The effect of phylogeographic history on species boundaries: a comparative framework in Hyla tree frogs.

Because it is indicative of reproductive isolation, the amount of genetic introgression across secondary contact zones is increasingly considered in species delimitation. However, patterns of admixture at range margins can be skewed by the regional dynamics of hybrid zones. In this context, we posit an important role for phylogeographic history: hybrid zones located within glacial refugia (putatively formed during the Late-Pleistocene) should be better defined than those located in post-glacial or introduced ranges (putatively formed during the Holocene and the Anthropocene). We test this hypothesis in a speciation continuum of tree frogs from the Western Palearctic (Hyla), featuring ten identified contacts between species spanning Plio-Pleistocene to Miocene divergences. We review the rich phylogeographic literature of this group and examine the overlooked transition between H. arborea and H. molleri in Western France using a multilocus dataset. Our comparative analysis supports a trend that contacts zones resulting from post-glacial expansions and human translocations feature more extensive introgression than those established within refugial areas. Integrating the biogeographic history of incipient species, i.e. their age since first contact together with their genetic divergence, thus appears timely to draw sound evolutionary and taxonomic inferences from patterns of introgression across hybrid zones.

Folk taxonomy and indigenous names for frogs in Zululand, South Africa.

BACKGROUND: We use taxonomy to organize the world into recognizable units. Folk taxonomy deals with the naming and classification of organisms through culture. Unlike its scientific counterpart, folk taxonomy is mostly undocumented, the Zoological Code of Nomenclature does not regulate it, and the resulting names are specific to each culture. A growing body of literature is steadily shedding light on the principles underlying this pre-scientific taxonomy. Vernacular names can be an instrument to increase participation of non-scientists in biodiversity matters. In South Africa, great strides have been made in standardizing and increasing relatability of vernacular amphibian names in English and Afrikaans. However, there is a need to achieve the same with the country's autochthonous languages which are used by a majority of the population. METHODS: This study investigates amphibian-related folk taxonomy using a semi-structured interview process in KwaZulu-Natal's Zululand region and  pilots methods of applying folk taxonomy principles to compile a comprehensive list of standardized indigenous frog names. RESULTS: Folk taxonomy in Zululand is systematic, developed, and bears similarities to other indigenous taxonomies around the world. Similarities also exist between folk and scientific taxonomy. Six uninomial indigenous names were found to be used for the 58 amphibian species occurring in the study area. The 58 species were assigned individual indigenous names using folk taxonomy guidelines supplemented with guidelines for modern taxonomies. CONCLUSIONS: There is a gap in the documentation and investigation of amphibian folk taxonomy in South Africa. Standardization of indigenous frog names is required to increase their universality. Similarities between folk and modern taxonomies allow for supplementation of indigenous guidelines when compiling a comprehensive indigenous species list. Through this study, social inclusion in wildlife matters is increased, indigenous knowledge systems are promoted, and a contribution is made to the development of an indigenous South African language.

Breeding Guild Determines Frog Distributions in Response to Edge Effects and Habitat Conversion in the Brazil's Atlantic Forest.

Understanding the response of species with differing life-history traits to habitat edges and habitat conversion helps predict their likelihood of persistence across changing landscape. In Brazil's Atlantic Forest, we evaluated frog richness and abundance by breeding guild at four distances from the edge of a reserve: i) 200 m inside the forest, ii) 50 m inside the forest, iii) at the forest edge, and iv) 50 m inside three different converted habitats (coffee plantation, non-native Eucalyptus plantation, and abandoned pastures, hereafter matrix types). By sampling a dry and a wet season, we recorded 622 individual frogs representing 29 species, of which three were undescribed. Breeding guild (i.e. bromeliad, leaf-litter, and water-body breeders) was the most important variable explaining frog distributions in relation to edge effects and matrix types. Leaf-litter and bromeliad breeders decreased in richness and abundance from the forest interior toward the matrix habitats. Water-body breeders increased in richness toward the matrix and remained relatively stable in abundance across distances. Number of large trees (i.e. DBH > 15 cm) and bromeliads best explained frog richness and abundance across distances. Twenty species found in the interior of the forest were not found in any matrix habitat. Richness and abundance across breeding guilds were higher in the rainy season but frog distributions were similar across the four distances in the two seasons. Across matrix types, leaf-litter species primarily used Eucalyptus plantations, whereas water-body species primarily used coffee plantations. Bromeliad breeders were not found inside any matrix habitat. Our study highlights the importance of primary forest for bromeliad and leaf-litter breeders. We propose that water-body breeders use edge and matrix habitats to reach breeding habitats along the valleys. Including life-history characteristics, such as breeding guild, can improve predictions of frog distributions in response to edge effect and matrix types, and can guide more effective management and conservation actions.

