Comparison of Brain Transcriptome of the Greater Horseshoe Bats (Rhinolophus ferrumequinum) in Active and Torpid Episodes

Hibernation is an energy-saving strategy which is widely adopted by heterothermic mammals to survive in the harsh environment. The greater horseshoe bat (Rhinolophus ferrumequinum) can hibernate for a long period in the hibernation season. However, the global gene expression changes between hibernation and non-hibernation season in the greater horseshoe bat remain largely unknown. We herein reported a comprehensive survey of differential gene expression in the brain between winter hibernating and summer active greater horseshoe bats using next-generation sequencing technology. A total of 90,314,174 reads were generated and we identified 1,573 differentially expressed genes between active and torpid states. Interestingly, we found that differentially expressed genes are over-represented in some GO categories (such as metabolic suppression, cellular stress responses and oxidative stress), which suggests neuroprotective strategies might play an important role in hibernation control mechanisms. Our results determined to what extent the brain tissue of the greater horseshoe bats differ in gene expression between summer active and winter hibernating states and provided comprehensive insights into the adaptive mechanisms of bat hibernation.     

Life expectancy of spruce needles under extremely high air pollution stress: performance of trees in the Ore Mountains

Summary Photosynthetic performance of Norway spruce needles [Picea abies (L.) Karst.] was measured over a 1-year period. The trees grew in an area of heavy air pollution and forest decline on a mountain ridge in the eastern Ore Mountains (Czech Republic). Photosynthetic capacity, as well as light use efficiency, decreased dramatically with time, starting in July (2 months after bud-break) to finally reach zero (respiration only) by February of the following year. Two months later all needles from upper crown parts were shed. Needles from lower crown parts, on the other hand, were undamaged. The chlorophyll and Mg content decreased transiently during the cold season, with Mg reaching deficiency thresholds during winter. However, total sulfur, as well as organic and sulfate S increased with time. The increase was higher in needles from the upper parts of the crown, which were exposed to windy air throughout the year, than in the lower parts of the crown, which were covered by grass during summer and by snow during most of the winter.     

Interactions between green spruce aphid (Elatobium abietinum (Walker)) and Norway and Sitka spruce under high and low nutrient conditions

1 Green spruce aphid (Elatobium abietinum) is a serious pest of spruce (Picea spp.) in north‐west Europe, causing defoliation of one‐year‐old and older needles. 2 Relationships between population development of E. abietinum, needle loss and tree growth were compared for five pure genotypes of Sitka spruce and mixed‐genotype material of Sitka and Norway spruce, grown under high and low nutrient conditions. 3 Despite wide differences in flushing date between spruce genotypes, E. abietinum populations peaked on the same date on each genotype and on the mixed‐genotype material, irrespective of nutrient supply. 4 Larger aphid populations developed on trees grown under high nutrient conditions than under low nutrients. However, more needles were lost per aphid in the low nutrient treatment and overall defoliation rates in the two nutrient treatments were similar. 5 Total aphid numbers differed significantly between Sitka spruce genotypes within nutrient treatments, but not in relation to bud‐burst or needle terpene content. Reductions in height growth caused by infestation were greater (15–44%), and related to mean aphid densities and defoliation, in the low nutrient treatment, but were smaller (11–27%) and not related to aphid density and defoliation in the high nutrient treatment. 6 Development of E. abietinum populations was similar on Norway and Sitka spruce, but Norway spruce lost fewer needles. However, the effects of infestation on tree growth were more closely related to aphid density and were similar for Norway and Sitka spruce. 7 Infestation caused a decrease in total root dry weight of Norway and Sitka spruce in proportion to the reductions observed in above‐ground growth.     

Seasonal variation in the monoterpenes in needles of Picea abies (L.) Karst.

