Nuclear DNA Regions Research Articles

Molecular and morphoanatomical characterization of Urocystis heteropogonis sp. nov.: a novel smut fungus infecting Heteropogon contortus.

A new species of smut fungus, Urocystis heteropogonis, was discovered infecting Heteropogon contortus in Shawar Valley, Swat district, Khyber Pakhtunkhwa, Pakistan. The study aimed to characterize this fungus based on its morpho-anatomical and molecular features and clarify its phylogenetic position within the genus Urocystis. Urocystis heteropogonis was identified as a novel species, distinct from other Urocystis species. Morphologically, it is characterized by larger spore balls (14-69 × 11-45μm) and central spores that are 14-28 × 11-20μm in size, with each spore containing1-8 central spores. The spore walls measure 0.9-2.5μm in thickness and the species differs in infection patterns compared to other Urocystis species. Phylogenetic analysis based on the ITS and LSU regions of nuclear ribosomal DNA (nrDNA) further confirmed the novelty of the species, placing it within a distinct clade alongside U. agropyri, U. occulta, U. piptatheri, and U. tritici. The discovery of Urocystis heteropogonis adds to the diversity of smut fungi infecting grasses and highlights the need for further research into its ecological and agricultural implications. Future studies should focus on the disease's spread, management, and potential impact on host populations.

Catalogue of fungi in China 2. Ramaria from northern China

ABSTRACT The species of Ramaria are coralloid-fungi with intricately branched and vividly coloured basidiomata. There are many studies on the genus Ramaria in China, but molecular phylogenetic analysis is rarely employed. In this study, we performed a multigene phylogenetic analysis to identify Ramaria species in Shanxi Province of northern China. Phylogenetic analyses based on four loci, the internal transcribed spacer region of nuclear ribosomal DNA (ITS), the nuclear large subunit ribosomal DNA (nrLSU), ATPase subunit 6 (atp6), and mitochondrial small subunit ribosomal DNA (mtSSU), revealed 13 Ramaria species from our collections. Combined with morphological examinations, 12 of them were identified as new species, plus a new report to China. The thirteen species were described and illustrated in this paper.

A new computational method to estimate adaptation time in Avicennia by using divergence time

Evolutionary studies of plant groups which are distributed in vast geographical regions and face different ecological and environmental conditions are important as they through light on different mechanisms of local adaptation and species divergence through time. The genus Avicennia is one of these plant groups which inspire of few species show interesting geographical distribution with some degree of species-specific geographical isolations. In general, very limited molecular phylogenetic investigations have been carried out in the genus Avicennia, and therefore we conducted the present study with the following aims:To estimate the species divergence time based on different nuclear and chloroplast DNA regions, separately. This will illustrate how different genetic regions evolved in this genus,To identify the sequences with potential adaptive value against geographical variable by latent factor mixed models (LFMM) analysis,To illustrate the phylogenetic signal of these DNA regions and their role in speciation within the genus and,To introducing a new computational strategy for estimating adaptive time for the sequences. The results showed that different genetic regions may produce different species divergent time, both the nuclear ribosomal internal transcribed spacer (ITS) region and chloroplast DNA sequences, contained potentially adaptive single nucleotide polymorphisms (SNPs).We could present a suggestive time for these adaptive sequences for the first time. In conclusion both local adaptation and independent mutations seem to have played role in Avicennia speciation and evolution.

Crinipellis siamensis (Marasmiaceae, Agaricales), a new macrofungus from northern Thailand

A new Crinipellis species, described herein as C. siamensis, was collected from northern Thailand during the rainy seasons in 2022 and 2023. This species was identified based on morphological characteristics and molecular data. Crinipellis siamensis is characterized by its crinipelloid basidiomata, grayish orange to brown pileus, elongated to cylindrical basidiospores, and hair-like terminal cells on the pileus surface. Phylogenetic analysis of combined internal transcribed spacer (ITS) and the large subunit (nrLSU) of nuclear ribosomal DNA (rDNA) regions revealed that C. siamensis is well-supported as a distinct monophyletic member within the genus Crinipellis. A full description, color photographs, illustrations, and a phylogenetic tree showing the position of C. siamensis are provided. Morphologically similar and related species to the new species are also discussed.

