlings

Increased intracellular sugar concentration is an important contributor to the increased cold tolerance of conifers in winter. This study examines the extent to which wintertime respiratory loss of sugars leads to premature dehardening. Two-year-old seedlings of Scots pine (Pinus sylvestris L.), grown and cold-hardened in the field, were exposed to different temperature regimes for 16 weeks while dormant. To minimize short-term carry-over effects, after the temperature treatments, all seedlings were conditioned to 5.5 degrees C and watered before the assessment of non-structural carbohydrates and cold tolerance. Needle sugar concentration was decreased by 54, 32, 21 and 9% following treatment at 5.5, 0, -1.5 and -8.5 degrees C, respectively. Sugar concentration did not decrease as much in root tissues as in needles because starch was mobilized in roots. Cold tolerance of needles was analyzed by controlled freezing, and the temperature causing an initial 10% damage (LT(10)) was plotted as a function of needle sugar concentration, revealing a strong, linear relationship. When one-third of the initial sugars had been consumed, LT(10) had increased from -24.5 to -16.5 degrees C, and when one half had been consumed, LT(10) had increased to -10 degrees C. Consequences of these findings for the field performance of conifers are discussed in relation to climatic variation and change.     

Chemical and physiological responses of four Turkish red pine (Pinus brutia Ten.) provenances to cold temperature treatments

Determining the adaptability to abiotic conditions and potential establishment success of tree species needs to be conducted before attempting to use a species in large-scale afforestation programs. In this study, the chemical and physiological performance of four Turkish red pine (Pinus brutia Ten.) provenances was investigated after exposure to artificial cold temperature treatments to determine their adaptability to cold environment for potential use in afforestation programs. Seeds were sown and raised for 24, 28, and 32 weeks and exposed to decreasing temperatures in an artificial freezer. Relative electrolyte leakage, chlorophyll fluorescence, and carbohydrate concentrations were measured to determine the variability between provenances. Results showed that diameter and height growth did not vary with origin for each of the three growth stages measured. Root electrolyte leakage values differed between provenances, confirming that cold stress was effectively causing physiological damages when plants were exposed to temperature at ?15 °C and below. The variability observed in the relationship between provenances and cold hardiness responses can be attributed to tree-to-tree variability within provenances and microsites conditions. There was generally no significant difference in chlorophyll fluorescence between provenances, also attributed to low genetic variation between provenances. Carbohydrate concentrations were also very variable and varied significantly among growth stages and provenances. High-altitude provenances had higher soluble carbohydrates concentrations in several cases, suggesting a relationship between altitude, soluble sugars, and cold hardiness. However, these trends were not consistent; therefore, we suggest that such hypotheses be confirmed through more comprehensive further studies.     

Seasonal fluctuations of low‐molecular‐weight sugars,starch and nitrogen in sapwood of Pinus sylvestris L.

The content of water‐soluble substances and starch in the living tree stem at the time of felling influences wood durability during further utilization. The aim of the study was to describe the annual and seasonal fluctuations in the contents of stored carbohydrates and nitrogenous compounds. The contents of soluble sugars, starch, and nitrogen were measured in the outer sapwood (0–15 mm from cambium) of 65‐yr‐old Scots pine {Pinus sylvestris L.) trees during an annual cycle. To study the influence of growth conditions, samples were taken from different stands in Sweden, in one stand from a control and a fertilized block. The effect of the age of the trees on the content of stored substances was also considered by comparing young (10–15 yrs old) and older trees (40–65 yrs old). Determination of the carbohydrates was carried out using enzymatic analysis. The outer pine sapwood contained a higher content of low‐molecular weight sugars during autumn and winter than during spring and summer. Starch content rose at the beginning of the growth period and decreased in autumn. The content of soluble sugars increased towards the cambium on all sampling occasions. Fructose and glucose were dominant sugars in all the stands studied. Seasonal changes of sucrose were different from those of glucose and fructose, in that the sucrose content was already decreasing in February. The variations in the nitrogen content of the sapwood were far smaller than those of the soluble sugars. No significant differences were found between the stands with regard to glucose, fructose and the sum of all three sugars. The fertilized block showed low content of sucrose and nitrogen. It is concluded that seasonal fluctuations are more important with regard to the sugar content than the effect of growth conditions. Data indicate a lower content of soluble sugars and nitrogen in the outermost sapwood of young trees as compared with older trees.     