Summary This investigation was conducted to obtain information about the fluctuations in composition and amount of needle monoterpenes during the development of spruce needles. Studies conducted with two Norway spruce clones clearly revealed the existence of fluctuations. In juvenile needles, the amounts of the oxygenated terpenes increase constantly with age during the first 2 months of needle growth. The hydrocarbon terpenes dominate within the first weeks, some of them even showing a very distinct first maximum within the first 3 weeks after bud burst. All terpenes, including the oxygenated ones, have a maximum in June/July, which favours the hypothesis of a substitution of the hydrocarbons later on. There are significant changes even in mature needles of Norway spruce. The terpene level of 1-year-old needles of the clonal trees increased from spring to early summer and then dropped again towards winter. In addition, fluctuations in mature needles were shown for a set of ten wild trees. Needles of the same age class, which emerged in 1986, were sampled 4 times from 1986 to 1988. The needle terpene concentrations of the 1 -year-old needles were considerably lower in spring at the time of bud burst than in autumn. The terpene level of older needles thus seems to be influenced by biosynthetic and catabolic activities.     

Seasonal changes in the distribution of green spruce aphid Elatobium abietinum (Walker) (Homoptera: Aphididae) in the canopy of Sitka spruce

1 Seasonal changes in the distribution of green spruce aphid Elatobium abietinum (Walker) within the canopy of 20–25‐year‐old Sitka spruce are described based on data from two low‐altitude sites (310–420 m above sea‐level), two mid‐altitude sites (500–550 m a.s.l) and one high‐altitude site (610 m a.s.l). 2 Aphids were counted throughout the canopy on shoots representative of all needle age‐classes present at each whorl of branches. Counts were made during the middle week of each month from September to July for 4 years (1999–2003), and mean E. abietinum densities at each canopy position were calculated separately for each month and for the low‐, mid‐ and high‐altitude sites. 3 During September to November, the highest densities of E. abietinum occurred on 3–4‐year‐old needles on branches low in the canopy. Over the winter and spring, the centre of the aphid’s distribution shifted outward and upward, so that by June of the next year the highest aphid densities occurred on current and 1‐year‐old needles on branches near the top of the tree. 4 The aphid distribution was re‐set each year during July, at the time when the nutrient quality of the host was in decline and E. abietinum populations were decreasing. Aphid densities decreased less on 3–4‐year‐old needles than on current and 1‐year‐old needles, suggesting that older needles were a superior food resource at this time of year and in the autumn. However, other factors, such as higher temperatures in the upper canopy during the summer or differential mortality caused by natural enemies, could also have contributed to the change in distribution. 5 The outward and upward shift in the aphid distribution over the winter period provided no evidence that aphids at positions lower and deeper in the canopy were better insulated from freezing temperatures and had higher over‐winter survival rates. Mean air temperatures at the top and bottom of the canopy during the winter were also found to differ by only 0.1–0.2 °C. 6 The percentage of the total aphids per tree that occurred on current or 1‐year‐old needles varied widely between seasons and between sites. Consequently, sampling programmes designed to estimate total population numbers of E. abietinum have little option but to sample needles throughout the canopy, and at regular intervals during the period when the aphid is abundant.     

Changes in Sulfur Metabolism During Needle Development of Norway Spruce*

In a field-study with approx. 150-year-old spruce trees, seasonal changes in thiol composition and content, sulfate content, and oxtractable activities of enzymes of sulfate reduction and assimilation were analyzed in needles and buds and were related to developmental processes during flushing. GSH was the predominant thiol in spruce needles throughout the year, with maximum contents of more than 400nmol g FW^?1 during winter and minimum contents of less than 200 nmol g FW^?1 during summer. Negative correlation of changes in GSI1 contents during winter and spring with air temperature showed a low correlation coefficient (r =– 0.55), suggesting minor significance of GSH in frost protection. Drastic changes in thiol contents during flushing suggested that GSH accumulated in the previous year's needles during winter supports the growing needles with reduced sulfur. Sulfate and other substrates for sulfate assimilation appeared to be available in buds and the new needles. An external supply with reduced sulfur may still be required because of insufficient activities of enzymes of sulfate reduction and assimilation, esp. ATP-sulfurylase and APS-sulfotransferase.     

