Phylogenetic relationships and genetic population structure inferred from nuclear genome
Among 595,163 de novocatalog loci generated with STACKS de novo pipeline and mapped to the B. barbus genome 251,956 loci had multiple alignments, 322,820 had one alignment and 20,387 were unmapped. The ‘stacks-integrate-alignments’ procedure filtered out 371,899 loci due to mapping quality, 97,561 due to alignment coverage, and 12,020 due to percent identity, resulting in 93,296 loci retained for downstream analyses (detailed statistics on each sample including raw reads is given in Supplementary material S10).
The phylogeny of Ethiopian Labeobarbus (Figure 6A) based on 15,820 SNPs is generally congruent with that based on mtDNA data (Levin et al., 2020 and Figure 6A). Sympatric generalized and thick-lipped ecomorphs clustered together and form monophyletic lineages with high support for all studied basins/subbasins (90-100 bootstrap values) except for the Gojeb (62). The Labeobarbus lineages subdivided into two clades representing i) Eastern (L. gananensis from Genale River, Indian Ocean basin) and ii) Western (L. cf.intermedius sensu lato from the Didessa, Gojeb, Sore, and Birbir Rivers) parts of the Ethiopian Highlands (Fig. 6A). The сlade of the Western Plateau is further subdivided for the Northern subclade (Didessa River in the Blue Nile basin) and the Southern subclade that combines the Gojeb (Omo-Turkana basin), Sore, and Birbir (White Nile basin) populations. The Southern subclade is further subdivided into lineages according to geographical basin belonging to the Gojeb (Omo-Turkana) lineage and White Nile lineage comprising the Sore and Birbir populations. Remarkably, the Sore and Birbir populations also form the sister monophyletic sublineages within White Nile lineage (Fig. 6A).
An analysis of the population genetic structure revealed an optimum of four (K ) clusters that correspond to the (i) Genale, (ii) Didessa, (iii) Gojeb, and iv) White Nile populations from the Sore and Birbir Rivers (Figure 6A). Five samples from the Gojeb River have a little admixture (~10%) from the White Nile cluster. When each basin with sympatric ecomorphs was analyzed independently, only one river with subdivision by ecomorphs was revealed – the Didessa in the Blue Nile basin (Fig. 6A). Individuals from White Nile basin are further subdivided by geographical populations from the Sore and Birbir Rivers (Fig. 6A). PCA of the 2,411 SNPs confirmed six well-defined clusters that correspond to the phylogenetic and population genomics results including subdivision of generalized and thick-lipped ecomorphs in the Didessa (Fig. 6B).
All ReichF ST pairwise comparisons between inferred genetic clusters were statistically significant with values ranging from 0.21 (0.18-0.25 95%CI) between Didessa generalized ecomorph and Didessa thick-lipped ecomorph to 0.66 (0.63-0.68) between Sore and Genale (Supplementary Data S11).F STpairwise comparisons of the pairs of sympatric ecomorphs within each basin (Supplementary Data S12) was significant only for Didessa, where ecomorphs constitute well segregated genetic pools.