Browsing by Author "Windham, MD"
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Item Open Access A molecular phylogeny of the fern family Pteridaceae: assessing overall relationships and the affinities of previously unsampled genera.(Molecular phylogenetics and evolution, 2007-09) Schuettpelz, E; Schneider, H; Huiet, L; Windham, MD; Pryer, KMThe monophyletic Pteridaceae accounts for roughly 10% of extant fern diversity and occupies an unusually broad range of ecological niches, including terrestrial, epiphytic, xeric-adapted rupestral, and even aquatic species. In this study, we present the results of the first broad-scale and multi-gene phylogenetic analyses of these ferns, and determine the affinities of several previously unsampled genera. Our analyses of two newly assembled data sets (including 169 newly obtained sequences) resolve five major clades within the Pteridaceae: cryptogrammoids, ceratopteridoids, pteridoids, adiantoids, and cheilanthoids. Although the composition of these clades is in general agreement with earlier phylogenetic studies, it is very much at odds with the most recent subfamilial classification. Of the previously unsampled genera, two (Neurocallis and Ochropteris) are nested within the genus Pteris; two others (Monogramma and Rheopteris) are early diverging vittarioid ferns, with Monogramma resolved as polyphyletic; the last previously unsampled genus (Adiantopsis) occupies a rather derived position among cheilanthoids. Interestingly, some clades resolved within the Pteridaceae can be characterized by their ecological preferences, suggesting that the initial diversification in this family was tied to ecological innovation and specialization. These processes may well be the basis for the diversity and success of the Pteridaceae today.Item Open Access A plastid phylogeny of the cosmopolitan fern family cystopteridaceae (Polypodiopsida)(Systematic Botany, 2013-06-01) Rothfels, CJ; Windham, MD; Pryer, KMAmong the novel results of recent molecular phylogenetic analyses are the unexpectedly close evolutionary relationships of the genera Acystopteris, Cystopteris, and Gymnocarpium, and the phylogenetic isolation of these genera from Woodsia. As a consequence, these three genera have been removed from Woodsiaceae and placed into their own family, the Cystopteridaceae. Despite the ubiquity of this family in rocky habitats across the northern hemisphere, and its cosmopolitan distribution (occurring on every continent except Antarctica), sampling of the Cystopteridaceae in phylogenetic studies to date has been sparse. Here we assemble a three-locus plastid dataset (matK, rbcL, trnG-R) that includes most recognized species in the family and multiple accessions of widespread taxa from across their geographic ranges. All three sampled genera are robustly supported as monophyletic, Cystopteris is strongly supported as sister to Acystopteris, and those two genera together are sister to Gymnocarpium. The Gymnocarpium phylogeny is deeply divided into three major clades, which we label the disjunctum clade, the robertianum clade, and core Gymnocarpium. The Cystopteris phylogeny, similarly, features four deeply diverged clades: C. montana, the sudetica clade, the bulbifera clade, and the fragilis complex. Acystopteris includes only three species, each of which is supported as monophyletic, with A. taiwaniana sister to the japonica/tenuisecta clade. Our results yield the first species-level phylogeny of the Cystopteridaceae and the first molecular phylogenetic evidence for species boundaries. These data provide an essential foundation for further investigations of complex patterns of geographic diversification, speciation, and reticulation in this family. © Copyright 2013 by the American Society of Plant Taxonomists.Item Open Access A step-by-step protocol for meiotic chromosome counts in flowering plants: A powerful and economical technique revisited.