Abstract Genomic variation within populations reflects both past and contemporary evolutionary processes. The genetic structure of the European peatland plant species is shaped by complex postglacial recolonization and recent habitat loss. Here, we investigate population genomic patterns in the vulnerable mire plant, round-leaved sundew ( Drosera rotundifolia L.). Using ddRAD sequencing of 311 individuals from 38 populations across Europe, we detected significant genetic differentiation (Fst = 0.02-0.44, p < 0.05) and a consistent deficit of heterozygosity (Fis = 0.248) across populations. In contrast, sequencing of 10,449 bp of chloroplast DNA revealed extremely low variation, with only one SNP detected. Clustering analyses (Admixture and DAPC) both identified pronounced genetic structure comprising three major clusters (Western, Northern, and Eastern) that broadly correspond to European biogeographic regions. Genetic differentiation was partially explained by geographic distance (3.3%; p = 0.0001; Mantel test), while climatic variables, particularly temperature and precipitation, accounted for 2.9% of genomic variation ( p < 0.0001; redundancy analysis). The observed pattern is consistent with polygenic responses to climatic gradients and is supported by 1,022 SNPs, distributed across 632 loci, that were significantly associated with environmental variables. Demographic reconstruction revealed distinct evolutionary trajectories among clusters, with the Western cluster showing a more recent expansion. Together, our results suggest that the genetic structure of D. rotundifolia is a result of postglacial recolonization, geographic isolation, and climate influence. We highlight the importance of conserving genetic diversity of D. rotundifolia across all three clusters and accounting for potential local adaptation and population vulnerability in future conservation strategies.

Abstract Root-associated fungal (RAF) communities can influence tree nutrition and performance, yet their environmental drivers and relationship with tree growth remain poorly understood, particularly near treelines. We characterized RAF communities, with a focus on ectomycorrhizal (ECM) fungi, associated with fine roots of white spruce ( Picea glauca [Moench] Voss) across paired forest and treeline plots representing two elevational and one drought-limited treeline ecotones in Alaska. Using ITS2 DNA metabarcoding, we assessed fungal community composition and alpha diversity and related these metrics to basal area increment (BAI) over 5–30 years of individual trees. Sampling site was the strongest predictor of RAF community composition, explaining 19.6% of variation, followed by soil pH (11.7%), whereas habitat explained only 2.1%. Treeline effects were context-dependent: RAF composition differed between forest and treeline only in the Alaska Range, alpha diversity was lower at the treeline in Interior Alaska, and ECM relative abundance was higher at the Brooks Range treeline. In contrast, RAF community composition did not differ between fast- and slow-growing trees within sites. Interestingly, alpha diversity indices were significantly associated with BAI over the previous 5, 10 and 15 years, with lower diversity associated with faster tree growth. These relationships weakened with increasing BAI averaging period. Our results indicate that, in mature P. glauca , tree growth is associated with lower RAF and ECM diversity but not with distinct fungal community composition, suggesting that growth may be linked to greater dominance by a subset of fungal partners rather than higher fungal diversity.

This study evaluates the effectiveness of Illumina-based genome skimming for barcoding myxomycete herbarium collections ranging from 29 to 91 years in age. We successfully retrieved partial sequences of the standard marker gene (nucSSU) in all cases, as well as additional markers (mtSSU, EF1a, and COI) for certain collections. Altogether, 28 genes were recognized in the studied material. In a 33-year-old specimen of Lindbladia tubulina, the assembly reached an N50 of 4.19 kb, enabling the recovery of extended functional loci. The input genomic DNA quantity emerges as the primary determinant of sequencing success. Samples with high DNA yields provide representative amounts of contigs coming confirmedly (matching sequences in the NCBI nucleotide database) or potentially (no-hit fraction) from myxomycetes, regardless of specimen age. In addition to target DNA, we revealed distinct signals of both anthropogenic contamination (human DNA and skin microflora) and natural substrate inhabitants, including oribatid mites and bacteria from dead wood, soil, and grass litter. Thus, even in old collections, metagenomic data still carry information regarding the substrate upon which the myxomycete developed. The results demonstrate that short-read genome skimming may help to integrate historical type material of myxomycetes into contemporary phylogenetic research. This method overcomes the length-dependent limitations of traditional Sanger sequencing, thus providing a roadmap for the future of museomics in myxomycetology.

