Dissertations / Theses on the topic 'Mosses'

The aim of this research was to assess how mosses took up nitrogen (N), assimilated this N and sensed N status. Research into the short term foliar uptake and assimilation of N investigated the rapid induction of nitrate reductase activity (NRA) in two species of moss Mnium hornum and Sphagnum fimbriatum. NRA was induced in both species in just 15 minutes. Tissue nitrate (NO3) content was measured over this period and it was shown that the species had contrasting NO3" storage capabilities. Both species also differed in their long term N use. S. fimbriatum was a less nitrophilous species and died after approximately 80 days N-deprivation but M. hornum had a tremendous capacity to sustain itself and survived for in excess of 450 days. It was suggested from electrophysiological measurements and microscopy that S. fimbriatum had a lower NO3" storage capacity due to the lack of a distinct vacuole or storage compartment for NO3". M. hornum had both cytosolic and vacuolar compartments and was more able to store NO3", even in the longer term, being more similar in structure to a higher plant. Ammonium (Nlt induced NRA in the absence of NO3" in both mosses. Electrophysiological measurements, using pH-selective microelectrodes, showed a depolarisation of the membrane potential and acidification of intracellular compartments in moss cells upon addition of NH/ an example being a pH change from pH 7.11 to 5.84 in the cytosol of M. hornum. N uptake mechanisms in these mosses supported the classical pH-stat model and current thinking on plant N transporter mechanisms in higher plants whereby the addition of NH/ caused an acidification of the cytosol, in turn inducing NRA to stabilise intracellular pH. This project brought together the biochemistry, physiology and localised changes in metabolite pools and pH to explain why these mosses behaved differently.

Our first aim was to identify and quantify Biological Volatile Organic Compound (BVOC) emissions, specifically emissions of isoprene, from the moss Polytrichum juniperinum during its earliest stage of life. Isoprene emission from mosses could be a significant component of the total global budget of BVOC emissions. Data concerning the spatial and temporal variability of these emissions are lacking due to poor characterization of the physical and biological factors controlling isoprene synthesis in both vascular and non-vascular plants. We found that P. juniperinum in its early life stage (protonema) can emit isoprene at detectable levels at day five after spore germination. These results suggest that protonema is capable of isoprene emission, shown here for the first time, and media affects emission rates. We saw a negative trend with leaf emergence and protonema growth. Chlorophyll fluorescence ratio had a positive correlation with isoprene emission, and isoprene emission was both temperature and light dependent in early protonema. Our second aim was to determine the effects of sex and sexual expression on isoprene emission in mosses. Sex is a biologically costly, but it can result in genetic adaptability and variety for resulting offspring. Using three Polytrichaceae species, we measured isoprene emission between plants sexually expressing and non-expressing and between male and female moss plants. We found that non-sexually expressing gametophytes had significantly higher isoprene emission than gametophytes expressing sex, suggesting that sex expression may be costly and that additional resources are allocated to isoprene emission when plants are not reproducing. Males emitted higher levels of isoprene than females, but surprisingly this difference occurred only when plants were not expressing sex. We found species and sex-specific differences in chlorophyll fluorescence ratio (CFR) and relative electron transport rate (RETR). Our third aim was to investigate and categorize the effect of nitrogen addition on isoprene emission of P. juniperinum by creating an artificial nitrogen gradient with ammonium nitrate addition (NH4NO3). Current rates of anthropogenic N deposition are altering many biogeochemical processes. In these changing environments, increased nitrogen availability alters plant phenology, physiology, and the allocation of resources, but no information is available on whether additional N increases isoprene emissions in mosses. We used a manipulative experiment to measure the effects of nitrogen addition on moss isoprene emission, as well as on moss morphology and reproductive effort. We found site-specific differences in our morphological and physiological measurements. Isoprene emission was site, sex, and N addition specific, with the highest isoprene emission seen from our mid-level N addition, in both female- and male-majority pots. We found significant treatment and sex-specific interactions among our sites and within sites. We saw the highest reproductive effort counts from non-sexual expressive gametophytes, followed by high sporophyte count from female-majority pots. We also saw chlorophyll fluorescence ratio (CFR) differences between sites and treatments, but not between sexes.

Substantial ambiguities still remain concerning the broad backbone of moss phylogeny. I surveyed 17 slowly evolving plastid genes from representative taxa to reconstruct phylogenetic relationships among the major lineages of mosses in the overall context of land-plant phylogeny. I first designed 78 bryophyte-specific primers and demonstrated that they permit straightforward amplification and sequencing of 14 core genes across a broad range of bryophytes (three of the 17 genes required more effort). In combination, these genes can generate sturdy and well-resolved phylogenetic inferences of higher-order moss phylogeny, with little evidence of conflict among different data partitions or analyses. Liverworts are strongly supported as the sister group of the remaining land plants, and hornworts as sister to vascular plants. Within mosses, besides confirming some previously published findings based on other markers, my results substantially improve support for major branching patterns that were ambiguous before. The monogeneric classes Takakiopsida and Sphagnopsida likely represent the first and second split within moss phylogeny, respectively. However, this result is shown to be sensitive to the strategy used to estimate DNA substitution model parameter values and to different data partitioning methods. Regarding the placement of remaining nonperistomate lineages, the [[[Andreaeobryopsida, Andreaeopsida], Oedipodiopsida], peristomate mosses] arrangement receives moderate to strong support. Among peristomate mosses, relationships among Polytrichopsida, Tetraphidopsida and Bryopsida remain unclear, as do the earliest splits within sublcass Bryidae. A Funariidae, [Timmiidae, [Dicranidae, Bryidae]]] arrangement is strongly supported, as are major relationships within subclasses Funariidae and Dicranidae. I also reconstructed the phylogeny of the nonperistomate moss family Andreaeaceae, with a focus on costate taxa, using two complementary sets of plastid markers and taxa. The major subgenera (Andreaea and Chasmocalyx) and sections of Andreaea (Andreaea and Nerviae) are rejected as monophyletic. Well-supported lineages include clades comprising: (1) Andreaea nivalis and A. rigida (northern hemisphere members of subgenus Chasmocalyx) and A. blyttii (section Nerviae); (2) most of the remainder of Nerviae; (3) a mixture of costate and ecostate species from Chasmocalyx, Nerviae, all sampled members of section Andreaea, and subgenus Acroschisma. Relationships among the major lineages, including the root of the family, are all well supported.

In arctic tundra ecosystems mosses dominant the vegetation in terms of productivity and diversity.  Despite this, mosses are often overlooked in studies of tundra ecology.  However, evidence from this thesis suggests that mosses maybe integral to the functioning of these systems.  Mosses insulate soil keeping it cooler than air temperature, an effect more apparent under deeper moss.  The effects of the moss layer on soil characteristics alter conditions for microbial populations resulting in higher nitrogen availability in soil under shallow moss.  This thesis shows that the role of mosses in determining vascular plant success may dictate many higher plant interactions.  There are both positive and negative effects of the moss layer on vascular plant growth.  The relationship between positive and negative impacts of the moss layer on vascular plants is species specific, meaning that moss cover may be a key determinate of vascular plant community structure.  Climatic warming and herbivory are important drivers of vegetation change in the Arctic.  This thesis shows that grazing by reindeer and grubbing by geese is detrimental to moss cover.  As mosses insulate the soil, a reduction in depth or integrity increases soil temperatures and enhances microbial activity and thus nitrogen availability.  This in conjunction with addition of nutrients from faeces enhances vascular plant productivity to the further detriment of mosses.  Warming increases soil temperature and accelerates decomposition, but has little affect on either biomass of moss or vascular plants.  Moss grubbing has a greater negative effect on mosses in a warmed environment.  This thesis concludes that mosses are integral to the current functioning of tundra heaths.

Bibliography: leaves 86-97.
Lepfodon smithii (Leptodontaceae) and Pterogonium gracile (Leucodontaceae) are widespread and disjunctly distributed moss species, that in South Africa are confined to forest patches believed to be relics of vast forests that existed before the Pleistocene glaciation period. These two species exhibit similar distribution and ecologies and frequently co-occur. Wardia hygrometrica is a southwestern Cape endemic that is restricted to streams within the kloofs that the forests generally occupy. In an attempt to trace the species histories and their subsequent colonization of forest patches, two molecular markers, internal transcribed spacer (ITS) nuclear rDNA and trnL-F cpDNA were employed.