(Applications in plant sciences, 2020-04-23) Windham, MD; Pryer, KM; Poindexter, DB; Li, F; Rothfels, CJ; Beck, JBPremise:Counting chromosomes is a fundamental botanical technique, yet it is often intimidating and increasingly sidestepped. Once mastered, the basic protocol can be applied to a broad range of taxa and research questions. It also reveals an aspect of the plant genome that is accessible with only the most basic of resources-access to a microscope with 1000× magnification is the most limiting factor. Methods and Results:Here we provide a detailed protocol for choosing, staining, and squashing angiosperm pollen mother cells. The protocol is supplemented by figures and two demonstration videos. Conclusions:The protocol we provide will hopefully demystify and reinvigorate a powerful and once commonplace botanical technique that is available to researchers regardless of their location and resources.Item Open Access A Unified Approach to Taxonomic Delimitation in the Fern Genus Pentagramma (Pteridaceae)(Systematic Botany, 2015-01-01) Schuettpelz, E; Pryer, KM; Windham, MD© Copyright 2015 by the American Society of Plant Taxonomists.The native goldback and silverback ferns of western North America, composing the genus Pentagramma, are phylogenetically isolated within the xeric-adapted cheilanthoid clade. Although species-poor compared to its sister group, Pentagramma encompasses a diverse array of morphotypes, cytotypes, and flavonoid chemotypes. Because the differences are generally cryptic, however, the various entities are usually recognized at an infraspecific level. In recent years, as many as five subspecies have been ascribed to P. triangularis, and only P. pallida has been considered sufficiently divergent to warrant recognition as a distinct species. In this study, we take a unified approach to taxonomic delimitation in Pentagramma. Combining spore studies with phylogenetic analyses of plastid and nuclear sequences, we identify six genetically and morphologically distinct diploid lineages, each of which is here treated as a species. A new species is described (P. glanduloviscida) and three new combinations are made (P. maxonii, P. rebmanii, and P. viscosa).Item Open Access Are there too many fern genera?(Taxon, 2018-06-01) Schuettpelz, E; Rouhan, G; Pryer, KM; Rothfels, CJ; Prado, J; Sundue, MA; Windham, MD; Moran, RC; Smith, ARItem Open Access Baja: A New Monospecific Genus Segregated from Cheilanthes s. l. (Pteridaceae)(Systematic Botany, 2019-09-06) George, LO; Pryer, KM; Kao, T; Huiet, L; Windham, MDItem Open Access Deciphering the origins of apomictic polyploids in the Cheilanthes yavapensis complex (Pteridaceae).(American journal of botany, 2009-09) Grusz, AL; Windham, MD; Pryer, KMDeciphering species relationships and hybrid origins in polyploid agamic species complexes is notoriously difficult. In this study of cheilanthoid ferns, we demonstrate increased resolving power for clarifying the origins of polyploid lineages by integrating evidence from a diverse selection of biosystematic methods. The prevalence of polyploidy, hybridization, and apomixis in ferns suggests that these processes play a significant role in their evolution and diversification. Using a combination of systematic approaches, we investigated the origins of apomictic polyploids belonging to the Cheilanthes yavapensis complex. Spore studies allowed us to assess ploidy levels; plastid and nuclear DNA sequencing revealed evolutionary relationships and confirmed the putative progenitors (both maternal and paternal) of taxa of hybrid origin; enzyme electrophoretic evidence provided information on genome dosage in allopolyploids. We find here that the widespread apomictic triploid, Cheilanthes lindheimeri, is an autopolyploid derived from a rare, previously undetected sexual diploid. The apomictic triploid Cheilanthes wootonii is shown to be an interspecific hybrid between C. fendleri and C. lindheimeri, whereas the apomictic tetraploid C. yavapensis is comprised of two cryptic and geographically distinct lineages. We show that earlier morphology-based hypotheses of species relationships, while not altogether incorrect, only partially explain the complicated evolutionary history of these ferns.Item Open Access Do asexual polyploid lineages lead short evolutionary lives? A case study from the fern genus Astrolepis.