Population structure of mycobionts and photobionts of the widespread lichen Cetraria aculeata

Lichens are symbioses between fungi (mycobionts) and photoautotrophic green algae or cyanobacteria (photobionts). Many lichens occupy large distributional ranges covering several climatic zones. So far, little is known about the large‐scale phylogeography of lichen photobionts and their role in shaping the distributional ranges of lichens. We studied south polar, temperate and north polar populations of the widely distributed fruticose lichen Cetraria aculeata. Based on the DNA sequences from three loci for each symbiont, we compared the genetic structure of mycobionts and photobionts. Phylogenetic reconstructions and Bayesian clustering methods divided the mycobiont and photobiont data sets into three groups. An amova shows that the genetic variance of the photobiont is best explained by differentiation between temperate and polar regions and that of the mycobiont by an interaction of climatic and geographical factors. By partialling out the relative contribution of climate, geography and codispersal, we found that the most relevant factors shaping the genetic structure of the photobiont are climate and a history of codispersal. Mycobionts in the temperate region are consistently associated with a specific photobiont lineage. We therefore conclude that a photobiont switch in the past enabled C. aculeata to colonize temperate as well as polar habitats. Rare photobiont switches may increase the geographical range and ecological niche of lichen mycobionts by associating them with locally adapted photobionts in climatically different regions and, together with isolation by distance, may lead to genetic isolation between populations and thus drive the evolution of lichens.     

Coalescence-based species delimitation using genome-wide data reveals hidden diversity in a cosmopolitan group of lichens

Although there is an abundance of species delimitation methods on the market, most approaches depend on predefined assignment of specimens to species or populations. Assignment-free methods, which can simultaneously infer boundaries and relationships among species, are of high importance in cases, when correct pre-assignment is difficult or not at all possible. In this study, we use assignment-free multispecies coalescent-based species delimitation (STACEY, tr2-delimitation, and BP&P), phylogenetic methods, and clustering algorithms to investigate the inter- and infraspecific relationships within a common and widespread group of lichens with contentious species boundaries. The Cetraria aculeata group presents a good example of extreme morphological variability and unclear species delimitation in lichens. Based on DNA-sequence data from 26 fungal loci and 10 microsatellite loci, as well as morphological and chemical data, our results provide evidence for the occurrence of five different taxa within the group and highlight the difficulties of morphologically distinguishing these species. We discovered a separate lineage (clade C) within C. aculeata s. str., which does not fully coincide with any of the a priori identified species C. aculeata, C. crespoae, or C. steppae and conclude that this clade constitutes a semi-cryptic, genetically isolated lineage within C. aculeata. We recognize this lineage at subspecific rank as C. aculeata subsp. steppae and synonymize Cetraria crespoae with C. aculeata subsp. aculeata. Epitypes are designated for all involved names to stabilize their usage. The PKS8 gene locus is recommended as a barcode for the separation of C. aculeata subsp. aculeata and subsp. steppae. We demonstrate the potential use of microsatellite data for species delimitation in lichens that might offer an alternative insight or be used to test species delimitation hypotheses, when dealing with closely related or potentially cryptic species. Our results also confirm the presence of an undescribed sister lineage to C. odontella previously misidentified as C. muricata and extend the known range of this lineage to Central Asia (Altay Mts.) and the Central European Alps (France, Switzerland), which calls for a critical reappraisal of records of C. aculeata and C. muricata from these mountain ranges.

     

Antioxidant and antimicrobial activity of some lichen species

The aim of the research is to explore the overall in vitro antioxidant activity, total phenol content, reduction power and antimicrobial activity of the methanol extracts of the lichens Cetraria pinastri, Cladonia digitata, Cladonia fimbriata, Fulgensia fulgens, Ochrolechia parella and Parmelia crinita. The methanol extract of the Cetraria pinastri showed a strong antioxidant activity, whereas the extracts of the species Fulgenesi fulgens, Cladonia fimbriata and Parmelia crinita showed the moderate one and the extract of the species Ochrolechia parella and Cladonia digitata the weak one. The methanol extract of the lichen Cetraria pinastri had the biggest total phenol content (32.9 mg/g of the dry extract). A certain correlation was established between the antioxidant activity and the total phenol content for the researched lichen extracts. The work also explores the antimicrobial activity of the methanol extracts of the mentioned species of lichens against six bacterial and eleven fungi species by the disc-diffusion method and by establishing the minimum inhibitory concentration (MIC). The methanol extracts of the lichens Cetraria pinastri and Parmelia crinita showed the strongest both antibacterial and antifungal activity against most of the tested microorganisms. These researches suggest that the lichens Cetraria prunastri can be used as new sources of the natural antioxidants and the substances with antimicrobial features.     

