Allergenicity and structural properties of new Cor a 1 isoallergens from hazel identified in different plant tissues.

The hazel allergen Cor a 1 is a PR-10 protein, closely related to the major birch pollen allergen Bet v 1. Hazel allergies are caused by cross-reactive IgE antibodies originally directed against Bet v 1. Despite the importance of PR-10 proteins in allergy development, their function and localization in the plant remain largely elusive. Therefore, the presence of Cor a 1 mRNA and proteins was investigated in different tissues, i.e., the female flower, immature and mature nuts, catkins, and pollen. Four yet unknown Cor a 1 isoallergens, i.e., Cor a 1.0501-1.0801, and one new Cor a 1.03 variant were discovered and characterized. Depending on the isoallergen, the occurrence and level of mRNA expression varied in different tissues, suggesting different functions. Interestingly, Cor a 1.04 previously thought to be only present in nuts, was also detected in catkins and pollen. The corresponding Cor a 1 genes were expressed in Escherichia coli. The purified proteins were analysed by CD and NMR spectroscopy. Immunoblots and ELISAs to determine their allergenic potential showed that the new proteins reacted positively with sera from patients allergic to birch, hazel and elder pollen and were recognized as novel isoallergens/variants by the WHO/IUIS Allergen Nomenclature Sub-Committee.

Comprehensive Transcriptome Analysis of Rare Carpinus putoensis Plants under NO2 stress.

We evaluated a transcriptome using high-throughput Illumina HiSeq sequencing and related it to the morphology, leaf anatomy, and physiological parameters of Carpinus putoensis putoensis under NO2 stress. The molecular mechanism of the C. putoensis NO2 stress response was evaluated using sequencing data. NO2 stress adversely affected the morphology, leaf anatomy, and total peroxidase (POD) activity. Through RNA-seq analysis, we used NCBI to compare the transcripts with nine databases and obtained their functional annotations. We annotated up to 2255 million clean Illumina paired-end RNA-seq reads, and 250,200 unigene sequences were assembled based on the resulting transcriptome data. More than 89% of the C. putoensis transcripts were functionally annotated. Under NO2 stress, 1119 genes were upregulated and 1240 were downregulated. According to the KEGG pathway and GO analyses, photosynthesis, chloroplasts, plastids, and the stimulus response are related to NO2 stress. Additionally, NO2 stress changed the expression of POD families, and the HPL2, HPL1, and POD genes exhibited high expression. The transcriptome analysis of C. putoensis leaves under NO2 stress supplies a reference for studying the molecular mechanism of C. putoensis resistance to NO2 stress. The given transcriptome data represent a valuable resource for studies on plant genes, which will contribute towards genome annotations during future genome projects.

Physiological and biochemical responses of two precious Carpinus species to high-concentration NO2 stress and their natural recovery.

Carpinus betulus and Carpinus putoensis are precious species in the world. Studies on the ecosystem function of the two species are rare. This study investigated the physiological and biochemical responses of C. betulus and C. putoensis to NO2 stress and their natural recovery. C. betulus and C. putoensis seedlings underwent fumigation with 12.0 mg/m3 NO2 for 0, 1, 6, 12, 24, 48, and 72 h, respectively. Then, the plants were allowed to recover at room temperature for 30 d. Physiological and biochemical changes in the leaves were compared between the two species. In terms of peroxidase (POD) activity, the damage response of C. betulus under NO2 stress appeared later than that of C. putoensis. The soluble protein content of C. betulus was noticeably higher than that of C. putoensis, and C. betulus exhibited more stable membrane lipoperoxidation. The tendency of the changes in nitrate reductase of C. betulus was less noticeable than that of C. putoensis. The variation amplitudes of N, K, Mg, Zn and Mn in the leaves of C. putoensis were greater than those of C. betulus. C. putoensis showed more sensitive metabolisms in response to NO2 stress compared with C. betulus. High-concentration NO2 caused damage to C. betulus and C. putoensis was reversible, and both species returned to normal growth via their own metabolism after 30-d recovery. The results of this study may provide useful reference data for quantitative assessment of the ecosystem function of C. betulus and C. putoensis and for their scientific application in urban greening.

