Timing of cold temperature exposure affects root and shoot frost hardiness of Picea Mariana container seedlings

Seventeen‐week‐old black spruce seedlings were hardened under short daylengths and one of three short day length environments, which were either warm (24/16°C, day/night) throughout a 10 week hardening period (WW), cool (10/5°C) throughout hardening (CC), or warm for three weeks followed by seven weeks of cool temperatures (WC). Greatest root and shoot frost hardiness resulted from the exposure of seedlings to three weeks of warm followed by seven weeks of cool temperatures. Seedlings receiving warm temperatures throughout hardening increased in root and shoot frost hardiness, but to a lesser extent than seedlings exposed to cool temperatures. The frost hardiness of woody roots was generally greater than that of fine roots, but the extent of the difference in frost hardiness depended on the time since bud initiation and on the hardening treatment.     

Temperature control of the development of frost hardiness in two populations of Leptospermum scoparium

Seedlings of Leptospermum scoparium J.R. et G. Forst (manuka) originating from seed from a low altitude coastal site (Auckland) and from a high altitude inland site (Desert Road) were grown for 96 days in four controlled environments to compare the relationship between growth temperature and frost hardening. Day/night temperature treatments were 12/6, 12/3, 12/0 and 12/-3 degrees C. Frost hardiness was determined at 14-day intervals by exposing whole seedlings to temperatures ranging from -2 to -8 degrees C. Frost damage differed significantly between the two populations: Desert Road seedlings were less affected than Auckland seedlings. At all growth temperatures, the time courses of frost hardiness of both populations followed curvilinear relationships reaching a maximum hardiness at about Day 50, after which the seedlings spontaneously dehardened. The rate of frost hardening increased linearly with decreasing temperature from 6 to 0 degrees C, but thereafter, no further increase occurred with decreasing temperature to -3 degrees C. The frost hardening process was more sensitive to temperature in the Desert Road seedlings than in the Auckland seedlings, and this difference may account for the intraspecific variation in frost hardening capacity of this species. Comparisons with Pinus radiata D. Don and Lolium perenne L. indicated that interspecific variation in frost hardening capacity can also be accounted for by differences in the sensitivity of the hardening process to temperature.     

The effect of photosynthetic photon flux density on development of frost hardiness in top and roots of Larix leptolepis seedlings

Seedlings of Larix leptolepis were grown in a growth chamber under short day conditions (8 h day) at 4 different photosynthetic photon flux densities (PPFD's). After a period of 8 weeks frost hardiness of the shoot tips and roots, dry matter content, dry weight (dw), content of glucose and starch were determined. The frost hardiness in the shoot tips increased from ‐15°C at a PPFD of 55μmol m^‐2s^‐1 to about ‐35°C at 440 μmol m^‐2s^‐1. No effect of PPFD was found on frost hardiness of the roots. A high PPFD results in a high dry matter content. The effect on dry matter content was most pronounced for the shoot tips and less pronounced for the roots. The total dry weight increased for both root and top with increasing PPFD. The height of the plants increased when the PPFD increased up to 220 μmol m^‐2s^‐1.     

Effects of nitrogen fertilization and temperature on frost hardiness of Aleppo pine (Pinus halepensis Mill.) seedlings assessed by chlorophyll fluorescence

In a 14-week study, 1-year-old Aleppo pine seedlings were grownin two growth chambers. Seedlings were artificially hardenedby decreasing photoperiod and temperature. In each chamber halfof the seedlings were fertilized with nitrogen (8.4 mg seedling^–1).In order to determine the relative importance of the hardeningenvironment versus fertilization, each chamber was programmedto decrease night temperatures down to a low of 8 or 4°C.Chlorophyll fluorescence and frost hardiness was measured fivetimes during the experiment. A sample of seedlings from eachtreatment was exposed to an artificial frost at –5°Cand the freezing effects were assessed by measurements of chlorophyllfluorescence and visual evaluation of needle damage. Seedlingsincreased their frost hardiness during the experiment in allthe treatments but the ratio of variable to maximal chlorophyllfluorescence (F_v/F_m) measured before freezing did not vary duringthe experiment. This indicates that Aleppo pine maintains itsphotosynthetic ability during hardening in contrast to otherconiferous species from colder climates. The effect of nitrogenfertilization on frost hardiness was small in comparison withchamber effect. Nitrogen fertilization slightly delayed theacquisition of hardening in the coldest chamber. Seedlings inthe warmest chamber did not become fully resistant to –5°C,but in the coldest chamber, where night temperature reached4°C, all the seedlings were resistant to the frost. Severedamage caused by frost could be related to a rapid rise of minimalfluorescence (F₀) but the best index of damage was the dropof F_v/F_m after freezing.     

