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Desiccation–rehydration experiments have been employed over the years to evaluate desiccation tolerance of bryophytes (Bryophyta, Marchantiophyta, and Anthocerotophyta). Researchers have applied a spectrum of protocols to induce desiccation and subsequent rehydration, and a wide variety of techniques have been used to study desiccation-dependent changes in bryophyte molecular, cellular, physiological, and structural traits, resulting in a multifaceted assortment of information that is challenging to synthesize. We analysed 337 desiccation–rehydration studies, providing information for 351 species, to identify the most frequent methods used, analyse the advances in desiccation studies over the years, and characterize the taxonomic representation of the species assessed. We observed certain similarities across methodologies, but the degree of convergence among the experimental protocols was surprisingly low. Out of 52 bryophyte orders, 40% have not been studied, and data are lacking for multiple remote or difficult to access locations. We conclude that for quantitative interspecific comparisons of desiccation tolerance, rigorous standardization of experimental protocols and measurement techniques, and simultaneous use of an array of experimental techniques are required for a mechanistic insight into the different traits modified in response to desiccation. New studies should also aim to fill gaps in taxonomic, ecological, and spatial coverage of bryophytes. [ABSTRACT FROM AUTHOR]

Accurate species delimitation has a pivotal role in conservation biology, and it is especially important for threatened species where decisions have political and economic consequences. Finding and applying appropriate character sets and analytical tools to resolve interspecific relationships remains challenging in lichenized fungi. The main aim of our study was to re-assess the species boundaries between Usnea subfloridana and Usnea florida , which have been phylogenetically indistinguishable until now, but are different in reproductive mode and ecological preferences, using fungal-specific simple sequence repeats (SSR), i.e. microsatellite markers. Bayesian clustering analysis, discriminant analysis of principal components (DAPC), minimal spanning network (MSN), and principal component analysis (PCA) failed to separate U. florida and U. subfloridana populations. However, a low significant differentiation between the two taxa was observed across all populations according to AMOVA results. Also, analysis of shared haplotypes and statistical difference in clonal diversity (M) supported the present-day isolation between the apotheciate U. florida and predominantly sorediate U. subfloridana. Our results do not provide a clear support either for the separation of species in this pair or the synonymization of U. florida and U. subfloridana. We suggest that genome-wide data could help resolve the taxonomic question in this species pair. • Species boundaries between Usnea florida and Usnea subfloridana were re-assessed. • Microsatellite markers were used for species delimitation purposes. • A high level of gene flow must have occurred between Usnea populations. • Our study does not support either for separation or synonymization of study taxa. [ABSTRACT FROM AUTHOR]

Summary: The popular dual definition of lichen symbiosis is under question with recent findings of additional microbial partners living within the lichen body. Here we compare the distribution and co‐occurrence patterns of lichen photobiont and recently described secondary fungus (Cyphobasidiales yeast) to evaluate their dependency on lichen host fungus (mycobiont).We sequenced the nuclear internal transcribed spacer (ITS) strands for mycobiont, photobiont, and yeast from six widespread northern hemisphere epiphytic lichen species collected from 25 sites in Switzerland and Estonia. Interaction network analyses and multivariate analyses were conducted on operational taxonomic units based on ITS sequence data.Our study demonstrates the frequent presence of cystobasidiomycete yeasts in studied lichens and shows that they are much less mycobiont‐specific than the photobionts. Individuals of different lichen species growing on the same tree trunk consistently hosted the same or closely related mycobiont‐specific Trebouxia lineage over geographic distances while the cystobasidiomycete yeasts were unevenly distributed over the study area – contrasting communities were found between Estonia and Switzerland.These results contradict previous findings of high mycobiont species specificity of Cyphobasidiales yeast at large geographic scales. Our results suggest that the yeast might not be as intimately associated with the symbiosis as is the photobiont. See also the Commentary on this article by Hawksworth & Grube, 227: 1281–1283. [ABSTRACT FROM AUTHOR]

