Systematics and biogeography of the Old World fern genus Antrophyum.

Antrophyum is one of the largest genera of vittarioid ferns (Pteridaceae) and is most diverse in tropical Asia and the Pacific Islands, but also occurs in temperate Asia, Australia, tropical Africa and the Malagasy region. The only monographic study of Antrophyum was published more than a century ago and a modern assessment of its diversity is lacking. Here, we reconstructed a comprehensively sampled and robustly supported phylogeny for the genus based on four chloroplast markers using Bayesian inference, maximum likelihood and maximum parsimony analyses. We then explored the evolution of the genus from the perspectives of morphology, systematics and historical biogeography. We investigated nine critical morphological characters using a morphometric approach and reconstructed their evolution on the phylogeny. We describe four new species and provide new insight into species delimitation. We currently recognize 34 species for the genus and provide a key to identify them. The results of biogeographical analysis suggest that the distribution of extant species is largely shaped by both ancient and recent dispersal events.

Plastome phylogenomic analysis reveals evolutionary divergences of Polypodiales suborder Dennstaedtiineae.

BACKGROUND: Polypodiales suborder Dennstaedtiineae contain a single family Dennstaedtiaceae, eleven genera, and about 270 species, and include some groups that were previously placed in Dennstaedtiaceae, Hypolepidaceae, Monachosoraceae, and Pteridaceae. The classification and phylogenetic relationships among these eleven genera have been poorly understood. To explore the deep relationships within suborder Dennstaedtiineae and estimate the early diversification of this morphologically heterogeneous group, we analyzed complete plastomes of 57 samples representing all eleven genera of suborder Dennstaedtiineae using maximum likelihood and Bayesian inference. RESULTS: The phylogenetic relationships of all the lineages in the bracken fern family Dennstaedtiaceae were well resolved with strong support values. All six genera of Hypolepidoideae were recovered as forming a monophyletic group with full support, and Pteridium was fully supported as sister to all the other genera in Hypolepidoideae. Dennstaedtioideae (Dennstaedtia s.l.) fell into four clades with full support: the Microlepia clade, the northern Dennstaedtia clade, the Dennstaedtia globulifera clade, and the Dennstaedtia s.s. clade. Monachosorum was strongly resolved as sister to all the remaining genera of suborder Dennstaedtiineae. Based on the well resolved relationships among genera, the divergence between Monachosorum and other groups of suborder Dennstaedtiineae was estimated to have occurred in the Early Cretaceous, and all extant genera (and clades) in Dennstaedtiineae, were inferred to have diversified since the Late Oligocene. CONCLUSION: This study supports reinstating a previously published family Monachosoraceae as a segregate from Dennstaedtiaceae, based on unique morphological evidence, the shady habitat, and the deep evolutionary divergence from its closest relatives.

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.

Genetic and morphological identification of a recurrent Dicksonia tree fern hybrid in New Zealand.

Hybridization is common in many ferns and has been a significant factor in fern evolution and speciation. However, hybrids are rare between the approximately 30 species of Dicksonia tree ferns world-wide, and none are well documented. In this study we examine the relationship of a newly-discovered Dicksonia tree fern from Whirinaki, New Zealand, which does not fit the current taxonomy of the three species currently recognized in New Zealand. Our microsatellite genotyping and ddRAD-seq data indicate these plants are F1 hybrids that have formed multiple times between D. fibrosa and D. lanata subsp. lanata. The Whirinaki plants have intermediate morphology between D. fibrosa and D. lanata subsp. lanata and their malformed spores are consistent with a hybrid origin. The Whirinaki plants-Dicksonia fibrosa × D. lanata subsp. lanata-are an example of hybridization between distantly related fern lineages, with the two parent species estimated to have diverged 55-25 mya. Our chloroplast sequencing indicates asymmetric chloroplast inheritance in the Whirinaki morphology with D. lanata subsp. lanata always contributing the chloroplast genome.

