nrDNA ITS Sequences Research Articles

Molecular phylogenetic studies on Trigonella section Ellipticae (Trifolieae, Fabaceae) from India utilising nrDNA (ITS) and cpDNA (trnL–trnF & petL–psbE)

This study highlights the critical importance of species delimitation in biodiversity conservation, focusing on the poorly understood Indian Trigonella species, with focus on T. sect. Ellipticae. Despite their ecological and socio-economic significance, these species, primarily found in the northwest Himalayas, have been inadequately catalogued and preserved. Adapted to harsh, dry environments, T. sect. Ellipticae exhibits morphological traits that complicate traditional species delimitation due to significant overlap among species boundaries. To address these challenges, we integrated molecular sequence data (nrDNA ITS and cpDNA (trnL–trnF & petL–psbE)) with morphological analysis, confirming the monophyly of Trigonella and revealing distinct evolutionary lineages within T. sect. Ellipticae. While nrDNA ITS sequences alone proved insufficient for resolving species relationships, phylogenetic investigations utilizing plastid data identified three distinct clades, enhancing taxonomic clarity. The study also suggests that observed morphological variations in T. cachemiriana may be due to phenotypic plasticity rather than distinct evolutionary lineages. We propose a new combination, T. emodi var. gracilis, to better represent the diversity within this group. Overall, this research underscores the necessity of combining molecular and morphological approaches for effective species delimitation, contributing to broader biodiversity conservation efforts in India, where many species remain inadequately catalogued. As the urgency for conservation increases, refining taxonomic frameworks is essential for ensuring that conservation strategies are based on accurate biological data.

A new taxonomic treatment and comprehensive evaluation of the genus Aegokeras (Apiaceae) endemic to Turkey

The systematic position of the species Pimpinella ibradiensis, a narrow endemic to Turkey, was evaluated on the basis of morphological data and nrDNA ITS sequences. The species Pimpinella ibradiensis has been classified within the genus Pimpinella, particularly due to the similarity of leaf characteristics. However, the presence of trifoliolate leaves has been observed in both the isotype specimens and newly collected specimens from the field. Field observations, detailed morphological comparison, mericarp anatomy, and nrITS sequences data have supported the inclusion of this species within the genus Aegokeras. Furthermore, the description of the species P. ibradiensis has been emended, and the diagnostic morphological characters of all Aegokeras species are discussed. Additionally, the geographical distribution of the genus Aegokeras species is mapped, and an identification key with a description of all species is provided.

Glycine tabacina, native to Australia, is an alternate host of Erysiphe diffusa causing powdery mildew on soybean

AbstractPowdery mildew, caused by Erysiphe diffusa, is an emerging pathogen in Australian soybean crops. Since its initial detection in 2012, the disease has been observed every season in soybean paddocks throughout Australia. It is not known how E. diffusa survives between soybean cropping seasons in the Australian environment. This study identified the native Glycine species, G. tabacina, as an alternate host for E. diffusa in Australia. G. tabacina specimens naturally infected with powdery mildew were collected and the pathogen was identified based on morphological characters and nrDNA ITS and MCM7 sequences. Cross‐inoculation experiments demonstrated that the E. diffusa isolates infecting G. tabacina in the field were pathogenic to soybean. This study is the first to report E. diffusa on G. tabacina in Australia. As a perennial native often found in the vicinity of the annual soybean crops, G. tabacina can easily serve as an alternate host for E. diffusa and could be an example of a host range expansion in this powdery mildew species. Weed control in soybean crops, with special attention to the removal of the native Glycine species, may be an option for powdery mildew management for Australian soybean growers.

MORPHOLOGICAL AND NRDNA-ITS SEQUENCES-BASED IDENTIFICATION AND DISTRIBUTION OF VOLVOPLUTEUS EARLEI IN PAKISTAN

Volvopluteus is represented by seven species distributed in America, Asia and Europe. They occur in the tropical regions growing as saprobes on wood debris and leaves, in grassy fields and forests. To explore the macrofungal diversity, different localities in Punjab were visited and collections were made. Among these several collections were found similar to Volvopluteus species. They were analyzed morphologically and phylogenetically using ITS sequences of nrDNA. Comparative analysis on the basis of morphological and anatomical features revealed that these collections represent V. earlei . Maximum likelihood phylogeny also showed the position of current sequences in a same clade with already known V. earlei sequences with 74% bootstrap value. Volvopluteus earlei is being described from different areas of Pakistan with variety of climatic conditions from warm semi-arid climate of Kasur, Lahore and Sheikhupura districts, humid subtropical climate of Sialkot district, in Punjab and warm semi-arid to humid subtropical climate of Swat district, Khyber Pakhtunkhwa. The occurrence of V. earlei in different climatic zones represents its wide distribution range and ecological amplitude in the country.