(Evolution; international journal of organic evolution, 2011-11) Beck, JB; Windham, MD; Pryer, KMA life-history transition to asexuality is typically viewed as leading to a heightened extinction risk, and a number of studies have evaluated this claim by examining the relative ages of asexual versus closely related sexual lineages. Surprisingly, a rigorous assessment of the age of an asexual plant lineage has never been published, although asexuality is extraordinarily common among plants. Here, we estimate the ages of sexual diploids and asexual polyploids in the fern genus Astrolepis using a well-supported plastid phylogeny and a relaxed-clock dating approach. The 50 asexual polyploid samples we included were conservatively estimated to comprise 19 distinct lineages, including a variety of auto- and allopolyploid genomic combinations. All were either the same age or younger than the crown group comprising their maternal sexual-diploid parents based simply on their phylogenetic position. Node ages estimated with the relaxed-clock approach indicated that the average maximum age of asexual lineages was 0.4 My, and individual lineages were on average 7 to 47 times younger than the crown- and total-ages of their sexual parents. Although the confounding association between asexuality and polyploidy precludes definite conclusions regarding the effect of asexuality, our results suggest that asexuality limits evolutionary potential in Astrolepis.Item Open Access Evidence for reciprocal origins in Polypodium hesperium (Polypodiaceae): a fern model system for investigating how multiple origins shape allopolyploid genomes.(American journal of botany, 2014-09-17) Sigel, EM; Windham, MD; Pryer, KM•Many polyploid species are composed of distinct lineages originating from multiple, independent polyploidization events. In the case of allopolyploids, reciprocal crosses between the same progenitor species can yield lineages with different uniparentally inherited plastid genomes. While likely common, there are few well-documented examples of such reciprocal origins. Here we examine a case of reciprocal allopolyploid origins in the fern Polypodium hesperium and present it as a natural model system for investigating the evolutionary potential of duplicated genomes.•Using a combination of uniparentally inherited plastid and biparentally inherited nuclear sequence data, we investigated the distributions and relative ages of reciprocally formed lineages in Polypodium hesperium, an allotetraploid fern that is broadly distributed in western North America.•The reciprocally derived plastid haplotypes of Polypodium hesperium are allopatric, with populations north and south of 42°N latitude having different plastid genomes. Incorporating biogeographic information and previously estimated ages for the diversification of its diploid progenitors, we estimate middle to late Pleistocene origins of P. hesperium.•Several features of Polypodium hesperium make it a particularly promising system for investigating the evolutionary consequences of allopolyploidy. These include reciprocally derived lineages with disjunct geographic distributions, recent time of origin, and extant diploid progenitors.Item Open Access Exploring Past and Future Distributions of the Rare Appalachian Oak Fern Using MaxEnt Modeling(American Fern Journal, 2023-06-16) Hay, NM; Akinwuntan, JV; Cai, V; Windham, MD; Pryer, KMAbstract . Anthropogenic climate change is projected to have an especially negative impact on the survival of plants that are dependent on limited microclimatic refugia or that already reside at their climatic extreme. Gymnocarpium appalachianum is a narrowly endemic fern restricted to cold mountaintops and algific vents in the central and southern Appalachian region of eastern North America. It is the much rarer of the two documented diploid parents of the circumboreal allotetraploid G. dryopteris - one of the most widespread fern species on the planet. Gymnocarpium appalachianum is a good case study for forecasting how evolutionarily significant, but rare, species might survive on a warming planet. We utilize an ecological niche modeling approach (MaxEnt) to explore the projected distribution of G. appalachianum under past (Last Glacial Maximum) and future climate models. All known verified herbarium records of G. appalachianum were georeferenced, for a total of 70 occurrence points. Nineteen standard bioclimatic variables extracted from WorldClim were used to model near-current climate projections; representative concentration pathways (RCPs 2.6 and 8.5) were used for future climate projections (2070). The temperature annual range, mean temperature of warmest quarter, precipitation of driest month, precipitation of coldest quarter, and mean diurnal range were identified as the key variables for shaping the distribution of G. appalachianum. An unanticipated result from our analyses is that G. appalachianum has past and current projected habitat suitability in Alaska. Because