The revision of collections, held in the Herbarium of the University of Alcal & aacute; (AH) reveals 94 specimens of the myxomycete genus Lycogala, mainly from Spain, but also from Mexico (3 specimens) and France (1). Morphological identification, based on recently developed criteria, allowed identification of 14 Lycogala species, 10 of which are new to Spain, further two are new for Mexico. All species found in Spain were already known from Europe, with the single exception of L. cf. nigroconfusum. The collections of L. paraconfusum and L. guttatum in Mexico represent the first records for the whole of North America. Due to the age of collections, the molecular identification was possible only for nine specimens, thus confirming the identification of three species. To provide reference material for further studies, representative specimens of all taxa were photographed, and the barcoding sequences were submitted to NCBI GenBank.

The 1654 publication by Thomas Pankow is traditionally considered to be the first mention of myxomycetes, and also serves as the source of the first illustration of these organisms. However, it remains largely unknown to the myxomycete community that Pankow was not the author of these illustrations nor even a contemporary of the printmaker who executed them. These in the period 1578-1584. We analysed Pankow's original publication, Herbarium Portatile, which explains the origin of the 1363 engravings that form the core of his book, as well as Thurneysser's Historia sive plantarum omnium. In our opinion, L. Thurneysser should be credited in the history of myxomycete studies as the conditional discoverer of the first myxomycete, and the date of discovery should be shifted back to the 16th century.

The plasmodial slime molds (Amoebozoa, class Myxomycetes) remain one of the few major eukaryotic groups lacking a classification based on robust multigene phylogenies. Extreme sequence divergence and the scarcity of universal primers have thus far impeded broad taxon sampling and the construction of deeply resolved phylogenies. Herein, we propose an updated system for the subclass Columellomycetidae (dark-spored myxomycetes). We employed a genome skimming approach to assemble a 22-gene matrix for 112 herbarium specimens spanning 101 species and integrated public transcriptomic and genomic data from three additional myxomycete species and two dictyostelid species. Different phylogeny inference methods and sequencing data from different loci (nuclear vs. mitochondrial) produced highly consistent and robust clades, most of which we propose as taxa at the level of families. The 22-gene backbone phylogeny was expanded by adding more than nine hundred accessions of dark-spored myxomycetes sequenced for 1–4 genes. This allowed us to produce a highly resolved and species-rich phylogeny of the Columellomycetidae and to revise the delimitation of several genera and families. We now recognize five orders and 12 families in the subclass Columellomycetidae. A new genus, Argentoderma, is described together with the new family Argentodermataceae and the new order Argentodermatales to accommodate three species forming an early-diverging lineage within the Columellomycetidae. The family Didymiaceae is split into four families that better reflect phylogenetic relationships within the Physarales; for this, three new families (Diacheaceae, Polyschismiaceae, and Didermataceae) are described alongside a more narrowly circumscribed Didymiaceae. Echinostelium australiense is transferred to Clastoderma, and Echinosteliopsis is placed within the Echinosteliaceae rather than in a separate order. Semimorula is synonymized with Echinostelium, Physarella is synonymized with Fuligo, Paradiacheopsis is synonymized with Comatricha, Collaria is restricted to the C. rubens clade within the Meridermataceae, and Stemonaria is synonymized with Stemonitis, herein redefined to include the clade centered on Stemonitis fusca. The genera Aethaliopsis, Angioridium, Carcerina, Claustria, and Scyphium are reinstated. In total, 35 new combinations and one nom. nov. (Stemonitis pinicola) are proposed. Extensive homoplasy in sporophore characters underlines the need to explore more fine-scale morphological characters that reflect the evolutionary history of myxomycetes more precisely. Our study demonstrates the utility of shallow genome sequencing of herbarium collections for resolving systematic problems and provides a phylogenomic foundation for comparative research on this neglected lineage of the Amoebozoa.

