Lara D. Shepherd

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. Hookers 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.

A molecular phylogeny for the New Zealand Blechnaceae ferns from analyses of chloroplast trnL‐trnF DNA sequences

Abstract The Blechnaceae is one of the most speciose fern families in New Zealand, with two genera represented: Blechnum and Doodia. We se‐quenced the chloroplast trnL‐trnF locus for all of the Blechnaceae species indigenous to New Zealand, plus several non‐indigenous species. Although deeper relationships were not well resolved by phylogenetic analyses of these DNA sequences, several groupings of species were consistently recovered. Some of these relationships have been previously suspected on the basis of morphological similarity and/or hybridisation (e.g., the B. procerum group), and are consistent with variation in base chromosome numbers, but others were unexpected (e.g., the relationship of B. fluviatile and B. vulcanicum). The species of Doodia sampled here were found to be monophyletic, and were nested within a paraphyletic Blechnum. Infraspecific variation in the trnL‐trnF locus was detected within six New Zealand species, and may prove useful for future phylogeographic and taxonomic studies.

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.

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.

Phylogeny of the fern family Aspleniaceae in Australasia and the south-western Pacific

Abstract. Aspleniaceae is one of the largest fern families. It is species-rich in Australasia and the south-western Pacific (ASWP), where approximately 115 species occur. In the current study, the chloroplast regions rbcL, trnL–trnF and rps4–trnS were sequenced for 100 Aspleniaceae samples from ASWP. These data were combined with published sequences for species from New Zealand and other regions for phylogenetic analyses. Species of Aspleniaceae from ASWP were placed in six of the eight previously identified inter-continental clades. The majority of species from ASWP were placed in two of these clades, with the remaining four clades each being represented by three or fewer species. Strong biogeographic affinities with South-east Asia were observed and immigration, rather than local radiations of endemic taxa, appears to have made a more important contribution to patterns of diversity in ASWP. This study supports the current taxonomic practice of recognising two genera, Asplenium L. and Hymenasplenium Hayata, in Aspleniaceae, and identifies future taxonomic work required for the family in this region, including potential synonymising of species, and revision of species complexes or widespread species that are demonstrably non-monophyletic.

An expanded phylogeny of the Dennstaedtiaceae ferns: Oenotrichia falls within a non-monophyletic Dennstaedtia, and Saccoloma is polyphyletic

Abstract. Recent studies have transferred several species previously attributed to the fern genus Oenotrichia to other genera, and even out of the Dennstaedtiaceae to other families. However, the relationship of the type species, O. maxima from New Caledonia, has not previously been investigated using DNA sequences. With phylogenetic analyses of chloroplast DNA sequences, we verify the placement of Oenotrichia within the Dennstaedtiaceae. Moreover, O. maxima actually nests along with Leptolepia in a clade of Dennstaedtia. Dennstaedtia itself is non-monophyletic, with a second clade being more closely related to Microlepia. We outline what is required to resolve the generic taxonomy of this group. We also find that samples attributed to Saccoloma are polyphyletic, with some falling inside the Dennstaedtiaceae and others outside.

Gleichenia inclusisora, a new and uncommon tangle fern from New Zealand

Abstract Gleichenia inclusisora is described here as a new and uncommon fern endemic to New Zealand. Several morphological features readily distinguish G. inclusisora from other Gleichenia species, and in the context of New Zealand species these features include: sori embedded in the lamina; bicolorous, orbicular to ovate, peltate scales on abaxial surfaces of the ultimate segments; glabrous stipes and glabrescent proximal costae; and unpouched and abaxially white ultimate segments. Phylogenetic analyses of New Zealand species using chloroplast DNA sequences support the distinctiveness of G. inclusisora. It is recorded from four disjunct regions, but no extant population is known for one of these. We suggest that G. inclusisora be treated in the threat classification system as Naturally Uncommon, with the Range Restricted qualifier.

Evidence of a Strong Domestication Bottleneck in the Recently Cultivated New Zealand Endemic Root Crop, Arthropodium cirratum (Asparagaceae)

We use chloroplast DNA sequencing to examine aspects of the pre-European Māori cultivation of an endemic New Zealand root crop, Arthropodium cirratum (rengarenga). Researching the early stages of domestication is not possible for the majority of crops, because their cultivation began many thousands of years ago and/or they have been substantially altered by modern breeding methods. We found high levels of genetic variation and structuring characterised the natural distribution of A. cirratum, while the translocated populations only retained low levels of this diversity, indicating a strong bottleneck even at the early stages of this species’ cultivation. The high structuring detected at four chloroplast loci within the natural A. cirratum range enabled the putative source(s) of the translocated populations to be identified as most likely located in the eastern Bay of Plenty/East Cape region. The high structuring within A. cirratum also has implications for the conservation of genetic diversity within this species, which has undergone recent declines in both its natural and translocated ranges.

A molecular investigation into the origin and relationships of karaka/kōpi (Corynocarpus laevigatus) in New Zealand

Corynocarpus is a small, morphologically and phylogenetically isolated genus of trees from the southwest Pacific. We examined the phylogeny of Corynocarpus using the nuclear WAXY and ITS loci. Our molecular dating, when examined in combination with other published molecular dating studies, indicates that the origin of the genus and the arrival of Corynocarpus in New Zealand are likely to be much more recent than previously suggested. A second focus of our study was to investigate patterns of pre-European putative translocation of C. laevigatus (karaka, kōpi) by Māori throughout New Zealand using diversity at these two loci. Our New Zealand-wide sampling revealed the distinctiveness of specimens from the Three Kings Islands at both loci, precluding the Three Kings Islands as the source population for translocated C. laevigatus. However, the low level of sequence variation at these loci prevented further insight into the human-mediated dispersal of this species.

Investigation of species boundaries and relationships in the Asplenium paleaceum complex (Aspleniaceae) using AFLP fingerprinting and chloroplast and nuclear DNA sequences

Abstract. Species boundaries and relationships were investigated in the Asplenium paleaceum (Aspleniaceae) species complex from eastern Australia, using AFLP fingerprinting and chloroplast trnL–trnF and rps4–trnS and nuclear pgiC DNA sequences. Phenetic analyses of AFLP data resulted in the following five distinct groups: (1) A. carnarvonense, (2) A. bicentenniale (including nearby collections originally identified as A. paleaceum), (3) A. paleaceum with both aborted and normal spores, (4) one population of putative tetraploid A. attenuatum var. indivisum from south Queensland, and (5) remaining octoploid A. attenuatum populations and several putative hybrids. Taxonomic revision of this complex will require morphological re-circumscription of the current species and recognition of a new species if these AFLP groups are taken to represent separate species. The chloroplast regions, morphology and pgiC together provide good evidence that an Asplenium of unconfirmed identity, A. sp. ‘Kroombit Tops’, is an allopolyploid with a species of the A. paleaceum chloroplast clade, probably A. paleaceum, and distantly related A. polyodon as parents. Further study is required to determine the complete ancestry of the other species of the A. paleaceum complex.