Ophioglossum Research Articles

Anti-inflammatory properties of ophioglonin derived from the fern Ophioglossum vulgatum L. via inactivating NF-κB and MAPK signaling pathways.

Medicinal plants contain bioactive compounds that have therapeutic effects on human health. Ophioglossum vulgatum L. is a representative species of the fern genus Ophioglossum that has anti-inflammatory properties as recognized in folk medicine. Herein, we performed a nitric oxide (NO) assay-guided screening in RAW264.7 cells to investigate the active components of the plant. We found that ophioglonin (OPN), a characteristic homoflavonoid of the genus Ophioglossum, is one of the bioactive components. Therefore, we performed a comparative analysis of the isolated compounds and found that OPN has effects similar to those of isolated dihydroquercetin and luteolin at the concentrations tested. The antioxidant and anti-inflammatory activities of OPN were extensively validated using lipopolysaccharide -stimulated RAW264.7 cells, mouse bone marrow-derived macrophages (BMDMs), and peritoneal exudate macrophages (PEMs). In vivo experiments with a carrageenan-induced mouse paw edema model further confirmed the anti-inflammatory effect of OPN. Additionally, we found that OPN and Ophioglossum vulgatum extracts inhibit the activation of signal transducers, NF-ĸB p65, IĸBα, ERK, p38, and JNK, consistent with the findings of pathway enrichment analysis. This work reinforces the anti-inflammatory properties of Ophioglossum vulgatum and indicates that OPN is a promising therapeutic agent for inflammation-associated disorders. Further clinical evaluations, including clinical trials, would be beneficial to validate the anti-inflammatory properties of OPN.

Sequencing of the complete plastome of the thermal adder’s-tongue fern, Ophioglossum thermale Kom. (Ophioglossaceae)

The Ophioglossaceae family, one of the oldest orders of extant ferns, exhibits diverse morphological and chromosomal characteristics. This study presents the first complete plastome sequence of thermal adder’s-tongue fern (Ophioglossum thermale), a species renowned for its antioxidant properties in traditional Chinese medicine. Our analyses revealed 27 simple sequence repeats (SSRs) in the plastome, with variations in SSR frequencies compared to related genera. Our phylogenetic analyses placed O. thermale within the Ophioglossum s.s. clade, supporting previous studies and suggesting polyphyly within the genus Ophioglossum based on the sensu PPG I system. The enlarged noncoding regions in fern organelles (ENRFOs) resulting from foreign DNA insertions in O. thermale were identified in the ycf2-trnH and trnT-trnfM regions, similar to other Ophioglossum species. ENRFOs were found at the LSC and SSC, but not in IRs in Ophioglossaceae. Consequently, foreign DNA insertions and lineage-specific SSRs shed light on plastome evolution in the Ophioglossaceae family.

Species of <I>Ophioglossum</I> in India: their Taxonomy and Phylogeny

The paper gives an account of the taxonomy of the genus Ophioglossum of which ten species are found in India. In doing so morphology of the leaf, stipules, spore and epidermal characters are taken into account and a key based on all characters is given to identify species. They are also summarised in a table. The question of confused taxonomy of species like O. reticulatum, O. vulgatum and O. aitchisoni has been specifically discussed and the probable phylogeny of Indian species is schematically represented. The characters of stipules which were overlooked so far are given in detail.

Organellar phylogenomics of Ophioglossaceae fern genera.

Previous phylogenies showed conflicting relationships among the subfamilies and genera within the fern family Ophioglossaceae. However, their classification remains unsettled where contrasting classifications recognize four to 15 genera. Since these treatments are mostly based on phylogenetic evidence using limited, plastid-only loci, a phylogenomic understanding is actually necessary to provide conclusive insight into the systematics of the genera. In this study, we have therefore compiled datasets with the broadest sampling of Ophioglossaceae genera to date, including all fifteen currently recognized genera, especially for the first time the South African endemic genus Rhizoglossum. Notably, our comprehensive phylogenomic matrix is based on both plastome and mitogenome genes. Inferred from the coding sequences of 83 plastid and 37 mitochondrial genes, a strongly supported topology for these subfamilies is presented, and is established by analyses using different partitioning approaches and substitution models. At the generic level, most relationships are well resolved except for few within the subfamily Ophioglossoideae. With this new phylogenomic scheme, key morphological and genomic changes were further identified along this backbone. In addition, we confirmed numerous horizontally transferred (HGT) genes in the genera Botrypus, Helminthostachys, Mankyua, Sahashia, and Sceptridium. These HGT genes are most likely located in mitogenomes and are predominately donated from angiosperm Santalales or non-Ophioglossaceae ferns. By our in-depth searches of the organellar genomes, we also provided phylogenetic overviews for the plastid and mitochondrial MORFFO genes found in these Ophioglossaceae ferns.

