Further evidence for the hybrid status of the Brazilian native fern Hypolepis ×paulistana (Dennstaedtiaceae).

Hypolepis ×paulistana was described in 2016 as a putative hybrid, known from a single gathering. The hybrid status of these plants was based solely on the intermediate morphology of the sporophyte, when compared to its presumed parent species. These were thought to be H. stolonifera and H. rugosula, but, H. rigescens (Kunze) T. Moore could not be explicitly ruled out either. In the present work, we tested the hybrid status of Hypolepis ×paulistana adding palynological evidence and by using chloroplast sequences to unambiguously identify the maternal progenitor of the species. We find that sporangia of Hypolepis ×paulistana contain both well-formed spores, as well as spores with morphological and developmental anomalies. The size of the regular spores and the abnormal spores suggest that H. ×paulistana is likely a diploid, and probably infertile hybrid. The ornamentation of the regular spores of H. ×paulistana is similar to that of H. stolonifera. The chloroplast sequences of H. ×paulistana are identical to those of H. stolonifera, as well as their sister position within the global phylogeny of the genus. Thus, we provide new evidence for the hybrid status of H. ×paulistana, and we corroborate the earlier finding that H. stolonifera is the maternal parent.

New insights into the evolution of the fern family Dennstaedtiaceae from an expanded molecular phylogeny and morphological analysis.

Dennstaedtiaceae has 270 species, a worldwide distribution, and an edge-colonizing habit that is unusual among ferns. Aneuploidy, polyploidy, and hybrids are common in the family. Combining morphology, anatomy, chromosome number, and geographical distributions with our newly generated molecular phylogeny, we provide new insights into the evolution of the family. We paid special attention to Hypolepis. Our molecular dataset of five cpDNA markers is the most comprehensive to date, comprising 72 species (and a total of 98 taxa), of which 33 are Hypolepis (45 taxa). We also generated divergence-time estimates through BEAST with four fossil calibrations. We recovered three sub-families in Dennstaedtiaceae: Monachosoroideae (monogeneric), Dennstaedtioideae, and Hypolepidoideae. Monachosoroideae has a chromosome base number of x = 28; Hypolepidoideae of x = 26; while in Dennstaedtioideae this is still obscure, with different numbers ranging from 30 to 47. Dennstaedtioideae genera require re-circumscriptions because Dennstaedtia is polyphyletic. In Hypolepidoideae, the six genera are monophyletic. Within Hypolepis, seven geographically distinct clades were recovered; but we found no strong morphological characters to define them. Within the family, the long-creeping rhizome evolved with a change in habit: from shade-tolerant to edge-colonizers, to thicket-formers. Short or extremely large leaves are derived conditions. Sorus shape and position, glandular hairs, and prickles are homoplastic. Hybridization/allotetraploidy in Hypolepis can be suggested by the combined data. In our phylogenetic hypothesis, Dennstaedtiaceae originated around 135 Ma, with the split of Monachosoroideae around 94 Ma, and the split between Dennstaedtioideae/Hypolepidoideae around 78 Ma. All extant genera are inferred to be relatively young. Hypolepis started to diversify around 10 Ma, and it probably originated in east Asia and/or Oceania. Hypolepis reached the Neotropics twice: through elements of the Hypolepis rugosula clade (which originated at 7 Ma), and through the ancestor of the Neotropical clade, which originated at 3.1 Ma and was prickly.

Hiya: A new genus segregated from Hypolepis in the fern family Dennstaedtiaceae, based on phylogenetic evidence and character evolution.

The relationship of Hypolepis brooksiae, H. nigrescens, and H. scabristipes to the remainder of Hypolepis (Dennstaedtiaceae) has been questioned by previous authors based on their unique combination of morphological characters and different base chromosome number. Using four chloroplast genes including rbcL, atpA, rpL6, and rps4-trnS intergenic spacer (IGS) from 32 samples, representing 24 species of Dennstaedtiaceae, we recovered a clade comprising H. brooksiae and H. nigrescens, distinct from the remaining species of Hypolepis. This clade is resolved as sister to the clade comprising Blotiella, Paesia and Histiopteris. We reconstructed ancestral states of 16 morphological characters and found that this clade is distinguished by indeterminate, scandent leaves exhibiting rhythmic growth, provided with recurved black-tipped prickles, and stipule-like pinnules that protect the emerging crosier and pinnae departures, rachis-costa architecture where the adaxial sulcus is confluent with the next lower order, and a base chromosome number of x = 29. In light of this molecular and morphological evidence, we describe a new genus, Hiya, and provide nomenclatural combinations to accommodate the three known species segregated from Hypolepis: Hiya brooksiae, Hiya nigrescens, and Hiya scabristipes.

