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A number of lichen-forming fungal species are widely distributed. Here, we investigate biogeographic patterns in a widely distributed isidiate taxon -Parmelinella wallichiana - using molecular sequence data. Our results revealed that Parmelinella wallichina, as currently circumscribed, is not monophyletic but falls into four clades, two of them represented by a sample only. A third clade, occurring in Africa and southern India is described as a new species, Parmelinella schimperiana Kirika & Divakar, sp. nov. Our study adds a further example of previously overlooked, geographically distinct, lineages that were discovered using molecular data. [ABSTRACT FROM AUTHOR]

Accurate specimen identification is challenging in groups with subtle or scarce taxonomically diagnostic characters, and the use of DNA barcodes can provide an effective means for consistent identification. Here, we investigate the utility of DNA barcode identification of species in a cosmopolitan genus of lichen-forming fungi, Parmelia (Parmeliaceae). Two hundred and two internal transcribed spacer (ITS) sequences generated from specimens collected from all continents, including Antarctica, were analysed, and DNA barcodes of 14 species of Parmelia s.s. are reported. Almost all species show a barcode gap. Overall, intraspecific divergence values were lower than the threshold previously established for Parmeliaceae. However, the mean and range were elevated by deep barcode divergences in three species, indicating the likely occurrence of overlooked specieslevel lineages. Here, we provide a DNA barcode reference library with well-identified specimens sampled worldwide and sequences from most of the type material to enable easy and fast accurate sample identification and to assist in uncovering overlooked species in Parmelia s.s. Further, our results confirm the efficiency of the ITS region in the identification of species of Parmelia s.s. [ABSTRACT FROM AUTHOR]

Two new species in the genus Trapelia, namely, T. calvariana Kantvilas & Lumbsch from the Tasmanian highlands and T. thieleana Kantvilas & Lumbsch from south-western Western Australia, are described. Examination of ascus types and analyses of DNA-sequence data confirmed the inclusion of these taxa in Trapelia sensu lato, but raised questions concerning the homogeneity of the genus as currently circumscribed and, in particular, the generic classification of T. lilacea Kantvilas & Elix. [ABSTRACT FROM AUTHOR]

Context: The emergence of antibiotic resistant pathogens is a serious global health threat. Hence, the search for new antibiotic drugs from various natural sources should be given high priority. Lichens produce a variety of low molecular weight metabolic compounds and many cultures have utilized these compounds in traditional medicine for centuries. Objective: Report the antibiotic properties of extracts from 34 North American lichens screened against four pathogenic bacteria. Materials and methods: The micro-well dilution method was used to determine the minimum inhibitory concentration (MIC) of acetone and methanol extracts of 34 lichen species against four bacterial strains. Major chemical compounds in each species were identified using thin layer chromatography (TLC). Results: Most of the lichen extracts demonstrated inhibitory effects against Staphylococcus aureus, Pseudomonas aeruginosa, and methicillin-resistant S. aureus (MRSA) with MIC values ranging from 3.9 to 500 µg/ml. In addition, extracts from three species, Letharia columbiana (Nutt.) J. W. Thomson (Parmeliaceae), Letharia vulpina (L.) Hue (Parmeliaceae), and Vulpicida canadensis (Räsänen) J.-E. Mattsson & M. J. Lai (Parmeliaceae) (MIC = 125-500 µg/ml) were also effective against Escherichia coli. Generally, acetone extractions were found to be more effective than methanol extractions. Discussion and conclusion: Results of this study show that lichen extracts provide significant antimicrobial activity against both Gram-positive and Gram-negative bacteria. These results suggest that lichens may be an important potential source of antibacterial drugs. [ABSTRACT FROM AUTHOR]

The inclusion of molecular data is increasingly an integral part of studies assessing species boundaries. Analyses based on predefined groups may obscure patterns of differentiation, and population assignment tests provide an alternative for identifying population structure and barriers to gene flow. In this study, we apply population assignment tests implemented in the programs STRUCTURE and BAPS to single nucleotide polymorphisms from DNA sequence data generated for three previous studies of the lichenized fungal genus Letharia. Previous molecular work employing a gene genealogical approach circumscribed six species-level lineages within the genus, four putative lineages within the nominal taxon L. columbiana (Nutt.) J.W. Thomson and two sorediate lineages. We show that Bayesian clustering implemented in the program STRUCTURE was generally able to recover the same six putative Letharia lineages. Population assignments were largely consistent across a range of scenarios, including: extensive amounts of missing data, the exclusion of SNPs from variable markers, and inferences based on SNPs from as few as three gene regions. While our study provided additional evidence corroborating the six candidate Letharia species, the equivalence of these genetic clusters with species-level lineages is uncertain due, in part, to limited phylogenetic signal. Furthermore, both the BAPS analysis and the ad hoc ΔK statistic from results of the STRUCTURE analysis suggest that population structure can possibly be captured with fewer genetic groups. Our findings also suggest that uneven sampling across taxa may be responsible for the contrasting inferences of population substructure. Our results consistently supported two distinct sorediate groups, ‘L. lupina’ and L. vulpina, and subtle morphological differences support this distinction. Similarly, the putative apotheciate species ‘L. lucida’ was also consistently supported as a distinct genetic cluster. However, additional studies will be required to elucidate the relationships of other L. columbiana s.l. populations with the two sorediate genetic clusters. [ABSTRACT FROM AUTHOR]