Spores Morphological Characteristics of Several Ophioglossaceae and Psilotaceae Fern in "Eka Karya" Botanical Garden - Bali

Ferns of Ophioglossaceae and Psilotaceae families can be found in the "Eka Karya" Botanical Garden, Bali as a collection or naturally growing in the botanical garden and those families are taxonomically classified as primitive ferns. The morphological characteristics of the family of Ophioglossaceae and Psilotaceae or even other types of ferns can be known through the morphological characteristics of the spores. This study aimed to characterize the spore morphology of the Ophioglossaceae and Psilotaceae fern of The "Eka Karya" Botanical Garden. Observation of the morphological characteristics of spores in the "Eka Karya" Botanical Garden was carried out by a simple method without acetolysis. The result of six types of fern plants from the family of Ophioglossaceae and Psilotaceae showed similarities to the spore unit, monads. Each family has unique characteristics that can be used to differentiate each other. The family Ophioglossaceae has a variety of spore colors, trilete spore types, mostly globose-shaped (except for one species the trilobate), the average polar diameter is 28.59±2.19 – 31.00±2.49 μm and the equatorial diameter is 32.85±1.74 – 37.55± 1.71 μm, the P/E ratio is 0.79-0.87, the shape based on the P/E ratio is suboblate, the spore size category is medium and has radial symmetry. For the family Psilotaceae, has a pale yellow-green spore color, monolete type, elongate-ellipsoidal shape, average polar diameter of 26.20±2.76 –29.81±2.96 μm and equatorial diameter of 52.98±2.55 – 63.31±4.65 μm, P/E ratio of 0.47-0.49, shape based on P/E ratio i.e. peroblate, large spore size category and has bilateral symmetry. Those characteristics are valuable taxonomic data that can be used as diagnostic characters for the identification of the families Ophioglossaceae and Psilotaceae.

Comparative spore morphology of ten species of the genus Ophioglossum L. from Kerala, India

The eusporangiate fern Ophioglossum brings some uncertainty among taxonomists while identifying the species. Therefore, a combined approach by considering the spore morphology and other morphological characters is more authentic for delimiting species of this genus. This study documented and compared the spore morphology of O. costatum, O. gramineum, O. lusitanicum, O. lusoafricanum, O. parvifolium, O. pendulum, O. petiolatum, O. raphaelianum, O. reticulatum and O. rubellum collected from Kerala, India by Scanning Electron Microscopy (SEM) analysis. SEM images of the spores for each species were provided, the P/E ratio was calculated, and the size classes of the spores were determined here. The occurance of O. lusitanicum in India was confirmed and O. madhusoodhananii was synonymised under O. costatum.

Ugonin L inhibits osteoclast formation and promotes osteoclast apoptosis by inhibiting the MAPK and NF-κB pathways

Bone loss is a major issue for patients with osteoporosis, arthritis, periodontitis, and bone metastasis; however, anti-resorption drugs used to treat bone loss have been linked to a variety of adverse effects. Helminthostachys zeylanica (L.) Hook, belonging to the family Ophioglossaceae, is commonly used in traditional Chinese medicine to treat inflammation and liver problems. In the current study, ugonin L extracted from H. zeylanica was shown to reduce the receptor activator of nuclear factor kappa beta ligand (RANKL)-induced osteoclastogenesis in RAW264.7 cells in a concentration-dependent manner. Ugonin L treatment also inhibited the mRNA expression of osteoclast markers. Ugonin L was also shown to promote cell apoptosis in mature osteoclasts and suppress RANKL-induced ERK, p38, JNK, and NF-κB activation. Taken together, ugonin L appears to be a promising candidate for the development of novel anti-resorption therapies.

