Genetic analysis of cavefish reveals molecular convergence in the evolution of albinism.

The genetic basis of vertebrate morphological evolution has traditionally been very difficult to examine in naturally occurring populations. Here we describe the generation of a genome-wide linkage map to allow quantitative trait analysis of evolutionarily derived morphologies in the Mexican cave tetra, a species that has, in a series of independent caves, repeatedly evolved specialized characteristics adapted to a unique and well-studied ecological environment. We focused on the trait of albinism and discovered that it is linked to Oca2, a known pigmentation gene, in two cave populations. We found different deletions in Oca2 in each population and, using a cell-based assay, showed that both cause loss of function of the corresponding protein, OCA2. Thus, the two cave populations evolved albinism independently, through similar mutational events.

Parallel speciation in Astyanax cave fish (Teleostei) in Northern Mexico.

We investigated differentiation processes in the Neotropical fish Astyanax that represents a model system for examining adaptation to caves, including regressive evolution. In particular, we analyzed microsatellite and mitochondrial data of seven cave and seven surface populations from Mexico to test whether the evolution of the cave fish represents a case of parallel evolution. Our data revealed that Astyanax invaded northern Mexico across the Trans-Mexican Volcanic Belt at least three times and that populations of all three invasions adapted to subterranean habitats. Significant differentiation was found between the cave and surface populations. We did not observe gene flow between the strongly eye and pigment reduced old cave populations (Sabinos, Tinaja, Pachon) and the surface fish, even when syntopically occurring like in Yerbaniz cave. Little gene flow, if any, was found between cave populations, which are variable in eye and pigmentation (Micos, Chica, Caballo Moro caves), and surface fish. This suggests that the variability is due to their more recent origin rather than to hybridization. Finally, admixture of the young Chica cave fish population with nuclear markers from older cave fish demonstrates that gene flow between populations that independently colonized caves occurs. Thus, all criteria of parallel speciation are fulfilled. Moreover, the microsatellite data provide evidence that two co-occurring groups with small sunken eyes and externally visible eyes, respectively, differentiated within the partly lightened Caballo Moro karst window cave and might represent an example for incipient sympatric speciation.

Population genetic patterns revealed by microsatellite data challenge the mitochondrial DNA based taxonomy of Astyanax in Mexico (Characidae, Teleostei).

Astyanax has become an important model system for evolutionary studies of cave animals. We investigated correlations of population genetic patterns revealed by microsatellite data and phylogeographic patterns shown by mitochondrial DNA sequences in Mexican cave and surface fish of the genus Astyanax (Characidae, Teleostei) to improve the understanding of the colonization history of this neotropical fish in Central and North America and to assess a recent taxonomic classification. The distribution of nuclear genotypes is not congruent with that of the mitochondrial clades. Admixture analyses suggest there has been nuclear gene flow between populations defined by different mitochondrial clades. The microsatellite data indicate that there was mitochondrial capture of a cave population from adjacent populations. Furthermore, gene flow also occurred between populations belonging to different nuclear genotypic clusters. This indicates that neither the nuclear genotypic clusters nor the mitochondrial clades represent independent evolutionary units, although the mitochondrial divergences are high and in a range usually characteristic for different fish species. This conclusion is supported by the presence of morphologically intermediate forms. Our analyses show that the Trans-Mexican Volcanic Belt limited gene flow, but has been crossed by Astyanax several times. In Yucatán, where obvious geographic barriers are missing, the incongruence between the distribution of nuclear and mitochondrial markers reflects random colonization events caused by inundations or marine transgressions resulting in random phylogeographic breaks. Thus, conclusions about the phylogeographic history and even more about the delimitation of species should not be based on single genetic markers.

Retinal oxygen supply shaped the functional evolution of the vertebrate eye.

The retina has a very high energy demand but lacks an internal blood supply in most vertebrates. Here we explore the hypothesis that oxygen diffusion limited the evolution of retinal morphology by reconstructing the evolution of retinal thickness and the various mechanisms for retinal oxygen supply, including capillarization and acid-induced haemoglobin oxygen unloading. We show that a common ancestor of bony fishes likely had a thin retina without additional retinal oxygen supply mechanisms and that three different types of retinal capillaries were gained and lost independently multiple times during the radiation of vertebrates, and that these were invariably associated with parallel changes in retinal thickness. Since retinal thickness confers multiple advantages to vision, we propose that insufficient retinal oxygen supply constrained the functional evolution of the eye in early vertebrates, and that recurrent origins of additional retinal oxygen supply mechanisms facilitated the phenotypic evolution of improved functional eye morphology.

Multiple photopigments from the Mexican blind cavefish, Astyanax fasciatus: a microspectrophotometric study.

