The dynamics of barrier locus accumulation during speciation with gene flow

Understanding how reproductive isolation (RI) evolves in the context of gene flow is central to explaining how new species arise. Theory predicts that the accumulation of barrier loci is shaped by the interplay between divergent selection, recombination, and genomic architecture, yet empirical tests spanning multiple stages of divergence at the intraspecific level remain rare. The pea aphid complex, comprising multiple sympatric biotypes specialised on different legume hosts and spanning a continuum of RI, provides an exceptional opportunity to examine how ecological adaptation drives reproductive isolation under ongoing gene flow. Using whole-genome sequencing data from 13 sympatric European biotypes, we constructed a dense, reference-genome anchored picture of the divergence continuum that controls for genetic background and recombination landscape. Joint patterns of genetic differentiation, absolute divergence and genetic diversity, as well as recombination and introgression rates, reveal a consistent signature of divergence with gene flow and allow robust identification of barrier loci. Across the divergence gradient, RI is highly polygenic, yet barrier loci are clustered and enriched in low-recombination regions and large chromosomal rearrangements, which promote strong linkage disequilibrium and coupling among barrier effects. While genome-wide FST and the number of barrier loci gradually accumulate as divergence increases, the dynamics of differentiation within barrier loci show stepwise increases consistent with theoretical "tipping points" in the accumulation of RI. Barrier loci are enriched in salivary effector, detoxification and chemosensory genes, highlighting their central role in host plant specialisation and RI. These candidate genes are dynamically recruited at different stages of divergence, and some of them are shared among independent biotype comparisons, suggesting generic functional determinants of host plant specialisation and reproductive isolation acquired via parallel evolution or introgression. Our study provides a comprehensive view of the genomic architecture and evolutionary dynamics underlying speciation with gene flow, illustrating how tightly linked, polygenic architectures can facilitate adaptation and diversification in natural populations.