Boundaries of Morphological and Molecular Variation and the Distribution of a Miniaturized Froglet, Brachycephalus nodoterga (Anura: Brachycephalidae)
Most miniaturized froglets of the genus Brachycephalus occur in isolation in slopes of mountain ranges at elevations varying from 600 to 1,800 m in the Atlantic Forest of eastern Brazil. For organisms such as Bracycephalus with spatially discontinuous distributions, a fundamental task is to determine whether observed patterns of variation are consistent with geographic differentiation among allopatric populations within a single species or are suggestive of a potential species boundary. We address this problem by focusing on continental and island population samples potentially assignable to Brachycephalus nodoterga (Anura: Brachycephalidae) from the perspective of variation in qualitative and quantitative morphological traits and DNA sequences. Population samples from continental and island populations share color characteristics, qualitative traits, and multivariate patterns of variation and covariation in cranial metric traits. Comparative analysis of DNA sequences showed the magnitude of molecular distances between B. nodoterga and Brachycephalus ephippium to be 1 order of magnitude larger than molecular distances within B. nodoterga and B. ephippium. We interpret the combined morphological and molecular evidence to indicate that continental and island population samples examined here are conspecific. Therefore, by defining species boundaries for B. nodoterga, we also established minimal estimates of its distribution.Abstract
Representative specimens assignable to Brachycephalus nodoterga from Ilha de São Sebastião, Salesópolis, and Serra da Cantareira, São Paulo, Brazil. (A) Live specimens. (B) Specimens cleared and double stained with alizarin red and Alcian blue. (C) Electron scanning micrographs. Scale bars = 1 mm.
Ordination of population samples assignable to Brachycephalus nodoterga from the localities of Salesópolis, Serra da Cantareira, and Ilha de São Sebastião in the state of São Paulo, Brazil, in the space of the first and second PCs. In these figures, each symbol represents the position of an individual from a given sample, and polygons define the limits of variation in cranial shape for each sample.
Pairwise molecular sequence comparisons calculated within samples of Brachycephalus ephippium (A–D) and within population samples assignable to B. nodogerga (E–H) for the rRNA 12S, rRNA 16S, Cyt b, and COI genes.
Pairwise molecular sequence comparisons between population samples of Brachycephalus ephippium and population samples assignable to B. nodoterga for the rRNA 12S, rRNA 16S, Cyt b, and COI genes.
Mean uncorrected molecular distances between population samples assignable to Brachycephalus nodoterga from the localities of Serra da Cantareira, Salesópolis, and Ilha de São Sebastião in the state of São Paulo, Brazil. Distances are given for the genes rRNA 12S, rRNA 16S, Cyt b, and COI.
Haplotype network for population samples assignable to Brachycephalus nodoterga for the genes (A) rRNA 12S, (D) rRNA 16S, (C) Cyt b, and (D) COI. Circle size is proportional to haplotype frequency. Each line represents one mutational step, and the cross-dashes are missing, that is, extinct or unsampled haplotypes.
(A) Molecular genealogy for the population samples assignable to Brachycephalus nodoterga from the localities of Salesópolis, Cantareira, and Ilha de São Sebastião in the state of São Paulo, Brazil. Blue horizontal bars represent the range for the node ages based on two different substitution rates for the mitochondrial genes obtained from previous date estimate (Heinicke et al., 2007). (B) Schematic global sea-level fluctuations during the past 350 kyr before present. Modified from Rohling et al. (1998).
Map showing present-day coastline, the continental island of Ilha de São Sebastião, and the adjacent mainland. The light-green contour represents the estimated landmass of the Atlantic shelf exposed by the sea-level drop (lowstand of 100 m) caused by glacial advance approximately 275 kyr ago.
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