Viability and integrity of Pinus densiflora seeds stored for 20 years at three different temperatures.

Storage temperature is one of the most important factors determining seed longevity in the genebank. This study aimed to investigate the effect of storage temperature on the seed viability and physiological integrity after a 20-year storage period of Pinus densiflora, a tree species of ecological and economic significance in South Korea. To this end, seeds were collected and stored dry for 20 years at -18°C, 4°C and 25°C. Germination tests were conducted to assess seed viability and vigour, electrolyte leakage analysis was performed to assess cell membrane integrity, and carbohydrate analysis was conducted to assess metabolic integrity during germination. The results revealed that over 20 years, seeds stored at -18°C maintained a high germination percentage (GP; 89%), comparable to initial GP (91%), whilst those stored at 4°C exhibited a decline in GP (44%) along with a decrease in vigour. Seeds stored at 25°C lost their viability entirely. Electrical conductivity of the leachate and leakage of inorganic compounds and soluble sugars were higher with elevated storage temperature, indicating increased imbibition damage. Additionally, changes in carbohydrate content during germination revealed that the loss of viability according to storage temperature is associated with reduced storage reserve utilization and altered carbohydrate metabolism during germination. These results enhance our understanding of the effect of seed storage temperature on longevity and physiological changes of aging in the genebank, serving as a reference for establishing conservation strategies for Pinus densiflora.

Canopy height affects the allocation of photosynthetic carbon and nitrogen in two deciduous tree species under elevated CO2.

Down-regulation of leaf N and Rubisco under elevated CO2 (eCO2) are accompanied by increased non-structural carbohydrates (NSC) due to the sink-source imbalance. Here, to investigate whether the canopy position affects the down-regulation of Rubisco, we measured leaf N, NSC and N allocation in two species with different heights at maturity [Fraxinus rhynchophylla (6.8 ± 0.3 m) and Sorbus alnifolia (3.6 ± 0.2 m)] from 2017 to 2019. Since 2009, both species were grown at three different CO2 concentrations in open-top chambers: ambient CO2 (400 ppm; aCO2); ambient CO2 × 1.4 (560 ppm; eCO21.4); and ambient CO2 × 1.8 (720 ppm; eCO21.8). Leaf N per unit mass (Nmass) decreased under eCO2, except under eCO21.8 in S. alnifolia and coincided with increased NSC. NSC increased under eCO2 in F. rhynchophylla, but the increment of NSC was greater in the upper canopy of S. alnifolia. Conversely, Rubisco content per unit area was reduced under eCO2 in S. alnifolia and there was no interaction between CO2 and canopy position. In contrast, the reduction of Rubisco content per unit area was greater in the upper canopy of F. rhynchophylla, with a significant interaction between CO2 and canopy position. Rubisco was negatively correlated with NSC only in the upper canopy of F. rhynchophylla, and at the same NSC, Rubisco was lower under eCO2 than under aCO2. Contrary to Rubisco, chlorophyll increased under eCO2 in both species, although there was no interaction between CO2 and canopy position. Finally, photosynthetic N content (Rubisco + chlorophyll + PSII) was reduced and consistent with down-regulation of Rubisco. Therefore, the observed Nmass reduction under eCO2 was associated with dilution due to NSC accumulation. Moreover, down-regulation of Rubisco under eCO2 was more sensitive to NSC accumulation in the upper canopy. Our findings emphasize the need for the modification of the canopy level model in the context of climate change.

Down-regulation of photosynthesis and its relationship with changes in leaf N allocation and N availability after long-term exposure to elevated CO2 concentration.

Down-regulation of photosynthesis under elevated CO2 (eCO2) concentrations could be attributed to the depletion of nitrogen (N) availability after long-term exposure to eCO2 (progressive nitrogen limitation, PNL) or leaf N dilutions due to excessive accumulation of nonstructural carbohydrates. To determine the mechanism underlying this down-regulation, we investigated N availability, photosynthetic characteristics, and N allocation in leaves of Pinus densiflora (shade-intolerant species, evergreen tree), Fraxinus rhynchophylla (intermediate shade-tolerant species, deciduous tree), and Sorbus alnifolia (shade-tolerant species, deciduous tree). The three species were grown under three different CO2 concentrations in open-top chambers, i.e., ambient 400 ppm (aCO2); ambient × 1.4, 560 ppm (eCO21.4); and ambient × 1.8, 720 ppm (eCO21.8), for 11 years. Unlike previous studies that addressed PNL, after 11 years of eCO2 exposure, N availability remained higher under eCO21.8, and chlorophyll and photosynthetic N use efficiency increased under eCO2. In the case of nonstructural carbohydrates, starch and soluble sugar showed significant increases under eCO2. The maximum carboxylation rate, leaf N per mass (Nmass), and ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) were low under eCO21.8. The ratio of RuBP regeneration to the carboxylation rate as well as that of chlorophyll N to Rubisco N increased with CO2 concentrations. Based on the reduction in Nmass (not in Narea) that was diluted by increase in nonstructural carbohydrate, down-regulation of photosynthesis was found to be caused by the dilution rather than PNL. The greatest increases in chlorophyll under eCO2 were observed in S. alnifolia, which was the most shade-tolerant species. This study could help provide more detailed, mechanistically based processes to explain the down-regulation of photosynthesis by considering two hypotheses together and showed N allocation seems to be flexible against changes in CO2 concentration.

