Evolutionary analyses of visual opsin genes in frogs and toads: Diversity, duplication, and positive selection.

Among major vertebrate groups, anurans (frogs and toads) are understudied with regard to their visual systems, and little is known about variation among species that differ in ecology. We sampled North American anurans representing diverse evolutionary and life histories that likely possess visual systems adapted to meet different ecological needs. Using standard molecular techniques, visual opsin genes, which encode the protein component of visual pigments, were obtained from anuran retinas. Additionally, we extracted the visual opsins from publicly available genome and transcriptome assemblies, further increasing the phylogenetic and ecological diversity of our dataset to 33 species in total. We found that anurans consistently express four visual opsin genes (RH1, LWS, SWS1, and SWS2, but not RH2) even though reported photoreceptor complements vary widely among species. The proteins encoded by these genes showed considerable sequence variation among species, including at sites known to shift the spectral sensitivity of visual pigments in other vertebrates and had conserved substitutions that may be related to dim-light adaptation. Using molecular evolutionary analyses of selection (dN/dS) we found significant evidence for positive selection at a subset of sites in the dim-light rod opsin gene RH1 and the long wavelength sensitive cone opsin LWS. The function of sites inferred to be under positive selection are largely unknown, but a few are likely to affect spectral sensitivity and other visual pigment functions based on proximity to previously identified sites in other vertebrates. We also found the first evidence of visual opsin duplication in an amphibian with the duplication of the LWS gene in the African bullfrog, which had distinct LWS copies on the sex chromosomes suggesting the possibility of sex-specific visual adaptation. Taken together, our results indicate that ecological factors, such as habitat and life history, as well as behavior, may be driving changes to anuran visual systems.

The roles of inter- and intra-sexual selection in behavioral isolation between native and invasive pupfishes.

Male-male competition and female mate choice may both play important roles in driving and maintaining reproductive isolation between species. When previously allopatric species come into secondary contact with each other due to introductions, they provide an opportunity to evaluate the identity and strength of reproductive isolating mechanisms. If reproductive isolation is not maintained, hybridization may occur. We examined how reproductive isolating mechanisms mediate hybridization between endemic populations of the Red River pupfish Cyprinodon rubrofluviatilis and the recently introduced sheepshead minnow C. variegatus. In lab-based dominance trials, males of both species won the same number of competitions. However, male C. rubrofluviatilis that won competitions were more aggressive than C. variegatus winners, and more aggression was needed to win against competitor C. variagatus than allopatric C. rubrofluviatilis. Duration of fights also differed based on the relatedness of the competitor. In dichotomous mate choice trials, there were no conspecific or heterospecific preferences expressed by females of either species. Our findings that male-male aggression differs between closely and distantly related groups, but female choice does not suggest that male-male competition may be the more likely mechanism to impede gene flow in this system.

Preference for conspecifics evolves earlier in males than females in a sexually dimorphic radiation of fishes.

Speciation by sexual selection is generally modeled as the coevolution of female preferences and elaborate male ornaments leading to behavioral (sexual) reproductive isolation. One prediction of these models is that female preference for conspecific males should evolve earlier than male preference for conspecific females in sexually dimorphic species with male ornaments. We tested that prediction in darters, a diverse group of freshwater fishes with sexually dimorphic ornamentation. Focusing on the earliest stages of divergence, we tested preference for conspecific mates in males and females of seven closely related species pairs. Contrary to expectation, male preference for conspecific females was significantly greater than female preference for conspecific males. Males in four of the 14 species significantly preferred conspecific females; whereas, females in no species significantly preferred conspecific males. Relationships between the strength of preference for conspecifics and genetic distance revealed no difference in slope between males and females, but a significant difference in intercept, also suggesting that male preference evolves earlier than females'. Our results are consistent with other recent studies in darters and suggest that the coevolution of female preferences and male ornaments may not best explain the earliest stages of behavioral isolation in this lineage.

Differences in spectral sensitivity within and among species of darters (genus Etheostoma).

We examined variation in the visual system both within and among seven species of darters, colorful freshwater fishes of the genus Etheostoma. Using microspectrophotometry, we found that darters possess rod photoreceptor cells, single cone photoreceptor cells containing middle wavelength sensitive (MWS) visual pigments, and twin photoreceptor cells containing (LWS) visual pigments. No variation in peak sensitivity was detected among species or individuals in the rod class. In the MWS class, significant variation was detected among species and a strong statistical trend suggests differences among individuals. By contrast, all differences in the LWS class could be attributed to variation among individuals. Patterns of variation detected among species, among individuals, and among cone classes suggest that complex patterns of selection may be shaping the visual system of these fishes. Further, differences among individuals may have important consequences for visually based behaviors.

Fin-mutant female zebrafish (Danio rerio) exhibit differences in association preferences for male fin length.

Females often choose to associate with males that have exaggerated traits. In fishes, this may reflect an overall preference for larger size in a potential mate. Female zebrafish (Danio rerio) prefer males with larger bodies but not longer fins. The availability of mutant and transgenic strains of zebrafish make this a unique model system in which to study the role of phenotypic variation in social and sexual behavior. We used mutant strains of zebrafish with truncated (short fin) and exaggerated (long fin) fins to further examine female preferences for fin length in dichotomous association tests. Wild type females showed no preferences between wild type males and short fin mutant males or between wild type males and long fin mutant males. short fin females also showed no preference for short fin males or wild type males while long fin females preferred to associate with long fin males over wild type males. These results suggest that the single gene long fin mutation that results in altered fin morphological may also be involved in a related female association preference.

Conservation and conflict between endangered desert fishes.

Conservation of naturally sympatric endangered species requires unique considerations. While impacts of invasive species garner much attention, interactions between endangered species must also be managed. The endangered Leon Springs pupfish, Cyprinodon bovinus, has suffered a population decline due to decreasing natural habitat. As breeding habitat is lost, C. bovinus is also adversely affected by the sympatric, endangered Pecos gambusia, Gambusia nobilis. Here, we document interactions between these species, finding significantly more G. nobilis accumulated at pupfish spawning events than randomly distributed on breeding grounds in the absence of spawning. As a known egg predator, our results suggest that G. nobilis presence at spawnings may further decrease pupfish numbers while also altering the evolutionary dynamics of C. bovinus breeding tactics. Habitat restoration may decrease Gambusia concentrations or influence C. bovinus breeding behaviour and increase the number of territorial males resulting in viable population sizes for both critically endangered fishes.