Transferring Peucedanum macilentum (Apioideae, Apiaceae) to Ligusticopsis based on morphological and molecular evidences

Peucedanum macilentum is a less-studied species endemic to China, previously assigned to the genus Kitagawia. To elucidate its taxonomic affinity, morphological and molecular analyses were conducted. Morphologically, Peucedanum macilentum resembles Ligusticopsis species in several diagnostic characters of the stem base clothed in fibrous sheaths, pinnate bracteoles, conspicuous calyx teeth, strongly dorsally compressed fruits, as well as more vittae on the commissure. Phylogenetic analyses of nrDNA ITS and plastome sequences further reveal that P. macilentum is distant from Kitagawia but nested within Ligusticopsis. Therefore, in the basis of the morphological and molecular evidences, Peucedanum macilentum is transferred into Ligusticopsis and a new combination, Ligusticopsis macilenta, is proposed.

Complete Plastomes of Ten Rorippa Species (Brassicaceae): Comparative Analysis and Phylogenetic Relationships

The genus Rorippa belongs to the family Brassicaceae, and its members usually have high medicinal value. The genus consists of approximately 75 species and mainly grows in the Northern Hemisphere, occurring in every continent except Antarctica. The taxonomy and phylogenetic relationships of Rorippa are still unsettled, largely due to complex morphological variations in Rorippa, which were caused by frequent hybridization events. Here, we sequenced four complete plastid genomes of Rorippa species by Illumina paired-end sequencing. The four new plastid genomes of Rorippa ranged in total size from 154,671 bp for R. palustris to 154,894 bp for R. sylvestris. There are 130 genes in the four plastomes, embodying 8 rRNA, 37 tRNA, and 85 protein-coding genes. Combining with six published plastid genomes, we carried on comparative and phylogenetic analyses. We found that the ten Rorippa plastid genomes were conservative in gene number and order, total size, genomic structure, codon usage, long repeat sequence, and SSR. Fourteen mutational hotspot regions could be selected as candidate DNA barcoding to distinguish Rorippa plants. The phylogenetic trees clearly identified that ten Rorippa species displayed monophyletic relationships within the tribe Cardamineae based on plastomes and nrDNA ITS sequences. However, there are significant cytonuclear discordances in the interspecific relationships within Rorippa, as well as the intergeneric relationships between Rorippa and its related genera. We inferred that the cytonuclear discordance is most likely a result of interspecific hybridization within Rorippa, as well as intergeneric hybridization with its related genera. These plastid genomes can offer precious information for studies of species authentication, evolutionary history, and the phylogeny of Rorippa.

Molecular phylogeny and spatio‐temporal diversification of the Millettioid/Phaseoloid clade (Fabaceae: Papilionoideae)

AbstractThe Millettioid/Phaseoloid (or the Millettioid) clade is a major lineage of the subfamily Papilionoideae (Fabaceae) that is poorly understood in terms of its diversification and biogeographic history. To fill this gap, we generated a time‐calibrated phylogeny for 749 species representing c. 80% of the genera of this clade using nrDNA ITS, plastid matK, and plastome sequence (including 38 newly sequenced plastomes). Using this phylogenetic framework, we explored the clade's temporal diversification and reconstructed its ancestral areas and dispersal events. Our phylogenetic analyses support the monophyly of the Millettioid/Phaseoloid clade and four of its tribal lineages (Abreae, Desmodieae, Indigofereae, and Psoraleeae), while two tribal lineages sensu lato millettioids and phaseoloids are polyphyletic. The fossil‐calibrated dating analysis showed a nearly simultaneous divergence between the stem node (c. 62 Ma) and the crown node (c. 61 Ma) of the Millettioid/Phaseoloid clade in the Paleocene. The biogeographic analysis suggested that the clade originated in Africa and dispersed to Asia, Europe, Australia, and the Americas at different periods in the Cenozoic. We found evidence for shifts in diversification rates across the phylogeny of the Millettioid/Phaseoloid clade throughout the Cenozoic, with a rapid increase in net diversification rates since c. 10 Ma. Possible explanations for the present‐day species richness and distribution of the Millettioid/Phaseoloid clade include boreotropical migration, frequent intra‐ and intercontinental long‐distance dispersals throughout the Cenozoic, and elevated speciation rates following the Mid‐Miocene Climatic Optimum. Together, these results provide novel insights into major diversification patterns of the Millettioid/Phaseoloid clade, setting the stage for future evolutionary research on this important legume clade.