The Complete Mitochondrial Genome of Ophioglossum vulgatum L. Is with Highly Repetitive Sequences: Intergenomic Fragment Transfer and Phylogenetic Analysis.

Many plant mitochondrial (mt) genomes have been sequenced but few in ferns. Ophioglossum vulgatum represents a typical species of fern genus Ophioglossum with medicinal and scientific value. However, its mt genome structure remains to be characterized. This study assembled and annotated the complete O. vulgatum mt genome and presented its structural characters and repeat sequences firstly. Its mt and chloroplast (cp) transfer sequences were explored, and the phylogenetic significance of both mt and cp genomes was also evaluated at the family level. Our results showed that the complete mt genome of O. vulgatum is a single circular genome of 369,673 bp in length, containing 5000 dispersed repetitive sequences. Phylogenetic trees reconstructed from cp and mt genomes displayed similar topologies, but also showed subtle differences at certain nodes. There exist 4818 bp common gene fragments between cp and mt genomes, of which more than 70% are located in tRNA intergenic regions (in mt). In conclusion, we assembled the complete mt genome of O. vulgatum, identified its remarkable structural characters, and provided new insights on ferns. The complementary results derived from mt and cp phylogeny highlighted that some higher taxonomic-level phylogenetic relationships among ferns remain to be resolved.

Phylogeny, character evolution, and systematics of the fern family Ophioglossaceae based on Sanger sequence data, plastomes, and morphology

Adder's tongue ferns or Ophioglossaceae are best known among evolutionary biologists and botanists for their highest chromosome count of any known organisms, the presence of sporophores, and simple morphology. Previous studies recovered and strongly supported the monophyly of the family and the two multi-generic subfamilies, Botrychioideae and Ophioglossoideae, but the relationships among these and two other subfamilies (Helminthostachyoideae and Mankyuoideae) are not well resolved preventing us from understanding the character evolution. The monophyly of and the relationships in the species-rich genus, Ophioglossum, have not well been understood. In this study, new phylogenetic trees are reconstructed based on four datasets: Sanger sequences of eight plastid markers of 184 accessions, 22 plastomes (12 are new), 29 morphological characters, and combined Sanger and morphological data. Our major results include: (1) the relationships among the four subfamilies are well resolved and strongly supported in Bayesian and parsimony analyses based on plastomes: Mankyua is sister to the rest, followed by Ophioglossoideae which are sister to Helminthostachys + Botrychioideae; (2) Sanger data, plastomes, and combined Sanger and morphological data recovered and strongly supported the monophyly of Ophioglossum in its current circumscription (sensu lato; s.l.) in Bayesian and/or parsimony analyses; (3) within Ophioglossum s.l., four deeply diverged clades are identified and the relationships among the four clades are well resolved; (4) evolution of 34 morphological characters is analyzed in the context of the new phylogeny, among which shape of rhizomes, germination time of spores, shape of early gametophytes, and a number of other characters are found to contain interesting phylogenetic signal; and (5) based on the new phylogeny and character evolution, we propose a new classification of Ophioglossaceae in which the currently circumscribed Ophioglossum is divided into four genera including three new ones: Goswamia, Haukia, and Whittieria considering their molecular, morphological, ecological, and biogeographical distinctiveness.

Discovery of a new species of Adder\u2019s tongue fern from India with comparative analysis of morphological and molecular attributes

Eusporangiate fern genus Ophioglossum L. is commonly known as Adder’s tongue fern as its fertile frond gives the appearance of snake tongue. A new species in this fern genus, O. trilokinathii belonging to Ophioglossaceae family has been discovered from the plateau region of Rajasthan State of northwestern India. The new species can be distinguished from other taxa of this genus by its smaller habit, subglobose-tuberous rhizome, basipetal emergence of young roots, aggregation of old decaying roots on rhizome apex, fertile stalk as well as spike short and thick, trophophylls in rosette, ovate or orbicular and a unique sporoderm sculpture pattern under SEM having broad reticulations with thick and raised muri enclosing large hexagonal or irregular areas on the distal and proximal faces of the spores hitherto unreported in any of the presently known taxa of Ophioglossum. In addition, comparative study of stomatal structure, foliar anatomy and nucleotide sequence data of its three chloroplast DNA markers (trnL-F, rbcL and psbA-trnH) was carried out. In view of all the attributes including habitat, ecology, morphology, foliar anatomy, stomatal features, palynology and molecular phylogenetic data, the present study suggests that the Ophioglossum specimen collected from plateau region of Rajasthan represents a hitherto undescribed species thereby warranting its establishment as O. trilokinathii sp. nov. A detailed comparative account of the new taxon with its allied species has also been provided.