Palynological analysis of Dennstaedtiaceae taxa from the Paranaense Phytogeografic Province that produce monolete spores and its systematic implications (I): Blotiella lindeniana , Histiopteris incisa and Paesia glandulosa.

The genera of Dennstaedtiaceae have sporophytes with very different morphological characteristics between each other, and this feature has made difficult the systematic circumscription of the family. This reason makes necessary the study of new characters that allow a better understanding of the relations within the group. The spore morphology and wall structure of Blotiella lindeniana, Histiopteris incisa and Paesia glandulosa from the Paranaense Phytogeographic Province were studied using light microscope, and scanning and transmission electron microscope. The exospore has two layers and, according to the species, the exospore surface bears pila, echinae, verrucae, bacula and tubercles. The perispore has two or three layers and its surface is psilate, baculate or rugulate. The variability found in the sculpture of the spores and their stratification and ultrastructure of perispore reflects the morphological differences observed in the sporophyte of the species studied. Additionally, while the stratification and ultrastructure of the exospore is shared by the Dennstaedtiaceae species, their ornamentation could be a character to distinguish species into the clade "hypolepidoide". The finding of spores with similar characteristics in phylogenetically unrelated families allows us to suggest that palynological features do not have an evolutionary value to establish relationships between groups above the genus level.

Palynological analysis of Dennstaedtiaceae taxa from the Paranaense Phytogeografic Province that produce monolete spores and its systematic implications (I): Blotiella lindeniana, Histiopteris incisa and Paesia glandulosa

ABSTRACT The genera of Dennstaedtiaceae have sporophytes with very different morphological characteristics between each other, and this feature has made difficult the systematic circumscription of the family. This reason makes necessary the study of new characters that allow a better understanding of the relations within the group. The spore morphology and wall structure of Blotiella lindeniana, Histiopteris incisa and Paesia glandulosa from the Paranaense Phytogeographic Province were studied using light microscope, and scanning and transmission electron microscope. The exospore has two layers and, according to the species, the exospore surface bears pila, echinae, verrucae, bacula and tubercles. The perispore has two or three layers and its surface is psilate, baculate or rugulate. The variability found in the sculpture of the spores and their stratification and ultrastructure of perispore reflects the morphological differences observed in the sporophyte of the species studied. Additionally, while the stratification and ultrastructure of the exospore is shared by the Dennstaedtiaceae species, their ornamentation could be a character to distinguish species into the clade "hypolepidoide". The finding of spores with similar characteristics in phylogenetically unrelated families allows us to suggest that palynological features do not have an evolutionary value to establish relationships between groups above the genus level.

Palynological analysis of Dennstaedtiaceae taxa from the Paranaense Phytogeographic Province that produce Trilete spores II: Microlepia speluncae and Pteridium arachnoideum.

The spore morphology and wall ultrastructure of Microlepia speluncae and Pteridium arachnoideum from the Paranaense Province were analyzed with LM, SEM and TEM and a comparative analysis was carried out. In both species the spores are covered by a three-dimensional network of threads branched and fused, tangentially arranged to the surface, and some free-end threads are also seen. The species were differentiated by morphology and the frequency of threads fusion and the networks distribution on the surface of the spores. In both species the exospore is two-layered in section, both layers are traversed by single or branched channels. The perispore is three-layered in section: the inner layer is adhered to the exospore, the middle layer is formed of a three-dimensional network of threads and the outer layer is discontinuous. The perispore ultrastructure of Microlepia speluncae was interpreted as formed of helical subunits displayed around a central channel. The spore morphology and perispore ultrastructure allow differentiating Microlepia from Pteridium but also to the other Dennstaedtiaceae genera that grow in the Paranaense Province. The results obtained allow establishing relationships that let us recognize different groups and gave a new reference to get a better knowledge of the family.

