The Northern Andes, acting both as barrier and corridor, exert strong influence in Neotropical biodiversity. Still, the diversification of foothill taxa inhabiting ecotonal zones between lowland rainforest and montane environments in this region remains poorly understood. We investigated the evolutionary history of Eutoxeres aquila, a hermit hummingbird distributed across Central America, the Chocó, and both Andean slopes. Using mitochondrial DNA (ND2 and cytb), morphometrics, resistance surface analyses, and ecological niche models, we examined the potential roles of topography, connectivity, and climatic oscillations in shaping lineage divergence. Phylogenetic analyses of 59 individuals of Eutoxeres aquila revealed two major clades: (i) an eastern clade (eastern Andean slope–upper Magdalena valley + mid Magdalena valley–Tamá), and (ii) a western clade (western slope–N Central cordillera + Panama). Divergence from E. condamini occurred ~4.3 million years ago (Mya), while crown E. aquila diversification began ~0.9 Mya, coinciding with Pleistocene climate cycles. The Panama lineage diverged from the western slope–N Central Cordillera ~0.4 Mya, suggesting post-Isthmus dispersal. Morphometric analyses of 69 museum specimens showed significant bill shape differentiation among regions, broadly consistent with genetic structure. Genetic distances among individuals were weakly correlated with both resistance and geographic (Euclidean) distances. Ecological niche projections indicated expanded connectivity during the Mid-Holocene and LGM, but contraction and fragmentation during the LIG, with downward elevational shifts during colder periods (LGM and mid-Holocene) and upward shifts during warmer conditions (LIG and present). These models also revealed that ~227,500 km² of long-term climatic stability areas concentrated mostly in Amazonia and the Chocó. These results highlight recurrent isolation and reconnection of mountain environments as key drivers of foothill diversification and emphasize the importance of ecotonal taxa for understanding Andean evolutionary processes.

The methods associated to this data submission are the Procrustes coordinates used for the geometrics morphometric analyses of the study. To evaluate potential continuity in the morphological traits of Eutoxeres aquila, we measured bill shape on specimens across the species' range. We focused on the bill because this structure is known to respond even to subtle differences in selective pressures (Grant et al. 1976, Krugler et al. 2025), potentially revealing discontinuities in the environmental conditions experienced by different populations. Moreover, given the complexity of the Eutoxeres bill, capturing its shape provides a two-dimensional assessment that is more informative than standard linear measurements.

The morphological sampling consisted of 69 museum specimens of E. aquila across the distribution of the species, which were classified by region according to the groups discovered in the genetic analyses (see Results): eastern slope–upper Magdalena (38 samples), mid Magdalena valley–Tamá (6 samples), western slope–N central Cordillera (22 samples), and Panama (3 samples). Collection dates ranged from 1987 to 2019, and individuals were not separated by sex due to the scarcity of specimens with sex information on museum labels. Although sexual dimorphism in bill curvature has been documented in several species of hermit hummingbirds (Phaethornithinae), E. aquila exhibits a markedly reduced degree of bill curvature dimorphism compared to other members of the subfamily, with an estimated ~4 % decrease in female-biased curvature (Temeles et al., 2010). Thus, we would not expect a substantial increase in bill shape variance driven by sexual dimorphism.

To capture the bill profile, each specimen was placed in a supine position on a flexible foam base covered with black material painted with matching watercolor to reduce bill glare. Pins were positioned around the body to immobilize it at a 90° angle to the lateral head-bill profile. Once the bill was in focus, photographs were taken in Program (P) mode without flash, with a ruler (in cm) included as a scale reference. Images were captured using a Nikon D7000 camera equipped with an AF-S MICRO NIKKOR 105 mm F/2.8G ED lens, with a minimum focusing distance of 0.50 m.

We analyzed bill shape variation using landmark-based geometric morphometric methods. From the bill profile photographs, we used the thin plate spline (TPS) series (Rohlf 2006) to digitize landmarks: three fixed points (bill tip, proximal limit of the nares, mouth gap) and two curves of 50 semi-landmarks each, traced along the dorsal outline of the maxilla and ventral outline of the mandible, respectively (Bookstein 1991, Foster et al. 2008, Zelditch et al. 2018). Semi-landmarks were allowed to slide along their respective curves during alignment to minimize bending energy. A generalized Procrustes analysis was applied to remove the effects of orientation, scale, and position, centering configurations at the origin and scaling them to unit centroid size (Shao et al. 2016).

References

Bookstein, F. L. 1991. Morphometric tools for landmark data. Cambridge Univ. Press.

Foster, D. J., Podos, J. and Hendry, A. P. 2008. A geometric morphometric appraisal of beak shape in Darwin’s finches. J. Evol. Biol. 21: 263–275.

Grant, P. R., Grant, B. R., Smith, J. N., Abbott, I. J. and Abbott, L. K. 1976. Darwin's finches: population variation and natural selection. – Proc. Natl Acad. Sci. USA 73: 257–261.

Krugler, J. D., Benham, P. M. and Bowie, R. C. 2025. Shared temporal increases in bill size among songbirds of the San Francisco Bay Area provide evidence for different seasonal selective pressures. – Ecography: e07494.

Rohlf, F. J. 2006. TpsDig 2.10. – Department of Ecology and Evolution, State Univ. of New York, Stony Brook, NY.

Shao, S., Quan, Q., Cai, T., Song, G., Qu, Y. and Lei, F. 2016. Evolution of body morphology and beak shape revealed by a morphometric analysis of 14 Paridae species. – Front. Zool. 13: 30.

Temeles, E. J., Miller, J. S. and Rifkin, J. L. 2010. Evolution of sexual dimorphism in bill size and shape of hermit hummingbirds (Phaethornithinae): a role for ecological causation. Philos. Trans. R. Soc. B 365: 1053–1063.

Zelditch, M. L., Swiderski, L. D. and Sheets, H. D. 2018. A practical companion to geometric morphometrics for biologists: running analyses in freely-available software, 2nd edn. Univ. of Michigan.