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Lichens and phototolerant poikilohydric mosses differ from spinach leaves, fern fronds or photosensitive mosses in that they show strongly decreased Fo chlorophyll fluorescence after drying. This desiccation-induced fluorescence loss is rapidly reversible under rehydration. Fluorescence emission from Photosystem II at 685 nm was decreased more strongly by dehydration than 720 nm emission. Reaction centers of Photosystem II lose activity on dehydration and regain it on hydration. Heating of desiccated lichens increased Fo chlorophyll fluorescence. The activation energy for the reversible part of the temperature-dependent fluorescence increase was 0.045 eV, which corresponds to the energy difference between the 680 and 697 nm absorption bands. In desiccated chlorolichens such as Parmelia sulcata, heating induces the appearance of positive variable fluorescence related to the reversible reduction of QA due to overcoming the energy barrier. This is interpreted to provide information on the mechanism of photoprotection: energy is dissipated by changing Chl680 or P680 into a chlorophyll form, which absorbs at 700 nm and emits light at 720 nm (Chl-720 or P680(700)) with a low quantum yield. Dissipation of light energy in this trap is activated by desiccation.

Plant species capable of hyper-accumulating heavy metals are of considerable interest for phytoremediation, and differ in their ability to accumulate metals from the environment. This work aims to examine (i) arsenic accumulation in three fern species [Chinese brake fern (Pteris vittata L.), slender brake fern (Pteris ensiformis Burm. f.), and Boston fern (Nephrolepis exaltata L.)], which were exposed to 0, 150, or 300 muM of arsenic (Na(2)HAsO(4).7H(2)O), and (ii) the role of anti-oxidative metabolism in arsenic tolerance in these fern species. Arsenic accumulation increased with an increase in arsenic concentration in the growth medium, the most being found in P. vittata fronds showing no toxicity symptoms. In addition, accumulation was highest in the fronds, followed by the rhizome, and finally the roots, in all three fern species. Thiobarbituric acid-reacting substances, indicators of stress in plants, were found to be lowest in P. vittata, which corresponds with its observed tolerance to arsenic. All three ferns responded differentially to arsenic exposure in terms of anti-oxidative defence. Higher levels of superoxide dismutase, catalase, and ascorbate peroxidase were observed in P. vittata than in P. ensiformis and N. exaltata, showing their active involvement in the arsenic detoxification mechanism. However, no significant increase was observed in either guaiacol peroxides or glutathione reductase in arsenic-treated P. vittata. Higher activity of anti-oxidative enzymes and lower thiobarbituric acid-reacting substances in arsenic-treated P. vittata correspond with its arsenic hyper-accumulation and no symptoms of toxicity.

The fern Athyrium yokoscense is known to be highly tolerant to lead toxicity, and is a lead hyperaccumulator that can accumulate over 1,000 microg g(-1) of lead in its dry matter. In this work, we examined whether the gametophytic generation of A. yokoscense also resists lead toxicity like the sporophytic generation. Spore germination in A. yokoscense was more tolerant to Pb2+, compared to that in other fern species, such as Pteridium aquilinum, Lygodium japonicum and Pteris vittata. In addition, the early gametophyte development of A. yokoscense was not much affected by 10 microM Pb2+, as evaluated from the prothallial growth and rhizoid development. We also showed that Athyrium gametophytes could accumulate more than 10,000 microg g(-1) of lead, and that the lead was localized in the cytosol and vacuole of rhizoidal cells, as determined by a transmission electron micrograph. These results indicate that Athyrium gametophytes have the ability to accumulate lead in the rhizoids. Furthermore, the gametophytes were found to include a large amount of proanthocyanidins (condensed tannins). Because proanthocyanidins have a latent ability to complex with lead ions, the possible roles of proanthocyanidins in the lead tolerance and accumulation of Athyrium gametophytes are discussed.