Frost hardening in first-year eastern larch (Larix laricina) container seedlings

Eastern larch (Larix laricina [du Roi] K. Koch) container seedlings were tested to determine shoot frost hardiness development under short or long days and warm (15 to 25 °C) or cool (10/5 °C, day/night) temperatures, to aid in the development of greenhouse hardening strategies. Seedlings were sampled sequentially over time (25 seedlings per week) from a population of 1000 trees. Frost hardiness increased significantly after one week of fluctuated over the next 6 weeks, and increased thereafter through week 14. Seven weeks of warm, intermittent short days, followed by 6 weeks of cool, continuous short days, resulted in greater frost hardiness than 13 weeks of warm, intermittent short days. In contrast, seedlings exposed to 7 weeks of warm, intermittent short days, followed by six weeks of warm, long days were significantly less frost hardy. Stems with needles attached had lower Index of Injury than stems without needles.     

Germination and seedling growth in Abies lasiocarpa (Hook.) Nutt. as affected by provenance,seed pretreatment,and temperature regime

Abies lasiocarpa (Hook.) Nutt. seeds germinated and the seedlings grew on under controlled environments (18 h photoperiod and 16/8 h thermoperiods; 10/10^oC, 20/20°C, 20/10°C, and 25/10°C). The experiment was concluded after 180 days, and the seedlings were scored for stem diameter and length. Four provenances were included: Colorado “high elevation”; (CO), Oregon 1 600 m a.s.l. (OR), British Columbia 1200 m a.s.l. (BC), and Finland (BCF; originally from British Columbia 1800 m a.s.l.). A cold‐moist pretreatment for 30 days improved germination and seedling growth in the BC and BCF seed lots, while the CO and OR seed lots were less affected. Daily fluctuating temperatures improved germination in the BC and BCF seed lots, the widest fluctuation 25/10°C producing the greatest germination. The average length of the seedlings was 34 mm at 20/20°C, 33 mm at 20/10°C, and 37 mm at 25/10°C; differences in hypocotyl length being most important. There were small but significant differences between the provenances in their reaction to temperature regime. The major part of the seedlings produced 2 or 3 flushes of growth in the 180 days period, the CO provenance having the most stabile buds and the OR provenance being the most labile.     

Phenotypic Differences Between Natural and Selected Populations of Picea Abies. I. Frost Hardiness

The frost hardiness of non-juvenile Norway spruce [Picea abies (L.) Karst.] populations growing in northern Sweden (63°54' N) was monitored during 1996-1997. The investigated progenies originated from 12 natural populations and six seed orchards located between 58° N and 68° N in Sweden. Frost hardiness of needles was assessed by measuring chlorophyll fluorescence and electrolyte leakage after freezing. The loss of frost hardiness in 1-yr-old needles during spring occurred slightly earlier in populations originating north of 63°30' N than in those originating further south. Dehardening was slightly delayed in selected populations compared with natural populations of similar origin. The level of frost hardiness during autumn was higher in populations originating north of 63°30' N than in those originating south of this latitude, but there were no clear differences in frost hardiness between selected and natural populations of similar origin. The results are discussed in relation to climatic factors and inherent growth rhythms.     

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.     

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.     

国外木本植物抗寒性测定方法综述

文中对目前国外采用的不同抗寒性测定方法的测定原理、测定程序以及优缺点等作了较为详细的介绍。这些测定方法有全株冰冻测试法（组织褐变法）、电解质渗出率法、叶绿素荧光法、热分析法（主要用差热分析法）、电阻抗图谱法以及核磁共振显微镜图谱法和可视＋近红外线光谱法等。并简介了这些测定方法的适用性和抗寒性测定方法的研究展望。     

Morphological and physiological differences in Scots pine seedlings of six seed origins