Fungal selectivity (range of potential partners) for algal symbionts has been found to vary in lichen associations. Although a large number of studies have focused on the taxonomy and evolution of fungi in the speciose parmelioid clade (Parmeliaceae, Ascomycota), fungal-algal interactions in this ecologically and evolutionarily diverse group remain largely unexplored. In this study we investigated the algal symbionts of Xanthoparmelia fungi in western North America. We generated sequence data from a total of 58 rock shield lichens (Xanthoparmelia fungi + Trebouxia algae) collected across three sites in a subalpine community in southern Utah, USA. We explored the role of substrates, secondary metabolite variation, phenotype-based Xanthoparmelia species, mycobiont genetic population clusters, and sitespecific differences in structuring assemblages of algae. Our results indicate that Xanthoparmelia fungi associate with a broad range of Trebouxia lineages in a local, subalpine habitat. Most algae sampled form part of the diverse Trebouxia 'arboricola/gigantea' clade, while a small number of algae from outside of this clade were also found to associate with the Xanthoparmelia species investigated here. Our results also revealed multiple Trebouxia lineages within the T. 'arboricola/gigantea' clade that have not been previously recognized. Furthermore, accumulation curves suggested that additional algal diversity in rock shield lichen communities in the sampled subalpine habitat may be recovered with increased sampling. Overall, we found no clear association of algal clades with traditional phenotype-based Xanthoparmelia species, mycobiont genetic population clusters, distinct extrolites, or substrates. However, our data revealed significant genetic structuring of Trebouxia communities in separate subalpine meadow areas in a relatively homogeneous subalpine community. Significant site-specific differences in Trebouxia diversity in rock shield lichen communities and low selectivity suggest that photobiont flexibility may play an important role in overall successful colonization of rock shield lichens across a wide array of habitats in western North America. [ABSTRACT FROM AUTHOR]

The relationship of Aspicilia uxoris within Megasporaceae is assessed within a phylogenetic context. ‘Aspicilia’ uxoris and other related species are recovered as sister to the genus Lobothallia s. str. and described here as a new genus. Teuvoa (Ascomycota, Megasporaceae) is erected based on nuclear ITS and LSU sequence data and morphological characters. In addition to Teuvoa uxoris, a second species, T. junipericola, is added to the new genus based on material collected from North America. Teuvoa junipericola, T. uxoris and T. tibetica form a group with 8-spored asci, absence of extrolites, rather short-sized conidia and ascospores, lack of a subhypothecial algal layer, and different substratum preferences (on organic substratum) with a sister relationship to genus Lobothallia s. lat. (Aspicilia subgenus Pachyothallia Clauzade & C. Roux). Based on spore measurements of the holotypes, Lecanora ferganensis Tomin from central Asia (Kyrgyzstan, Tajikistan and Uzbekistan), Lecanora atrodiscata Gintovt, from Tajikistan and Lecanora takyroides Dzhur. from Turkmenistan are new synonyms to T. uxoris. A lectotype for Lecanora ferganensis is designated, expanding the known distribution of T. uxoris from Algeria, Morocco and Spain, into Central Asia. [ABSTRACT FROM AUTHOR]

Recent studies using sequence data from multiple loci and coalescent-based species delimitation have revealed several species-level lineages within the phenotypically circumscribed taxon Rhizoplaca melanophthalma sensu lato. Here, we formally describe five new species within this group, R. occulta, R. parilis, R. polymorpha, R. porterii, and R. shushanii, using support from the coalescent-based species delimitation method implemented in the program Bayesian Phylogenetics and Phylogeography (BPP) as the diagnostic feature distinguishing new species. We provide a reference DNA sequence database using the ITS marker as a DNA barcode for identifying species within this complex. We also assessed intraspecific genetic distances within the six R. melanophthalma sensu lato species. While intraspecific genetic distances within the phenotypically cryptic lineages within this broadly distributed lineage. Overall, our results demonstrate the potential for accurately identifying species within the R. melanophthalma group by using molecularbased identification methods.five new species were less than or equal to the lowest interspecific pairwise comparison values, an overlap in genetic distances within the R. melanophthalma sensu stricto clade suggests the potential for additional phenotypically cryptic lineages within this broadly distributed lineage. Overall, our results demonstrate the potential for accurately identifying species within the R. melanophthalma group by using molecular-based identiication methods. [ABSTRACT FROM AUTHOR]