Environmental pollution with metals and xenobiotics is a global problem, and the development of phytoremediation technologies for the plant-based clean-up of contaminated soils is therefore of significant interest. Phytoremediation technologies are currently available for only a small subset of pollution problems, such as arsenic. Arsenic removal employs naturally selected hyperaccumulator ferns, which accumulate very high concentrations of arsenic specifically in above-ground tissues. Elegant two-gene transgenic approaches have been designed for the development of mercury or arsenic phytoremediation technologies. In a plant that naturally hyperaccumulates zinc in leaves, approximately ten key metal homeostasis genes are expressed at very high levels. This outlines the extent of change in gene activities needed in the engineering of transgenic plants for soil clean-up. Further analysis and discovery of genes for phytoremediation will benefit from the recent development of segregating populations for a genetic analysis of naturally selected metal hyperaccumulation in plants, and from comprehensive ionomics data--multi-element concentration profiles from a large number of Arabidopsis mutants.

There has been an interest in using hyperaccumulating plants for the removal of heavy metals and metalloids. High selenium (Se) concentrations in the environment are detrimental to animals, humans, and sustainable agriculture, yet selenium is also an essential nutrient for humans. This experiment was conducted to screen fern plants for their potential to accumulate selenium. Eleven fern species, Pteris vittata, P. quadriaurita, P. dentata, P. ensiformis, P. cretica, Dryopteris erythrosora, Didymochlaena truncatula, Adiantum hispidulum, Actiniopteris radiata, Davallia griffithiana, and Cyrtomium fulcatum, were grown under hydroponic conditions for one week at 20 mg L(-1) selenate or selenite. Root Se concentrations reached 245-731 and 516-1082 mg kg(-1) when treated with selenate and selenite, respectively. The corresponding numbers in the fronds were 153-745 and 74-1,028 mg kg(-1) with no visible toxicity symptoms. Only three fern species were able to accumulate more Se in the fronds than the roots, which were D. griffithiana when treated with selenate, P. vittata when treated with selenite, and A. radiata regardless of the forms of Se. A. radiata was the best species overall for Se accumulation. More research is needed to further determine the potential of the fern species identified in this study for phytoremediation of the Se contaminated soils and water.

Two hydroponic experiments were conducted to evaluate factors affecting plant arsenic (As) hyperaccumulation. In the first experiment; two As hyperaccumulators (Pteris vittata and P. cretica mayii) were exposed to 1 and 10 mg L(-1) arsenite (AsIII) and monomethyl arsenic acid (MMA) for 4 wk. Total As concentrations in plants (fronds and roots) and solution were determined In the second experiment P. vittata and Nephrolepis exaltata (a non-As hyperaccumulator) were exposed to 5 mgL(-1) arsenate (AsV) and 20 mgL(-1) AsIIIfor 1 and 15 d. Total As and AsIII concentrations in plants were determined Compared to P. cretica mayii, P. vittata was more efficient in arsenic accumulation (1075-1666 vs. 249-627mg kg(-1) As in the fronds) partially because it is more efficient in As translocation. As translocation factor (As concentration ratio in fronds to roots) was 3.0-5.6 for P. vittata compared to 0.1 to 4.8 for P. cretica. Compared to N. exaltata, P. vittata was significantly more efficient in arsenic accumulation (38-542 vs. 4.8-71 mg kg(-1) As in thefronds) as well asAs translocation (1.3-5.6 vs. 0.2-0.5). In addition, P. vittata was much more efficient in As reduction from AsV to AsIII (83-84 vs. 13-24% AsIII in the fronds). Little As reduction occurred after 1-d exposure to AsV in both species indicates that As reduction was not instantaneous even in an As hyperaccumulator. Our data were consistent with the hypothesis that both As translocation and As reduction are important for plant As hyperaccumulation.

Past reports of the water relations of epiphytes, particularly bromeliads, indicate that tissue osmotic potentials in these tropical and subtropical plants are very high (close to zero) and are similar to values for aquatic plants. This is puzzling because several ecophysiological studies have revealed a high degree of drought stress tolerance in some of these epiphytes. The goal of this study was two-fold: (1) to increase the number of epiphytic taxa sampled for tissue osmotic potentials; and (2) to explain the apparent discrepancy in the significance of the tissue water relations and tolerance of drought stress in epiphytes. Tissue osmotic potentials of 30 species of epiphytic ferns, lycophytes, and orchids were measured in a subtropical rain forest in northeastern Taiwan. Nearly all values were less negative than -1.0 MPa, in line with all previous data for epiphytes. It is argued that such high osmotic potentials, indicative of low solute concentrations, are the result of environmental constraints of the epiphytic habitat on productivity of these plants, and that low rates of photosynthesis and transpiration delay the onset of turgor loss in the tissues of epiphytes such that they appear to be very drought-stress tolerant. Maintenance of photosynthetic activity long into drought periods is ascribed to low rates of transpiration and, hence, delayed tissue desiccation, and hydration of the photosynthetic tissue at the expense of water from the water-storage parenchyma.