One-year-old seedlings of Scots pine (Pinus sylvestris L.) offour native seed origins (Loch Maree Islands, Glengarry/GlenMorriston, Glen Affric and Abernethy), a commercial Britishseedlot, and a seedlot from Hedesunda, in middle Sweden, werecompared at monthly intervals from October 1993 to April 1994.Seedling morphology, root condition, root frost hardiness andbud dry matter were determined at each date. There were clear morphological differences among seed origins.Seedlings raised from the commercial seedlot (A70) were largerbut had a poorer root:shoot ratio than the other seed origins.Of the native pines tested, the Loch Maree Islands origin allocateda larger proportion of its photosynthate to fine roots and needlesand smaller proportion to woody structures. Seedlings raised from the commercial British seedlot tendedto have poorer bud lignification than the other origins andalso, in autumn, higher electrolyte leakage rates from its fineroots. During winter, the Swedish origin had the lowest fineroot electrolyte leakage. Seedlings of all origins showed aprogressive increase in fine root hardiness towards mid-winterwith maximum hardiness (–7°C) in January. Dehardeningoccurred over subsequent months reaching –3°C in April.Differences among origins were evident. The Swedish seedlotdeveloped greater frost resistance than the other origins, hardeningbegan earlier in autumn and dehardening began later in spring.The commercial seedlot hardened later than the other originsbut reached a similar level of frost hardiness by January. Ofthe native pines, seedlings of the Loch Maree Islands originwere slowest to develop root hardiness.     

Elevated temperature during reproductive development affects cone traits and progeny performance in Picea glauca x engelmannii complex

Two temperature regimes were applied during reproductive development of seed and pollen cones of interior spruce (Picea glauca (Moench) Voss and Picea engelmannii (Parry) complex) to determine temperature effects on the adaptive traits of progeny. In Experiment 1, identical crosses were made on potted interior spruce using untreated pollen followed by exposure to a day/night temperature of 22/8 or 14/8 degrees C with a 12-h photoperiod during the stages of reproductive development from post-pollination to early embryo development. Frost hardiness and growth of progeny from seed produced in the two temperature treatments were measured over a 4-year period. Elevated temperature significantly affected both seed-cone development and the adaptive properties of the progeny. Seed cones exposed to the 22/8 degrees C treatment reached the early embryo stage in 53 days versus 92 days in the 14/8 degrees C treatment. Seed yields, cotyledon emergence and percent germination were also significantly enhanced by the 22/8 degrees C treatment. Progeny from seed produced in the higher temperature treatment showed significantly reduced spring and fall frost hardiness, but the elevated temperature treatment had no significant effects on time of bud burst, growth patterns or final heights. In Experiment 2, single ramets of the same clone were subjected to a day/night temperature of 20/8 or 10/8 degrees C during pollen cone development, starting from meiosis and ending at pollen shedding. The two populations of pollen were then crossed with untreated seed cones. Compared with pollen cones exposed to the 10/8 degrees C treatment, pollen cones exposed to the 20/8 degrees C treatment during development reached the shedding stage 2-4 weeks earlier, whereas pollen yields, in vitro viability and fertility (seed set) were significantly lower; however, the resulting progeny displayed no treatment differences in frost hardiness or growth after 1 year. Results suggest that seed orchard after-effects could be caused by temperature differences between orchard site and parent tree origin and that this effect acts on maternal development. Gametophytic (pollen or megagametophyte or both) and early embryo (sporophytic) selection are possible mechanisms that may explain the observed results. Although the effects are biologically significant, they are relatively small and do not justify changes in current deployment strategies for seed orchard seed.     

Seasonal Changes in the Frost Hardiness of Provenances of Picea sitchensis in Scotland

Changes in the natural level of frost hardiness of shoots offour provenances of Picea sitchensis were monitored over twogrowing seasons by detaching shoots from 7 to 10-year-old treesgrowing in a nursery in Scotland, and subjecting them to freezingtemperatures under conditions which simulated night frosts. Six seasonal phases of frost hardiness were identified (Fig.3).

1. During each autumn, killing temperatures (the level of hardiness)decreased from –5°C to below –20°C, beginningseveral weeks after shoot elongation ceased. Alaskan provenanceshardened in September, apparently in response to shorteningday lengths alone, whereas an Oregon provenance did not hardenuntil November, after repeated frosts. Queen Charlotte Islandsprovenances were intermediate.
2. From November to March allprovenances were hardy to below –20°C,which is adequateto prevent direct freezing injury at mostplantation sites.
3. In March-April, several weeks before bud-burst, old shootsdehardenedto killing temperatures of about –10°Cin responseto warm temperatures, and southerly provenancesdid so beforenortherly ones.
4. During bud-burst the newly-emergingshoots were hardy to only–3°C to –5°C untilthey were about 3.5 cmlong. All provenances burst bud at thesame time and were equallyfrost susceptible at this time.
5. DuringMay-July the elongating shoots fluctuated in hardinessbetween–5°C and –10°C apparently in responsetofluctuating ambient temperatures.
6. In August 1980 there wasa period of late summer dehardeningto killing temperaturesof about –3°C.

