Cell wall-bound ultraviolet-screening compounds explain the high ultraviolet tolerance of the Antarctic moss, Ceratodon purpureus

* Studies of ultraviolet (UV) light-induced DNA damage in three Antarctic moss species have shown Ceratodon purpureus to be the most UV tolerant, despite containing lower concentrations of methanol-soluble UV-screening compounds than the co-occurring Bryum pseudotriquetrum. * In this study, alkali extraction of cell wall-bound phenolics, combined with methanol extraction of soluble phenolics, was used to determine whether cell wall-bound UV screens explain the greater UV tolerance of C. purpureus. * The combined pool of UV screens was similar in B. pseudotriquetrum and C. purpureus, but whilst B. pseudotriquetrum had almost equal concentrations of MeOH-soluble and alkali-extractable cell wall-bound UV-screening compounds, in C. purpureus the concentration of cell wall-bound screening compounds was six times higher than the concentration of MeOH-soluble UV screens. The Antarctic endemic Schistidium antarctici possessed half the combined pool of UV screens of the other species but, as in C. purpureus, these were predominantly cell wall bound. Confocal microscopy confirmed the localization of UV screens in each species. * Greater investment in cell wall-bound UV screens offers C. purpureus a more spatially uniform, and potentially more effective, UV screen. Schistidium antarctici has the lowest UV-screening potential, indicating that this species may be disadvantaged under continuing springtime ozone depletion. Cell wall compounds have not previously been quantified in bryophytes but may be an important component of the UV defences of lower plants.     

冻融作用对冻土区微生物生理和生态的影响

不同时空尺度的冻融过程导致冻土温度变化及水的相变和迁移,改变着微生物生境的物理和化学性质。冻融过程可改变细胞内外渗透压平衡,且冰晶生长过程能损伤细胞膜和细胞器。限于营养、氧气等其它极端不利条件,不少细胞逐渐转入休眠状态以度过难关;微生物DNA、蛋白质的合成和能量代谢仅用于维持细胞生存。冻融作用通过改变细胞代谢模式而影响微生物参与的寒区碳氮元素的生物地球化学循环。多年冻土层保存了不同地质时期微生物种群的多样性,作为物理和生物地球化学屏障,可有效削弱地表过程和地壳本底辐射对微生物的影响。在长期的适应过程中,微生物发展了相应的耐受机制,从结构和功能方面,细胞和分子水平上应对冻土环境和冻融过程。这可为寻找地外寒冷星球上可能的生命形式提供一些线索。     

Surface reflectance properties of Antarctic moss and their relationship to plant species, pigment composition and photosynthetic function

In this study the variations in surface reflectance properties and pigment concentrations of Antarctic moss over species, sites, microtopography and with water content were investigated. It was found that species had significantly different surface reflectance properties, particularly in the region of the red edge (approximately 700 nm), but this did not correlate strongly with pigment concentrations. Surface reflectance of moss also varied in the visible region and in the characteristics of the red edge over different sites. Reflectance parameters, such as the photochemical reflectance index (PRI) and cold hard band were useful discriminators of site, microtopographic position and water content. The PRI was correlated both with the concentrations of active xanthophyll‐cycle pigments and the photosynthetic light use efficiency, F_v/F_m, measured using chlorophyll fluorescence. Water content of moss strongly influenced the amplitude and position of the red‐edge as well as the PRI, and may be responsible for observed differences in reflectance properties for different species and sites. All moss showed sustained high levels of photoprotective xanthophyll pigments, especially at exposed sites, indicating moss is experiencing continual high levels of photochemical stress.     

Impact of changes in natural ultraviolet radiation on pigment composition, physiological and morphological characteristics of the Antarctic moss, Grimmia antarctici