Diurnal changes in gas exchange and carbon partitioning in needles of fast- and slow-growing families of loblolly pine (Pinus taeda)

We investigated diurnal and seasonal changes in carbon acquisition and partitioning of recently assimilated carbon in fast- and slow-growing families of loblolly pine (Pinus taeda L.) to determine whether fast-growing families exhibited greater carbon gain at the leaf level. Since planting on a xeric infertile site in Scotland County, NC, USA in 1993, five Atlantic Coastal Plain (ACP) and five "Lost Pines" Texas (TX) families have been grown with either optimal nutrition or without fertilization (control). In 1998 and 1999, gas exchange parameters were monitored bimonthly in four families and needles were analyzed bimonthly for starch and soluble sugar concentrations. Although diurnal and seasonal effects on net photosynthesis (A(net)) and maximum rate of light-saturated photosynthesis (A(max)) were significant, few family or treatment differences in gas exchange characteristics were observed. The A(net) peaked at different times during the day over the season, and A(max) was generally highest in May. Instantaneous water-use efficiency (WUE(i)), derived from gas exchange parameters, did not differ among families, whereas foliage stable isotope composition (delta(13)C) values suggested that TX families exhibited lower WUE than more mesic ACP families. Although there were no diurnal effects on foliar starch concentrations, needles exhibited pronounced seasonal changes in absolute concentrations of total nonstructural carbohydrates (TNC), starch and soluble sugars, and in partitioning of TNC to starch and sugars, mirroring seasonal changes in photosynthesis and shoot and root growth. In all families, foliar starch concentrations peaked in May and decreased to a minimum in winter, whereas reducing sugar concentrations were highest in winter. Some family and treatment differences in partitioning of recently assimilated carbon in needles were observed, with the two TX families exhibiting higher concentrations of TNC and starch and enhanced starch partitioning compared with the ACP families. We conclude that growth differences among the four families are not a function of differences in carbon acquisition or partitioning at the leaf level.     

The Influence of Desiccation, Rough Handling and Cold Storage on the Quality and Establishment of Sitka Spruce Planting Stock

In January 3-year-old Sitka spruce were lifted and treated eithercarefully or roughly. Plants in each treatment were either transferredto the laboratory for testing, or transferred to cold storageat –1°C. In the laboratory, half of the plants ineach treatment were desiccated to shoot water potentials ofless than –2.0 MPa. Plants were then tested for root growthpotential (RGP), bud dormancy, frost hardiness and carbohydratecontent before transplanting at a field site, Additional plantswere removed from the nursery and cold store at approximatelymonthly intervals until late April, and then treated as above.In April, plants in each treatment were transferred to on-sitefield storage (sheughs), for 2 or 4 weeks prior to testing asabove. Carefully handled plants maintained large RGP which was positivelycorrelated with plant water potential, root water content, shootrelative growth rate and field survival. Overall RGP was reduced: 59 per cent by desiccation; 85 percent by rough handling; and 98 per cent by desiccation and roughhandling in combination. On site storage for 4 weeks in April/May reduced RGP. Whereplants produced more than 30 new roots > 1 cm long in RGPtests, field survival exceeded 90 per cent. Cold storage at–1°C maintained RGP, (of carefully handled plants),bud dormancy and shoot frost hardiness at approximately pre-storagevalues. In contrast, total nonstructural carbohydrate contentdeclined in cold store but increased in March/April for plantsin the nursery.     

Carbohydrate reserve accumulation and depletion in Engelmann spruce (Picea engelmannii Parry): effects of cold storage and pre-storage CO(2) enrichment

The effects of pre-storage CO(2) enrichment on growth, non-structural carbohydrates and post-storage root growth potential of Engelmann spruce (Picea engelmannii Parry) seedlings were studied. Seedlings were grown from seed for 202 days in growth chambers with ambient (340 micro l l(-1)) or CO(2) enriched (1000 micro l l(-1)) air. Some seedlings were transferred between CO(2) treatments at 60 and 120 days. Photoperiod was reduced at 100 days to induce bud set and temperature was reduced at 180 days to promote frost hardiness development for storage at -5 degrees C for 2 or 4 months. Stored seedlings were planted in a growth chamber after thawing for one week at +5 degrees C. At 80, 120, 140 and 202 days, and at each planting time after storage, seedlings were harvested for growth measurements and analysis of starch and soluble sugar concentrations. Planted seedlings were assessed for bud break every two days and new roots > 5 mm long were counted after four weeks. Carbon dioxide enrichment increased root collar diameter and almost doubled seedling biomass, with the most obvious effects occurring after bud set. Stem height was affected only slightly and shoot/root ratios were not affected at all. Carbon dioxide enrichment increased the rate of reserve carbohydrate accumulation, but did not influence the final concentration attained before storage (accounting for 32% of seedling dry weight). Needles were the major storage organ for soluble sugars, whereas roots were the major storage organ for starch. Soluble sugars were not strongly affected by two or four months of storage, but starch was reduced by more than 50% in all plant parts. None of the CO(2) treatments had an impact on bud break or root growth potential.     

