Donald B. Shepard

LTR Retrotransposons Contribute to Genomic Gigantism in Plethodontid Salamanders

Among vertebrates, most of the largest genomes are found within the salamanders, a clade of amphibians that includes 613 species. Salamander genome sizes range from ∼14 to ∼120 Gb. Because genome size is correlated with nucleus and cell sizes, as well as other traits, morphological evolution in salamanders has been profoundly affected by genomic gigantism. However, the molecular mechanisms driving genomic expansion in this clade remain largely unknown. Here, we present the first comparative analysis of transposable element (TE) content in salamanders. Using high-throughput sequencing, we generated genomic shotgun data for six species from the Plethodontidae, the largest family of salamanders. We then developed a pipeline to mine TE sequences from shotgun data in taxa with limited genomic resources, such as salamanders. Our summaries of overall TE abundance and diversity for each species demonstrate that TEs make up a substantial portion of salamander genomes, and that all of the major known types of TEs are represented in salamanders. The most abundant TE superfamilies found in the genomes of our six focal species are similar, despite substantial variation in genome size. However, our results demonstrate a major difference between salamanders and other vertebrates: salamander genomes contain much larger amounts of long terminal repeat (LTR) retrotransposons, primarily Ty3/gypsy elements. Thus, the extreme increase in genome size that occurred in salamanders was likely accompanied by a shift in TE landscape. These results suggest that increased proliferation of LTR retrotransposons was a major molecular mechanism contributing to genomic expansion in salamanders.

Lineage diversification and historical demography of a sky island salamander, Plethodon ouachitae, from the Interior Highlands

Sky islands provide ideal opportunities for understanding how climatic changes associated with Pleistocene glacial cycles influenced species distributions, genetic diversification, and demography. The salamander Plethodon ouachitae is largely restricted to high‐elevation, mesic forest on six major mountains in the Ouachita Mountains. Because these mountains are separated by more xeric, low‐elevation valleys, the salamanders appear to be isolated on sky islands where gene flow among populations on different mountains may be restricted. We used DNA sequence data along with ecological niche modelling and coalescent simulations to test several hypotheses related to diversifications in sky island habitats. Our results revealed that P. ouachitae is composed of seven well‐supported lineages structured across six major mountains. The species originated during the Late Pliocene, and lineage diversification occurred during the Middle Pleistocene in a stepping stone fashion with a cyclical pattern of dispersal to a new mountain followed by isolation and divergence. Diversification occurred primarily on an east–west axis, which is likely related to the east–west orientation of the Ouachita Mountains and the more favourable cooler and wetter environmental conditions on north slopes compared to south‐facing slopes and valleys. All non‐genealogical coalescent methods failed to detect significant population expansion in any lineages. Bayesian skyline plots showed relatively stable population sizes over time, but indicated a slight to moderate amount of population growth in all lineages starting approximately 10 000–12 000 years ago. Our results provide new insight into sky island diversifications from a previously unstudied region, and further demonstrate that climatic changes during the Pleistocene had profound effects on lineage diversification and demography, especially in species from environmentally sensitive habitats in montane regions.

Phylogeographic and demographic effects of Pleistocene climatic fluctuations in a montane salamander, Plethodon fourchensis.

Climatic changes associated with Pleistocene glacial cycles profoundly affected species distributions, patterns of interpopulation gene flow, and demography. In species restricted to montane habitats, ranges may expand and contract along an elevational gradients in response to environmental fluctuations and create high levels of genetic variation among populations on different mountains. The salamander Plethodon fourchensis is restricted to high‐elevation, mesic forest on five montane isolates in the Ouachita Mountains. We used DNA sequence data along with ecological niche modelling and coalescent simulations to test several hypotheses related to the effects of Pleistocene climatic fluctuations on species in montane habitats. Our results revealed that P. fourchensis is composed of four well‐supported, geographically structured lineages. Geographic breaks between lineages occurred in the vicinity of major valleys and a narrow high‐elevation pass. Ecological niche modelling predicted that environmental conditions in valleys separating most mountains are suitable; however, interglacial periods like the present are predicted to be times of range expansion in P. fourchensis. Divergence dating and coalescent simulations indicated that lineage diversification occurred during the Middle Pleistocene via the fragmentation of a wide‐ranging ancestor. Bayesian skyline plots showed gradual decreases in population size in three of four lineages over the most recent glacial period and a slight to moderate amount of population growth during the Holocene. Our results not only demonstrate that climatic changes during the Pleistocene had profound effects on species restricted to montane habitats, but comparison of our results for P. fourchensis with its parapatric, sister taxon, P. ouachitae, also emphasizes how responses can vary substantially even among closely related, similarly distributed taxa.