Nathaniel Hawthorne’s collection of short stories, Mosses from an Old Manse, serves as his contribution to the philosophical discussions on Transcendentalism in Concord, MA in the early 1840s. While Ralph Waldo Emerson, Henry David Thoreau, and the other individuals involved in the Transcendental club often seem to readily accept the positions presented in Emerson’s work, it is never so simple for Hawthorne. Repeatedly, Hawthorne’s stories demonstrate his difficulty in trying to identify his own opinion on the subject. Though Hawthorne seems to want to believe in the optimistic potential of the spiritual and intellectual ideal presented in Emersonian Transcendentalism, he consistently dwells on the evil and blackness that may be contained in the human heart. The collection of short stories written while Hawthorne lived in Concord and surrounded himself with those dominant literary figures represents the clearest articulation of his ambivalent position on Transcendentalism.

The microorganisms colonizing plants can have a significant effect on host phenotype, mediating such processes as pathogen resistance, stress tolerance, nutrient acquisition, growth, and reproduction. Research regarding plant-microbe interactions has focused almost exclusively on vascular plants, and we know comparatively little about how bryophytes -- including mosses, liverworts, and hornworts -- are influenced by their microbiomes. Ceratodon purpureus is a dioecious, cosmopolitan moss species that exhibits sex-specific fungal communities, yet we do not know whether these microbes have a differential effect on the growth and physiology of male and female genotypes. Using a common-garden design, we reared ten axenic genotypes of C. purpureus in a controlled environmental chamber. Clonal C. purpureus replicates, with and without the addition of a microbial inoculation, were used to test the effect of a mixed microbial community on vegetative growth, sex expression, photosynthetic efficiency (Fv/Fm and ETR), and chlorophyll content (CFR) for male and female mosses. We found that microbes had a negative impact on the growth and photosynthesis efficiency of C. purpureus, and this effect varied among genotypes of C. purpureus for ETR and growth. Microbes also had a positive, sex-specific effect on chlorophyll content in C. purpureus, with males exhibiting lower CFR values in the absence of microbes. C. purpureus sex expression was marginally negatively affected by microbe addition, but gametangia production was low overall in our experiment. We also conducted preliminary surveys using direct counts from moss ramets to assess the community composition of epiphytic algae associated with our microbe addition and control C. purpureus. These surveys identified three algal morphospecies in association with the microbe addition C. purpureus genotypes, as well as cyanobacteria, nematodes, rotifers, and testate amoeba. No algae, cyanobacteria, or micro-fauna were observed in the control plants. Transplantation of a mixed microbial community from field-to-laboratory conditions may be applied to other bryophyte species under varying environmental conditions to provide insight into how these diminutive yet important ecosystems will respond to environmental perturbation.

This thesis examines the relative importance of local conditions, dispersal and dynamics of the trees on epiphyte distribution patterns and colonization-extinction dynamics. Study species are the mosses Orthotrichum speciosum and O. obtusifolium, and the red-listed Neckera pennata. The thesis also includes an attempt to parameterize a model for a lichen metapopulation (Lobaria pulmonaria) in a dynamic landscape, based on only presence/absence data of the epiphyte and its host trees.

The results show that epiphyte colonization of trees is affected by both local conditions, and by connectivity to occupied trees. The positive effect of connectivity, implying a restricted dispersal range, was established by both demographic and genetic studies. The important local conditions were tree diameter and vitality, and shade. Local extinctions from trees occurred among small trees with low local epiphyte abundance, but more often, were the results of tree fall.

The observed importance of connectivity on epiphyte colonization agrees with the assumptions of the classic metapopulation model. However, the classic metapopulation model assumes that the landscape is static, and that local extinctions occur for stochastic reasons. The dynamics of epiphytes are different. A new conceptual model is therefore suggested, the patch-tracking metapopulation model. It differs from the classic metapopulation model in that it includes dynamics of the patches, and in that local extinctions only occur as patches are destroyed.

Simulations of the dynamics of N. pennata showed that its future metapopulation size will be overestimated unless the dynamics of the trees are accounted for. The simulation results further suggest that the dynamics of N. pennata can be characterised by the patch-tracking metapopulation model.

The attempt to parameterize the L. pulmonaria metapopulation model showed that more information are required for rigorous parameterization, preferably of the past historic fire regime.

Peat mosses (Sphagnum) are widely distributed and are a major component of mire vegetation and peat throughout the boreal and temperate regions. Most boreal Sphagnum species regularly produce sporophytes, but the ecological role of the spore has been questioned. This study shows that the spores can form a spore bank and have the ability to germinate and contribute to moss establishment whenever suitable conditions occur. The results suggest that spore production is important for explaining the wide distribution and omnipresence of Sphagnum in nutrient-poor wetlands. The results further imply that initial recruitment from spores predominates in Sphagnum after disturbance or formation of suitable habitats.

A series of experiments showed that addition of phosphorus-containing substrates, such as fresh plant litter or moose dung, resulted in spore establishment on bare, moist peat. A field experiment indicated establishment rates of about 1% of sown, germinable spores on peat with added substrates. Plant litter on moist soil, without a closed cover of bryophytes, is an important safe site for the establishment of Sphagnum spores. The results fit the observed pattern of colonisation by Sphagnum beneath Eriophorum vaginatum tussocks in mires severely disturbed by peat extraction. Successful long-distance dispersal was indicated by the occurrence of several regionally new or rare Sphagnum species in disturbed mires.

Spore number per sporophyte ranged among Sphagnum species from 18 500 to 240 000, with a trade-off between spore number and spore size. Annual spore production was estimated at 15 million spores per square metre on two investigated mires. Sporophyte production showed a large interannual variation. Sporophyte production was positively related to the amount of precipitation the preceding summer. This was probably because a high water level promoted gametangium formation. Spore dispersal occurred in July and August. The earlier timing of spore dispersal in the more drought-sensitive, hollow-inhabiting sphagna should reduce the risk of sporophytes drying out prematurely during summer droughts.

Spores kept refrigerated up to 13 years retained high germinability. A field experiment showed that Sphagnum can form a persistent spore bank, with a potential longevity of several decades.

Moss dominated ecosystems are an important part of the global terrestrial carbon cycle. Over large areas, remote sensing can be useful to provide an improved understanding of these ecosystems. Two boreal mossess (Pleurozium and Sphagnum) were assessed using remote sensing based spectral vegetation indices for estimating biochemical capacity and photosynthetic efficiency by varying net photosynthesis rate via changes in water content. In the laboratory, changes in the normalized difference vegetation index (NDVI) and chlorophyll index coincided with declining photosynthetic capacity due to desiccation. This effect was more dramatic in Sphagnum. The photochemical reflectance index (PRI) did not vary with changes in CO2 supply as anticipated, possibly due to overriding effects of changing water content. The water band index (WBI) was strongly related to water content but this relationship showed an uncoupling in the field. Bi-directional reflectance measurements indicated what WBI was sensitive to sensor, sun, and moss surface slope angles.
xi, 110 leaves : ill. (some col.) ; 29 cm.

Mosses are small avascular bryophytes with a haploid dominant gametophyte and a diploid sporophyte stage. The gametophyte cells are single layered and lack a protective cuticle, which is the first line of defense in vascular plants. These factors would render them highly susceptible to stress but on the contrary, mosses have flourished on land for the past 450 million years with tolerance to both abiotic and biotic stress. Occurrence of unique lipids in bryophytes was considered as an adaptive means to survive harsh terrestrial condition. A recent study identified a lipid metabolite, anandamide in the Physcomitrella patens. Anandamide (NAE 20:4) belongs to a group of fatty acid ethanolamides or N –acylethanolamines (NAEs). In eukaryotes, NAEs were shown to play an important role in mediating stress responses. In plants, NAE 14:0 has been implicated in biotic stress response; its levels increased up to 50-fold in elicitor-treated tobacco plants, along with induction of defense gene expression and inhibition of alkalization. In animals anandamide acts as an endocannabinoid ligand and mediates several physiological responses including stress. This study aims to use P. patens as the model system because of its available genomic database and prior studies on biotic stress, to examine if NAE 20:4 contributes to their ability to tolerate biotic stress. It is hypothesized that the occurrence of anandamide will play a role in mediating biotic stress tolerance in P. patens. To test this hypothesis, three specific aims are proposed. They are to determine the effect of 1) elicitor-treatment on NAE and fatty acid profile in the moss, 2) anandamide on elicitor-induced morphological and physiological changes in the moss and 3) anandamide on elicitor-induced defense gene expression in moss. Mosses utilize similar defense mechanisms as flowering plants and disease symptoms can easily be studied using microscopy because of their haploid dominant gametophyte stage with monolayer cells. The induction of defense gene expression will be studied by quantitative PCR and changes in lipid profile by selective lipidomics. This study is expected to provide novel insights into the role of anandamide in early land plants, specifically in response to biotic stress.

