Can field botany be effectively taught as a distance course? Experiences and reflections from the COVID-19 pandemic.

The COVID-19 pandemic that started in 2020 forced a rapid change in university teaching, with large numbers of courses switching to distance learning with very little time for preparation. Courses involving many practical elements and field excursions required particular care if students were to fulfil planned learning outcomes. Here, we present our experiences in teaching field botany in 2020 and 2021. Using a range of methods and tools to introduce students to the subject, promote self-learning and reflection and give rapid and regular feedback, we were able to produce a course that allowed students to achieve the intended learning outcomes and that obtained similarly positive student evaluations to previous years. The course and its outcomes were further improved in 2021. We describe how we structured field botany as a distance course in order that we could give the best possible learning experience for the students. Finally, we reflect on how digital tools can aid teaching such subjects in the future, in a world where public knowledge of natural history is declining.

Tree species identity and composition shape the epiphytic lichen community of structurally simple boreal forests over vast areas.

Greatly simplified ecosystems are often neglected for biodiversity studies. However, these simplified systems dominate in many regions of the world, and a lack of understanding of what shapes species occurrence in these systems can have consequences for biodiversity and ecosystem services at a massive scale. In Fennoscandia, ~90% of the boreal forest (~21Mha) is structurally simplified with little knowledge of how forest structural elements shape the occurrence and diversity of for example epiphytic lichens in these managed forests. One form of structural simplification is the reduction of the number and frequency of different tree species. As many lichen species have host tree preferences, it is particularly likely that this simplification has a huge effect on the lichen community in managed forests. In a 40-70 years old boreal forest in Sweden, we therefore related the occurrence and richness of all observed epiphytic lichens to the host tree species and beta and gamma lichen diversity at the forest stand level to the stand's tree species composition and stem diameter. Picea abies hosted the highest lichen richness followed by Pinus sylvestris, Quercus robur, Alnus glutinosa, Betula spp., and Populus tremula. However, P. tremula hosted twice as many uncommon species as any of the other tree species. Stand level beta and gamma diversity was twice as high on stands with four compared to one tree species, and was highest when either coniferous or deciduous trees made up 40-50% of the trees. The stem diameter was positively related to lichen richness at the tree and stand level, but negatively to beta diversity. For biodiversity, these findings imply that leaving a few trees of a different species during forest thinning is unlikely as effective as combining life-boat trees for endangered species with an even tree species mixture.

Refining the picture: new records to the lichen biota of Italy.

Based on the analysis of both historical and recent collections, this paper reports an annotated list of taxa which are new to the lichen biota of Italy or of its administrative regions. Specimens were identified using a dissecting and a compound microscope; routine chemical spot tests and standardized thin-layer chromatography (TLC or HPTLC). The list includes 225 records of 153 taxa. Twenty taxa are new to Italy, the others are new to one or more administrative regions, with 15 second records and 5 third records for Italy. Some of the species belong to recently-described taxa, others are poorly known, sterile or ephemeral lichens which were largely overlooked in Italy. Several species are actually rare, either because of the rarity of their habitats (e.g. old-growth forests), or because in Italy they are at the margins of their bioclimatic distribution. The picture of the lichen biota of Italy has now new pixels, but its grain is still coarse. Further analysis of historical collections, increased efforts in the exploration of some areas, and the taxonomic revision of critical groups are still necessary to provide more complete distributional data for new biogeographic hypotheses, taxonomic and ecological research, and biodiversity conservation.

The Plot Thickens: Haploid and Triploid-Like Thalli, Hybridization, and Biased Mating Type Ratios in Letharia.

The study of the reproductive biology of lichen fungal symbionts has been traditionally challenging due to their complex lifestyles. Against the common belief of haploidy, a recent genomic study found a triploid-like signal in Letharia. Here, we infer the genome organization and reproduction in Letharia by analyzing genomic data from a pure culture and from thalli, and performing a PCR survey of the MAT locus in natural populations. We found that the read count variation in the four Letharia specimens, including the pure culture derived from a single sexual spore of L. lupina, is consistent with haploidy. By contrast, the L. lupina read counts from a thallus' metagenome are triploid-like. Characterization of the mating-type locus revealed a conserved heterothallic configuration across the genus, along with auxiliary genes that we identified. We found that the mating-type distributions are balanced in North America for L. vulpina and L. lupina, suggesting widespread sexual reproduction, but highly skewed in Europe for L. vulpina, consistent with predominant asexuality. Taken together, we propose that Letharia fungi are heterothallic and typically haploid, and provide evidence that triploid-like individuals are hybrids between L. lupina and an unknown Letharia lineage, reconciling classic systematic and genetic studies with recent genomic observations.