Microsatellite DNA analyses of a highly disjunct New Zealand tree reveal strong differentiation and imply a formerly more continuous distribution.

Although New Zealand is a biodiversity hotspot, there has been little genetic investigation of why so many of its threatened and uncommon plants have naturally disjunct distributions. We investigated the small tree Pseudopanax ferox (Araliaceae), which has a widespread but highly disjunct lowland distribution within New Zealand. Genotyping of nuclear microsatellites and a chloroplast locus revealed pronounced genetic differentiation and four principal genetic clusters. Our results indicate that the disjunct distribution is a product of vicariance rather than long-distance dispersal. This highlights the need to preserve multiple populations when disjunct distributions are the result of vicariance, rather than focusing conservation efforts on a core area, in order to retain as much as possible of a species' evolutionary legacy and potential. Additionally, based on our genetic findings and the ecology of P. ferox, we hypothesize that it was more continuously distributed during the drier (but not maximally colder) interstadials of glacial periods and/or on the fertile soils available immediately postglacial. We further hypothesize that P. ferox belongs to a suite of species of drought-prone and/or fertile habitats whose distributions are actually restricted during warmer and wetter interglacial periods, despite being principally of the lowlands. Our genetic data for P. ferox are also the first consistent with the survival during the Last Glacial Maxima of a lowland tree at high latitudes in the south-eastern South Island.

Parallel polyploid speciation: distinct sympatric gene-pools of recurrently derived allo-octoploid Asplenium ferns.

Although polyploidy is widespread, its significance to the generation of biodiversity remains unclear. Many polyploids have been derived recurrently. For a particular polyploid, gene-flow between the products of independent origin is typical where they come into contact. Here, we use AFLP DNA-fingerprinting and chloroplast DNA sequences to demonstrate parallel polyploid speciation within both of the ferns Asplenium cimmeriorum and A. gracillimum. Both of these taxa comprise at least two allopolyploids, recurrently derived from the same progenitor pair. Each of these allopolyploids remain genetically distinguishable even with extensive sympatry, and could therefore be considered distinct species. To our knowledge, parallel speciation on this scale amongst recurrent polyploids has not been previously reported. With their parallel origins, these 'evolutionary replicates' provide an unrivalled opportunity to investigate how the reproductive barriers and ecological differentiation necessary for speciation arise following polyploidy.

Reconstructing the species phylogeny of Pseudopanax (Araliaceae), a genus of hybridising trees.

Pseudopanax (Araliaceae) comprises 12 tree species of diverse morphology and ecology, and is endemic to New Zealand. It is notable for the hybridisation that occurs between P. crassifolius and P. lessonii, which have very different leaves and habits. To provide context for the study of this hybridisation and other investigations, we examined the phylogeny of Pseudopanax using chloroplast DNA sequences (c.5900 base-pairs) and AFLP DNA-fingerprinting. Both approaches resolve two principal groups within Pseudopanax--the Arboreus group and the Crassifolius+Lessonii union--but how they are related to other genera remains unclear. There is, nevertheless, little compelling evidence against the monophyly of Pseudopanax, making unnecessary the recent re-segregation of the Arboreus group as Neopanax. The chloroplast data provided minimal additional resolution, although the position of P. linearis was discordant compared to other data. Analyses of the AFLP data strongly recovered each species, aside from the morphologically heterogeneous P. colensoi, and the two mainland species (P. arboreus and P. crassifolius) that each contained a nested island-endemic (P. kermadecensis and P. chathamicus, respectively). However, relationships amongst species within the two principal groups were poorly resolved. An example was the uncertainty of whether P. crassifolius grouped with P. lessonii and its allies, or the morphologically similar species it had been previously placed with. This in turn raises the issue of how hybridisation might affect phylogenies and the ability to reconstruct them, even when using multiple, independent markers.

Low-copy nuclear DNA sequences reveal a predominance of allopolyploids in a New Zealand Asplenium fern complex.