Drought stress significantly affects plant physiology and growth, yet the molecular mechanisms underlying drought responses remain poorly understood. In this study, we investigate how tetraploid and octoploid Phragmites australis (common reed), a key species in wetland ecosystems and paludiculture, respond to drought at the transcriptional and epigenetic levels. Using RNA-seq, we identify changes in gene expression after 20 and 30 days of drought and assess methylation-sensitive amplification polymorphism (MSAP) over 50 days of drought. Transcriptomic analysis reveals that key drought-response genes are shared between ploidy levels, including those involved in the saccharopine pathway, water deprivation response, cell wall remodelling, and the mevalonate pathway. Drought supresses photosynthesis, with a pronounced down-regulation of the photosynthetic gene PsbP . Ploidy level influences gene expression under both drought and non-stress conditions, highlighting distinct adaptive strategies. In control samples, gene expression differed between ploidy levels, with octoploids up-regulating genes related to translation and metabolism, while tetraploids activate genes involved in cell wall modification and transmembrane transport. Prolonged drought increases DNA methylation variability, though no significant correlation is found between methylation levels and drought duration. Methylation differences are more pronounced between ploidy levels, with octoploids exhibiting lower overall methylation. These findings highlight the complex interactions between gene expression, epigenetic modifications, and polyploidy in drought response and provide a theoretical framework for future selection, hybridization, and conservation initiatives. Main Conclusion Key drought-response genes regulate saccharopine and mevalonate pathways, and cell wall remodelling. Ploidy level influences gene expression under drought and non-stress conditions. Octoploids overall exhibit lower methylation than tetraploids. ### Competing Interest Statement The authors have declared no competing interest.

In June 2017, a systematic survey for nivicolous myxomycetes was carried out in two regions of the Kamchatka Peninsula (Russian Federation), both belonging to the National Park "Vulkany Kamchatki": in southeastern Kamchatka around the Avachinsky volcano and in central Kamchatka, near the village of Esso. Over 90% of all collections were barcoded (18S rDNA, 279 sequences), and a BLAST search revealed many new sequences: only 104 barcodes (37.2%) were 100.0% identical to sequences in GenBank, further 152 were similar to published ones (54.5%, >= 97.9% ID), but 23 barcodes (8.2%, < 97.9% ID) were highly divergent. The ASAP algorithm, which basically provides barcode-based species hypotheses, supported the empirically found similarity threshold of 2.1% dissimilarity: the lowest number of contradictions between morphologically identified and barcode-defined species were found for this barcode gap. However, our initial morphological identification was confirmed by BLAST in only 115 of 256 cases (44.9%), where sufficiently similar sequences were found in GenBank. This highlights the necessity of a DNA barcode-assisted species identification, especially in quantitative surveys involving numerous weathered or maldeveloped colonies. We present a workflow for implementing this approach. The survey covered three vegetation belts: stone birch forests (400-800 m a.s.l.), subalpine shrub thickets of mountain alder and/or dwarf pine (700-1000 m), and mountain tundra (900-1700 m). Thirty morphospecies from 307 collections were recorded. Twenty-eight morphologically identified species were previously unknown for Kamchatka, and one species, Lamproderma zonatopulchellum, is a new record for Russia. Many (13) taxa are classified as rare (a species represents < 0.5% of all records). Only Didymium dubium, Lamproderma echinosporum, L. ovoideum, and Badhamia albescens were frequently occurring in both areas and all three vegetation belts.

The myxomycete Lycogala fuscoviolaceum was described by P. Onsberg in 1972 based on a single specimen collected in Nepal. The status of this species remains ambiguous, as no additional findings have been reported since its description. In this study, we re-examined the holotype stored in Copenhagen to determine the true systematic position of L. fuscoviolaceum. Morphological observations revealed features inconsistent with Lycogala, including the dense, cartilaginous cortex, the pseudocapillitium forming vertical bundles, and the thick-walled spores with elongated pore. These characteristics suggest that L. fuscoviolaceum is either a member of the family Reticulariaceae, possibly related to Reticularia or Siphoptychium, or it is not a myxomycete at all. Cross-sections of the cortical structures of L. fuscoviolaceum revealed the presence of cell-like elements arranged in distinct layers. Such elements were absent in Reticularia splendens, Siphoptychium reticulatum, and S. violaceum, but present in L. flavofuscum. Attempts to obtain molecular data from the holotype of L. fuscoviolaceum, including high-throughput sequencing of short fragments, were unsuccessful due to DNA degradation. New findings are required to clarify the taxonomic position of the species.

