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Funder: Georges and Antoine Claraz Foundation grant, Funder: University Research Priority Program “Evolution in Action” of the University of Zurich, Land plants comprise two large monophyletic lineages, the vascular plants and the bryophytes, which diverged from their most recent common ancestor approximately 480 million years ago. Of the three lineages of bryophytes, only the mosses and the liverworts are systematically investigated, while the hornworts are understudied. Despite their importance for understanding fundamental questions of land plant evolution, they only recently became amenable to experimental investigation, with Anthoceros agrestis being developed as a hornwort model system. Availability of a high-quality genome assembly and a recently developed genetic transformation technique makes A. agrestis an attractive model species for hornworts. Here we describe an updated and optimized transformation protocol for A. agrestis, which can be successfully used to genetically modify one more strain of A. agrestis and three more hornwort species, Anthoceros punctatus, Leiosporoceros dussii, and Phaeoceros carolinianus. The new transformation method is less laborious, faster, and results in the generation of greatly increased numbers of transformants compared with the previous method. We have also developed a new selection marker for transformation. Finally, we report the development of a set of different cellular localization signal peptides for hornworts providing new tools to better understand the hornwort cell biology.

An important step for plant diversification was the transition from freshwater to terrestrial habitats. The bryophytes and all vascular plants share a common ancestor that was probably the first to adapt to life on land. A polysaccharide-rich cell wall was necessary to cope with newly faced environmental conditions. Therefore, some pre-requisites for terrestrial life have to be shared in the lineages of modern bryophytes and vascular plants. This review focuses on hornwort and liverwort cell walls and aims to provide an overview on shared and divergent polysaccharide features between these two groups of bryophytes and vascular plants. Analytical, immunocytochemical, and bioinformatic data were analysed. The major classes of polysaccharides—cellulose, hemicelluloses, and pectins—seem to be present but have diversified structurally during evolution. Some polysaccharide groups show structural characteristics which separate hornworts from the other bryophytes or are too poorly studied in detail to be able to draw absolute conclusions. Hydroxyproline-rich glycoprotein backbones are found in hornworts and liverworts, and show differences in, for example, the occurrence of glycosylphosphatidylinositol (GPI)-anchored arabinogalactan-proteins, while glycosylation is practically unstudied. Overall, the data are an appeal to researchers in the field to gain more knowledge on cell wall structures in order to understand the changes with regard to bryophyte evolution.

Several paleoecological studies over the last decade examining plant microfossils in Late Quaternary wetland sediment cores and dryland soil samples from multiple sites on Rapa Nui found three dist...

Main conclusion Two isoforms of phenylalanine ammonia-lyase (PAL) have been isolated as cDNA sequences from the hornwort Anthoceros agrestis. The encoded enzymes convert l-phenylalanine and to lower extents l-tyrosine and l-histidine. Thus, the functional presence of the general phenylpropanoid pathway in one of the earliest land plant groups is established. Abstract The hornwort Anthoceros agrestis has an elaborated phenolic metabolism resulting in phenolic compounds, such as rosmarinic acid or megacerotonic acid. The general phenylpropanoid pathway is involved in the biosynthesis of these compounds. Two phenylalanine ammonia-lyase (PAL) genes, AaPAL1 and AaPAL2, have been identified in Anthoceros agrestis and the protein with an N-terminal 6xHis-tag heterologously synthesized in Escherichia coli for a full biochemical characterization. Both PAL proteins accept l-phenylalanine, l-tyrosine as well as l-histidine as substrates, although the activity is explicitly the highest with l-phenylalanine. Km values as well as catalytic efficiencies were determined for phenylalanine (Km AaPAL1 39 µM, AaPAL2 18 µM) and tyrosine (Km AaPAL1 3.3 mM, AaPAL2 3.5 mM). In suspension cultures of Anthoceros agrestis, PAL genes were transcribed in parallel to rosmarinic acid (RA) accumulation and both showed highest abundance in the early growth phase. In a phylogenetic tree, both AaPAL amino acid sequences grouped within a clade with PAL amino acid sequences of diverse origin ranging from non-vascular to vascular plants, while most PALs from eudicots and monocots were mainly found in two other clades. The similarity of the hornwort PAL amino acid sequences to PAL sequences from vascular plants is more than 80% showing a strong conservation within the land plants. With this characterization of PALs from Anthoceros agrestis together with former investigations concerning cinnamic acid 4-hydroxylase and 4-coumaric acid CoA-ligase, the functional presence of the general phenylpropanoid pathway in this hornwort is proven.

