James P. Bogart

Ridges and rivers: a test of competing hypotheses of Amazonian diversification using a dart-poison frog ( Epipedobates femoralis )

Mitochondrial DNA cytochrome b sequence data from a dart–poison frog, Epipedobates femoralis, were used to test two hypotheses of Amazonian diversification: the riverine barrier and the ridge hypotheses. Samples were derived from sites located on both banks of the Rio Juruá and on both sides of the Iquitos Arch in western Amazonia. The phylogeographic structure was inconsistent with predictions of the riverine barrier hypothesis. Haplotypes from opposite river banks did not form monophyletic clades in any of our phylogenetic analyses, nor was the topology within major clades consistent with the riverine hypothesis. Further, the greatest differentiation between paired sites on opposite banks was not at the river mouth where the strongest barrier to gene flow was predicted to occur. The results instead were consistent with the hypothesis that ancient ridges (arches), no longer evident on the landscape, have shaped the phylogeographic relationships of Amazonian taxa. Two robustly supported clades map onto opposite sides of the Iquitos Arch. The mean haplotypic divergence between the two clades, in excess of 12%, suggests that this cladogenic event dates to between five and 15 million years ago. These estimates span a period of major orogenesis in western South America and presumably the formation of these ancient ridges.

Evolutionary implications of polyploidy in amphibians and reptiles.

In plants, Polyploidy is recognized to be a wide-spread phenomenon and of considerable practical and evolutionary importance, whereas polyploid animal species have been relegated for the most part, to insignificance in terms of their existence or evolutionary importance. Evolutionary and genetic authorities have adhered mostly to Mullers’s 1925 (1) contention that sexual imbalance in polyploids would not permit bisexual polyploids to exist as natural entities in animals as they do in plants, which are capable of vegetative reproduction. Asexual polyploids are also condemned, in animals, by the commonly held, and mathematically “proven” (2) viewpoint that this method of reproduction reduces genetic recombination and is tantamount to phylogenetic suicide (3–5). It is evident, however, that an increasing number of polyploid amphibians and reptiles are being encountered in natural populations living in North America, South America, Europe, Asia, and Africa. To ignore their existence or to pass judgement on their evolutionary significance without adequate study is incomprehensible. In spite of the theoretical dogma surrounding animal polyploids, there is a growing bank of data which suggests that naturally occurring animal polyploids may play an interesting and significant role in population genetics and speciation.

Unisexual salamanders (genus Ambystoma) present a new reproductive mode for eukaryotes.

To persist, unisexual and asexual eukaryotes must have reproductive modes that circumvent normal bisexual reproduction. Parthenogenesis, gynogenesis, and hybridogenesis are the modes that have generally been ascribed to various unisexuals. Unisexual Ambystoma are abundant around the Great Lakes region of North America, and have variously been described as having all 3 reproductive modes. Diploid and polyploid unisexuals have nuclear genomes that combine the haploid genomes of 2 to 4 distinct sexual species, but the mtDNA is unlike any of those 4 species and is similar to another species, Ambystoma barbouri. To obtain better resolution of the reproductive mode used by unisexual Ambystoma and to explore the relationship of A. barbouri to the unisexuals, we sequenced the mitochondrial control and highly variable intergenic spacer region of 48 ambystomatids, which included 28 unisexuals, representatives of the 4 sexual species and A. barbouri. The unisexuals have similar sequences over most of their range, and form a close sister group to A. barbouri, with an estimated time of divergence of 2.4-3.9 million years ago. Individuals from the Lake Erie Islands (Kelleys, Pelee, North Bass) have a haplotype that demonstrates an isolation event. We examined highly variable microsatellite loci, and found that the genetic makeup of the unisexuals is highly variable and that unisexual individuals share microsatellite alleles with sexual individuals within populations. Although many progeny from the same female had the same genotype for 5 microsatellite DNA loci, there was no indication that any particular genome is consistently inherited in a clonal fashion in a population. The reproductive mode used by unisexual Ambystoma appears to be unique; we suggest kleptogenesis as a new unisexual reproductive mode that is used by these salamanders.

Identification of intergenomic recombinations in unisexual salamanders of the genus Ambystoma by genomic in situ hybridization (GISH)

Unisexual salamanders in the genus Ambystoma (Amphibia, Caudata) are endemic to eastern North America and are mostly all-female polyploids. Two to four of the bisexual species, A. laterale, A. jeffersonianum, A. texanum and A. tigrinum, contribute to the nuclear genome of unisexuals and more than 20 combinations that range from diploid to pentaploid have been identified in this complex. Because the karyotypes of the four bisexual species are similar, homologous and homoeologous chromosomes in the unisexuals can not be distinguished by conventional or banded karyotypes. We chose two widespread unisexual genomic combinations (A.laterale–2 jeffersonianum [or LJJ] and A. 2 laterale–jeffersonianum [or LLJ]) and employed genomic in situ hybridization (GISH) to identify the genomes in these unisexuals. Under optimum conditions, GISH reliably distinguishes the respective chromosomes attributed to both A.laterale and A. jeffersonianum. Of four populations examined, two were found to have independently evolved homoeologous recombinants that persist in both LJJ and LLJ individuals. Our results refute the previous hypothesis of clonal integrity and independent evolution of the genome combinations in these unisexuals. Our data provide evidence for intergenomic interactions between maternal chromosomes during meiosis in unisexuals and help to explain previously observed non-homologous bivalents and/or quadrivalents among lampbrush chromosomes that were possibly initiated by partial homosequential pairing among the homo(eo)logues. To explore the utility of GISH in other members of the complex, probes developed from A. laterale were also applied to unisexuals that contained A. tigrinum and A. texanum genomes. GISH is an effective tool that can be used to identify and to quantify genomic constituents and to investigate intergenomic interactions in unisexual salamanders. GISH also has potential application to examine possible genomic evolution in other unisexuals.