* Premise of the study: In spite of the recent advances in generic and species circumscriptions and in recognizing species diversity in lichen-forming fungi, the timing of speciation and the factors that promote diversification in lichens remain largely unexplored. We used brown parmelioids as a model to assess the timing of divergence and explore the impact of geological and climatic events on lineage divergence and diversification in lichenized fungi. Additionally, to clarify the phylogenetic position of the species currently placed in Melanelia disjuncta group, we evaluated the taxonomic status and phylogenetic relationships within Parmeliaceae. * Methods: Phylogenetic relationships and divergence time estimates were inferred from a four-loci data set. Alternative hypotheses were tested using Shimodaira-Hasegawa and expected likelihood weights tests. * Key results: The M. disjuncta group forms a strongly supported, monophyletic lineage independent from Melanelia s.s. The M. disjuncta clade arose ca. 23.1 million years ago (Ma). Our results suggest that most of the lineages within the clade diversified during the Miocene (17.6 to 11.2 Ma). The split of other brown parmelioids, such as Emodomelanelia-Melanelixia occurred ca. 41.70 Ma, and the radiation of Melanelixia began during the Eocene-Oligocene transition (ca. 33.75 Ma). * Conclusions: Montanelia is described here as a new genus to accommodate species of the Melaneliadisjuncta group. Further, the study indicates that the current species delimitation within the newly described genus requires revision. We provide evidence of lineage divergence of Montanelia at the Oligocene-Miocene boundary. Our results indicate that the diversification during Miocene would have happened during major mountain uplifts. [ABSTRACT FROM AUTHOR]

Premise of the study: Diversification in neotropical regions has been attributed to both Tertiary geological events and Pleistocene climatic fl uctuations. However, the timing and processes driving speciation in these regions remain unexplored in many important groups. Here, we address the timing of diversification in the neotropical lichenized fungal genus Oropogon (Ascomycota) and assess traditional species boundaries. Methods: We analyzed sequence data from three loci to assess phenotypically circumscribed Oropogon species from the Oaxacan Highlands, Mexico. We provide a comparison of dated divergence estimates between concatenated gene trees and a calibrated multilocus species-tree using substitution rates for two DNA regions. We also compare estimates from a data set excluding ambiguously aligned regions and a data set including the hyper-variable regions in two ribosomal markers. Key results: Phylogenetic reconstructions were characterized by well-supported monophyletic clades corresponding to traditionally circumscribed species, with the exception of a single taxon. Divergence estimates indicate that most diversification of the sampled Oropogon species occurred throughout the Oligocene and Miocene, although diversification of a single closely related clade appears to have occurred during the late Pliocene and into the Pleistocene. Divergence estimates calculated from a data set with ambiguously aligned regions removed were much more recent than those from the full data set. Conclusions: Overall, our analyses place the majority of divergence events of Oropogon species from the Oaxacan Highlands within the Neogene and provide strong evidence that climatic changes during the Pleistocene were not a major factor driving speciation in the lichenized genus Oropogon in neotropical highlands. [ABSTRACT FROM AUTHOR]

In North America, the widespread sorediate lichen known as Hyperphyscia adgiutinata is shown to be heterogeneous. It contains two distinct species, one with primarily laminal soralia and another with marginal soralia. The latter taxon is here described as the new species H. confusa. Hyperphyscia adgiutinata and H. confusa have broadly overlapping distributions in North America, especially in the Midwest and Great Plains states. Where they are sympatric, H. confusa is generally the more common of the two. Descriptions and discussions for H. adgiutinata and the two other sorediate North American species, H. minor and H. pyrithrocardia, are also included, along with a key for separation of the four species. Hyperphyscia pyrithrocardia is reported for the first time from North America. [ABSTRACT FROM AUTHOR]

• Premise of the study: Accurate species delimitation is important for understanding the diversification of biota and has critical implications for ecological and conservation studies. However, a growing body of evidence indicates that morphology-based species circumspection in lichenized fungi misrepresents fungal diversity. The foliose lichen genus Xanthoparmelia includes over 800 species displaying a complex array of morphological and secondary melabolite diversity. • Methods: We used a multifaceted approach, applying phylogenetic, population genetic, and genealogical analyses to delimit species in a single well-supported monophyletic clade containing 10 morphologically and chemically divcrsc Xanthoparntelia species in western North America. Sequence data from four ribosomal and two low-copy, protein-coding markers, along with chemical and morphological data were used to assess species diversity. • Key results: We found that traditionally circumscribed species are not supported by molecular data. Rather, all sampled taxa were better represented by three polymorphic population clusters. Our results suggest that secondary metabolite variation may have limited utility in diagnosing lineages within this group, while identified populations clusters did not rellect major phylogeographic or ecological patterns. • Conclusions: In contrast to studies revealing previously undiscovered fungal lineages masked within lichen species circumscribed by traditional morphological and chemical concepts, the present study suggests that species diversity has been overestimated in the species-rich genus Xanthoparmelia. [ABSTRACT FROM AUTHOR]