Lecanoric acid,a secondary lichen substance with antioxidant properties from <Emphasis Type="Italic">Umbilicaria antarctica</Emphasis> in maritime Antarctica (King George Island)

Eight lichen species, Cetraria aculeata, Cladonia furcata, Pseudephebe pubescens, Sphaerophorus globosus, Stereocaulon alpinum, Umbilicaria antarctica, Usnea antarctica and Usnea aurantiacoatra, were collected from King George Island, maritime Antarctica, for the evaluation of antioxidant activities. Anti-linoleic acid peroxidation activity, free radical scavenging activity, reducing power and superoxide anion scavenging activity were assessed of methanol and acetone extract of the lichens in vitro. Extract of Umbilicaria antarctica, Cladonia furcata, Sphaerophorus globosus and Usnea antarctica were found to have strong in vitro antioxidant properties. In general, acetone extract exhibited stronger activities than methanol extract. The activity-guided bioautographic TLC and HPLC analysis demonstrated that lecanoric acid was the main antioxidant compound in the acetone extract of Umbilicaria antarctica, the most potent antioxidant lichen species among the test species. The results suggested that several Antarctic lichens and their substances can be used as novel bioresources of natural antioxidants.     

Phylogeny of cetrarioid lichens (Parmeliaceae) inferred from ITS and b-tubulin sequences,morphology, anatomy and secondary chemistry

Phylogenetic relationships within the family Parmeliaceae (lichenized ascomycetes) with emphasis on the heterogeneous group of cetrarioid lichens are reconstructed. The results are based on cladistic analyses of DNA-sequences, morphological and chemical data. Almost all currently recognized cetrarioid genera were included in the analyses together with parmelioid and alectorioid members of the presumably monophyletic family Parmeliaceae. We tried to sample taxonomic diversity of the family as widely as possible. The ITS1-5.8S-ITS2 region of the rDNA and a partial β-tubulin gene from 126 samples representing 82 species were analysed. Cetrarioid lichens were identified as a monophyletic group, supported by both ITS and β-tubulin characters. This group was reanalysed using 47 morphological, anatomical and secondary chemistry characters combined with the DNA data matrix. ITS and β-tubulin sequences provide congruent information, and a clear correlation between DNA-data and conidial shape is observed. The current taxonomy of the cetrarioid lichens is discussed and compared with the phylogenetic trees obtained here. A comprehensive study of the phylogeography of some bipolar or subcosmopolitic species with representatives from both hemispheres was performed. Cetraria aculeata is the only taxon where correlation between DNA-data and geographic origin is observed.     

Lipid and Fatty Acid Composition of Tree-Growing and Terrestrial Lichens

A comparison of lipid and fatty acid composition was made of the tree-growing lichens Evernia prunastri (L.) Ach., Parmelia saxatilis (L.) Ach. and Hypogymnia physodes (L.) Ach. and the terrestrial species Cetraria islandica (L.) Ach. and Cladonia impexa Harm. In the terrestrial species the total lipid content varied strongly during spring, while the tree-growing species showed much less variation. Phospholipid and sterol content of all lichens was unusually low. Monoglycosyl diglyceride was absent from Parmelia saxatilis. Fatty acids common to higher plants as palmitic, stearic, oleic, linoleic and linolenic acid were invariably present in all lichen species. In addition large quantities of extra-long chain fatty acids like behenic acid, eicosadienoic acid and cyclic aliphatic lichen acids were present in the terrestrial species. The degree of (poly) unsaturation decreased in the order Evernia prunastri, Parmelia saxatilis. Fatty acids common to higher plants as palmitic, stearic, impexa, which decrease was compensated by an increase in extralong chain fatty acids and lichen acids. It is suggested that the lichen acids are of adaptive value for lichen species growing in the terrestrial habitats, which were characterized by extreme diurnal temperature variations. Just as the polyunsaturated fatty acids, lichen acids guarantee at lower temperatures a high flexibility of the membranes involved, at the same time as they are less susceptible to photo-oxidation at the high daytime temperatures.     

