Ashley B. Morris

Comparative phylogeography of unglaciated eastern North America

Regional phylogeographical studies involving co‐distributed animal and plant species have been conducted for several areas, most notably for Europe and the Pacific Northwest of North America. Until recently, phylogeographical studies in unglaciated eastern North America have been largely limited to animals. As more studies emerge for diverse lineages (including plants), it seems timely to assess the phylogeography across this region: (i) comparing and contrasting the patterns seen in plants and animals; (ii) assessing the extent of pseudocongruence; and (iii) discussing the potential applications of regional phylogeography to issues in ecology, such as response to climatic change. Unglaciated eastern North America is a large, geologically and topographically complex area with the species examined having diverse distributions. Nonetheless, some recurrent patterns emerge: (i) maritime — Atlantic vs. Gulf Coast; (ii) Apalachicola River discontinuity; (iii) Tombigbee River discontinuity; (iv) the Appalachian Mountain discontinuity; (v) the Mississippi River discontinuity; and (vi) the Apalachicola River and Mississippi River discontinuities. Although initially documented in animals, most of these patterns are also apparent in plants, providing support for phylogeographical generalizations. These patterns may generally be attributable to isolation and differentiation during Pleistocene glaciation, but in some cases may be older (Pliocene). Molecular studies sometimes agree with longstanding hypotheses of glacial refugia, but also suggest additional possible refugia, such as the southern Appalachian Mountains and areas close to the Laurentide Ice Sheet. Many species exhibit distinct patterns that reflect the unique, rather than the shared, aspects of species’ phylogeographical histories. Furthermore, similar modern phylogeographical patterns can result from different underlying causal factors operating at different times (i.e. pseudocongruence). One underemphasized component of pseudocongruence may result from the efforts of researchers to categorize patterns visually — similar patterns may, in fact, not fully coincide, and inferring agreement may obscure the actual patterns and lead to erroneous conclusions. Our modelling analyses indicate no clear spatial patterning and support the hypothesis that phylogeographical structure in diverse temperate taxa is complex and was not shaped by just a few barriers.

Stratified analysis of the soil seed bank in the cedar glade endemic Astragalus bibullatus: evidence for historical changes in genetic structure

Persistent seed banks may provide information on historical changes in the genetic composition of populations. We used stratified sampling of the soil seed bank of Astragalus bibullatus (Pynes ground plum) to assess levels of temporal variation in population genetic structure and to infer historical changes in the levels of inbreeding and relative gene flow. This species has an extremely limited distribution in the Central Basin of Tennessee, where it is found in open areas and along the edges of cedar glades. Protein electrophoresis was conducted on seedlings grown from seeds that had been recovered from three successive 1 cm thick layers of soil sampled from six sites. Analyses of seven polymorphic allozyme loci indicated that there were substantial levels of genetic differentiation among soil layers and sites. Higher levels of genetic diversity were found in seed than in vegetative populations that had been sampled in a previous study. Seed populations from the uppermost soil layer had higher heterozygote deficiencies, displayed higher levels of differentiation among sites, and had higher private allele frequencies than seed populations from the lower two layers. The change in heterozygosity and distribution of genetic variation among sites for the youngest soil layer is consistent with a pattern of increased selfing, sib mating, and decreased gene flow among populations. These changes in inbreeding and relative levels of gene flow are corroborated by information on historical land use practices in the region and support the hypothesis that loss of appropriate habitat has led to smaller population sizes and a more fragmented distribution of this cedar glade endemic.

Phylogeographical structure and temporal complexity in American sweetgum (Liquidambar styraciflua; Altingiaceae)

Eastern North American plant biogeography has traditionally focused on two primary issues: (i) the location of temperate Pleistocene refugia and their proximity to the southern margin of the ice sheet during the last glacial maximum, and (ii) the origin of the temperate element of northern Latin America. While numerous population genetic and phylogeographical studies have focused on the first issue, few (if any) have considered the second. We addressed these issues by surveying 117 individuals from 24 populations of Liquidambar styraciflua (American sweetgum; Altingiaceae) across the southeastern USA, eastern Mexico, and Guatemala, using more than 2200 bp of chloroplast DNA sequence data. To specifically address the issue of timing, we estimated intraspecific divergence times on the basis of multiple fossil‐based calibration points, using taxa from Altingiaceae (Liquidambar and Altingia) and Hammamelidaceae (Hamamelis) as outgroups. More than half of the sampled localities exhibited multiple haplotypes. Remarkably, the greatest variation was observed within the USA, with Mexico and Guatemala sharing widespread haplotypes with Texas, Mississippi, Kentucky, Ohio, and northern Virginia. This lack of differentiation suggests shared ancestral polymorphisms, and that the genetic signal we observed is older than the disjunction itself. Our data provide support for previously proposed hypotheses of Pleistocene refugia in peninsular Florida and along the eastern Atlantic, but also for deeper divergences (~8 million years ago) within the USA. These patterns reflect a dynamic biogeographical history for eastern North American trees, and emphasize the importance of the inclusion of a temporal component in any phylogeographical study.