In the United States, research natural areas (RNAs) are permanently established to maintain areas of natural ecosystem significance. The Upper Sand Creek Research Natural Area (USCRNA) in the Escalante River watershed in southern Utah was established nearly a quarter century ago. However, the botanical diversity has remained largely unknown. To provide data to aid in preserving and maintaining biological and genetic diversity in USCRNA, we surveyed vascular plant, bryophyte, and lichen diversity within the RNA in 2021. We document high vascular plant (187 species) and lichen (152 species / candidate species) diversity, in addition to 15 bryophyte species. These preliminary inventories will be critical for investigating ecosystem function and integrity, providing a baseline for future research. [ABSTRACT FROM AUTHOR]

Simple Summary: This paper reports on two new species and two new records of the lichen-forming fungal genus Peltula Nyl., highlighting progress in our investigation of Chinese Peltula spp. The external morphology, anatomy, and molecular systematics were comprehensively analyzed to support the species delimitation. The results contribute to the knowledge of species diversity and geographical distributions of Peltula species in China. In this study, two new species, Peltula confusa Q.X. Yang & X.L. Wei, sp. nov., growing in a dry microenvironment within a semi-humid area, and Peltula subpatellata Q.X. Yang & X.L. Wei, sp. nov., occurring in arid and semi-arid regions, are described. In addition, two species, P. polyspora (Tuck.) Wetmore and P. obscuratula (Nyl.) Poelt ex Egea, are recorded for the first time in China. All four species are described based on morphological, anatomical and molecular data. Peltula confusa is characterized by a lighter, brighter, and more yellowish upper surface than other species of this genus, with discs concentrated in the central part of squamules, and a thick lower cortex. Peltula subpatellata can be distinguished from P. patellata (Bagl.) Swinscow & Krog by its non-thickened and sometimes darkened margins and sometimes the presence of peltate squamules. Phylogenetic analysis based on DNA sequences of four loci (ITS, nrSSU, nrLSU, and RPB2) demonstrates the placement of these species within Peltula, and supports current species delimitations. We suggest that the growth substrate should be considered as an additional characteristic for species delimitation. [ABSTRACT FROM AUTHOR]

The article focuses on the proposal to conserve the name Lichen muralis (Lecanora muralis, Protoparmeliopsis muralis) with a conserved type (Ascomycota: Lecanorales: Lecanoraceae). Topics discussed include Schreber's name for the fungi being illegitimate as he cited Lichen pallescens L. as a synonym, and the corticolous species is now being called Ochrolechia pallescens and is protected with a conserved type.

Recent work has suggested exceptional species-level diversity in the lichen-forming Lecanora polytropa complex (Lecanoraceae, Ascomycota). However, biogeographic patterns and the spatial structuring of this diversity remains poorly known. To investigate diversity across multiple spatial scales, we sampled members of this species complex from two distinct regions—the Pacific Coast Ranges in southern Alaska, USA, and montane habitats in Spain. We also included sequence data from several species within this complex that were recently described from populations in France. Using the standard DNA barcoding marker and a sequence-based species delimitation approach (ASAP), we inferred a total of 123 candidate species (SHs) within the Lecanora polytropa complex, 32 of which were sampled for the first time here. Of 123 SHs, 21 had documented intercontinental distributions, while the vast majority were found at much smaller spatial scales. From our samples collected from Alaska, USA, and Spain, representing 36 SHs, we found high genetic diversity occurring within each sampled site, but limited overlap among all sites. Mountain ranges in both regions had high proportions of endemic lineages, with the highest diversity and endemism occurring in mountain ranges in Spain. Our sequence data generally support the recent taxonomic proposals, and an integrative taxonomy may help partly resolve the taxonomic conundrums within this hyper-diverse lineage. [ABSTRACT FROM AUTHOR]

Species of lichen-forming fungi (LFF) display an array of geographical distribution patterns. Among the broadly distributed lichen-forming fungal species, the degree of reproductive isolation and genetic substructure among populations varies widely, in some cases masking unrecognized diversity or meaningful biogeographical patterns. Lecidea atrobrunnea (Raymond ex Lam. & DC.) Schaer. s. lat. (Lecideaceae) is a widespread species complex that has been studied for over two centuries since its initial description. The diversity of the L. atrobrunnea group is highest in western North America, where a dizzying array of morphologies and chemistry can occur at local scales. Here we investigate whether the assumed cosmopolitan distribution of L. atrobrunnea s. lat. is an artifact of taxonomic limitations and masks biogeographical patterns in this species complex. To address these questions, we compiled sequence data from the standard fungal barcoding marker (ITS) for over 100 specimens within this complex, in addition to genome-scale data from a subset of these representing over 1600 single-copy nuclear genes spanning over 3 Mb of the genome. Our study corroborates the perspective that the morphologically and chemically variable Lecidea atrobrunnea group reflects a complex of distinct species-level lineages, with 42–83 candidate species inferred from the ITS region and high levels of diversity inferred from a subset of specimens using genome-scale data. However, both phenotype- and molecular-based species boundaries remained unsettled, with the most common nominal taxa recovered as highly polyphyletic and with conflict among different molecular species delimitation approaches. Our study also highlights the potential for geographically restricted species, with fascinating biogeographical patterns, challenging, in part, the assumed cosmopolitan distribution of L. atrobrunnea s. lat. This study provides valuable direction for future research that will be crucial in understanding diversification and establishing a robust taxonomy for this well-known species complex. [ABSTRACT FROM AUTHOR]

