A permineralized Early Cretaceous lycopsid from China and the evolution of crown clubmosses.

Lycopodiaceae are one of three surviving families of lycopsids, a lineage of vascular plants with a fossil history dating to at least the Early Devonian or perhaps the Late Silurian (c. 415 Ma). Many fossils have been linked to crown Lycopodiaceae, but the lack of well-preserved material has hindered definitive recognition of this group in the paleobotanical record. New, exceptionally well-preserved permineralized lycopsid fossils from the Early Cretaceous (125.6 ± 1.0 Ma) of Inner Mongolia, China, were examined in detail using acetate peel and micro-computed tomography techniques. The anatomy of extant Lycopodiaceae was analyzed for comparison using fluorescence microscopy. Phylogenetic relationships of the new fossil to extant Lycopodiaceae were evaluated using parsimony and maximum likelihood analyses. Lycopodicaulis oellgaardii gen. et sp. nov. provides the earliest unequivocal and best-documented evidence of crown Lycopodiaceae and Lycopodioideae, based on anatomically-preserved fossil material. Recognition of Lycopodicaulis in Asia during the Early Cretaceous indicates the presence of crown Lycopodiaceae at this time, and striking similarities of stem anatomy with extant species provide a framework for the understanding of the interaction of branching and vascular anatomy in crown-group lycopsids.

First insights on the biogeographical history of Phlegmariurus (Lycopodiaceae), with a focus on Madagascar.

We explored the biogeographical history of a group of spore-bearing plants focusing on Phlegmariurus (Lycopodiaceae), a genus of lycophytes comprising ca. 250 species. Given its wide distribution in the Southern Hemisphere, Phlegmariurus provides a good model to address questions about the biogeographical processes underlying southern distributions, notably in Madagascar and surrounding islands, also called the Western Indian Ocean (WIO). Our aims were (i) to discuss the systematics of the Malagasy species in the light of molecular phylogenetic results, (ii) to provide the first dating analysis focused on Phlegmariurus and (iii) to understand the relative role of vicariance, dispersal and diversification in the origin of the Malagasy Phlegmariurus species. The phylogenetic relationships were inferred based on three plastid DNA regions (rbcL, trnH-psbA and trnL+trnL-trnF) and on a dataset comprising 93 species, including 16 Malagasy species (80% of the total Malagasy diversity for the genus). Our results highlighted the need to combine Malagasy Huperzia species in Phlegmariurus, as well as the polyphyly of widely distributed species: Phlegmariurus phlegmaria, P. squarrosus and P. verticillatus with the Malagasy species not belonging with the types of P. phlegmaria or P. squarrosus. This led us to propose new circumscriptions of Phlegmariurus species, especially in the WIO. Our dating analysis, relying on fossil calibrations, showed that Phlegmariurus would have originated in the Late Cretaceous and diversified in the Early Eocene. The biogeographical analysis highlighted uncertainties about the biogeographical origins of Phlegmariurus: the genus would have started to diversify in an ancestral range covering at least the Neotropics and Australasia. Hypotheses on the biogeographical history of Phlegmariurus were discussed, especially the roles of long distance dispersal, migration via Antarctica and via the Boreotropics. Six long distance dispersal events over the last 40 Ma would explain the Malagasy species diversity of Phlegmariurus, in combination with an important in situ diversification starting in the Miocene.

Molecular phylogenetics and the morphology of the Lycopodiaceae subfamily Huperzioideae supports three genera: Huperzia, Phlegmariurus and Phylloglossum.

