Species distributed across vast continental areas and across major biomes provide unique model systems for studies of biotic diversification, yet also constitute daunting financial, logistic and political challenges for data collection across such regions. The tree frog Dendropsophus minutus (Anura: Hylidae) is a nominal species, continentally distributed in South America, that may represent a complex of multiple species, each with a more limited distribution. To understand the spatial pattern of molecular diversity throughout the range of this species complex, we obtained DNA sequence data from two mitochondrial genes, cytochrome oxidase I (COI) and the 16S rhibosomal gene (16S) for 407 samples of D. minutus and closely related species distributed across eleven countries, effectively comprising the entire range of the group. We performed phylogenetic and spatially explicit phylogeographic analyses to assess the genetic structure of lineages and infer ancestral areas. We found 43 statistically supported, deep mitochondrial lineages, several of which may represent currently unrecognized distinct species. One major clade, containing 25 divergent lineages, includes samples from the type locality of D. minutus. We defined that clade as the D. minutus complex. The remaining lineages together with the D. minutus complex constitute the D. minutus species group. Historical analyses support an Amazonian origin for the D. minutus species group with a subsequent dispersal to eastern Brazil where the D. minutus complex originated. According to our dataset, a total of eight mtDNA lineages have ranges >100,000 km². One of them occupies an area of almost one million km² encompassing multiple biomes. Our results, at a spatial scale and resolution unprecedented for a Neotropical vertebrate, confirm that widespread amphibian species occur in lowland South America, yet at the same time a large proportion of cryptic diversity still remains to be discovered.

Stomach-flushing is a gentle treatment in analysing the diet of vertebrates. We applied this method in the study of seasonal nutrition of anurans of 15 species at an Araucaria forest habitat on the Serra Geral of Rio Grande do Sul, Brazil. Over 500 frogs and toads of different body size were handled in order to evaluate an improved protocol without narcosis using soft infusion tubes in order to avoid negative effects of the flushing procedure. The specimens were treated soon after capture and then returned to the sampling sites. Our results are discussed with reference to the few studies on stomach flushing in amphibians. The improved technique is recommended to avoid killing of numerous animals for nutritional studies, in particular because of the worldwide threatening of amphibian populations.

A new conservation strategy incorporates functional, phylogenetic, and taxonomic diversity for improved cost-effective outcomes.