Impact of the combination of sourdough fermentation and hop extract addition on baking properties, antioxidant capacity and phenolics bioaccessibility of rice bran-enhanced bread.

The effect of hop sourdough fermentation combined with rice bran (RB) addition was investigated for improving technological, bioactive and sensory properties of wheat bread by optimizing the quantities of RB, yeast, sourdough and hop extract contents based on response surface methodology. The bread produced according to the optimized formula was compared with control bread, sourdough or RB-enhanced bread with regard to quality attributes, bioaccessible phenolics and antioxidant activity. Sourdough fermentation combined with RB flour increased volume of loaves and consumer acceptance with no significant variations in crumb moisture content and crumb firmness during storage. RB darkened crumb while no effects on colour were observed for sourdough bread. Sourdough technology allowed increasing phenolic content of RB bread with no significant differences in its antioxidant activity. The bioaccessible ferulic and sinapic acids, the main phenolics released in vitro digestion of optimized bread were lower compared to those of control bread.

Leaf shedding increases the photosynthetic rate of the canopy in N2-fixing and non-N2-fixing woody species.

It has long been hypothesized that timing of leaf shedding is critical for plant fitness but there is little experimental evidence to support the hypothesis. According to an optimality theory, shedding of old leaves increases canopy photosynthesis despite some nitrogen (N) being lost as litterfall, when the ratio of daily photosynthesis to leaf N (N-use efficiency, ε) in old leaves, expressed as a fraction of ε in new leaves, becomes lower than the fraction of leaf N that is resorbed before shedding (RN). This was shown to be true for N-poor plants but not for N-rich plants in a pot experiment; however, the use of planting pots imposes a variety of physical, chemical and biological constraints that could change the experimental results. Here we conducted a 3-year field survey in a cool temperate deciduous forest to examine whether Alnus sieboldiana Matsum. (N2-fixing) and Carpinus tschonoskii Maxim. (non-N2-fixing) shed their leaves to increase canopy photosynthesis in accord with the above criterion. These species often grow sympatrically and were chosen as representatives of N-rich and N-poor plants, respectively. Overall, daily photosynthesis decreased with leaf age, accompanied by small changes in leaf N, resulting in a decrease in ε. In both species, ε of leaves at shedding expressed as a fraction of ε in new leaves was nearly equal to RN in all years, implying that the old leaves were shed to increase canopy photosynthesis. Our results, together with those of previous field surveys, suggested that the timing of leaf shedding is explained by N use in maximizing canopy photosynthesis across broad groups of species.

Taxanes content and cytotoxicity of hazel cells extract after elicitation with silver nanoparticles.

The toxicity of silver nanoparticles (AgNPs) has been attributed to the generation of Ag+ ions as well as production of ROS. The latter can elicit defensive response of plant cells in different ways e.g., enhancement of secondary metabolite productions. In the present study this hypothesis was evaluated by assessment of taxanes production by suspension-cultured hazel (Corylus avellana L.) cells after treatment with AgNPs. The cells were treated with different concentrations of AgNPs (0, 2.5, 5, and 10 ppm), in their logarithmic growth phase (d7) and were harvested after 1 weak. The growth of cells and their membrane integrity decreased but extracellular electro conductivity and total dissolved solids increase by AgNPs (probably due to loosening of cell membrane). Treatment of hazel cells with AgNPs (in particular of 5 ppm) rapidly and remarkably increased the yields of two major taxanes, i.e., Taxol and baccatin III; so that 24 h of the treatment their contents reached to 378% and 163% of the control, respectively. Increase of Taxanes was accompanied by the increase of total soluble phenols. The extracts of AgNPs-treated cells were able to inhibit the growth of cancerous HeLa cells and reduce their viability to 60% of the control. The results suggest the elicitation of suspension-cultured hazel cells with AgNPs as a procedure for rapid enhancement of anticancer taxanes biosynthesis by the cells.

Synthesis of siderophores by plant-associated metallotolerant bacteria under exposure to Cd(2.).