Les bases moléculaires des réponses aux excès de lumière chez les organismes photosynthétiques appartenant à des lignées évolutives distinctes ne sont toujours pas complètement caractérisées. Par conséquent, j'ai caractérisé des antennes photoprotectrices dans les algues vertes, les mousses et les diatomées et j'ai exploré la fonction de deux cycles de xanthophylles chez les diatomées.J'ai étudié les protéines Light Harvesting Complex Stress-Related (LHCSR) dans tous ces organismes. Chez l'algue verte Chlamydomonas reinhardtii, j’ai identifié par mutagénèse dirigée, complémentation fonctionnelle et par une approche biochimique les acides aminés responsables de l'activation de LHCSR3, une protéine importante pour le NPQ.Dans le modèle de mousse Physcomitrella patens, j'ai etudié in vitro les caractéristiques spectroscopique ainsi que le quenching de différents mutants de liaison de pigment sur la protéine LHCSR1.Les protéines LHCSR dans les diatomées sont nommées LHCXs, et dans Phaeodactylum tricornutum j'ai montré que l'expansion de la famille des gènes LHCX reflète une diversification fonctionnelle de ces protéines permettant de répondre à des environnements marins très variables.L'autre acteur principal de la photoprotection dans les diatomées est le cycle des xanthophylles. J’ai trouvé que l'accumulation d'une grande quantité de viola- et zéaxanthin a un effet négatif sur le NPQ montrant que la zéaxanthin ne participe pas au NPQ chez diatomées.Grâce à ces études effectuées, nous avons acquis une connaissance plus approfondie sur les caractéristiques communes et les spécificités de la photoprotection.chez différents organismes
The molecular bases of responses to light excess in photosynthetic organisms having different evolutionary histories and belonging to different lineages are still not completely characterized. Therefore I explored the functions of photoprotective antennae in green algae, mosses and diatoms, together with the role of the two xanthophyll cycles present in diatoms.I studied the Light Harvesting Complex Stress-Related (LHCSR) proteins in different organisms. In the green alga Chlamydomonas reinhardtii, LHCSR3 is a protein important for photoprotection. I used site-specific mutagenesis in vivo and in vitro and identified three residues of LHCSR3 that are responsible for its activation.With the moss Physcomitrella patens I studied the in vitro spectroscopic and quenching characteristics of different pigment-binding mutants of the protein LHCSR1, focusing in particular on chlorophylls A2 and A5.LHCSRs in diatoms are named LHCXs, and in Phaeodactylum tricornutum I found that multiple abiotic stress signals converge to regulate the LHCX content of cells, providing a way to fine-tune light harvesting and photoprotection.The other main driver of photoprotection in diatoms is the xanthophyll cycle. Here I found that the accumulation of viola- and zeaxanthin in P. tricornutum have a negative effect in the development of NPQ, showing that zeaxanthin does not participate in the enhancing of NPQ in diatoms.Thanks to these studies done on different organisms, we gained a deeper knowledge on the shared characteristics and on the peculiar features about photoprotection in green algae, mosses and diatoms

The molecular bases of responses to light excess in photosynthetic organismshaving different evolutionary histories and belonging to different lineages are still not completely characterized.Therefore I explored the functions of photoprotective antennae in green algae, mosses and diatoms, together withthe role of the two xanthophyll cycles present in diatoms.I studied the Light Harvesting Complex Stress-Related (LHCSR) proteins in different organisms. In thegreen alga Chlamydomonas reinhardtii, LHCSR3 is a protein important for photoprotection. I used site-specificmutagenesis in vivo and in vitro and identified three residues of LHCSR3 that are responsible for its activation.With the moss Physcomitrella patens I studied the in vitro spectroscopic and quenching characteristics ofdifferent pigment-binding mutants of the protein LHCSR1, focusing in particular on chlorophylls A2 and A5.LHCSRs in diatoms are named LHCXs, and in Phaeodactylum tricornutum I found that multiple abioticstress signals converge to regulate the LHCX content of cells, providing a way to fine-tune light harvesting andphotoprotection.The other main driver of photoprotection in diatoms is the xanthophyll cycle. Here I found that the accu-mulation of viola- and zeaxanthin in P. tricornutum have a negative effect in the development of NPQ, showingthat zeaxanthin does not participate in the enhancing of NPQ in diatoms.Thanks to these studies done on different organisms, we gained a deeper knowledge on the shared character-istics and on the peculiar features about photoprotection in green algae, mosses and diatoms.

Les bases moléculaires des réponses aux excès de lumière chez les organismes photosynthétiques appartenant à des lignées évolutives distinctes ne sont toujours pas complètement caractérisées. Par conséquent, j'ai caractérisé des antennes photoprotectrices dans les algues vertes, les mousses et les diatomées et j'ai exploré la fonction de deux cycles de xanthophylles chez les diatomées.J'ai étudié les protéines Light Harvesting Complex Stress-Related (LHCSR) dans tous ces organismes. Chez l'algue verte Chlamydomonas reinhardtii, j’ai identifié par mutagénèse dirigée, complémentation fonctionnelle et par une approche biochimique les acides aminés responsables de l'activation de LHCSR3, une protéine importante pour le NPQ.Dans le modèle de mousse Physcomitrella patens, j'ai etudié in vitro les caractéristiques spectroscopique ainsi que le quenching de différents mutants de liaison de pigment sur la protéine LHCSR1.Les protéines LHCSR dans les diatomées sont nommées LHCXs, et dans Phaeodactylum tricornutum j'ai montré que l'expansion de la famille des gènes LHCX reflète une diversification fonctionnelle de ces protéines permettant de répondre à des environnements marins très variables.L'autre acteur principal de la photoprotection dans les diatomées est le cycle des xanthophylles. J’ai trouvé que l'accumulation d'une grande quantité de viola- et zéaxanthin a un effet négatif sur le NPQ montrant que la zéaxanthin ne participe pas au NPQ chez diatomées.Grâce à ces études effectuées, nous avons acquis une connaissance plus approfondie sur les caractéristiques communes et les spécificités de la photoprotection.chez différents organismes
The molecular bases of responses to light excess in photosynthetic organisms having different evolutionary histories and belonging to different lineages are still not completely characterized. Therefore I explored the functions of photoprotective antennae in green algae, mosses and diatoms, together with the role of the two xanthophyll cycles present in diatoms.I studied the Light Harvesting Complex Stress-Related (LHCSR) proteins in different organisms. In the green alga Chlamydomonas reinhardtii, LHCSR3 is a protein important for photoprotection. I used site-specific mutagenesis in vivo and in vitro and identified three residues of LHCSR3 that are responsible for its activation.With the moss Physcomitrella patens I studied the in vitro spectroscopic and quenching characteristics of different pigment-binding mutants of the protein LHCSR1, focusing in particular on chlorophylls A2 and A5.LHCSRs in diatoms are named LHCXs, and in Phaeodactylum tricornutum I found that multiple abiotic stress signals converge to regulate the LHCX content of cells, providing a way to fine-tune light harvesting and photoprotection.The other main driver of photoprotection in diatoms is the xanthophyll cycle. Here I found that the accumulation of viola- and zeaxanthin in P. tricornutum have a negative effect in the development of NPQ, showing that zeaxanthin does not participate in the enhancing of NPQ in diatoms.Thanks to these studies done on different organisms, we gained a deeper knowledge on the shared characteristics and on the peculiar features about photoprotection in green algae, mosses and diatoms

Investigation of morphology, karyotype, isozymes, cultivation, geography and habitat confirmed differences between Limbella tricostata (Sull.) CM. and Sciaromium fryei Williams. Significant differences were observed experimentally in branch bud frequency, leaf insertion angle, leaf areolation, chromosome length, mobility of superoxide dismutase and shoot orientation. Sciaromium fryei is considered a subspecies of Limbella tricostata. Chromosome numbers of both subspecies is n=11. L. tricostata ssp. tricostata has a Bryum type sporeling and a hypnaceous peristome, and is thought to have derived from L. pachyloma (Mont.) C.M. or its antecedent after long-distance dispersal from South America to Hawaii no later than the mid-Tertiary. L. tricostata ssp. fryei is thought to have derived from L. tricostata ssp. tricostata after long-distance dispersal from Hawaii to North America in the mid- to late Tertiary.
Science, Faculty of
Botany, Department of
Graduate