Subalpine habitats in sky islands in the Southwestern USA are currently facing large-scale transformations. Lichens have widely been used as bioindicators of environmental change. On the Colorado Plateau, fruticose lichens occur in patchy, disconnected populations, including unique lichen-draped conifer sites in subalpine forests in the La Sal Mountains in southeastern Utah. Here, we document the distribution and fungal diversity within these lichen communities. We find that lichen-draped conifer sites in the La Sal Mountains are restricted to only three known, small areas in Picea englemannii forests above 3000 m above sea level, two of which have recently been impacted by wildfire. We document 30 different species of lichen-forming fungi in these communities, several which represent the first reports from the Colorado Plateau. We also characterize mycobiont haplotype diversity for the fruticose lichens Evernia divaricata, Ramalina sinensis, and multiple Usnea species. We also report a range of diverse fungi associated with these lichens, including genetic clusters representing 22 orders spanning seven classes of Ascomycetes and fewer clusters representing Basidiomycetes. Our results provide a baseline for ongoing monitoring and help to raise awareness of unique lichen communities and other biodiversity in the region. [ABSTRACT FROM AUTHOR]

Biodiversity inventories are important for informing land management strategies, conservation efforts, and for biomonitoring studies. For many organismal groups, including lichens, comprehensive, accurate inventories are challenging due to the necessity of taxonomic expertise, limitations in sampling protocols, and the commonplace occurrence of morphologically cryptic species and other undescribed species. Lichen communities in arid regions are often dominated by crustose lichens, which have been particularly difficult to incorporate into biodiversity inventories. Here, we explore the utility of DNA metabarcoding for characterizing the diversity of lichen-forming fungi at a typical crustose lichen-dominated site on the Colorado Plateau in the southwestern USA. We assessed the consistency of independent sampling efforts to comprehensively document lichen diversity, evaluated the capability of minimally trained technicians to effectively sample the lichen communities, and provide a metagenomic-based inventory of lichen diversity, including representative sequence data, for a diverse, crustose-dominate lichen community on the Colorado Plateau. Our results revealed that crustose lichen communities in the southwestern USA are more diverse than traditionally thought, and community metabarcoding is a promising strategy for characterizing the lichen-forming fungal diversity more thoroughly than other methods. However, consistently sampling the diversity of crustose lichen communities, even at small spatial scales, remains difficult. Interpreting these results within a traditional taxonomic context remains challenging without the use of vouchers. [ABSTRACT FROM AUTHOR]

[Display omitted] • Wanderlust lichens occur in isolated populations in steppe habitats in North America. • A 19 Mb alignment revealed high, spatially structured genetic diversity. • R. robusta , a sexually reproducing taxon, is nested within this asexual lineage. • Wanderlust lichens don't fully occupy the areas of highest distribution probability. The northern North American Cordillera is a globally significant center of endemism. In western North America, imperiled arid steppe habitats support a number of unique species, including several endemic lichens. However, processes driving diversification and endemism in this region remain unclear. In this study, we investigate diversity and phylogeography of the threatened wanderlust lichens (mycobiont = Rhizoplaca species) which occur unattached on calcareous soils in steppe habitats. Wanderlust lichens comprise three species of lichen-forming fungi (LFF) – Rhizoplaca arbuscula , R. haydenii , and R. idahoensis (endangered, IUCN Red List) – which occur in fragmented populations in Idaho and Wyoming, with more limited populations in southern Montana and northern Utah. These lichens reproduce almost exclusively via large, asexual vegetative propagules. Here, our aims were to (i) assess the evolutionary origin of this group and identify phylogeographic structure, (ii) infer ancestral geographic distributions for lineages within this clade, and (iii) use species distribution modeling to better understand the distribution of contemporary populations. Using a genome-skimming approach, we generated a 19.1Mb alignment, spanning ca. half of the complete LFF genome, from specimens collected throughout the entire range of wanderlust lichens. Based on this phylogeny, we investigated phylogeographic patterns using RASP. Finally, we used MaxEnt to estimate species distribution models for R. arbuscula and R. haydenii. We inferred a highly structured topology, with clades corresponding to distinct geographic regions and morphologies collected throughout the group's distribution. We found that R. robusta , a sexually reproducing taxon, is clearly nested within the vagrant Rhizoplaca clade. Phylogeographic analyses suggest that both dispersal and vicariance played significant roles throughout the evolutionary history of the vagrant Rhizoplaca clade, with most of the dispersal events originating from the Salmon Basin in eastern Idaho – the center of diversity for this group. Despite the fact that wanderlust lichens are dispersal limited due to large, unspecialized vegetative propagules, we inferred multiple dispersal events crossing the Continental Divide. Comparing herbarium records with species distribution models suggests that wanderlust lichens don't fully occupy the areas of highest distribution probability. In fact, documented records often occur in areas predicted to be only marginally suitable. These data suggest a potential mismatch between contemporary habitats outside of the center of diversity in eastern Idaho with the most suitable habitat, adding to the vulnerability of this imperiled complex of endemic lichens. [ABSTRACT FROM AUTHOR]