No need for stepping stones: Direct,joint dispersal of the lichen‐forming fungus Mastodia tessellata (Ascomycota) and its photobiont explains their bipolar distribution

Aim

The hypotheses proposed to explain the high percentage of bipolar lichens in Antarctica have never been explicitly tested. We used the strictly bipolar, coastal lichenized fungus Mastodia tessellata (Verrucariaceae, Ascomycota) and its photobionts (Prasiola, Trebouxiophyceae, Chlorophyta) as model species to discern whether this extraordinary disjunction originated from vicariance or long‐distance dispersal.

Location

Coasts of Antarctica, Tierra del Fuego (Chile), Alaska (USA) and British Columbia (Canada).

Methods

Based on a comprehensive geographical (315 specimens and 16 populations from Antarctica, Tierra del Fuego and North America) and molecular sampling (three and four loci for the fungus and algae respectively), we implemented explicit Bayesian methods to compare alternative hypotheses of speciation and migration, and performed dating analyses for the fungal and algal partner, in order to infer the timing of the colonization events and the direction of gene flow among distant, disjunct areas.

Results

Mastodia tessellata comprises two fungal species which in turn associate with three photobiont lineages along the studied distribution range. Independent estimation of divergence ages for myco‐ and photobionts indicated a middle to latest Miocene species split in the Southern Hemisphere, and a late Miocene to Pleistocene acquisition of the bipolar distribution. Comparison of migration models and genetic diversity patterns suggested an austral origin for the bipolar species.

Main conclusions

The complex evolutionary history of Mastodia tessellata s.l. can be explained by a combination of vicariant and long‐distance dispersal mechanisms. We provide novel evidence of a pre‐Pleistocene long‐term evolution of lichens in Antarctica as well as for bipolar distributions shaped by Southern to Northern Hemisphere migratory routes without the need for stepping stones.     

Diversity and succession of epiphytic macrolichen communities in low‐elevation managed conifer forests in Western Oregon

Abstract. We examined epiphytic macrolichen communities in Pseudotsuga menziesii (Douglas‐fir) forests across the western Oregon landscape for relationships to environmental gradients, stand age and structure, and commercial thinning. We used a retrospective, blocked design through the Coast and the western Cascade ranges of Oregon. Each of our 17 blocks consisted of a young, unthinned stand (age 50–110 yr); an adjacent, thinned stand of equivalent age; and an old‐growth stand (age > 200 yr). We found 110 epiphytic macrolichen taxa in the stands. Forage‐providing alectorioid lichens and the nitrogen‐fixing cyanolichen Lobaria oregana associated strongly with old‐growth stands and remnant old trees in younger stands (unthinned + thinned). Relative to unthinned stands, thinned stands had a slightly higher abundance of alectorioid lichens and a greater presence of Hypogymnia imshaugii. However, thinned stands hosted a lower landscape‐level (γ) diversity, lacking many species that occurred infrequently in the unthinned stands. Patterns in the lichen community composition correlated strongly with climatic gradients; the greatest variation in composition was between the Coast and Cascade ranges. The difference in communities between mountain ranges was greatest among stands 70–110 yr old, suggesting a difference in lichen successional dynamics between the ranges.     

Predicting growth of mat‐forming lichens on a landscape scale – comparing models with different complexities