Cytomorphological studies of the development of young fern gametophytes (Anemia phyllitidis) have been used to investigate combined effects of gibberellic acid and ethylene on male sex expression. ACC (the key by-product in ethylene biosynthesis pathway) was found to exert a synergetic effect on the gibberellic acid-induced antheridia formation, and this phenomenon could be related with the specific stimulation of cell growth and activity of their differentiation. To complete and verify those observations male sex expression in the fern gametophytes treated with ACC-biosynthesis inhibitor was reinvestigated. Aminooxyacetic acid (AOA) restrained antheridia formation via inhibition of cell divisions. AOA influenced the arrangement and flexibility of cellulose microfibrils in the antheridial zone cells, thus affecting cell expansion. On the other hand, the level of DNA synthesis was not reduced. Transient increase in the number of S-phase cells, followed by the accumulation of G2-phase cells led to the enhancement of cell polyploidization. All these findings correspond with the previous observations and support participation of ethylene in gibberellic acid-induced male sex expression in ferns.

Plastocyanin (Pc) is a soluble copper protein that transfers electrons from cytochrome b(6)f to photosystem I (PSI), two protein complexes that are localized in the thylakoid membranes in chloroplasts. The surface electrostatic potential distribution of Pc plays a key role in complex formation with the membrane-bound partners. It is practically identical for Pcs from plants and green algae, but is quite different for Pc from ferns. Here we report on a laser flash kinetic analysis of PSI reduction by Pc from various eukaryotic and prokaryotic organisms. The reaction of fern Pc with fern PSI fits a two-step kinetic model, consisting of complex formation and electron transfer, whereas other plant systems exhibit a mechanism that requires an additional intracomplex rearrangement step. The fern Pc interacts inefficiently with spinach PSI, showing no detectable complex formation. This can be explained by assuming that the unusual surface charge distribution of fern Pc impairs the interaction. Fern PSI behaves in a similar way as spinach PSI in reaction with other Pcs. The reactivity of fern Pc towards several soluble c-type cytochromes, including cytochrome f, has been analysed by flavin-photosensitized laser flash photolysis, demonstrating that the specific surface motifs for the interaction with cytochrome f are conserved in fern Pc.

In this study, we characterized lead (Pb2+) accumulation and storage by the aquatic fern Azolla filiculoides. Lead precipitates were detected in the vacuoles of mesophyll cells of Azolla plants cultured for 6 d in rich growth medium containing 20 mg l(-1) Pb2+. Energy dispersive spectroscopy (EDS) analysis of the relative element content of leaves collected from these plants revealed a 100% increase in the levels of P, S, Na and Ca and a 40% decrease in Mg and Cl compared to the untreated plants. Both Azolla whole plants and isolated apoplasts were incubated for 6 d in 20 mg l(-1) Pb2+. Lead content in the whole plant composed 0.37%, 2.3% and 1.8% of the dry weight after 2, 4 and 6 d of growth, respectively, while the isolated Azolla apoplast contained 0.125%, 1.22% and 1.4% Pb2+, respectively. Lead content in Azolla whole plant increase by 200%, 100% and 22% after 2, 4 and 6 d of growth, respectively, when compared to Azolla apoplast. Dark, electron dense deposits of lead were observed in light and transmission electron microscope in leaf cells treated with lead. All the observed lead deposits were localized in vacuoles while larger lead deposits were found in mature leaves than in young leaves. No lead deposits were found in cells of the cyanobiont Anabaena when the plants were exposed to similar conditions. Activity and content of V-H+-ATPase were studied in Azolla plants grown in the presence of 20, 40 and 80 mg l(-1) of lead for a period of 4 d. Activity of V-H+-ATPase was increased by 190%, 210% and 220%, respectively, but the content of V-H+-ATPase was reduced by all lead concentrations., (Copyright 2004 Elsevier SAS)