Influence of geology and human activity on the genetic structure and demography of the Oriental fire-bellied toad (Bombina orientalis).

The Oriental fire-bellied toad (Bombina orientalis) is a commonly used study organism, but knowledge of its evolutionary history is incomplete. We analyze sequence data from four genetic markers (mtDNA genes encoding cytochrome c oxidase subunit I, cytochrome b, and 12S-16S rRNA; nuDNA gene encoding recombination activating gene 2) from 188 individuals across its range in Northeast Asia to elucidate phylogeographic patterns and to identify the historic events that shaped its evolutionary history. Although morphologically similar across its range, B. orientalis exhibits phylogeographic structure, which we infer was shaped by geologic, climatic, and anthropogenic events. Phylogenetic and divergence-dating analyses recover four genetically distinct groups of B. orientalis: Lineage 1-Shandong Province and Beijing (China); Lineage 2-Bukhan Mountain (Korea); Lineage 3-Russia, Northeast China, and northern South Korea; and Lineage 4-South Korea. Lineage 2 was previously unknown. Additionally, we discover an area of secondary contact on the Korean Peninsula, and infer a single dispersal event as the origin of the insular Jeju population. Skyline plots estimate different population histories for the four lineages: Lineages 1 and 2 experienced population decreases, Lineage 3 remained stable, while Lineage 4 experienced a sharp increase during the Holocene. The timing of the population expansion of Lineage 4 coincides with the advent of rice cultivation, which may have facilitated the increase in population size by providing additional breeding habitat.

Acute toxicity and sublethal effects of the mixture glyphosate (Roundup Active) and Cosmo-Flux 411F to anuran embryos and tadpoles of four Colombian species.

Glyphosate is the most widely used herbicide in the world with application in agriculture, forestry, industrial weed control, garden and aquatic environments. However, its use is highly controversial for the possible impact on not-target organisms, such as amphibians, which are vanishing at an alarming and rapid rate. Due to the high solubility in water and ionic nature, the glyphosate requires of surfactants to increase activity. In addition, for the control of coca (Erythroxylum coca) and agricultural weeds in Colombia, formulated glyphosate is mixed and sprayed with the adjuvant Cosmo-Flux 411F to increase the penetration and activity of the herbicide. This study evaluates the acute toxic and sublethal effects (embryonic development, tadpole body size, tadpole swimming performance) of the mixture of the formulated glyphosate Roundup Active and Cosmo-Flux 411F to anuran embryos and tadpoles of four Colombian species under 96h laboratory standard tests and microcosms, which are more similar to field conditions as they include soil, sand and macrophytes. In the laboratory, embryos and tadpoles of Engystomops pustulosus were the most tolerant (LC50 = 3904 microg a.e./L; LC50=2 799 pg a.e./L, respectively), while embryos and tadpoles of Hypsiboas crepitans (LC50=2 203 microg a.e./L; LC50=1424 microgg a.e./L, respectively) were the most sensitive. R. humboldti and R. marina presented an intermediate toxicity. Embryos were significantly more tolerant to the mixture than tadpoles, which could be likely attributed to the exclusion of chemicals by the embryonic membranes and the lack of organs, such as gills, which are sensitive to surfactants. Sublethal effects were observed for the tadpole body size, but not for the embryonic development and tadpole swimming performance. In microcosms, no toxicity (LC50 could not be estimated), or sublethal responses were observed at concentrations up to fourfold (14.76 kg glyphosate a.e./ha) the highest field application rate of 3.69 kg glyphosate a.e./ha. Thus, toxicity was less in the microcosms than in laboratory tests, which may be attributed to the presence of sediments and organic matter which rapidly adsorb glyphosate and surfactants such as POEA. It is concluded that the mixture of glyphosate (Roundup Active) and Cosmo-Flux*411F, as used in the field, has a negligible toxic effect to embryos and tadpoles of the species tested in this study.