Hiddensee, a small island in the Baltic Sea, is characterized by a rather dry, windy, and sunny climate, resembling a periodic desert. We studied epiphytic and fimicolous myxomycetes on the island using the moist chamber method for 101 substrate samples. A total of 37 myxomycete species were identified from 124 records, including 4 species newly recorded in Germany. Molecular barcoding revealed that 67 % of the obtained DNA sequences were new, differing by more than 1 % from their closest matches in the GenBank database. We obtained the first molecular data for Didymium megalosporum (found to be related to the aethaloid species D. spongiosum and D. yulii) and C. elegans var. microspora (new data for both the species and the variety). For Trichia rapa, described in 2023 based on a single barcoded collection, we found three different ribotypes, including one already known. Presumably undescribed taxa within the morphospecies Comatricha nigra, Didymium squamulosum, Enerthenema papillatum, and Trichia contorta were identified by molecular barcoding. Substrate preferences of myxomycetes, categorized into four substrate types (bark of living trees, leaf litter, twigs, and dung), showed distinct patterns of occurrence, with each substrate type associated with a characteristic assemblage of myxomycetes. The species composition on the bark of living trees showed a well-known dependence on bark pH and hardness, with differing pH optima and tolerance ranges among the studied species. Echinostelium minutum occurred across a broad pH spectrum (6.1-8.0; 11 records), whereas Didymium leptotrichum was restricted to a narrow pH range (7.9-8.1; 7 records). Trichia munda preferred relatively acidic substrates (6.4-7.2; 9 records), while Perichaena luteola (7.4-8.0, 5 records) was more commonly found in slightly alkaline conditions.

We report a systematic survey for nivicolous myxomycetes (Amoebozoa, Myxomycetes) carried out between April 30 and May 6 in the French Pyrenees (Hautes-Pyrénées, 900-2000 m). The 738 specimens were barcoded for the nuclear small subunit ribosomal gene (nucSSU, 652, 88.3 % successful). Trichia alpina, the only bright-spored species found, was not sequenced. Additionally, a section of the translation elongation factor 1-alpha gene (EF1A) was successfully sequenced for 496 specimens (67.2 %). The nucSSU phylogeny showed 31 dark-spored species as genetically distinct, yet not always monophyletic lineages. Two species, Polyschismium fallax and P. peyerimhoffii, were grouped in one clade, although differing in barcode sequences. These separations were confirmed by EF1A in all cases except Didymium pseudodecipiens, where EF1A sequences could not be obtained. The resolution for the species pair P. fallax and P. peyerimhoffii increased, and for Polyschismium chailletii two distinct clades were found, indicating a cryptic species complex. Based on the molecular clades, we describe in detail the corresponding morphological differences in four taxa (Didymium dubium and Didymium pseudodecipiens; Polyschismium chailletii groups a and C). The study confirms the reliability of barcoding via nucSSU with an independent second marker and delivers a barcoded, quality-checked comprehensive data set for the Pyrenees.

Throughout evolution, distinct plant lineages independently established mutualistic relationships with various fungal taxa. However, the extent to which these relationships are conserved across different plant and fungal lineages remains unclear. In this study, we compared fungal communities associated with the fine roots of three phylogenetically distant yet cohabiting plant species: Diphasiastrum complanatum, a member of lycophytes, the most basal extant vascular plant lineages; Pinus sylvestris, a gymnosperm; and Vaccinium myrtillus, an angiosperm, an evolutionary relatively young lineage. To minimize environmental variability, fine roots of three species were collected from each of 19 five-square-meter plots within a Scots pine forest in Lithuania. Using metabarcoding and microscopic techniques, we observed significant differences in the fungal community composition and diversity among the three plant species. We detected no signs of arbuscular mycorrhiza in any species. Samples of D. complanatum showed significantly higher taxonomical diversity, while P. sylvestris showed lowest diversity, with ectomycorrhizal fungi being most abundant. Samples of V. myrtillus had a prevalence of putative ericoid mycorrhiza taxa, classes Sebacinales and Trechisporales, likely forming hyphal coils detected through microscopy. In contrast, no mycorrhiza was detected in D. complanatum sporophytes. This, along with the presence of well-developed root hairs and similarity to the fungal community inhabiting soil, suggest a low dependency of D. complanatum sporophytes on mycorrhizal associations and a more opportunistic fungi-plant relationship. This is the first study of fungi associated with the sporophytes of D. complanatum. Our findings provide valuable insights into the complex interactions between fungi and plants from diverse phylogenetic lineages in natural environments.