During the 20th century, forestry practices has adversely affected lichen‐rich habitats. Mat‐forming lichens are important components of the vegetation of boreal and arctic ecosystems and are the main reindeer forage during the winter. To support the long‐term management of lichens in such habitats we developed models for predicting the growth of two common species. The lichens were transplanted across northern Scandinavia along a west‐east gradient varying in precipitation, temperature and irradiance. Growth was recorded seasonally over 16 months and ranged from ?4.8 to 34.6% and ?12.7 to 34.7% dry weight change for Cetraria stellaris and Cladina islandica, respectively. Growth was light limited below canopies with more than ca 60% cover and highest at the more humid sites when light levels were optimal. The models were based on various meteorological parameters, irradiance, physiological data and lichen hydration status; the latter was derived from a recently developed lichen hydration model. Our models' abilities to predict growth, both annually and seasonally (i.e. in summer), were evaluated in relation to their complexity and their potential usefulness from a management perspective. One parameter related to irradiance (the logarithm of site openness) was valuable in the prediction of annual growth for both species and could, in combination with precipitation, explain 52% of the variation in annual growth for C. stellaris and, in combination with total wet time and the irradiance received while wet, explain 66% of the variation in annual growth for C. islandica. The best simplified model explained 43% of the variation in annual growth for C. stellaris, using stem basal area and the annual normal temperature, and 24% for C. islandica using basal area alone. It is concluded that ensuring sufficient irradiance below the forest canopy is of crucial importance in the long‐term management of mat‐forming lichens and that simplified models can be used to identify appropriate habitats.     

HOW AGRICULTURE AFFECTS LICHEN VEGETATION IN CENTRAL SWITZERLAND

The results from several floristic and bioindication projects in Central Switzerland (1986–1998), focusing on the effects of agriculture, particularly from nitrogen and agrochemical emissions, are summarized. The abundance of nutrient-tolerant lichen species and the decreased occurrence of nitrophytic lichens are both correlated with agricultural land use and high atmospheric deposition. The impact could be demonstrated not only by the distribution patterns of nutrient-tolerant species, but also by the high N content of lichens such asPhyscia caesia. Positive growth response ofCetraria islandicato application of mineral nutrients and increased CO₂in open-top chambers at high altitudes was also observed. The methods and a survey of the results from studies conducted within the framework of the lichen research programme of the Natural History Museum of Lucernce are presented here. A discussion of the issues shows the need for further studies concenring the influence of introduced nutrients on the biodiverisyt of lichens.     

Screening of antioxidant potential of Arctic lichens

Antioxidants are compounds that scavenge the free radicals produced in living organisms. The antioxidant potential of eight Arctic lichen species was evaluated in vitro using free radical scavenging activity (FRS), inhibition of lipid peroxidation (ILP), and Trolox equivalent antioxidant capacity assay (TEAC). FRS activities of lichen species in various organic solvents such as methanol, ethanol, acetone, and dimethyl sulphoxide (DMSO) were in the range 9.6–51.77%, while ILP activities in these solvents ranged from 32.5 to 82.43%. Pseudophebe pubescens showed the highest ILP (82.43%) and FRS (51.77%) activities as compared to other lichen species and the standard antioxidants butylated hydroxyanisole (BHA) and butylated hydroxytoluene (BHT). The TEAC value was also found to be higher in all species compared to the standard water soluble vitamin E analog Trolox (3.9 mM). The order of antioxidative activities in lichen species was Pseudophebe pubescens > Cladonia amaurocraea > Cladonia mediterranea > Physcia caesia > Flavocetraria nivalis > Cetraria fastigata > Xanthoria elegans > Umbilicaria hyperborea. This is the first report of the measurement of antioxidant potential in Arctic lichens.     

Reindeer (<Emphasis Type="Italic">Rangifer tarandus</Emphasis>) avoidance of a highway as revealed by lichen measurements

Reindeer and caribou Rangifer tarandus are reported to avoid human infrastructure such as roads, high-voltage power lines, pipelines, and tourist resorts. Lichens are important forage for reindeer during winter, and their relatively slow growth rates make them vulnerable to overgrazing. Height and volume of lichens are often used as an indicator of grazing pressure by reindeer and, thus, as an indirect measure of Rangifer avoidance of human infrastructure. We sampled lichen height in Cetraria nivalis-dominated communities along 4 and 3 parallel transects located on two parallel mountain ridges in Hardangervidda, south central Norway. The lichen measurements were analyzed in relation to altitude and the distance from four tourist cabins in the area and a highway (Rv7) running perpendicular to the 7 transects. The mountain ridge with 4 transects is part of a much used migratory corridor for wild reindeer R. tarandus tarandus. Along the nonmigratory ridge, lichen height decreased 35% over an 8-km distance from Rv7 and a tourist cabin, indicating reindeer aversion toward Rv7 and/or a tourist cabin. No similar relationship was found for the migration ridge in relation to distance from Rv7 or the tourist cabins. Our results suggest that avoidance of human infrastructure by wild reindeer might be limited where reindeer use of winter pastures is influenced by herd traditions and/or motivation to follow established migration corridors. This has important implications for addressing the use of similar pasture measurements when testing for Rangifer aversion toward human disturbances.     