Characterization of the stem anatomy of the Eocene fern Dennstaedtiopsis aerenchymata (Dennstaedtiaceae) by use of confocal laser scanning microscopy.

PREMISE OF THE STUDY: Permineralization provides the most faithful known mode of three-dimensional preservation of the morphology and cellular anatomy of fossil plants. Standard optical microscopic documentation of such structures can provide only an approximation of their true three-dimensional form and is incapable of revealing fine-structural (<300 nm) details, deficiencies that can be addressed by the use of confocal laser scanning microscopy (CLSM). METHODS: To demonstrate the usefulness of CLSM in such studies, we compare confocal laser scanning micrographs and optical photomicrographs of the permineralized tissues of rhizomes and petioles of the Eocene fern Dennstaedtiopsis aerenchymata preserved in cherts of the Clarno Formation of Oregon, USA, and the Allenby Formation (Princeton chert) of British Columbia, Canada. KEY RESULTS: The laser-induced fluorescence detected by CLSM produces crisp high-resolution images of the three-dimensionally permineralized tissues of Dennstaedtiopsis aerenchymata. Tissues analyzed include the epidermis and epidermal hairs, cortex, aerenchyma, endodermis, vascular tissue, and pith-for each of which, CLSM yields results superior to those of standard optical microscopy. CLSM and previous Raman spectroscopic analyses of the same specimens provide evidence consistent with original biochemistry. CONCLUSIONS: Use of CLSM to characterize the morphology and cellular anatomy of permineralized fossil plants can provide accurate data in two and three dimensions at high spatial resolution, information that can be critically important to taxonomic, taphonomic, and developmental interpretations. Results presented here from this first detailed CLSM-based study of permineralized plant axes indicate that this nonintrusive, nondestructive technique should be widely applicable in paleobotany.

Four new antioxidant phenylpropanoid glycosides from Microlepia pilosissima.

Four new phenylpropanoid glycosides, 9-O-[ß-D-glucopyranosyl-(1→2)-α-L-rhamnopyranosyl]-3,4-dimethoxy-cinnamic acid (1), 9-O-[ß-D-glucopyranosyl-(1→2)-α-L-rhamnopyranosyl]-4-methoxycinnamic acid (2), 9-O-α-L-rhamnopyranosyl-3,4-dimethoxy-cinnamic acid (3), and 9-O-[6-Oacetyl-ß-D-glucopyranosyl]-4-methoxy-cinnamic acid (4), together with three known compounds 9-O-α-L-rhamnopyranosyl-4-hydroxy-cinnamic acid (5), 9-O-ß-D-glucopyranosyl-4-methoxycinnamic acid (6), and 9-O-ß-D-glucopyranosyl-3,4-dimethoxy-cinnamic acid (7) were isolated from the 70% EtOH extract of the dry fronds of Microlepia pilosissima. Their chemical structures were elucidated by spectroscopic analysis. Moreover, 1 and 2 exhibited comparable scavenging activities with (±)-α-tocopherol against DPPH radicals, while compounds 3-7 displayed moderate antioxidant activities.

Three new amides from Microlepia pilosissima.

Phytochemical investigation of 70% EtOH extract of the dry fronds of Microlepia pilosissima has resulted in isolation of three new amides, (7E)-N-(3'-hydroxyl-4'-methoxy)-phenylethyl-4-hydroxyl-cinnamamide (1), (7E)-N-(3',4',5'-trihydroxyl)-phenylethyl-4-hydroxyl-cinnamamide (2), and (7E)-N-(3',4',5'-trihydroxyl)-phenylethyl-4-methoxy-cinnamamide (3). Their structures were characterized by spectroscopic analysis and chemical method, including 1D NMR, 2D NMR, and HR-ESI-MS.

Polyphenol content and modulatory activities of some tropical dietary plant extracts on the oxidant activities of neutrophils and myeloperoxidase.