National Red Lists are useful tools in establishing local conservation priorities. The threat status of Estonian lichens based on the IUCN system has been assessed twice, in 2008 and 2023. In the latest Red List, the proportion of species of elevated conservation concern, that is taxa belonging to the Near Threatened and threatened categories or having become regionally extinct in Estonia, was high (58%) while Least Concern (LC) species represented one-third (36%) of all taxa. Macrolichens were more threatened than microlichens. The Red List Index (RLI), illustrating the trends of species in their projected extinction risk, was calculated. The values were low (< 0.7 in 2023), thus indicating a heightened risk both for the set of all species and for macrolichens. More than half of all Estonian lichens are associated with woodlands and 54% of these species are of elevated conservation concern. Lichens preferring broad-leaved deciduous trees included more threatened than LC species, while among lichens preferring other deciduous or coniferous trees the proportion of LC species was higher than that of threatened taxa. Lichens inhabiting calcareous grasslands had the highest share (69%) of taxa of elevated conservation concern. Comparisons of national red-listed data with four selected countries (Czech Republic, Denmark, Finland and Sweden) revealed that the biggest overlaps of Estonian taxa of elevated conservation concern were with the Czech Republic and Finland. [ABSTRACT FROM AUTHOR]

Abstract 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. Highlights • A well-resolved multilocus phylogeny was recovered in lichen genus Cetrelia. • Chemotypes based on secondary compounds clearly correlated with gene genealogy. • Species of the same reproductive mode were dispersed throughout phylogenetic tree. • Delimiting Cetrelia species by combining morpho- and chemotypes is partly supported. • Chemical evolution in Cetrelia has probably been towards more complex substances. [ABSTRACT FROM AUTHOR]

Desiccation-rehydration studies in cryptogams constitute an important tool to understand the relation of key physiological traits with species stress tolerance and environmental adaptability. Real-time monitoring of responses has been limited by the design of commercial or custom measuring cuvettes and difficulties in experimental manipulation. We developed a within-chamber rehydration method that allows to rewater the samples rapidly, without the need to open the chamber and take out the sample for manual rehydration by the investigator. Data is collected in real-time and simultaneously with an infrared gas-analyzer (LICOR-7000), a chlorophyll fluorometer (Maxi Imaging-PAM) and a proton transfer reaction time-of-flight mass-spectrometer (PTR-TOF-MS) for volatile organic compound emissions. The system was tested on four cryptogam species with contrasting ecological distributions. No major errors or kinetics disruptions were found during system testing and measurements. Our within-chamber rehydration method improved accuracy, as measurement periods were not lacking, and repeatability of the protocol by reducing error variance in sample manipulation. This method provides an improved technique to conduct desiccation-rehydration measurements, contributing to the standardization and accuracy of current existing methodologies. A close real-time and simultaneous monitoring of photosynthesis, chlorophyll fluorescence and volatile organic compound emission data, offers a novel perspective in the analysis of the cryptogam stress responses that is yet to be fully explored. • We present a custom-built gas-exchange system for desiccation-rehydration measurements. • Continuous real-time measurements of photosynthesis, Chl fluorescence and VOC data. • In-chamber rehydration reduces gaps in measurement data and sample manipulation. • We observe contrasting stress responses from ecologically different mosses and lichens. [ABSTRACT FROM AUTHOR]