Rhizosphere and endophytic bacteria are well known producers of siderophores, organic compounds that chelate ferric iron (Fe(3+)), and therefore play an important role in plant growth promotion in metalliferous areas, thereby improving bioremediation processes. However, in addition to their primary function in iron mobilization, siderophores also have the capacity to chelate other heavy metals, such as Al(3+), Zn(2+), Cu(2+), Pb(2+) and Cd(2+), that can affect homeostasis and the heavy metal tolerance of microorganisms. The main goal of our study was to select the most efficient siderophore-producing bacterial strains isolated from the roots (endophytes) and rhizosphere of Betula pendula L. and Alnus glutinosa L. growing at two heavy metal contaminated sites in southern Poland. Siderophore biosynthesis of these strains in the presence of increasing concentrations of Cd(2+) (0, 0.5, 1, 2 and 3 mM) under iron-deficiency conditions was analysed using spectrophotometric chemical tests for hydroxamates, catecholates and phenolates, as well as the separation of bacterial siderophores by HPLC and characterization of their structure by UHPLC-QTOF/MS. We proved that (i) siderophore-producing bacterial strains seems to be more abundant in the rhizosphere (47%) than in root endophytes (18%); (ii) the strains most effective at siderophore synthesis belonged to the genus Streptomyces and were able to secrete three types of siderophores under Cd(2+) stress: hydroxamates, catecholates and phenolates; (iii) in general, the addition of Cd(2+) enhanced siderophore synthesis, particularly ferrioxamine B synthesis, which may indicate that siderophores play a significant role in tolerance to Cd(2+) in Streptomyces sp.

Coordination between growth, phenology and carbon storage in three coexisting deciduous tree species in a temperate forest.

In deciduous trees growing in temperate forests, bud break and growth in spring must rely on intrinsic carbon (C) reserves. Yet it is unclear whether growth and C storage occur simultaneously, and whether starch C in branches is sufficient for refoliation. To test in situ the relationships between growth, phenology and C utilization, we monitored stem growth, leaf phenology and stem and branch nonstructural carbohydrate (NSC) dynamics in three deciduous species: Carpinus betulus L., Fagus sylvatica L. and Quercus petraea (Matt.) Liebl. To quantify the role of NSC in C investment into growth, a C balance approach was applied. Across the three species, >95% of branchlet starch was consumed during bud break, confirming the importance of C reserves for refoliation in spring. The C balance calculation showed that 90% of the C investment in foliage (7.0-10.5 kg tree(-1) and 5-17 times the C needed for annual stem growth) was explained by simultaneous branchlet starch degradation. Carbon reserves were recovered sooner than expected, after leaf expansion, in parallel with stem growth. Carpinus had earlier leaf phenology (by ∼25 days) but delayed cambial growth (by ∼15 days) than Fagus and Quercus, the result of a competitive strategy to flush early, while having lower NSC levels.

Seasonal variation of pheophorbide a and flavonoid in different organs of two Carpinus species and its correlation with immunosuppressive activity.

The genus Carpinus of Betulaceae is the most widely distributed in the European landscape. This study reports a comparative study based on the pheophorbide a and flavonoid content from the two main species of the genus Carpinus, Carpinus betulus and Carpinus turczaninowii, respectively, in Nanjing, China. The pheophorbide a and flavonoid content depends on the organ, species, and season. HPLC analysis showed that the pheophorbide a and flavonoid levels were the highest in May and June, respectively, from the leaves of C. betulus 'Fastigiata.' In contrast, the content of pheophorbide a and flavonoid in the stems of C. betulus 'Fastigiata' or in other species was low. The immunosuppressive effects of the ethyl acetate extracts and methanol extracts from the two Carpinus species were also evaluated. The ethyl acetate extracts of C. betulus 'Fastigiata' in May and the methanol extracts of C. betulus 'Fastigiata' in June showed better immunosuppressive activity than in other seasons, which coincided with the content of pheophorbide a and flavonoid, respectively. Our findings indicated that C. betulus 'Fastigiata' can serve as a medicinal plant against inflammation because of its pheophorbide a and flavonoid content.

Fruit production in three masting tree species does not rely on stored carbon reserves.