this overlaps with the current range of G. disjunctum, the other diploid parent of G. dryopteris, it suggests a possible region of origin for this circumboreal tetraploid descendent of G. appalachianum - a research avenue to be pursued in the future. Our study envisions a dire fate for G. appalachianum; its survival will likely require an urgent contingency plan that includes human-mediated population relocation to cooler, northern locations. Understanding the long-term sustainability of narrowly endemic plants such as G. appalachianum is critical in decisions about their management and conservation.Item Open Access Expression level dominance and homeolog expression bias in recurrent origins of the allopolyploid fern polypodium hesperium(American Fern Journal, 2019-07-01) Sigel, EM; Der, JP; Windham, MD; Pryer, KM© 2019 American Fern Society. All rights reserved. Allopolyploidization is a common mode of speciation in ferns with many taxa having formed recurrently from distinct hybridization events between the same parent species. Each hybridization event marks the union of divergent parental gene copies, or homeologs, and the formation of an independently derived lineage. Little is known about the effects of recurrent origins on the genomic composition and phenotypic variation of allopolyploid fern taxa. To begin to address this knowledge gap, we investigated gene expression patterns in two naturally formed, independently derived lineages of the allotetraploid fern Polypodium hesperium relative to its diploid progenitor species, Polypodium amorphum and Polypodium glycyrrhiza. Using RNA-sequencing to survey total gene expression levels for 19194 genes and homeolog-specific expression for 1073 genes, we found that, in general, gene expression in both lineages of P. hesperium was biased toward P. amorphum - both by mirroring expression levels of P. amorphum and preferentially expressing homeologs derived from P. amorphum. However, we recovered substantial expression variation between the two lineages at the level of individual genes and among individual specimens. Our results align with similar transcriptome profile studies of angiosperms, suggesting that expression in many allopolyploid plants reflects the dominance of a specific parental subgenome, but that recurrent origins impart substantial expression, or phenotypic, variation to allopolyploid taxa.Item Open Access Gaga, a new fern genus segregated from cheilanthes (pteridaceae)(Systematic Botany, 2012-10-01) Li, F; Pryer, KM; Windham, MDOngoing molecular phylogenetic studies of cheilanthoid ferns confirm that the genus Cheilanthes (Pteridaceae) is polyphyletic. A monophyletic group of species within the hemionitid clade informally called the "C. marginata group" is here shown to be distinct from its closest relatives (the genus Aspidotis) and phylogenetically distant from the type species of Cheilanthes. This group is here segregated from Cheilanthes as the newly described genus, Gaga . In this study, we use molecular data from four DNA regions (plastid: matK, rbcL, trnG-R; and nuclear: gapCp) together with spore data to circumscribe the morphological and geographical boundaries of the new genus and investigate reticulate evolution within the group. Gaga is distinguished from Aspidotis by its rounded to attenuate (vs. mucronate) segment apices, minutely bullate margins of mature leaves (vs. smooth at 40 ×), and less prominently lustrous and striate adaxial blade surfaces. The new genus is distinguished from Cheilanthes s. s. by its strongly differentiated, inframarginal pseudoindusia, the production of 64 small or 32 large spores (vs. 32 small or 16 large) per sporangium, and usually glabrous leaf blades. A total of nineteen species are recognized within Gaga; seventeen new combinations are made, and two new species, Gaga germanotta and Gaga monstraparva , are described. © Copyright 2012 by the American Society of Plant Taxonomists.Item Open Access Infraspecific diversification of the star cloak fern (Notholaena standleyi) in the deserts of the United States and Mexico.