Climate change leads to more extreme weather events. Therefore, high stress tolerance is becoming increasingly critical for plants. A higher ploidy level has been reported to lead to higher stress tolerance in plants. Phragmites australis is a species well known for its many ploidy levels and is a target species for paludiculture, i.e. the wet use of peatlands. We expected octoploid genotypes of P. australis to outperform tetraploid ones in a 15-month mesocosm experiment including a gradient of 0–100 days of drought. We used pairs of genotypes differing in ploidy from three different geographic regions. Increasing drought length led to a decrease in growth, biomass, morphological and ecophysiological traits in both ploidy levels. A ploidy level of 4x outperformed 8x in almost all traits under constant water supply, leading to more than doubled biomass production, and up to moderate drought (approximately 50 days). Under severe and prolonged drought, both ploidy levels performed equally poorly. This study suggests that lower ploidy levels can outperform higher ploidy levels of P. australis. In this species, ploidy alone may not explain performance, but the genotype can be as or more important than ploidy.

Myxomycetes are a unique branch of life, recognisable by sporophores showing a fungus-like dispersal biology. These structures bear nearly all diagnostic characters for species identification and develop by rapid transformation of plasmodia. During this short period of time, external factors can significantly influence the formation of morphological characters. Therefore, the description of a new species must be carried out with utmost care. Over the last 50 years, approximately 10-15 new species of myxomycetes have been described per year and only some of the latest publications underpin this with molecular data. In this paper, we discuss a set of recommendations for the description of myxomycete species new to science, striving for the following goals: (i) to minimise the number of erroneous descriptions of the species, whose names later have to be put into synonymy; (ii) to make all respective data easily accessible for the scientific community; and (iii) to comply with existing rules of nomenclature. We recommend (1) whenever possible not to describe a new taxon from a single specimen; however, an exception could be made only if supported by molecular data and by unique morphological characters which are unlikely to fall in the range of infraspecific variation of related species; (2) preparing detailed descriptions, including data on developmental stages, microhabitats, ecology, phenology and associated species; (3) providing at least two independent diagnostic characters that tell the new species apart from all others; (4) obtaining a molecular barcode and, whenever possible, providing proof for reproductive isolation of the new species from related taxa; and (5) depositing type specimens in public herbaria. To comply with nomenclatural rules, (6) the new name must be registered in a recognised repository, (7) all published names should be checked for usability before proposing a new name and (8) a unique name should be chosen, preferably highlighting a distinct character of the new species.

Key drought-response genes regulate saccharopine, mevalonate, water-stress pathways, and cell wall remodeling. Ploidy level influences gene expression under drought and non-stress conditions. Octoploids overall exhibit lower methylation than tetraploids. Drought stress significantly affects plant physiology and growth, yet the molecular mechanisms underlying drought responses remain poorly understood. In this study, we investigate how tetraploid and octoploid Phragmites australis (common reed), a key species in wetland ecosystems and paludiculture, respond to drought at the transcriptional and epigenetic levels. Using RNA-seq, we identify changes in gene expression after 20 and 30 days of drought and assess methylation-sensitive amplification polymorphism (MSAP) over 50 days of drought. Transcriptomic analysis reveals that key drought-response genes are shared between ploidy levels, including those involved in the saccharopine pathway, water deprivation response, cell wall remodeling, and the mevalonate pathway. Drought suppresses photosynthetic genes, with PsbP downregulated by up to 32-fold. Ploidy level influences gene expression under both drought and non-stress conditions, highlighting distinct adaptive strategies. In control samples, gene expression differed between ploidy levels, with octoploids upregulating genes related to translation and metabolism, while tetraploids activate genes involved in cell wall modification and transmembrane transport. Prolonged drought increases DNA methylation variability, though no significant correlation was detected between methylation levels and drought duration. Methylation differences are more pronounced between ploidy levels, with octoploids exhibiting lower overall methylation. These findings highlight the complex interactions between gene expression, epigenetic modifications, and polyploidy in drought response and provide a theoretical framework for future selection, hybridization, and conservation initiatives.

We present the first survey of nivicolous myxomycetes (plasmodial slime molds, Amoebozoa) conducted in Kazakhstan, specifically from the Ile-Alatau mountain range near Almaty. A total of 82 specimens were collected, and 16 species were identified using a comparative morphological approach. Except for Didymium dubium, all identified species represent first records for Kazakhstan. DNA barcoding confirmed the morphology-based identification of 70 specimens, revealing 26 distinct barcode sequence variants. Among these, nine 18S rDNA barcode variants were novel and have not been previously reported in the GenBank database.