Wood discrimination of six commonly traded Phoebe and Machilus species using high-resolution plastid and nuclear DNA barcodes

Abstract Phoebe and Machilus trees, whose wood is known as Nanmu in China, face endangerment owing to long-term manmade destruction and climate change. Therefore, effective and accurate forensic tools to identify Nanmu species are required. This study developed a method for DNA identification of woods from six commonly traded Phoebe and Machilus species. DNA was efficiently extracted from oil cell-rich wood tissues, DNA barcodes were screened, species were compared, and a Nanmu DNA database was constructed. Comparison and analysis involved 26 sets of plastid genomes of Phoebe and Machilus. The five highest variable regions (Pi>0.009; psbA-trnH, psbJ, psbJ-petA, rpl32-trnL, and ycf 1) were identified, and three nuclear regions (ITS, LEAFY, and RPB2) were compared. Among single DNA fragments, the nuclear DNA region LEAFY proved optimal for identifying Nanmu species, while psbA-trnH+psbJ+ycf1 was the optimal plastid combination locus. Compared with distance-based (TaxonDNA) and machine-learning analyses, the tree-based method had a lower success rate for species identification. Four labelled Nanmu samples were selected to validate the method and clustered with P. zhennan and P. sheareri data based on the LEAFY region and plastid combination locus. These findings will contribute to the conservation, classification, architecture maintenance, and trade monitoring of Nanmu natural resources.

Out of and in East Asia: Phylogeny, biogeography and diversification of Thalictroideae (Ranunculaceae) in the Northern Hemisphere.

Understanding the biogeographical patterns and processes underlying the distribution of diversity within the Northern Hemisphere has fascinated botanists and biogeographers for over a century. However, as a well-known centre of species diversity in the Northern Hemisphere, whether East Asia acted as a source and/or a sink of plant diversity of the Northern Hemisphere remains unclear. Here, we used Thalictroideae, a subfamily widely distributed in the Northern Hemisphere with the majority of species in East Asia, to investigate the role of East Asia in shaping the biogeographical patterns of the Northern Hemisphere and to test whether East Asia acted as a museum or a cradle for herbaceous taxa. Based on six plastid and one nuclear DNA regions, we generated the most comprehensive phylogeny for Thalictroideae including 217 taxa (ca. 66% species) from all ten of the currently recognized genera. Within this phylogenetic framework, we then estimated divergence times, ancestral ranges, and diversification rates. The monophyletic Thalictroideae contains three major clades. All genera with more than one species are strongly supported as monophyletic except for Isopyrum, which is nested in Enemion. The most recent common ancestor of Thalictroideae occurred in East Asia in the late Eocene (ca. 36 Ma). From the Miocene onwards, at least 46 dispersal events were inferred to be responsible for the current distribution of this subfamily. East Asian Thalictroideae lineages experienced a rapid accumulation at ca. 10 Ma. The biogeographical patterns of Thalictroideae support the "out of and in East Asia" hypothesis, i.e., East Asia is both a source and a sink of biodiversity of the Northern Hemisphere. The global cooling after the middle Miocene Climatic Optimum, combined with the exposed land bridges due to sea-level decline, might have jointly caused the bidirectional plant exchanges between East Asia and other Northern Hemisphere regions. East Asia serves as evolutionary museums and cradles for the diversity of Thalictroideae and likely for other herbaceous lineages.

First report of wheat leaf spot disease caused by Curvularia inaequalis in China.