The aim of this paper was to investigate the capacity of a small water fern, Azolla caroliniana Willd. (Azollaceae), to purify waters polluted by Hg and Cr. Many plants are capable of accumulating heavy metals (called hyperaccumulators) and one of them is the water fern A. caroliniana. During 12 days of the experiment the fern was grown on the nutrient solution containing Hg2+, Cr3+ and CrO4(2-) ions, each in a concentration 0.1, 0.5 and 1.0 mg dm(-3). The presence of these ions caused a 20-31% inhibition of A. caroliniana growth, the highest in the presence of Hg(II) ions, in comparison to the control. After day 12 of the experiment, metal contents the solution decreased to 0-0.25 mg dm(-3), and this decrease comprised between 74 (Cr3+ 1.0 mg dm(-3) treatment) and 100% (CrO4(2-) 0.1 mg dm(-3) treatment). The fern took a lesser quantity of the metals from 0.1 mg dm(-3) treatments compared to 0.5 and 1.0 mg dm(-3) treatments. In the A. caroliniana tissues the concentration of heavy metals under investigation ranged from 71 to 964 mg kg(-1) dm; the highest level being found for Cr(III) containing nutrient solution.

Borate ester cross-linking of the cell wall pectic polysaccharide rhamnogalacturonan II (RG-II) is required for the growth and development of angiosperms and gymnosperms. Here, we report that the amounts of borate cross-linked RG-II present in the sporophyte primary walls of members of the most primitive extant vascular plant groups (Lycopsida, Filicopsida, Equisetopsida, and Psilopsida) are comparable with the amounts of RG-II in the primary walls of angiosperms. By contrast, the gametophyte generation of members of the avascular bryophytes (Bryopsida, Hepaticopsida, and Anthocerotopsida) have primary walls that contain small amounts (approximately 1% of the amounts of RG-II present in angiosperm walls) of an RG-II-like polysaccharide. The glycosyl sequence of RG-II is conserved in vascular plants, but these RG-IIs are not identical because the non-reducing L-rhamnosyl residue present on the aceric acid-containing side chain of RG-II of all previously studied plants is replaced by a 3-O-methyl rhamnosyl residue in the RG-IIs isolated from Lycopodium tristachyum, Ceratopteris thalictroides, Platycerium bifurcatum, and Psilotum nudum. Our data indicate that the amount of RG-II incorporated into the walls of plants increased during the evolution of vascular plants from their bryophyte-like ancestors. Thus, the acquisition of a boron-dependent growth habit may be correlated with the ability of vascular plants to maintain upright growth and to form lignified secondary walls. The conserved structures of pteridophyte, lycophyte, and angiosperm RG-IIs suggests that the genes and proteins responsible for the biosynthesis of this polysaccharide appeared early in land plant evolution and that RG-II has a fundamental role in wall structure.

Chloromethane (CH(3)Cl) is the most abundant halocarbon in the atmosphere. Although largely of natural origin it is responsible for around 17% of chlorine-catalysed ozone destruction. Sources identified to date include biomass burning, oceanic emissions, wood-rotting fungi, higher plants and most recently tropical ferns. Current estimates reveal a shortfall of around 2 million ty(-1) in sources versus sinks for the halocarbon. It is possible that emissions from green plants have been substantially underestimated. A potentially valuable tool for validating emission flux estimates is comparison of the delta13C value of atmospheric CH(3)Cl with those of CH(3)Cl from the various sources. Here we report delta13C values for CH(3)Cl released by two species of tropical ferns and show that the isotopic signature of CH(3)Cl from pteridophytes like that of CH(3)Cl from higher plants is quite different from that of CH(3)Cl produced by biomass burning, fungi and industry. delta13C values for CH(3)Cl produced by Cyathea smithii and Angiopteris evecta were respectively -72.7 per thousand and -69.3 per thousand representing depletions relative to plant biomass of 42.3 per thousand and 43.4 per thousand. The characteristic isotopic signature of CH(3)Cl released by green plants should help constrain their contribution to the atmospheric burden when reliable delta13C values for all other major sources of CH(3)Cl are obtained and a globally averaged delta13C value for atmospheric CH(3)Cl is available.