Nothoceros renzaglianus J.C.Villarreal, L.V.Campos & Uribe, Syst. Bot. 37(1): 32. 2012, (renzagliensis) TYPE:— COLOMBIA, Nariño: Pasto, La Divina Pastora, 2968 m., 19 April 2010, J.C. Villarreal et al. 1080 (Holotype: COL)

The relationships among the four major embryophyte lineages (mosses, liverworts, hornworts, vascular plants) and the timing of the origin of land plants are enigmatic problems in plant evolution. Here, we resolve the monophyly of bryophytes by improving taxon sampling of hornworts and eliminating the effect of synonymous substitutions. We then estimate the divergence time of crown embryophytes based on three fossil calibration strategies, and reveal that maximum calibration constraints have a major effect on estimating the time of origin of land plants. Moreover, comparison of priors and posteriors provides a guide for evaluating the optimal calibration strategy. By considering the reliability of fossil calibrations and the influences of molecular data, we estimate that land plants originated in the Precambrian (980–682 Ma), much older than widely recognized. Our study highlights the important contribution of molecular data when faced with contentious fossil evidence, and that fossil calibrations used in estimating the timescale of plant evolution require critical scrutiny.

Main conclusion Anthoceros agrestis hydroxycinnamoyltransferase accepts shikimic and 3-hydroxyanthranilic acids while hydroxycinnamoylester/amide 3-hydroxylase (CYP98A147) preferred p-coumaroyl-(3-hydroxy)anthranilic acid compared to the shikimic acid derivative. Alternative pathways towards rosmarinic acid have to be considered. Abstract Rosmarinic acid (RA) is a well-known ester of caffeic acid and 3,4-dihydroxyphenyllactic acid. In the search for enzymes involved in RA biosynthesis in the hornwort Anthoceros agrestis, the hydroxycinnamoyltransferase sequence with the highest similarity to rosmarinic acid synthase from Lamiaceae has been amplified and heterologously expressed in Escherichia coli. In parallel, the single cytochrome P450 sequence belonging to the CYP98 group in Anthoceros agrestis was isolated and expressed in Saccharomyces cerevisiae which did not result in protein formation. Codon optimization and co-expression with NADPH:cytochrome P450 reductase (CPR) from Coleus blumei resulted in the formation of active enzymes. Both, the hydroxycinnamoyltransferase and CYP98 were characterized with respect to their temperature and pH optimum as well as their substrate acceptance. The hydroxycinnamoyltransferase (AaHCT6) readily accepted p-coumaroyl- and caffeoyl-CoA with a slightly higher affinity towards p-coumaroyl-CoA. The best acceptor substrate was shikimic acid (Km 25 µM with p-coumaroyl-CoA) followed by 3-hydroxyanthranilic acid (Km 153 µM with p-coumaroyl-CoA). Another accepted substrate was 2,3-dihydroxybenzoic acid. Anthranilic acid and 4-hydroxyphenyllactic acid (as precursor for RA) were not used as substrates. p-Coumaroylesters and -amides are substrates hydroxylated by CYP98 hydroxylases. The only CYP98 sequence from Anthoceros agrestis is CYP98A147. The best substrates for the NADPH-dependent hydroxylation were p-coumaroylanthranilic and p-coumaroyl-3-hydroxyanthranilic acids while p-coumaroylshikimic and p-coumaroyl-4-hydroxyphenyllactic acids were poor substrates. The biosynthetic pathway towards rosmarinic acid thus still remains open and other enzyme classes as well as an earlier introduction of the 3-hydroxyl group to afford the caffeic acid substitution pattern must be taken into consideration.

Extant land plants consist of two deeply divergent groups, tracheophytes and bryophytes, which shared a common ancestor some 500 million years ago. While information about vascular plants and the two of the three lineages of bryophytes, the mosses and liverworts, is steadily accumulating, the biology of hornworts remains poorly explored. Yet, as the sister group to liverworts and mosses, hornworts are critical in understanding the evolution of key land plant traits. Until recently, there was no hornwort model species amenable to systematic experimental investigation, which hampered detailed insight into the molecular biology and genetics of this unique group of land plants. The emerging hornwort model species, Anthoceros agrestis, is instrumental in our efforts to better understand not only hornwort biology but also fundamental questions of land plant evolution. To this end, here we provide an overview of hornwort biology and current research on the model plant A. agrestis to highlight its potential in answering key questions of land plant biology and evolution.