As non-vascular, early land plants with an aquatic ancestry, mosses do not regulate internal water conditions separate from that of their environment and as a result, evolved mechanisms to survive in a terrestrial world out of water. Yet, there is a widely accepted dogma that moss reproductive success is solely dependent on rainfall events carrying swimming, bi-flagellate sperm across the landscape to reproductively mature and receptive female mosses--but this classic view of moss reproduction may be too simplistic. In this dissertation I test the assumptions of reproductive limitation in mosses and present novel findings in a basal, yet understudied terrestrial plant mating system. I find evidence of environmental desiccation tolerance in moss sperm, thus offering the potential for stress-tolerant gametes on the landscape possibly suited for various transport vectors, reminiscent of a pollen grain. To investigate the broad evolutionary implications of this newfound complexity in moss sexual reproduction, I tested classic tenets of plant-pollinator theory on the ancient mosses and their ubiquitous microarthropod inhabitants. Experimental results show that mosses and microarthropods are engaged in a "pollination-like" syndrome guided by sex-specific volatile cues that differentially affect microarthropod behavior. These data indicate an existing complex mutualistic relationship and provides new evidence of sexually dimorphic investment by male and female mosses into reproductive success. Further, these data put forth the idea that female mosses challenge an inherent mate limitation by investing into reproductive assurance via maintaining a relationship with microarthropods. Experimental work further confirms a role for invertebrates in moss reproductive success and tests mutualism theory through ongoing experiments. Such experiments include an assessment of moss genetic diversity, paternity, and male fitness traits as it relates to mosses with or without the presence of microarthropods, thereby testing for fitness benefits gained by mosses possibly engaged in a transport mutualism with microarthropods. I further tested mutualism, community ecology and moss sexual reproduction concepts in extreme geothermal moss populations living at the edges of inhabitable Earth, and results show that even geothermal moss canopies are diverse and host differential and abundant life. In a first field test of mutualism I found that although extreme heat stress may constrain sexual reproduction in mosses, a correlation between within-population moss genetic diversity and microarthropod abundances exists. To further examine mosses in extreme environments, and how these environments may constrain sexual reproductive success, I evaluated the effects of simulated warming on Antarctic moss physiology and reproductive biology. Data indicates that simulated warming relieves mosses of physiological stress, and results in a greater investment into primary productivity and sexual reproduction. These data support the hypothesis that with less stress, sexual reproduction is increased. Mosses are an ideal system by which to understand organisms that exist in environments ranging from the mesic to the extreme, in the laboratory and the field and even in the classroom, where the small functioning ecosystem of mosses can be used for discovery-based biology education as described in the Mosscosms curriculum. This work contributes significantly to the field of bryophyte and plant biology by revealing novel insights into the biotic and abiotic drivers of sexual reproduction in mosses.

In plants, change in lipid composition is a common response to various abiotic stresses. Lipid constituents of bryophytes are of particular interest as they differ from that of flowering plants. Unlike higher plants, mosses have high content of very long-chain polyunsaturated fatty acids. Such lipids are considered to be important for survival of nonvascular plants. Here, using abscisic acid (ABA )-induced changes in lipid composition in Physcomitrella patens as an example, a protocol for total lipid extraction and quantification by gas chromatography (GC) coupled with flame ionization detector (FID) is described.

As one of the first land plant groups to diversify, mosses are central in understanding the origin, diversification, and early function of stomata. Unlike tracheophytes that have stomata on anatomically complex leaves and stems, mosses bear stomata exclusively on spore-bearing organs (capsules). However, stomata do not occur in all mosses and, indeed, are absence in the earliest-divergent mosses (Takakia, Andreaea, Andreaeobryum and Sphagnum), suggesting that stomata originated in mosses independently of other plants. The occurrence of structurally unique pseudostomata in Sphagnum further confounds the resolution of homology of moss stomata with those of other plants. The five studies included in this dissertation are aimed at clarifying the structure, development and evolution of moss stomata. The first study focuses on the sporophyte anatomy and stomatal ultrastructure in two structurally and phylogenetically divergent mosses, Oedipodium and Ephemerum. Oedipodium is the sister to peristomate mosses and the first extant moss with true stomata. This monospecific genus has an elaborated capsule with an extended apophysis bearing numerous long-pored stomata. In contrast, Ephemerum nests within the peristomate mosses and has a reduced capsule that lacks an apophysis and has a few round-pored stomata. Ultrastructure of stomata is similar in these two mosses and comparable to that of tracheophytes, except that the stomata of mosses are not as structurally distinct from epidermal cells as are tracheophyte stomata. Anatomical features such as the presence of a cuticle, water-conducting cells, and spongy tissues with large areas for gas exchange are more pronounced in Oedipodium sporophytes and support the role of stomata in gas exchange and water transport during development and maturation. The second study examines changes in pectin composition during development in the model moss Funaria. Stomatal movement in tracheophytes requires guard cell walls to be strong, yet flexible, because they have to undergo reversible deformation to open and close the pore. Pectins are necessary for wall flexibility and proper stomatal functioning in seed plants. In this study of Funaria, immunogold-labeling using five antibodies to pectin epitopes was conducted on guard cell walls during development to relate these features to the limited movement of stomata in moss. Movement of Funaria stomata coincides with capsule expansion when guard cell walls are thin and pectinaceous. Walls dramatically increase in thickness after pore formation and the pectin content significantly decreases in mature guard cell walls, suggesting that a decrease in flexibility is responsible for the inability to open a close previously reported in older moss guard cells. Because this was the first study to demonstrate changes in pectin composition during stomatal development in any plant, a similar study was done on Arabidopsis to identify the main types of pectins in guard cell walls. Localization of pectins in guard cell walls of Arabidopsis is similar to mosses in the stage they can move, with homogeneous walls rich in arabinan pectins that are required for wall flexibility. This study extends knowledge of pectin composition from stomata of the moss Funaria with limited stomatal movement to an angiosperm in which stomatal activity is crucial to the physiological health of the plant. The fourth study describes stomata development and internal changes in sporophyte anatomy that lead to formation of air spaces in the moss Funaria. Developing sporophytes at different stages were examined using light, fluorescence and electron microscopy; immunogold-labeling was used to investigate the presence of pectin in the newly formed cavities. Stomata in mosses do not develop from a self-generating meristemoid like in Arabidopsis, but instead they originate from a protodermal cell that differentiates directly into a guard mother cell. Epidermal cells develop from protodermal or other epidermal cells, i.e., there are no stomatal lineage ground cells. This developmental pattern is congruent with the presence of a gene ortholog of FAMA, but not SPCH and MUTE, in Physcomitrella. The final study in this dissertation focuses on the enigmatic Sphagnum. Although true stomata are absent in early-divergent mosses, Sphagnum has specialized epidermal cells, pseudostomata, that partially separate but do not open to the inside. To further understand the structure, function and evolution of pseudostomata, capsule anatomy and ultrastructure of pseudostomata were detailed. As in moss stomata, pseudostomata wall architecture and behavior facilitate capsule dehydration, shape change, and dehiscence, supporting this common function. Unlike other moss stomata, pseudostomata collapse along their ventral walls and they lack a substomatal cavity. Similarities to true stomata include two modified epidermal cells with specialized cell walls that separate by cuticle deposition and respond to drying. Pseudostomata may be interpreted as modified stomata that suppressed substomatal cavity formation, which in turn eliminated pore development. However, clarification of the homology of pseudostomata and moss stomata will require genomic studies integrated with physiological and structural data. The studies described in this dissertation significantly advance our understanding of moss stomatal development and structure, and provide a comparison point to better evaluate the evolution of stomata. Moss capsule anatomy coupled with the exclusive existence of stomata on capsules supports the concept that stomata in moss are involve in gas exchange but also facilitate drying and dispersal of spores. Changes in wall architecture coupled with a decrease in total pectin explain the inability of mature stomata to move. Development and distribution of stomata in Funaria provides evidence of a direct and less elaborated mechanism for stomatal development than described in Arabidopsis. Resolving relationships among early land plants, especially hornworts and mosses, the only bryophyte groups with stomata, is critical to understanding stomata evolution. Evaluated together, the results of this dissertation are consistent with a single origin of stomata in land plants.

