Alex Richter-Boix

Local selection modifies phenotypic divergence among Rana temporaria populations in the presence of gene flow

In ectotherms, variation in life history traits among populations is common and suggests local adaptation. However, geographic variation itself is not a proof for local adaptation, as genetic drift and gene flow may also shape patterns of quantitative variation. We studied local and regional variation in means and phenotypic plasticity of larval life history traits in the common frog Rana temporaria using six populations from central Sweden, breeding in either open‐canopy or partially closed‐canopy ponds. To separate local adaptation from genetic drift, we compared differentiation in quantitative genetic traits (QST) obtained from a common garden experiment with differentiation in presumably neutral microsatellite markers (FST). We found that R. temporaria populations differ in means and plasticities of life history traits in different temperatures at local, and in FST at regional scale. Comparisons of differentiation in quantitative traits and in molecular markers suggested that natural selection was responsible for the divergence in growth and development rates as well as in temperature‐induced plasticity, indicating local adaptation. However, at low temperature, the role of genetic drift could not be separated from selection. Phenotypes were correlated with forest canopy closure, but not with geographical or genetic distance. These results indicate that local adaptation can evolve in the presence of ongoing gene flow among the populations, and that natural selection is strong in this system.

A comparative study of predator-induced phenotype in tadpoles across a pond permanency gradient

In a field survey the distribution of pond-breeding anuran species and their potential large predators was investigated along a freshwater habitat gradient, ranging from ephemeral pools to permanent ponds. In a laboratory experiment predator-induced plasticity was examined for all tadpole species to test whether the plastic response of ephemeral and temporary pond species differs from that of permanent pond species. Desiccation and predation pose conflicting demands; reduced activity lowers the risk of death by predation but increases the risk of death by desiccation. It was expected that species from time-constrained habitats would display a morphotype that would reduce vulnerability to invertebrate predators, thus allowing these species to maintain a high level of activity, whereas species from permanent ponds would avoid predation both morphologically and behaviourally. Species distribution and predator composition along the hydroperiod gradient differed. Variations between ephemeral and temporary ponds can be attributed to hydroperiod differences and the presence of large invertebrate predators in temporary ponds, whereas the contrasts between temporary and permanent ponds can only be attributed to the hydroperiod, since the presence and abundance of top predators are similar in both habitat types. With the exception of bufonids, all species showed predator-induced plasticity in agreement with previous studies. Tadpole species differed in the integration of the phenotypic traits measured, but differences observed between species could not be attributed only to habitat. Species from temporary habitats showed an expected response, with a low reduction of activity in comparison with the rest of the species. The lack of general patterns in the morphological changes suggests that species within the same habitat type did not converge on similar phenotypes, perhaps due to functional constraints on differences in microhabitat use in the water column.

Evolution and plasticity of anuran larval development in response to desiccation. A comparative analysis

Anurans breed in a variety of aquatic habitats with contrasting levels of desiccation risk, which may result in selection for faster development during larval stages. Previous studies suggest that species in ephemeral ponds reduce their developmental times to minimize desiccation risks, although it is not clear how variation in desiccation risk affects developmental strategies in different species. Employing a comparative phylogenetic approach including data from published and unpublished studies encompassing 62 observations across 30 species, we tested if species breeding in ephemeral ponds (High risk) develop faster than those from permanent ponds (Low risk) and/or show increased developmental plasticity in response to drying conditions. Our analyses support shorter developmental times in High risk, primarily by decreasing body mass at metamorphosis. Plasticity in developmental times was small and did not differ between groups. However, accelerated development in High risk species generally resulted in reduced sizes at metamorphosis, while some Low risk species were able compensate this effect by increasing mean growth rates. Taken together, our results suggest that plastic responses in species breeding in ephemeral ponds are constrained by a general trade-off between development and growth rates.