(American journal of botany, 2020-04-07) Kao, T; Rothfels, CJ; Melgoza‐Castillo, A; Pryer, KM; Windham, MDPREMISE:Not all ferns grow in moist and shaded habitats. One well-known example is Notholaena standleyi, a species that thrives in deserts of the southwestern United States and Mexico. This species exhibits several "chemotypes" that differ in farina (flavonoid exudates) color and chemistry. By integrating data from molecular phylogenetics, cytology, biochemistry, and biogeography, we circumscribed the major evolutionary lineages within N. standleyi and reconstructed their diversification histories. METHODS:Forty-eight samples were selected from across the geographic distribution of N. standleyi. Phylogenetic relationships were inferred using four plastid and five nuclear markers. Ploidy levels were inferred using spore sizes calibrated by chromosome counts, and farina chemistry was compared using thin-layer chromatography. RESULTS:Four clades are recognized, three of which roughly correspond to previously recognized chemotypes. The diploid clades G and Y are found in the Sonoran and Chihuahuan deserts, respectively; they are estimated to have diverged in the Pleistocene, congruent with the postulated timing of climatological events separating these two deserts. Clade P/YG is tetraploid and partially overlaps the distribution of clade Y in the eastern Chihuahuan Desert. It is apparently confined to limestone, a geologic substrate rarely occupied by members of the other clades. The cryptic (C) clade, a diploid group known only from southern Mexico and highly disjunct from the other three clades, is newly recognized here. CONCLUSIONS:Our results reveal a complex intraspecific diversification history of N. standleyi, traceable to a variety of evolutionary drivers including classic allopatry, parapatry with or without changes in geologic substrate, and sympatric divergence through polyploidization.Item Open Access Low-copy nuclear data confirm rampant allopolyploidy in the cystopteridaceae (Polypodiales)(Taxon, 2014-10-01) Rothfels, CJ; Johnson, AK; Windham, MD; Pryer, KM© International Association for Plant Taxonomy (IAPT) 2014. Here we present the first nuclear phylogeny for Cystopteridaceae (Polypodiales), using the single-copy locus gapCp “short”. This phylogeny corroborates broad results from plastid data in demonstrating strong support for the monophyly of the family’s three genera—Cystopteris, Acystopteris, and Gymnocarpium—and of the major groups within Cystopteris (C. montana, the sudetica and bulbifera clades, and the C. fragilis complex). In addition, it confirms the rampant hybridization (allopolyploidy) that has long been suspected within both Cystopteris and Gymnocarpium. In some cases, these data provide the first DNA-sequence-based evidence for previous hypotheses of polyploid species origins (such as the cosmopolitan G. dryopteris being an allotetraploid derivative of the diploids G. appalachianum and G. disjunctum). Most of the allopolyploids, however, have no formal taxonomic names. This pattern is particularly strong within the C. fragilis complex, where our results imply that the eight included accessions of “C. fragilis” represent at least six distinct allopolyploid taxa.Item Open Access Low-copy nuclear sequence data confirm complex patterns of farina evolution in notholaenid ferns (Pteridaceae).(Molecular phylogenetics and evolution, 2019-09) Kao, T; Pryer, KM; Freund, FD; Windham, MD; Rothfels, CJNotholaenids are an unusual group of ferns that have adapted to, and diversified within, the deserts of Mexico and the southwestern United States. With approximately 40 species, this group is noted for being desiccation-tolerant and having "farina"-powdery exudates of lipophilic flavonoid aglycones-that occur on both the gametophytic and sporophytic phases of their life cycle. The most recent circumscription of notholaenids based on plastid markers surprisingly suggests that several morphological characters, including the expression of farina, are homoplasious. In a striking case of convergence, Notholaena standleyi appears to be distantly related to core Notholaena, with several taxa not before associated with Notholaena nested between them. Such conflicts can be due to morphological homoplasy resulting from adaptive convergence or, alternatively, the plastid phylogeny itself might be misleading, diverging from the true species tree due to incomplete lineage sorting, hybridization, or other factors. In this study, we present a species phylogeny for notholaenid ferns, using four low-copy nuclear loci and concatenated data from three plastid loci. A total of 61 individuals (49 notholaenids and 12 outgroup taxa) were sampled, including 31 out of 37 recognized notholaenid species. The