Extended Low Temperature and Cryostorage Longevity of Salix Seeds with Desiccation Control.

Salix xerophila, S. maximowiczii, and S. koreensis are species of willow native to Korea that are important for bioenergy production. However, the native range of these species has narrowed in recent years due to the impact of climate change. Seeds of these Salix species lose viability within 4 weeks at ambient temperature, and within 4 months at -4°C. Preservation techniques are urgently needed to protect these valuable resources. The effects of seed water content (SWC; 3%, 6%, 9%, 12%, 18%, and 24%) and temperature (ambient, 4°C, -18°C, -80°C, and -196°C) on storage stability were investigated for up to 48, 52, or 60 months, depending on species. Optimal storage temperature and SWC varied between species. S. xerophila seed could be stored without deterioration for 60 months with 9% SWC at -80°C, but rapidly lost viability when stored at -18°C. In S. maximowiczii and S. koreensis, 100% and 90% of normal germination, respectively, was maintained with 18% SWC at -18°C or -80°C. Thus, for some Salix species, storage at -18 and -80°C may provide an economical alternative to cryopreservation or medium-term storage for the maintenance of seedbanks or breeding stocks.

Direct Effects on Seed Germination of 17 Tree Species Under Elevated Temperature and CO2 Conditions.

Effects on seed germination characteristics of 17 tree species were investigated under elevated temperature and CO2. Seeds of 5 needle-leaf and 12 broad-leaf species were germinated under four conditions: 24°C + 400 µmol CO2 mol air-1, 24°C + 750 µmol CO2 mol air-1, 27°C + 400 µmol CO2 mol air-1, and 27°C + 750 µmol CO2 mol air-1. The elevated temperature and CO2 affected germination percent (GP) of 7 tree species seeds.GPs of Pinus densiflora, P. thunbergii, Betula ermanii, and Maackia amurensisseeds were affected by the elevated temperature, while only that of P.jezoensis seed was influenced by the elevated CO2. GPs of Malus baccata and Zelkova serrataseeds were influenced by both the elevated temperature and CO2. In addition, the elevated temperature and CO2also affected mean germination time (MGT) of 12 tree species seeds. Particularly, MGTs of P. thunbergii and Rhododendron tschonoskii seeds were influenced by both factors. In conclusion, elevated temperature and CO2 affected seed germination characteristics, which were reflected by significant differences among tree species. Specifically, these two factors exerted stronger influence on germination pattern such as MGT rather than seed germination percent.

Expression of both CuZnSOD and APX in chloroplasts enhances tolerance to sulfur dioxide in transgenic sweet potato plants.

We have previously reported that transgenic sweet potato (Ipomoea batatas) plants overexpressing both CuZn superoxide dismutase (CuZnSOD) and ascorbate peroxidase (APX) under the control of a stress-inducible SWPA2 promoter in chloroplasts (referred to as SSA plants) showed increased resistance to methyl viologen-mediated oxidative stress and chilling. To investigate whether SSA plants show enhanced tolerance to air pollutants, they were exposed to 500ppb of sulfur dioxide (SO2). SO2 caused visible damage to the leaves of sweet potato, but damage in the leaves of non-transgenic (NT) plants was more severe than in those of SSA plants. The photosynthetic activity (Fv/Fm) of the SSA plants decreased by only 7% on the 5th day after the treatment, whereas that of NT plants severely decreased by 63% after 5days of recovery. Moreover, the chlorophyll content in the oldest leaf of NT plants decreased by 69%, whereas that of SSA plants remained at a high level. APX activity in NT plants increased about three times under an SO2 stress, and in SSA plants about five times compared to the case with no stress conditions. These results suggest that the overexpression of both CuZnSOD and APX in chloroplasts reduces the oxidative stress derived from SO2.

Three-dimensional surface topography of the needle stomatal complexes of Pinus rigida and its hybrid species by complementary microscopy.

Three-dimensional surface topography of needle stomatal complexes was investigated in Pinus rigida, Pinus taeda, and their interspecific hybrid Pinus rigitaeda. The stomatal complexes of P. rigida appeared to be sunken and ca. 15 microm deep by white light scanning interferometry. Stomatal grooves were evident in P. taeda along the stomata and amounted to ca. 5 microm deep. The centers of stomata maintained the similar height to the stomatal apertures. Meanwhile, the stomatal complexes of P. rigitaeda (ca. 15 microm deep) were characterized by distinct stomatal grooves and sunken stomatal chambers. In addition, field emission scanning electron microscopy revealed the stomatal complexes of P. rigida partially filled with epicuticular waxes. It was common to observe distinct stomatal grooves and chamber-filled stomata on P. taeda needles. The stomatal complexes of P. rigitaeda had the distinct stomatal grooves and were partially filled with wax tubules and rodlets. Surface roughness measurements of stomatal complexes showed higher levels of roughness from P. rigida and P. rigitaeda than that from P. taeda. These results indicate that the hybrid species P. rigitaeda showed intermediacy in surface characteristics between the parent species, suggesting the genetic control of needle stomatal complexes in the hybrid species.

