Plumage coloration analyses
To objectively describe plumage colour variation within T. ruficapillus , we obtained reflectance spectra measurements from museum skins deposited at the MACN (Table S3). We measured five males and four females of T. r. cochabambae and 30 males and 18 females ofT. r. ruficapillus , which are the only two subspecies present in Argentina and available in the MACN collection. We used only adult specimens that were in excellent preservation condition for reflectance measurements.
Male plumage in this species is mostly brown (T. r. cochabambaeand T. r. ruficapillus ) or dark gray (T. r. jaczewskii ,T. r. marcapatae and T. r. subfasciatus ), with rufous wings, a rufous or chestnut cap and barred underparts and tail. The species is sexually dichromatic, with brown females in all subspecies (i.e., no gray plumage) and either lack or very faint barring in their underparts and no white marking in their tales (Figure S2). Taking into account this colour patterning, we measured reflectance in six plumage patches on each specimen: throat, breast, belly, crown, nape and back. All reflectance measurements were performed with an Ocean Optics USB 2000 spectrometer (Ocean Optics Inc., Dunedin, Florida) with a PX-2 pulsed xenon light source (effective range of emission from 220 to 750 nm), calibrated against a WS-1 diffuse reflectance white standard (Ocean Optics, Inc., Dunedin, Florida, USA). Plumage was illuminated and reflectance data were collected with a bifurcated probe housed in a prismatic holder that was held against the chosen region on the study skin. The probe was held with an angle of 90º to the surface of the plumage patch. The diameter of the circular measured area was approximately 6 mm, and the distance between the probe and the plumage was 23 mm.
We performed analyses using the pavo package in R (Maia et al. 2013). Because this species is sexually dichromatic, males and females were analyzed separately. We evaluated colour differentiation in the chromatic component between subspecies for each plumage patch by estimating a perceptual distance (∆S) using the Vorobyev and Osorio (1998) colour discrimination model. Colour perceptual distances are expressed in terms of just noticeable differences (jnd) and a value of 1.0 jnd represents the theoretical threshold for discrimination of two colours, meaning that it is the distance in the perceptual colour space at which two colours would be visually discernible (Barreira et al. 2021). Thus, ∆S > 1.0 jnd indicates significant colour discrimination by the birds in all light conditions (Vorobyev et al. 1998, Siddiqi et al. 2004). In addition, we determined the statistical significance of the estimated ∆S values for each plumage patch comparison between subspecies (Maia and White 2018). For this, we estimated the geometric average of ∆S values for all comparisons performed between subspecies and calculated the 95% confidence interval (CI) through a bootstrap analysis with the bootcoldist function of pavo in R. We also tested the significance of these comparisons through a PERMANOVA analysis with the adonis function of the vegan package in R (Oksanen et al. 2008).