DOI: 10.5061/dryad.b2rbnzsm1
title: The role of glaciations in the evolutionary history of a widely distributed Neotropical open habitat bird

description: Aim: The Neotropics constitute the most biodiverse region of the world,
 yet its patterns of diversification and speciation differ among
 Neotropical areas and are not equally well understood. Particularly, avian
 evolutionary processes are understudied in the open habitats of temperate
 South America, where the role of glacial cycles is not clear. We analyzed
 the evolutionary history of a Neotropical widespread bird species as a
 case study to evaluate its continental-scale patterns and processes of
 diversification, with a focus on Patagonia. Location: Open
 habitats of the Neotropics. Taxon: Vanellus chilensis (Aves,
 Charadriiformes). Methods: We obtained reduced representation
 genomic and mitochondrial data from the four subspecies of V. chilensis to
 perform a phylogenetic/phylogeographic analysis and study the evolutionary
 history of the species. We complemented these analyses with the study of
 vocalizations, a reproductive signal in birds. Results: The initial
 diversification event within V. chilensis, approximately 600,000 years
 ago, split a Patagonian lineage from one containing individuals from the
 rest of the Neotropics. We found considerable gene flow between these two
 lineages and a contact zone in northern Patagonia and showed that genomic
 admixture extends to northwestern Argentina. Shallower divergence was
 detected between the two non-Patagonian subspecies, which are separated by
 the Amazon River. Vocalizations were significantly different between the
 two main lineages and were intermediate in their temporal and frequency
 characteristics in the contact zone. Main conclusions: Patagonian
 populations of V. chilensis are clearly differentiated from those of the
 rest of the Neotropics, possibly as a consequence of Pleistocene
 glaciations. A secondary contact zone in northern Patagonia with extensive
 gene flow among lineages appears to be the consequence of post-glacial,
 northward expansion of the Patagonian populations. Future analyses focused
 on the dynamics of the contact zone will allow us to establish whether the
 species continues to diverge or is homogenizing. 
description: We performed double-digest
 restriction site-associated DNA sequencing (ddRADseq; Peterson et al.,
 2012) following the protocol detailed in Thrasher et al. (2018). Genomic
 DNA from fresh tissue and blood samples was isolated using the DNeasy
 tissue extraction kit (Qiagen, CA, USA). For each individual, we isolated
 ~500 ng of genomic DNA at a standardized concentration of 20 ng/µl
 (quantified using a Qubit fluorometer; Life Technologies, NY, USA). DNA
 was digested with the restriction enzymes SbfI High Fidelity (8 base bp
 recognition site; 5’-CCTGCAGG-3’) and MspI (4 base bp recognition site;
 5’-CCGG-3’) (New England BioLabs, MA, USA). The 5' and 3' ends
 of the digested genomic DNA were ligated to P1 and P2 adapters,
 respectively. Sequences of these adapters are available in Peterson et al.
 (2012). Samples with unique P1 barcodes were pooled into different
 indexing groups postdigestion/ligation and cleaned up using 1.59 Agencourt
 AMPure XP beads (Beckman Coulter, CA, USA). Samples were ordered in these
 groups according to their DNA concentration, independently of their
 geographic location or taxonomy. We size-selected fragments between 450
 and 700 bp using Blue Pippin (Sage Science, MA, USA). To incorporate the
 full Illumina TruSeq primer sequences and unique indexing primers into
 each library, we performed low cycle number PCR with Phusion High-Fidelity
 DNA Polymerase (New England BioLabs), with the following thermocycling
 profile: 98°C for 30 sec followed by 11 cycles at 98°C for 5 sec, 60°C for
 25 sec, and 72°C for 10 sec with a final extension at 72°C for 5 min. We
 visualized the product of this amplification on a 1% agarose gel and
 performed a final 0.73 AMPure to eliminate DNA fragments smaller than 200
 bp. Finally, the index groups were combined at equimolar ratios and
 sequenced on an Illumina HiSeq 2500 lane at the Cornell University
 Biotechnology Resource Center, obtaining single-end 150-bp sequences.
 We
 demultiplexed raw reads and applied standard quality filters in ipyrad
 0.7.28 (Eaton & Overcast, 2020). We only exported loci that were
 present in at least 80% of the samples to generate the final nuclear DNA
 dataset used for analyses.
description: Vanellus chilensis ddRAD data \-\-\- LOCI file containing the ddRAD data
 of 45 samples of the Southern Lapwing (Vanellus chilensis) collected in
 Brazil, Argentina, Uruguay and Chile. Title of the manuscript: The role of
 the glaciations in the evolutionary history of a widely distributed
 Neotropical open habitat bird. Authors: Belén Bukowski, Leonardo Campagna,
 María José Rodríguez-Cajarville, Gustavo S. Cabanne, Pablo L. Tubaro and
 Darío A. Lijtmaer.