Seasonal changes in hardiness are discussed in relation to changesin shoot growth and environmental factors. The main opportunitiesfor selecting frost hardy genotypes seem to be in the rate ofautumn hardening, the time of pre-bud burst dehardening, andthe time of bud-burst.     

Influence of environment, fertilizer and genotype on shoot morphology and subsequent rooting of birch cuttings

Differences in rooting ability of birch (Betula pubescens J.F. Ehrh.) cuttings were observed as a result of differences in genotype and physiology of the stock plants. The uniformity in response among cuttings from micropropagated plants compared with cuttings from seed plants confirmed the advantage of using micropropagated plants to study environmental effects. Shoot morphology of the seed stock plants was influenced by both photoperiod and thermoperiod. A day/night temperature of 15/25 degrees C reduced stem elongation compared with a day/night temperature of 25/15 degrees C regardless of photoperiod, and a continuous light regime resulted in more shoots per plant in both temperature regimes than a 16-h photoperiod. A reduction in the supply of macronutrients did not influence shoot morphology, but increased rooting substantially and seemed to override the effects of environmental factors. In cuttings of seed plants, the highest rooting percentage and number of roots were obtained in a 16-h photoperiod with a day/night temperature of 15/25 degrees C. In micropropagated stock plants, there was a positive correlation between shoot length and number of leaves per shoot and topographical distribution of light within the plants, but there was no correlation between these parameters and rooting ability of the cuttings. A rooting temperature of 16 degrees C delayed the rate of root production compared with the rate at higher temperatures, but the final rooting percentage was the same over the range from 16 to 28 degrees C. Root branching increased with temperature. At all temperatures, there was a large increase in sucrose content at the base of the cuttings during rooting, whereas the concentration of nontranslocated sugars remained constant. The carbohydrate content at the base of cuttings from micropropagated stock plants was three times higher than at the base of cuttings from seed stock plants, but the higher carbohydrate content was not correlated with a higher rooting potential.     

Genotype and environment: two factors related to autumn cold hardiness on Persian walnut (Juglans regia L.)

Context

J. regia timber is appreciated for high-value wood products. In new plantations, biotic or abiotic events which could affect wood quality should be monitored. Autumn frosts could affect annual shoot development, with consequent loss of timber value or even tree death. In southern Europe, climate change forecasts include erratic and severe autumn frost events.

Aims

The relationship between genotype and environment regarding susceptibility to autumn frost damage was examined in four provenances of Juglans regia L., planted at two ecologically different sites, one subject to Mediterranean and the other Atlantic weather conditions.

Methods

Annual budsticks from eight trees per provenance were collected in November 2010 from each site. The samples were then submitted to freeze–thaw cycles down to ?8, ?13 and ?18 °C, plus a control treatment (keeping samples at 5 °C). Damage to the stem was assessed using the index of freezing injury calculated from relative electrolyte leakage at each temperature considered. Frost damage to stem, apical and lateral buds was recorded by visual scoring.

Results

Differences in cold acclimation between sites were detected, with provenances exhibiting differences on senescence. A ‘provenance?×?site’ interaction was found in some of the analyses, but the same ranking of susceptibility was detected for all provenances at both sites and in all the tissues analysed. The differences between provenances could be related to their geographical origins, where an altitude gradient was observed.

Conclusions

The genetic component was important in the expression of autumn cold hardiness and, together with productive traits, should be considered in new afforestation projects.     