Young leaves of Manihot esculenta Crantz (Euphorbiaceae), Abelmoschus esculentus (Malvaceae), Hibiscus acetosella (Malvaceae) and Pteridium aquilinum (Dennstaedtiaceae) are currently consumed as green vegetables by peoples in sub-Saharan Africa, Latin America, Asia and their migrants living in Western Europe. Sub-Saharan peoples use Manihot, Abelmoschus and Hibiscus also in the folk medicine to alleviate fever and pain, in the treatment of conjunctivitis, rheumatism, hemorrhoid, abscesses, ... The present study investigates the effects of aqueous extracts of those plants on the production of reactive oxygen species (ROS) and the release of myeloperoxidase (MPO) by equine neutrophils activated with phorbol 12-myristate 13-acetate (PMA). The ROS production was measured by lucigenin-enhanced chemiluminescence (CL), and the release of total MPO by an ELISA method. The study also investigates the effect of the extracts on the activity of MPO by studying its nitration activity on tyrosine and by using a new technique called SIEFED (Specific Immunological Extraction Followed by Enzymatic Detection) that allows studying the direct interaction of compounds with the enzyme. In all experiments, the aqueous extracts of the plants developed concentration-dependent inhibitory effects. A moderate heat treatment did not significantly modify the inhibitory capacity of the extracts in comparison to not heated ones. Total polyphenol and flavonoid contents were determined with an HPLC-UV/DAD analysis and a spectroscopic method using Folin-Ciocalteu reagent. Some polyphenols with well-known antioxidant activities (caffeic acid, chlorogenic acid, hyperoside, rosmarinic acid and rutin) were found in the extracts and may partly explain the inhibitory activities observed. The role of those dietary and medicinal plants in the treatment of ROS-dependent inflammatory diseases could have new considerations for health.

Two new sesquiterpene glucosides from Dennstaedtia scabra (Wall.) Moore.

Two new sesquiterpene glucosides onitioside A (1) and dennstoside B (2), were isolated from the 95% EtOH extract of Dennstaedtia scabra (Wall.) Moore, together with seven known compounds, onitisin (3), pterosin A (4), pinocembrin (5), pinocembrin 7-rutinoside (6), kaempferol (7), nicotiflorin (8), and galangin (9). Their structures were determined by extensive spectroscopic analysis.

Fase sexual de los helechos Odontosoria schlechtendalii y Odontosoria scandens (Dennstaedtiaceae) / Sexual phase of the ferns Odontosoria schlechtendalii and Odontosoria scandens (Dennstaedtiaceae)

The results of morphogenesis studies of the sexual phase of Odontosoria schlechtendalii and O. scandens are presented in this paper. In O. schlechtendalii and O. scandens the spores are triletes, non-chlorophyllous, and germinación is of the Vittaria-type, devoid of perine, the exine is smooth and sometimes coarsely ridged to reticulate. The development pattern is of the Adiantum-type. The adult gametophyte is cordate-spathulate, with probable presence of anteridiogen in O. schlechtendalii, both species are glabrous. Sex organs are of the common type of the leptosporangiate ferns. The first leaves appeared after 56-92 days of culture, with petiole and plate divide narrow, trichomes bifurcate and stomate anomocytic.

[Sexual phase of the ferns Odontosoria schlechtendalii and Odontosoria scandens (Dennstaedtiaceae)]. / Fase sexual de los helechos Odontosoria schlechtendalii y Odontosoria scandens (Dennstaedtiaceae).

The results of morphogenesis studies of the sexual phase of Odontosoria schlechtendalii and O. scandens are presented in this paper. In O. schlechtendalii and O. scandens the spores are triletes, non-chlorophyllous, and germinación is of the Vittaria-type, devoid of perine, the exine is smooth and sometimes coarsely ridged to reticulate. The development pattern is of the Adiantum-type. The adult gametophyte is cordate-spathulate, with probable presence of anteridiogen in O. schlechtendalii, both species are glabrous. Sex organs are of the common type of the leptosporangiate ferns. The first leaves appeared after 56-92 days of culture, with petiole and plate divide narrow, trichomes bifurcate and stomate anomocytic.

[New species of Dennstaedtia (Filicales: Dennstaedtiaceae) to Mexico]. / Una especie nueva de Dennstaedtia (Filicales: Dennstaedtiaceae) para México.

Dennstaedtia gracilis A. Rojas et Tejero (Dennstaedtiaceae) is herein described and illustrated as a new species endemic to Mexico. Its belongs to the group of winged adaxial secondary costae species but it differs by the combination of characters as smaller fronds, deltate pinnules and lobed segment apex.