Transpiration and root water uptake by olive trees

While the cultivated olive tree (Olea europaea L.) is known to be sclerophyllous and effective at tolerating drought, little is known of its short-term water-use dynamics for most studies have been based on longer-term, water-balance information. We present here, for the first time, heat-pulse measurements of the sap flux measured not only within the semi-trunk of an olive tree, but also within a root excavated close to the stump. One tree in the olive grove near Seville in Spain had regularly received basin irrigation during the summer, whereas the other, growing on this deep silt loam, had been without water for over 3 months. Following a flood irrigation of 730 L to a dyked area around the tree, the regularly-irrigated olive maintained a transpiration rate of 1.65 mm³ mm^–2 d^–1, on a leaf area basis, for only 3 days following the irrigation. This rate was maintained for a total consumption of 110 L. It then began again to limit its rate of water use with transpiration falling below that predicted for well-watered conditions by the Penman-Monteith equation. The flow of sap in the near-surface root dropped concomitantly. Meanwhile the unirrigated tree was using water at just 0.78 mm d^–1. Yet following an irrigation of 870 L it only lifted its consumption to 1.12 mm d^–1, on a leaf area basis. Neither did it recover its leaf water potential following this wetting because of an inability to refill cavitated vessels. These data again show olive to be a parsimonious and cautious consumer of soil water.     

Osmoregulation and osmoprotection in the leaf cells of two olive cultivars subjected to severe water deficit

In this study, we compared the efficacy of defense mechanisms against severe water deficit in the leaves of two olive (Olea europaea L.) cultivars, ‘Chemlali’ and ‘Meski’, reputed drought resistant and drought sensitive, respectively. Two-year old plants growing in sand filled 10-dm³ pots were not watered for 2 months. Changes in chlorophyll fluorescence parameters and malondialdehyde content as leaf relative water content (RWC) decreased showed that ‘Chemlali’ was able to maintain functional and structural cell integrity longer than ‘Meski’. Mannitol started to accumulate later in the leaves of ‘Chemlali’ but reached higher levels than in the leaves of ‘Meski’. The latter accumulated several soluble sugars at lower dehydration. ‘Chemlali’ leaves also accumulated larger quantities of phenolic compounds which can improve its antioxidant response. Furthermore, the activity of three antioxidant enzymes catalase (CAT), peroxidase (POD) and ascorbate peroxidase (APX) increased as leaf RWC decreased. However, differences were observed between the two cultivars for CAT and POD but not for APX. The activity of the first two enzymes increased earlier in ‘Meski’, but reached higher levels in ‘Chemlali’. At low leaf hydration levels, ‘Chemlali’ leaves accumulated mannitol and phenolic compounds and had increased CAT and POD activities. These observations suggest that ‘Chemlali’ was more capable of maintaining its leaf cell integrity under severe water stress because of more efficient osmoprotection and antioxidation mechanisms.     

In-vitro regeneration of olive tree by somatic embryogenesis

We induced somatic embryogenesis from the cotyledon segments ofOlea europaea (L) cvs. ‘Chetoui’, ‘Chemleli’, and ‘Arbequina’. Calli were established from all three cultvars on OMc media supplemented with IBA and 2i-R The greatest success was obtained with media that contained zero or low concentrations of growth regulators. High levels of hormones (i.e.,>0.5 mgL^-1 IBA and 2i-P) inhibited embryogenesis. Embryos at different maturation stages were observed with continuously proliferating secondary embryogenesis. Abnormally shaped embryos and teratoma were also noted. Four weeks was the optimal incubation period for inducing embryogenesis on the auxin-containing medium. In addition, 30 to 40 gL^-1 sucrose was more effective than glucose in stimulating the growth and maturation of somatic embryos. Embryogeic efficiency was also higher when multivariate combinations of nitrogen sources (inorganic and organic nitrogen forms) were used. The plantlets that were derived from our germinating somatic embryos were similar to those obtained from axillary buds.     