Breeding phenology of an amphibian community in a Mediterranean area

We studied the temporal breeding patterns and strategies of anuran assemblages in the Mediterranean region over five consecutive years. We collected monthly data on the number of clutches, tadpoles and juveniles presence of six species in 98 ponds. The data showed a great temporal segregation of species. Species using permanent ponds have a breeding peak that is related to temperature whereas reproductive success in temporary pond breeders is determined by rainfall pulses. Many species showed great plasticity of reproduction with two peaks: one in spring and the other in autumn. In spite of this temporal segregation, a large overlap was observed among species during the larval phase period. Three species (Alytes obstetricans, Pelodytes punctatus and Rana perezi) have over-wintering tadpoles. We discuss temporal segregation, differences between species in their breeding strategies and variable conditions between years as factors that favour the temporal coexistence of species in the Mediterranean region.

Do anuran larvae respond behaviourally to chemical cues from an invasive crayfish predator? A community-wide study.

Antipredator behaviour is an important fitness component in most animals. A co-evolutionary history between predator and prey is important for prey to respond adaptively to predation threats. When non-native predator species invade new areas, native prey may not recognise them or may lack effective antipredator defences. However, responses to novel predators can be facilitated by chemical cues from the predators’ diet. The red swamp crayfish Procambarus clarkii is a widespread invasive predator in the Southwest of the Iberian Peninsula, where it preys upon native anuran tadpoles. In a laboratory experiment we studied behavioural antipredator defences (alterations in activity level and spatial avoidance of predator) of nine anurans in response to P. clarkii chemical cues, and compared them with the defences towards a native predator, the larval dragonfly Aeshna sp. To investigate how chemical cues from consumed conspecifics shape the responses, we raised tadpoles with either a tadpole-fed or starved crayfish, or dragonfly larva, or in the absence of a predator. Five species significantly altered their behaviour in the presence of crayfish, and this was largely mediated by chemical cues from consumed conspecifics. In the presence of dragonflies, most species exhibited behavioural defences and often these did not require the presence of cues from predation events. Responding to cues from consumed conspecifics seems to be a critical factor in facilitating certain behavioural responses to novel exotic predators. This finding can be useful for predicting antipredator responses to invasive predators and help directing conservation efforts to the species at highest risk.

Genetic analysis of differentiation among breeding ponds reveals a candidate gene for local adaptation in Rana arvalis

One of the main questions in evolutionary and conservation biology is how geographical and environmental features of the landscape shape neutral and adaptive genetic variation in natural populations. The identification of genomic polymorphisms that account for adaptive variation can aid in finding candidate loci for local adaptation. Consequently, a comparison of spatial patterns in neutral markers and loci under selection may help disentangle the effects of gene flow, genetic drift and selection at the landscape scale. Many amphibians breed in wetlands, which differ in environmental conditions and in the degree of isolation, enhancing the potential for local adaptation. We used microsatellite markers to measure genetic differentiation among 17 local populations of Rana arvalis breeding in a network of wetlands. We found that locus RC08604 deviated from neutral expectations, suggesting that it is a good candidate for directional selection. We used a genetic network analysis to show that the allele distribution in this locus is correlated with habitat characteristics, whereas this was not the case at neutral markers that displayed a different allele distribution and population network in the study area. The graph approach illustrated the genomic heterogeneity (neutral loci vs. the candidate locus for directional selection) of gene exchange and genetic divergence among populations under directional selection. Limited gene flow between wetlands was only observed at the candidate genomic region under directional selection. RC08604 is partially located inside an up‐regulated thyroid‐hormone receptor (TRβ) gene coordinating the expression of other genes during metamorphosis and appears to be linked with variation in larval life‐history traits found among R. arvalis populations. We suggest that directional selection on genes coding larval life‐history traits is strong enough to maintain the divergence in these genomic regions, reducing the effective recombination of locally adapted alleles but not in other regions of the genome. Integrating this knowledge into conservation plans at the landscape scale will improve the design of management strategies to preserve adaptive genetic diversity in wetland networks.

