Crossroads of assembling a moss genome: navigating contaminants and horizontal gene transfer in the moss Physcomitrellopsis africana.

The first chromosome-scale reference genome of the rare narrow-endemic African moss Physcomitrellopsis africana is presented here. Assembled from 73x nanopore long reads and 163x BGI-seq short reads, the 414 Mb reference comprises 26 chromosomes and 22,925 protein-coding genes (BUSCO: C:94.8%[D:13.9%]). This genome holds two genes that withstood rigorous filtration of microbial contaminants, have no homolog in other land plants and are thus interpreted as resulting from two unique horizontal gene transfers from microbes. Further, Physcomitrellopsis africana shares 176 of the 273 published HGT candidates identified in Physcomitrium patens, but lacks 98 of these, highlighting that perhaps as many as 91 genes were acquired in P. patens in the last 40 million years following its divergence from its common ancestor with P. africana. These observations suggest rather continuous gene gains via HGT followed by potential losses, during the diversification of the Funariaceae. Our findings showcase both dynamic flux in plant HGTs over evolutionarily "short" timescales, alongside enduring impacts of successful integrations, like those still functionally maintained in extant Physcomitrellopsis africana. Furthermore, this study describes the informatic processes employed to distinguish contaminants from candidate HGT events.

Evolutionary dynamism in bryophytes: Phylogenomic inferences confirm rapid radiation in the moss family Funariaceae.

Rapid diversifications of plants are primarily documented and studied in angiosperms, which are perceived as evolutionarily dynamic. Recent studies have, however, revealed that bryophytes have also undergone periods of rapid radiation. The speciose family Funariaceae, including the model taxon Physcomitrella patens, is one such lineage. Here, we infer relationships among major lineages within the Entosthodon-Physcomitrium complex from virtually complete organellar exomes (i.e., 123 genes) obtained through high throughput sequencing of genomic libraries enriched in these loci via targeted locus capture. Based on these extensive exonic data we (1) reconstructed a robust backbone topology of the Funariaceae, (2) confirmed the monophyly of Funaria and the polyphyly of Entosthodon, Physcomitrella, and Physcomitrium, and (3) argue for the occurrence of a rapid radiation within the Entosthodon-Physcomitrium complex that began 28 mya and gave rise more than half of the species diversity of the family. This diversification may have been triggered by a whole genome duplication and coincides with global Eocene cooling that continued through the Oligocene and Miocene. The Funariaceae join a growing list of bryophyte lineages whose history is marked by at least one burst of diversification, and our study thereby strengthens the view that bryophytes are evolutionarily dynamic lineages and that patterns and processes characterizing the evolution of angiosperms may be universal among land plants.

Topography as a driver of cryptic speciation in the high-elevation cape sedge Tetraria triangularis (Boeck.) C. B. Clarke (Cyperaceae: Schoeneae).

Since some speciation mechanisms are more likely to generate morphological disparity than others, the general failure of vascular plant taxonomists to recognize cryptic diversity may bias perceptions about speciation process in plants. While the exceptional floristic richness of the South African Cape has largely been attributed to adaptive divergence ('ecological' speciation), a combination of climatic dynamism and complex topography has likely provided ample opportunities for 'non-ecological' vicariant speciation, a mechanism which is perhaps more likely to produce cryptic species. We explore the role of topography as a driver of 'non-ecological' speciation in the high-elevation sedge Tetraria triangularis. Within this species, molecular and morphological data reveal five cryptic or semi-cryptic lineages of Miocene-Pliocene age which qualify as evolutionary species. At least three of these maintain their distinctness in sites of sympatry, identifying them as biological species. Negligible range overlap, and the identification of topography as a significant predictor of range turnover, identifies speciation as allopatric and a result of impeded gene flow across low-elevation topographic features. Weak morphological and ecological divergence implies a limited role for adaptive divergence in powering speciation, with character displacement in sympatry possibly arising as a consequence of interspecific competition. Although we cannot exclude a role for disruptive selection in species differentiation, we identify isolation of populations on topographically separated mountains as the principal motor of speciation. We suggest that the importance of topography in the genesis of Cape floristic diversity has been inadequately acknowledged.