Changes in gas exchange, proline accumulation and antioxidative enzyme activities in three olive cultivars under contrasting water availability regimes

Changes in photosynthetic performance, osmolyte accumulation and the activities of superoxide dismutase (SOD), ascorbate peroxidase (APX), catalase (CAT) and polyphenol oxidase (PPO) were investigated in one-year-old olive cultivars (Chemlali, Meski and Picholine) subjected to contrasting water availability regimes under arid climatic conditions in Tunisia. Shoot elongation rates (SER) and photosynthetic performance were markedly reduced by the water deficit regime (WD) in all cultivars except for Chemlali, which proved to be superior to the other two cultivars with respect to drought tolerance. Higher photosynthetic performance (net photosynthesis (P_n), stomatal conductance (g_s) and transpiration rates (E)) in the Chemlali and Meski cvs. compared to Picholine olive allowed them to maintain better plant water status and shoot elongation rates. Under WD conditions, Chemlali showed a greater capability for proline accumulation. Leaves grown under WD conditions showed signs of oxidative stress such as reduced chlorophyll and carotenoid concentrations. Nevertheless, different cultivars developed certain antioxidative defense mechanisms, including elevated SOD, APX and CAT activities. In contrast, PPO activity decreased under WD circumstances. Comparatively, Chemlali olive displayed better antioxidative enzyme activity, and thus better protection against oxidative stress. These results show that the ability of olive trees to up-regulate the enzymatic antioxidative system might be an important attribute linked to drought tolerance. These findings demonstrate that the association of higher P_n, proline accumulation and antioxidative defenses could be effective in a water-limited environment and may be useful selection criteria in breeding programs with the objective of improving drought tolerance and growth of olive trees, at least under the described environmental conditions.     

Ionic relations of aeroponically-grown olive genotypes,during salt stress

Two olive (Olea europaea L.) genotypes, Frantoio and Leccino, were exposed to increasing concentrations of NaCl (0-30-60-120 mM) in an aeroponic cultivation system for 60 days. Dry weights and sodium and potassium contents of apical and basal leaves, new and old wood, and roots were measured to determine Na uptake rate, Na translocation rate and K-Na selectivity ratio (S_K,Na). Frantoio showed a higher salt resistance than Leccino. Frantoio and Leccino had a similar Na uptake rate, but largely differed for Na translocation to the shoot. Furthermore Frantoio exhibited a higher K-Na selectivity than Leccino at both whole plant level and above all at the level of shoot system. Resistance mechanism of Frantoio is probably related to Na esclusion by roots and to the ability to maintain an appropriate K/Na ratio in actively growing tissues.Research supported by National Research Council of Italy, Special project RAISA.     

The effect of deficit irrigation on seasonal variations of plant water use in Olea europaea L.

A field experiment on olive trees (Olea europaea L.) was designed with the objective to search for an optimum irrigation scheduling by analyzing the possible effects of deficit irrigation. Treatments were: a non-irrigated control (rainfed) and three treatments that received seasonal water amount equivalent to 33 and 66% of crop evapotranspiration (ET_C) in the period August–September (respectively 33II and 66II), and 66% of (ET_C) from late May to early October (66I-II). Atmospheric evaporative demand and soil moisture conditions were regularly monitored. Irrigation effects on plant water relations were characterized throughout a growing season. Whole-plant water use, in deficit irrigated (66I-II) and rainfed olive trees, was determined using a xylem sap flow method (compensation heat-pulse technique). The magnitude of variations in water use and the seasonal dynamic of water relations varied among treatments, suggesting that olive trees were strongly responsive to both irrigation amount and time. Physiological parameters responded to variations in tree water status, soil moisture conditions and atmospheric evaporative demand. All measurements of tree water status were highly correlated with one another. There was a considerable degree of agreement between daily transpiration deduced from heat-pulse velocity and that determined by calibration using the water balance technique. Deficit irrigation during the whole summer (66I-II) resulted in improved plant water relations with respect to other watering regimes; while, severe regulated deficit irrigation differentiated only slightly 33II treatment from rainfed plants. Nevertheless, regulated deficit irrigation of olive trees after pit hardening (66II) could be recommended, at least in soil, cultivar and environmental conditions of this study.     

Stomatal and photosynthetic responses of olive (Olea europaea L.) leaves to water deficits

The leaf gas exchange of mature olive trees (Olea europaea L.) was characterized over a wide range of water deficits in the field during 1998, in Cordoba, Spain. Leaf photosynthesis (A) and stomatal conductance (g_l) responded diurnally and seasonally to variations in tree water status and evaporative demand. In the absence of water stress, A and g_l were generally high during autumn and low in days of high vapour pressure deficits (VPD). Leaf A varied between 0 and 2 µmol m^?2 s^?1 under severe water deficits that lowered the stem water potential (Ψ_x) to ?8·0 MPa, but recovered rapidly following rehydration. Transpiration efficiency (TE) was curvilinearly related to VPD and not influenced by water deficits except in cases of severe water stress, where low TE values were observed at Ψ_x below ?4 MPa. Three models of leaf conductance were calibrated and validated with the experimental data; two were based on the model proposed by Leuning (L) and the other was derived from the widely used Jarvis (J) model. The L models performed better than the J model in two validation tests. The scatter of the predictions and the limited accuracy of all three models suggest that, in addition to the physiological and environmental variables considered, there are additional endogenous factors influencing the g_l of olive leaves.     