Wheat (Triticum aestivum) is an economically important crop widely cultivated in China. In August 2022, brown oval leaf spots with yellow halos were observed on approximately 10% wheat seedlings over an area of about 1 hectare in Xining City, Qinghai Province, which adversely affected wheat growth and production. Six diseased leaves were collected from the field in Huangyuan county (101°69' E, 37°04' N). The 0.5 cm × 0.5 cm pieces were cut from the border between healthy and diseased regions of the sampled leaves, surface sterilized for 10 s in 75% ethanol, followed by a 1% NaClO for 90 s, and rinsed three times with distilled sterile water. The pieces of leaf tissue were dried with sterile tissue, and plated on potato dextrose agar (PDA) amended with streptomycin (0.02 g/L) and ampicillin sulfate (0.05 g/L) to eliminate bacterial contamination. The dishes were placed in an incubator at 25°C for 72 h in dark. Three isolates, WGC201, WGC202 and WGC203, were obtained by a single-spore culture method. Fungal colonies on PDA media were dark green (Fig. 1A and 1B). Conidiophores were septate and geniculate terminals, while conidia exhibited straight or slightly curved forms with four transverse septa, the central cell being notably longer and wider than the others. The size of such conidia were 27.34 µm to 40.62 µm× 11.61 µm to 15.97 µm (number = 50) (av. 32.71 µm× 13.11 µm) (Fig. 1C and 1D) (Moubasher et al. 2010). The internal transcribed spacer (ITS) region of nuclear ribosomal DNA and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene were amplified and sequenced using universal primers ITS1/ITS4 and GPDF/GPDR (White et al. 1990; Berbee et al. 1999). DNA sequences were deposited into the NCBI database (ITS, PP789629, PP801333, and PP801574; GAPDH, PP849124, PP849125, and PP849126). Phylogenetic analysis with a neighbor-joining method based on the concatenated sequences of ITS and GAPDH genes showed that the three isolates clustered within a C. inaequalis branch (Fig. 2). Based on morphological and molecular identification, the fungal isolates were identified as C. inaequalis. The pathogenicity test was conducted in a greenhouse at 25°C using a spore suspension method and three isolates were used. Conidia were produced on PDA media (25℃) for 14 days. Plates were washed with sterilized distilled water and filtered with cheese cloth. Conidial suspension was adjusted to a concentration of 1×107 conidia/mL. Fifteen healthy seedlings of a wheat cultivar Xiaoyan-6 at a 3-4 leaf stage were inoculated by evenly spraying a 100mL spore suspension. Plants inoculated with sterile water served as a control. All plants were covered with plastic bags for 3 days. At 7 days after inoculation, all pathogen-inoculated plants showed similar symptoms (brown leaf oval spots with yellow halos) with those observed in the field, while all plants inoculated with sterile water showed no symptoms (Fig. 1E and 1F). The pathogen was reisolated from the symptomatic leaves and proved to be C. inaequalis. Morphological, molecular and pathogenic results indicated that C. inaequalis is the pathogen causing wheat leaf disease in China. The results are consistent with a previous report in Azerbaijan (Özer et al. 2020). To our knowledge, this is the first report of C. inaequalis causing spot disease on wheat in China. The occurrence, spread and economic importance to different wheat cultivars of the emerging disease in China will be further investigated and evaluated.

Molecular evidence for the origin of the little-known hybrid Potamogeton ×vilnensis

A combined study of morphology, stem anatomy, and the genetic markers (ITS1-ITS4 region of nuclear ribosomal DNA and rpl20-rps12 intergenic spacer of chloroplast DNA) was used to verify the identity of hybrid Potamogeton ×vilnensis Galinis previously recognized as P. gramineus × P. praelongus. Authentic herbarium material and newly collected specimens morphologically similar to the hybrid from the locus classicus (SE Lithuania) were used for comparative morphological, anatomical and molecular analyses. The study proved the paternity of P. lucens and P. praelongus and excluded the involvement of P. gramineus in hybridization and identity with P. ×angustifolius. This hybrid formula is shared with plants recorded in the Czech Republic and Denmark under the name P. ×jutlandicus Zalewska-Galosz. The site in Lithuania, known since 1961, is the first natural habitat of long-lived hybrid between P. lucens and P. praelongus whose origin has been confirmed by molecular markers. The scarce records of this taxon indicate that the knowledge of the hybrid species of Potamogeton and their distribution is insufficient and that more extensive regional studies, including molecular methods, are needed.