The evolutionary species pool concept does not explain occurrence patterns of dead-wood-dependent organisms: implications for logging residue extraction.

Emulation of natural disturbances is often regarded as a key measure to make forestry biodiversity-oriented. Consequently, extraction of logging residues is assumed to have little negative effect in comparison to extraction of dead wood mainly formed at natural disturbances. This is consistent with the evolutionary species pool hypothesis, which suggests that most species are evolutionary adapted to the naturally most abundant habitats. We tested this hypothesis for dead-wood-dependent macrofungi, lichens, and beetles in a boreal forest landscape in central Sweden, assuming that species are adapted to conditions similar to today's unmanaged forest. No occurrence patterns, for the species groups which we investigated, were consistent with the hypothesis. Overall, stumps and snags had the highest habitat quality (measured as average population density with equal weight given to each species) and fine woody debris the lowest, which was unexpected, since stumps were the rarest dead-wood type in unmanaged forest. We conclude that the evolutionary species pool concept did not explain patterns of species' occurrences, and for two reasons, the concept is not reliable as a general rule of thumb: (1) what constitute habitats harbouring different species communities can only be understood from habitat-specific studies and (2) the suitability of habitats is affected by their biophysical characteristics. Thus, emulation of natural disturbances may promote biodiversity, but empirical studies are needed for each habitat to understand how natural disturbances should be emulated. We also conclude that stump extraction for bioenergy is associated with larger risks for biodiversity than fine woody debris extraction.

Lichen chemistry is concordant with multilocus gene genealogy in the genus Cetrelia (Parmeliaceae, Ascomycota).

The lichen genus Cetrelia represents a taxonomically interesting case where morphologically almost uniform populations differ considerably from each other chemically. Similar variation is not uncommon among lichenized fungi, but it is disputable whether such populations should be considered entities at the species level. Species boundaries in Cetrelia are traditionally delimited either as solely based on morphology or as combinations of morpho- and chemotypes. A dataset of four nuclear markers (ITS, IGS, Mcm7, RPB1) from 62 specimens, representing ten Cetrelia species, was analysed within Bayesian and maximum likelihood frameworks. Analyses recovered a well-resolved phylogeny where the traditional species generally were monophyletic, with the exception of Cetrelia chicitae and Cetrelia pseudolivetorum. Species delimitation analyses supported the distinction of 15 groups within the studied Cetrelia taxa, dividing three traditionally identified species into some species candidates. Chemotypes, distinguished according to the major medullary substance, clearly correlated with clades recovered within Cetrelia, while samples with the same reproductive mode were dispersed throughout the phylogenetic tree. Consequently, delimiting Cetrelia species based only on reproductive morphology is not supported phylogenetically. Character analyses suggest that chemical characters have been more consistent compared to reproductive mode and indicate that metabolite evolution in Cetrelia towards more complex substances is probable.

Two Basidiomycete Fungi in the Cortex of Wolf Lichens.

Since the late 1800s, mycologists have been detecting fungi above and beyond the assumed single fungus in lichen thalli [1-6]. Over the last century, these fungi have been accorded roles ranging from commensalists to pathogens. Recently, Cyphobasidiales yeasts were shown to be ubiquitous in the cortex layer of many macrolichens [7], but for most species, little is known of their cellular distribution and constancy beyond visible fruiting structures. Here, we demonstrate the occurrence of an additional and distantly related basidiomycete, Tremella, in 95% of studied thalli in a global sample of one of the most intensively studied groups of lichens, the wolf lichens (genus Letharia). Tremella species are reported from a wide range of lichen genera [8], but until now, their biology was deduced from fruiting bodies (basidiomata) formed on lichen thalli. Based on this, they have been thought to be uncommon to rare, to occur exclusively in a hyphal form, and to be parasitic on the dominant fungal partner [9, 10]. We show that, in wolf lichens, Tremella occurs as yeast cells also in thalli that lack basidiomata and infer that this is its dominant stage in nature. We further show that the hyphal stage, when present in Letharia, is in close contact with algal cells, challenging the assumption that lichen-associated Tremella species are uniformly mycoparasites. Our results suggest that extent of occurrence and cellular interactions of known fungi within lichens have historically been underestimated and raise new questions about their function in specific lichen symbioses.