Daily variations in water relations of apricot trees under different irrigation regimes

Mature apricot (Prunus armeniaca L. cv. Búlida) trees, growing under field conditions, were submitted to two drip irrigation treatments: a control (T1), irrigated to 100 % of seasonal crop evapotranspiration (ETc), and a continuous deficit (T2), irrigated to 50 % of the control throughout the year. The behaviour of leaf water potential and its components, leaf conductance and net photosynthesis were studied at three different times during the growing season, when they revealed a diurnal and seasonal pattern in response to water stress, evaporative demand of the atmosphere and leaf age. The deficit-irrigated trees showed, among other effects, a pronounced decrease in leaf water potential (ψ_w), decreased in leaf conductance (g_s) and no osmotic adjustment. For this reason, g_l and ψ_w can be considered good indicators of mature apricot tree water status and can therefore be used for irrigation scheduling.     

Comparative study of the discriminating capacity of RAPD,AFLP and SSR markers and of their effectiveness in establishing genetic relationships in olive

RAPDs, AFLPs and SSRs were compared in terms of their informativeness and efficiency in a study of genetic diversity and relationships among 32 olive cultivars cultivated in Italy and Spain. SSRs presented a higher level of polymorphism and a greater information content, as assessed by the expected heterozygosity, than AFLPs and RAPDs. The lowest values of expected heterozygosity were obtained for AFLPs, which, nevertheless were the most efficient marker system due to their capacity to reveal the highest number of bands per reaction and because of the high values achieved for a considerable number of indexes. All three techniques discriminated the genotypes very effectively, but only SSRs were able to discriminate the cultivars Frantoio and Cellina. The correlation coefficients of similarity were statistically significant for all three marker systems used but were lower for the SSR data than for RAPDs and AFLPs. For all markers a high similarity in dendrogram topologies was obtained although some differences were observed. All the dendrograms, including that obtained by the combined use of all the marker data, reflect some relationships for most of the cultivars according to their geographic diffusion. AMOVA analysis detected greater genetic differentiation among cultivars within each country than it did between the two countries.Communicated by H.F. Linskens     

Stem and leaf water potentials, gas exchange, sap flow, and trunk diameter fluctuations for detecting water stress in lemon trees

The sensitivity of continuous (on a whole-day basis) and discretely (at midday) measured indicators of the plant water status in adult lemon trees in response to a cycle of water deprivation and recovery, and the feasibility of obtaining baselines for tree water status indicators was investigated in 30-year-old Fino lemon trees (Citrus limon (L.) Burm. fil.) grafted on sour orange (C. aurantium L.) rootstocks. Control plants (T0) were irrigated daily above their crop water requirements in order to obtain non-limiting soil water conditions, while T1 plants were subjected to water stress by withholding irrigation for 50 days, after which time irrigation was restored and plant recovery was studied for 16 days. In T0 plants the water relations and the plant symptoms confirmed that they had not suffered waterlogging. In contrast, T1 plants showed a substantial degree of water stress, which developed very slowly. Maximum daily trunk shrinkage (MDS) increased in response to water stress during the first 15 days of the experiment, but when the stem water potential (Ψ_stem) fell below −1.8 MPa, the MDS signal intensity decreased. However, Ψ_stem and sap flow (SF) signal intensities progressively increased during the water stress period. The results showed that MDS is a very suitable plant-based indicator for precise irrigation scheduling in adult lemon trees. Reference or baseline relationships for MDS, Ψ_stem, and SF measurements as a function of several parameters related to the evaporative demand of the atmosphere were obtained. This fact open up the possibility of considering a plant-based indicator measurement at a given time relative to the expected value under non-limiting water conditions, which can be calculated from the reference relationships.     