homeologous/allelic nuclear sequences were retrieved using PacBio sequencing and the PURC bioinformatics pipeline. Each dataset was first analyzed individually using maximum likelihood and Bayesian inference, and the species phylogeny was inferred using *BEAST. Although we observed several incongruences between the nuclear and plastid phylogenies, our principal results are broadly congruent with previous inferences based on plastid data. By mapping the presence of farina and their biochemical constitutions on our consensus phylogenetic tree, we confirmed that the characters are indeed homoplastic and have complex evolutionary histories. Hybridization among recognized species of the notholaenid clade appears to be relatively rare compared to that observed in other well-studied fern genera.Item Open Access Natural hybridization between genera that diverged from each other approximately 60 million years ago.(Am Nat, 2015-03) Rothfels, CJ; Johnson, AK; Hovenkamp, PH; Swofford, DL; Roskam, HC; Fraser-Jenkins, CR; Windham, MD; Pryer, KMA fern from the French Pyrenees-×Cystocarpium roskamianum-is a recently formed intergeneric hybrid between parental lineages that diverged from each other approximately 60 million years ago (mya; 95% highest posterior density: 40.2-76.2 mya). This is an extraordinarily deep hybridization event, roughly akin to an elephant hybridizing with a manatee or a human with a lemur. In the context of other reported deep hybrids, this finding suggests that populations of ferns, and other plants with abiotically mediated fertilization, may evolve reproductive incompatibilities more slowly, perhaps because they lack many of the premating isolation mechanisms that characterize most other groups of organisms. This conclusion implies that major features of Earth's biodiversity-such as the relatively small number of species of ferns compared to those of angiosperms-may be, in part, an indirect by-product of this slower "speciation clock" rather than a direct consequence of adaptive innovations by the more diverse lineages.Item Open Access Patterns of diversification in the xeric-adapted fern genus myriopteris (Pteridaceae)(Systematic Botany, 2014-01-01) Grusz, AL; Windham, MD; Yatskievych, G; Huiet, L; Gastony, GJ; Pryer, KMStrong selective pressures imposed by drought-prone habitats have contributed to extensive morphological convergence among the 400 + species of cheilanthoid ferns (Pteridaceae). As a result, generic circumscriptions based exclusively on macromorphology often prove to be non-monophyletic. Ongoing molecular phylogenetic analyses are providing the foundation for a revised classification of this challenging group and have begun to clarify its complex evolutionary history. As part of this effort, we generated and analyzed DNA sequence data for three plastid loci (rbcL, atpA, and the intergenic spacer trnG-trnR) for the myriopterid clade, one of the largest monophyletic groups of cheilanthoid ferns. This lineage encompasses 47 primarily North and Central American taxa previously included in Cheilanthes but now placed in the recircumscribed genus Myriopteris. Here, we infer a phylogeny for the group and examine key morphological characters across this phylogeny. We also include a brief discussion of the three well-supported Myriopteris subclades, along with a review of reproductive mode and known ploidy levels for members of this early diverging lineage of cheilanthoid ferns. © Copyright 2014 by the American Society of Plant Taxonomists.Item Open Access Phylogeny, divergence time estimates, and phylogeography of the diploid species of the polypodium vulgare complex (Polypodiaceae)(Systematic Botany, 2014-01-01) Sigel, EM; Windham, MD; Haufler, CH; Pryer, KM© 2014 by the American Society of Plant Taxonomists. The Polypodium vulgare complex (Polypodiaceae) comprises a well-studied group of fern taxa whose members are cryptically differentiated morphologically and have generated a confusing and highly reticulate species cluster. Once considered a single species spanning much of northern Eurasia and North America, P. vulgare has been segregated into 17 diploid and polyploid taxa as a result of cytotaxonomic work, hybridization experiments, and isozyme studies conducted during the 20th century. Despite considerable effort, however, the evolutionary relationships among the diploid members of the P. vulgare complex remain poorly resolved. Here we infer