The effect of needle age on the acceptability of Sitka spruce needles to the aphid,Elatobium abietinum (Walker)

Summary A comparison of the feeding behaviour of E. abietinum on current year needles and previous year needles of P. sitchensis revealed that during the summer months the aphids preferentially settled on previous year needles, this response not being evident in winter. Aphids on current year needles in summer took a much longer period of time to commence sap uptake than in winter, intake ceasing following a very short feeding period. On previous year needles sap uptake in December commenced after a shorter feeding period than in June. Analysis of total and soluble nitrogen levels in Sitka spruce needles showed that current year needles had initially higher levels during shoot elongation in May and early June, but that previous year needles had higher levels for most of the remainder of the year. Quantitative analyses of amino acids revealed that in current year needles the levels were generally lower than in previous year needles. Less marked proportional differences were observed between previous year needles in May and in July/August when the needles were unsuitable. Addition of amino acids in solution into cut current year shoots resulted in increased longevity on shoots containing introduced iso-leucine, histidine and methionine and revealed a general imbalance of the amino acids.     

Winter at the alpine timberline. Why does embolism occur in norway spruce but not in stone pine?

Conifers growing at the alpine timberline are exposed to frost drought and freeze-thaw cycles during winter-stress factors known to induce embolism in tree xylem. The two dominant species of the European Central Alps timberline were studied: Norway spruce (Picea abies [L.] Karst) and stone pine (Pinus cembra), which usually reaches higher altitudes. We hypothesized to find embolism only at the timberline and to observe less embolism in stone pine than in Norway spruce due to avoidance mechanisms. Seasonal courses of embolism and water potential were studied at 1,700 and 2,100 m during two winter seasons and correlated to vulnerability (to drought-induced embolism), leaf conductance, and micrometeorological data. Embolism was observed only at the timberline and only in Norway spruce (up to 49.2% loss of conductivity). Conductivity losses corresponded to low water potentials (down to -3.5 MPa) but also to the number of freeze-thaw events indicating both stress factors to contribute to embolism induction. Decreasing embolism rates-probably due to refilling-were observed already in winter. Stone pine did not exhibit an adapted vulnerability (50% loss of conductivity at -3.5 MPa) but avoided critical potentials (minimum -2.3 MPa): Cuticulare conductance was 3.5-fold lower than in Norway spruce, and angles between needles and axes were found to decrease in dehydrating branches. The extent of conductivity losses in Norway spruce and the spectrum of avoidance and recovery mechanisms in both species indicates winter embolism to be relevant for tree line formation.     

Effects of prolonged drought on the anatomy of sun and shade needles in young Norway spruce trees

Predicted increases in the frequency and duration of drought are expected to negatively affect tree vitality, but we know little about how water shortage will influence needle anatomy and thereby the trees’ photosynthetic and hydraulic capacity. In this study, we evaluated anatomical changes in sun and shade needles of 20‐year‐old Norway spruce trees exposed to artificial drought stress. Canopy position was found to be important for needle structure, as sun needles had significantly higher values than shade needles for all anatomical traits (i.e., cross‐sectional needle area, number of tracheids in needle, needle hydraulic conductivity, and tracheid lumen area), except proportion of xylem area per cross‐sectional needle area. In sun needles, drought reduced all trait values by 10–40%, whereas in shade needles, only tracheid maximum diameter was reduced by drought. Due to the relatively weaker response of shade needles than sun needles in drought‐stressed trees, the difference between the two needle types was reduced by 25% in the drought‐stressed trees compared to the control trees. The observed changes in needle anatomy provide new understanding of how Norway spruce adapts to drought stress and may improve predictions of how forests will respond to global climate change.     