Genetic diversity and morphological variability of Iranian Silene aucheriana populations inferred from nrDNA ITS sequences and morphological analysis

Abstract We conducted a comprehensive study on the diversity of Silene aucheriana Boiss. populations by analyzing both morphological data and nrDNA ITS sequences. Maximum parsimony and Bayesian methods on representative material from 15 populations throughout Iran demonstrated that they did not form a monophyletic group. Strong positive correlation occurred between epipetalous filaments length to claw (EFLC), basal leaf length (BLL), basal leaf width (BLW), and plant height (PLH) with elevation. While negative correlation achieved between anthophore length (AnL) with the average air temperature and rainfall. Canonical Correspondence Analysis (CCA) showed that most morphological traits were more correlated to elevation than to average rainfall and temperature. Multivariate analysis of morphological traits with ITS analysis displayed a slight divergence between two types of regions based on their elevation.

Effect of Different Edible Trichosanthes Germplasm on Its Seed Oil to Enhance Antioxidant and Anti-Aging Activity in Caenorhabditis elegans

The seeds of various Trichosanthes L. plants have been frequently used as snacks instead of for traditional medicinal purposes in China. However, there is still a need to identify the species based on seeds from Trichosanthes germplasm for the potential biological activities of their seed oil. In this study, 18 edible Trichosanthes germplasm from three species were identified and distinguished at a species level using a combination of seed morphological and microscopic characteristics and nrDNA-ITS sequences. Seed oil from the edible Trichosanthes germplasm significantly enhanced oxidative stress tolerance, extended lifespan, delayed aging, and improved healthspan in Caenorhabditis elegans. The antioxidant activity of the seed oil exhibits a significant positive correlation with its total unsaturated fatty acid content among the 18 edible Trichosanthes germplasm, suggesting a genetic basis for this trait. The biological activities of seed oil varied among species, with T. kirilowii Maxim. and T. rosthornii Harms showing stronger effects than T. laceribractea Hayata.

Tragopogon davutii‐turanii: a new species of Tragopogon from Anatolia

A new salsify species, Tragopogon davutii‐turanii, is described from Anatolia based on micro‐/macro‐morphological and molecular evidence. The new species is morphologically similar to T. vvedenskyi and T. abbreviatus, but differs from T. vvedenskyi mainly in ligule colour and from T. abbreviatus by achene characters and number of phyllaries. In contrast to morphological similarities, based on the phylogenetic tree obtained from nrDNA ITS sequences T. davutii‐turanii is closely related to T. stenophyllus and T. coelesyriacus. Diagnostic characters, description, and conservation status of the new species, as well as micro‐ and macro‐morphological features of its achenes and pollen grains, are provided.

A revised genus-level classification for Cerrenaceae (Polyporales, Agaricomycetes)-SUPP

Supplementary files:<br />Fig. S1. Bayesian consensus tree of Cerrenaceae based on nrDNA ITS and LSU sequences. Countries (and states) are denoted with ISO 3166 codes.<br />Fig. S2. Bayesian consensus tree of the Cerrenaceae based on mitochondrial SSU sequences.<br />Fig. S3. Bayesian consensus tree of the Cerrenaceae based on mitochondrial ATP6 sequences.<br />Fig. S4. Bayesian consensus tree of the Cerrenaceae based on nuclear RPB1 sequences.<br />Fig. S5. Bayesian consensus tree of the Cerrenaceae based on nuclear TEF1 sequences.

Phylogeny and taxonomic review of Oreocomopsis with description of a new genus Oreocomoides (Apioideae, Apiaceae)

A revision of the genus Oreocomopsis was carried out based on the study of morphological, anatomical and molecular phylogenetic data based on ITS nrDNA sequences, of which the scope and composition were specified. The identity of O. xizangensis with the previously described O. dochenensis is shown; on the contrary, O. aromatica is singled out as an independent taxon. Ostericum scaberulum var. longiinvolucellatum is included in the genus Oreocomopsis as an independent species. Taxonomic features of the genus and differences from related genera are analysed. The species Oreocomopsis stelliphora has been assigned to the new genus Oreocomoides. The new genus Oreocomoides markedly differs from Oreocomopsis in many morphological characters, such as heterophyllous leaf blades: bipinnate in lower part and narrow-linear pinnate in upper part; bracts polymorphic, linear or pinnate with linear lobules; calyx teeth developed; petals with oval plate, at base with well-developed claw; pedicels rounded; mericarps ribs similar, narrowly-winged, slightly widened at the base.

