Amazonian rivers function as important barriers to dispersal of Amazonian birds. Studying population genetics of lineages separated by rivers may help us to uncover the dynamics of biological diversification in the Amazon. We reconstructed the phylogeography of the Wedge-billed Woodcreeper, Glyphorynchus spirurus (Furnariidae) in the Amazon basin. Sampling included 134 individuals from 63 sites distributed in eight Amazonian areas of endemism separated by major Amazonian rivers. Nucleotide sequences were generated for five genes: two mtDNA genes (1047 bp for cyt b and 1002 bp for ND2) and three nuclear genes (647 bp from the sex-linked gene ACO, 319 bp from the intron of G3PDH, and 619 bp from intron 2 of MYO). In addition, 37 individuals were randomly selected from the Rondônia and Inambari areas of endemism for genomic fingerprinting, using five ISSR primers. Our results reveal allopatric and well-supported lineages within G. spirurus with high levels of genetic differentiation (p-distances 0.9-6.3%) across opposite banks of major Amazonian rivers. The multilocus phylogenetic reconstructions obtained reveal several incongruences with current subspecies taxonomy. Within currently recognized subspecies, we found high levels of both paraphyly and genetic differentiation, indicating deep divergences and strong isolation consistent with species-level differences. ISSR fingerprinting supports the existence of genetically differentiated populations on opposite sides of the Madeira River. Molecular dating suggests an initial vicariation event isolating populations from the Guiana center of endemism during the Late Miocene/Early Pliocene, while more recent events subdivided Brazilian Shield populations during the Lower Pleistocene.
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