The impact of ambient ultraviolet (UV)‐B radiation on the endemic bryophyte, Grimmia antarctici, was studied over 14 months in East Antarctica. Over recent decades, Antarctic plants have been exposed to the largest relative increase in UV‐B exposure as a result of ozone depletion. We investigated the effect of reduced UV and visible radiation on the pigment concentrations, surface reflectance and physiological and morphological parameters of this moss. Plexiglass screens were used to provide both reduced UV levels (77%) and a 50% decrease in total radiation. The screen combinations were used to separate UV photoprotective from visible photoprotective strategies, because these bryophytes are growing in relatively high light environments compared with many mosses. G. antarctici was affected negatively by ambient levels of UV radiation. Chlorophyll content was significantly lower in plants grown under near‐ambient UV, while the relative proportions of photoprotective carotenoids, especially β‐carotene and zeaxanthin, increased. However, no evidence for the accumulation of UV‐B‐absorbing pigments in response to UV radiation was observed. Although photosynthetic rates were not affected, there was evidence of UV effects on morphology. Plants that were shaded showed fewer treatment responses and these were similar to the natural variation observed between moss growing on exposed microtopographical ridges and in more sheltered valleys within the turf. Given that other Antarctic bryophytes possess UV‐B‐absorbing pigments which should offer better protection under ambient UV‐B radiation, these findings suggest that G. antarctici may be disadvantaged in some settings under a climate with continuing high levels of springtime UV‐B radiation.     

Effect of freezing and thawing rates on denaturation of proteins in aqueous solutions

The freeze denaturation of model proteins, LDH, ADH, and catalase, was investigated in absence of cryoprotectants using a microcryostage under well-controlled freezing and thawing rates. Most of the experimental data were obtained from a study using a dilute solution with an enzyme concentration of 0.025 g/l. The dependence of activity recovery of proteins on the freezing and thawing rates showed a reciprocal and independent effect, that is, slow freezing (at a freezing rate about 1 degrees C/min) and fast thawing (at a thawing rate >10 degrees C/min) produced higher activity recovery, whereas fast freezing with slow thawing resulted in more severe damage to proteins. With minimizing the freezing concentration and pH change of buffer solution by using a potassium phosphate buffer, this phenomenon could be ascribed to surface-induced denaturation during freezing and thawing process. Upon the fast freezing (e.g., when the freezing rate >20 degrees C/min), small ice crystals and a relatively large surface area of ice-liquid interface are formed, which increases the exposure of protein molecules to the ice-liquid interface and hence increases the damage to the proteins. During thawing, additional damage to proteins is caused by recrystallization process. Recrystallization exerts additional interfacial tension or shear on the entrapped proteins and hence causes additional damage to the latter. When buffer solutes participated during freezing, the activity recovery of proteins after freezing and thawing decreased due to the change of buffer solution pH during freezing. However, the patterns of the dependence on freezing and thawing rates of activity recovery did not change except for that at extreme low freezing rates (<0.5 degrees C/min). The results exhibited that the freezing damage of protein in aqueous solutions could be reduced by changing the buffer type and composition and by optimizing the freezing-thawing protocol.     

Effect of rapid freezing and thawing on cellular integrity of honey bee sperm

Abstract. Direct observations on the effect of rapid freezing and thawing on honey bee ( Apis mellifera L.) sperm were made by light, scanning and transmission electron microscopy. Rapid freezing of honey bee ejaculated sperm, suspended in freezing diluent, in liquid nitrogen followed by rapid thawing can cause cellular injuries which lead to the death of the sperm. The frozen-thawed sperm, supravitally stained, showed a significant decrease in cell viability compared with that of the control fresh sperm ( P <0.001). Significant uptake of the stain in the dead sperm resulted from damage in the cell membrane. The scanning electron micrographs of frozen-thawed sperm further demonstrated that the injury of cell membrane can lead to the splitting of mitochondrial derivatives from the flagellar axoneme. More cellular injuries including the release of acrosomal content and membrane damage at the acrosome, nucleus and the tail regions were further revealed by transmission electron microscopy. The impact of cellular injuries on the quality of honey bee sperm cryopreserved for artificial insemination of honey bee queens is discussed.     