A neutral cubane with a Zn(4)O(4) core: tetra-benzoato-tetra-kis(µ(3)-hydroxydi-2-pyridylmethano-lato)tetra-zinc(II)-acetone-methanol (1/2/1).

In the title compound, [Zn(4)(C(11)H(9)N(2)O(2))(4)(C(7)H(5)O(2))(4)]·2(CH(3))(2)CO·CH(3)OH, the tetra-nuclear mol-ecule lies on a fourfold inversion axis. Zn(II) ions and µ(3)-O atoms in the cubane core occupy alternating vertices, forming two inter-penetrating tetra-hedra. Each Zn(II) ion is further coordinated by two N atoms from two different (py)(2)C(OH)O ligands (py is pyrid-yl) and one O atom from a monodentate benzoate ligand, forming a distorted octa-hedral environment. The (py)(2)C(OH)O ligand acts in an η(1):η(3):η(1):µ(3) manner, forming two five-membered ZnNCCO chelating rings with two different Zn(II) atoms sharing a common C-O bond, and an alkoxide-type bond to a third Zn(II) ion. There are four symmetry-related intra-molecular O-H⋯O hydrogen bonds between the two types of ligands. In the asymmetric unit, there is a half-occupancy acetone solvent mol-ecule and a half-occupancy methanol solvent molecule that lies on a twofold rotation axis.

Differential expression of 10 sweetpotato peroxidases in response to sulfur dioxide, ozone, and ultraviolet radiation.

Secretory class III plant peroxidase (POD, EC 1.11.1.7) is believed to function in diverse physiological processes, including responses to various environmental stresses. To understand the function of each POD in terms of air pollutants and UV radiation, changes in POD activity and expression of 10 POD genes isolated from cell cultures of sweetpotato (Ipomoea batatas) were investigated in the leaves of sweetpotato after treatment with sulfur dioxide (SO(2) 500ppb, 8h/day for 5 days), ozone (O(3) 200ppb, 8h/day for 6 days), and ultraviolet radiation (UV-B 0.6mWm(-2) for 24h, UV-C 0.16mWm(-2) for 24h). All treatments significantly reduced the PSII photosynthetic efficiency (F(v)/F(m)). POD-specific activities (units/mg protein) were increased in leaves treated with SO(2) and O(3) by 5.2- and 7.1-fold, respectively, compared to control leaves. UV-B and UV-C also increased POD activities by 3.0- and 2.4-fold, respectively. As determined by RT-PCR analysis, 10 POD genes showed differential expression patterns upon treatment with air pollutants and UV radiation. Among the POD genes, swpa1, swpa2, and swpa4 were strongly induced following each of the treatments. Interestingly, basic POD genes (swpb1, swpb2, and swpb3) were highly expressed following SO(2) treatment only, whereas neutral swpn1 was highly induced following O(3) treatment only. These results indicated that some specific POD isoenzymes might be specifically involved in the defense mechanism against oxidative stress induced by air pollutants and UV radiation in sweetpotato plants.

Alleviation of Cd toxicity by composted sewage sludge in Cd-treated Schmidt birch (Betula schmidtii) seedlings.

We investigated alleviation of Cd toxicity and changes in the physiological characteristics of Betula schmidtii seedlings following application of composted sewage sludge to Cd-treated plants. Plants were grown under four test conditions: control, Cd treatment, sludge amendment, and Cd treatment with sludge amendment. B. schmidtii treated with Cd only accumulated the greatest amount of Cd in the leaves, but absorbed Cd was also highly concentrated in the roots. In contrast, Cd concentrations in the Cd and sludge amendment treated seedlings were the lowest in the roots. Since sludge amendment increased the growth of seedlings, it may have alleviated toxicity by dilution of Cd. Additionally, the absorbed Cd was more widely distributed since it was transported from the roots and accumulated in the stems and leaves of Cd and sludge treated plants. Cd treatment inhibited the growth and physiological functions of B. schmidtii seedlings, but sludge amendment compensated for these effects and improved growth and physiological functions in both Cd-treated and control plants. SOD activity in the leaves of seedlings was increased in the Cd-treated plants, but not in the Cd and sludge amendment treated seedlings. In conclusion, alleviation of Cd toxicity in response to sludge amendment may be related to a dilution effect, in which the Cd concentration in the tissues was effectively lowered by the improved growth performance of the seedlings.