Fungal-plant interaction: a pathogenic relationship between Ganoderma segmentatum sp. nov. and Vachellia nilotica.

The diversity of Ganoderma remains largely unexplored, with little information available due to fungiphobia and the morphological plasticity of the genus. To address this gap, an ongoing study aims to collect and identify species with this genus using nuclear ribosomal DNA regions called the "Internal Transcribed Spacer" (ITS1-5.8S-ITS2 = ITS). In this study, a new species, Ganoderma segmentatum sp. nov., was discovered on the dead tree trunk of the medicinal plant, Vachellia nilotica. The species was identified through a combination of morpho-anatomical characteristics and phylogenetic analyses. This new species was closely related to Ganoderma multipileum, G. mizoramense, and G. steyaertanum, with a 99% bootstrap value, forming a distinct branch in the phylogenetic tree. Morphologically, G. segmentatum can be distinguished by its frill-like appearance on the margin of basidiome. Wilt or basal stem rot, a serious disease of trees caused by Ganoderma species and V. nilotica, is brutally affected by this disease, resulting in substantial losses in health and productivity. This Ganoderma species severely damages V. nilotica through deep mycelial penetration in the upper and basal stems of the host species. Pathogenic observational descriptions of G. segmentatum on dead tree trunks showed the exudation of viscous reddish-brown fluid from the basal stem portion, which gradually extended upward. Symptoms of this disease include decay, stem discoloration, leaf drooping, and eventual death, which severely damaged the medicinal tree of V. nilotica.

Taxonomic Re-Evaluation of the Genus Fuscoporia in the Republic of Korea Including Three Unrecorded Species

The genus Fuscoporia (Hymenochaetaceae, Basidiomycota) comprises poroid white-rot fungi characterized by dark brown hymenial setae, a dimitic hyphal system, and encrusted generative hyphae. Despite the ecological and commercial significance of Fuscoporia species, their identification has been challenging owing to their morphological overlap with other genera of Hymenochaetaceae and to the limited resolution of nuclear ribosomal DNA markers. With the advances in molecular research, Fuscoporia has been revised to include species from Inonotus sensu lato and Phellinus sensu lato, and 71 new species have been reported over the past decade. In Korea, a comprehensive taxonomic study elucidating the true diversity of Fuscoporia is yet to be conducted. Among the 11 Fuscoporia species reported in Korea, two were identified solely based on morphological characteristics, and four were identified based on nuclear ribosomal DNA regions, which have limited resolution for species identification in Fuscoporia. To investigate the current status of Fuscoporia species native to Korea, we conducted a phylogenetic study using four genetic markers (ITS + nrLSU + RPB2 + TEF1), along with morphological characteristics, and re-analyzed the GenBank records deposited from Korea. Ten Fuscoporia species were identified, including three previously unrecorded species. A detailed description of the unrecorded species and a list of proposed Korean names for all Fuscoporia species in Korea are provided. This study will guide further taxonomic and applied research of Fuscoporia by providing a species identification key and a verified multigenetic database, in addition to confirming the sequences in public database.

Poaceascoma zborayi sp. nov. and Agrorhizomyces patris gen. et spec. nov. – Two novel dark septate endophytes colonizing wheat (Triticum aestivum) roots from a cropland in Hungary

In this study, we investigated two distinct new phylogenetic lineages of root-colonizing dark septate endophytic fungi colonizing wheat (Triticum aestivum) roots from a long-term agricultural experimental site in Hungary. According to four-locus (internal transcribed spacer, partial large and small subunit regions of nuclear ribosomal DNA, and partial translation elongation factor 1-alpha) phylogenetic analyses, the isolates belong to the Lentitheciaceae and Didymosphaeriaceae of the Pleosporales (Dothideomycetes). We studied the morphology and culture characteristics of the strains. We carried out in vitro resynthesis pot experiments with their original hosts and found no overall negative effect of the inoculation with different isolates of the new taxa. One of the lineages belonged to the genus Poaceascoma (Lentitheciaceae) and represented a novel species described here as Poaceascoma zborayi. We could describe conidia-like structures from this species. Isolates of the other lineage represented a monotypic novel genus in the Didymosphaeriaceae. Accordingly, the new genus, Agrorhizomyces, represented by the species A. patris, is introduced. Sterile, globose structures resembling immature sporocarps were detected. Sequence similarity searches indicated that P. zborayi might be widely distributed, while no sequence similar to A. patris was found outside the sampling area.