Photobiont switching causes changes in the reproduction strategy and phenotypic dimorphism in the Arthoniomycetes.

Phylogenetic analyses using mtSSU and nuITS sequences of Buellia violaceofusca (previously placed in Lecanoromycetes), a sterile, sorediate lichen having a trebouxioid photobiont, surprisingly prove that the species is conspecific with Lecanographa amylacea (Arthoniomycetes), a fertile, esorediate species with a trentepohlioid photobiont. These results suggest that L. amylacea and B. violaceofusca are photomorphs of the same mycobiont species, which, depending on the photobiont type, changes the morphology and the reproduction strategy. This is the first example of a lichenized fungus that can select between Trebouxia (Trebouxiophyceae) and trentepohlioid (Ulvophyceae) photobionts. Trebouxia photobionts from the sorediate morphotype belong to at least three different phylogenetic clades, and the results suggest that Lecanographa amylacea can capture the photobiont of other lichens such as Chrysothrix candelaris to form the sorediate morphotype. Phylogenetic analyses based on rbcL DNA data suggest that the trentepohlioid photobiont of L. amylacea is closely related to Trentepohlia isolated from fruticose lichens. The flexibility in the photobiont choice enables L. amylacea to use a larger range of tree hosts. This strategy helps the lichen to withstand changes of environmental conditions, to widen its distribution range and to increase its population size, which is particularly important for the survival of this rare species.

Consequences of bioenergy wood extraction for landscape-level availability of habitat for dead wood-dependent organisms.

Stumps and slash resulting from forest clearcutting is used as a source of low-net-carbon energy, but there are concerns about the consequences of biofuel extraction on biodiversity. Logging residues constitute potentially important habitats, since a large part of forest biodiversity is dependent on dead wood. Here we used snapshot field data from a managed forest landscape (25 000 ha) to predict landscape scale population changes of dead wood dependent organisms after extraction of stumps and slash after clearcutting. We did this by estimating habitat availability for all observed dead wood-dependent beetles, macrofungi, and lichens (380 species) in the whole landscape. We found that 53% of species occurred in slash or stumps. For most species, population declines after moderate extraction (≤30%) were small (<10% decline) because they mainly occur on other dead wood types. However, some species were only recorded in slash and stumps. Red listed species were affected by slash and stump extraction (12 species), but less often than other species. Beetles and fungi were more affected by stump extraction, while lichens were more affected by slash extraction. For beetles and lichens, extraction of a combination of spruce, pine and birch resulted in larger negative effects than if only extracting spruce, while for fungi tree species had little effect. We conclude that extensive extraction decreases the amount of habitat to such extent that it may have negative consequences on species persistence at the landscape level. The negative consequences can be limited by extracting only slash, or only logging residues from spruce stands.

Basidiomycete yeasts in the cortex of ascomycete macrolichens.

For over 140 years, lichens have been regarded as a symbiosis between a single fungus, usually an ascomycete, and a photosynthesizing partner. Other fungi have long been known to occur as occasional parasites or endophytes, but the one lichen-one fungus paradigm has seldom been questioned. Here we show that many common lichens are composed of the known ascomycete, the photosynthesizing partner, and, unexpectedly, specific basidiomycete yeasts. These yeasts are embedded in the cortex, and their abundance correlates with previously unexplained variations in phenotype. Basidiomycete lineages maintain close associations with specific lichen species over large geographical distances and have been found on six continents. The structurally important lichen cortex, long treated as a zone of differentiated ascomycete cells, appears to consistently contain two unrelated fungi.

A Pine Is a Pine and a Spruce Is a Spruce--The Effect of Tree Species and Stand Age on Epiphytic Lichen Communities.