Background The amphibian, non-vascular, gametophyte-dominant, bio-indicator class, bryophytes; with their wide ranges of habitat have attained importance due to their promising medicinal attributions and therapeutic role; mostly aided by presence of aromatic bibenzyl and bisbybenzyl class of compounds. Bibenzyls are steroidal ethane derivatives, resembling the structural moiety of bioactive dihydro-stilbenoids or iso-quinoline alkaloids. These stress triggered secondary metabolites are the by-products of the flavonoid biosynthetic pathway. Different classes of bryophytes (Bryophyta, Marchantiophyta and Anthocerotophyta) possess different subtypes of bibenzyls and dimeric bisbibenzyls. Among the liverwort, hornwort and mosses, former one is mostly enriched with bibenzyl type constituents as per the extensive study conducted for phytochemical deposit. Considering macrocyclic and acyclic group of bibenzyls and bisbybenzyls, generally marchantin type compounds are reported vividly for significant biological activity that includes neuro-nephro-cardio-protection besides anti-allergic, anti-microbial, anti-apoptotic and cytotoxic activities studied on in-vitro and in-vivo models or on cell lines. Result The critical analysis of reported chemical and pharmaceutical attributions of bibenzyls and bis-bibenzyls yielded detailed report on this compound class along with their application, mode of action, natural source, techniques of synthesis, extraction procedure, isolation and characterization. Further, the structure activity relationship studies and bioactivity of bibenzyls derived from non-bryophytic origin were also summarized. Conclusion This review encompasses prospective biological application of botanical reservoir of this primarily ignored, primeval land plant group where recent technical advances has paved the way for qualitative and quantitative isolation and estimation of novel compounds as well as marker components to study their impact on environment, as bio-control agents and as key leads in future drug designing. Graphical abstract

The genus Notothylas Sull. ex A.Gray is reviewed for Brazil based on type material, herbarium specimens and recent collections. Five species of the genus are recognized, Notothylas vitalii is synonymized with N. javanica and two species are new to science: Notothylas granulata sp. nov and N. vermiculata sp. nov. A diagnostic key, descriptions, illustrations and taxonomic comments are provided.

Angiosperms represent one of the most spectacular terrestrial radiations on the planet1, but their early diversification and phylogenetic relationships remain uncertain2–5. A key reason for this impasse is the paucity of complete genomes representing early-diverging angiosperms. Here, we present high-quality, chromosomal-level genome assemblies of two aquatic species—prickly waterlily (Euryale ferox; Nymphaeales) and the rigid hornwort (Ceratophyllum demersum; Ceratophyllales)—and expand the genomic representation for key sectors of the angiosperm tree of life. We identify multiple independent polyploidization events in each of the five major clades (that is, Nymphaeales, magnoliids, monocots, Ceratophyllales and eudicots). Furthermore, our phylogenomic analyses, which spanned multiple datasets and diverse methods, confirm that Amborella and Nymphaeales are successively sister to all other angiosperms. Furthermore, these genomes help to elucidate relationships among the major subclades within Mesangiospermae, which contain about 350,000 species. In particular, the species-poor lineage Ceratophyllales is supported as sister to eudicots, and monocots and magnoliids are placed as successively sister to Ceratophyllales and eudicots. Finally, our analyses indicate that incomplete lineage sorting may account for the incongruent phylogenetic placement of magnoliids between nuclear and plastid genomes., Genome assemblies of two aquatic species, prickly waterlily and rigid hornwort, clarify the early diversification and phylogeny of plants. Multiple independent polyploidization events are inferred in each of the five major angiosperm clades.

Key message Cinnamic acid 4-hydroxylase from the hornwort Anthoceros agrestis (AaC4H) was functionally expressed in the moss Physcomitrella patens and characterized at biochemical and molecular levels. Abstract Cinnamic acid 4-hydroxylase (C4H), a cytochrome P450-dependent hydroxylase, catalyzes the formation of 4-coumaric acid (=4-hydroxycinnamic acid) from trans-cinnamic acid. In the hornwort Anthoceros agrestis (Aa), this enzyme is supposed to be involved in the biosynthesis of rosmarinic acid (a caffeic acid ester of 3-(3,4-dihydroxyphenyl)lactic acid) and other related compounds. The coding sequence of AaC4H (CYP73A260) was expressed in the moss Physcomitrella patens (Pp_AaC4H). Protein extracts from the transformed moss showed considerably increased C4H activity driven by NADPH:cytochrome P450 reductase of the moss. Since Physcomitrella has own putative cinnamic acid 4-hydroxylases, enzyme characterization was carried out in parallel with the untransformed Physcomitrella wild type (Pp_WT). Apparent Km-values for cinnamic acid and NADPH were determined to be at 17.3 µM and 88.0 µM for Pp_AaC4H and 25.1 µM and 92.3 µM for Pp_WT, respectively. Expression levels of AaC4H as well as two Physcomitrella patens C4H isoforms were analyzed by quantitative real-time PCR. While PpC4H_1 displayed constantly low levels of expression during the whole 21-day culture period, AaC4H and PpC4H_2 increased their expression during the first 6–8 days of the culture period and then decreased again. This work describes the biochemical in vitro characterization of a cytochrome P450-dependent enzyme, namely C4H, heterologously expressed in the haploid model plant Physcomitrella patens.

