Supplementary information of: Beyond basic genomics: morphology, DNA barcodes, and genomic gene flow analyses reject multiple speciations within a highly structured Iberian ant-eating spider

Genomic data boost our understanding of evolution, shedding new light on long-standing debates, including species delimitation. However, we still lack a solid framework to perform reliable genome-based species delineation. A common way is through a combination of phylogenetic inference, genomic clustering, and multispecies coalescent species delimitation (PCD approach). Here we assess the species limits and the phylogeography of <i>Zodarion alacre</i>, an ant-eating spider widely distributed across the Iberian Peninsula. We first performed species delimitation based on the PCD approach and then validated its results using ecological and morphological information, DNA barcodes, and gene flow analyses. The PCD approach detected four distinct lineages with poor signs of admixture, which present allopatric distributions. These lineages showed ecological differentiation, although no morphological divergence, a low phylogenetic signal in a nuclear barcode, and evidence of introgression in a mitochondrial barcode. Additionally, phylogenomic networks found evidence of substantial gene flow between three of the four lineages. Finally, the phylogeographic methods highlighted remarkable isolation by distance and detected evidence of range expansion of <i>Z. alacre</i> from south-central Portugal to central-north Spain. We conclude that despite their deep genomic differentiation, the lineages of <i>Z. alacre</i> did not show evidence of complete speciation. Our results support the PCD approach as a solid and straightforward starting point for genome-wide species delimitation, but also that additional evidence is needed to maximize the reliability of the delineations. If genomic data are still used as standalone evidence, gene flow analyses could help detect signs of conspecificity.