Extreme phenotypic variation in Cetraria aculeata (lichenized Ascomycota): adaptation or incidental modification?

Background and Aims

Phenotypic variability is a successful strategy in lichens for colonizing different habitats. Vagrancy has been reported as a specific adaptation for lichens living in steppe habitats around the world. Among the facultatively vagrant species, the cosmopolitan Cetraria aculeata apparently forms extremely modified vagrant thalli in steppe habitats of Central Spain. The aim of this study was to investigate whether these changes are phenotypic plasticity (a single genotype producing different phenotypes), by characterizing the anatomical and ultrastructural changes observed in vagrant morphs, and measuring differences in ecophysiological performance.

Methods

Specimens of vagrant and attached populations of C. aculeata were collected on the steppes of Central Spain. The fungal internal transcribed spacer (ITS), glyceraldehyde-3-phosphate dehydrogenase (GPD) and the large sub-unit of the mitochondrial ribosomal DNA (mtLSUm), and the algal ITS and actin were studied within a population genetics framework. Semi-thin and ultrathin sections were analysed by means of optical, scanning electron and transmission electron microscopy. Gas exchange and chlorophyll fluorescence were used to compare the physiological performance of both morphs.

Key Results and Conclusions

Vagrant and attached morphs share multilocus haplotypes which may indicate that they belong to the same species in spite of their completely different anatomy. However, differentiation tests suggested that vagrant specimens do not represent a random sub-set of the surrounding population. The morphological differences were related to anatomical and ultrastructural differences. Large intercalary growth rates of thalli after the loss of the basal–apical thallus polarity may be the cause of the increased growth shown by vagrant specimens. The anatomical and morphological changes lead to greater duration of ecophysiological activity in vagrant specimens. Although the anatomical and physiological changes could be chance effects, the genetic differentiation between vagrant and attached sub-populations and the higher biomass of the former show fitness effects and adaptation to dry environmental conditions in steppe habitats.     

Neuroprotective activity and cytotoxic potential of two Parmeliaceae lichens: Identification of active compounds

BackgroundLichens are symbiotic organisms capable of producing unique secondary metabolites, whose pharmacological activities are attracting much interest.PurposeThe present study aimed to investigate the in vitro neuroprotective effects and anticancer potential of methanol extracts of two Parmeliaceae lichens: Cetraria islandica and Vulpicida canadensis. The chemical composition of the two lichens was also determined.MethodsNeuroprotective activity was studied with respect to the antioxidant properties of the extracts; radical scavenging tests (ORAC and DPPH assays) were performed and oxidative stress markers (intracellular ROS production, caspase-3 activity, MDA and glutathione levels) were assessed in a hydrogen peroxide-induced oxidative stress model in astrocytes. Cytotoxic activity was tested against human HepG2 (hepatocellular carcinoma) and MCF-7 (breast adenocarcinoma) cell lines.ResultsCell viability studies identified a single concentration for each extract that was subsequently used to measure oxidative stress markers. Lichen extracts were able to reverse the oxidative damage caused by hydrogen peroxide, thus promoting astrocyte survival. Both lichen extracts also had anticancer activity in the cell lines, with IC₅₀ values of 19.51–181.05 µg/ml. The extracts had a high total phenolic content, and the main constituents identified by HPLC were fumarprotocetraric acid in Cetraria islandica, and usnic, pinastric and vulpinic acids in Vulpicida canadensis. The biological activities of the lichen extracts can be attributed to these secondary metabolites.ConclusionThe lichen species studied are promising sources of natural compounds with neuroprotective activity and cytotoxic potential, and warrant further research.     