Robust species delimitations provide a foundation for investigating speciation, phylogeography, and conservation. Here we attempted to elucidate species boundaries in the cosmopolitan lichen-forming fungal taxon Lecanora polytropa. This nominal taxon is morphologically variable, with distinct populations occurring on all seven continents. To delimit candidate species, we compiled ITS sequence data from populations worldwide. For a subset of the samples, we also generated alignments for 1209 single-copy nuclear genes and an alignment spanning most of the mitochondrial genome to assess concordance among the ITS, nuclear, and mitochondrial inferences. Species partitions were empirically delimited from the ITS alignment using ASAP and bPTP. We also inferred a phylogeny for the L. polytropa clade using a four-marker dataset. ASAP species delimitations revealed up to 103 species in the L. polytropa clade, with 75 corresponding to the nominal taxon L. polytropa. Inferences from phylogenomic alignments generally supported that these represent evolutionarily independent lineages or species. Less than 10% of the candidate species were comprised of specimens from multiple continents. High levels of candidate species were recovered at local scales but generally with limited overlap across regions. Lecanora polytropa likely ranks as one of the largest species complexes of lichen-forming fungi known to date. [ABSTRACT FROM AUTHOR]

Species delimitation among closely related species is challenging because traditional phenotype‐based approaches, for example, using morphology, ecological, or chemical characteristics, may not coincide with natural groupings. With the advent of high‐throughput sequencing, it has become increasingly cost‐effective to acquire genome‐scale data which can resolve previously ambiguous species boundaries. As the availability of genome‐scale data has increased, numerous species delimitation analyses, such as BPP and SNAPP+Bayes factor delimitation (BFD*), have been developed to delimit species boundaries. However, even empirical molecular species delimitation approaches can be biased by confounding evolutionary factors, for example, hybridization/introgression and incomplete lineage sorting, and computational limitations. Here, we investigate species boundaries and the potential for micro‐endemism in a lineage of lichen‐forming fungi, Niebla Rundel & Bowler, in the family Ramalinaceae by analyzing single‐locus and genome‐scale data consisting of (a) single‐locus species delimitation analysis using ASAP, (b) maximum likelihood‐based phylogenetic tree inference, (c) genome‐scale species delimitation models, e.g., BPP and SNAPP+BFD, and (d) species validation using the genealogical divergence index (gdi). We specifically use these methods to cross‐validate results between genome‐scale and single‐locus datasets, differently sampled subsets of genomic data and to control for population‐level genetic divergence. Our species delimitation models tend to support more speciose groupings that were inconsistent with traditional taxonomy, supporting a hypothesis of micro‐endemism, which may include morphologically cryptic species. However, the models did not converge on robust, consistent species delimitations. While the results of our analysis are somewhat ambiguous in terms of species boundaries, they provide a valuable perspective on how to use these empirical species delimitation methods in a nonmodel system. This study thus highlights the challenges inherent in delimiting species, particularly in groups such as Niebla, with complex, relatively recent phylogeographic histories. [ABSTRACT FROM AUTHOR]

Background: Regions within the nuclear ribosomal operon are a major tool for inferring evolutionary relationships and investigating diversity in fungi. In spite of the prevalent use of ribosomal markers in fungal research, central features of nuclear ribosomal DNA (nrDNA) evolution are poorly characterized for fungi in general, including lichenized fungi. The internal transcribed spacer (ITS) region of the nrDNA has been adopted as the primary DNA barcode identification marker for fungi. However, little is known about intragenomic variation in the nrDNA in symbiotic fungi. In order to better understand evolution of nrDNA and the utility of the ITS region for barcode identification of lichen-forming fungal species, we generated nearly complete nuclear ribosomal operon sequences from nine species in the Rhizoplaca melanophthalma species complex using short reads from high-throughput sequencing. Results: We estimated copy numbers for the nrDNA operon, ranging from nine to 48 copies for members of this complex, and found low levels of intragenomic variation in the standard barcode region (ITS). Monophyly of currently described species in this complex was supported in phylogenetic inferences based on the ITS, 28S, intergenic spacer region, and some intronic regions, independently; however, a phylogenetic inference based on the 18S provided much lower resolution. Phylogenetic analysis of concatenated ITS and intergenic spacer sequence data generated from 496 specimens collected worldwide revealed previously unrecognized lineages in the nrDNA phylogeny. Conclusions: The results from our study support the general assumption that the ITS region of the nrDNA is an effective barcoding marker for fungi. For the R. melanophthalma group, the limited amount of potential intragenomic variability in the ITS region did not correspond to fixed diagnostic nucleotide position characters separating taxa within this species complex. Previously unrecognized lineages inferred from ITS sequence data may represent undescribed species-level lineages or reflect uncharacterized aspects of nrDNA evolution in the R. melanophthalma species complex. [ABSTRACT FROM AUTHOR]

