Asymmetric Response of Costa Rican White-Breasted Wood-Wrens (Henicorhina leucosticta) to Vocalizations from Allopatric Populations.

Divergence in song between allopatric populations can contribute to premating reproductive isolation in territorial birds. Song divergence is typically measured by quantifying divergence in vocal traits using audio recordings, but field playback experiments provide a more direct way to behaviorally measure song divergence between allopatric populations. The White-breasted Wood-Wren (Henicorhina leucosticta; hereafter "WBWW") is an abundant Neotropical species with four mitochondrial clades (in Central America, the Darién, the Chocó and the Amazon) that are deeply divergent (~5-16% sequence divergence). We assessed the possibility that the WBWW as currently defined may represent multiple biological species by conducting both statistical analysis of vocal characters and field playback experiments within three clades (Central America, Chocó and Amazon). Our analysis of vocal traits revealed that Central American songs overlapped in acoustic space with Chocó songs, indicating vocal similarity between these two populations, but that Central American songs were largely divergent from Amazonian songs. Playback experiments in the Caribbean lowlands of Costa Rica revealed that Central American WBWWs typically responded aggressively to songs from the Chocó population but did not respond to playback of songs from the Amazonian population, echoing the results of the vocal trait analysis. This marked difference in behavioral response demonstrates that the songs of Central American and Amazonian WBWWs (but not Central American and Chocó WBWWs) have diverged sufficiently that Central American WBWWs no longer recognize song from Amazonian WBWWs as a signal to elicit territorial defense. This suggests that significant premating reproductive isolation has evolved between these two populations, at least from the perspective of the Central American population, and is consistent with the possibility that Central American and Amazonian populations represent distinct biological species. We conclude by advocating for the further use of field playback experiments to assess premating reproductive isolation (and species limits) between allopatric songbird populations, a situation where behavioral systematics can answer questions that phylogenetic systematics cannot

Loading scores for the first two principal components (PC1 and PC2, which explained 47% and 19% of variation, respectively) from the vocal trait analysis.

<p>Loading scores for the first two principal components (PC1 and PC2, which explained 47% and 19% of variation, respectively) from the vocal trait analysis.</p

Spectrograms showing representative song phrases from White-breasted Wood-Wren populations.

<p>(A) Central American (Costa Rica, ML3922); (B) Amazon (ML46962); and (C) Chocó (XC17276).</p

Percent of individuals approaching the speaker in each experiment.

<p>Responses to the Amazon experiment are on the left and responses to the Chocó experiment are on the right. Note that sympatric trials serve as positive controls (wrens approached the speaker in all sympatric trials).</p

Range map of the White-breasted Wood-Wren.

<p>This map illustrates the distributions of four deeply divergent mitochondrial clades (A = Central American, B = Darién, C = Chocó, D = Amazon; the precise distribution of the Darién clade is unknown and marked in dashed lines). The location of La Selva Biological Station in the Caribbean lowlands of Costa Rica, where fieldwork for this study was completed, is marked by a star. Map from [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0144949#pone.0144949.ref023" target="_blank">23</a>].</p

Summary of vocal traits for three populations of WBWWs.

<p>Vocal trait values are presented as mean +/- standard deviation.</p

Behavioral response to playback experiments.

<p>Responses to the Amazon experiment are on the left and responses to the Chocó experiment are on the right. Latency to approach (A), closest approach to speaker (B), total number of songs (C) and latency to vocalize (D). Boxplots illustrate median (horizontal black bar), first and third quartiles (boxes), and minimum and maximum values (points and dotted lines). Raw data are plotted as points in front of the boxplots, with points offset slightly to better display values. Sympatric treatments elicited aggressive responses (fast approaches, close approaches, many songs, low latency to vocalize) in both experiments. Allopatric treatments elicited asymmetric responses: individuals typically responded aggressively to playback of Chocó songs but not to playback of Amazonian songs.</p

PCA plot of acoustic space.

<p>Larger values on PC1 represent songs that are longer, that have more notes (that are shorter) and that cover a wider range of frequencies. Larger values on PC2 represent songs with faster note rates and smaller frequency ranges. Populations, shown in yellow (Costa Rica), red (Amazon) and blue (Chocó), are somewhat differentiated in acoustic space.</p

Summary of subspecies and their clades based on [21].

<p>Birds from western Colombia are presumed to be within the Chocó clade pending further evidence.</p