Hornworts, liverworts and mosses are three early diverging clades of land plants, and together comprise the bryophytes. Here, we report the draft genome sequence of the hornwort Anthoceros angustus. Phylogenomic inferences confirm the monophyly of bryophytes, with hornworts sister to liverworts and mosses. The simple morphology of hornworts correlates with low genetic redundancy in plant body plan, while the basic transcriptional regulation toolkit for plant development has already been established in this early land plant lineage. Although the Anthoceros genome is small and characterized by minimal redundancy, expansions are observed in gene families related to RNA editing, UV protection and desiccation tolerance. The genome of A. angustus bears the signatures of horizontally transferred genes from bacteria and fungi, in particular of genes operating in stress-response and metabolic pathways. Our study provides insight into the unique features of hornworts and their molecular adaptations to live on land., A draft genome sequence of the hornwort Anthoceros augustus confirms the phylogenetic relationships among the three clades of bryophytes and provides insight into the unique characteristics of hornworts and their adaptations to live on land.

As a result of studies of the flora of the liverworts and hornworts of the Darwinskiy Reserve, conducted in the Cherepovets District of the Vologda Region in 2018, and literature data, an annotated list including 64 species of liverworts and 1 species of hornworts was compiled. 19 species were recorded for the first time for the reserve, including Cephalozia curvifolia, Chiloscyphus fragilis, and Riccia rhenana, recorded for the first time for the Vologda Region. For every species data on frequency, habitats, reproduction, and, if necessary, taxonomic comments are provided. For rare and new species for the Vologda Region and the reserve the coordinates of collecting sites and examined specimens are listed. The specimens are kept in the Bryological Herbarium of Komarov Botanical Institute of the Russian Academy of Sciences (LE). The findings of 13 species were not confirmed by the collections of 2018 due to the natural transformation of the territory or random factors. In the examined part of the reserve Lophozia ascendens, Odontoschisma denudatum, Phaeoceros carolinianus (as P. laevis s. l.) and Riccia canaliculata, protected in the Vologda Region, were found. Calypogeia suecica, Geocalyx graveolens, and Syzygiella autumnalis are in need of biological control of their condition in the Vologda Region.