Acute toxicity and sublethal effects of the mixture glyphosate (Roundup® Active) and Cosmo-Flux®411F to anuran embryos and tadpoles of four Colombian species / Toxicidad aguda y efectos subletales de la mezcla glifosato (Roundup® Activo) y Cosmo-Flux®411F en embriones y renacuajos de cuatro especies de anuros colombianos

Glyphosate is the most widely used herbicide in the world with application in agriculture, forestry, industrial weed control, garden and aquatic environments. However, its use is highly controversial for the possible impact on not-target organisms, such as amphibians, which are vanishing at an alarming and rapid rate. Due to the high solubility in water and ionic nature, the glyphosate requires of surfactants to increase activity. In addition, for the control of coca (Erythroxylum coca) and agricultural weeds in Colombia, formulated glyphosate is mixed and sprayed with the adjuvant Cosmo-Flux®411F to increase the penetration and activity of the herbicide. This study evaluates the acute toxic and sublethal effects (embryonic development, tadpole body size, tadpole swimming performance) of the mixture of the formulated glyphosate Roundup® Active and Cosmo-Flux®411F to anuran embryos and tadpoles of four Colombian species under 96h laboratory standard tests and microcosms, which are more similar to field conditions as they include soil, sand and macrophytes. In the laboratory, embryos and tadpoles of Engystomops pustulosus were the most tolerant (LC50=3 904µg a.e./L; LC50=2 799µg a.e./L, respectively), while embryos and tadpoles of Hypsiboas crepitans (LC50=2 203µg a.e./L; LC50=1 424µg a.e./L, respectively) were the most sensitive. R. humboldti and R. marina presented an intermediate toxicity. Embryos were significantly more tolerant to the mixture than tadpoles, which could be likely attributed to the exclusion of chemicals by the embryonic membranes and the lack of organs, such as gills, which are sensitive to surfactants. Sublethal effects were observed for the tadpole body size, but not for the embryonic development and tadpole swimming performance. In microcosms, no toxicity (LC50 could not be estimated), or sublethal responses were observed at concentrations up to fourfold (14.76kg glyphosate a.e./ha) the highest field application rate of 3.69kg glyphosate a.e./ha. Thus, toxicity was less in the microcosms than in laboratory tests, which may be attributed to the presence of sediments and organic matter which rapidly adsorb glyphosate and surfactants such as POEA. It is concluded that the mixture of glyphosate (Roundup® Active) and Cosmo-Flux®411F, as used in the field, has a negligible toxic effect to embryos and tadpoles of the species tested in this study.