Hysterium madraspatanum (Hysteriaceae), a new species from Tropical Dry Evergreen Forest of Tamil Nadu, India

A new species of Hysterium, is described from tropical dry evergreen forest region of Chennai, Tamil Nadu, India, based on both morphological characteristics and molecular phylogenetic analysis using the Internal Transcribed Spacers (ITS) region of nuclear ribosomal DNA. The key morphological characters of the ascomata include the presence of a longitudinal slit remaining closed even at maturity. The ascospores biseriate having three septa with all the cells evenly pigmented including the terminal cells. The ascus measuring 115–122 × 20–22 μm. Ascospores initially two celled and finally become four celled, hyaline first, later changing to yellow and then dark brown, oblong to ellipsoid, 28–39 × 7–12 μm. The nrITS-based phylogenetic tree inferred from the Bayesian posterior probabilities indicate that H. madraspatanum is closely related to Hysterium pulicare, H. angustatum and H. rhizophorae. However, H. madraspatanum is morphologically distinct from them by having relatively larger spore size, shallow groove in the ascomata, spores with concolorous terminal cells. Therefore, is a novel species under the genus Hysterium. Azadirachta indica is reported in the study as a new host for the genus Hysterium.

Myxotrichum persicum sp. nov., and two new combinations in the genus Myxotrichum

We describe Myxotrichum persicum sp. nov. as a seed endophyte of Aegilops triuncialis in Lorestan province of Iran, using morphological traits and sequences of the internal transcribed spacer regions 1 and 2 including the intervening 5.8S nuclear ribosomal DNA (ITS) and partial nuclear 28S ribosomal DNA (LSU) regions. Myxotrichum persicum is differentiated from its closest relative, M. deflexum, by possessing smaller ascomata bearing much shorter, usually unbranched appendages, and smaller, ellipsoidal to fusiform, smooth-walled ascospores. Based on morphology and the multilocus phylogeny of ITS, LSU, MCM7 and RPB1, we herein propose to combine two species of Malbranchea, namely M. circinata and M. flavorosea, into the genus Myxotrichum as Myxotrichum circinatum and M. flavoroseum, respectively.

Prevalence and molecular characterization of Hysterothylacium species infecting Pandora (Pagellus erythrinus) in the Mediterranean Sea of Egypt

Species of the genus Hysterothylacium are aquatic roundworms (nematodes) belonging to the family Raphidascarididae. Some species in this family are known to be associated with zoonotic diseases in humans after they consume their parasitic larvae in raw or undercooked fish. The aim of this research was to report the prevalence, morphology, and molecular characteristics of Hysterothylacium species in Pagellus erythrinus. A total of Two hundred fish were purchased from the fish market in Damanhour, Beheira Province, between December 2021 and November 2022 and subjected to examination. For molecular characterization, the internal transcribed spacer (ITS) region of nuclear ribosomal DNA and the mitochondrial cytochrome oxidase subunit 2 (COX-2) gene were used. Hysterothylacium species were morphologically described and identified from the intestine of Pagellus erythrinus in Beheira Province, Egypt. The PCR amplified 1087 bp and 629 bp of the target sequences of the ITS region and COX-2 gene, respectively. Sequence analysis revealed the Hysterothylacium thalassini species. The identified species provided novel biological data for the Hysterothylacium nematode in Pagellus erythrinus. The prevalence of Hysterothylacium species recovered from the intestine was 55%. The highest prevalence of 72% has been reported in summer compared to the lowest prevalence of 38% in the winter. Females had a higher prevalence of 61.8% than males, with 44.2%. The first detection, prevalence, and molecular characterization of H. thalassini in Pagellus erythrinus from Beheira Province, Egypt, was presented in this study.