The cave-dwelling (hypogean) form of the teleost Astyanax fasciatus is blind, having only subdermal eye rudiments, but nevertheless maintains intact opsin genes. Second generation offspring of a cross between these and the normally sighted surface (epigean) form inherit opsin genes from both ancestries. A study of the expressed hypogean opsins of the hybrids, in comparison to the epigean forms, was undertaken by microspectrophotometry. The hybrid population showed considerable variation in the visual pigments of double cones, with evidence for two groups of cells with lambda(max) intermediate to those of the epigean pigments. Possible explanations for these intermediate pigments are discussed, including the hypothesis that they may represent hybrid genes similar to the genes for anomalous cone pigments in humans. Evidence was also found for ultraviolet-sensitive single cones and for an additional MWS pigment.

Regressive evolution of an eye pigment gene in independently evolved eyeless subterranean diving beetles.

Regressive evolution, the reduction or total loss of non-functional characters, is a fairly common evolutionary phenomenon in subterranean taxa. However, the genetic basis of regressive evolution is not well understood. Here we investigate the molecular evolution of the eye pigment gene cinnabar in several independently evolved lineages of subterranean water beetles using maximum likelihood analyses. We found that in eyeless lineages cinnabar has an increased rate of sequence evolution, as well as mutations leading to frame shifts and stop codons, indicative of pseudogenes. These results are consistent with the hypothesis that regressive evolution of eyes proceeds by random mutations, in the absence of selection, that ultimately lead to the loss of gene function in protein-coding genes specific to the eye pathway.

[Eye structure of cavernicolous populations ofAstyanax fasciatus : Contribution to the problems of degenerative evolution]. / Der Bau des Auges cavernicoler Sippen vonAstyanax fasciatus (Characidae, Pisces) : Beitrag zur Problematik degenerativer Evolutionsprozesse.

1. The characidAstyanax fasciatus, which has good, fully pigmented eyes is the ancestral form of a series of cave descendents, whose melanin pigment and eyes are reduced almost completely. Eye rudiments of two cave populations ("Anoptichthysantrobius" = population of Pachon and "A. hubbsi" = population of Sabinos) were examined anatomically. 2. Variabilities of eye types of both populations do not differ. Those developed best show a retina, which consists of all layers typical for teleostian eyes, except for the outer nuclear and outer reticular layers. The optic nerve is connected with the brain. The most degenerate eyes of all are those which have lost the optic nerve and show some tissue of unidentified character. 3. Within the eye itself there are correlations: large ones are less degenerate than small ones. 4. Size and degree of differentiation of the eyes of these cave fish are obviously dependent on genetic and inductive correlations, but also on environmental factors, such as light. 5. The E1-hybrids of the two cave forms demonstrate that they are genetically different because of separate evolution. This difference cannot be seen in the morphology of eye rudiments: there are more similarities than differences. The phenotypical divergences of eye rudiments between the two populations as observed by Peters and Peters (1966) are non-existent, and were obviously caused by an unexpected degree of modificability. 6. The genetic basis of eyes results developmentally physiologically in a special pattern of organisation. Phenotypically it reacts on all degenerative mutations alike. Therefore eyes degenerate in a trend. This pattern of organisation seems to be different in every species. Cavernicolous Amblyopsidae, for example, show similarities of structures of degenerate eyes, but they differ notably from those ofAstyanax cave populations.

Phylogeography of surface and cave Astyanax (Teleostei) from Central and North America based on cytochrome b sequence data.

Astyanax fasciatus has become a model organism for the study of regressive and adaptive evolution in cave animals. To fully understand these processes, it is important to have background information on the systematics and phylogeography of surface and cave populations of this species. Here we investigate the phylogeography of A. fasciatus in North and Central America and also the historical biogeography of this region. Phylogenetic analysis of part of the mtDNA cytochrome b gene from 26 surface and nine cave A. fasciatus populations revealed seven major clades, which, in principle, represent geographical patterns of distribution. However, the four strongly eye and pigment reduced cave populations, Piedras, Sabinos, Tinaja, and Curva, form a separate cluster, which is not sister group to the surface populations from the same locality. Similarly the Belizean populations do not cluster with their geographic neighbors from the Yucatan. The analyses indicate that there have been recurrent invasions of surface Astyanax from the south, that were most likely influenced by major climate changes during the Pleistocene. During this period, ancestors of the strongly eye and pigment reduced cave populations were able to survive underground as thermophilic relics when the surface populations became extinct. The high level of genetic divergence among the different clades shows that differing haplotype lineages must have reinvaded the surface waters from the south and/or back-colonized them from residual habitats and also penetrated into the caves. Nested clade analyses show that recurrent gene flow as well as historic processes like past fragmentation and range expansion have influenced current populations of A. fasciatus in Central and North America. Different haplotype clades of the phylogeny are not compatible with the present taxonomy of Astyanax and, therefore, we propose the application of a single systematic unit, called A. fasciatus.