Anthoceros subtilis Stephani (1916: 1003). Fig. 2, 3A–B. Type:— INDIA. South Canara, Mangalore, August 1907, R. Pfleiderer 6127 (holotype G-G00060882). For synonyms see Hasegawa (1998), Peng & Zhu (2013) and Villarreal et al. (2015). Monoicous. Plants growing on soil in rosettes. Thallus light to dark green, up to 2.6 mm in length and 3.5 mm in width. Dorsal lamellae absent or scarce. Margins of thallus highly crenulate. Thallus up to 22 cells thick in solid regions (106– 599 μm), cavernous, schizogenous cavities 75–150 × 115–155 μm, 1–3 layers of schizogenous cavities in transverse section. Single chloroplast per cell with central pyrenoid. Rhizoids hyaline to brown; unbranched. Ventral spheric Nostoc colonies abundant. Antheridia scattered. Antheridial chamber with up to 15 antheridia per cavity, antheridia yellowish to dark brown, 102–125 μm μm in length, with four-tiered jacket cell arrangement. Archegonia scattered, with five neck cells covered by mucilage. Involucres erect, cylindrical, up to 1.5 mm long, with smooth surface or with very few lamellae, 3–6 cells thick in solid regions (28–159 μm), cavernous, schizogenous cavities 15–30 × 22–47 μm, one layer of schizogenous cavities in transverse section. Up to three sporophytes per plant. Sporophyte foot with a layer of palisade-like cells. Capsules erect, up to 10 mm long, opening by two longitudinal valves. Epidermal layer of capsule one cell thick in transverse section. Epidermal cells of capsule rectangular, elongated, 7.5–19 × 37–85 μm and thick-walled, stomata abundant (17 per mm 2), stomata 24–40 × 42–58 μm. Assimilative layer 4–5 cells (26.4–95.8 μm) thick in transverse section. Columella present within the sporophyte, extending to the tip, rigid and brown, 16 cells in cross section. Sporogenous layer one spore tetrad thick. Pseudoelaters short, thin-walled, light brown to dark brown at maturity, 42.8–81.6 μm long (65.1±26.9, n=22), 3.9–33.4 μm in width, and 1–5 cells at maturity. Spores light brown to brown at maturity, 26.6–42.1 μm (34.9±3.5, n=31) in equatorial diameter. Distal spore surface with numerous papillate to tuberculate projections, up to 3 μm in height and 2.2 μm in width, sometimes confluent at their bases. Proximal spore surface with a distinct trilete mark, 0.89–1.5 μm in width, irregularly covered with verrucae; triangular areas covered with numerous papillate to tuberculate projections, up to 2.8 μm in height and 2.5 μm in width, sometimes confluent at their bases. The entire spore surface is covered with minute papillae, up to 0.8 μm in height and 0.4 μm in width. Habitat: —Growing on the sides of roads and around coffee plantations in tropical montane cloud forest, with other hornworts such as A. telaganus Stephani (1916:1005) and Phaeoceros sp., ca. 1300 m. Distribution:— China, Central African Republic, India, Japan, Korean Peninsula, Nepal, Thailand, Vietnam (Asthana & Srivastava 1991, Hasegawa 1998, Peng & Zhu 2013, Villarreal et al. 2015, Choi et al. 2021) and Mexico. Affinities and differentiation:—Mexican populations of A. subtilis are characterized by small, brown spores (26.6–42.1 μm), with papillate to tuberculate projections covering both proximal and distal surfaces. Within Mexican taxa, A. aff sambesianus Stephani (1916: 996) shares with A. subtilis the small sized plants (Ibarra-Morales et al. 2015), up to 7 mm in diameter in A. aff sambesianus and up to 2.6 mm in length and 3.5 mm in width in A. subtilis, but can be differentiated by spore ornamentation pattern. Anthoceros aff sambesianus possess a low reticulated network with foramens and central sub-globose tubercles in triangular areas of proximal face (Fig. 3 C-D), while A. subtilis presents irregular tuberculate projections and minute papillae (Fig. 3 A-B). According to Peng & Zhu (2013), A. subtilis resembles A. angustus Stephani (1916: 1001) in spore morphology, nevertheless, they can be differentiated by the presence of marginal spongy gemmae on the thallus margins of A. angustus, absent in A. subtilis; and by the size of the pseudoelaters, which are considerably longer in A. angustus (40–496 µm) and somewhat shorter in A. subtilis (42–199 µm). Anthoceros angustus is restricted to Asia (Peng & Zhu 2013). Specimens examined:— MEXICO. Chiapas: Unión Juarez, roadside, 1438 m, 15°04’03” N 92°04’53” W, 26 July 2019, Ariadna Ibarra-Morales 2019 -14b (FCME). Remarks: —Mexican populations of this species were always found growing with other bryophytes. In one of the most crowded patches, some rosettes of A. subtilis were formed by very small plants of less than 1 mm in capsule height and thallus diameter. Some also exhibited an unusual erect morphology of the thallus, similar to the thallus morphology that has been reported in A. erectus Kashyap (1915: 9) (Asthana & Srivastava 1991). In these dwarf plants, the surface of the thallus was barely large enough for the base of the involucre to fit. This dwarf morphotype was probably a result of space and resource availability and may have also been affected by a fungal infection, since red hyphae were found within the gametophyte of some of these plants. Previous reports of this species from Asia (Asthana & Srivastava 1991, Hasegawa, 1998; Peng & Zhu 2013) document slightly larger thallus size (5–6 mm in diameter) and larger spores (35–45 µm) than those of Mexican plants (thallus up to 3.5 mm and spores of 26.6–42.1 μm), but spore morphology is very similar to that illustrated by Peng & Zhu (2013) from China., Published as part of Ibarra-Morales, Ariadna, 2021, Anthoceros subtilis and A. telaganus (Anthocerotaceae): two new records for America from the hornwort flora of Mexico, pp. 93-104 in Phytotaxa 529 (1) on pages 94-97, DOI: 10.11646/phytotaxa.529.1.7, http://zenodo.org/record/5814330, {"references":["Stephani, F. (1916) Species Hepaticarum Vol. V. Geneve, 991 - 1008.","Hasegawa, J. (1998) Taxonomic status of Anthoceros subbrevis Haseg. and taxonomy of its allied taxa. Bryological Research 7 (6): 173 - 177. https: // doi. org / 10.24474 / bryologicalresearch. 7.6 _ 173","Peng, T. & Zhu, R. - L. (2013) A revision of the genus Anthoceros (Anthocerotaceae, Anthocerotophyta) in China. Phytotaxa 100 (1): 21 - 35. https: // doi. org / 10.11646 / phytotaxa. 100.1.3","Villarreal, J. C., Cargill, D. C., Soderstrom, L., Hagborg, A., von Konrat, M. (2015) Notes on early land plants today. 70. Nomenclatural notes in hornworts (Anthocerotophyta). Phytotaxa 208 (1): 92 - 96. https: // doi. org / 10.11646 / phytotaxa. 208.1.9","Asthana, A. K. & Srivastava, S. C. (1991) Indian hornworts (A taxonomic study). Bryophytorum Bibliotheca 42: 1 - 158.","Choi, S. S., Bakalin, V. & Park, S. J. (2021) Integrating continental mainland and islands in temperate East Asia: liverworts and hornworts of the Korean Peninsula. PhytoKeys 176: 131 - 226. https: // doi. org / 10.3897 / phytokeys. 176.56874","Ibarra-Morales, A., Muniz, M. E. & Valencia, S. (2015): The genus Anthoceros (Anthocerotaceae, Anthocerotophyta) in Central Mexico. Phytotaxa 205 (4): 215 - 228. http: // dx. doi. org / 10.11646 / phytotaxa. 205.4.1","Kashyap, S. R. (1915) Morphological and biological notes on new and little known West-Himalayan liverworts III. New Phytologist 14 (1): 1 - 18."]}