With an increasing demand for forest-based products, there is a growing interest in introducing fast-growing non-native tree species in forest management. Such introductions often have unknown consequences for native forest biodiversity. In this study, we examine epiphytic lichen species richness and species composition on the trunks of non-native Pinus contorta and compare these to the native Pinus sylvestris and Picea abies in managed boreal forests in northern Sweden across a chronosequence of age classes. Overall, we recorded a total of 66,209 lichen occurrences belonging to 57 species in the 96 studied forest stands. We found no difference in species richness of lichens between stands of P. contorta and P. sylvestris, but stands of P. abies had higher total species richness. However, species richness of lichens in stands of P. abies decreased with increasing stand age, while no such age effect was detected for P. contorta and P. sylvestris. Lichen species composition progressively diverged with increasing stand age, and in 30-year-old stands all three tree species showed species-specific assemblages. Epiphytic lichen assemblages in stands of 30-year-old P. contorta were influenced by greater basal area, canopy closure, and average diameter at breast height, P. abies stands by higher branch density and canopy closure, and stands of P. sylvestris by greater bark crevice depth. Differences in lichen species richness and composition were mainly explained by canopy closure and habitat availability, and the greater canopy closure in mature P. abies stands promoted the colonization and growth of calicioid lichen species. Our results indicate that the non-native P. contorta have similar species richness as the native P. sylvestris. The main difference in lichen species richness and composition is between P. abies and Pinus spp. in managed forests of boreal Sweden.

Morphological, chemical and species delimitation analyses provide new taxonomic insights into two groups of Rinodina.

The genus Rinodina (Physciaceae), with approximately 300 species, has been subject to few phylogenetic studies. Consequently taxonomic hypotheses in Rinodina are largely reliant on phenotypic data, while hypotheses incorporating DNA dependent methods remain to be tested. Here we investigate Rinodina degeliana/R. subparieta and the Rinodina mniaraea group, which previously have not been subjected to comprehensive molecular and phenotypic studies. We conducted detailed morphological, anatomical, chemical, molecular phylogenetic and species delimitation studies including 24 newly sequenced specimens. We propose that Rinodina degeliana and R. subparieta are conspecific and that chemical morphs within the R. mniaraea group should be recognized as distinct species. We also propose the placement of the recently described genus Oxnerella in Physciaceae.

Diagnostics for a troubled backbone: testing topological hypotheses of trapelioid lichenized fungi in a large-scale phylogeny of Ostropomycetidae (Lecanoromycetes).

Trapelioid fungi constitute a widespread group of mostly crust-forming lichen mycobionts that are key to understanding the early evolutionary splits in the Ostropomycetidae, the second-most species-rich subclass of lichenized Ascomycota. The uncertain phylogenetic resolution of the approximately 170 species referred to this group contributes to a poorly resolved backbone for the entire subclass. Based on a data set including 657 newly generated sequences from four ribosomal and four protein-coding gene loci, we tested a series of a priori and new evolutionary hypotheses regarding the relationships of trapelioid clades within Ostropomycetidae. We found strong support for a monophyletic group of nine core trapelioid genera but no statistical support to reject the long-standing hypothesis that trapelioid genera are sister to Baeomycetaceae or Hymeneliaceae. However, we can reject a sister group relationship to Ostropales with high confidence. Our data also shed light on several long-standing questions, recovering Anamylopsoraceae nested within Baeomycetaceae, elucidating two major monophyletic groups within trapelioids (recognized here as Trapeliaceae and Xylographaceae), and rejecting the monophyly of the genus Rimularia. We transfer eleven species of the latter genus to Lambiella and describe the genus Parainoa to accommodate a previously misunderstood species of Trapeliopsis. Past phylogenetic studies in Ostropomycetidae have invoked "divergence order" for drawing taxonomic conclusions on higher level taxa. Our data show that if backbone support is lacking, contrasting solutions may be recovered with different or added data. We accordingly urge caution in concluding evolutionary relationships from unresolved phylogenies.

Occurrence patterns of lichens on stumps in young managed forests.