Improved and reproducible cell viability in the superflash freezing method using an automatic thawing apparatus

Cell cryopreservation stops the biological activity of cells by placing them in the frozen state, and can be used to preserve cells without subculturing, which can cause contamination and genetic drift. However, the freezing process used in cryopreservation can injure or damage the cells due to the cytotoxicity of cryoprotecting agents (CPAs). We have previously reported a CPA-free cryopreservation method based on inkjet technology. In this method, the vitrified cells were exposed to the room temperature atmosphere during the transport of the cells using tweezers, which caused devitrification due to the increased temperature and often lowered the cell viability. In the present study, we developed an automatic thawing apparatus that transports the vitrified cells rapidly into a prewarmed medium using a spring hinge. Observations with a high-speed camera revealed that the spring hinge drops the cells into the prewarmed medium within 20 ms. All heat-transfer simulations for the apparatuses with different designs and rotation speeds showed that the cells remained below the glass-transition temperature during the transport. Finally, the apparatus was evaluated using mouse fibroblast 3T3 cells. The cell viability was improved and its reproducibility was enhanced using this apparatus. The results indicate that the combination of superflash freezing with the rapid thawing process represents a promising approach to circumvent the problems typically associated with the addition of CPAs.     

Accumulation of DNA damage in Antarctic mosses: correlations with ultraviolet-B radiation, temperature and turf water content vary among species

The susceptibility of three East Antarctic moss species to UV-B radiation was examined by measuring accumulation of cyclobutane pyrimidine dimers under natural sunlight during the austral summer season of 2002/03. The 2002/03 season was characterized by unusually low springtime ozone depletion and as such our results likely underestimate the DNA damage possible in a more typical UV-B radiation season. Despite this all three species accumulated significant DNA photoproducts. We also found a positive association between photoproduct accumulation and incident UV-B radiation in the two cosmopolitan species, Bryum pseudotriquetrum and Ceratodon purpureus , with more DNA damage in samples collected early in the season compared with later in the summer. For B. pseudotriquetrum , negative associations were also observed between photoproduct accumulation and both turf water content and the 10-day mean air temperature. Photoproduct accumulation in the endemic species Schistidium antarctici was similarly high across the season and no significant association with environmental variables was found. Our results are consistent with the two cosmopolitan species having somewhat higher UV-B-screening capabilities and possibly more efficient mechanisms for repairing DNA damage than the endemic S. antarctici .     

Photoinhibition at chilling temperatures and effects of freezing stress on cold acclimated spinach leaves in the field. A fluorescence study

The role of high light stress in a natural environment was studied on spinach plants ( Spinacia oleracea L. cv. Wolter) grown in the field during the winter season. Fluorescence induction (at 293 K and 77 K) of leaves was used to characterize the stress effects. Night frost with minimum temperatures between – 1.5°C and –7.5°C (i.e. above the'frost killing point'at ca. –11.5°C) led to impaired photosynthesis. This was seen as increased initial fluorescence (F_o), decreased ratio of variable to maximum fluorescence (F_V/F_M) and lowered rates of O₂ evolution. The freezing injury was reversible within several frostless days. Exposure to high light (about 900 mol m_–2 s_–1) at chilling temperatures in the field caused photoinhibition, manifested as decreased variable fluorescence (F_V) and F_V/F_M ratio without changes in F_O. The photoinhibitory fluorescence quenching was not stronger after frost than after frostless nights; synergism between light stress and preceding freezing stress was not observed. Fluorescence induction signals at 77 K showed that F_V of photosystems I and II decreased to the same extent, indicating increased thermal deactivation of excited chlorophyll. Photoinhibition was fully reversible at +4°C within 1 h in low light, but only partially in moderate light. Preceding night frosts did not affect the recovery. The photoinhibition observed here is regarded as a protective system of thermal dissipation of excess light energy.     

Establishment of a novel method for cryopreservation and thawing of the mouse ovary

During cryopreservation of ovarian tissue, the conditions of freezing and thawing are big factors controlling the survival rate of oocytes obtained. However, the conditions and procedures as they pertain to ovarian follicles and oocytes have not been established. Thus, we tried to determine the appropriate freeze-thaw times using the vitrification method with ethylene glycol and DMSO as cryoprotective agents and dd Y female mouse ovaries. The maturity rate from GV to the metaphase-II (MII) stage was 62.8% with ethylene glycol and 69.3% using DMSO, while the controls (GV oocytes obtained from a fresh ovary) showed a maturation rate of 83.6% (46/55). MII oocytes obtained by culturing GV oocytes in vitro showed a 64.3% (18/28) fertility rate via in vitro fertilization and a developmental rate into a 2 cell stage embryo of 35.7% (10/28) and into a 4-cell stage, 7.1% (2/28). However, development beyond the 8 cell stage embryo was not observed. A significant difference was not recognized between control (fresh) and ovarian tissues that had been frozen/thawed with respect to their ability to produce hormones. It is concluded that the vitrification method was effective for both freezing ovarian tissues and preserving its functional ability (maturation and capacitation).     