Fruiting is typically considered to massively burden the seasonal carbon budget of trees. The cost of reproduction has therefore been suggested as a proximate factor explaining observed mast-fruiting patterns. Here, we used a large-scale, continuous (13)C labeling of mature, deciduous trees in a temperate Swiss forest to investigate to what extent fruit formation in three species with masting reproduction behavior (Carpinus betulus, Fagus sylvatica, Quercus petraea) relies on the import of stored carbon reserves. Using a free-air CO2 enrichment system, we exposed trees to (13)C-depleted CO2 during 8 consecutive years. By the end of this experiment, carbon reserve pools had significantly lower δ(13)C values compared to control trees. δ(13)C analysis of new biomass during the first season after termination of the CO2 enrichment allowed us to distinguish the sources of built-in carbon (old carbon reserves vs. current assimilates). Flowers and expanding leaves carried a significant (13)C label from old carbon stores. In contrast, fruits and vegetative infructescence tissues were exclusively produced from current, unlabeled photoassimilates in all three species, including F. sylvatica, which had a strong masting season. Analyses of δ(13)C in purified starch from xylem of fruit-bearing shoots revealed a complete turn-over of starch during the season, likely due to its usage for bud break. This study is the first to directly demonstrate that fruiting is independent from old carbon reserves in masting trees, with significant implications for mechanistic models that explain mast seeding.

Climate change effects on macrofaunal litter decomposition: the interplay of temperature, body masses and stoichiometry.

Macrofauna invertebrates of forest floors provide important functions in the decomposition process of soil organic matter, which is affected by the nutrient stoichiometry of the leaf litter. Climate change effects on forest ecosystems include warming and decreasing litter quality (e.g. higher C : nutrient ratios) induced by higher atmospheric CO(2) concentrations. While litter-bag experiments unravelled separate effects, a mechanistic understanding of how interactions between temperature and litter stoichiometry are driving decomposition rates is lacking. In a laboratory experiment, we filled this void by quantifying decomposer consumption rates analogous to predator-prey functional responses that include the mechanistic parameters handling time and attack rate. Systematically, we varied the body masses of isopods, the environmental temperature and the resource between poor (hornbeam) and good quality (ash). We found that attack rates increased and handling times decreased (i) with body masses and (ii) temperature. Interestingly, these relationships interacted with litter quality: small isopods possibly avoided the poorer resource, whereas large isopods exhibited increased, compensatory feeding of the poorer resource, which may be explained by their higher metabolic demands. The combination of metabolic theory and ecological stoichiometry provided critically important mechanistic insights into how warming and varying litter quality may modify macrofaunal decomposition rates.

Short-term dynamics of nonstructural carbohydrates and hemicelluloses in young branches of temperate forest trees during bud break.

Nonstructural carbohydrates (NSC) are the most important C reserves in the tissues of deciduous and evergreen tree species. Besides NSC, cell-wall hemicelluloses as the second most abundant polysaccharides in plants have often been discussed to serve as additional mobile carbon (C) reserves during periods of enhanced carbon-sink activities. To assess the significance of hemicelluloses as mobile carbon reserves, branches of two deciduous (Carpinus betulus L. and Fagus sylvatica L.) and two evergreen (Picea abies L. and Pinus sylvestris L.) tree species were sampled in a mature mixed forest stand in short intervals before and during bud break to assess NSC and hemicellulose concentrations in response to the increased carbon demand during bud break. Starch concentrations in branch sapwood of deciduous trees strongly decreased immediately before bud break and increased after bud break. In both evergreen species, only small changes of NSC were found in branch sapwood. However, 1-year-old needles exhibited a significant increase in starch concentration shortly before bud break which declined again after flushing. Hemicellulose concentrations (on an NSC-free dry matter basis) in branch sapwood of Carpinus decreased significantly shortly before bud break, but increased again after bud break. Contrarily, in Fagus branch sapwood, hemicellulose concentrations remained constant during bud break. Moderate increases of total hemicellulose concentrations before bud break were found in 1-year-old needles of both conifers, which could be explained by an accumulation of glucose units within the hemicellulose fraction. Overall, cell-wall hemicelluloses appeared to respond in a species-specific manner to the enhanced carbon demand during bud break. Hemicelluloses in branch sapwood of Carpinus and in 1-year-old needles of conifers likely act as additional carbon reserves similar to starch.

13C labelling reveals different contributions of photoassimilates from infructescences for fruiting in two temperate forest tree species.

