DOI: 10.5061/dryad.1c59zw46k
title: Hidden white and black feather layers enhance plumage coloration in tanagers and other songbirds

description: Birds are renowned for their diverse and colorful plumage. Here, we
 demonstrate that vibrant plumage in the tanager
 genus Tangara is dramatically intensified by a “hidden”
 layer of achromatic (white or black) plumage concealed beneath the
 outermost colorful layer. Using hyperspectral imaging, multispectral
 photography, microspectrophotometry, reflectance spectrophotometry, and
 optical modeling, we show that hidden white and black feather layers are
 systematically distributed on the body to enhance the brightness and
 saturation of carotenoid-pigmented and structurally colored plumage,
 respectively, by increasing or decreasing the amount of backscattered
 light that interacts with pigments or nanostructures. We compare male and
 female coloration and show that sexual dichromatism in
 some Tangara carotenoid plumage stems primarily from
 white layers in males and black layers in females rather than from
 differences in carotenoid pigmentation. Finally, we find that white and
 black hidden feather layers are widespread in colorful passerines. Hidden
 feather layers likely play a previously overlooked but critical role in
 colorful plumage evolution in birds.
description: Datasets in this project include reflectance spectra collected
 using UV-vis reflectance spectrophotometry, hyperspectral imaging, and
 microspectrophotometry. Additional datasets include distributions of hue,
 saturation, and lightness values from regions-of-interest of feathers from
 multispectral images.
description: # Hidden white and black feather layers enhance plumage coloration in
 tanagers and other songbirds Dataset DOI:
 [10.5061/dryad.1c59zw46k](10.5061/dryad.1c59zw46k) ## Description of the
 data and file structure This dataset contains all data and code to
 reproduce the Main and Supplementary figures and analyses in Price-Waldman
 et al. (2025), which demonstrates that hidden layers of achromatic (white
 or black) plumage are systematically distributed on the body to enhance
 the color of carotenoid-pigmented and structurally colored plumage in
 *Tangara* tanagers and other songbirds. Broadly, this dataset contains
 three types of data: 1) Measurements of color collected with reflectance
 spectrophotometry, microspectrophotometry (MSP), hyperspectral imaging
 (HI), and multispectral photography (MP); 2) code and supporting files for
 optical modeling; and 3) measurements of morphological features of
 feathers. Data from MSP, HI, and MP capture the experimental changes in
 reflectance of the colorful regions of individual feathers placed on white
 or black achromatic backgrounds. We additionally used HI to measure the
 reflectance of white and black hidden achromatic feather layers on a
 tanager specimen with the colorful tips removed, MP to measure the
 reflectance of the achromatic regions of individual feathers, and
 spectrophotometry to measure the reflectance of the synthetic achromatic
 backgrounds that we used in experiments. ### Files and variables ####
 File: Final\_folder\_Dryad.zip **Description:**  **Essential_code/**
 contains: 1. An R script (*LayeringAnalyses_final_Dryad.R*) which contains
 all code to reproduce the Main and Supplemental figures and analyses in
 the manuscript. 2. An R source file (*model_functions.R*) which contains
 functions for optical modeling. **Essential_data/** contains
 subdirectories for data collected with hyperspectral imaging,
 multispectral photography, multispectrophotometry, reflectance
 spectrophotometry, as well as a subdirectory with files relevant to
 optical modeling. **Essential_data/** also contains a CSV with feather
 length measurements. 1. **Hyperspectral/** contains three CSV files with
 reflectance spectra (400 - 700 nm) *achromatic_feather_spectra.csv*
 contains reflectance spectra for the exposed white and black achromatic
 layers from a *Tangara chilensis* specimen with the outermost colorful
 layer of feather tips removed. *blue_feather_spectra.csv* contains
 reflectance spectra for a blue structurally colored feather placed on the
 exposed white and black layers from the *T. chilensis* specimen.
 *red_feather_spectra.csv* contains reflectance spectra for a red
 carotenoid pigmented feather placed on the exposed white and black layers
 from the *T. chilensis* specimen. Each file contains 500 reflectance
 spectra for each condition (for example, 500 reflectance spectra for the
 blue feather on the white background) and the mean reflectance.