The barriers to oceanic island radiation in bryophytes: insights from the phylogeography of the moss Grimmia montana

Aim In contrast to angiosperms, bryophytes do not appear to have radiated in Macaronesia and the western Mediterranean. We evaluate if: (1) the apparent lack of radiation in bryophytes reflects our failure to recognize cryptic endemic species; (2) bryophytes are characterized by extremely low evolutionary rates; or (3) bryophytes have a high dispersal ability, which prevents genetic isolation. Location Worldwide, with a special emphasis on Macaronesia and the western Mediterranean. Methods Three chloroplast regions were sequenced from samples of the moss Grimmia montana from its entire distribution range. Network analyses, Fst and Nst statistics were used to describe and interpret the phylogeographical signal in the data. Results Despite significant phylogeographical signal in the chloroplast genome, which demonstrates limits to gene flow at the continental scale, repeated sister group relationships observed among accessions from different geographical areas suggest recurrent colonization patterns. These observations are consistent with mounting evidence that intercontinental distributions exhibited by many bryophyte species result from long‐distance dispersal rather than continental drift. Madeiran and western Mediterranean island haplotypes are either shared by, or closely related to, European and North American ones. Fst values between Madeira, western Mediterranean islands, North America and Europe are not significantly different from zero, and suggest that Madeira and the south‐western Mediterranean are subject to strong transatlantic gene flow. By contrast, haplotypes found in the Canary Islands are shared or closely related to those of populations from south‐western Europe or southern Africa. Main conclusions Multiple origins and colonization events are not consistent with the hypothesis of a relictual origin of the Macaronesian moss flora. One possible reason for the failure of taxa that experienced multiple colonization events to radiate is niche pre‐emption. We suggest that strong gene flow, coupled with the occupancy of all suitable niches, either by earlier conspecific colonizers or by other species, could be the mechanism preventing island radiation in G. montana and other cryptogams with high long‐distance dispersal abilities.     

Methodological aspects of moss sample preparation. Effects of freezing and duration of washing on the cellular distribution of elements in Fontinalis squamosa Hedw.

Aquatic bryophytes are widely used in biomonitoring studies, mainly because of their capacity to act as bioaccumulators. As different methods can be used to preserve and process moss samples, the results of elemental analyses may also vary, particularly if the contents of different cellular compartments are analyzed separately. In the present study, we compared the total concentrations of some elements and the concentrations of these in different cellular locations in frozen-thawed and fresh samples of the aquatic moss Fontinalis squamosa that were also washed for different lengths of time before analysis. We used the sequential elution technique (SET) to extract the different fractions, and we determined the concentrations of K, Mg, Ca, Cd, Co, Cu, Al and Zn in the extracts. The results varied depending on the element, cellular location and moss sampling site. In the moss samples processed after freezing, the greatest differences were in the intracellular concentrations of K, Mg and Cd, which were lower than in the fresh samples. Minor differences were found in the concentrations of elements in other cellular locations and in the total concentrations of elements. The increase in the duration of the initial washing step, carried out to remove soluble and particulate intercellular elements, may also cause the release of elements (e.g. K and Mg) bound to extracellular exchange sites. The concentrations in the other cellular fractions and the total concentrations were less affected by the washing duration. Neither freezing nor the use of long washing times are recommended for processing moss samples prior to extraction of elements by the SET. Further studies are needed to confirm and clarify the observations.     