Abstract Previous reports on the effects of enhanced UV-B radiation on bryophytes have been equivocal. This study shows that mosses not only respond to enhanced UV-B, but they are affected by changes in ambient radiation. The studies were conducted with two model species common in northern environments; red-stemmed feather moss (Pleurozium schreberi) and juniper haircap moss (Polytrichum juniperinum). Both species showed high concentrations of methanol-extractable UV-absorbing compounds (UACs) with high spring-time and early-summer UV, whereas in P. juniperinum, the concentration was affected by early-summer drought. The UACs of P. juniperinum increased again towards autumn suggesting a role in winter hardening. The (spring-time) cell wall-bound UV screen was important to both species. The fundamental adaptation of P. juniperinum to open and exposed environments was reflected in relatively higher concentrations of total UACs compared to P. schreberi. The enhanced UV-B experiments in situ were conducted over two years in Oulu and six years at the FUVIRC site in Sodankylä. Some of the effects of UV-B were seen within the first years of the experiments, or even within hours, while others were observed after several years. Five or six years of enhanced UV-B treatment increased the methanol-extractable UACs of P. schreberi and decreased the green shoot growth of P. juniperinum. The immediate light environment was proposed to have an impact on the varying UAC concentrations. Some mitigating effects of UV-A were observed as well. Off-site measured, reconstructed and modelled UV radiation data was used for comparisons of light environment in situ, or when performing a reconstructive research with historical samples. The environmental sample banks can provide a useful tool to study past environmental conditions, and even reconstruct past radiation levels. It was shown in this study that UACs in P. schreberi and P. juniperinum have fundamental roles as UV-B screens in the cell walls, but there is also a variable response with the soluble fraction that reacts and adapts to the changes in UV radiation. The responses to increasing UV-B radiation vary in magnitude and in time. As P. schreberi and P. juniperinum possess circumboreal and cosmopolitan distributions, the effects of UV-B on these species and consequently on ecosystems has a broad application.

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Bachelors
Arts and Sciences
Liberal Studies

Alpine and arctic tree line expansion depends on the establishment of tree seedlings above the current tree line, which is expected to occur with climate warming. However, tree lines often fail to respond to higher temperatures. Other environmental factors are therefore likely important for tree seedling establishment. Above the tree line, establishing seedlings encounter existing vegetation such as bryophytes, which often dominate in arctic and alpine tundra. Bryophytes modify their environment in various ways and may mediate climate change effects on establishing tree seedlings, and with that tree line expansion. The aim of this thesis was to understand if and how the environment, in particular bryophytes, mediates the impact of climate change on tree seedling establishment at the alpine and arctic tree line. This was explored by reviewing literature on tree seedling establishment at alpine and arctic tree lines globally. In addition, tree seedling survival and growth of Betula pubescens and Pinus sylvestris were assessed experimentally. Here, individuals were planted into mono-specific mats of different bryophytes species and exposed to warming and different precipitation regimes. The literature review revealed that besides from temperature, tree seedling establishment is affected by a wide range of abiotic and biotic factors including water, snow, nutrients, light, disturbance and surrounding vegetation. Furthermore the review revealed that for example vegetation can change tree seedling responses to climate change. The experiments showed that especially tree seedling survival was adversely affected by the presence of bryophytes and that the impacts of bryophytes were larger than those of the climate treatments. Seedling growth, on the other hand, was not hampered by the presence of bryophytes, which is in line with earlier findings that seedling survival, growth and seed germination do not respond similarly to changes in environmental conditions. Moreover, we found several indications that vegetation above the tree line, including bryophytes, mediated tree seedling responses to warming and precipitation or snow cover. This thesis shows that temperature alone should not be used to predict future tree seedling establishment above the alpine and arctic tree line and that extrapolations from climate envelope models could strongly over or under estimate tree line responses to warming. This underlines the value of multi-factorial studies for understanding the interplay between warming and other environmental factors and their effects on tree seedling establishment across current tree lines.

Different histories for forests in Southern Africa have been hypothesized from vegetation biogeography and pollen analysis. However, the history of forests is still controversial. Phylogeography uses gene genealogies to infer history of distributions. Two forest endemic moss species were sampled: Leptodon smithii; and Neckera valentiniana. Two gene regions were used, trnLF (chloroplast genome) and ITS1 (nuclear genome). Neckera valentiniana showed no variation from the populations sampled. Results from Leptodon smithii based on the trn and ITS region suggest that forests once were widespread, but then became fragmented. Dispersal corridors still exist between Southern Cape populations and Western Cape populations.

In boreal and northern temperate forests, the moss layer and ectomycorrhizal (ECM) fungi play important roles in carbon and nutrient cycling. ECM mycelium is present in the lower parts of the moss layer, but little is currently known about the interaction between these two key components of northern forest ecosystems. This thesis aims to address this knowledge gap and to improve our understanding of the mechanisms through which the moss layer and ECM fungi influence carbon and nutrient cycling. Nutrient transfer between litter and Scots pine seedlings in symbiosis with the ECM fungus Paxillus involutus (Batsch) Fr. was investigated in highly controlled microcosm experiments using, for the first time, intact moss and pine litter. In addition, moss removal plots were established in a Scots pine forest which allowed measurement of processes involved in carbon (C) and nutrient turnover and related variables. There was a close, reciprocal exchange of carbon and nutrients between the host plant and ECM mycelium colonising moss and pine litter (Chapter 2). This was greatly enhanced by intensive colonisation of moss litter, suggesting that mosses provide a key source of nutrients for ECM fungi and may facilitate transfer of photosynthetic C belowground. During almost 2 years of decomposition, moss tissue released more nitrogen (N) but retained more C than pine litter (Chapter 3), further highlighting the importance of the moss layer in providing nutrients for overstorey trees, and in the accumulation of recalcitrant C in soil. In addition to contributing directly to C cycling through inputs of recalcitrant C in litter, the moss layer can influence C cycling indirectly, by increasing soil microbial activity; CO2 efflux was on average 1.4 times greater from soil under the moss layer than from soil covered only in pine litter (Chapter 3). The results suggest that an indirect influence can occur via two pathways: through an insulating effect of the moss layer on soil temperature, and through inputs of dissolved organic carbon (DOC) leached from moss (Chapter 4), both of which may stimulate activity of soil microbes. These findings demonstrate the importance of both the moss layer and ECM fungi in carbon and nutrient cycling in boreal and northern temperate forests, and indicate that mosses provide a key pathway through which nutrients may bypass sequestration in saprotrophic microbial biomass and be transferred directly from plant tissue to ECM fungi and overstorey trees.

Mosses are some of the first colonizers to disturbed sites, yet their roles in early plant community structuring are not well understood. The primary succession zones of volcanoes provide opportunities to conduct natural experiments into how mosses contribute to early plant community formation, as well as how the unique environments found in such zones affect plant traits, particularly those associated with stress tolerance. Though plant community changes have been well-documented since Mount St. Helens (MSH) volcano erupted in 1980, the volcano's moss assemblages, their influence on other plants, and their potential roles in chemical-mediated competition and biogeochemical cycling have garnered little attention. Using a natural stress gradient from primary to secondary succession zones on MSH, and in control and nutrient manipulated test plots, I sought to elucidate how populations of three dominant moss species, Polytrichum juniperinum, Ceratodon purpureus, and Racomitrium canescens, respond to abiotic stress, as well as to provide life history and interaction data on establishment stages of these stress tolerant taxa. I first analyzed possible tradeoffs in survival strategies of four moss communities in test plots along an abiotic stress gradient. In P. juniperinum, seta specific density (mg/mm) increased significantly in response to nitrogen (N) addition. Differences in both vegetative and sexual reproductive morphological measurements were dependent on site and did not correlate with abiotic stress. In C. purpureus, the percentage of total spores germinated increased with N addition. Site dependent responses in nutrient allocation to vegetative and reproductive structures may be a result of phenotypic plasticity alone or may be a result of local adaptation. In mosses adapted to environmental stress, the allocation of nitrogen must be balanced between growth and survival. Efficient nitrogen uptake confers a competitive advantage if allocated to the higher dispersal of quickly germinating spores. Second, my results show the moss R. canescens may be able to inhibit the germination rate of co-occurring moss spores when spores were germinated in moss gametophyte infusions. R. canescens may also inhibit the germination of the co-occurring vascular plant Lupinus lepidus when seeds are germinated within intact moss patches. By uncovering chemical-mediated interactions between mosses on the germination and initial growth of neighboring mosses and vascular plants we can gain a better understanding of the mechanisms stress tolerant plants may use to limit resource competition. Such advantages offer insight into how mosses effectively colonize and affect primary succession landscapes.