Northern hemisphere biogeography of Cerastium (Caryophyllaceae): insights from phylogenetic analysis of noncoding plastidnucleotide sequences

Phylogenetic relationships and biogeography of the genus Cerastium were studied using sequences of three noncoding plastid DNA regions (trnL intron, trnL-trnF spacer, and psbA-trnH spacer). A total of 57 Cerastium taxa was analyzed using two species of the putative sister genus Stellaria as outgroups. Maximum parsimony analyses identified four clades that largely corresponded to previously recognized infrageneric groups. The results suggest an Old World origin and at least two migration events into North America from the Old World. The first event possibly took place across the Bering land bridge during the Miocene. Subsequent colonization of South America occurred after the North and South American continents joined during the Pliocene. A more recent migration event into North America probably across the northern Atlantic took place during the Quaternary, resulting in the current circumpolar distribution of the Arctic species. Molecular clock dating of major biogeographic events was internally consistent on the phylogenetic trees. The arctic high-polyploid species form a polytomy together with some boreal and temperate species of the C. tomentosum group and the C. arvense group. Lack of genetic variation among the arctic species probably indicates a recent origin. The annual life form is shown to be of polyphyletic origin.

Phylogeny and Divergence Time Estimation in Illicium with Implications for New World Biogeography

Abstract Illiciaceae comprise the single genus, Illicium, and approximately 30–40 species distributed in southern and eastern Asia, the southeastern United States, Mexico, and the Greater Antilles. On the basis of perianth morphology, current taxonomic treatments recognize two sections that do not correspond to geographic distributions. A recent phylogenetic study using ITS sequences found strong support for separate Old and New World clades, but did not include all New World species. We elaborate on this previous study by including all known New World species of Illicium, as well as additional Asian species, using ITS and the plastid loci trnL, the trnL-F spacer, and trnG. We estimated divergence times within Illicium using penalized likelihood and multiple fossil calibration points. Our results support separate New and Old World clades, and we re-circumscribe current sectional groupings based on these data. The Illicium crown group appears to have arisen during the Cretaceous, with New World extant taxa diversifying as recently as the late Miocene or early Pliocene. These dates are consistent with recent fossil evidence and molecular estimates for minimum ages of the angiosperms, and for timing of diversification events in other ancient groups, such as Nymphaeales and Chloranthaceae.

Variation in Frequency of Clonal Reproduction Among Populations of Fagus grandifolia Ehrh. in Response to Disturbance

Abstract Asexual reproduction has been observed to be more frequently associated with marginal habitats or with higher levels of disturbance. Here we examined frequencies of clonal reproduction across populations of Fagus grandifolia with varying levels of disturbance and habitat quality. We used ISSRs to identify clonal genotypes at each of five sites. Three of the sites were typical temperate habitats with relatively low densities of stems and were below 1,100 m, and two sites were above 1,370 m, possessed a high density of stems, and were characteristic of sub-alpine beech gap populations. The frequency of clonal reproduction varied substantially across sites and appears to be greater where disturbance is more severe.

Digitization Workflows for Flat Sheets and Packets of Plants, Algae, and Fungi

Effective workflows are essential components in the digitization of biodiversity specimen collections. To date, no comprehensive, community-vetted workflows have been published for digitizing flat sheets and packets of plants, algae, and fungi, even though latest estimates suggest that only 33% of herbarium specimens have been digitally transcribed, 54% of herbaria use a specimen database, and 24% are imaging specimens. In 2012, iDigBio, the U.S. National Science Foundations (NSF) coordinating center and national resource for the digitization of public, nonfederal U.S. collections, launched several working groups to address this deficiency. Here, we report the development of 14 workflow modules with 7–36 tasks each. These workflows represent the combined work of approximately 35 curators, directors, and collections managers representing more than 30 herbaria, including 15 NSF-supported plant-related Thematic Collections Networks and collaboratives. The workflows are provided for download as Portable Document Format (PDF) and Microsoft Word files. Customization of these workflows for specific institutional implementation is encouraged.