Recent generalisations about polyploidy in plants have been largely based on studies of angiosperms. A compelling group to compare with angiosperms is ferns, because of their high polyploidy. The bi-parental inheritance of nuclear DNA sequence markers makes them advantageous for investigating polyploid complexes, but few such markers have been available for ferns. We have used DNA sequences from the low-copy nuclear LFY locus to study an Asplenium polyploid complex. The New Zealand species of this Austral group comprise seven tetraploids and eight octoploids. LFY sequences indicate that allopolyploidy is much more predominant than previously thought, being implicated in the origins of seven of the octoploids. One of the tetraploids has had a central role, being a progenitor for five of the octoploids. All of the octoploids appear to have relatively recent origins, with the dynamic environmental conditions of the Pleistocene possibly playing a role in their formation and/or establishment.

Conflict amongst chloroplast DNA sequences obscures the phylogeny of a group of Asplenium ferns.

A previous study of the relationships amongst three subgroups of the Austral Asplenium ferns found conflicting signal between the two chloroplast loci investigated. Because organelle genomes like those of chloroplasts and mitochondria are thought to be non-recombining, with a single evolutionary history, we sequenced four additional chloroplast loci with the expectation that this would resolve these relationships. Instead, the conflict was only magnified. Although tree-building analyses favoured one of the three possible trees, one of the alternative trees actually had one more supporting site (six versus five) and received greater support in spectral and neighbor-net analyses. Simulations suggested that chance alone was unlikely to produce strong support for two of the possible trees and none for the third. Likelihood permutation tests indicated that the concatenated chloroplast sequence data appeared to have experienced recombination. However, recombination between the chloroplast genomes of different species would be highly atypical, and corollary supporting observations, like chloroplast heteroplasmy, are lacking. Wider taxon sampling clarified the composition of the Austral group, but the conflicting signal meant analyses (e.g., morphological evolution, biogeographic) conditional on a well-supported phylogeny could not be performed.

Fire and ice: volcanic and glacial impacts on the phylogeography of the New Zealand forest fern Asplenium hookerianum.

In the Southern Hemisphere there has been little phylogeographical investigation of forest refugia sites during the last glacial. Hooker's spleenwort, Asplenium hookerianum, is a fern that is found throughout New Zealand. It is strongly associated with forest and is a proxy for the survival of woody vegetation during the last glacial maximum. DNA sequence data from the chloroplast trnL-trnF locus were obtained from 242 samples, including c. 10 individuals from each of 21 focal populations. Most populations contained multiple, and in many cases unique, haplotypes, including those neighbouring formerly glaciated areas, while the predominant inference from nested clade analysis was restricted gene flow with isolation by distance. These results suggest that A. hookerianum survived the last glacial maximum in widespread populations of sufficient size to retain the observed phylogeography, and therefore that the sheltering woody vegetation must have been similarly abundant. This is consistent with palynological interpretations for the survival in New Zealand of thermophilous forest species at considerably smaller distances from the ice sheets than recorded for the Northern Hemisphere. Eastern and central North Island populations of A. hookerianum were characterized by a different subset of haplotypes to populations from the remainder of the country. A similar east-west phylogeographical pattern has been detected in a diverse array of taxa, and has previously been attributed to recurrent vulcanism in the central North Island.

Genetic variation is not concordant with morphological variation in the fern Asplenium hookerianum sensu lato (Aspleniaceae).

Species status cannot be adequately determined when partitions are based on only a single morphological character. For instance, the sympatry of plants with broad and narrow pinnules in the fern Asplenium hookerianum sensu lato from New Zealand creates the impression that two entities are present. The narrow-pinnuled plants are sometimes segregated as a distinct species, A. colensoi. However, this variation in pinnule morphology could equally be infraspecific, and only additional data can resolve this uncertainty. Analyses using AFLP DNA-fingerprinting and DNA sequencing of the chloroplast trnL-trnF region indicate that genetic variation in A. hookerianum sensu lato is not concordant with pinnule morphology. Consequently, the recognition of A. colensoi is not supported.