• Comprehensive molecular phylogeny of the Holarctic clade of lichen-forming fungi genus Xanthoparmelia. • Despite reconstructing relatively consistent phylogenetic trees, rampant phylogenetic conflict was revealed. • Widespread hybridization was detected among species in the most diverse genus of lichen-forming fungi using three independent tests for hybridization. While advances in sequencing technologies have been invaluable for understanding evolutionary relationships, increasingly large genomic data sets may result in conflicting evolutionary signals that are often caused by biological processes, including hybridization. Hybridization has been detected in a variety of organisms, influencing evolutionary processes such as generating reproductive barriers and mixing standing genetic variation. Here, we investigate the potential role of hybridization in the diversification of the most speciose genus of lichen-forming fungi, Xanthoparmelia. As Xanthoparmelia is projected to have gone through recent, rapid diversification, this genus is particularly suitable for investigating and interpreting the origins of phylogenomic conflict. Focusing on a clade of Xanthoparmelia largely restricted to the Holarctic region, we used a genome skimming approach to generate 962 single-copy gene regions representing over 2 Mbp of the mycobiont genome. From this genome-scale dataset, we inferred evolutionary relationships using both concatenation and coalescent-based species tree approaches. We also used three independent tests for hybridization. Although different species tree reconstruction methods recovered largely consistent and well-supported trees, there was widespread incongruence among individual gene trees. Despite challenges in differentiating hybridization from ILS in situations of recent rapid radiations, our genome-wide analyses detected multiple potential hybridization events in the Holarctic clade, suggesting one possible source of trait variability in this hyperdiverse genus. This study highlights the value in using a pluralistic approach for characterizing genome-scale conflict, even in groups with well-resolved phylogenies, while highlighting current challenges in detecting the specific impacts of hybridization. [ABSTRACT FROM AUTHOR]

DNA barcoding approaches provide powerful tools for characterizing fungal diversity. However, DNA barcoding is limited by poor representation of species-level diversity in fungal sequence databases. Can the development of custom, regionally focused DNA reference libraries improve species-level identification rates for lichen-forming fungi? To explore this question, we created a regional ITS database for lichen-forming fungi (LFF) in the Intermountain West of the United States. The custom database comprised over 4800 sequences and represented over 600 formally described and provisional species. Lichen communities were sampled at 11 sites throughout the Intermountain West, and LFF diversity was characterized using high-throughput ITS2 amplicon sequencing. We compared the species-level identification success rates from our bulk community samples using our regional ITS database and the widely used UNITE database. The custom regional database resulted in significantly higher species-level assignments (72.3%) of candidate species than the UNITE database (28.3-34.2%). Within each site, identification of candidate species ranged from 72.3-82.1% using the custom database; and 31.5-55.4% using the UNITE database. These results highlight that developing regional databases may accelerate a wide range of LFF research by improving our ability to characterize species-level diversity using DNA barcoding.

Thirty years after its designation as a federally endangered species, the Florida Perforate Cladonia (FPC) remains imperiled in isolated populations in the Florida scrub in the southeastern USA. For threatened and endangered species, such as FPC, reference genomes provide critical insight into genomic diversity, local adaptations, landscape-level genetics, and phylogenomics. Using high-throughput sequencing, we assemble the first draft nuclear and mitochondrial genomes for the FPC mycobiont- Cladonia perforata . We also assess genetic diversity within and among populations in southeastern Florida using genome-scale data and investigate diversity across the entire nuclear ribosomal cistron, including the standard DNA barcoding marker for fungi. The draft nuclear genome spanned 33.6 Mb, and the complete, circular mitochondrial genome was 59 Kb. We also generated the first chloroplast genome, to our knowledge, for the photobiont genus associated with FPC, an undescribed Asterochloris species. We inferred the presence of multiple, distinct mycobiont parental genotypes (genets) occurring at local scales in southeastern Florida, and strikingly, no genets were shared among even the closest sample sites. All sampled thalli shared identical mitochondrial genomes, while the nuclear ribosomal cistron showed limited variability-highlighting the genetic resolution provided by nuclear genome-scale datasets. The genomic resources generated here provide critical resources for informed conservation efforts for the FPC.