Enhanced ethylene emissions from red and norway spruce exposed to acidic mists

Acidic cloudwater is believed to cause needle injury and to decrease winter hardiness in conifers. During simulations of these adverse conditions, rates of ethylene emissions from and levels of 1-aminocyclopropane-1-carboxylic acid (ACC) in both red and Norway spruce needles increased as a result of treatment with acidic mists but amounts of 1-malonyl(amino)cyclopropane-1-carboxylic acid remained unchanged. However, release of significant quantities of ethylene by another mechanism independent of ACC was also detected from brown needles. Application of exogenous plant growth regulators such as auxin, kinetin, abscisic acid and gibberellic acid (each 0.1 millimolar) had no obvious effects on the rates of basal or stress ethylene production from Norway spruce needles. The kinetics of ethylene formation by acidic mist-stressed needles suggest that there is no active inhibitive mechanism in spruce to prevent stress ethylene being released once ACC has been formed.     

Damage caused by the Green Spruce Aphid to Norway and Sitka spruce needles

Three-month-old needles of Sitka spruce were less susceptible to Elatobium abietinutn than 15-month-old needles. Symptoms appeared after longer aphid feeding times but only a proportion of damaged needles fell. After short feeding periods symptoms appeared in more Norway spruce needles than in Sitka spruce, whereas longer feeding periods resulted in more needles producing symptoms in the Sitka spruce. The symptoms took 4—6 days longer to appear in Norway spruce, and needle fall followed a longer feeding period than on Sitka spruce. Following 72 h feeding, needle fall occurred more quickly on Sitka spruce than on Norway spruce. The time taken for needle fall to occur was inversely related to the feeding time in Sitka spruce but such a response was not evident in Norway spruce. The results are discussed in relation to the differences exhibited in the probing behaviour of the aphid on the two spruces.     

The effect of previously infested spruce needles on the growth of the green spruce aphid, Elatobium abietinum, and the effect of the aphid on the amino acid balance of the host plant

The effect of chlorosis induced in needles of Sitka and Norway spruce by the green spruce aphid on growth of the aphid is investigated, and the effect of infestation of the aphid on amino acid levels in Sitka spruce foliage is reported. On both Sitka and Norway spruce green spruce aphids were heavier when reared on chlorotic (previously infested) needles than when reared on green (previously uninfested) needles. The effect was more pronounced on Sitka than on Norway spruce. Infestation of the aphid altered the amino acid balance of Sitka spruce foliage but not the concentration of total amino acids. Possible causes of chlorosis, the influence of individual amino acids on aphid growth, the potential effect of chlorosis on outbreaks of the aphid and the differences in susceptibility of Sitka and Norway spruce to damage by the aphid are discussed.     

Genetic and environmental contributions to the winter hardiness of conifers

Summary

Both genetic and environmental components are involved in the processes of winter hardening and dehardening which permit needles of conifers like Norway spruce to survive very low temperatures (< - 30°C) over winter and to recover fully in time for the following and subsequent seasons. One of the major environmental effects of increasing concentrations of atmospheric ozone (O₃) in recent summers has been to affect detrimentally the ability of conifer needles to harden properly and at the correct rate the following autumn. Part of the mechanism by which this occurs in the cytoplasm of needle cells has been traced to detrimental effects on both the Δ¹² fatty acid desaturase and the unusual Δ⁵ desaturase which appears to be part of the low temperature survival mechanism of conifers. The genetic component of winter hardening also involves needle lipids. Studies of lipids in the needles of Norway spruce trees of different provenances growing for many years in the UK have shown that they change during winter hardening as though they were still on trees in the original sites throughout Europe from which the seeds of these trees were initially collected.     