Legumin- and vicilin-like proteins have been isolated from spores of the fern Matteuccia struthiopteris. Their relationship with seed legumin and vicilin was demonstrated by cross-reactivities of antibodies directed against respective storage globulins from Vicia faba as evidenced by Western blotting. The Matteuccia legumin-like protein was characterised as a 300-340 kDa holoprotein preferentially consisting of a 32 kDa alpha-chain and a 24 kDa beta-chain. Patterns of limited proteolysis of the spore legumin-like protein and seed legumins were similar as well. In contrast to seed legumins, the Matteuccia legumin-like protein is devoid of disulfide bridges between alpha- and beta-chains. A 52 kDa polypeptide of the Matteuccia vicilin-like protein, first detected by SDS gel electrophoresis, is probably encoded by a vicilin-like gene specifically expressed in Matteuccia struthiopteris spores (Shutov et al. 1998). The vicilin-like holoprotein was found to form a complex of 600 kDa apparent molecular mass, presumably composed of four vicilin-like trimers.

The distribution of ptaquiloside (PTA) was studied in four Danish bracken populations in order to evaluate the transfer of PTA from ferns to soil. Populations showed statistically significant differences in PTA contents of fronds and rhizomes despite large in-site variations. The highest concentrations were encountered in fronds with concentrations ranging between 213 and 2145 microg/g, while rhizomes had concentrations between 11 and 902 microg/g. PTA was present in soil materials in amounts of 0.22-8.49 microg/g but apparently with no correlation with PTA contents of fronds or rhizomes. Laboratory tests showed that water could leach PTA from bracken fronds, which is in support of the high soil contents observed at sites exposed to heavy showers just before sampling. The observed soil contents correspond to estimated soil solution concentrations of 200-8500 microg/liter, demonstrating a substantial risk of PTA contamination of surface water and groundwater.

Methyl chloride is the largest natural source of ozone-depleting chlorine compounds, and accounts for about 15 per cent of the present atmospheric chlorine content. This contribution was likely to have been relatively greater in pre-industrial times, when additional anthropogenic sources-such as chlorofluorocarbons-were absent. Although it has been shown that there are large emissions of methyl chloride from coastal lands in the tropics, there remains a substantial shortfall in the overall methyl chloride budget. Here we present observations of large emissions of methyl chloride from some common tropical plants (certain types of ferns and Dipterocarpaceae), ranging from 0.1 to 3.7 microg per gram of dry leaf per hour. On the basis of these preliminary measurements, the methyl chloride flux from Dipterocarpaceae in southeast Asia alone is estimated at 0.91 Tg yr-1, which could explain a large portion of missing methyl chloride sources. With continuing tropical deforestation, natural sources of chlorine compounds may accordingly decrease in the future. Conversely, the abundance of massive ferns in the Carboniferous period may have created an atmosphere rich in methyl chloride.

Expansins are a group of extracellular proteins that directly modify the mechanical properties of plant cell walls, leading to turgor-driven cell extension. Within the completely sequenced Arabidopsis genome, we identified 38 expansin sequences that fall into three discrete subfamilies. Based on phylogenetic analysis and shared intron patterns, we propose a new, systematic nomenclature of Arabidopsis expansins. Further phylogenetic analysis, including expansin sequences found here in monocots, pine (Pinus radiata, Pinus taeda), fern (Regnellidium diphyllum, Marsilea quadrifolia), and moss (Physcomitrella patens) indicate that the three plant expansin subfamilies arose and began diversifying very early in, if not before, colonization of land by plants. Closely related "expansin-like" sequences were also identified in the social amoeba, Dictyostelium discoidium, suggesting that these wall-modifying proteins have a very deep evolutionary origin.

A method for quantitative assessment of 137Cs availability to plants in forest ecosystems on the basis of soil properties has been developed. It is shown that the experimental dependencies of 137Cs soil-to-plant transfer factor (TFag) for fern and bilberry on the bioavailability factor calculated on the basis of soil characteristics of root layer: 137Cs exchangeability, exchangeable Ca, effective selectivity coefficient, were satisfactory described by linear function. The advantage of the proposed method is that the necessary soil characteristics can be taken from the reference literature, evaluated using empirical correlations or determined with standard agrochemical procedures.