Effects of compaction and simulated root channels in the subsoil on root development, water uptake and growth of radiata pine

Effects of subsoil compaction and simulated root channels (perforations) through the compacted layer on root growth, water uptake, foliar nutrient concentration and growth of radiata pine (Pinus radiata D. Don) were studied in a field experiment where a range of treatments were applied in reconstituted soil profiles. Subsoil compaction adversely affected root penetration in deeper parts of the soil and consequently caused greater water stress in trees. However, the effect of compaction was largely overcome when the subsoil was perforated to render 0.2% of the soil volume into vertical channels. Roots showed a remarkable ability to reach the points of low penetration strength and to travel through them to deeper parts of the profile. Perforations through compacted soil layers at a relatively low frequency may be a practical solution to allow root development into deeper parts of the soil and allow greater soil water exploration by roots.     

Identification and quantitative analysis of stage-specific carbohydrates in loblolly pine (Pinus taeda) zygotic embryo and female gametophyte tissues

Stage-specific analyses of starch and 18 sugars, including pentoses, hexoses, disaccharides, trisaccharides, oligosaccharides and sugar alcohols, were made throughout seed development for zygotic embryo and female gametophyte (FG) tissues of loblolly pine (Pinus taeda L.). Tissue was most often analyzed in triplicate from two open-pollinated families grown in different locations and sampled in different years. Carbohydrates were analyzed by enzymatic assay, high performance liquid chromatography or gas chromatography/mass spectrometry. For all carbohydrates quantified, peak concentrations were higher in embryo tissue than in FG tissue. Significant changes in starch and sugar concentrations occurred over time, with both seed collections showing similar trends in temporal changes. Although concentrations were not always similar, embryo and FG tissues generally showed similar patterns of change in starch and sugar concentrations over time. Total starch concentration was highest during early seed development and decreased as development progressed. The major sugars contributing to osmotic potential during early seed development were D-pinitol, sucrose, fructose and glucose. During mid-seed development, D-pinitol, sucrose, fructose, glucose, melibiose and raffinose provided major contributions to the osmotic environment. During late seed development, sucrose, raffinose, melibiose, stachyose and fructose were the major contributors to osmotic potential. These data suggest stage-specific media composition for each step in the somatic embryogenesis protocol.     

Comparison of three cold hardiness tests for conifer seedlings

Greenhouse-cultured, container-grown ponderosa pine (Pinus ponderosa var. scopulorum Engelm.), interior Douglas-fir (Pseudotsuga menziesii var. glauca (Beissn.) Franco), and Engelmann spruce (Picea engelmannii (Parry) Engelm.) were cold acclimated and deacclimated in growth chambers over 19 weeks. Cold hardiness was measured weekly by a whole-plant freeze test and by two quick tissue tests: freeze-induced electrolyte leakage of needles, and differential thermal analysis of buds. The whole-plant freeze test provided results in 7 days, and indicated differences in cold hardiness among stems, buds, and needles. Although the whole-plant freeze test could accurately measure cold hardiness, it was not precise, and it required destructive sampling. Results from freeze-induced electrolyte leakage and differential thermal analysis were available in 2 days and 1 hour, respectively. The freeze-induced electrolyte leakage test was a precise, sensitive and objective predictor of changes or differences in tissue cold hardiness. To determine actual cold hardiness, results could be calibrated to the response of the same tissue in the whole-plant freeze test. The speed and objectivity of differential thermal analysis made this test useful for rapid, general assessment of cold hardiness status, but calibration was difficult, and precision varied.     