El glifosato es el herbicida más usado en el mundo con aplicaciones para la agricultura, control de malezas forestales, industriales, en jardines y ambientes acuáticos. Sin embargo, su uso es altamente controversial por el posible impacto sobre organismos no blanco, como los anfibios, los cuales están desapareciendo a una tasa alarmantemente rápida. Debido a su alta solubilidad en agua y naturaleza iónica, el glifosato requiere de surfactantes para incrementar su actividad. Además, para el control de Erythroxylum coca y de malezas en la agricultura en Colombia, el glifosato formulado es mezclado y rociado con el coadyuvante Cosmo-Flux®411F para incrementar la penetración y actividad del herbicida. Este estudio evalúa los efectos tóxicos agudos y subletales (desarrollo embrionario, tamaño corporal y desempeño natatorio de los renacuajos) de la mezcla del glifosato formulado Roundup® Activo con el Cosmo-Flux®411F en embriones y renacuajos de cuatro especies de anuros colombianos, bajo pruebas de 96h en condiciones estándar de laboratorio y microcosmos, que son más similares a las condiciones de campo al incluir tierra, arena y macrófitas. En laboratorio, los embriones y renacuajos de Engystomops pustulosus fueron los más tolerantes (CL50=3 904µg a.e./L; CL50=2 799µg a.e./L, respectivamente), mientras que los embriones y renacuajos de Hypsiboas crepitans fueron los más sensibles (CL50=2 203µg a.e./L; CL50=1 424µg a.e./L, respectivamente). R. humboldti y R. marina presentaron una toxicidad intermedia. Los embriones fueron más tolerantes a la mezcla que los renacuajos, lo cual podría ser atribuido a la exclusión de los químicos por las membranas embrionarias y a la falta de órganos, como las branquias, que son más sensibles a los surfactantes. Se observaron efectos subletales en el tamaño corporal de los renacuajos, pero no en el desarrollo embrionario ni el desempeño natatorio de los renacuajos. En microcosmos no se observaron efectos tóxicos ni respuestas subletales a concentraciones hasta cuatro veces (14.67kg glifosato a.e./ha) la tasa de aplicación más alta de 3.69kg glifosato a.e./ha. Por lo tanto, la toxicidad fue menor en los microcosmos que en las pruebas de laboratorio, lo que puede ser atribuido a la presencia de sedimentos y materia orgánica que absorbe rápidamente el glifosato y surfactantes como el POEA. Se concluye que la mezcla del glifosato (Roundup® Activo) y Cosmo-Flux®411F, como se aplica en campo, tiene un efecto tóxico bajo en los embriones y renacuajos de las especies estudiadas.

Components of the peptidome and transcriptome persist in lin wa pi: the dried skin of the Heilongjiang brown frog (Rana amurensis) as used in traditional Chinese medicine.

Although the ancient practice of traditional Chinese medicine (TCM) utilizes predominantly herbal ingredients, many of which are now the subject of intense scientific scrutiny, significant quantities of animal tissue-derived materials are also employed. Here we have used contemporary molecular techniques to study the material known as lin wa pi, the dried skin of the Heilongjiang brown frog, Rana amurensis, that is used commonly as an ingredient of many medicines, as a general tonic and as a topical antimicrobial/wound dressing. Using a simple technology that has been developed and validated over several years, we have demonstrated that components of both the skin granular gland peptidome and transcriptome persist in this material. Interrogation of the cDNA library constructed from the dried skin by entrapment and amplification of polyadenylated mRNA, using a "shotgun" primer approach and 3'-RACE, resulted in the cloning of cDNAs encoding the precursors of five putative antimicrobial peptides. Two (ranatuerin-2AMa and ranatuerin-2AMb) were obvious homologs of a previously described frog skin peptide family, whereas the remaining three were of sufficient structural novelty to be named amurins 1-3. Mature peptides were each identified in reverse phase HPLC fractions of boiling water extracts of skin and their structures confirmed by MS/MS fragmentation sequencing. Components of traditional Chinese medicines of animal tissue origin may thus contain biologically active peptides that survive the preparation procedures and that may contribute to therapeutic efficacy.

Comparative study of immature erythroid cells of the diploid Bufo ictericus and the tetraploid Odontophrynus americanus (Amphibia, Anura): ultrastructural cytochemical detection of nucleic acids and polysaccharides, and mapping of the element phosphorus.

In the present study, we used ultrastructural cytochemistry to analyze the distribution of nuclear and cytoplasmic nucleic acids and polysaccharides, and electron spectroscopic imaging to map the element phosphorus in immature erythroid cells taken from two amphibians, the diploid Bufo ictericus and the tetraploid Odontophrynus americanus. In the cytoplasm of cells from the tetraploid species, we detected numerous inclusions containing a material that was similar to the dispersed chromatin seen in the nucleus of these cells. The RNase-gold complex labeled both the dispersed nuclear chromatin and the cytoplasmic inclusions. The Thiéry technique showed that glycoconjugates were present in all the membranous complexes of the erythroid cells of both types of amphibians under study, although they were absent within or around the cytoplasmic RNA inclusions. Electron spectroscopic imaging revealed the presence of phosphorus in these inclusions. These data suggest that an increase in RNA synthesis occurs in tetraploid amphibian cells, probably as a result of an alteration in the mechanisms of gene regulation.