In the 1850s when the timber floating began in Sweden many streams were channelized, which meant that the streams were straightened out, side channels were closed, and stones, trees and boulders were moved out to the banks of the streams. These measures altered the morphology of the streams, causing impediments to water moving in from the stream to the riparian zone, and reduced flooding frequency. When the era of timber floating was over humans began to restore the channelized streams to restore the damaged systems, mainly to improve fish habitats. Channelization and restoration involve major disruptions to the stream ecosystem, and the purpose of this study was to investigate how the bryophyte communities in the riparian zone differed in diversity, species richness and areal coverage, depending on the form of disturbance they were exposed to and the time passed since the disturbance occurred. This study was conducted along tributaries to the Vindel River in northern Sweden. There were three types of tributaries: partly channelized streams, best-practice restored streams and demonstration restored streams. At all tributaries included in the study, bryophyte samples were collected at two heights from the water surface, 0 and 40 cm. These samples were then identified to family or species level and their areal coverage was estimated. The results show that the demonstration restored streams had the lowest species richness and, there were no significant differences in diversity and species richness between the channelized streams and the best-practice restored streams. Regarding areal coverage, there was no significant difference between the stream types. The results also show that the time plays an important role for recovery of the bryophyte communities since diversity and species richness increased with the time since the disturbance occurred.
Under 1850-talet då flottning av timmer påbörjades i Sverige startade kanaliseringen av många vattendrag, vilket innebar att vattendragen rätades ut, sidokanaler stängdes, och stenar, träd och stenblock flyttades ut till vattendragens kanter. Dessa åtgärder förändrade vattendragens morfologi, orsakade fördämningar mellan vatten och strandzon och minskade översvämningsfrekvensen. När timmerflottningens era var slut började människan restaurera de kanaliserade vattendragen, främst för att det visat sig att fisket försämrats. Kanalisering och restaurering innebär stora störningar för ekosystemet de utförs i och denna studie syftar till att titta hur mossamhällen i strandzonen skiljer sig åt i diversitet, artrikedom och täckning beroende på vilken form av störning de utsatts för och tiden sedan de utsattes för störningen. Studien är gjord längs biflöden till Vindelälven i norra Sverige. Biflödena är dels kanaliserade vattendrag, bästa-praxis restaurerade vattendrag och demonstrationsåtgärdade vattendrag. Vid samtliga sträckor som ingår i studien har mossprover samlats in på två höjder från vattenytan, 0 och 40 centimeter. Dessa prover har sedan identifierats till lägsta möjliga taxonomiska nivå (art eller familj), och deras täckningsgrad har uppskattats. Resultaten visade att de demonstrationsåtgärdade vattendragen hade lägst artrikedom och diversitet och mellan kanaliserade vattendrag och bästa-praxis vattendrag kunde ingen signifikant skillnad i diversitet eller artrikedom påvisas.  Gällande täckningsgraden fanns ingen signifikant skillnad mellan vattendragstyperna. Resultatet visade också att tiden spelar en viktig roll för återhämtningen hos mossamhällena då artrikedom och diversitet ökar med tiden sedan inträffad störning.

Conselho Nacional de Desenvolvimento Científico e Tecnológico
A radiação ultravioleta (UV) induz diversos efeitos nocivos nos organismos e a quantidade desta radiação que atinge a biosfera é afetada pela concentração de ozônio, latitude, altitude, clima e reflexão especular. As respostas de briófitas em relação aos efeitos da radiação UV e a presença de compostos que absorvem esta radiação têm sido estudadas. Sanionia uncinata, Holomitriopsis laevifolia e Leucobryum laevifolium são espécies de musgos encontrados em locais expostos a alta incidência de radiação UV e com habitats distintos. Considerando que as respostas de musgos contra os efeitos da radiação UV e seus mecanismos de proteção ainda são pouco caracterizados, o objetivo deste estudo foi investigar o potencial fotoprotetor e possíveis riscos toxicológicos associados aos extratos dos musgos S. uncinata, proveniente da Antártica e H. laevifolia e L. laevifolium, proveniente do Amazonas. Seus extratos metanólico (EM), aquoso (EA), hidroalcoólico (EH) e etanólico (EE) foram estudados com a caracterização química por absorção ao UV e visível e pela cromatografia líquida de alta eficiência; quantificação do índice total de compostos fenólicos; determinação da capacidade captadora do radical 2,2-difenil-1-picril-hidrazila a fim de avaliar as atividades antioxidantes; avaliação do potencial de fotoproteção cutânea pela determinação do fator de proteção solar; avaliações do potencial mutagênico e citototóxico, através do ensaio de Salmonella/microssoma, utilizando as cepas TA97, TA98, TA100, TA102 e TA104; do potencial fotomutagênico através do ensaio de fotomutagenicidade, usando as cepas TA102 e TA104; e investigação dos efeitos genotóxicos e fotogenotóxicos, pelo ensaio de micronúcleo e fotomicronúcleo, respectivamente, usando diferentes linhagens celulares estabelecidas. Foram encontradas atividades fotoprotetoras e antioxidantes e observou-se que os extratos se apresentaram singulares devido a sua composição química. Os resultados fotoprotetores, além dos mutagênicos/fotomutagênicos, genotóxicos/fotogenotóxicos e suas respectivas avaliações citotóxicas também permitiram selecionar extratos e suas concentrações, como promissores candidatos em fotoproteção Assim, os EA e EH de H. laevifolia e L. laevifolium apresentam, no geral, os resultados mais significativos, tornando-se potenciais para avaliações refinadas em fotoproteção e na separação de componentes que possam levar a futuras aplicações como antioxidantes e protetores solares ou como adjuvantes.
The ultraviolet radiation (UV) induces many harmful effects in all living organisms and the amount of this radiation that reaching the ground is affected by many factors including ozone concentration, latitude, altitude, climate and specular reflection. The responses of bryophytes against the effects of UV radiation and the presence of compounds that absorb the UV region have been studied. Mosses Sanionia uncinata, Holomitriopsis laevifolia and Leucobryum laevifolium are found in locations exposed to UV at high levels of radiation and in different habitats. Whereas that the responses of mosses against the effects of UV radiation and their protection systems are poorly characterized yet, the aim of this study was to investigate photoprotective potential and possible toxicological risks associated with extracts of mosses S. uncinata (from Antarctica) and H. laevifolia and L. laevifolium (from Amazônia). Methanol (ME), aqueous (AE), hydroalcoholic (HE) and ethanolic (EE) were studied by: chemical characterization by UV/visible spectrophotometry and by High performance liquid chromatography; phenolic content estimation; 2,2-diphenyl-1-picrylhydrazyl scavenging activity; potential of skin photoprotection by in vitro determination of sun protection factor; the mutagenic potential, and cytotoxic by Salmonella/microsome assay, using the TA97, TA98, TA100, TA102 and TA104 strains; photomutagenic potential by photomutagenicity test, using TA102 and TA104 strains and; investigation of genotoxic and photogenotoxic effects by micronucleus test and photo-micronucleous assay, respectively, using different established cell lines. Photoprotective and antioxidant activities were found and it was observed that the extracts showed strong uniqueness due to its chemical composition. From the photoprotective, mutagenic/photomutagenic and genotoxic/photogenotoxic results and their cytotoxic evaluations it was possible to select extracts and their concentrations as promising candidates for photoprotection. Thus, the EA and EH of H. laevifolia and L. laevifolium demonstrated the most significant results, becoming potential for refined evaluations in photoprotection and separating components that can lead to future applications such as sunscreens and antioxidants or as adjuvants.