Carexqingyuanensis (Cyperaceae), a new species from Guangdong, China.

Carexqingyuanensis, a new species of Cyperaceae from Guangdong Province, China, is described and illustrated. The new species is morphologically similar to Carexpeliosanthifolia F. T. Wang & Tang ex P. C. Li, but it can be distinguished by the racemose inflorescence branches appearing single (rarely binate or ternate) (vs. binate or ternate), one (rarely two or three) (vs. 1-3) spiked, male part of linear-cylindrical spikes much longer than the female part (vs. just male part short-cylindrical and slightly longer than female part), style base thickened (vs. not thickened) and perigynium horizontally patent with a short (vs. long and excurved) beak. Phylogenetic analysis, based on the two nuclear DNA regions (ETS 1f and ITS) and three chloroplast DNA regions (matK, ndhF and rps16), suggests that the new species belongs to sect. Siderostictaes.s. of subg. Siderosticta and shows a closer phylogenetic relationship to Carexscaposa C. B. Clarke.

Phylogenetics, character evolution, and historical biogeography of the Neotropical genus Besleria (Gesneriaceae)

Abstract Besleria, a genus of perennial herbs, shrubs, or small trees growing in the understorey of rainforests, is one of the largest genera of neotropical Gesneriaceae, with over 165 species. Despite the ecological importance and ubiquity of Besleria in rainforest ecosystems, taxonomic and evolutionary studies of Besleria are limited. Here, we generated a phylogenetic analysis of Besleria using four nuclear and chloroplast DNA regions (ITS, matK, rps16, and trnL-trnF) covering more than 50% of the recognized species, along with two secondary calibration points to infer divergence times. Our results support the monophyly of Besleria and allowed us to revise the infrageneric classification and biogeographical history of the genus. We identified five major clades that do not correspond to sections or subsections in previous classifications. These clades are well circumscribed geographically but remain difficult to characterize using previously hypothesized morphological characters. Biogeographical reconstructions indicate an origin in the northern Andes during the Middle Miocene (ca. 15 Mya). The current distribution patterns of this plant group have been significantly shaped by geological and climatic events, particularly Andean uplift and the formation of the Panama Isthmus.

Phylogenetics of Cruckshanksia and Oreopolus (Rubiaceae) based on nuclear and plastid DNA sequences

Cruckshanksia Hook. & Arn. and Oreopolus Schltdl. Rubiaceae (Rubioideae – Coussareeae), endemic genera of Chile and Andean Argentina, have historically been highly taxonomically unstable. Molecular analyses have confirmed Oreopolus as the sister group of Cruckshanksia; however, the relationships among its species are still not resolved because previous studies have not considered all species of the genus and the published topologies have many unsupported nodes. For this reason, we carried out a phylogenetic study with all the species currently recognized by recent revisions, using two nuclear DNA regions and five plastid regions, to elucidate the relationships between species. In addition, the evolutionary history of the group was estimated based on divergence times, and a character reconstruction was performed. The results corroborate that Oreopolus is the sister group of Cruckshanksia. Cruckshanksia is monophyletic and composed of two principal clades. Clade I is composed of Subclade I (C. pumila and C. verticillata) associated with C. montiana; Clade II is composed of Subclade II (C. palmae and C. macrantha) and Subclade III (C. lithiophila and C. hymenodon). With 81% probability the common ancestor of the genus Cruckshanksia had petaloid appendices associated with two independent lines of evolution. The probability that the common ancestor of the Oreopolus‒Cruckshanksia clade had equal calyx lobes is 50%.

