Divergence in mating signals predicts behavioral responses among three closely related species of wrens in the genus Campylorhynchus

Vocal divergence among closely related species is a critical factor in species recognition and reproductive isolation, both of which play a fundamental role in avian speciation. To understand this process, playback experiments have provided direct evidence of species recognition. Such experiments serve as an effective tool to assess whether song divergence prevents recognition and interaction between populations, offering indirect evidence of their potential status as distinct biological species. In this study, we investigated whether divergent songs function as premating barriers by measuring the responses of three closely related wren species—Campylorhynchus humilis, C. rufinucha, and C. capistratus Campylorhynchus humilis (Russet-naped Wren), C. rufinucha (Veracruz Wren), and C. capistratus (Rufous-backed Wren)—to conspecific and heterospecific song playbacks. Unlike many playback studies that infer premating barriers based on male responses alone, we quantified both male and female territorial responses, as well as pair-level behaviors, providing a more robust behavioral dataset than typical male-focused playback studies. According to our expectations, we found species-specific responses in all three taxa: wrens responded strongly to conspecific songs, and weakly or intermediately to heterospecific signals demonstrating that song differences are crucial in species recognition. These findings align with previously reported patterns of genetic, morphological, and vocal (songs and duets) divergence among these species. The observed responses suggest that vocalizations potentially function as premating barriers, particularly in allopatric populations, reinforcing reproductive isolation and helping to delineate species boundaries. Our results highlight the evolutionary importance of vocal divergence in the speciation process and support the use of acoustic signals to assess species limits in closely related taxa.