Anthoceros telaganus Stephani. Fig. 4, 5. Type:— Indonesia, Java, I’rov. Preanger, Telaga bodas, Schiffner #19 (holotype G-G00061328, isotype NY-NY00231473). Monoicous. Plants growing on soil in rosettes. Thallus dark green, growing up to 15 mm in length and 15 mm in width. Dorsal lamellae are scarce to abundant. Margins of thallus lobulate or irregularly crenulate. Thallus up to 15 cells (368–845 μm) thick in solid regions, cavernous, schizogenous cavities 113–286 × 159–481 μm, 1–2 layers of schizogenous cavities in transverse section. One chloroplast per cell with a central pyrenoid. Rhizoids hyaline and unbranched. Ventral spheric Nostoc colonies are abundant. Antheridia scattered. Antheridial chamber with up to 10 antheridia per cavity, yellowish to brown, 59–106 μm in length, and with four-tiered jacket cell arrangement. Archegonia not observed. Involucres erect and cylindrical, up to 5 mm long, surface smooth to abundantly lamellate, 7–9 cells (154–262 μm) thick in solid regions, cavernous, schizogenous cavities 32–86 × 45–154 μm, 1 (–2) layer of schizogenous cavities in transverse section. Up to seven sporophytes per gametophyte. Sporophyte foot with a layer of palisade-like cells. Capsules erect, up to 30 mm long, opening by two longitudinal valves. Epidermal layer of capsule one cell thick in transverse section. Epidermal cells of capsule rectangular, elongated, 7–14 × 57–121 μm and thick-walled, stomata abundant (10 per mm 2), 34–46 × 44–64 μm.Assimilative layer 3–5 cells (32.89–114.4 μm) thick in transverse section. Columella present within sporophyte, extending to the tip, rigid and brownish to blackish, 16 cells in cross section. Sporogenous layer one spore tetrad thick. Pseudoelaters elongate, thin-walled, dark brown at maturity, 39.8–104.9 μm (73.2±16.8, n=19) long, 3.8– 26.7 μm in width, 1–2 cells at maturity. Spores black at maturity, 32.7–53.21 μm (40.48±3.8, n=44) in diameter. Distal spore surface has numerous spinose-lamellate projections, up to 3.7 μm in height, frequently irregularly confluent at their bases, often tipped by a papilla of 0.53–2.98 μm in diameter. Proximal spore surface has a distinct trilete mark, 0.9–3.66 μm in width, irregularly covered with verrucae, a smooth strip along either side of trilete mark, up to 3.98 μm in width (2.89 μm in average), triangular areas covered with numerous spinose-lamellate projections, up to 3.8 μm in height, frequently irregularly confluent at their bases, tipped by a papilla of 0.64–3.25 μm in diameter. Habitat:—Growing on the sides of roads and around coffee plantations in tropical montane cloud forest, with other hornworts such as A. subtilis and Phaeoceros sp., 1200–1300 m. Distribution:— Indonesia (Java and Sumatra) (Meijer 1957, Hasegawa 1998, Söderström et al. 2010) and Mexico. Affinities and differentiation:—Only one species known from Mexico, Anthoceros orizabensis (Stephani 1916: 965) Hässel de Menéndez (1990: 211) (SEM spore micrographs of this species can be found in Hässel de Menéndez (1990), plate 4, fig. 9 on page 212), resembles the spore ornamentation of A. telaganus, sharing the smooth strip along the trilete mark. Nevertheless, the smooth strip in A. orizabensis is incomplete, not reaching the equatorial girdle of the spores; moreover, the distal face of the spore has spines in A. orizabensis, while A. telaganus has a spinose-lamellate reticulated distal face. FIGURE 5. Scanning electron microscopy (SEM) micrographs of spores from Mexican populations of Anthoceros telaganus. A-B. From A. Ibarra-Morales 2019-11; C-D. From A. Ibarra-Morales 2019-20. A & C distal surface, B & D proximal surface. Other American species that share the smooth strip along the spore trilete mark of the proximal face are Anthoceros cavernosus Stephani (1916: 998), A. caucasicus Stephani (1923: 427) and A. fusiformis Austin (1875: 28) (Hässel de Menéndez 1990, Sérgio et al. 2020). Anthoceros fusiformis frequently presents a strap shaped thallus and lacks a pyrenoid in the chloroplasts (Hasegawa 1993), both characters differentiate it from A. telaganus. Regarding A. cavernosus and A. caucasicus, both species have spores with spinose outgrows reticulated at the base with ridges enclosing both lumina and pits on the distal face, and the proximal face with scattered spinose outgrowths, sometimes confluent at their bases (SEM spore micrographs of these species can be found in Hässel de Menéndez 1990 and Sérgio et al. 2020). In A. telaganus the spores present blunter papillate projections and an incomplete reticulum on the distal face, while the proximal face presents spinose-lamellate projections that are tipped by a papilla. In the location here reported, A. subtilis can be differentiated from A. telaganus by spore ornamentation, mature spore color and size, light brown to brown in the first species and black in the second, and smaller spores in A. subtilis. Plants not bearing sporophytes are difficult to differentiate. Thalli of Anthoceros subtilis are smaller and paler in color than those of A. telaganus. Anthoceros telaganus also presents an abundantly lamellate thallus morphotype that could be easily identified in this location. Specimens examined:— MEXICO. Chiapas: Unión Juarez, roadside, 1260 m, 15°03’35”N 92°04’44”W, 24 July 2019, Ariadna Ibarra-Morales 2019 -11 (FCME); 1438 m, 15°04’03”N 92°04’53”W, 26 July 2019, Ariadna Ibarra-Morales 2019-20 (FCME). Remarks: —Within the mature region of the sporophyte, colorless and shrunken (probably aborted) spores were observed amongst black mature regular-size spores in all plants examined of this species. The percentage of these inviable spores could not be estimated, but it was observed frequently that at least one spore of almost every tetrad displayed this condition and remain attached to a fully mature spore. This phenomenon was not observed in any other hornwort species growing in the same locations. Morphological variation within Mexican plants of this species was observed mainly in two characters: dorsal lamellae on the thallus and spore ornamentation. Dorsal lamellae on the thallus presented a variation range from nearly absent to abundant and plants expressing different ranges of this character could be found in the same population, but generally not close together in the same rosette. Regarding spore ornamentation, spores of northern populations (A. Ibarra -Morales #2019-20) were, in most of the individuals examined, more spinose and forming a more reticulated pattern (Fig. 5C–D) when compared to the ornamentation of southern populations (A. Ibarra -Morales #2019-11) (Fig. 5A–B); nevertheless, both spore morphologies were present in all collections. It is noteworthy that the type (Stephani 1916) and the collections here described, present similar ecological preferences, both being collected at or near coffee plantations. Meijer (1957) also states that this species grows on cultivated areas at 700–2000 m, and on earth walls and roadsides., Published as part of Ibarra-Morales, Ariadna, 2021, Anthoceros subtilis and A. telaganus (Anthocerotaceae): two new records for America from the hornwort flora of Mexico, pp. 93-104 in Phytotaxa 529 (1) on pages 97-100, DOI: 10.11646/phytotaxa.529.1.7, http://zenodo.org/record/5814330, {"references":["Meijer, W. (1957) Notes on some Malayan species of Anthoceros L. (Hepaticae) - II. The Journal of the Hattori Botanical Laboratory 18: 411 - 423. https: // doi. org / 10.18968 / jhbl. 18.0 _ 1","Hasegawa, J. (1998) Taxonomic status of Anthoceros subbrevis Haseg. and taxonomy of its allied taxa. Bryological Research 7 (6): 173 - 177. https: // doi. org / 10.24474 / bryologicalresearch. 7.6 _ 173","Soderstrom, L., Gradstein, S. R. & Hagborg, A. (2010) Checklist of the hornworts and liverworts of Java. Phytotaxa 9: 53 - 149. https: // doi. org / 10.11646 / phytotaxa. 9.1.7","Stephani, F. (1916) Species Hepaticarum Vol. V. Geneve, 991 - 1008.","Hassel de Menendez, G. (1990) Las especies de Anthoceros y Folioceros (Anthocerotophyta) de America del Norte, Sud y Central; la ornamentacion de sus esporas y taxonomia. Candollea 45 (1): 201 - 219.","Stephani, F. (1923) Species Hepaticarum Vol. VI. Geneve, 425 - 430.","Austin, C. F. (1875) Notes on the Anthocerotaceae of North America, with descriptions of several new species. Bulletin of the Torrey Botanical Club 6 (4): 25 - 29.","Sergio, C., Draper, D. & Porley, R. D. (2020) Three new records of Anthocerotophyta for Western Africa (Sierra Leone) based on spore ornamentation of a specimen collected by A. Harrington, with an emphasis on Anthoceros sect. Fusiformes Grolle. Journal of Bryology 44 (2): 160 - 168. https: // doi. org / 10.1080 / 03736687.2019.1709022","Hasegawa, J. (1993) Taxonomical studies on Asian Anthocerotae V. A short revision of Taiwanese Anthocerotae. Acta Phytotaxonomica et Geobotanica 44 (2): 97 - 112."]}