Lichens are well-known examples of complex symbiotic associations between organisms from different Kingdoms. Microfungi in particular, establish diverse associations with the hosting lichen thallus, as species-specific parasites or transient co-inhabitants. The whole community of lichen-associated fungi constitute the 'lichen mycobiome' comprising both ascomycetes and basidiomycetes, including filamentous and yeast taxa. Metabarcoding results and microscopy analyses show that in some thalli, basidiomycetes are frequent lichen-associated fungi but still only a few species could be axenically isolated and morphologically characterized. Within a broad project aiming at characterizing the mycobiome diversity by culture-dependent and independent approaches in two lichen species selected as reference models - Rhizoplaca melanophthalma and Tephromela atra, we succeed in isolating and culturing 76 new strains of basidiomycetous yeasts. The lichen thalli were collected in different mountain regions worldwide and at relatively high elevation. The yeast strains were isolated on different growth media and were studied for their morphological and genetic diversity. Nuclear internal transcribed spacer (ITS) and ribosomal large subunit (LSU) sequence analyses identified them to belong to ten families within the orders Agaricostilbomycetes, Cystobasidiomycetes, Microbotryomycetes, Tremellomycetes and Ustilaginomycetes. The yeasts here detected showed patterns of host-preference in a few cases and they are potentially related to the ecological conditions., Competing Interests: Declaration of competing interest None., (Copyright © 2022 British Mycological Society. Published by Elsevier Ltd. All rights reserved.)

Fungal-algal relationships-both across evolutionary and ecological scales-are finely modulated by the presence of the symbionts in the environments and by the degree of selectivity and specificity that either symbiont develop reciprocally. In lichens, the green algal genus Trebouxia Puymaly is one of the most frequently recovered chlorobionts. Trebouxia species-level lineages have been recognized on the basis of their morphological and phylogenetic diversity, while their ecological preferences and distribution are still only partially unknown. We selected two cosmopolitan species complexes of lichen-forming fungi as reference models, i.e., Rhizoplaca melanophthalma and Tephromela atra , to investigate the diversity of their associated Trebouxia spp. in montane habitats across their distributional range worldwide. The greatest diversity of Trebouxia species-level lineages was recovered in the altitudinal range 1,000-2,500 m a.s.l. A total of 10 distinct Trebouxia species-level lineages were found to associate with either mycobiont, for which new photobionts are reported. One previously unrecognized Trebouxia species-level lineage was identified and is here provisionally named Trebouxia "A52." Analyses of cell morphology and ultrastructure were performed on axenically isolated strains to fully characterize the new Trebouxia "A52" and three other previously recognized lineages, i.e., Trebouxia "A02," T. vagua "A04," and T. vagua "A10," which were successfully isolated in culture during this study. The species-level diversity of Trebouxia associating with the two lichen-forming fungi in extreme habitats helps elucidate the evolutionary pathways that this lichen photobiont genus traversed to occupy varied climatic and vegetative regimes., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 De Carolis, Cometto, Moya, Barreno, Grube, Tretiach, Leavitt and Muggia.)

Lichen associations are overwhelmingly supported by carbon produced by photosynthetic algal symbionts. These algae have diversified to occupy nearly all climates and continents; however, we have a limited understanding of how their climatic niches have evolved through time. Here we extend previous work and ask whether phylogenetic signal in, and the evolution of, climatic niche, varies across climatic variables, phylogenetic scales, and among algal lineages in Trebouxia- the most common genus of lichen-forming algae. Our analyses reveal heterogeneous levels of phylogenetic signal across variables, and that contrasting models of evolution underlie the evolution of climatic niche divergence. Together these analyses demonstrate the variable processes responsible for shaping climatic tolerance in Trebouxia , and provide a framework within which to better understand potential responses to climate change-associated perturbations. Such predictions reveal a disturbing trend in which the pace at which modern climate change is proceeding will vastly exceed the rate at which Trebouxia climatic niches have previously evolved., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Nelsen, Leavitt, Heller, Muggia and Lumbsch.)

Advancements in molecular genetics have revealed that hybridization may be common among plants, animals, and fungi, playing a role in evolutionary dynamics and speciation. While hybridization has been well-documented in pathogenic fungi, the effects of these processes on speciation in fungal lineages with different life histories and ecological niches are largely unexplored. Here we investigated the potential influence of hybridization on the emergence of morphologically and reproductively distinct asexual lichens. We focused on vagrant forms (growing obligately unattached to substrates) within a clade of rock-dwelling, sexually reproducing species in the Rhizoplaca melanophthalma (Lecanoraceae, Ascomycota) species complex. We used phylogenomic data from both mitochondrial and nuclear genomes to infer evolutionary relationships and potential patterns of introgression. We observed multiple instances of discordance between the mitochondrial and nuclear trees, including the clade comprising the asexual vagrant species R. arbuscula, R. haydenii, R. idahoensis, and a closely related rock-dwelling lineage. Despite well-supported phylogenies, we recovered strong evidence of a reticulated evolutionary history using a network approach that incorporates both incomplete lineage sorting and hybridization. These data suggest that the rock-dwelling western North American subalpine endemic R. shushanii is potentially the result of a hybrid speciation event, and introgression may have also played a role in other taxa, including vagrant species R. arbuscula, R. haydenii and R. idahoensis. We discuss the potential roles of hybridization in terms of generating asexuality and novel morphological traits in lichens. Furthermore, our results highlight the need for additional study of reticulate phylogenies when investigating species boundaries and evolutionary history, even in cases with well-supported topologies inferred from genome-scale data.