Fine‐grained adaptive divergence in an amphibian: genetic basis of phenotypic divergence and the role of nonrandom gene flow in restricting effective migration among wetlands

Adaptive ecological differentiation among sympatric populations is promoted by environmental heterogeneity, strong local selection and restricted gene flow. High gene flow, on the other hand, is expected to homogenize genetic variation among populations and therefore prevent local adaptation. Understanding how local adaptation can persist at the spatial scale at which gene flow occurs has remained an elusive goal, especially for wild vertebrate populations. Here, we explore the roles of natural selection and nonrandom gene flow (isolation by breeding time and habitat choice) in restricting effective migration among local populations and promoting generalized genetic barriers to neutral gene flow. We examined these processes in a network of 17 breeding ponds of the moor frog Rana arvalis, by combining environmental field data, a common garden experiment and data on variation in neutral microsatellite loci and in a thyroid hormone receptor (TRβ) gene putatively under selection. We illustrate the connection between genotype, phenotype and habitat variation and demonstrate that the strong differences in larval life history traits observed in the common garden experiment can result from adaptation to local pond characteristics. Remarkably, we found that haplotype variation in the TRβ gene contributes to variation in larval development time and growth rate, indicating that polymorphism in the TRβ gene is linked with the phenotypic variation among the environments. Genetic distance in neutral markers was correlated with differences in breeding time and environmental differences among the ponds, but not with geographical distance. These results demonstrate that while our study area did not exceed the scale of gene flow, ecological barriers constrained gene flow among contrasting habitats. Our results highlight the roles of strong selection and nonrandom gene flow created by phenological variation and, possibly, habitat preferences, which together maintain genetic and phenotypic divergence at a fine‐grained spatial scale.

Hierarchical competition in pond-breeding anuran larvae in a Mediterranean area

The anuran larval guild is frequently characterised by the co-occurrence, with high niche overlap, of distinct species in the same pond at variables densities during development. Anuran larvae have therefore been widely studied as a model system for competition. Body size and activity level are considered the most important factors that influence the outcome of competition between tadpoles. As species from temporary ponds normally show higher activity levels in order to achieve rapid growth and thus reduce the risk of desiccation, these species are often considered superior competitors. We designed several laboratory experiments to examine the intra- and interspecific effects on growth rate, mass at metamorphosis and survival to metamorphosis of six species in a Mediterranean area. Body size and activity level were used as explanatory covariables to determine competitive ability among species. An asymmetric and hierarchical relationship was found among the six species. Larger tadpole species were more successful in competitive interactions than smaller ones, but no relationship was found between activity level and competition effects. Species typically found in temporary ponds (Pelodytes punctatus and Bufo calamita) were considered poor competitors in contrast with other communities studied. Species with low competitive ability can persist by using refuges in which competition is reduced (e.g. ephemeral ponds).

Local divergence of thermal reaction norms among amphibian populations is affected by pond temperature variation

Although temperature variation is known to cause large‐scale adaptive divergence, its potential role as a selective factor over microgeographic scales is less well‐understood. Here, we investigated how variation in breeding pond temperature affects divergence in multiple physiological (thermal performance curve and critical thermal maximum [CTmax]) and life‐history (thermal developmental reaction norms) traits in a network of Rana arvalis populations. The results supported adaptive responses to face two main constraints limiting the evolution of thermal adaptation. First, we found support for the faster–slower model, indicating an adaptive response to compensate for the thermodynamic constraint of low temperatures in colder environments. Second, we found evidence for the generalist–specialist trade‐off with populations from colder and less thermally variable environments exhibiting a specialist phenotype performing at higher rates but over a narrower range of temperatures. By contrast, the local optimal temperature for locomotor performance and CTmax did not match either mean or maximum pond temperatures. These results highlight the complexity of the adaptive multiple‐trait thermal responses in natural populations, and the role of local thermal variation as a selective force driving diversity in life‐history and physiological traits in the presence of gene flow.

Transgenerational phenotypic plasticity links breeding phenology with offspring life‐history

The timing of seasonal life-history events is assumed to evolve to synchronize life cycles with the availability of resources. Temporal variation in breeding time can have severe fitness consequences for the offspring, but the interplay between adult reproductive decisions and offspring phenotypes remains poorly understood. Transgenerational plasticity (TGP) is a potential mechanism allowing rapid responses to environmental change. Here, we investigated if experimentally delayed breeding induces TGP in larval life-history traits in the moor frog (Rana arvalis). We found clear evidence of TGP in response to changes in breeding phenology: delayed breeding increased offspring development and growth rates in the absence of external cues. This constitutes the first unequivocal evidence for TGP in response to changes in breeding phenology in vertebrates. TGP can play an important role in adjusting offspring life-history strategies to the environment they are most likely to encounter, and may constitute an important mechanism for coping with climate change.