Reptile road mortality around an oasis in the Illinois corn desert with emphasis on the endangered eastern Massasauga

Abstract Roads have numerous negative ecological effects on terrestrial fauna, and vehicular mortality can have significant demographic consequences for some species. We studied road mortality of reptiles around Carlyle Lake, Clinton County, Illinois, USA, from April 2000 through November 2002, to assess the impact of vehicular traffic and identify influential factors. Carlyle Lake, a popular tourism/recreation area, is situated in a larger agricultural landscape and is home to the largest Illinois population of the endangered Eastern Massasauga (Sistrurus catenatus). We documented 321 cases of reptile road mortality (84 individuals of six turtle species and 237 individuals of nine snake species) while driving our approx. 46 km study route roundtrip daily. Turtle road mortality was highest in May and June, and positively associated with precipitation and minimum daily temperature. Colubrid snake road mortality was highest in April and October, and positively associated with minimum daily temperature. We recorded 42 cases of road mortality of S. catenatus with the highest number occurring from mid-August to mid-September. Road mortality in S. catenatus was biased toward adult males, which show an increase in movement in August, coinciding with the peak of the mating season and a period of high tourist visitation. The traffic intensity on a road segment did not significantly affect the level of road mortality, but segments through high quality habitats had higher levels of mortality than segments through lower quality habitats. Based on our study on the ecology of S. catenatus, we make recommendations to reduce road mortality that should aid in the conservation of the Carlyle Lake population.

Body size, age, growth and alternative mating tactics in toads: satellite males are smaller but not younger than calling males

Body size–age relationships may have important implications for the evolution and maintenance of alternative mating tactics, but these relationships remain largely unexplored in anuran amphibians (frogs and toads). In a single territorial anuran species, smaller noncalling ‘satellite’ males are younger than calling males. These results have led to the general assumption that such an age difference also holds for other anuran species with body size differences among behavioural phenotypes. However, individuals of many organisms show different developmental and/or growth trajectories that correspond to variation in adult morphology and behaviour. To test the generality of smaller satellite males being younger than larger calling males in anurans, we examined body size–age–behaviour relationships in Woodhouse’s toads, Bufo woodhousii, and Great Plains toads, B. cognatus. Age estimates, based on skeletochronologic analysis of lines of arrested growth (LAGs) from phalanges, indicated that males sampled while adopting a noncalling satellite tactic were smaller, but not younger, than males sampled while adopting a calling tactic. Furthermore, examination of inter-LAG areas of bone deposition indicated that smaller satellite male B. woodhousii showed lower growth rates relative to calling males. A similar trend occurred for B. cognatus .W e discuss our results in the context of developmental patterns associated with alternative mating tactics in other species and endocrine regulation of behavioural tactics in anurans.

Ecology of Anolis nitens brasiliensis in Cerrado Woodlands of Cantao

Abstract We studied the ecology of Anolis nitens brasiliensis during late-dry and early-wet season 2005 in a Cerrado habitat in Tocantins state, Brazil. Most lizards were found on tree trunks or leaf litter in non-flooded igapó forest. Most were found in shade or filtered sun on both cloudy and sunny days. Body temperatures (Tbs) averaged 30.6°C and did not vary among microhabitats. Microhabitats exposed to direct sun consistently reached extremely high temperatures whereas microhabitats in shade or filtered sun provided temperatures throughout the day allowing lizard activity. Nineteen prey categories were found in lizard stomachs, but the diet was dominated by spiders, crickets/grasshoppers, ants, and beetles. Although lizards that ate large prey ate fewer prey, no correlation existed between size or number of prey and lizard body size (SVL). Males were larger in SVL and mass than females, and males had relatively longer hind limbs than females. Females were variable but larger in body width. In general, the ecology of Anolis n. brasiliensis is similar to that of its Amazonian relatives, with the exception that it lives in a more thermally extreme environment and is active at slightly higher Tbs. Ecological traits of this lizard, particularly its reliance on relatively low Tb for activity, suggest that it might be particularly vulnerable to local extinction if its habitat is altered. The presence and apparent widespread distribution of A. n. brasiliensis in the Cerrado provides further evidence that the “vanishing refuge” theory cannot account for geographical patterns of distribution in the A. nitens complex.

Molecular systematics and historical biogeography of tree boas (Corallus spp.)

Inferring the evolutionary and biogeographic history of taxa occurring in a particular region is one way to determine the processes by which the biodiversity of that region originated. Tree boas of the genus Corallus are an ancient clade and occur throughout Central and South America and the Lesser Antilles, making it an excellent group for investigating Neotropical biogeography. Using sequenced portions of two mitochondrial and three nuclear loci for individuals of all recognized species of Corallus, we infer phylogenetic relationships, present the first molecular analysis of the phylogenetic placement of the enigmatic C. cropanii, develop a time-calibrated phylogeny, and explore the biogeographic history of the genus. We found that Corallus diversified within mainland South America, via over-water dispersals to the Lesser Antilles and Central America, and via the traditionally recognized Panamanian land bridge. Divergence time estimates reject the South American Caribbean-Track as a general biogeographic model for Corallus and implicate a role for events during the Oligocene and Miocene in diversification such as marine incursions and the uplift of the Andes. Our findings also suggest that recognition of the island endemic species, C. grenadensis and C. cookii, is questionable as they are nested within the widely distributed species, C. hortulanus. Our results highlight the importance of using widespread taxa when forming and testing biogeographic hypotheses in complex regions and further illustrate the difficulty of forming broadly applicable hypotheses regarding patterns of diversification in the Neotropical region.