The pathways of currently fixed carbon in fruit bearing branchlets were investigated in two temperate forest tree species (CARPINUS BETULUS and FAGUS SYLVATICA), which differ in texture of their vegetative infructescence tissues (leaf-like in CARPINUS vs. woody in FAGUS). During late spring, (13)C pulse-labelling was conducted on girdled, defoliated, girdled plus defoliated and untreated fruiting branchlets of mature trees IN SITU, to assess changes in C relations in response to the introduced C source-sink imbalances. At harvest in early August, 75 - 100 % of the recovered (13)C label was bound to infructescences (either fruits or vegetative infructescence tissue), revealing them as the prime C sinks for current photoassimilates. Leaves on girdled branchlets were not stronger labelled than on ungirdled ones in both species, indicating no upregulation of the leaves' photosynthetic capacity in response to the prevention of phloemic transport, which was also supported by measurements of light saturated photosynthesis. In contrast, (13)C labels tended to be higher after complete defoliation in the vegetative infructescence tissues of CARPINUS, suggesting enhanced net photosynthesis of green infructescence parts as compensation for the loss of regular leaves. The total labelling-derived (13)C content of whole infructescences was very similar between foliated and defoliated CARPINUS branchlets. Cupulae of FAGUS, on the other hand, remained almost unlabelled on defoliated branchlets, indicating the photosynthetic inactivity of this woody infructescence tissue. Consequently, CARPINUS still produced relatively high fruit masses on girdled plus defoliated branchlets, while in FAGUS fruit development ceased almost completely at this most severe treatment. Our results highlight that green vegetative infructescence tissue assimilates substantial amounts of C and can partly substitute regular leaves as C sources for successful fruit development.

Comparison of leaf life span, photosynthesis and defensive traits across seven species of deciduous broad-leaf tree seedlings.

BACKGROUND AND AIMS: Leaf life span, photosynthetic parameters and defensive traits were compared across seven species of deciduous broad-leaved tree seedlings native to northern Japan to test the "cost-benefit hypothesis" that more productive leaves are more susceptible to herbivore attack than less productive leaves. METHODS: Studies were made on three early successional species, Alnus hirsuta, Betula maximowicziana and Betula platyphylla "japonica"; one mid-successional species, Ostrya japonica, and three late-successional species, Carpinus cordata, Quercus mongolica 'grosseserrata' and Acer mono. Photosynthetic parameters and defensive traits (total phenolics, condensed tannin and toughness) of leaves were measured for each species, and a bioassay test with Eri silkmoth larvae (Samia cynthia ricini) was undertaken to evaluate differences between species in susceptibility to herbivore attack. KEY RESULTS: Early successional species have a shorter leaf life span (62-88 d) than late successional species (155-187 d). Leaf nitrogen content and light-saturated photosynthetic rate per unit leaf area (P(sat)-area) and per unit leaf mass (P(sat)-mass) were negatively correlated with leaf life span. The nitrogen content of early successional species was about 30 mg g(-1) and that of late successional species was about 16 mg g(-1). Leaf toughness and the C/N ratio were positively correlated with leaf life span, although condensed tannin was not correlated with leaf life span. The bioassay test showed that the number of days the larvae survived was negatively correlated with leaf life span. Average survival of larvae feeding on leaves of A. hirsuta, which has the shortest leaf life span, was 14.4 d and that of Q. mongolica, which has the longest leaf life span, was 6.6 d. The number of days of larval survival was positively correlated with leaf nitrogen content. There was no correlation between days of larval survival and defensive traits. CONCLUSIONS: These results indicate that species with a shorter leaf life span have higher photosynthetic productivity and are more susceptible to herbivore attack than species with a longer leaf life span. This supports the "cost-benefit hypothesis".

Impact of short-term and long-term elevated CO2 on emission of carbonyls from adult Quercus petraea and Carpinus betulus trees.

The study aimed to elucidate the effect of elevated CO2 in short- and long-term experiments on the emission potential of carbonyls from mature oak (Quercus petraea) and hornbeam (Carpinus betulus). The trees were investigated using a canopy crane established in a mixed forest in Switzerland. Short-term CO2 experiments were performed on single leaves with a gas exchange measuring unit by increasing CO2 in the enclosure before and during the measurements. Long-term CO2 fumigation was performed for one vegetation period by a webFACE design. Short-term as well as long-term exposure of leaves resulted in no significant changes in carbonyl exchange in neither Q. petraea nor C. betulus. Carbonyl emission was not affected by stomatal closure under elevated CO2 levels. In contrast to the emission of isoprenoids from vegetation which is thought to be reduced at elevated atmospheric CO2, the exchange of carbonyls seems to be generally unaffected under these conditions.