Oxygenic photosynthesis is a crucial process for life on earth as it enables plants, algae and cyanobacteria to convert sunlight into chemical energy, generating molecular oxygen as a byproduct. Light can also be harmful and excess light intensities can drive to photosystems over-excitation, production of reactive oxygen species (ROS) and cell damage. Thus, plants and algae evolved several photoprotective processes to survive in a variable environment. The fastest one among them is called Non Photochemical Quenching (NPQ), which consists in the thermal dissipation of excess energy absorbed triggered by the generation of a ΔpH gradient across thylakoid membranes. The main and fastest component of NPQ is called “Energy quenching” or “feedback de-excitation” (qE) and it is activated in a few seconds upon an increase in light irradiation. Although widespread among oxygenic photosynthetic organisms, NPQ shows important differences in its machinery: in vascular plants as Arabidopsis thaliana it depends on the presence of the Lhc-like protein PSBS, while a different polypeptide, LHCSR (previously called Li818), is required in algae as the Chlorophyta Chlamydomonas reinhardtii and diatoms. In this work we used the moss Physcomitrella patens as a new model organism for the study of photosynthesis and NPQ in particular. The choice of this moss is justified by several reasons. First of all, Physcomitrella patens diverged from higher plants soon after land colonization and its analysis can provide information on adaptation to terrestrial environment and evolution of NPQ process from unicellular algae to higher plants. Consistent with its peculiar position in evolution of photosynthetic organisms, in Physcomitrella genome both PSBS and LHCSR were identified. A further reason to study this moss is that Physcomitrella is able to integrate efficiently foreign DNA by homologous recombination (gene targeting), which is a unique feature among plants. Thus, it is relatively simple to obtain targeted knock-out plants with higher precision and frequency compared with seed plants, such as Arabidopsis. We generated P. patens knock-out (KO) mutants depleted in PSBS and/or LHCSR proteins (the ones involved in vascular plants and algae NPQ, respectively) exploiting Physcomitrella ability of doing homologous recombination. We obtained single KO mutants depleted in PSBS (psbs KO) and in each of the two isoforms of LHCSR present in Physcomitrella genome (lhcsr1KO and lhcsr2KO), as well as double KO mutants (psbs lhcsr1 KO, psbs lhcsr2 KO and lhcsr1 lhcsr2 KO) and the triple KO mutant, depleted in all the three proteins analyzed (psbs lhcsr1 lhcsr2 KO). All single and double KO mutants showed a decreased NPQ capacity compared with WT mosses, but all of them retain a residual NPQ capacity, still identifiable as qE, demonstrating that both PSBS and LHCSR play a role in Physcomitrella NPQ, with LHCSR1 having the largest effect. Only the triple KO depleted in all PSBS and LHCSR isoforms showed a completely abolished NPQ capacity. Its phenotype is very close to the one of Arabidopsis plants depleted in PSBS, again clarifying that both PSBS- and LHCSR-dependent NPQ mechanisms are indeed active in this moss. The different KO mutants were also grown in different light and temperature conditions to evaluate the effect of the NPQ reduction. The ones having the strongest reduction in NPQ also showed the strongest photoinhibition, demonstrating that NPQ capacity is important for Physcomitrella photoprotection. When grown in high light conditions for several days WT mosses showed an enhanced NPQ which was correlated with the increased expression of both PSBS and LHCSR. The modulation of PSBS or LHCSR accumulation is thus a method for modulation NPQ and provide optimal photoprotection according to the growth conditions. We also over-expressed (OE) PSBS and LHCSR proteins in the KO lines previously obtained, and we were able to show that in P. patens PSBS and LHCSR are alone capable of activating a strong NPQ. Their activity appears to be largely independent, suggesting a different and autonomous activation mechanism. The carotenoid zeaxanthin has a fundamental role in several protection mechanisms and its synthesis also enhances NPQ. We also showed that although a part of NPQ is induced also in the absence of zeaxanthin, the synthesis of this carotenoid is fundamental for the activation of a strong response in Physcomitrella. Zeaxanthin presence enhance similarly both PSBS and LHCSR–dependent NPQ but it is unable to induce any quenching in their absence. In this thesis we exploited Physcomitrella ability of gene targeting also to express mutated form of PSBS protein. We obtained two deleted isoforms, one with a N-terminal deletion while the other truncated in the C-terminus, as these regions have conserved basic and acidic residues, respectively. PSBS function is likely driven by interaction with other proteins and these charged residues may be essential for these protein-protein interactions and thus for PSBS activity. The expression of mutated PSBS proteins in psbs KO genotype complemented the effect of PSBS knocking-out, however, demonstrating that the deleted regions are not required for PSBS function. As mentioned above, the study of P. patens can provide information on the adaptation to terrestrial environment. To obtain additional information we also analyzed NPQ in some Streptophyta algae (also known as Charophyta). Only the groups which diverged later with respect to plants, Coleochaetales and Charales, show the presence of a PSBS-dependent NPQ. The analysis of these algae, together with the one of Physcomitrella patens and literature data allows proposing an hypothesis for the evolution of NPQ. In correspondence to land colonization a new mechanism for NPQ, dependent on PSBS, evolved. This was at first in addition to the one dependent on LHCSR already present in green algae, which was subsequently lost by vascular plants.
La fotosintesi riveste un’importanza fondamentale per la biosfera: grazie a questo processo l’energia solare è utilizzata per fissare l’anidride carbonica atmosferica in biomassa, liberando ossigeno. Piante e alghe, però, sono molto spesso esposte a condizioni d’illuminazione molto variabili. In condizioni di elevata illuminazione è fondamentale che l’energia in eccesso venga dissipata, pena la formazione di specie radicaliche dell’ossigeno (ROS) molto dannose per la cellula. Uno dei principali sistemi di protezione dall’eccesso di luce viene attivato pochi secondi dopo una variazione di illuminazione ed è definito Non Photochemical Quenching (NPQ). L’NPQ permette di dissipare l’eccesso di stati eccitati della clorofilla come calore ed è attivato a seguito dell’acidificazione del pH lumenale. L’NPQ comprende diverse componenti, la più rapida delle quali è definita qE (da “Energy quenching” o “Feedback de-excitation”). L’NPQ è presente in tutti gli organismi fotosintetici anche se meccanismi di attivazione e proteine coinvolte sono diversi. Nelle piante vascolari (quali la pianta modello Arabidopsis thaliana) è noto che l’NPQ dipende da una proteina presente nelle membrane dei tilacoidi chiamata PSBS. Nelle alghe, quali diatomee o la Clorofita Chlamydomonas reinhartii, che pure sono in grado di attivare NPQ, PSBS è assente e l’attivazione dipende da una proteina diversa, detta LHCSR. In questo lavoro è stato utilizzato il muschio Physcomitrella patens come organismo modello per lo studio della regolazione della fotosintesi. Questa scelta è dettata da molteplici ragioni. Le briofite presentano un’interessante posizione evolutiva, dal momento che si sono separate dalle piante vascolari poco dopo la colonizzazione delle terre emerse: il loro studio può quindi fornire informazioni su quelli che sono i primi adattamenti degli organismi fotosintetici alla vita terrestre. Per quanto riguarda i meccanismi di fotoprotezione, Physcomitrella patens ha nel proprio genoma sia PSBS che LHCSR, le proteine indispensabili per l’NPQ nelle piante vascolari e nelle alghe. Physcomitrella, inoltre, presenta la capacità unica tra le piante analizzate fino ad ora di fare ricombinazione omologa, facilitando così la possibilità di ottenere mutanti specifici per le proteine di interesse. Nel particolare, in questo lavoro abbiamo ottenuto dei mutanti specifici per PSBS e LHCSR, le proteine caratteristiche dell’NPQ di piante ed alghe rispettivamente. Sfruttando la capacità di ricombinazione omologa di Physcomitrella, sono stati generati mutanti knock-out (KO) per PSBS (psbs KO) e le due isoforme di LHCSR (LHCSR1 e LHCSR2; lhcsr1 KO e lhcsr2 KO) presenti nel genoma del muschio. Queste mutazioni sono poi stati combinate in doppi KO (psbs lhcsr1 KO, psbs lhcsr2 KO and lhcsr1 lhcsr2 KO) ed in un triplo KO (psbs lhcsr1 lhcsr2 KO), genotipo in cui è stato eliminato il gene di tutte tre le proteine in analisi. La risposta di NPQ nelle varie linee di singoli e doppi KO ha dimostrato come tutti questi mutanti diminuiscano la loro capacità di indurre NPQ rispetto a muschi WT. Ciò nonostante, tutte queste linee sono comunque in grado di attivare una risposta qE, dimostrando che queste proteine non solo sono presenti nel genoma ma sono entrambe attive nell’NPQ. Inoltre, l’isoforma LHCSR1 risulta essere la maggiore responsabile dell’NPQ in questa specie. Al contrario, mutanti privi dei tre polipeptidi (triplo KO) non sono in grado di indurre alcuna risposta qE, con un fenotipo simile ai mutanti di Arabidopsis privi di PSBS. In Arabidopsis PSBS è l’unica proteina coinvolta nel qE, dal momento che le piante vascolari non possiedono geni per LHCSR, confermando ulteriormente che in Physcomitrella sia PSBS che LHCSR sono attive nell’NPQ. Gli effetti di una diminuita capacità di indurre NPQ sulla fotoprotezione di P. patens sono stati ulteriormente approfonditi analizzando la capacità di acclimatazione ad alta luce o basse temperature nel muschio WT e nelle varie linee di mutanti KO ottenute. I mutanti che già in condizioni di controllo mostravano la maggiore diminuzione di NPQ in queste condizioni di crescita mostrano anche la maggiore fotoinibizione, dimostrando l’importanza dell’NPQ per la fotoprotezione. Nei muschi WT, l’acclimatazione ad alta luce o basse temperature, infatti, è accompagnata da un aumento della capacità di attivare la risposta di NPQ che correla con un aumento dell’espressione sia di PSBS che LHCSR. Questi dati dimostrano che Physcomitrella è in grado di modulare la sua capacità di indurre NPQ modificando l’espressione delle proteine chiave di questo processo, PSBS e LHCSR, per ottenere una fotoprotezione adeguata alle condizioni di crescita in cui si trova. In aggiunta a mutanti KO per PSBS e LHCSR, durante questo lavoro sono state generate anche delle linee che over-esprimono (OE) queste proteine. Questi mutanti hanno permesso di dimostrare che sia PSBS che LHCSR sono in grado di attivare una risposta NPQ in Physcomitrella in assenza dell’altra proteina. Questo risultato dimostra che PSBS e LHCSR sono attive in modo indipendente l’una dall’altra, ed hanno quindi verosimilmente dei meccanismi di attivazione differenti ed autonomi. Anche i carotenoidi, e la zeaxantina in particolare, hanno un ruolo fondamentale nella protezione delle cellule dagli stress ossidativi dal momento che sono coinvolti in diversi meccanismi di fotoprotezione. Tra gli altri, la zeaxantina è in grado di amplificare la risposta NPQ in piante quali Arabidopsis. In Physcomitrella abbiamo ottenuto lo stesso risultato, dimostrando che la presenza di questo carotenoide è fondamentale per una completa attivazione di NPQ sia PSBS che LHCSR dipendente. Se queste due proteine non sono presenti, invece, il solo accumulo di zeaxantina non è in grado di attivare alcuna risposta di fotoprotezione rapida qE. Durante questo progetto di Dottorato è stata sfruttata la capacità di ricombinazione omologa di Physcomitrella anche per l’espressione di isoforme mutate di PSBS. Nel particolare, sono state ottenute le sequenze codificanti per PSBS con una delezione all’N- o al C- terminale della proteina matura. Si tratta di regioni conservate in PSBS di diverse specie di piante, che presentano rispettivamente carica positiva e negativa, e potrebbero quindi essere coinvolte in interazioni proteina-proteina che sembrano essere necessarie per il meccanismo di induzione NPQ PSBS-dipendente. L’espressione di queste isoforme in muschi psbs KO, però, ha portato alla complementazione del fenotipo in entrambi i casi, dimostrando che le regioni delete non sono fondamentali per l’attività di PSBS. Nell’ultima parte del lavoro, inoltre, abbiamo focalizzato l’attenzione sull’evoluzione del meccanismo di NPQ da alghe a piante. Come accennato in precedenza, Physcomitrella presenta una posizione evolutiva chiave per ottenere informazioni sugli adattamenti necessari alla colonizzazione delle terre emerse ed abbiamo esteso l’analisi anche ad alghe della linea Streptofita (alghe Carofita) per ottenere maggiori informazioni sull’evoluzione dell’NPQ. I dati ottenuti hanno dimostrato che nelle specie evolutivamente più vicine alle piante terrestri (Coleochatales e Charales, identificati come i probabili “sister group” delle piante terrestri) PSBS è presente. Queste analisi, unitamente a quanto riportato in precedenza per P. patens e a dati di letteratura, permettono di formulare un’ipotesi sull’evoluzione dell’NPQ da alghe a piante: in corrispondenza della colonizzazione delle terre emerse è stato evoluto un nuovo meccanismo di fotoprotezione (PSBS dipendente), che dapprima si è affiancato, poi sostituito completamente a quello LHCSR-dipendente tipico delle alghe.