Compensation heat-pulse measurements of sap flow for estimating transpiration in young lemon trees

Potted two-year-old lemon trees [Citrus limon (L.) Burm. f.], cv. Verna grafted on sour orange (C. aurantium L.) rootstock, growing in greenhouse, were subjected to drought for 33 d. Control plants were daily irrigated at field capacity. Values of sap flow (SF) were compared with transpiration (E) rates measured gravimetrically. The results underlined the robustness and high sensitivity of the compensation heat-pulse technique for estimating transpiration on a wide range of SF. Good direct correlations between E and SF rates on an instantaneous and daily basis were obtained in both treatments. On a daily basis, a common calibration curve can be used for both irrigation treatments. On an instantaneous basis, changes in SF were matches by similar changes in E in both treatments, although the relationships between these parameters presented different intercepts in each treatment. Sap flow rates were influenced by weather conditions in trees growing in non-limiting soil water conditions. This makes it possible to evaluate the significance of any sap flow measurement in relation to the reference value calculated for the vapour pressure deficit at the time the measurement was taken.This research was supported by Ministerio de Educacion y Ciencia (MEC), (CICYT/FEDER AGL2003-9387-C05-02 and AGL2004-0794-C03-02), and PETRI (PTR1995-0693-OP-02-01) grants to the authors. M.F. Ortuno was a recipient of a Program I3P research fellowship from CSIC.     

Observation of eight ancient olive trees (Olea europaea L.) growing in the Garden of Gethsemane

For thousands of years, olive trees (Olea europaea L.) have been a significant presence and a symbol in the Garden of Gethsemane, a place located at the foot of the Mount of Olives, Jerusalem, remembered for the agony of Jesus Christ before his arrest. This investigation comprises the first morphological and genetic characterization of eight olive trees in the Garden of Gethsemane. Pomological traits, morphometric, and ultrastructural observations as well as SSR (Simple Sequence Repeat) analysis were performed to identify the olive trees. Statistical analyses were conducted to evaluate their morphological variability. The study revealed a low morphological variability and minimal dissimilarity among the olive trees. According to molecular analysis, these trees showed the same allelic profile at all microsatellite loci analyzed. Combining the results of the different analyses carried out in the frame of the present work, we could conclude that the eight olive trees of the Gethsemane Garden have been propagated from a single genotype.     

Influence of carbon source and concentration on the in vitro development of olive zygotic embryos and explants raised from them

The influence of sucrose or mannitol on in vitro zygotic embryo germination, seedling development and explant propagation of olive tree (Olea europaea L.) was compared. Embryos germinated without sucrose in the medium but for adequate development of the seedlings to yield viable plants, a carbohydrate supply was necessary; both sucrose and mannitol were equally suitable for this purpose. However, when explants obtained from in vitro germinated embryos were cultured with mannitol or sucrose, then the polyalcohol promoted significantly more growth than sucrose by increasing shoot length, pairs of leaves formed, and breaking apical dominance. This improved the in vitro culture of olive plant material, thus allowing new olive clonal lines to be obtained in shorter times. This will assist in future breeding experiments with the species.     

Vegetative growth response of young olive trees (Olea europaea L., cv. Arbequina) to soil salinity and waterlogging

High-density olive orchards are increasing around the world, many of which may be potentially affected by salinity and waterlogging (hypoxia), two important stresses common in irrigated fields in arid and semi-arid climates. However, the response of olive to these stresses under field conditions is not well established. Therefore, our objective was to evaluate the vegetative growth response of young olive trees (Olea europaea L., cv. Arbequina) grown in a spatially variable waterlogged, saline-sodic field. We monitored the growth in trunk diameter of 341, 3-year-old olives between September 1999 and September 2000. Field contour maps were developed delineating soil salinity (ECa), relative ground elevation (RGE) and water table depth (WTD). Soil samples were also collected and analyzed for ECe and SARe in order to characterize the salinity and sodicity profiles and develop the ECa-ECe calibration equation. The infiltration rate (IR) of the crusted and uncrusted soil and the penetration resistance (PR) were also measured. The field was characterized by spatially variable ECe (2–15 dS m^–1), SARe (3–40), RGE (–4 to +4 cm) and WTD (0.5–1.9 m, with corresponding ground water EC values between 12 and 6 dS m^–1). Steady-state IR of crusted soil was only 7% of the uncrusted soil. Since the field was heavily irrigated by flooding, waterlogging conditions were related to low RGE values. Soil salinity was negatively correlated (R ² = 0.83, P<0.001) with RGE (ponded water) and WTD (upward flux), due to the evapo-concentration of water and salts at the soil surface. Thus, inverted salinity profiles developed in high salinity areas. Fifty-five percent of the olives were dead 3.5 years after planted, and most of them were located in areas of high ECe (> 10 dS m^–1), low RGE (< – 1.5 cm) and low WTD (< 1.2 m). The surviving trees had vegetative salinity tolerance values of ECe threshold = 4 dS m^–1 and slope = –12% (i.e., percent decline per unit increase in ECe above the treshold), indicating that the Arbequina olive is moderately tolerant to salinity. The RGE and WTD thresholds for olive's survival were > 0.1 cm and > 1.6 m, respectively. Thus, very small changes in ground elevation had a significant effect on olive's survival or death. The coupled effects of salinity and waterlogging (hypoxia) stresses were most detrimental for olive's growth.     