a diploids-only phylogeny of the P. vulgare complex and related species to test previous hypotheses concerning relationships within Polypodium sensu stricto. Using sequence data from four plastid loci (atpA, rbcL, matK, and trnG-trnR), we recovered a monophyletic P. vulgare complex comprising four well-supported clades. The P. vulgare complex is resolved as sister to the Neotropical P. plesiosorum group and these, in turn, are sister to the Asian endemic Pleurosoriopsis makinoi. Using divergence time analyses incorporating previously derived age constraints and fossil data, we estimate an early Miocene origin for the P. vulgare complex and a late Miocene-Pliocene origin for the four major diploid lineages of the complex, with the majority of extant diploid species diversifying from the late Miocene through the Pleistocene. Finally, we use our node age estimates to reassess previous hypotheses, and to propose new hypotheses, about the historical events that shaped the diversity and current geographic distribution of the diploid species of the P. vulgare complex.Item Open Access rbcL and matK earn two thumbs up as the core DNA barcode for ferns.(PloS one, 2011-01) Li, F; Kuo, L; Rothfels, CJ; Ebihara, A; Chiou, W; Windham, MD; Pryer, KMBACKGROUND: DNA barcoding will revolutionize our understanding of fern ecology, most especially because the accurate identification of the independent but cryptic gametophyte phase of the fern's life history--an endeavor previously impossible--will finally be feasible. In this study, we assess the discriminatory power of the core plant DNA barcode (rbcL and matK), as well as alternatively proposed fern barcodes (trnH-psbA and trnL-F), across all major fern lineages. We also present plastid barcode data for two genera in the hyperdiverse polypod clade--Deparia (Woodsiaceae) and the Cheilanthes marginata group (currently being segregated as a new genus of Pteridaceae)--to further evaluate the resolving power of these loci. PRINCIPAL FINDINGS: Our results clearly demonstrate the value of matK data, previously unavailable in ferns because of difficulties in amplification due to a major rearrangement of the plastid genome. With its high sequence variation, matK complements rbcL to provide a two-locus barcode with strong resolving power. With sequence variation comparable to matK, trnL-F appears to be a suitable alternative barcode region in ferns, and perhaps should be added to the core barcode region if universal primer development for matK fails. In contrast, trnH-psbA shows dramatically reduced sequence variation for the majority of ferns. This is likely due to the translocation of this segment of the plastid genome into the inverted repeat regions, which are known to have a highly constrained substitution rate. CONCLUSIONS: Our study provides the first endorsement of the two-locus barcode (rbcL+matK) in ferns, and favors trnL-F over trnH-psbA as a potential back-up locus. Future work should focus on gathering more fern matK sequence data to facilitate universal primer development.Item Open Access Rediscovery of Polypodium calirhiza (Polypodiaceae) in Mexico(Brittonia, 2014) Sigel, EM; Windham, MD; Smith, AR; Dyer, RJ; Pryer, KMThis study addresses reported discrepancies regarding the occurrence of Polypodium calirhiza in Mexico. The original paper describing this taxon cited collections from Mexico, but the species was omitted from the recent Pteridophytes of Mexico. Originally treated as a tetraploid cytotype of P. californicum, P. calirhiza now is hypothesized to have arisen through hybridization between P. glycyrrhiza and P. californicum. The tetraploid can be difficult to distinguish from either of its putative parents, but especially so from P. californicum. Our analyses show that a combination of spore length and abaxial rachis scale morphology consistently distinguishes P. calirhiza from P. californicum, and we confirm that both species occur in Mexico. Although occasionally found growing together in the United States, the two species are strongly allopatric in Mexico: P. californicum is restricted to coastal regions of the Baja California peninsula and neighboring Pacific islands, whereas P. calirhiza grows at high elevations in central and southern Mexico. The occurrence of P. calirhiza in Oaxaca, Mexico, marks the southernmost extent of the P. vulgare complex in the Western Hemisphere. © 2014 The New York Botanical Garden.