Abstract The thesis consists of regional forest condition studies, using different biomonitors. Heavy metal deposition was investigated in 1985–2000 on the basis of the heavy metal concentrations (As, Cd, Cr, Cu, Fe, Hg, Ni, Pb, V, Zn) in mosses in Finland. A comparison on the suitability of mosses, epiphytic lichens and pine bark as biomonitors of heavy metals was also carried. Bark was also used to study the dispersal of emissions from the Kola Peninsula into northern Finland. The occurrence of green algae on conifers in Finland was investigated in 1985 and 1995. Regional and temporal differences were found in the heavy metal concentrations of mosses in Finland. The concentrations of most metals were the highest in southern Finland, and they decreased towards the north. Some of the major emission source had a noticeable effect on the Cu, Ni and Cr concentrations of mosses in the surroundings of the emission sources. The Pb, Cd and V concentrations decreased the most during the study period. Mosses, lichens and bark gave a relatively similar result for heavy metal deposition in Finland. However, the comparisons indicated that mosses are better suited as biomonitors for regional surveys than epiphytic lichens, because the regional differences in heavy metal deposition were more readily reflected by concentrations in mosses than in lichens. Bark is relatively unsuitable for regional surveys due to the small range of variation in the concentrations. Emissions from the Kola Peninsula had a clear effect on the sulphur and heavy metal concentrations of pine bark. The concentrations in bark were at very high levels close to the smelters, but they rapidly decreased on moving towards the west. The effects of emissions were still clearly visible in north-eastern Lapland. There was strong increase in the abundance of green algae on conifers in southern and central Finland during the period 1985–1995. The increase is probably due to following factors: climate warming, and an increase in nitrogen and a decrease in sulphur in their habitats. Half of each biomonitor sample collected in the surveys has been stored in the specimen bank at Paljakka. The storage of samples offers advantages for monitoring purposes. The availability of long-term sample series makes it possible to construct retrospective time series of the pollutants. The specimen bank is to be further developed in the future by establishing a reputation as a storage facility for samples related to forest ecosystems.

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Made available in DSpace on 2016-09-16T14:48:23Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) Comunidade vegetais de áreas de degelo da Ilha Half Moon, Arquipélago das Shetlands do Sul, Antártica.pdf: 769779 bytes, checksum: 794a3b4b95c7a64c38ab5708c8c281b2 (MD5) Previous issue date: 2015-07-28
Este trabalho apresenta os resultados do estudo das comunidades vegetais de áreas de degelo realizado durante os verões austrais de 2013/2014 e 2014/2015, nos meses de fevereiro e março, na Ilha Half Moon, no Arquipélago Shetlands do Sul, Antártica Marítima, com o objetivo de avaliar a distribuição e abundância de musgos e líquens, bem como para descrever e mapear as comunidades de plantas que ocorrem na área. As plantas foram correlacionadas com a ocorrência de aves marinhas, relevo e umidade para entender sua influência sobre a distribuição de plantas. Para aferir as comunidades existentes na ilha através da fitossociologia e quantificar as espécies com maior significância ecológica (IES) foi utilizada a metodologia de quadrados (20 x 20cm) de Braun-Blanquet adaptado às condições Antárticas. Para o mapeamento da área foi utilizado o DGPS Astech Promark II®, com precisão sub-metrica e posteriormente processados no software Astech Solutions®. Foram identificadas 37 espécies de briófitas, 59 liquens, uma angiosperma, a Deschampsia antarctica Desv., e duas espécies de algas macroscópicas. Foram dispostos 358 quadrados e identificadas cinco comunidades vegetais na ilha, distribuídas de acordo com a ocorrência de animais e a influência marítima. As comunidades descritas foram as seguintes: Comunidade Liquens fruticulosos, Comunidade carpete de musgos, Comunidade de Liquens Muscicolas, Comunidade de Liquens crustosos e Comunidade Turfa de musgos. A espécie que apresentou o IES mais elevado da ilha foi Sanionia uncinata (Hedw.) Loeske aparecendo em 71.78% dos quadrados amostrados. A comunidade que apresentou maior número de espécies foi a Carpete de musgos e a mais diversa foi a comunidade liquens crustosos, que tem sua maior parte localizada na área onde estão as colônias de pingüins.
This paper presents the results of the study of plant communities of free ice areas conducted during the austral summer of 2013/2014 and 2014/2015, in February and March, on Half Moon Island in the South Shetlands Archipelago, Maritime Antarctica, aiming to evaluate the distribution and abundance of mosses and lichens, as well as to describe the plant communities occurring in the area, with mapping of the vegetation communities. To assess existing communities on the island through the phytosociology and quantify the species with the highest ecological significance (IES) was used the methodology square (20 x 20cm) of Braun-Blanquet adapted to Antarctic conditions. . The area was mapped using an Astech Promark II® DGPS, yielding sub-metric precision after post-processing with Astech Solutions® software. The number of species totalized 38 bryophyte species, 59 lichens, only one flowering plant (Deschampsia antarctica Desv.) and two macroscopic terrestrial algae. Five plant communities were identified in the island, distributed accordingly to animal occurrence and to sea influence. The plant communities were described as follows: Fruticose lichens community, Moss-carpet community, Muscicolous Lichens community, Crustose lichens community and Moss turf community. The species with the highest IES of the island was Sanionia uncinata (Hedw.) Loeske appearing in 71.78% of sampled squares. The community with the highest richness species was the Moss Carpet, and the most diverse community was the crustosos lichens, which has mostly located in the area where the penguin colonies.