Psilotum nudum Griseb is a pteridophyte and belongs to the single family (Psilotaceae) of the division, Psilophyta. Being the only living species of a once populated division, P. nudum is the most primitive vascular plant. Chalcone synthase (CHS; EC 2.3.1.74) superfamily enzymes are responsible for biosyntheses of diverse secondary metabolites, including flavonoids and stilbenes. Using a reverse transcription-polymerase chain reaction strategy, four CHS-superfamily enzymes (PnJ, PnI, PnL and PnP) were cloned from P. nudum, and heterologously expressed in Escherichia coli. These four enzymes of 396-406 amino acids showed sequence identity of > 50% among themselves and to other higher-plant CHS-superfamily enzymes. PnJ and PnP preferred p-coumaroyl-CoA and isovaleryl-CoA respectively, as starter CoA and catalyzed CHS-type ring formation, indicating that they are CHS and phlorisovalerophenone synthase, respectively. On the other hand, PnI and PnL preferred cinnamoyl-CoA as starter CoA and catalyzed stilbene synthase-type cyclization and thus were determined to be pinosylvin synthases (EC 2.3.1.146). In addition, PnE, which uniquely contains a glutamine in place of otherwise strictly conserved histidine, had no apparent in vitro catalytic activity. Phylogenetic analysis indicated that these P. nudum clones form a separate cluster together with Equisetum arvense CHS. This cluster of pteridophytes is located next to the cluster formed by pine (gymnosperm) enzymes, in agreement with their evolutionary relationships. Psilotum nudum represents a plant with the most diverse CHS-superfamily enzymes and this ability to diverge may have provided a survival edge during evolution.

Cd uptake capacity of a group of floating macrophytes (Salvinia herzogii, Pistia stratiotes, Hydromistia stolonifera and Eichhornia crassipes) was determined in outdoors experiments during the lowest temperature period of the year. Although all studied species were highly efficient in the Cd uptake, Pistia stratiotes was selected for further research because of its superior performance and its higher average relative growth rate. Cadmium% removal by Pistia stratiotes was greater in the first 24 h of the experiments (63, 65, 72 and 74% of the added Cd for 1, 2, 4 and 6 mg Cd 1(-1), respectively). After 31 days of growth, Pistia statiotes efficiently removed Cd at the studied concentrations. The macrophyte was able to keep its capacity for Cd removal even though some toxicity symptoms appeared at 4 and 6 mg Cd 1(-1). The greater the initial concentration, the greater Cd bioaccumulation rates. The increase of Cd concentration in plant tissues occurred especially in roots and was linearly related to the quantity of Cd added. Cd sorption by roots is faster than translocation to the plant aerial part and it occurs mainly during the first 24h.

The equipment of using Azolla for O2-supplementation and food-production in future space station was developed and tested. Dog was used as the O2-consuming animal. The design of this device considered both the requirement of Azolla growth, such as illumination, temperature, humidity, nutrition and biomass harvesting, and also the food supplement, excretion draining and temperature controlling for the dog under the condition of an airtight chamber for a relatively long duration. This device was preliminarily tested for O2-release by Azolla, and data about O2-supplement by Azolla were obtained.

This study was conducted to characterize the responses of selected nitrogen-fixing biocatalysts to various environmental parameters that are likely to be encountered in a scaled-up nitrogen-fixing bioregenerative unit for use in a biologically based life support system (BLSS). The results showed that while both Azolla filiculoides and Azolla nilotica thrived on standard nitrogen-free liquid media, only Azolla filiculoides maintained its growth rate in the presence of 50% by volume of aerobic-bioreactor potato effluent (ABPE) in its liquid medium. The growth rate and Anabaena cavity population of Azolla filiculoides also remained undiminished in the presence of as much as 10 mM of NO3- in its liquid medium. The tolerance of Azolla filiculoides for NO3- however, lay somewhere between 10 and 20 mM of NO3-. Azolla filiculoides failed to grow at 5 mM or greater concentrations of NH4+. The growth rate of the Oscillatoria-dominated microbial mat was significantly elevated by increased silage loading density (16 g/L) as well as by adding 5 mM of NO3- into its liquid medium. Neither the growth rate of Azolla filiculoides nor that of the microbial mat was affected by three levels of photosynthetic photon flux (PPF) (520, 265, and 125 micromoles m-2 s-1) under high-pressure sodium or cool fluorescent lamps.