The generic classification of huperzioid Lycopodiaceae was tested using Bayesian inference and Maximum likelihood phylogenetic analyses of DNA sequences from four chloroplast loci for 119 taxa and optimisation of 29 morphological characteristics onto the phylogeny. Consistent with previous studies, the subfamilies Lycopodioideae and Huperzioideae are monophyletic and diagnosable by synapomorphies that correlate with differences in their life-histories. Within the Huperzioideae, the monophyly of the widely adopted genus Huperzia (excl. Phylloglossum) is poorly supported. Three clades of huperzioid Lycopodiaceae were recovered in all analyses of molecular data: Phylloglossum drummondii, Huperzia sensu stricto and Phlegmariurus sensu lato. These clades are strongly supported by morphological characters, including differences in spores, gametophytes, sporophyte macro-morphology, as well as growth habit and life-histories. Our findings indicate that either a one-genus (Huperzia s.l.) or a three-genus (Phylloglossum, Huperzia s.s. and Phlegmariurus s.l.) classification of huperzioid Lycopods are equally supported by molecular evidence, but a two-genus system (Huperzia s.l.+Phylloglossum) is not. We recommend recognising three genera in the huperzioid Lycopodiaceae, as this classification best reflects evolutionary, ecological, and morphological divergence within the lineage.

Characterization of microsatellite loci for an Australian epiphytic orchid, Dendrobium calamiforme, using Illumina sequencing.

PREMISE OF THE STUDY: Microsatellite loci were developed for the epiphytic pencil orchid Dendrobium calamiforme for population genetic and phylogeographic investigation of this Australian taxon. METHODS AND RESULTS: Nineteen microsatellite loci were identified from an Illumina paired-end shotgun library of D. calamiforme. Polymorphism and genetic diversity were assessed in 24 individuals from five populations separated by a maximum distance of ∼80 km. All loci were polymorphic with two to 14 alleles per locus, expected heterozygosity ranging from 0.486 to 0.902, and probability of identity values ranging from 0.018 to 0.380. CONCLUSIONS: These novel markers will serve as valuable tools for investigation of levels of genetic diversity as well as patterns of gene flow, genetic structure, and phylogeographic history.

New and existing combinations in Palaeotropical Phlegmariurus (Lycopodiaceae) and lectotypification of the type species Phlegmariurus phlegmaria (L.) T.Sen & U.Sen.

The genus Phlegmariurus Holubis recognised more widely than originally proposed and is circumscribed to include both Neotropic and Palaeotropic epiphytic and terrestrial species of Huperzioid Lycopodiaceae that have isotomous shoots, lack bulbils in their sporophyllous shoots and have spores with convex lateral margins and foveolate-fossulate sculpture restricted to their distal surfaces. New combinations with Phlegmariurus are proposed for 81 species and existing combinations identified for 33 species originating from the Palaeotropics. This installs a generic circumscription that is consistent between the Neotropics and Palaeotropics. A lectotype is designated for the type species of the genus, Phlegmariurus phlegmaria (L.) T.Sen & U.Sen.

Huperzine alkaloids from Australasian and southeast Asian Huperzia.

CONTEXT: The pharmaceutical alkaloid huperzine A (HupA), currently used in herbal supplements and medicines worldwide, is predominantly sourced from the Chinese lycopod Huperzia serrata (Thunb. ex Murray) Trev. (Lycopodiaceae), which on average contains only 0.08 mg HupA g(-1) dry weight, and is experiencing a rapid decline in China due to over-harvesting. OBJECTIVE: To find a high-yielding, natural source of HupA and/or the related huperzine B (HupB) that could potentially be used as the starting material in a commercial propagation program. MATERIALS AND METHODS: We surveyed 17 Huperzia species (15 indigenous to Australia and southeast Asia) for their foliar HupA and HupB concentrations. We also studied intra-specific variation for the huperzines in four species that were available in sufficient numbers, and determined tissue-specific accumulation in larger specimens. RESULTS: HupA was detected in 11 Australasian and southeast Asian species, with eight also containing HupB, albeit at much lower concentrations. A H. elmeri (Herter) Holub plant from the Philippines had one of the highest HupA concentrations recorded (1.01 mg g(-1) dry wt) and it also had the highest HupB content of all plants surveyed (0.34 mg g(-1) dry wt). Intra-specific HupA and HupB concentrations were extremely variable, and at the intra-plant level, reproductive strobili were found to accumulate the highest HupA concentrations. DISCUSSION AND CONCLUSION: Select Huperzia species from Australia and southeast Asia have potential as the starting material for establishing commercial HupA plantations, but the high intra-specific variability observed suggests that detailed screening is needed to isolate high huperzine-yielding individuals.