毛尖紫萼藓(Grimmia pilifera P.Beauv) PSII光化学效率对脱水和复水的响应

利用快速叶绿素荧光动力学技术研究了毛尖紫萼藓脱水和复水过程中叶绿素荧光变化,结果显示在脱水过程中毛尖紫萼藓PSⅡ的最大光化学效率（Fv/Fm）、光合机构电子传递的量子产额（ETo／ABS）、捕获的激子将电子传递到电子传递链中超过QA的其它电子受体的概率（ETo／TRo）、单位叶面积的反应中心的数量（RC／CSo）以及PSⅡ受体库（Area）对叶片含水量的响应等均存在相对含水量阈值。在阈值范围内脱水,对以上荧光参数影响不大,低于阈值后,各荧光参数值迅速下降,直至PSⅡ反应中心完全关闭以及光化学过程结束。再复水后,毛尖紫萼藓光合机构的最大捕光效率、实际光化学效率、PSⅡ反应中心受体侧的电子传递链以及反应中心均能得到快速而有效的恢复。表明一定时间内脱水不会对毛尖紫萼藓的光合器官造成严重伤害,光合系统仍维持在可恢复状态。     

Supercooling points of insects and mites on the Antarctic Peninsula

Abstract. 1. Mean supercooling points of eleven species of arthropods (three Collembola, seven Acari and one Diptera) ranged from -6.2 to -9.4°C (high group), and from -17.7 to -31.0°C (low group). The majority of individuals in the high group had food in their gut systems.
2. Freezing was lethal to all species examined except larval Belgica antarctica Jacobs (Chironomidae).
3. Glucose, glycerol and mannitol were found in low concentrations in extracts of Ctyptopygus antarcticus Willem (Collembola) and larvae of B. antarcrica; it is udikely that these substances had a major effect on the supercooling of either species.
4. Two Collembola species possessed significantly ( P< 0.05) lower supercooling points at locations on the Antarctic Pensinsula than at Signy Island, South Orkney Islands. The converse was observed for two species of Acari.
5. It is suggested that whilst gross climatic and also micro-habitat conditions may influence the cold hardiness of such arthropods, especially seasonally, their full supercooling ability is rarely tested.     

Effect of thawing time, cooling rate and boron nutrition on freezing point of the primordial shoot in norway spruce buds

BACKGROUND: Effects of cooling rates on bud frost hardiness have been studied but there is little information on bud responses to thawing. Since the cell wall pore size has been found to increase with boron (B) deficiency, B deficiency may affect the supercooling ability of buds in winter. METHODS: The effects of duration of thawing time and rate of cooling on bud frost hardiness of Norway spruce (Picea abies) were studied in a B fertilization trial in February 2003 and March 2005. Frost hardiness of apical buds was determined by differential thermal analysis (DTA) and visual scoring of damage. KEY RESULTS: In 2003, the freezing point of primordial shoots of buds (T(f)), i.e. the low-temperature exotherm (LTE), was, on average, -39 degrees C when buds were thawed for less than 3 h and the T(f) increased to -21 degrees C after 18 h of thawing. During the first 4 h of thawing, the rate of dehardening was 6 degrees C h(-1). In 2005, buds dehardened linearly from -39 degrees C to -35 degrees C at a rate of 0.7 degrees C h(-1). In 2003, different cooling rates of 1-5 degrees C h(-1) had a minor effect on T(f) but in 2005 with slow cooling rates T(f) decreased. In both samplings, at cooling rates of 2 and 1 degrees C h(-1), T(f) was slightly higher in B-fertilized than in non-fertilized trees. By contrast, at very short thawing times in 2003, T(f) was somewhat lower in B-fertilized trees. CONCLUSIONS: There was little evidence of reduced frost hardiness in trees with low B status. This study showed that buds deharden rapidly when exposed to above-freezing temperatures in winter, but if cooled again they reharden more slowly. According to this study, rapid dehardening of buds has to be taken into account in assessments of frost hardiness.     

