DOI: 10.5061/dryad.0zpc866x8
title: Evolutionary drivers of sexual signal variation in Amazon slender anoles

description: Phenotypic variation among populations, as seen in the signaling traits of
 many species, provides an opportunity to test whether similar factors
 generate repeated phenotypic patterns in different parts of a species’
 range. We investigated whether genetic divergence, abiotic gradients, and
 sympatry with closely related species explain variation in the dewlap
 colors of Amazon Slender Anoles, Anolis fuscoauratus. To this aim, we
 characterized dewlap diversity in the field with respect to population
 genetic structure and evolutionary relationships, assessed whether dewlap
 phenotypes are associated with climate or landscape variables, and tested
 for non-random associations in the distributions of A. fuscoauratus
 phenotypes and sympatric Anolis species. We found that dewlap colors vary
 among but not within sites in A. fuscoauratus. Regional genetic clusters
 included multiple phenotypes, while populations with similar dewlaps were
 often distantly related. Phenotypes did not segregate in environmental
 space, providing no support for optimized signal transmission at a local
 scale. Instead, we found a negative association between certain phenotypes
 and sympatric Anolis species with similar dewlap color attributes,
 suggesting that interactions with closely related species promoted dewlap
 divergence among A. fuscoauratus populations. Amazon Slender Anoles emerge
 as a promising system to address questions about parallel trait evolution
 and the contribution of signaling traits to speciation.
description: <em>Brief description of the methods for data
 collection:</em> <strong>Dewlap color
 variation data</strong> To characterize geographic dewlap
 color variation in <em>Anolis fuscoauratus</em>, we used data
 from our comprehensive herpetofaunal inventories in Amazonia and the
 Atlantic Forest over the last two decades. To this purpose, we sampled
 individuals by hand or pitfall traps. We only included in our
 environmental and species co-occurrence analyses 32 sites for which dewlap
 color information was available
 (pictures or field notes) from the 63 sites that were included in genetic
 analyses.
 We also obtained data on the presence of
 all other anole species sympatric with <em>A.
 fuscoauratus</em> at a given site based on our field inventory data.
 To reduce the chance of undetected species, we only included data from
 surveys that lasted a minimum of one week and involved at least three
 herpetologists searching for animals both night and day. The final dataset
 included occurrence data for 11 other anole species at the 32 sites
 for which <i>A. fuscoauratus</i> dewlap
 coloration data were available: <i>Anolis auratus</i>,
 <i>A. chrysolepis</i>, <i>A. dissimilis</i>,
 <i>A. nasofrontalis</i>, <i>A. ortonii</i>,
 <i>A. planiceps</i>, <i>A. punctatus</i>,
 <i>A. scypheus</i>, <i>A. tandai</i>, <i>A.
 trachyderma</i>, and <i>A.
 transversalis.</i>
 <strong>Genetic data</strong> A
 double-digest restriction site associated DNA library (ddRAD) was
 generated at the University of Wisconsin Biotechnology Center. Briefly,
 DNA extractions were digested with the restriction enzymes PstI and MspI,
 and the resulting fragments were tagged with individual barcodes,
 PCR-amplified, multiplexed, and sequenced in a single lane on an Illumina
 HiSeq 2500 platform. The number of paired-end reads ranged from ~1.15 to
 8.85 million per individual, with a read length of 100 base pairs.
 De-multiplexed raw sequence data were deposited in the Sequence Read
 Archive (BioProject PRJNA492310; BioSample accessions
 SAMN18340748-18340924). We used Ipyrad v. 0.7.30 to
 de-multiplex and assign reads to individuals based on sequence barcodes
 (allowing no mismatches from individual barcodes), perform de novo read
 assembly (minimum clustering similarity threshold = 0.95), align reads
 into loci, and call single nucleotide polymorphisms (SNPs). A minimum
 Phred quality score (= 33), sequence coverage (= 6x), read length (= 35
 bp), and maximum proportion of heterozygous sites per locus (= 0.5) were
 enforced, while ensuring that variable sites had no more than two alleles
 (i.e., a diploid genome). Moreover, for inclusion in the final datasets,
 we ensured that each locus was present in at least 70% of the sampled
 individuals. To estimate population genetic structure
 and admixture in <em>Anolis fuscoauratus</em>, we generated in
 Ipyrad a final dataset composed of 118,434 SNPs at 16,368 loci (including
 no outgroups). A single SNP was then extracted from each locus to minimize
 sampling of linked SNPs. We used VCFtools v. 0.1.16 to filter out SNPs
 whose minor allele frequency (MAF) was lower than 0.05.
 To estimate phylogenetic relationships,
 we generated in Ipyrad a second dataset composed of 135,952 SNPs at 17,302 RAD loci
 (now including outgroup taxa and linked SNPs), ensuring that
 each locus was present
 in at least 70% of the sampled individuals.
 
 <strong>Environmental data</strong>
 We used 17 variables in
 environmental analyses: cover of
 evergreen broadleaf trees, deciduous broadleaf trees, shrubs, herbaceous
 vegetation, and regularly flooded vegetation, annual cloud cover,
 elevation, slope, terrain roughness, and terrain
 ruggedness, all
 obtained from the EarthEnv database. As climatic variables, we used annual mean
 temperature, maximum temperature of the warmest month, mean temperature of
 the warmest quarter, annual precipitation, precipitation of the wettest
 month, and precipitation of the wettest quarter, obtained
 from the Chelsa database, as well as the climatic moisture index, a metric of relative
 wetness, obtained
 from the ENVIREM
 database.
 Values were
 extracted for each environmental variable from the 32 sites for which
 <i>A. fuscoauratus</i> dewlap color information was available
 in QGIS v.
 3.4.5. Please
 refer to the manuscript's Material and Methods for details on the
 analyses. Please send questions to ivanprates [at]
 gmail [dot] com
description: This repository contains information associated with the
 manuscript entitled: <strong>Evolutionary drivers
 of sexual signal variation in Amazon slender
 anoles.</strong> By Ivan Prates, Annelise
 D'Angiolella, Paulo R. Melo-Sampaio, Miguel T. Rodrigues, Kevin de
 Queiroz, and Rayna C. Bell. The manuscript is currently
 (March 2021) in press in Evolution. The following
 information is provided: 1. R and Unix shell scripts
 used to prepare and filter the data and perform all analyses;
 2. Filtered and assembled genetic (ddRAD) data;
 3. Outputs from phylogenetic, genetic structure, environmental,
 and co-occurrence analyses; 4. Supplementary tables,
 figures, and text. Raw read data were deposited in the
 Sequence Read Archive and can be retrieved at: https://www.ncbi.nlm.nih.gov/sra/PRJNA492310 Supplementary files as follows: <b>Figure S1. </b>Violin plots depicting the ranges of all 17 environmental variables. <b>Table S1. </b>Locality information for <i>Anolis fuscoauratus</i> and sympatric <i>Anolis</i> species used in the co-occurrence analyses. <b>Table S2. </b>Specimen and locality information of individuals used in the genetic analyses. <b>Table S3. </b>Locality information and data used in the environmental analyses. <b>Table S4.</b> Loadings of variables used in environmental principal component analyses. <b>Text S1.</b> Protein precipitation protocol used for genomic DNA extraction. <b>Text S2.</b> Phylogenetic tree including node support values and outgroup taxa.