An interactive, microcomputer-based karyotype analysis system for phylogenetic cytotaxonomy

Abstract A versatile and practical semi-automatic computerized system of karyotype analysis has been developed for phylogenetic cytotaxonomy for under

GENETIC AND MORPHOLOGICAL VARIATION IN VANZOLINIUS DISCODACTYLUS : A TEST OF THE RIVER HYPOTHESIS OF SPECIATION

4,000. The modular software system is designed for use solely with a small 8-bit microcomputer and requires virtually no custom made hardware. The system interacts with the operator and greatly augments the speed and accuracy of cytotaxonomic investigations.

Multi-character perspectives on the evolution of intraspecific differentiation in a neotropical hylid frog

Allozyme and morphological variation among populations of an Amazonian frog were analyzed to test for a riverine effect on population differentiation. High levels of genetic diversity within populations and genetic diversification among populations were found. Although correlations between genetic and geographic distances of population pairs were positive, they were not significant and indicated that sites that were close geographically were not genetically more similar. Only one of the 15 polymorphic loci showed any variation in allozyme frequency attributable to the presence of the river, but none of the overall among-population component of allozyme frequency variation (FST = 0.368) can be attributed to the river (FRT = 0.000). Significant differences in morphology (corrected for size differences) were found among the 11 populations in canonical variate space. In univariate Nested ANOVAs, five of the 12 morphological variables showed significant differences due to the presence of the river. However, cluster analysis performed on all morphological variables failed to detect obvious groups based on river bank locality. We could detect no congruence of pattern between morphological and genetic variation among populations. These results suggest that morphological characters may evolve at different rates than detectable allozyme differentiation.

An unexpected recent ancestor of unisexual Ambystoma.

BackgroundMulti-character empirical studies are important contributions to our understanding of the process of speciation. The relatively conserved morphology of, and importance of the mate recognition system in anurans, combined with phylogenetic tools, provide an opportunity to address predictions about the relative role of each in the process of speciation. We examine the relationship among patterns of variation in morphology, call characters, and 16S gene sequences across seven populations of a neotropical hylid frog, Hyla leucophyllata, to infer their relative importance in predicting the early stages of population differentiation.ResultsMultivariate analyses demonstrate that both morphological and call characteristics were significantly variable among populations, characterized by significantly lower intra-population dispersion in call space than morphological space, and significantly greater among-population variation in call structure. We found lack of concordance between a 16S DNA phylogeny of Hyla leucophyllata and the significant population-level differentiation evident in both external morphology and male advertisement call. Comparisons of the reconstructed gene trees to simulated lineages support the notion that variation in call cannot be simply explained by population history.ConclusionDiscordance among traits may reflect sampling biases (e.g. single genetic marker effects), or imply a decoupling of evolution of different suites of characters. Diagnostic differences among populations in call structure possibly reflect local selection pressures presented by different heterospecific calling assemblages and may serve as a precursor of species-wide differentiation. Differentiation among populations in morphology may be due to ecophenotypic variation or to diversifying selection on body size directly, or on frequency attributes of calls (mediated by female choice) that show a strong relationship to body size.

Vocalizations of primary forest frog species in the Central Amazon.

Previous research has shown that members of the unisexual hybrid complex of the genus Ambystoma possess a mitochondrial genome that is unrelated to their nuclear parental species, but the origin of this mitochondrion has remained unclear. We used a 744‐bp fragment of the mitochondrial gene cytochrome b within a comparative phylogenetic framework to infer the maternal ancestor of this unisexual lineage. By examining a broader range of species than has previously been compared, we were able to uncover a recent maternal ancestor to this complex. Unexpectedly, Ambystoma barbouri, a species whose nuclear DNA has not been identified in the unisexuals, was found to be the recent maternal ancestor of the individuals examined through the discovery of a shared mtDNA haplotype between the unisexuals and A. barbouri. Based on a combination of sequence data and glacial patterning, we estimate that the unisexual lineage probably originated less than 25 000 years ago. In addition, all unisexuals examined showed extremely similar mtDNA sequences and the resultant phylogeny was consistent with a single origin for this lineage. These results confirm previous suggestions that the unisexual Ambystoma complex was formed from a hybridization event in which the nuclear DNA of the original maternal species was subsequently lost.

Polyploidy in Amphibia

The calls of 18 species of Amazonian forest frogs were recorded in 3 localities: the Tapajos National Park near Itaituba, the Reserva Ducke near Manaus, and the INPA-WWF reserves near Manaus. Structural and time parameters and sonographs of these calls including previously undescribed vocalization by 10 species are presented. Unlike open habitat species, several forest frong species characteriscally demonstrated one on more of the following temporal parameters: very low call rates, sporadic intervals, infrequent nights of calling and synchronized chorusing. It is hypothesized that predation has influenced the evolution of vocal behaviour in Amazonian forest frogs.