基于GIS和MaxEnt比较中国砂藓属与紫萼藓属植物地理分布

基于19个生物气候因子和紫萼藓属(Grimmia) 172个、砂藓属(Racomitrium) 181个国内分布记录,应用MaxEnt模型和ArcGis 9.3软件,定量预测了紫萼藓属与砂藓属植物在“属”水平上在我国各省区的生境适应性特点.预测结果表明,紫萼藓属植物在浙江(0.7099,综合生境适宜性指数,下同)、江苏(0.6212)、北京(0.5987)、天津(0.5648)、云南(0.5532)、辽宁(0.5515)、台湾(0.5422)、安徽(0.5398)和吉林(0.4945)有较高的气候适应性,而砂藓属植物在浙江(0.889)、上海(0.6564)、香港(0.5897)、台湾(0.5858)、贵州(0.5354)、湖北(0.5039)、云南(0.4885)、重庆(0.4871)、江苏(0.4793)和安徽(0.4552)具有较高的生境适宜性.对砂藓属与紫萼藓属的分布预测范围进行比较发现,砂藓属植物在香港、重庆、贵州、广东、广西、湖北、海南和台湾的生境适应性比紫萼藓属的总体上要高,而在江西、福建和湖南,两属的分布概率比较接近,其余省区则紫萼藓属植物的生境适宜性比砂藓的要高.紫萼藓属植物属于典型的温带性种类,其分布主要在高海拨的寒冷地段.     

Phase Transition of Thylakoid Membranes Modulates Photoinhibition in the Cyanobacterium Anabaena siamensis

A shift of the growth temperature from 40 degrees C to 18 degrees C promoted an increase in the degree of fatty acids unsaturation and a decrease, from 26 degrees C to 0 degrees C, of the phase transition temperature of thylakoid membranes in Anabaena siamensis. The pattern of photoinhibition of photosynthesis at distinct temperatures varied as a function of the phase transition temperature. In the absence of streptomycin, a pronounced photoinhibition at temperatures near the phase transition (26 degrees C) was observed in cells grown at 40 degrees C, while protection from photodamage was observed at chilling temperatures (15 degrees C to 5 degrees C). In this same range of temperature, such a protection was not verified if cells were grown at 18 degrees C. In both types of cells, however, the rate of photoinactivation in the presence of streptomycin was progressively decreased by lowering the temperature of photoinhibition. When recovery from photoinhibition was followed at the respective temperature in which cells were grown, the restoration profile of the photosynthetic O(2) evolution to initial levels was essentially the same in both types of cells. The protective effect of low temperatures against photoinhibition was attributed to a decreased solubility and diffusion of oxygen in the thylakoid membranes due to an increase of the membrane viscosity that would avoid the photogeneration of reactive oxygen species around PS II.     

下辽河平原典型农田融化期氧化亚氮和甲烷排放通量研究

应用静态箱／气相色谱法对下辽河平原典型农田（大豆地、玉米地、水稻田）土壤融化期N2O和CH4排放通量进行了研究。结果表明,在融化期间,3种农田N20排放量均较大,这段时期的农田是大气N2O的一个重要源;3种农田CH4排放不明显,成为大气CH4的汇。在融化期间,农田N2O排放量,旱田CH4排放量与箱内温度间均无显著相关性。而水稻田CH4排放量与箱内温度呈显著负相关,相对于生长季来说,这是土壤融化期间的特定现象。     

Historical ozone concentrations and flavonoid levels in herbarium specimens of the Antarctic moss Bryum argenteum

Depletion of stratospheric ozone since the mid 1970s has led to significant increases in ultraviolet B (UVB) irradiation over Antarctica. The detrimental effects of UVB on plants are many, but plants produce photoprotective flavonoids that reduce cellular damage. We used herbarium samples of the moss Bryum argenteum collected in Antarctica to compare the levels of flavone aglycones in plants collected before and after the formation of the ozone hole. The interpretation of historical data is difficult, because environmental conditions immediately before sample collection are unknown. Factors such as cloud cover can have a significant influence on UVB dose at ground level, modifying the flavonoid content of the specimen and adding considerable variability to the results. Nevertheless, our results revealed significant relationships between total flavone concentration and pooled year classes ( P = 0.001). Furthermore, regression analysis showed a significant negative relationship of total flavone concentration and the level of ozone immediately before the time of collection ( P = 0.016). In addition, the ratio of luteolin (an ortho -dihydroxylated flavone) to apigenin (a monohydroxylated flavone) increased significantly with several environmental parameters. These included (a) increasing modelled midday UVB radiation ( P = 0.002), (b) increasing modelled midday UVB/PAR ratio ( P < 0.001), and (c) decreasing ozone concentration ( P < 0.001). We emphasise the utility of this ratio in interpreting the historical ozone trends rather than relying on changes in total flavone concentrations alone. These results illustrate that herbarium specimens may reveal historical levels of UVB radiation.