Inferring the evolution of reproductive isolation in a lineage of fossil threespine stickleback, Gasterosteus doryssus.

Darwin attributed the absence of species transitions in the fossil record to his hypothesis that speciation occurs within isolated habitat patches too geographically restricted to be captured by fossil sequences. Mayr's peripatric speciation model added that such speciation would be rapid, further explaining missing evidence of diversification. Indeed, Eldredge and Gould's original punctuated equilibrium model combined Darwin's conjecture, Mayr's model and 124 years of unsuccessfully sampling the fossil record for transitions. Observing such divergence, however, could illustrate the tempo and mode of evolution during early speciation. Here, we investigate peripatric divergence in a Miocene stickleback fish, Gasterosteus doryssus. This lineage appeared and, over approximately 8000 generations, evolved significant reduction of 12 of 16 traits related to armour, swimming and diet, relative to its ancestral population. This was greater morphological divergence than we observed between reproductively isolated, benthic-limnetic ecotypes of extant Gasterosteus aculeatus. Therefore, we infer that reproductive isolation was evolving. However, local extinction of G. doryssus lineages shows how young, isolated, speciating populations often disappear, supporting Darwin's explanation for missing evidence and revealing a mechanism behind morphological stasis. Extinction may also account for limited sustained divergence within the stickleback species complex and help reconcile speciation rate variation observed across time scales.

Inferred genetic architecture underlying evolution in a fossil stickleback lineage.

Inferring the genetic architecture of evolution in the fossil record is difficult because genetic crosses are impossible, the acquisition of DNA is usually impossible and phenotype-genotype maps are rarely obvious. However, such inference is valuable because it reveals the genetic basis of microevolutionary change across many more generations than is possible in studies of extant taxa, thereby integrating microevolutionary process and macroevolutionary pattern. Here, we infer the genetic basis of pelvic skeleton reduction in Gasterosteus doryssus, a Miocene stickleback fish from a finely resolved stratigraphic sequence that spans nearly 17,000 years. Reduction in pelvic score, a categorical measure of pelvic structure, resulted primarily from reciprocal frequency changes of two discrete phenotypic classes. Pelvic vestiges also showed left-side larger asymmetry. These patterns implicate Pitx1, a large-effect gene whose deletion generates left-side larger asymmetry of pelvic vestiges in extant, closely related Gasterosteus aculeatus. In contrast, reductions in the length of the pelvic girdle and pelvic spines resulted from directional shifts of unimodal, continuous trait distributions, suggesting an additional suite of genes with minor, additive pelvic effects, again like G. aculeatus. Similar genetic architectures explain shared but phyletically independent patterns across 10 million years of stickleback evolution.

Evolution toward a new adaptive optimum: phenotypic evolution in a fossil stickleback lineage.

Natural selection has almost certainly shaped many evolutionary trajectories documented in fossil lineages, but it has proven difficult to demonstrate this claim by analyzing sequences of evolutionary changes. In a recently published and particularly promising test case, an evolutionary time series of populations displaying armor reduction in a fossil stickleback lineage could not be consistently distinguished from a null model of neutral drift, despite excellent temporal resolution and an abundance of indirect evidence implicating natural selection. Here, we revisit this case study, applying analyses that differ from standard approaches in that: (1) we do not treat genetic drift as a null model, and instead assess neutral and adaptive explanations on equal footing using the Akaike Information Criterion; and (2) rather than constant directional selection, the adaptive scenario we consider is that of a population ascending a peak on the adaptive landscape, modeled as an Orstein-Uhlenbeck process. For all three skeletal features measured in the stickleback lineage, the adaptive model decisively outperforms neutral evolution, supporting a role for natural selection in the evolution of these traits. These results demonstrate that, at least under favorable circumstances, it is possible to infer in fossil lineages the relationship between evolutionary change and features of the adaptive landscape.

Directional asymmetry of pelvic vestiges in threespine stickleback.

Extensive reduction of the size and complexity of the pelvic skeleton (i.e., pelvic reduction) has evolved repeatedly in Gasterosteus aculeatus. Asymmetrical pelvic vestiges tend to be larger on the left side (i.e., left biased) in populations studied previously. Loss of Pitx1 expression is associated with pelvic reduction in G. aculeatus, and pelvic reduction maps to the Pitx1 locus. Pitx1 knockouts in mice have reduced hind limbs, but the left limb is larger. Thus left-biased directional asymmetry of stickleback pelvic vestiges may indicate the involvement of Pitx1 in pelvic reduction. We examined 6,356 specimens from 27 Cook Inlet populations of G. aculeatus with extensive pelvic reduction. Samples from 20 populations exhibit the left bias in asymmetrical pelvic vestiges expected if Pitx1 is involved, and three have a slight, non-significant left bias. However, samples from three populations have a significant right bias, and one large sample from another population has equal frequencies of specimens with larger vestiges on the left or right side. A sample of fossil threespine stickleback also has significantly left-biased pelvic vestiges. These results suggest that silencing of Pitx1 or the developmental pathway in which it functions in the pelvis is the usual cause of pelvic reduction in most Cook Inlet populations of G. aculeatu, and that it caused pelvic reduction at least 10 million years ago in a stickleback population. A different developmental genetic mechanism is implicated for three populations with right-biased pelvic vestiges and for the population without directional asymmetry.

Correlated evolution and dietary change in fossil stickleback.

The importance of trophic ecology in adaptation and evolution is well known, yet direct evidence that feeding controls microevolution over extended evolutionary time scales, available only from the fossil record, is conspicuously lacking. Through quantitative analysis of tooth microwear, we show that rapid evolutionary change in Miocene stickleback was associated with shifts in feeding, providing direct evidence from the fossil record for changes in trophic niche and resource exploitation driving directional, microevolutionary change over thousands of years. These results demonstrate the potential for tooth microwear analysis to provide powerful insights into trophic ecology during aquatic adaptive radiations.

Hybridization, transgressive segregation, genetic covariation, and adaptive radiation.

Analysis of adaptive radiation has had a central role in the development of evolutionary theory, but it is not clear why some groups radiate and others do not. Two recent papers by Albertson and colleagues on the genetic architecture of East African cichlid fishes implicate hybridization, transgressive segregation and genetic covariation in the early stages of adaptive radiation. Transgressive segregation and genetic covariation might be key innovations in genetic architecture that favor adaptive radiation.