Idiosyncratic responses of evergreen broad-leaved forest constituents in China to the late Quaternary climate changes

Subtropical evergreen broad-leaved forest (EBLF) is one of the most important vegetation types in China. Inferences from palaeo-biome reconstruction (PBR) and phylogeography regarding range shift history of EBLF during the late Quaternary are controversial and should be reconciled. We compared phylogeographic patterns of three EBLF constituents in China, Castanopsis tibetana, Machilus thunbergii and Schima superba. Contrary to a chorus of previous phylogeographic studies and the results of species distribution modelling (SDM) of this study (in situ survival during the LGM), the three species displayed three different phylogeographic patterns that conform to either an in situ survival model or an expansion-contraction model. These results are partially congruent with the inference of PBR that EBLF was absent to the north of 24° N at the LGM. This study suggests that the constituents of EBLF could have responded idiosyncratically to climate changes during the Late Quaternary. The community assemblages of EBLF could have been changing over time, resulting in no palaeo-analogs to modern-day EBLF, which may be the main reason responsible for the failure of PBR to detect the occurrence of EBLF north of 24° N at the LGM.

Phylogenetic Relationships in the North American Genus Pseudemys (Emydidae) Inferred from Two Mitochondrial Genes

Abstract Pseudemys turtles are an important component of southeastern North American aquatic ecosystems, but the relationships within the genus are poorly understood. Convergent morphology and apparent hybridization have complicated the identification of species boundaries and have resulted in numerous conflicting taxonomic treatments. We used mitochondrial DNA sequence data from the control region and cytochrome-b gene to address 1 ) the monophyly of currently recognized subgeneric clades (the cooters and the red-bellies), and 2) relationships within these two groups. A total of 91 specimens representing 8 Pseudemys and 3 outgroup taxa were sampled, and 36 distinct haplotypes were recovered. Pseudemys forms a well-supported monophyletic group, but relationships among species were not well resolved, such that support for the two subgeneric groupings was lacking. Furthermore, most taxa do not appear to be monophyletic, with the exception of P. gorzugi (Rio Grande Cooter) and P. texana (Texas Cooter), suggesting the possibility of mitochondrial introgression as a result of historic or continuing hybrid swarms across the range of the genus, or the lack of resolution may reflect a pattern of recent speciation. In light of recent molecular surveys in turtles, the utility of mitochondrial DNA in turtle systematics is also discussed .

INVESTIGATIONS OF INTERNAL INTERACTIONS BETWEEN THE PARASITIC BARNACLE LOXOTHYLACUS TEXANUS (RHIZOCEPHALA: SACCULINIDAE) AND ITS HOST CALLINECTES SAPIDUS (BRACHYURA: PORTUNIDAE) USING PCR TECHNIQUES

We describe techniques that enable the preservation of tissues from the rhizocephalan barnacle, Loxothylacus texanus, inside the body cavities of blue crab hosts, Callinectes sapidus, in a manner that minimizes the degradative activities of hepatopancreatic enzymes. These procedures allow the extraction and amplification of both parasite and host 18S rDNA within the same sample and enable one to distinguish between parasitized and unparasitized crab tissue in as little as two weeks after infection, well before any external manifestations of the parasites. Two PCR-based approaches were taken to identify the presence of L. texanus. In the first approach, a set of primers specific for L. texanus was used to specifically amplify 18S sequence in a background of C. sapidus DNA or the DNA of other barnacle species. In the second approach, a set of general primers was used to amplify 18S sequence from C. sapidus and a variety of barnacle species. The products of this PCR were then digested with an enzyme that recognizes a restriction site present only in the L. texanus PCR product to yield a unique pattern of fragments. With these techniques, we could detect as few as five parasitic cypris larvae in water samples, as well as L. texanus in the tissue of a small crab collected from the field and in the four anterior periopods of a crab bearing the external stage of the parasite. In experiments with potential hosts of varying sizes and molt stages, we confirmed that the parasite was significantly more effective in infecting crabs less than 30 mm carapace width than larger individuals. These techniques will facilitate future investigations of ecological and physiological interactions between these important crustacean parasites and their hosts and will help to determine the economic impact of this parasite on blue crab fisheries.