Climate change leads to more extreme weather events. Therefore, a high stress tolerance is becoming more critical for plants, with higher ploidy being reported to lead to higher stress tolerance. Phragmites australis (P. australis) is a target species for paludiculture, i.e. the wet use of peatlands, and well known for its many ploidy levels. We expected octoploid genotypes of P. australis to outperform tetraploid ones in a 15-month mesocosm experiment including a gradient of 0 to 100 days of drought. We used pairs of genotypes differing in ploidy from three different geographic regions. Increasing drought length led to a decrease in growth, biomass, morphological and ecophysiological traits in both ploidy levels, but 4x outperformed 8x in almost all traits under constant water supply (e.g., 2.5-fold more biomass production) and up to moderate drought (about 50 days). Under severe and prolonged drought, both ploidy levels performed equally poorly. Our study suggests that higher ploidy levels do not necessarily outperform lower ploidy levels of P. australis under stressful conditions. In this species, ploidy alone may not explain performance, but the genotype can be as or more important than ploidy. ### Competing Interest Statement The authors have declared no competing interest.

Provenances show a high phenotypic plasticity and the ability to grow beyond the cold treeline. Local is best can still be applied. Boreal forests situated in high latitudes face heightened susceptibility to climate extremes and global warming. Understanding the relative influence of adaptation mechanisms like phenotypic plasticity or local adaptation on key traits is crucial to better understand and project species distribution, forest growth and vitality. To address this, we conducted a reciprocal transplant experiment featuring two white spruce (Picea glauca [Moench] Voss) provenances in Alaska, representing cold and dry treelines. Trees from each provenance were reciprocally transplanted across a gradient spanning from dry bluff sites, dry treelines via old-growth forests to cold-limited treelines and beyond. From 2015 to 2022, we monitored survival, vitality, growth, and various needle morphology traits. Results showed that the dry provenance had a superior performance in its home environment. Whereas both provenances performed similarly at the cold site. Survival and vitality rates indicated that elevated temperatures favoured tree growth. Seedling survival and growth are possible beyond the current cold treeline. Further, needle morphology traits were more influenced by the current environment than by origin, thus showing a high phenotypic plasticity. Nevertheless, significant differences in needle morphology among provenances hinted at a genetic base of these traits. Results suggested that local is best can still be applied.

The examination of new collections of Alwisia repens and the isotypes of A. repens and A. morula has revealed that these species exhibit a vestigial capillitium in the form of smooth tubules. These tubules are either directed downward or embedded parallel to the peridium; in A. morula they may be long enough to extend from the top to the base of the sporotheca. In both species the upper part of the peridium dehisces into polygonal platelets, which are delimited by preformed fissures. This type of sporotheca opening has not been documented in the Reticulariales before and is known to occur only in certain species within Echinosteliales, Liceales and Trichiales. Newly obtained specimens of A. repens display well-developed, erect stalks. This contradicts the original diagnosis which describes the species as having creeping stalks. We propose emended descriptions for A. repens and A. morula to incorporate these newly discovered characteristics.

The nivicolous species of the genus Diderma are challenging to identify, and there are several competing views on their delimitation. We analyzed 102 accessions of nivicolous Diderma spp. that were sequenced for two or three unlinked genes to determine which of the current taxonomic treatments is better supported by molecular species delimitation methods. The results of a haplotype web analysis, Bayesian species delimitation under a multispecies coalescent model, and phylogenetic analyses on concatenated alignments support a splitting approach that distinguishes six taxa: Diderma alpinum, D. europaeum, D. kamchaticum, D. meyerae, D. microcarpum and D. niveum. The first two approaches also support the separation of Diderma alpinum into two species with allopatric distribution. An extended dataset of 800 specimens (mainly from Europe) that were barcoded with 18S rDNA revealed only barcode variants similar to those in the species characterized by the first data set, and showed an uneven distribution of these species in the Northern Hemisphere: Diderma microcarpum and D. alpinum were the only species found in all seven intensively sampled mountain regions. Partial 18S rDNA sequences serving as DNA barcodes provided clear signatures that allowed for unambiguous identification of the nivicolous Diderma spp., including two putative species in D. alpinum.