Circumscription of the East Asia clade (Apiaceae subfamily Apioideae) and the taxonomic placements of several problematic genera

The East Asia (or Physospermopsis) clade was recognized in previous molecular phylogenetic investigations into the higher-level relationships of Apiaceae subfamily Apioideae. The composition of this clade, the phylogenetic relationships among its constituent taxa, and the placement of species previously determined to be problematic have yet to be resolved. Herein, nrDNA ITS sequences were obtained for 150 accessions of Apioideae, representing species whose distributions are in East Asia or genera having one or more species included within the East Asia clade. These data, along with published ITS sequences from other Apioideae (for 3678 accessions altogether), were subjected to maximum likelihood and Bayesian inference analyses. The results show that the East Asia clade contains representatives of 11 currently recognized genera: Hansenia, Hymenolaena, Keraymonia, Sinolimprichtia, Acronema, Hymenidium, Physospermopsis, Pimpinella, Sinocarum, Tongoloa, and Trachydium. However, the latter seven genera have members falling outside of the East Asia clade, including the generic types of all except Tongoloa. Within the clade, the species comprising these seven genera are widely intermingled, greatly increasing confusion among relationships than previously realized. The problematic species Physospermopsis cuneata is confirmed as falling within the East Asia clade, whereas P. rubrinervis allies with the generic type in tribe Pleurospermeae. Physospermopsis kingdon-wardii is confirmed as a member of the genus Physospermopsis, whereas the generic attributions of P. cuneata and Tongoloa stewardii remain unclear. Two species of Sinocarum (S. filicinum and S. wolffianum) are transferred into the genus Meeboldia. This is the most comprehensive molecular phylogenetic investigation of the East Asia clade to date, and while the results increase systematic understanding of the clade, they also highlight the need for further studies of one of the most taxonomically intractable groups in Apioideae.

Exploring diversity within the genus Tulostoma (Basidiomycota, Agaricales) in the Pannonian sandy steppe: four fascinating novel species from Hungary

Steppe vegetation on sandy soil in Hungary has recently been revealed as one of the hot spots in Europe for the stalked puffballs (genus Tulostoma). In the framework of the taxonomic revision of gasteroid fungi in Hungary, four Tulostoma species are described here as new to science: T. dunense, T. hungaricum, T. sacchariolens and T. shaihuludii. The study is based on detailed macro- and micromorphological investigations (including light and scanning electron microscopy), as well as a three-locus phylogeny of nrDNA ITS, nrDNA LSU and tef1-α sequences. The ITS and LSU sequences generated from the type specimen of T. cretaceum are provided and this resolved partly the taxonomy of the difficult species complex of T. aff. cretaceum.

Sequence Analysis of nrDNA ITS Region in Paşa and Şah Apple (Malus × domestica) Genotypes

In this study, phylogenetic analysis of Şah and Paşa apple genotypes was performed using nrDNA ITS sequences. After the plant leaves were brought to the laboratory, gDNAs were obtained by genomic DNA isolation method. PCR amplification was performed using primers ITS4 and ITS5A. ITS sequences of some apple and Rosaceae species were retrieved from NCBI, bioinformatics analyzes were made with Bioedit 7.2.3, Finch TV 1.4.0 and MEGA 6.0 programs and phylogenetic trees were constructed. In the study, ITS sequence length of the Paşa apple genotype was 656 bp, and the ITS sequence length of the Şah apple genotype was 649 bp. Only in the maximum likelihood phylogenetic tree constructed using sequences of apple genotypes, Paşa and Şah apple genotypes appeared in the same clade as Malus domestica cultivar="Casciana'' and Malus domestica cultivar="Rotella''. In the phylogenetic tree generated including other species belonging to the Rosaceae family; apple genotypes Pyrus, Cotoneaster, Crataegus, Sorbus, Eriobotrya and Prunus were detected in a same clade. Overall results clearly suggested that the ITS sequences were both suitable for differentiation between the selected genera and were compatible with previous phylogenetic studies.

<i>SIDERA TIBETICA</i> (<i>HYMENOCHAETALES</i>, <i>BASIDIOMYCOTA</i>), A NEW SPECIES TO RUSSIA

The first findings of recently described polypore fungus Sidera tibetica are revealed in the Caspian lowland forests of the Samursky National Park in the Republic of Dagestan, Russia. Basidiomata collected on Pinus brutia var. eldarica were studied by both light microscopy and molecular methods. Detailed information on new localities and habitats of Sidera tibetica as well as the collection numbers of specimens deposited in the Mycological Herbarium of the Komarov Botanical Institute of RAS (LE) are provided. Two complete sequences of ITS1–5.8S–ITS2 nuclear ribosomal DNA for Caucasian specimens of S. tibetica have been obtained and submitted to the GenBank database. Newly generated sequences are formed a separate well-supported clade in the Maximum Likelihood phylogenetic analysis together with all available ITS nrDNA sequences of S. tibetica originated from Belarussian and Chinese collections including the reference sequence from a holotype.