Constructing phylogenies in the presence of intra-individual site polymorphisms (2ISPs) with a focus on the nuclear ribosomal cistron.

Nuclear DNA is widely used to estimate phylogenetic and phylogeographic relationships. Nuclear gene variants may be present in an individual's genome, and these result in Intra-Individual Site Polymorphisms (2ISP; pronounced "twisp") in direct-PCR or individual-consensus sequences based on a sample of clones or fragment sequences from next generation sequencing (NGS). 2ISPs can occur fairly often, especially within, but not restricted to, high-copy-number regions such as the widely used internal transcribed spacers of the nuclear ribosomal cistron. Dealing with 2ISPs has been problematic as phylogeny reconstruction optimality criteria generally do not take account of this variation. Here we test whether an approach that treats 2ISPs as additional (termed "informative"), rather than ambiguous, characters offers improved support in three common criteria used for phylogenetic inference: Minimum Evolution (via Neighbour Joining), Maximum Parsimony, and Maximum Likelihood. We demonstrate significant improvements using the 2ISP-informative treatment with simulated, real-world, and case-study data sets. We envisage that this 2ISP-informative approach will greatly aid phylogenetic inference using any nuclear DNA regions that contain polymorphisms within individuals (including consensus sequences generated from NGS), especially at the intrageneric or intraspecific level.

Preserving the evolutionary potential of floras in biodiversity hotspots.

One of the biggest challenges for conservation biology is to provide conservation planners with ways to prioritize effort. Much attention has been focused on biodiversity hotspots. However, the conservation of evolutionary process is now also acknowledged as a priority in the face of global change. Phylogenetic diversity (PD) is a biodiversity index that measures the length of evolutionary pathways that connect a given set of taxa. PD therefore identifies sets of taxa that maximize the accumulation of 'feature diversity'. Recent studies, however, concluded that taxon richness is a good surrogate for PD. Here we show taxon richness to be decoupled from PD, using a biome-wide phylogenetic analysis of the flora of an undisputed biodiversity hotspot--the Cape of South Africa. We demonstrate that this decoupling has real-world importance for conservation planning. Finally, using a database of medicinal and economic plant use, we demonstrate that PD protection is the best strategy for preserving feature diversity in the Cape. We should be able to use PD to identify those key regions that maximize future options, both for the continuing evolution of life on Earth and for the benefit of society.

Phylogenetics, biogeography and classification of, and character evolution in, gamebirds (Aves: Galliformes): effects of character exclusion, data partitioning and missing data.

The phylogenetic relationships, biogeography and classification of, and morpho-behavioral (M/B) evolution in, gamebirds (Aves: Galliformes) are investigated. In-group taxa (rooted on representatives of the Anseriformes) include 158 species representing all suprageneric galliform taxa and 65 genera. The characters include 102 M/B attributes and 4452 nucleic acid base pairs from mitochondrial cytochrome b (CYT B), NADH dehydrogenase subunit 2 (ND2), 12S ribosomal DNA (12S) and control region (CR), and nuclear ovomucoid intron G (OVO-G). Analysis of the combined character data set yielded a single, completely resolved cladogram that had the highest levels of jackknife support, which suggests a need for a revised classification for the phasianine galliforms. Adding 102 M/B characters to the combined CYT B and ND2 partitions (2184 characters) decisively overturns the topology suggested by analysis of the two mtDNA partitions alone, refuting the view that M/B characters should be excluded from phylogenetic analyses because of their relatively small number and putative character state ambiguity. Exclusion of the OVO-G partition (with > 70% missing data) from the combined data set had no effect on cladistic structure, but slightly lowered jackknife support at several nodes. Exclusion of third positions of codons in an analysis of a CYT B + ND2 partition resulted in a massive loss of resolution and support, and even failed to recover the monophyly of the Galliformes with jackknife support. A combined analysis of putatively less informative, "non-coding" characters (CYT B/ND2 third position sites + CR +12S + OVO-G sequences) yielded a highly resolved consensus cladogram congruent with the combined-evidence cladogram. Traditionally recognized suprageneric galliform taxa emerging in the combined cladogram are: the families Megapodiidae (megapodes), Cracidae (cracids), Numididae (guineafowls), Odontophoridae (New World quails) and Phasianidae (pheasants, pavonines, partridges, quails, francolins, spurfowls and grouse) and the subfamilies Cracinae (curassows, chachalacas and the horned guan), Penelopinae (remaining guans), Pavoninae sensu lato (peafowls, peacock pheasants and argus pheasants), Tetraoninae (grouse) and Phasianinae (pheasants minus Gallus). The monophyly of some traditional groupings (e.g., the perdicinae: partridges/quails/francolins) is rejected decisively, contrasted by the emergence of other unexpected groupings. The most remarkable phylogenetic results are the placement of endemic African galliforms as sisters to geographically far-distant taxa in Asia and the Americas. Biogeographically, the combined-data cladogram supports the hypothesis that basal lineages of galliforms diverged prior to the Cretaceous/Tertiary (K-T) Event and that the subsequent cladogenesis was influenced by the break-up of Gondwana. The evolution of gamebirds in Africa, Asia and the Americas has a far more complicated historical biogeography than suggested to date. With regard to character evolution: spurs appear to have evolved at least twice within the Galliformes; a relatively large number of tail feathers (≥ 14) at least three times; polygyny at least twice; and sexual dimorphism many times.