Ferns at the digital herbarium of the Central Siberian Botanical Garden SB RAS.

BackgroundAccording to the data in Index Herbariorum as of 1 December 2020, there are 3426 active herbaria in the world, containing 396,204,891 specimens and 124 herbaria in Russia with more than 16,175,000 specimens. The Central Siberian Botanical Garden of the Siberian Branch of the Russian Academy of Sciences (CSBG SB RAS, Novosibirsk), founded in 1946, historically has two herbarium collections (NS and NSK). Currently these collections contain about 800,000 herbarium specimens comprising vascular plants, mosses, lichens and fungi gathered from all over the world. Digitisation of the NSK type specimens of vascular plants began in 2014 by using the special scanner Herbscan. In 2018, we started digitisation of the NS and NSK collections by using ObjectScan 1600.Pteridophytes (ferns, lycophytes and their extinct free-sporing relatives) are a diverse group of plants that today comprises approximately 12,900 species and plays a major role in terrestrial ecosystems. All herbarium specimens of ferns, collected over 170 years between 1851 and 2021 and stored in the NS and NSK collections, were digitised in 2021, placed at the CSBG SB RAS digital Herbarium (http://herb.csbg.nsc.ru:8081) and published through GBIF. Twenty families of Polypodiopsida, but not Equisetaceae, were included in this dataset. Family Ophioglossaceae was digitised and published in GBIF as a separate dataset.New informationBy August 2021, more than 62,600 specimens with good quality images and fully-captured label transcriptions had been placed at CSBG SB RAS Digital Herbarium. A total of 7,758 records of fern occurrences of 363 taxa in the world with 92% geolocations including 5100 records from Russia with 98.7% geolocations that are new for GBIF.org in 2021 were entered. In the dataset specimens from 43 countries of Europe, Asia, America, Africa and Australia (Oceania), 89% of them from Russia, are presented.

Transcriptome-wide SNPs for Botrychium lunaria ferns enable fine-grained analysis of ploidy and population structure.

Ferns are the second most diverse group of land plants after angiosperms. Extant species occupy a wide range of habitats and contribute significantly to ecosystem functioning. Despite the importance of ferns, most taxa are poorly covered by genomic resources and within‐species studies based on high‐resolution markers are entirely lacking. The genus Botrychium belongs to the family Ophioglossaceae, which includes species with very large genomes and chromosome numbers (e.g., Ophioglossum reticulatum 2n = 1520). The genus has a cosmopolitan distribution with 35 species, half of which are polyploids. Here, we establish a transcriptome for Botrychium lunaria (L.) Sw., a diploid species with an extremely large genome of about ~19.0–23.7 Gb. We assembled 25,677 high‐quality transcripts with an average length of 1,333 bp based on deep RNA‐sequencing of a single individual. We sequenced 11 additional transcriptomes of individuals from two populations in Switzerland, including the population of the reference individual. Based on read mapping to reference transcript sequences, we identified 374,463 single nucleotide polymorphisms (SNPs) segregating among individuals for an average density of 14 SNPs per kilobase. We found that all 12 transcriptomes were most likely from diploid individuals. The transcriptome‐wide markers provided unprecedented resolution of the population genetic structure, revealing substantial variation in heterozygosity among individuals. We also constructed a phylogenomic tree of 92 taxa representing all fern orders to ascertain the placement of the genus Botrychium. High‐quality transcriptomic resources and SNP sets constitute powerful population genomic resources to investigate the ecology, and evolution of fern populations.