Acarospora americana (Acarosporaceae), a new species for Eurasia from the European part of Russia

A lichen species Acarospora americana H. Magn., new to Russia and Eurasia, was discovered on the territory of the Pavlovsk granite quarry in the Voronezh Region. A detailed description of morphological and anatomical features is given, chemotaxonomic analysis is carried out. A comparison is made with closely related species: A. fuscata (Ach.) Arnold and A. veronensis A. Massal. ITS nrDNA sequence is obtained.

Megalaria crispisulcans, a new isidiate species of Megalaria (Ramalinaceae) from Aotearoa / New Zealand

Megalaria crispisulcans .J. Marshall, Blanchon & de Lange (Ramalinaceae) is described as a new species from populations in Te Ika a Maui / North Island of Aotearoa / New Zealand, and on Rēkohu (Chatham Island), Aotearoa / New Zealand. The new species is morphologically closely allied to Megalaria orokonuiana from which it is distinguished by its nrDNA ITS sequence and morphologically by the usual presence of a white prothallus, bright green (when fresh) isidiate thallus and dark black apothecia whose rims are usually crinkled / undulose at maturity. A phylogeny based on nrDNA ITS sequences is presented which recovers Megalaria crispisulcans as sister to M. orokonuiana. The ecology, habitats and conservation status of M. crispisulcans is provided and an update on the conservation status of M. orokonuiana provided. A revised key to the Aotearoa / New Zealand species of Megalaria is provided.

Investigating DNA barcodes of plants growing in some areas of Iran with high crime rate: Quercus brantii, Curpressus arizonica, Crataegus pentagyna, Ziziphus Spina-chtista, and Buxus hyrcana

According to criminal botany, the offender unknowingly carries plant samples from the crime scene. Therefore, studying the genetic data of plants native to the crime scene can solve many ambiguities in the criminal files. In this regard, the aim of this study was to investigate the genome of 5 endemic plants in some areas of Iran with high crime rate. Quercus brantii, Curpressus arizonica, Crataegus pentagyna, Ziziphus Spina-chtista, and Buxus hyrcana were assessed using 1 genetic fragment on plastid regions (trnH-psbA) as well as 1 gene on nuclear chromosome called ITS. The alignment of DNA sequences of trnH-psbA and ITS genes was done using BioEdit, Clustal X, and Muscle v4.0 software programs. The phylogenetic analysis was performed on aligned data using Maximum Parsimony (MP) and the Bayesian methods. The Splits Tree v.4.14.4 software program was used for phylogenetic network analysis. Finally, the data combinability test was conducted using the Incongruence Length Difference (ILD) test by PAUP* software program. All data from nrDNA ITS and trnH-psbA sequences were consistent with Information Compatibility Test (ICT) results. Moreover, the nrDNA ITS indicated more resolved relationship than trnH-psbA. The results from MP and Bayesian analyses did not differ significantly between singular and combined forms, except for a slight variance in confidence interval of branches. As the phylogenetic trees provide more thorough and deeper conception of species relations, it is hoped that they would be useful to illuminate some forensic gaps in regions with high crime rates enriched by these plants, not only in Iran, but also in all areas over the world with this vegetation.

Gelasia attariana (Scorzonerinae, Cichorieae, Asteraceae), a new species from NW of Iran, inferred from morphological and molecular data

Gelasia attariana is described as a new species from northwest of Iran, near the border to Azerbaijan. The new species is morphologically similar to G. cinerea and G. wendelboi, mainly because of sharing caulescent perennial life form, entire lanceolate leaves with 3–7 parallel veins, tomentose indumentum of phyllaries and glabrous achenes without carpopodium, but differs in plant height, indumentum type of stems and leaves, length/width ratio of leaves, leaves color, apex of leaves and length of achenes and pappus. Morphological comparisons and taxonomic relationships of G. attariana with its closely related taxa are discussed. Consistent with morphological data, molecular evidences based on nrDNA ITS sequences confirmed that the new species can be recognized as a distinct taxon from its close relatives. Description, images of living plants and distribution map of the new species are also provided.