 Reflectance spectra are smoothed (using pavo::procspec with span set to
 0.3) but otherwise unmodified. 2. **Microspectrophotometry/** contains
 *all_msp_specs_visible.csv,* which contains reflectance spectra (300 - 700
 nm) for nine barbs from each female and male feather placed on white or
 black backgrounds, as well as the average reflectance across nine barbs.
 Reflectance spectra are smoothed (using pavo::procspec with span set to
 0.3) but otherwise unmodified. *msp_transmission_measurements.csv*, which
 contains transmission measurements through the white achromatic, black
 achromatic, and blue structurally colored regions of three stacked
 feathers. Reflectance spectra are smoothed (using pavo::procspec with span
 set to 0.3) and are the average of transmission spectra across nine barbs
 (for structurally colored region) or six barbs and six barbules (for
 achromatic regions). 3. **Model/** contains
 *normalized_astaxanthin_spectra_smooth_JH.csv*, the normalized and
 smoothed absorbance spectrum for astaxanthin.
 *tangara_velia_epaulet_spec.csv*, a reflectance spectrum from the
 structurally colored epaulet of *Tangara* *velia.*
 *keratin_reflectance.csv*, a reflectance spectrum from the exposed white
 achromatic layer of *T. chilensis* that we used to approximate the
 reflectance of unpigmented keratin. Note that this spectrum is normalized
 and rescaled to a maximum reflectance of 40%. 4. **Multispectral/**
 contains  *multispectral_achromatic_barbs_HSL.csv*, measurements of hue,
 saturation, and lightness values from individual barbs of feather
 achromatic regions imaged on black backgrounds. We used these values to
 compare the reflectance of male and female feather achromatic regions for
 plumage patches where achromatic layers were sexually dichromatic.
 *multispectral_achromatic_HSL.csv*, measurements of hue, saturation, and
 lightness values from feather achromatic regions imaged on contrasting
 achromatic backgrounds (for example, white achromatic feather regions on
 black backgrounds). *multispectral_colorful_HSL.csv,* measurements of hue,
 saturation, and lightness values from feather colorful regions imaged on
 white and black synthetic achromatic backgrounds. *patch_mechanisms.csv*,
 classifications of whether the colorful feather layer for each patch in
 the dataset is colored by carotenoid pigments or structural colors. 5.
 **Spectrophotometry/** contains  *all_patch_spectra.csv*, reflectance
 spectra (300 - 700 nm) for all focal patches in this study collected with
 reflectance spectrophotometry from museum specimens. Note that these
 reflectance spectra capture the overall reflectance of each patch,
 including colorful, achromatic, and downy layers. Reflectance spectra in
 this file are the average of three repeated measurements for each patch
 and have been smoothed (using pavo::procspec with span set to 0.4).
 *Calibrite_background_spectra.csv,* measurements of the white and black
 photography standards that we used as achromatic backgrounds in our
 experiments. Columns contain reflectance spectra for the white and black
 squares on a Calibrite Colorchecker Grayscale ("CCG") and the
 second-brightest white and black squares a Calibrite Colorchecker Classic
 ("CCC"). We also include reflectance spectra of the exposed
 white and black achromatic layers on a *Tangara chilensis* specimen for
 comparison. 6. *feather_length_measurements.csv* contains measurements (in
 mm) of the length of each feather region (colorful, achromatic, and downy)
 for every feather in our dataset (we sampled three feathers from 12 focal
 patches for males and females). Each measurement is the mean of two
 measurements made independently by different authors. **Figures/**
 contains empty subdirectories *Main/* and *Supplement/*. Running the code
 in *Essential_code/LayeringAnalyses_final_Dryad.R* will populate these
 subdirectories with all the plots and tables in the Main Text and
 Supplement (note that figures were further modified in Illustrator and
 tables were slightly reorganized, but all data and results are identical).
 ## Code/software All data can be viewed with standard CSV viewing software
 (for example, Microsoft Excel). We wrote all code in R v. 4.2.0.