Enhancement of frost tolerance in olive shoots in vitro by cold acclimation and sucrose increase in the culture medium

The evaluation of frost tolerance in olive shoots in vitro has been successfully accomplished. The behavior of in vitro shoots at freezing temperatures was comparable to that of intact plants. Cold acclimation was found to increase frost tolerance in cv. Moraiolo and the LT₅₀ was about 4 °C lower compared to nonacclimated shoots. Damage in acclimated shoots occurred at –15 °C, whereas control shoots were damaged at –10 °C. Olive shoots were unable to withstand freezing temperatures of –20 °C, even when acclimated. The effects of sucrose were also determined. 6% (w/v) sucrose in the medium conferred the highest frost tolerance in both acclimated and nonacclimated plants.     

遮阴对骆驼刺叶性状和水分生理的影响

以塔克拉玛干沙漠南缘荒漠绿洲过渡带优势种骆驼刺为试验材料,研究正常光照(NL)、中度遮阴(MS)和重度遮阴(SS)3种不同遮阴环境对骆驼刺(Alhagi sparsifoliashap.)叶厚、单叶面积、比叶面积、比叶质量等叶性状参数和相对含水量、水势、失水速率、气孔导度、蒸腾速率以及水分利用效率等水分生理参数的影响。结果显示:随着光照强度的降低,骆驼刺的叶片厚度(LT)、比叶质量(LMA)、净光合速率(Pn)、气孔导度(Gs)、蒸腾速率(Tr)和水分利用效率(WUE)明显降低,而单叶面积(LA)、比叶面积(SLA)、相对含水量(RWC)、失水速率(RWL)和水势(WP)明显升高;各叶性状参数与水分生理参数之间均有极显著的相关性。说明:骆驼刺在遮阴环境下,其叶性状特征主要通过单叶面积和单叶干物质积累的变化来响应遮阴环境;其水分生理特征主要是牺牲对水分胁迫具有较好抗逆性的水分生理特征并通过气孔调节和更多的水分消耗用于维持一定的光合能力来响应遮阴环境。叶性状与水分生理参数相关性分析表明,遮阴环境下骆驼刺叶性状变化主要通过影响RWL、WP、Tr和WUE进而影响其水分生理特征的变化。因此,建议可利用遮阴措施对极端干旱区荒漠植物骆驼刺进行植被恢复,但其遮光度应设置在70%—80%自然光之间。     

A comparison of sap flux and water relations of leaves of various isolated trees with special reference to foundation movement in clay soil

Diurnal variation in sap flux (S) through stems of six trees, two each of Ulmus procera SALISB., Melaleuca styphelioides SM. and Prunus cerasifera EHRH. ‘Nigra’ (referred to hereafter by their generic names), were estimated from measurements of heat pulse velocities. Leaf water potential (ψ), stomatal conductance (g _s ) and transpiration from leaves (T) of all replicate trees were measured at 1300–1500h, once during the summer. On two separate occasions measurements were made of S, ψ, (g _s ) and T for one each of Ulmus and Melaleuca trees to study diurnal variations in these parameters. A 12×12 m² area around each tree was kept covered to simulate the condition of trees growing on pavements adjacent to residential properties. Sap flux for these tree species was in the order Melaleuca>Ulmus>Prunus. It is suggested that the smaller canopy and sapwood area in Prunus compared to the other two species is responsible for lower water potential and lower transpiration rate than the other species. Detailed analysis of the diurnal variation in sap flux and water relation of leaves of Melaleuca and Ulmus indicated sap flux of Melaleuca to be greater than that of Ulmus at the same transpiration rate per unit leaf area although the sapwood area of the two species was marginally different. This may have been due either to the difference in canopy conductance or in leaf area between the two species. With the assumption that sap flux closely resembles the rate of soil water extraction for both species, results indicate that Melaleuca is likely to extract soil water at a higher rate than Ulmus and hence is capable of causing greater shrinkage and soil movement than Ulmus.     