The effect of stand age on throughfall chemistry in spruce stands in the Potok Dupniañski catchment in the Silesian Beskid Mountains, southern Poland

The chemical composition of throughfall depends on the age of the Norway spruce (Picea abies Karst) stands and season of the year. The pH of throughfall decreased and the amount of hydrogen ion in throughfall deposited to the soil increased with increasing age of spruce stands, especially in the winter season. Concentrations of K+, H+, SO4(2-), Mn2+, and NH4(+) in throughfall were higher than bulk precipitation for the whole year and K+, H+, and Mn2+ concentrations were higher in throughfall in winter and the growing season. This indicates that these ions were washed out or washed from the surface of needles and/or the bark, and that NO3(-), NH4(+), Ca2+, Mg2+, Fe2+, and Zn2+ were absorbed in the canopy. The effect of high nitrogen deposition, above critical loads, and an increase in the amount of sulfur and in the sum of the strong acids (S-SO4(2-) and N-NO3(-)) that reached the soil with throughfall may have implications for the vitality of spruce stands, especially in older age classes. The application of Principal Component Analysis (PCA) has led to identification of five factors responsible for the data structure ("mineral dust", "acidic emissions", "heavy metals-dust particles", "ammonium [NH4(+)]", and "H+"). They explain more than 60% of the total variance system. The strong positive correlation between stand age class and ionic concentrations in throughfall occurs for all year and the winter period for ions within the following categories: "acidic emissions", SO4(2-) + NO3(-); "heavy metals-dust particles", Fe2+ + Mn2+ + Zn2+; "mineral dust", Na+ + K+ + Ca2+ + Mg2+; "NH4(+)"; and "H+". The strength of the relationship decreases in the growing period, probably due to processes occurring in the canopy (adsorption, leaching, etc.).     

Regulation of photochemical efficiency of photosystem 2 in Norway spruce at the beginning of winter and in the following spring

Based on the analysis of fluorescence quenching, the nonphotochemical dissipative processes were investigated in Norway spruce needles during acclimation to winter and spring conditions. The maximum nonphotochemical fluorescence quenching (q_Nmax) was reached at lower irradiances in winter (up to 310 μmol m-2 s-1) than in spring (about 1130 μmol m-2 s-1), but its values were nearly the same (q_Nmax = 0.91±0.01) during both winter and spring measurements. In early winter the pronounced initial fluorescence quenching (q0) suggested that nonradiative energy dissipation in the antennae complexes dominated. Significantly lower q0 (by 40-60 % compared to winter needles) during acclimation of needles to spring conditions supported a significant contribution of quenching in the reaction centres. These findings support the hypothesis that the antennae systems and reaction centres cooperate in the protective dissipation of excess excitation energy. This revised version was published online in September 2006 with corrections to the Cover Date.     

Inhibition of photosynthesis in current-year needles of unfertilized and fertilized Norway spruce [Picea abies (L.) Karst.] during autumn and early winter

Inhibition of photosynthesis was followed during autumn and early winter in current-year sun and shade needles of unfertilized and fertilized Norway spruce [Picea abies (L.) Karst.] by simultaneous measurements of photosynthetic O₂ evolution and chlorophyll a fluorescence at 20 °C. The CO₂-saturated rate of O₂ evolution was generally higher in sun needles of fertilized trees than in those of unfertilized trees over a wide range of incident photon flux densities (PFDs). Furthermore, the maximum photo-chemical efficiency of photosystem (PS) II, as indicated by the ratio of variable to maximum fluorescence (F_V/F_M) was generally higher for sun needles of fertilized trees. The depression of f_v/f_m during frost periods was more pronounced in sun needles than in shade needles, indicating that winter inhibition in Norway spruce is strongly light-dependent. However, the inhibition of the rate of O₂ evolution at high PFDs in needles of fertilized trees during early winter was partly independent of the light regime experienced by those needles in the field, which appeared to result in a pronounced decrease in the proportion of oxidized PS II reaction centres in shade needles. A nearly identical linear relationship between the quantum yield of PS II electron transport determined by chlorophyll fluorescence and the quantum yield of O₂ evolution (gross rate of O₂ evolution/PFD) was obtained for the investigated types of needles during autumn and early winter. Except for shade needles of fertilized trees, this appeared to be largely achieved by adjustments in thermal energy dissipation within PS II.