Diversity of fungi and dynamics of ethanol concentration during fermentation of porang (Amorphophallus oncophyllus)

Abstract. Helmi H, Kusmiadi R, Mahardika RG, Karsiningsih E. 2024. Diversity of fungi and dynamics of ethanol concentration during fermentation of porang (Amorphophallus oncophyllus). Biodiversitas 25: 542-552. Fermentation can be an alternative to process porang (Amorphophallus oncophyllus). As one of the fermentation products, ethanol can degrade starch and consequently increase the purity of glucomannan in porang flour. However, ethanol can also inhibit the growth of microorganisms that play a role in fermentation. This study aimed to determine the ethanol concentration and investigate the fungal diversity during the fermentation of porang tubers. Ethanol analysis was analyzed by Headspace Gas Chromatography coupled with flame ionization detection, and Starch content was analyzed by colorimetric method. Fungal metagenome was amplified at the Internal transcribed spacer (ITS) region of nuclear DNA (rDNA) and followed by sequencing using Oxford Nanopore Technology (ONT). The result showed that the ethanol concentration increased during 48 hours of fermentation, followed by decreased starch content. The highest fungal diversity index occurred at 48 hours of fermentation. Ethanol and fungal enzymatic activity can degrade the starch. During fermentation, the Candida parapsilosis population increased until 24 hours after fermentation. Cyberlindnera subsufficiens population increased at 48 hours fermentation until 72 hours fermentation. Fungal diversity and ethanol concentration had decreased at 72 hours of fermentation, so porang fermentation should be stopped at 48 hours. Candida parapsilosis could be the candidate of starter for fermentation porang due to the capability of this species to produce ethanol to degrade starch.

DNA barcoding of some endangered plant species of the Altai Mountains based on five genetic markers

The analysis of DNA nucleotide sequences is an important source of information in the identification of plant taxa or plant components in the composition of medicines. In this work, we studied the possibility of using universal molecular markers for DNA barcoding of rare plant species distributed in the Altai Biosphere Reserve, located in the Altai Republic (South Siberia, Russia). In order to identify commercially-used endangered species, universal markers recommended for DNA barcoding were applied: ITS (Internal Transcribed Spacers) region of nuclear ribosomal DNA and four regions of chloroplast DNA: trnL-F, trnH-psbA rpl32-trnL, and rpl16. Among 61 species of rare flowering plants protected in the Altai Biosphere Reserve, ten species belonging to eight families were studied. The results of sequencing showed that ITS, rpl32-trnL and trnH-psbA core plant barcodes are the most effective markers for discriminating rare species from the Altai Biosphere Reserve.

Glacial history of Saxifragawahlenbergii (Saxifragaceae) in the context of refugial areas in the Western Carpathians.

Despite the wealth of data available for mountain phylogeography, local-scale studies focused on narrow endemic species remain rare. Yet, knowledge of the genetic structure of such species biogeographically linked to a restricted area is of particular importance to understand the history of the local flora and its diversity patterns. Here, we aim to contribute to the phylogeographical overview of the Western Carpathians with a genetic study of Saxifragawahlenbergii, one of the most characteristic endemic species of this region. We sampled populations from all discrete parts of the species' distribution range to apply sequencing of selected non-coding cpDNA and nuclear ribosomal DNA (ITS) regions, as well as Amplified Fragment Length Polymorphism (AFLP) fingerprinting. First, while ITS sequences showed weak diversification, the genetic structure based on cpDNA sequences revealed two well-differentiated groups of haplotypes. One of them is restricted to the main center of the distribution range in the Tatra Mountains (Mts), while the second group included a series of closely related haplotypes, which in most cases were unique for particular isolated groups of populations in peripheral mountain ranges and in the south-eastern part of the Tatra Mts. AFLP fingerprinting also revealed a pattern of divergence among populations, while only partly corroborating the division observed in cpDNA. Taking into account all the data, the pattern of genetic structure, supported by the high levels of unique genetic markers in populations, may reflect the historical isolation of populations in several local refugia during the last glacial period. Not only the center of the range in the Tatra Mts, but also other, neighboring massifs (Malá Fatra, Nízke Tatry, Chočské vrchy, Muránska planina), where populations are characterized by separate plastid DNA haplotypes, could have acted as separate refugia.