Rising atmospheric CO2 reduces sequestration of root-derived soil carbon.

Forests have a key role as carbon sinks, which could potentially mitigate the continuing increase in atmospheric carbon dioxide concentration and associated climate change. We show that carbon dioxide enrichment, although causing short-term growth stimulation in a range of European tree species, also leads to an increase in soil microbial respiration and a marked decline in sequestration of root-derived carbon in the soil. These findings indicate that, should similar processes operate in forest ecosystems, the size of the annual terrestrial carbon sink may be substantially reduced, resulting in a positive feedback on the rate of increase in atmospheric carbon dioxide concentration.

Carbon flux and growth in mature deciduous forest trees exposed to elevated CO2.

Whether rising atmospheric carbon dioxide (CO2) concentrations will cause forests to grow faster and store more carbon is an open question. Using free air CO2 release in combination with a canopy crane, we found an immediate and sustained enhancement of carbon flux through 35-meter-tall temperate forest trees when exposed to elevated CO2. However, there was no overall stimulation in stem growth and leaf litter production after 4 years. Photosynthetic capacity was not reduced, leaf chemistry changes were minor, and tree species differed in their responses. Although growing vigorously, these trees did not accrete more biomass carbon in stems in response to elevated CO2, thus challenging projections of growth responses derived from tests with smaller trees.

Responses of deciduous forest trees to severe drought in Central Europe.

In 2003, Central Europe experienced the warmest summer on record combined with unusually low precipitation. We studied plant water relations and phenology in a 100-year- old mixed deciduous forest on a slope (no ground water table) near Basel using the Swiss Canopy Crane (SCC). The drought lasted from early June to mid September. We studied five deciduous tree species; half of the individuals were exposed to elevated CO(2) concentration ([CO(2)]) (530 ppm) using a free-air, atmospheric CO(2)-enrichment system. In late July, after the first eight weeks of drought, mean predawn leaf water potential about 30 m above ground was -0.9 MPa across all trees, dropping to a mean of -1.5 MPa in mid-August when the top 1 m of the soil profile had no plant accessible moisture. Mean stomatal conductance and rates of maximum net photosynthesis decreased considerably in mid-August across all species. However, daily peak values of sap flow remained surprisingly constant over the whole period in Quercus petraea (Matt.) Liebl., and decreased to only about half of the early summer maxima in Fagus sylvatica L. and Carpinus betulus L. (stomatal down- regulation of flux). Although we detected no differences in most parameters between CO(2)-treated and control trees, predawn leaf water potential tended to be less negative in trees exposed to elevated [CO(2)]. Leaf longevity was greater in 2003 compared with the previous years, but the seasonal increase in stem basal area reached only about 75% of that in previous years. Our data suggest that the investigated tree species, particularly Q. petraea, did not experience severe water stress. However, an increased frequency of such exceptionally dry summers may have a more serious impact than a single event and would give Q. petraea a competitive advantage in the long run.

Differences in nitrogen economy of temperate trees.

Twenty-four temperate tree species were classified into three groups based on cluster analysis of relative growth rate, nitrogen concentration, nitrogen-production efficiency, nitrogen-distribution ratio and nitrogen-use efficiency as follows: Group I (Asteridae and Rosidae), Group II (Dilleniidae and Hamamelidae) and Group III (Coniferopsidae). Relative growth rate (RGR) was high in Group II, moderate in Group I and low in Group III. The regression coefficient for the relationship between RGR and leaf nitrogen concentration was higher in Group II than in Group I, and no relationship was observed in Group III. Parameter analysis of RGR indicated that RGR per unit leaf nitrogen was important for all three groups, but that the allocation of nitrogen to leaves was particularly important in Groups I and II. The ratio of dark respiratory rate (R) to net photosynthetic rate (A) was higher in Group I than in Group II. Neither A nor R was measured in the Group III species. A linear relationship was observed between leaf nitrogen concentration and A in Group II, but this relationship was not evident in Group I.