PREMISE Nitrogen-fixing endosymbioses with cyanobacteria have evolved independently in five very different plant lineages. Expanding knowledge of these symbioses promises to improve the understanding of symbiosis evolution and broaden the toolkit for agricultural engineering to reduce artificial fertilizer use. Here we focused on hornworts, a bryophyte lineage in which all members host cyanobacteria, and investigated factors shaping the diversity of their cyanobiont communities. METHODS We sampled hornworts and adjacent soils in upstate New York throughout the hornwort growing season. We included all three sympatric hornwort species in the area, allowing us to directly compare partner selectivity. To profile cyanobacteria communities, we established a metabarcoding protocol targeting rbcL-X with PacBio long reads. RESULTS The hornwort cyanobionts detected were phylogenetically diverse, including clades that do not contain other known plant symbionts. We found significant overlap between hornwort cyanobionts and soil cyanobacteria, a pattern not previously reported in other plant-cyanobacteria symbioses. Cyanobiont communities differed between host plants only centimeters apart, but we did not detect an effect of sampling time or host species on the cyanobacterial community structure. CONCLUSIONS This study expands the phylogenetic diversity of known symbiotic cyanobacteria. Our analyses suggest that hornwort cyanobionts have a tight connection to the soil background, and we found no evidence that time within growing season, host species, or distance at the scale of meters strongly govern cyanobacteria community assembly. This study provides a critical foundation for further study of the ecology, evolution, and interaction dynamics of plant-cyanobacteria symbiosis.