Mineral nutrition,resin flow and phloem phytochemistry in loblolly pine

Southern pine beetles and associated pathogenic fungi represent the largest biotic threat to pine forests in the southeastern USA. The two primary defensive mechanisms of the tree to the beetle-fungal complex are the primary oleoresin flow and the concentrations of preformed and induced secondary compounds. We compared oleoresin flow and concentrations of phloem nutrients, soluble sugars, starch, total phenolics and proanthocyanidins in Pinus taeda L. trees in fertilized and control plots in the Sandhills region of North Carolina. Four blocks of 10 trees per treatment were sampled on five dates from May to November 1995. Phloem nitrogen and potassium concentrations were elevated in trees on fertilized plots, whereas phloem calcium concentrations were decreased. Fertilization significantly enhanced (10-20%) concentrations of phloem phenolics and proanthocyanidins. In contrast, phloem soluble sugars and starch concentrations were up to 30% lower in fertilized trees than in control trees. Increased phenolic concentrations and lower nonstructural carbohydrates should correlate with reduced tissue palatability and decreased pathogen susceptibility in fertilized trees; however, resin flows were significantly lower (30-100%) in fertilized trees compared with control trees, which may facilitate pine bark beetle establishment. Furthermore, fertilization-induced increases in phloem nitrogen concentration may be more important than tissue carbohydrate or phenolic content in determining tissue palatability.     

Carbohydrates in individual poplar fine roots: effects of root age and defoliation

Late-summer starch accumulation in fine roots of poplars (Populus x canadensis Moench.) defoliated by gypsy moth (Lymantria dispar L.) lagged behind that in fine roots of undefoliated trees. If starch concentration declines with age, defoliation-induced changes in root system age structure could be partly responsible for this difference. To test this hypothesis, we measured fine-root starch and soluble sugar concentrations in roots of known age from trees in defoliated and undefoliated plots. There was a significant interaction between the effects of defoliation and root type (white, brown, or dead) on fine root soluble sugar concentration because of the high concentration of soluble sugars in white roots from trees in undefoliated plots. Both root starch and soluble sugar concentrations were variable among individuals of each age class. The population frequency distributions for starch and soluble sugar concentrations were both right-skewed, and fit by exponential functions. These data are most consistent with direct defoliation effects on a labile and dynamic pool of carbohydrates in poplar fine roots, rather than indirect defoliation effects on root system age structure.     

Foliage re-establishment of <Emphasis Type="Italic">Pinus palustris</Emphasis> Mill. saplings after spring or fall prescribed fire

Repeated fire is key to the viability of longleaf pine (Pinus palustris Mill.) ecosystems, but its acceptance as a management tool may depend on satisfactory longleaf pine growth. This is because longleaf pine establishment often has the dual-purpose of ecosystem restoration and stemwood production. Timely recovery of scorched foliage among longleaf pine seedlings and saplings supports maximum juvenile growth. We identified two means of foliage re-establishment in the growing season after prescribed fire regardless of the season of fire application. New foliage growth after spring or fall fire was correlated with un-scorched foliage biomass and the presence of lateral branches. After prescribed fire in spring, foliage biomass recovery also appeared to benefit from the mobilization of starch. The high carbon demand of foliage recovery after fall prescribed fire was associated with interruption of seasonal starch accumulation in the stem and taproot. The implication of low starch accumulation in stem and taproot tissues during the growing season after fall prescribed fire is unknown and warrants further investigation. Our results demonstrate a positive influence of residual foliage, lateral branches, and stored starch on timely foliage recovery of young longleaf pines after fire. Together with knowledge of longleaf pine development and fuel and climate conditions at the time of prescribed fire, this information will aid prescribed fire practitioners charged with maintaining longleaf pine stands of high vigor.     

The effect of late summer fertilization on the frost hardening of second-year Scots pine seedlings

In this study the effect of summer fertilization on the initiation of frost hardening of containerized second-year Scots pine (Pinus sylvestris L.) seedlings is studied. During the second growing season three different fertilization programs (water soluble NPK with micronutrients) determined by electrical conductivity of peat water extract (0.2, 0.5 and 1.2 mS cm^-1) were initiated. The growth and nutrient concentrations of needles were monitored during the fertilization period. The frost hardiness of seedlings was assessed on four separate occasions at two week intervals from August 7 to September 18. This assessment was based on artificial freezing tests and visual damage scoring of tissue browning on current-year needles. Clear differences in foliar N, P and K concentrations were observed between the fertilization treatments. Fertilization prolonged the growing period of needles and increased root collar diameter. In all the tests, the highest fertilization level resulted in the highest level of frost hardiness. The difference between the fertilization treatments ranged from 1 °C to 2.2 °C. Frost hardiness increased with an increase in foliar nitrogen concentration and slightly less consistently with increases in foliar phosphorus and potassium concentrations.