干旱胁迫下油橄榄品种光合特性研究

为揭示油橄榄(Olea europaea L.)耐旱性与光合特性之间的关系,以筛选出的适宜于半干旱川西南地区种植的7个引进油橄榄品种为供试材料,采用盆栽模拟干旱胁迫的方法,研究持续干旱胁迫对其光合特性的影响。结果表明:(1)随着干旱胁迫程度加剧,7个油橄榄品种叶片相对含水量均显著降低,至干旱胁迫后期(25d),各品种叶片均出现大幅失水,其中品种‘科拉蒂’失水率最高(45.79%),而品种‘小苹果’失水率最低(25.52%),说明‘小苹果’叶片在干旱胁迫下较其他油橄榄品种具有更高保水能力。(2)随着干旱胁迫程度加剧,7个油橄榄品种叶片光合色素含量均不同程度降低,表明光合色素分解量大于合成量;干旱胁迫持续25d时,品种‘豆果’的叶绿素a和叶绿素b含量下降幅度最大(P0.05),品种‘皮削利’类胡萝卜素含量下降幅度最大(P0.05),而品种‘小苹果’叶绿素a含量下降幅度最小。(3)随着干旱胁迫的持续进行,各油橄榄品种叶片净光合速率(Pn)、蒸腾速率(Tr)、气孔导度(Gs)和胞间CO2浓度(Ci)均不同程度降低,而水分利用效率(WUE)则呈上升趋势;干旱胁迫期间,品种‘佛奥’的Pn、Tr和Ci以及‘皮削利’的Gs降幅均高于其他品种,而‘小苹果’的Pn、Gs和Ci降幅均为最小且WUE上升幅度最大。研究发现,在持续干旱胁迫条件下,油橄榄幼苗叶片均大幅失水,光合色素结构被破坏、色素分解、含量降低,同时气孔关闭蒸发减少,光合作用减弱,而供试油橄榄品种中‘小苹果’对干旱胁迫的适应性最强,适宜于在半干旱的川西地区种植。     

Morphological and anatomical evaluation of adult and juvenile leaves of olive plants

The olive tree (Olea europaea L.), like many other woody plants, has a long juvenile period in which the plant is not able to produce flowers. Knowledge of the moment when the plant is capable of flowering is important for breeding programs and also for determining the physiological basis for sexual reproductive behavior, but currently the only indicator of that moment is the actual flowering. In many species, the juvenile-to-adult phase shift includes changes in leaf structure known as heteroblasty, that is, varied form of successive leaves on the same plant. Some differences have been observed between juvenile and adult olive leaves, particularly in size and form, but to our knowledge, no complete systematic study has been carried out. In this research, we measured size, morphology and anatomy for juvenile and adult leaves of olive plants grown from seeds. Differences were found in most of the parameters studied, including leaf size, form, mesophyll thickness, layers of palisade parenchyma and quantity of peltate trichomes, which were generally significant but overlapping between the two leaf types. The most consistent and striking difference was the presence of an organized layer of subepidermal cells only in the abaxial mesophyll of adult leaves. This characteristic could be a simple and effective criterion of phase change in the olive tree.     

Comparing image (fractal analysis) and electrochemical (impedance spectroscopy and electrolyte leakage) techniques for the assessment of the freezing tolerance in olive

Olive growth and productivity are limited by low temperatures mainly during winter, but sometimes also in spring and fall. The most effective way to avoid these damages in areas subjected to these climatic conditions is to select least susceptible varieties, but the choice of the right method to determine cold hardiness is extremely difficult. The aims of the work were (1) to assess LT₅₀ (lethal temperature at which 50% of damage in plants subjected to low temperatures occurs) of some olive varieties in two seasons (summer and winter) and (2) to assess the reliability of different methods to evaluate cold hardiness. LT₅₀ was determined on 21 different olive (Olea europaea L.) Italian varieties by leaf and shoot electrolyte leakage, shoot impedance spectroscopy and leaf color determination of fractal spectrum. All the experiments were conducted on non-acclimated and cold-acclimated plants. Our results showed that all the three methods were able to detect damages on olive plants after exposure to low temperatures, with leaves appearing more sensitive to cold stress than shoots. Among these methods, fractal analysis could be very useful in assessing cold hardiness of plants on the basis of visible injury, without sophisticated or expensive instruments and in a reliable and cost-effective way, using only a scanning device, a personal computer and dedicated freeware software.