Although lichens (lichen-forming fungi) play an important role in the ecological integrity of many vulnerable landscapes, only a minority of lichen-forming fungi have been barcoded out of the currently accepted ∼18 000 species. Regular Sanger sequencing can be problematic when analyzing lichens since saprophytic, endophytic, and parasitic fungi live intimately admixed, resulting in low-quality sequencing reads. Here, high-throughput, long-read 454 pyrosequencing in a GS FLX+ System was tested to barcode the fungal partner of 100 epiphytic lichen species from Switzerland using fungal-specific primers when amplifying the full internal transcribed spacer region (ITS). The present study shows the potential of DNA barcoding using pyrosequencing, in that the expected lichen fungus was successfully sequenced for all samples except one. Alignment solutions such as BLAST were found to be largely adequate for the generated long reads. In addition, the NCBI nucleotide database-currently the most complete database for lichen-forming fungi-can be used as a reference database when identifying common species, since the majority of analyzed lichens were identified correctly to the species or at least to the genus level. However, several issues were encountered, including a high sequencing error rate, multiple ITS versions in a genome (incomplete concerted evolution), and in some samples the presence of mixed lichen-forming fungi (possible lichen chimeras). [ABSTRACT FROM AUTHOR]

Recent taxonomic and DNA sequence-based studies in several groups of lichen-forming fungi have revealed incongruence between the morphological and molecule-based circumscriptions of species. While the cosmopolitan genus Usnea is well-known and easily recognized by the yellowish beard-like thallus with central cord, delimitation of many Usnea species is difficult due to the high variation and complexity of diagnostic characters. In this study, we assessed the monophyly of 18 species from section Usnea occurring in North America and Europe, including sorediate and sexually reproducing taxa with both pendent and shrubby thalli. Six nuclear markers (ribosomal internal transcribed spacer (ITS) and intergenic spacer (IGS), and protein-coding beta-tubulin, MCM7, RPB1 and RPB2) were sequenced for 144 samples. All analyzed loci show weak genetic structure and short branch lengths in single-locus topologies, suggesting recent diversification history of the sampled taxa. Concatenated, multi-locus analyses conducted in Bayesian and maximum likelihood frameworks, as well as coalescent-based species delimitation and species tree methods, recover several distinct clades, some represent traditional morphology-based species ( Usnea cavernosa, U. praetervisa, U. silesiaca, U. wasmuthii), while others form clusters of two or more species ( Usnea florida- U. subfloridana, U. fulvoreagens- U. glabrescens, U. barbata- U. chaetophora- U. dasopoga- U. diplotypus, U. barbata- U. intermedia- U. lapponica- U. substerilis). We propose synonymization of U. substerilis under U. lapponica. The status of several other species within intermixed clusters requires further evaluation with more extensive sampling and the inclusion of more variable markers before taxonomic consequences can be considered. A new species, Usnea parafloridana is described from Wisconsin, USA. [ABSTRACT FROM AUTHOR]

Premise of the study: Species boundaries in many organism groups are still in a state of flux, and for empirical species delimitation, finding appropriate character sets and analytical tools are among the greatest challenges. In the lichenized fungal genus Vulpicida, six morphologically circumscribed species have been distinguished, but phenotypic characters partly overlap for three of these and intermediate forms occur. We used a combination of phylogenetic strategies to delimit the species in this genus. • Methods: Five DNA loci were sequenced and analyzed. Single-locus gene trees and a five-locus concatenated phylogeny were constructed to assess current Vulpicida species. Species boundaries were inferred from molecular data using two coalescentbased species delimitation methods (BP&P and Brownie) and from species trees reconstructed with three different algorithms (BEAST, BEST, and STEM). • Key results: The two species restricted to North America, Vulpicida canadensis and V. viridis, are clearly distinct in all analyses. The four other traditionally accepted species form two strongly supported, closely related species-level lineages within the core group of the genus. On the basis of these results, we propose four instead of the current six species in the genus: V. canadensis, V. juniperinus, V. pinastri, and V. viridis, while V. tilesii and V. tubulosus are reduced to synonymy under V. juniperinus. • Conclusions: Coalescent species delimitation and tree inference give consistent results for fully distinct Vulpicida species but not for diverging populations. Even the inconsistent results were informative, revealing developing isolation despite a complex history of recombination and incomplete lineage sorting. [ABSTRACT FROM AUTHOR]