Diversity of Pteridophytes in Nipani Taluk, Karnataka, India

Considering diverse ecological and topographical conditions, Nipani Taluk is close to the Eastern ranges of Western Ghats. Adi Hill and Stavanidhi Ranges are the extended branches of the Western Ghats. An attempt was undertaken to enumerate the ferns from Nipani Taluk to fill the gap of a valid document of the ferns from the study area. During the study, 14 species from 9 different genera of 6 different families were recorded. The distribution of some species is found to be unique from Adi Hill and Stavanidhi Ranges. The study revealed that the number of species from family Pteridaceae was the highest, common to both the ranges followed by family Ophioglossaceae, Marsileaceae, Lomariopsidaceae, Salviniaceae and Polypodiaceae. Adiantum philippense L. was dominantly found in both the studied areas, while Adiantum incisum Forssk. and Actiniopteris radiata (Sw.) Link are less dominant. Whereas Microsorum membranaceum (D. Don) Ching is least populated and found very rarely at both the study area.

Diversity, distribution and conservation status of the Adder’s-tongue ferns in Goa, India

The cosmopolitan fern genus Ophioglossum (Ophioglossaceae) is distributed from low to high altitude plateaux, coastal plains and forest floors of India. This genus has received special attention from pteridologists worldwide since the discovery that Ophioglossum reticulatum possesses the largest number of chromosomes. There are, however, no reported studies of Ophioglossum in Goa, hence the present investigation was undertaken to study the diversity, distribution and conservation status of Ophioglossum in that state. A total of six species were collected from different localities, of which four (O. nudicaule, O. lusitanicum, O. parvifolium, and O. reticulatum) are reported as new distributional records for Goa State. A detailed morpho-taxonomy, illustration and photographs of all collected species are given, along with a key to the species.

<p><strong><em>Ophioglossum chaloneri</em></strong><strong>: A new species of <em>Ophioglossum </em>(Ophioglossaceae: Pteridophyta) from India</strong></p>

In the present study, we describe Ophioglossum chaloneri as a new species from the Hurdulu forest area, near Ranchi (Jharkhand, India), on the basis of morphological, palynological, cytological, and molecular analyses. The new species can be distinguished from all other congeners of genus Ophioglossum in having their unusually small linear rhizome with a long stalk of trophophore, a sporophore arising from a distance from the base of the trophophore hence not always adnate or appearing inserted at the base. Triradiate, round spores show small tubercles towards the proximal pyramidal area and irregular smooth elevated exine striations on both the proximal and distal faces. The ventral face of the trophophore is unusually thin all around margin thereby showing thin whitish margin from the lower side, a situation that may be due to lack of chloroplast all around the margin at the lower face of the trophophore. This feature is not observed in any other species of the genus. Molecular study based on three chloroplast DNA sequences (cpDNA) rbcL, trnL-F, and psbA-trnH also supports O. chaloneri as a distinct species.

Comparative phylogenetics of some species of Ophioglossum L. (Ophioglossaceae: Pteridophyta) in India with comments on evolutionary significance of high palaeoploidy and rare morphological traits

The genomic constitution of the genus Ophioglossum is complicated because of its being geologically ancient and of long evolutionary history. A lot of plants of the same and different species resemble each other as well as show significant differences among morphological characters among themselves. Many of the new species described have exhibited several unknown features not observed in any species of the genus so far. In this paper, we wish to emphasize that mere observations on shape and size of trophophyll (leaf) and rhizome, should not be taken reliably in delimiting different species of the genus Ophioglossum because of several variations already reported for many species. Preference should be given to exine ornamentation of spores as emphasized by the present study. Detailed observations on exine ornamentation of spores under SEM have offered precious and distinctive features; semi-tuberculate, beaded strings, and stripped exines are a few forms of spore ornamentations offering clear distinction among species. Long chains of circulate-alete (round alete) spores, sometimes linear tetrads as described hereunder in Ophioglossum aletum are novel features exclusively confined to this species. Comparative chromosome profiles of many species described earlier do not offer any significant support to the identity except in O. eliminatum, which has the lowest chromosome count (n = 90). This appears that since almost all species possess intraspecific polyploidy, the intragenomic variability must be loaded with interspecific hybridization contents including chromosomal aberrations. Quite possibly, this may be one of the reasons that homologous chromosomes rarely form trivalents and multivalents during early meiotic stages, despite the fact that palaeoautoploidy must have resulted in copying homologous chromosomes. In recent years, repeated phylogenetic analysis of chloroplast rbcL have revealed that many species in India appear to be offshoots of a vulgatum clade which are likely to be the hybridization products involving O. costatum, O. vulgatum, O. reticulatum, and also including of other interspecific hybridizations. It is remarkable to mention that some of the new taxa published by us, viz. O. eliminatum, O. indicum, O. aletum and O. chalonerii (unpublished), explicitly show distinct phylogenetic relationship maintaining independent genomic identity. Furthermore, each new species shows some rare uniquely confined trait not possessed by any other species of the genus. These morphological traits also suggest that the plants may be expressing extremely rare genic-combinations which might have been brought forth due to autopolyploidy and/or allopolyploidy (hybridizations) during the past few decades. Obviously, we will need a multidisciplinary approach (morphological, anatomical, biochemical) in order to identify different species of Ophioglossum with new and unusual genomic identity and also to understand possible evolutionary mechanisms undergoing within the genome.