Ecophysiology and genetic structure of polar versus temperate populations of the lichen Cetraria aculeata

We studied polar and temperate samples of the lichen Cetraria aculeata to investigate whether genetical differences between photobionts are correlated with physiological properties of the lichen holobiont. Net photosynthesis and dark respiration (DR) at different temperatures (from 0 to 30 °C) and photon flux densities (from 0 to 1,200 μmol m^?2 s^?1) were studied for four populations of Cetraria aculeata. Samples were collected from maritime Antarctica, Svalbard, Germany and Spain, representing different climatic situations. Sequencing of the photobiont showed that the investigated samples fall in the polar and temperate clade described in Fernández-Mendoza et al. (Mol Ecol 20:1208–1232, 2011). Lichens with photobionts from these clades differ in their temperature optimum for photosynthesis, maximal net photosynthesis, maximal DR and chlorophyll content. Maximal net photosynthesis was much lower in Antarctica and Svalbard than in Germany and Spain. The difference was smaller when rates were expressed by chlorophyll content. The same is true for the temperature optima of polar (11 °C) and temperate (15 and 17 °C) lichens. Our results indicate that lichen mycobionts may adapt or acclimate to local environmental conditions either by selecting algae from regional pools or by regulating algal cell numbers (chlorophyll content) within the thallus.     

Environmental factors and ground disturbance affecting the composition of species and functional traits of ground layer lichens on grey dunes and dune heaths of Estonia

A unique, species‐rich and endangered lichen biota can be found on European coastal and inland sand dunes. However, it is increasingly affected by natural succession as well as by anthropogenic disturbances. We studied lichen diversity on the grey dunes and dune heaths of coastal and inland regions of Estonia. A total of 28 study plots were investigated; in each 0.1 ha study plot general environmental variables and anthropogenic disturbances were described and all epigeic lichen species were identified. We found 66 lichenized fungus (lichen) species, including several rare and ten red‐listed lichens. Multivariate analysis (DCA, CCA) was performed to examine gradients in species composition and to relate variation in species data to environmental factors. In addition, we used redundancy analysis (RDA) to relate variation in species’ trait composition to environmental factors. Species composition on grey dunes differed significantly from that on dune heaths. The characteristic species for grey dunes are, besides several Cladonia species, foliose lichens, e.g. Hypogymnia physodes, Parmelia sulcata and Peltigera spp. Also species’ traits composition was different for either habitat, indicating that sorediate lichens, foliose lichens, lichens with cyanobacterium as the main photobiont, and sparsely branched Cladonia species dominate on grey dunes, while esorediate, green‐algal, crustose and richly branched fruticose lichens are common on dune heaths. Soil pH was the most essential environmental variable for determining both species composition and species’ traits composition. The composition of lichen species was also significantly influenced by forest closeness, soil Mg content and cover of bare sand; the effect of ground disturbances was low compared to the effect of these environmental factors. To protect and conserve the species‐rich lichen biota, it is necessary to protect the dune habitats from building activity, to avoid overtrampling in recreation areas and to regularly remove shrubs and trees.     

Disturbance‐induced changes in a high‐alpine cushionfield community,south‐central New Zealand

Long‐term effects of two nearby disturbances, mechanical blading 34 years ago and an approximately century‐old sheep graveyard, in high‐alpine cushionfield share certain vegetation features despite their contrasting soil nutrient status. Secondary succession following blading has continued beyond that recorded over the first 24 years with continued trends towards the adjacent undisturbed cushionfield: increased cover of several cushion species and the lichen Cetraria islandica ssp. antarctica. Several mid‐seral species remain prominent while bare soil and stone pavement continue to decrease. Additional shelter, facilitated by tall snow tussocks planted locally at near‐natural density on part of the bladed site, allowed establishment of some larger, thin‐leaved species. Soil factors remain little affected by the mechanical disturbance compared with the general enrichment persisting at the graveyard. Here three small indigenous grasses, which dominated 50 years earlier (Trisetum spicatum, Agrostis muelleriana, Poa colensoi), remain prominent among the relatively diverse community (50 species, including several calciphilic lichens on the degrading bones). By contrast, frequency of the cushionfield dominant, Dracophyllum muscoides and the lichen Alectoria nigricans, are negatively correlated with the presence of bones. The post‐disturbance temporal community changes are assessed in the context of the initial productivity of the habitat, competitive exclusion and the intermediate disturbance hypothesis.     