Phenotypic species recognition in the genus I Usnea i is particularly complicated; the species are delimited by distinctive combinations of diagnostic morphological traits (Clerc 1998, 2011) which may, however, in certain cases be poorly developed or even absent (Clerc 2011). Clerc & Naciri (2021) conclude that I Usnea barbata i and I U. dasopoga i are morphologically and anatomically truly distinct species for which molecular data do not support their conspecificity; however, phylogenetic analyses of molecular data (including the analyses performed in Mark I et al i . Clerc & Naciri (2021) revise the traditional identification of 35 samples used in our analyses (table 1 in Clerc & Naciri (2021)) and present the details of their morphological and chemical characters. With Usnea it is truly a very complicated art and, as demonstrated by Clerc & Naciri (2021), in solitary cases the traditional morphological identification could not even be confirmed by the discriminant a posteriori analysis based on anatomical measures (table 1 in Clerc & Naciri (2021)). [Extracted from the article]

The delimitation of two morphologically similar and not easily separable Vulpicida species, V. juniperinus and V. tubulosus, is analyzed using nuclear ITS and Mcm7, and mitochondrial SSU DNA sequences. Seventy-nine Vulpicida specimens, most from the two focal taxa, are included in the three-locus gene tree. The results from Bayesian and parsimony analyses are presented. There are strong conflicts between the single locus gene trees. Vulpicida juniperinus and V. tubulosus are divided into two clearly distinguished groups in the ITS and concatenated B/MCMC tree. However, these species are mixed in both clades, appearing polyphyletic. Currently accepted V. juniperinus and V. tubulosus are not distinct according to the loci studied. Vulpicida pinastri appears monophyletic based on the available sequences. [ABSTRACT FROM AUTHOR]

Due to the importance of Rubisco in the biosphere, its kinetic parameters have been measured by different methodologies in a large number of studies over the last 60 years. These parameters are essential to characterize the natural diversity in the catalytic properties of the enzyme and they are also required for photosynthesis and cross-scale crop modeling. The present compilation of Rubisco kinetic parameters in model species revealed a wide intraspecific laboratory-to-laboratory variability, which was partially solved by making corrections to account for differences in the assay buffer composition and in the acidity constant of dissolved CO2, as well as for differences in the CO2 and O2 solubilities. Part of the intraspecific variability was also related to the different analytical methodologies used. For instance, significant differences were found between the two main methods for the determination of the specificity factor (S c/o), and also between Rubisco quantification methods, Rubisco purification versus crude extracts, and single-point versus CO2 curve measurements for the carboxylation turnover rate (k catc) determination. Causes of the intraspecific laboratory-to-laboratory variability for Rubisco catalytic traits are discussed. This study provides a normalized kinetic dataset for model species to be used by the scientific community. Corrections and recommendations are also provided to reduce measurement variability, allowing the comparison of kinetic data obtained in different laboratories using different assay conditions. [ABSTRACT FROM AUTHOR]

Summary: Photosynthesis in bryophytes and lycophytes has received less attention than terrestrial plant groups. In particular, few studies have addressed the nonstomatal diffusion conductance to CO2gnsd of these plant groups.Their lower photosynthetic rate per leaf mass area at any given nitrogen concentration compared with vascular plants suggested a stronger limitation by CO2 diffusion. We hypothesized that bryophyte and lycophyte photosynthesis is largely limited by low gnsd. Here, we studied CO2 diffusion inside the photosynthetic tissues and its relationships with photosynthesis and anatomical parameters in bryophyte and lycophyte species in Antarctica, Australia, Estonia, Hawaii and Spain.On average, lycophytes and, specially, bryophytes had the lowest photosynthetic rates and nonstomatal diffusion conductance reported for terrestrial plants. These low values are related to their very thick cell walls and their low exposure of chloroplasts to cell perimeter.We conclude that the reason why bryophytes lie at the lower end of the leaf economics spectrum is their strong nonstomatal diffusion conductance limitation to photosynthesis, which is driven by their specific anatomical characteristics. [ABSTRACT FROM AUTHOR]