Male Movement and Body Size Affect Mate Acquisition in the Eastern Massasauga (Sistrurus Catenatus)

Abstract Knowledge of mating system characteristics can elucidate forces driving sexual selection. In male pitvipers, both male movement tactics and body size are predicted to be important determinants of reproductive success. We used radio telemetry to monitor free-ranging Sistrurus catenatus (Eastern Massasauga) from 2000 through 2002 to determine whether male movement tactics and body size affect mate acquisition. Reproductive behaviors peaked in late July to early August. Females were accompanied by multiple males per season (up to seven); however, male mate acquisition varied considerably with only three of 17 (18%) males located more than one female during a single mating season. During the mating season, male mean daily distance moved (21.8 m) and movement frequency (77%) were higher than during the nonmating season (13.3 m, 63%). Male movement rate and body mass were positively related to the number of females acquired, and heavier males were observed accompanying females as the mating season progressed. Our results indicate that both movement tactics and body size are important in the mating system of S. catenatus; however, direct measures of reproductive success will be necessary before assessing the intensity of sexual selection.

Gastrointestinal helminths of six sympatric species of Leptodactylus from Tocantins state, Brazil.

Abstract A total of 107 leptodactylid frogs representing 6 species, Leptodactylus fuscus (n  =  15), Leptodactylus leptodactyloides (n  =  9), Leptodactylus mystaceus (n  =  2), Leptodactylus ocellatus (n  =  31), Leptodactylus petersii (n  =  31), and Leptodactylus pustulatus (n  =  19), collected in the Brazilian state of Tocantins, were examined for helminths. One species of Digenea (Catadiscus marinholutzi), 6 species of Nematoda (adults of Cosmocerca podicipinus, Ochoterenella scalaris, Oswaldocruzia proencai, and Schrankiana larvata and larvae of Brevimulticaecum sp. and Physaloptera sp.), and 1 species of Acanthocephala (as cystacanths) were found. The mean number of helminth species infecting each frog species was 3.3 ± 2.1 SD, with a range of 1–7. Ten new host records are reported.

From Foam to Free-living: Ecology of Larval Leptodactylus Labyrinthicus

Abstract Members of the genus Leptodactylus deposit eggs in foam nests; however, species vary in nest deposition sites, which may be aquatic or terrestrial, and the amount of time larvae remain in the nest. Life history characteristics are hypothesized to have a phylogenetic basis, but data to evaluate this idea are lacking for most species of Leptodactylus. We studied the ecology of tadpoles of Leptodactylus labyrinthicus in the Brazilian Cerrado to gain insight into the reproductive ecology of this species. Foam nests of L. labyrinthicus were deposited on land generally along the periphery of small, shallow ponds. Internal nest temperatures were lower, less variable, and less influenced by the environment than nest surface temperatures. Only a small percentage of eggs in each nest were fertilized, and tadpoles grew and developed within the foam while feeding on unfertilized eggs. Heavy rainfall washed the majority of tadpoles out of nests and into ponds. Tadpoles collected from the ponds had eggs in their guts, and feeding experiments confirmed that free-living tadpoles of L. labyrinthicus eat eggs of other frog species. The reproductive mode of L. labyrinthicus is intermediate between two previously described modes for closely related species. This mode may be adaptive in the unpredictable and seasonal environment in which this species occurs. Todos os membros do gênero Leptodactylus depositam ovos em ninhos de espuma; se os ninhos são aquáticos ou terrestres e até qual estágio as larvas desenvolvem-se neles, contudo, varia entre espécies. Acredita-se que os modos reprodutivos de anuros têm uma base filogenética, mas para a maioria das espécies não existem dados suficientes para avaliar essa hipótese. Nós estudamos a ecologia dos girinos de Leptodactylus labyrinthicus no Cerrado brasileiro com o intuito de tentar compreender melhor sua ecologia reprodutiva. Os ninhos de espuma de L. labyrinthicus foram depositados sobre o chão ao longo da periferia de poças pequenas e rasas. As temperaturas internas dos ninhos foram mais baixas e variaram menos do que as temperaturas externas. Apenas uma pequena porcentagem dos ovos em cada ninho foi fertilizada e os girinos permaneceram dentro da espuma, crescendo e desenvolvendo-se através da ingestão de ovos. Chuvas fortes dissolveram os ninhos e lavaram os girinos, carregando-os para as poças. Girinos coletados nas lagoas possuiam ovos em seus intestinos, e experimentos confirmaram que girinos de L. labyrinthicus alimentam-se dos ovos de outras espécies após abandonarem os ninhos. O modo reprodutivo de L. labyrinthicus é intermediário a dois outros modos descritos para espécies próximas. Esse modo pode ser adaptativo em ambientes sazonais e imprevisiveis onde essa espécie ocorre.