Frost Hardiness of Subalpine Eucalypts in Britain

Laboratory freezing tests were used to determine seasonal changesin the frost hardiness of detached shoots of young trees ofEucalyptus gunnii (from central Tasmania), and E. niphophilaand E. debeuzevillei (‘snow gums’ from the SnowyMountains, Australian Capital Territory). The trees were growingat the Bush Estate. No difference was found between the speciesor between seedlots, all of which were from high altitudes nearthe tree line. In midwinter (February) the shoots tolerated –16°Cwithout suffering damage, and many shoots survived temperaturesas low as –18°C to –22°C. This result agreedwith Evans' (1986) observation that some trees within theseseedlots survived temperatures in the range –19°Cto –23°C in field planting during the winter 1981/82.In their native habitats the trees rarely experience temperaturesbelow –20°C. The shoots did not harden appreciably before they experiencedfrosts (in late October/early November) and so may be proneto autumn frost damage. By contrast, they were slow to dehardenin late winter and spring and did not seem prone to spring frostdamage. There were no killing air frosts during the period of this study(winter 1985/6), but many trees died, possibly as a result ofground freezing, producing root injury and/or shoot desiccation.In subalpine regions of Tasmania and the Snowy Mountains theground is covered by snow throughout the winter. Variation in frost hardiness within these hardy seedlots couldbe exploited.     

Cold hardiness and transplant response of Juglans nigra seedlings subjected to alternative storage regimes

? Effects of overwinter storage regimes on seedling cold hardiness and physiological vigor are relatively unexplored, particularly for temperate deciduous forest tree species.

? We evaluated influence of storage duration (0, 66, 119, or 175 d) on electrolyte leakage of stem and root collar tissues following exposure to a series of freeze-test temperatures in black walnut (Juglans nigra L.) seedlings sampled from cold (3 °C) or freezer (?2 °C) storage. Seedlings were subsequently transplanted into a controlled growth chamber environment for two months.

? Regardless of storage temperature, mean LT₅₀ was lowest for seedlings stored for 66 d (≤ ?34 °C) and increased dramatically after 119 d (≥ ?13 °C).

? Root collar tissue had lower LT₅₀ than stem tissue after 119 d for cold-stored seedlings, reflecting importance of evaluative tissue type. Days to bud break shortened with increasing storage duration up to 119 d and stabilized thereafter for both storage regimes. Root growth potential was maximized after 119 d of storage, and subsequently declined for cold-stored seedlings. Height growth increased following storage, regardless of duration.

? To promote stress resistance and transplant growth response, we recommend that black walnut seedlings from this genetic source be outplanted after approximately 66–119 d of storage.

     

Growth and visible responses of Scots pine (Pinus sylvestris L.) seedlings to simulated summer frost

Scots pine seedlings, 20–30 days old, were exposed to simulated summer frost in controlled environment growth chambers. The responses observed showed extreme variation between individual seedlings; seedlings suffering from needle-necrosis, healthy-looking, and dead seedlings were found in the same treatment. Four days exposure to temperatures below –4.5°C caused multiple-leaders maximally in 7.7 % of the pine seedlings and resulted in decreased shoot and root dry weight and shoot length.     

Fast, nondestructive measurement of frost hardiness in conifer seedlings by VIS+NIR spectroscopy

Frost hardiness development from mid-August to mid-November was evaluated in seedlings of three provenances of Norway spruce (Picea abies (L.) Karst.) and three provenances of Scots pine (Pinus sylvestris L.) raised at nurseries in north, central and south Sweden. Measurements of the visible + near infrared (VIS+NIR) spectra of shoots were made simultaneously with estimates of frost hardiness based on electrolyte leakage following artificial freezing. Nine physiological variables known to influence frost hardiness were measured throughout the experiment. Multivariate analysis showed that VIS+NIR spectra explained 69% and 72% of the variation in frost hardiness in Scots pine and Norway spruce, respectively. Stem lignification, dry weight fraction, and starch, glucose, fructose, galactose, sucrose, raffinose and stachyose concentrations together explained 80% and 85% of the variation in frost hardiness in Scots pine and Norway spruce, respectively when used as independent X variables in a partial least squares model. These physiological variables could be related to varying degrees with variation in the VIS+NIR spectra. We conclude that VIS+NIR spectroscopy provides a rapid nondestructive technique for measuring frost hardiness in conifer seedlings based on causal relationships between the spectra and the physiology of seedling frost hardiness.     

Effects of nutritional factors on frost hardening in Larix leptolepis (Sieb & Zucc.) Gord.

A method for estimating frost hardiness in seedlings of Larix leptolepis in their first stage of acclimation is demonstrated. Cuttings of the shoot tip were frozen linearly from +2°C to ‐36°C. The viability was determined by TTC‐assay. Data were fitted to a logistic regression model, and hardiness was calculated from the fitting parameters. It was shown that hardiness is influenced by the duration of the fertilizer application period. No effect of the nutrient status of N, P or K in the shoots was found.