Revealing a new species of Ophioglossum (Ophioglossaceae-Pteridophyta) from India with palynological and phylogenetic implications

ABSTRACTIn recent years, Indian Ophioglossum L. taxonomy and systematic are receiving increasing interest from pteridologists as India has very rich germplasm of the genus. We present hereunder a new species of the genus Ophioglossum (Ophioglossaceae) from the marshy areas of southern region of Gujarat state, India. Ophioglossum hitkishorei sp. nov. can be diagnosed from other congeners by a combination of its smaller size, globose to subglobose, non-stoloniferous rhizome, number of trophophylls up to six, the absence of persistent leaf bases at top of rhizome and by very unique structure of spores. Spores are typically characterized by possessing granules deposition as exine strings bound around spores on the distal face and granulate assemblage on the proximal face of triradiate spores observed under SEM and also clearly distinct under light microscopic observations. As far as known, beaded strings of exine ornamentation have never been observed in any species of Ophioglossum in the world flora. Molecular study, based on two chloroplast DNA sequences trnL-F and psbA-trnH, also strongly supports O. hitkishorei sp. nov. as a distinct new species.

Genus Ophioglossum L., from Western Part of India with Special Reference to Gujarat State

The genus Ophioglossum L. is revised for the Gujarat state and there were reported eight species viz. Ophioglossum costatum R. Br., O. gramineum Willd., O. gujaratense SM Patil, RN Kachhiyapatel, R Patel and KS Rajput, O. lusitanicum L., O. reticulatum L., O. parvifolium Grev. & Hook., O. nudicaule L. f. and O. thermale Kom., from different parts of the state. Amongst these, Ophioglossum thermale Kom. and O. lusitanicum L. were collected for the first time from Jambughoda Wildlife Sanctuary and this result is documented as a new distributional record for Gujarat state.

Taxonomic significance of morphological characters of spores in the family Ophioglossaceae (Psilotopsida)

Primary and secondary ornamentation of spores of ferns of the family Ophioglossaceae are important characters, used in the taxonomy of this group. Considering the small number of published data on those characters in the Ophioglossaceae from Central and Eastern Europe, this study aimed (1) to describe morphological characters of spores of Botrychium and Ophioglossum species and to assess their taxonomic significance; (2) to analyse variation in spore size between and within species of these genera, based on specimens from various habitats and geographic locations; and (3) to create a key to species identification based on the diagnostic characters of the spore ornamentation. We examined spores of 6 species from 16 localities in Central and Eastern Europe. Results of cluster analysis based on morphological characters of spores indicate that the species form well-defined groups, partly reflecting the systematics of the Ophioglossaceae. A comparison of our results with data from North-West Europe and North America shows differences in some species. The broad range of variation in the size of spores from European populations, in comparison with American ones, may be linked with polyploidy of the studied species. Our findings will help in identification of spores of the family Ophioglossaceae in palynological and palaeobotanical studies.

Prodromus of a fern flora for Bolivia. V. Ophioglossaceae

We provide a synopsis to the family Ophioglossaceae in Bolivia, comprising seven species in four genera, Botrypus, Cheiroglossa and Sceptridium with one species each, and Ophioglossum with four species.