New genome sequences for early-branching, aquatic flowering plants provide fresh insights into angiosperm phylogeny as well as key resources for deciphering their genome adaptive landscapes.

Dendroceros is one of the most diverse genera of hornworts, with 41 species and a widespread distribution. The genus is highly distinctive by its epiphytic habitat, gametophytes with perforated win...

The potential of oil palm ash (OPA) to enhance H

SUMMARY The placenta of the anthocerote, Dendroceros tubercularis Hatt., consists of long and branched haustorial cells, that arise from the foot and gametophyte transfer cells. Both cell types contain electron-dense vacuolar deposits that were digested by pronase and therefore are assumed to be protein. These deposits were negative to the PATAg test for carbohydrates. Protein bodies were also found in the parenchyma cells of the foot and younger meristematic cells at the base of the capsule. Vacuolar deposits of osmiophilic material in the gametophyte cells external to the placenta were stained non-specifically with PATAg method and were not affected by pronase. The haustorial cells have pleomorphic plastids lacking starch and a thylakoid system, whereas the transfer cells have well developed chloroplasts. No pronase-sensitive material was detected in the apoplastic space separating gametophyte and sporophyte cells. These results suggest that protein is synthesized in the haustorial cells, perhaps from precursors provided by transfer cells, and is then transferred, via plasmodesmata, to the parenchyma cells of the foot and eventually to the cells of the growing capsule.

Main conclusion Tyrosine aminotransferase (AaTAT) from the hornwort Anthoceros agrestis Paton (Anthocerotaceae) was amplified and expressed in E. coli. The active enzyme is able to accept a wide range of substrates with distinct preference for l-tyrosine, therefore, possibly catalysing the initial step in rosmarinic acid biosynthesis. Abstract The presence of rosmarinic acid (RA) in the hornwort A. agrestis is well known, and some attempts have been made to clarify the biosynthesis of this caffeic acid ester in lower plants. Parallel to the biosynthesis in vascular plants, the involvement of tyrosine aminotransferase (EC 2.6.1.5; TAT) as the initial step was assumed. The amplification of a nucleotide sequence putatively encoding AaTAT (Genbank MN922307) and expression in E. coli were successful. The enzyme proved to have a high acceptance of l-tyrosine (Km 0.53 mM) whilst slightly preferring 2-oxoglutarate over phenylpyruvate as co-substrate. Applying l-phenylalanine as a potential amino donor or using oxaloacetate or pyruvate as a replacement for 2-oxoglutarate as amino acceptor resulted in significantly lower catalytic efficiencies in each of these cases. To facilitate further substrate search, two methods were introduced, one using ninhydrin after thin-layer chromatography and the other using derivatisation with o-phthalaldehyde followed by HPLC or LC–MS analysis. Both methods proved to be well applicable and helped to confirm the acceptance of further aromatic and aliphatic amino acids. This work presents the first description of a heterologously expressed TAT from a hornwort (A. agrestis) and describes the possible entry into the biosynthesis of RA and other specialised compounds in a so far neglected representative of terrestrial plants and upcoming new model organism.