Phylogeny of the Polytrichales (Bryophyta) based on simultaneous analysis of molecular and morphological data.

Phylogenetic analyses of Polytrichales were conducted using morphology and sequence data from the chloroplast genes rbcL and rps4 plus the trnL-F gene region, part of the mitochondrial nad5 and the nuclear-encoded 18S rDNA. Our analyses included 46 species representing all genera of Polytrichales. Phylogenetic trees were constructed with simultaneous parsimony analyses of all sequences plus morphology and separate combinations of sequence data only. Results lend support for recognition of Polytrichales as a monophyletic entity. Oedipodium griffithianum appears as a sister taxon to Polytrichales or as a sister taxon of all mosses excluding Sphagnales and Andreaeles. Within Polytrichales, Alophosia and Atrichopsis, species without the adaxial lamellae (in Atrichopsis present but poorly developed on male gametophyte) otherwise typical of the group are sister to the remaining species followed by a clade including Bartramiopsis and Lyellia, species with adaxial lamellae covering only the central portion of the leaves. Six taxa with an exclusively Southern Hemisphere distribution form a grade between the basal lineages and a clade including genera that are mostly confined to the Northern Hemisphere.

The Bryophyta (Mosses): Systematic and Evolutionary Inferences from an rps4 Gene (cpDNA) Phylogeny.

Phylogenetic analyses of nucleotide and amino acid sequences of the chloroplast protein coding gene rps4 were performed for 225 species of mosses, representing 84% of families recognized by Vitt (1984. In: Schuster RM, ed. New manual of bryology , vol 2 . Nichinan: Hattori Botanical Laboratory), under the criterion of maximum parsimony with Takakia and Sphagnum as outgroups. Most parsimonious topologies converge to a scenario wherein the Andreaeidae are monophyletic and sister to the Bryidae (peristomate mosses), the Nematodonteae and the Buxbaumiaceae form a monophyletic lineage, the Diphysciaceae are sister to the Arthrodonteae and, within the latter, the Funarineae-Encalyptineae-Timmiaceae-Haplolepideae compose a monophyletic clade sister to remaining diplolepideous mosses. This hypothesis suggests that early in the evolution of the Arthrodonteae, two major lineages diverged, with opposite and alternate peristomes, respectively. Bootstrap support for the deep dichotomies is poor or lacking but increases when protein translations of rps 4 sequences are included in the analysis. Several novel systematic hypotheses are raised, including ( a ) a diplolepideous rather than haplolepideous origin of the Pleurophascaceae; ( b ) an affinity of the Catascopiaceae with the Funariineae rather than the Bryineae; and ( c ) a close relationship of the Calomniaceae and Mitteniaceae to the Rhizgoniaceae. The advantages and disadvantages of a single gene phylogeny are discussed with respect to the identification of polyphyletic familial or suprafamilial taxa.