Lake Tanganyika is the oldest and phenotypically most diverse of the three East African cichlid fish adaptive radiations. It is also the cradle for the younger parallel haplochromine cichlid radiations in Lakes Malawi and Victoria. Despite its evolutionary significance, the relationships among the main Lake Tanganyika lineages remained unresolved, as did the general timescale of cichlid evolution. Here, we disentangle the deep phylogenetic structure of the Lake Tanganyika radiation using anchored phylogenomics and uncover hybridization at its base, as well as early in the haplochromine radiation. This suggests that hybridization might have facilitated these speciation bursts. Time-calibrated trees support that the radiation of Tanganyika cichlids coincided with lake formation and that Gondwanan vicariance concurred with the earliest splits in the cichlid family tree. Genes linked to key innovations show signals of introgression or positive selection following colonization of lake habitats and species’ dietary adaptations are revealed as major drivers of colour vision evolution. These findings shed light onto the processes shaping the evolution of adaptive radiations.

Lake Tanganyika, the oldest of the East African Great Lakes, harbors the ecologically, morphologically, and behaviorally most complex of all assemblages of cichlid fishes, consisting of about 200 described species. The evolutionary old age of the cichlid assemblage, its extreme degree of morphological differentiation, the lack of species with intermediate morphologies, and the rapidity of lineage formation have made evolutionary reconstruction difficult. The number and origin of seeding lineages, particularly the possible contribution of riverine haplochromine cichlids to endemic lacustrine lineages, remains unclear. Our phylogenetic analyses, based on mitochondrial DNA sequences of three gene segments of 49 species (25% of all described species, up to 2,400 bp each), yield robust phylogenies that provide new insights into the Lake Tanganyika adaptive radiation as well as into the origin of the Central- and East-African haplochromine faunas. Our data suggest that eight ancient African lineages may have seeded the Tanganyikan cichlid radiation. One of these seeding lineages, probably comprising substrate spawning Lamprologus -like species, diversified into six lineages that evolved mouthbrooding during the initial stage of the radiation. All analyzed haplochromines from surrounding rivers and lakes seem to have evolved within the radiating Tanganyikan lineages. Thus, our findings contradict the current hypothesis that ancestral riverine haplochromines colonized Lake Tanganyika to give rise to at least part of its spectacular endemic cichlid species assemblage. Instead, the early phases of the Tanganyikan radiation affected Central and East African rivers and lakes. The haplochromines may have evolved in the Tanganyikan basin before the lake became a hydrologically and ecologically closed system and then secondarily colonized surrounding rivers. Apparently, therefore, the current diversity of Central and East African haplochromines represents a relatively young and polyphyletic fauna that evolved from or in parallel to lineages now endemic to Lake Tanganyika.

Background: The tribe Lamprologini is the major substrate breeding lineage of Lake Tanganyika's cichlid species flock. Among several different life history strategies found in lamprologines, the adaptation to live and breed in empty gastropod shells is probably the most peculiar. Although shell-breeding arose several times in the evolutionary history of the lamprologines, all obligatory and most facultative shell-breeders belong to the so called "ossified group", a monophyletic lineage within the lamprologine cichlids. Since their distinctive life style enables these species to live and breed in closest vicinity, we hypothesized that these cichlids might be particularly prone to accidental hybridization, and that introgression might have affected the evolutionary history of this cichlid lineage. Results: Our analyses revealed discrepancies between phylogenetic hypotheses based on mitochondrial and nuclear (AFLP) data. While the nuclear phylogeny was congruent with morphological, behavioral and ecological characteristics, several species – usually highly specialized shell-breeders – were placed at contradicting positions in the mitochondrial phylogeny. The discordant phylogenies strongly suggest repeated incidents of introgressive hybridization between several distantly related shell-breeding species, which reticulated the phylogeny of this group of cichlids. Long interior branches and high bootstrap support for many interior nodes in the mitochondrial phylogeny argue against a major effect of ancient incomplete lineage sorting on the phylogenetic reconstruction. Moreover, we provide morphological and genetic (mtDNA and microsatellites) evidence for ongoing hybridization among distantly related shell-breeders. In these cases, the territorial males of the inferred paternal species are too large to enter the shells of their mate, such that they have to release their sperm over the entrance of the shell to fertilize the eggs. With sperm dispersal by water currents and wave action, trans-specific fertilization of clutches in neighboring shells seem inevitable, when post-zygotic isolation is incomplete. Conclusion: From the direct observation of hybrids we conclude that hybridization between distantly related gastropod-shell-breeding cichlids of Lake Tanganyika follows inevitably from their ecological specialization. Moreover, the observed incongruence between mtDNA and nuclear multilocus phylogeny suggests that repeated hybridization events among quite distantly related taxa affected the diversification of this group, and introduced reticulation into their phylogeny.