Tremella macrobasidiata and Tremella variae have abundant and widespread yeast stages in Lecanora lichens

Dimorphism is a widespread feature of tremellalean fungi in general, but a little-studied aspect of the biology of lichen-associated Tremella. We show that Tremella macrobasidiata and Tremella variae have an abundant and widespread yeast stage in their life cycles that occurs in Lecanora lichens. Their sexual filamentous stage is restricted to a specific lichen: T. macrobasidiata only forms basidiomata on Lecanora chlarotera hymenia and T. variae only on Lecanora varia thalli. However, the yeast stage of T. macrobasidiata is less specific and can occur in L. varia lichens, whilst all life stages of T. variae may be specific to L. varia. Contrary to the hyphal stages, the yeasts are distributed across the thalli and hymenia of Lecanora lichens, and not limited to specimens with basidiomata. Tremella macrobasidiata was present in all studied L. chlarotera, and in 59% of L. varia specimens. Only in 8% of the L. varia thalli could none of the two Tremella species be detected. Our results indicate that lichen-associated Tremella may be much more abundant and widespread than previously assumed leading to skewed estimations about their distribution ranges and lichen specificity, and raise new questions about their biology, ecology and function in the symbiosis.     

Epiphytic lichen synusiae and functional trait groups in boreo‐nemoral deciduous forests are influenced by host tree and environmental factors

Deciduous forests with temperate broad‐leaved tree species are particularily important in terms of biodiversity and its protection, but are threatened habitats in northern Europe. Using multivariate analyses we studied the effect of forest site type, environmental variables and host tree properties on epiphytic lichen synusiae as well as on the composition of species‐specific functional traits. Epiphytic lichens were examined on Acer platanoides, Fraxinus excelsior, Quercus robur, Tilia cordata, Ulmus glabra and U. laevis in two types of forests: Humulus‐type floodplain forests and Lunaria‐type boreo‐nemoral forests on the talus slopes of limestone escarpment (klint forests). Klint forests located near the seashore were under greater maritime influence compared to floodplain forests located in inland Estonia which experience stronger air temperature contrasts. In addition to stand level and climatic variables, tree level factors (bark pH, trunk circumference and cover of bryophytes) considerably affected the species composition of the lichen synusiae. Overall, 137 lichen species were recorded, including 14 red‐listed species characteristic of deciduous trees. We defined 13 lichen societies and showed their preference to forests of a specific site type and/or host tree properties. In forests of both types, most of the epiphytic lichens were crustose, and had apothecia as the fruit bodies and chlorococcoid algae as the photobiont. However, the proportion of lichens with a foliose or fruticose growth form, as well as the proportion of lichens with vegatative diaspores, were higher in floodplain forests. In klint forests with a stronger influence from the wind, crustose species completely dominated, while species with vegetative diaspores were rare and most species dispersed sexually. Lichens with Trentepohlia as the photobiont were characteristic of these forests, and lichens with lirellate ascomata were prevailing, indicating the great uniqueness of the kint forests for epiphytic lichens in the boreo‐nemoral region.     

Combined observational and experimental data provide limited support for facilitation in lichens

It is increasingly recognized that facilitative interactions can shape communities. One of the mechanisms through which facilitation may operate is when one species facilitates the colonization of another through the exchange of shared symbionts. Lichens are symbiotic associations composed of a mycobiont (lichenised‐fungus) and one or two photobionts (algae or cyanobacteria). Different lichen species may have overlapping specificity for photobionts, creating the possibility that facilitation drives lichen community assembly. To investigate whether facilitation occurs in lichens, we combined an observational study (a) with a manipulative field experiment (b). For (a), we quantified the effect of local patch conditions, facilitation and the size of the surrounding metapopulation on colonizations of an epixylic lichen species (Cladonia botrytes) in an area of managed boreal forest. This was done by twice surveying lichens on 293 stumps, located in stands of three age classes. For (b), we treated unoccupied surfaces of 56 cut stumps with algal mixtures of an Asterochloris photobiont and recorded C. botrytes colonizations over three years. In (a), colonization rates of C. botrytes increased with increasing abundance of other lichen species with specificity for Asterochloris photobionts, consistent with an effect of facilitation. However, in the field experiment (b), colonizations of the focal species did not provide support for facilitation. We conclude that our study provides limited support for facilitation in green‐algal lichens, underscoring the importance of combining observational studies with experiments when studying species interactions.