The increasing demand for forest-derived bio-fuel may decrease the amount of dead wood and hence also the amount of available substrate for saproxylic ( = dead-wood dependent) organisms. Cut stumps constitute a large portion of dead wood in managed boreal forests. The lichen flora of such stumps has received little interest. Therefore, we investigated which lichens that occur on stumps in young (4-19 years), managed forests and analyzed how species richness and occurrence of individual species were related to stump and stand characteristics. We performed lichen inventories of 576 Norway spruce stumps in 48 forest stands in two study areas in Central Sweden, recording in total 77 lichen species. Of these, 14 were obligately lignicolous, while the remaining were generalists that also grow on bark, soil or rocks. We tested the effect of characteristics reflecting successional stage, microclimate, substrate patch size, and the species pool in the surrounding area on (1) total lichen species richness, (2) species richness of obligately lignicolous lichens and (3) the occurrence of four obligately lignicolous lichen species. The most important variables were stump age, with more species on old stumps, and study area, with similar total species richness but differences in occupancy for individual species. Responses for total lichen species richness and species richness of obligately lignicolous lichens were overall similar, indicating similar ecological requirements of these two groups. Our results indicate that species richness measurements serve as poor proxies for the responses of individual, obligately lignicolous lichen species.

Estimating coextinction risks from epidemic tree death: affiliate lichen communities among diseased host tree populations of Fraxinus excelsior.

At least 10% of the world's tree species are threatened with extinction and pathogens are increasingly implicated in tree threats. Coextinction and threats to affiliates as a consequence of the loss or decline of their host trees is a poorly understood phenomenon. Ash dieback is an emerging infectious disease causing severe dieback of common ash Fraxinus excelsior throughout Europe. We utilized available empirical data on affiliate epiphytic lichen diversity (174 species and 17,800 observations) among 20 ash dieback infected host tree populations of F. excelsior on the island Gotland in the Baltic Sea, Sweden. From this, we used structured scenario projections scaled with empirical data of ash dieback disease to generate probabilistic models for estimating local and regional lichen coextinction risks. Average coextinction probabilities (A) were 0.38 (95% CI ± 0.09) for lichens occurring on F. excelsior and 0.14 (95% CI ± 0.03) when considering lichen persistence on all tree species. A was strongly linked to local disease incidence levels and generally increasing with lichen host specificity to F. excelsior and decreasing population size. Coextinctions reduced affiliate community viability, with significant local reductions in species richness and shifts in lichen species composition. Affiliates were projected to become locally extirpated before their hosts, illuminating the need to also consider host tree declines. Traditionally managed open wooded meadows had the highest incidence of ash dieback disease and significantly higher proportions of affiliate species projected to go extinct, compared with unmanaged closed forests and semi-open grazed sites. Most cothreatened species were not previously red-listed, which suggest that tree epidemics cause many unforeseen threats to species. Our analysis shows that epidemic tree deaths represent an insidious, mostly overlooked, threat to sessile affiliate communities in forested environments. Current conservation and management strategies must account for secondary extinctions associated with epidemic tree death.

Phylogenetic analysis of multiple loci reveal the population structure within Letharia in the Caucasus and Morocco.

The sequence variation within the genus Letharia in the Caucasus and Morocco was investigated. Twelve thalli from each area were sequenced at eight different loci. Phylogenetic analysis of the multiple loci data revealed the cryptic species Letharia 'lupina' in Morocco, previously known only from western North America. The two cryptic species L. vulpina and L. 'lupina' locally share the same ecology but are genetically isolated from each other. In the Caucasus, five different haplotypes of L. vulpina were detected, and in Morocco four L. vulpina haplotypes and six L. 'lupina' haplotypes were found. For L. vulpina, allelic differences were detected at five of the eight loci in the Caucasus and Morocco. The populations of L. vulpina in both the Caucasus and Morocco contain more genetic variation than those previously investigated in Europe, which indicates that the Caucasus and Morocco acted as refugia during quaternary glaciations, and that central and northern Europe may have been recolonised from one or both of these areas.

Reproductive mode and genetic variation suggest a North American origin of European Letharia vulpina.

Our data on the intercontinental population biology of Letharia vulpina show an unexpected shift from a recombining North American population with unique haplotypes to genetically depauperate Swedish and Italian populations, each with many representatives of a single repeated haplotype. Analysis of eight loci in 47 individuals supported recombination in North American populations and showed almost no variation among European populations. We infer that a genetic bottleneck caused by limited long-distance dispersal accounts for the lack of genetic variation found in marginal populations. This lack of variation in the European populations makes it impossible to use population genetic means to distinguish clonal reproduction from self-fertilization or even outcrossing, but phenotype indicates that reproduction in the marginal populations is by clonal spread, via soredia and isidioid soredia.