Parallel phenotypic diversification in closely related species is a rigorous framework for testing the role of natural selection in evolution. Do parallel phenotypes always diversify by parallel genetic bases or does selection pave many alternative genomic routes to the same phenotypic ends? In this review, we show that the advent of next-generation sequencing technologies and the growing use of genomic approaches make it increasingly feasible to answer these fundamental questions using ecological and evolutionary 'non-model' populations of vertebrates in nature. While it is generally expected, and often observed, that closely related populations or species have parallel genetic bases to parallel phenotypes, exceptions are not rare and show that alternative genetic routes can result in similar phenotypes. Ultimately, this framework may illuminate the ecological conditions, evolutionary histories and genetic architectures that result in recurrent phenotypes and rapid adaptation.

The polychromatic and trophically polymorphic Midas cichlid fish species complex (Amphilophus cf. citrinellus) is an excellent model system for studying the mechanisms of speciation and patterns of phenotypic diversification in allopatry and in sympatry. Here, we first review research to date on the species complex and the geological history of its habitat. We analyse body shape variation from all currently described species in the complex, sampled from six crater lakes (maximally 1.2 -23.9 kyr old) and both great lakes in Nicaragua. We find that Midas cichlid populations in each lake have their own characteristic body shape. In lakes with multiple sympatric species of Midas cichlid, each species has a distinct body shape. Across the species complex, most body shape change relates to body depth, head, snout and mouth shape and caudal peduncle length. There is independent parallel evolution of an elongate limnetic species in at least two crater lakes. Mitochondrial genetic diversity is higher in crater lakes with multiple species. Midas cichlid species richness increases with the size and age of the crater lakes, though no such relationship exists for the other syntopic fishes. We suggest that crater lake Midas cichlids follow the predicted pattern of an adaptive radiation, with early divergence of each crater lake colonization, followed by intralacustrine diversification and speciation by ecological adaptation and sexual selection.

Fundamental to understanding how biodiversity arises and adapts is whether evolution is predictable in the face of stochastic genetic and demographic factors. Here we show rapid parallel evolution across two closely related but geographically isolated radiations of Nicaraguan crater lake cichlid fishes. We find significant morphological, ecological and genetic differentiation between ecomorphs in sympatry, reflected primarily in elongated versus high-bodied shape, differential ecological niche use and genetic differentiation. These eco-morphological divergences are significantly parallel across radiations. Based on 442,644 genome-wide single nucleotide polymorphisms, we identify strong support for the monophyly of, and subsequent sympatric divergence within, each radiation. However, the order of speciation differs across radiations; in one lake the limnetic ecomorph diverged first while in the other a benthic ecomorph. Overall our results demonstrate that complex parallel phenotypes can evolve very rapidly and repeatedly in similar environments, probably due to natural selection, yet this evolution can proceed along different evolutionary genetic routes.

Crater lakes provide a natural laboratory to study speciation of cichlid fishes by ecological divergence. Up to now, there has been a dearth of transcriptomic and genomic information that would aid in understanding the molecular basis of the phenotypic differentiation between young species. We used next-generation sequencing (Roche 454 massively parallel pyrosequencing) to characterize the diversity of expressed sequence tags between ecologically divergent, endemic and sympatric species of cichlid fishes from crater lake Apoyo, Nicaragua: benthic Amphilophus astorquii and limnetic Amphilophus zaliosus. We obtained 24 174 A. astorquii and 21 382 A. zaliosus highquality expressed sequence tag contigs, of which 13 106 pairs are orthologous between species. Based on the ratio of nonsynonymous to synonymous substitutions, we identified six sequences exhibiting signals of strong diversifying selection (K a ⁄ K s > 1). These included genes involved in biosynthesis, metabolic processes and development. This transcriptome sequence variation may be reflective of natural selection acting on the genomes of these young, sympatric sister species. Based on Ks ratios and p-distances between 3¢-untranslated regions (UTRs) calibrated to previously published species divergence times, we estimated a neutral transcriptome-wide substitutional mutation rate of 1.25 · 10 )6 per site per year. We conclude that next-generation sequencing technologies allow us to infer natural selection acting to diversify the genomes of young species, such as crater lake cichlids, with much greater scope than previously possible.