Background: Cichlid fishes are notorious for their wealth of intra- and interspecific colour pattern diversity. In Lake Tanganyika, the endemic genus Tropheus represents the most impressive example for geographic variation in the pattern and hue of integument colouration, but the taxonomy of the over 100 mostly allopatric colour morphs remains to a large degree unresolved. Previous studies of mitochondrial DNA sequence data revealed polyphyly of the six nominally described species and complex phylogeographic patterns influenced by lake level fluctuations and population admixture, and suggested the parallel evolution of similar colour patterns in divergent evolutionary lineages. A gene tree of a rapidly radiating group may be subject to incomplete and stochastic lineage sorting, and to overcome this problem we used multi-locus, nuclear AFLP data in comparison with mtDNA sequences to study diversification, migration and introgression in Tropheus colour morphs in Lake Tanganyika. Results: Significant incongruence between phylogenetic reconstructions from mitochondrial and AFLP data suggested incomplete sorting of mitochondrial haplotypes as well as frequent introgression between differentiated lineages. In contrast to the mitochondrial phylogeny, the AFLP phenogram was largely congruent with species classifications, colour pattern similarities, and in many cases also with the current geographic distribution of populations, and did not produce evidence of convergent colour pattern evolution. Homoplasy in the AFLP data was used to identify populations that were strongly affected by introgression. Conclusion: Different evolutionary processes were distinguished by the combination of mitochondrial and AFLP data. Mitochondrial phylogeographic patterns retained signals of large-scale migration events triggered by historical, major lake level fluctuations, whereas AFLP data indicated genetic cohesion among local groups of populations resulting from secondary contact of adjacent populations in the course of the more frequently occurring, minor lake level fluctuations. There was no support for the parallel evolution of similar colour patterns in the AFLP data. Genetic signatures of introgression and hybridisation detected in several populations suggest that lake level fluctuations drove the stunning diversification of Tropheus morphs not only through population fragmentation, but also by promoting hybridisation between differentiated morphs in secondary contact.

Species diverge eco-morphologically through the continuous action of natural selection on functionally important structures, producing alternative adaptive morphologies. In cichlid fishes, the oral and pharyngeal jaws are such key structures. Adaptive variation in jaw morphology contributes to trophic specialization, which is hypothesized to fuel their rapid speciation in the East African Great Lakes. Much is known about the genes involved in cichlid jaw and craniofacial development. However, it is still unclear what salient sources of variation gave rise to trophic-niche specialization, facilitating adaptive radiation. Here, we explore two sources of transcriptional variation that may underlie species-specific disparities in jaw morphology. Using whole transcriptome RNA-sequencing, we analyze differences in gene expression and alternative splicing, at the end of postlarval development, in fully functional jaws of six species of cichlids from the Lake Tanganyika tribe Tropheini. Our data reveal a surprisingly high degree of alternative splicing events compared with gene expression differences among species and trophic types. This suggests that differential trophic adaptation of the jaw apparatus may have been shaped by transcriptional rewiring of splicing as well as gene expression variation during the rapid radiation of the Tropheini. Specifically, genes undergoing splicing across most species were found to be enriched for pharyngeal jaw gene ontology terms. Overall, jaw transcriptional patterns at postlarval developmental stage were highly dynamic and species-specific. In conclusion, this work indicates that shifts in alternative splicing could have played a more important role in cichlid adaptive radiation, and possibly adaptive radiation in general, than currently recognized.