Quantifying individual variation in reaction norms: Mind the residual

Phenotypic plasticity is a central topic in ecology and evolution. Individuals may differ in the degree of plasticity (individual‐by‐environment interaction (I × E)), which has implications for the capacity of populations to respond to selection. Random regression models (RRMs) are a popular tool to study I × E in behavioural or life‐history traits, yet evidence for I × E is mixed, differing between species, populations, and even between studies on the same population. One important source of discrepancies between studies is the treatment of heterogeneity in residual variance (heteroscedasticity). To date, there seems to be no collective awareness among ecologists of its influence on the estimation of I × E or a consensus on how to best model it. We performed RRMs with differing residual variance structures on simulated data with varying degrees of heteroscedasticity and plasticity, sample size and environmental variability to test how RRMs would perform under each scenario. The residual structure in the RRMs affected the precision of estimates of simulated I × E as well as statistical power, with substantial lack of precision and high false‐positive rates when sample size, environmental variability and plasticity were small. We show that model comparison using information criteria can be used to choose among residual structures and reinforce this point by analysis of real data of two study populations of great tits (Parus major). We provide guidelines that can be used by biologists studying I × E that, ultimately, should lead to a reduction in bias in the literature concerning the statistical evidence and the reported magnitude of variation in plasticity.     

Differences in developmental strategies between long‐settled and invasion‐front populations of the cane toad in Australia

Phenotypic plasticity can enhance a species’ ability to persist in a new and stressful environment, so that reaction norms are expected to evolve as organisms encounter novel environments. Biological invasions provide a robust system to investigate such changes. We measured the rates of early growth and development in tadpoles of invasive cane toads (Rhinella marina) in Australia, from a range of locations and at different larval densities. Populations in long‐colonized areas have had the opportunity to adapt to local conditions, whereas at the expanding range edge, the invader is likely to encounter challenges that are both novel and unpredictable. We thus expected invasion‐vanguard populations to exhibit less phenotypic plasticity than range‐core populations. Compared to clutches from long‐colonized areas, clutches from the invasion front were indeed less plastic (i.e. rates of larval growth and development were less sensitive to density). In contrast, those rates were highly variable in clutches from the invasion front, even among siblings from the same clutch under standard conditions. Clutches with highly variable rates of growth and development under constant conditions had lower phenotypic plasticity, suggesting a trade‐off between these two strategies. Although these results reveal a strong pattern, further investigation is needed to determine whether these different developmental strategies are adaptive (i.e. adaptive phenotypic plasticity vs. bet‐hedging) or instead are driven by geographic variation in genetic quality or parental effects.     

The phenotypic variance gradient – a novel concept

Evolutionary ecologists commonly use reaction norms, which show the range of phenotypes produced by a set of genotypes exposed to different environments, to quantify the degree of phenotypic variance and the magnitude of plasticity of morphometric and life‐history traits. Significant differences among the values of the slopes of the reaction norms are interpreted as significant differences in phenotypic plasticity, whereas significant differences among phenotypic variances (variance or coefficient of variation) are interpreted as differences in the degree of developmental instability or canalization. We highlight some potential problems with this approach to quantifying phenotypic variance and suggest a novel and more informative way to plot reaction norms: namely “a plot of log (variance) on the y‐axis versus log (mean) on the x‐axis, with a reference line added”. This approach gives an immediate impression of how the degree of phenotypic variance varies across an environmental gradient, taking into account the consequences of the scaling effect of the variance with the mean. The evolutionary implications of the variation in the degree of phenotypic variance, which we call a “phenotypic variance gradient”, are discussed together with its potential interactions with variation in the degree of phenotypic plasticity and canalization.     

Behavioral plasticity and G × E of reproductive tactics in Nicrophorus vespilloides burying beetles

Phenotypic plasticity is important in the evolution of traits and facilitates adaptation to rapid environmental changes. However, variation in plasticity at the individual level, and the heritable basis underlying this plasticity is rarely quantified for behavioral traits. Alternative behavioral reproductive tactics are key components of mating systems but are not often considered within a phenotypic plasticity framework (i.e., as reaction norms). Here, using lines artificially selected for repeated mating rate, we test for genetic (G × E) sources of variation in reproductive behavior of male Nicrophorus vespilloides burying beetles (including signaling behavior), as well as the role of individual body size, in responsiveness to changes in social environment. The results show that body size influences the response of individuals’ signaling behavior to changes in the social environment. Moreover, there was G × E underlying the responses of males to variation in the quality of social environment experienced (relative size of focal male compared to his rival). This shows that individual variation in plasticity and social sensitivity of signaling behavior can evolve in response to selection on investment in mating behavior, with males selected for high mating investment having greater social sensitivity.     

Stressful environments induce novel phenotypic variation: hierarchical reaction norms for sperm performance of a pervasive invader

Genetic variation for phenotypic plasticity is ubiquitous and important. However, the scale of such variation including the relative variability present in reaction norms among different hierarchies of biological organization (e.g., individuals, populations, and closely related species) is unknown. Complicating interpretation is a trade‐off in environmental scale. As plasticity can only be inferred over the range of environments tested, experiments focusing on fine tuned responses to normal or benign conditions may miss cryptic phenotypic variation expressed under novel or stressful environments. Here, we sought to discern the presence and shape of plasticity in the performance of brown trout sperm as a function of optimal to extremely stressful river pH, and demarcate if the reaction norm varies among genotypes. Our overarching goal was to determine if deteriorating environmental quality increases expressed variation among individuals. A more applied aim was to ascertain whether maintaining sperm performance over a wide pH range could help explain how brown trout are able to invade diverse river systems when transplanted outside of their native range. Individuals differed in their reaction norms of phenotypic expression of an important trait in response to environmental change. Cryptic variation was revealed under stressful conditions, evidenced through increasing among‐individual variability. Importantly, data on population averages masked this variability in plasticity. In addition, canalized reaction norms in sperm swimming velocities of many individuals over a very large range in water chemistry may help explain why brown trout are able to colonize a wide variety of habitats.     

An empirical test for a zone of canalization in thermal reaction norms

Theoretical models on the evolution of phenotypic plasticity predict a zone of canalization where reaction norms cross, and genetic variation is minimized in the environment a population most frequently encounter. Empirical tests of this prediction are largely missing, in particular for life‐history traits. We addressed this prediction by quantifying thermal reaction norms of three life‐history traits (somatic growth rate, age and size at maturation) of a Norwegian population of Daphnia magna and testing for the occurrence of an intermediate temperature (T_m) at which genetic variance in the traits is minimized. Size at maturation changed relatively little with temperature compared to the other traits, and there was no genetic variance in the shape of the reaction norm. Consequently, age at maturation and somatic growth rate were strongly negatively correlated. Both traits showed a strong genotype–environment interaction, and the estimated T_m was 14 °C for both age at maturation and growth rate. This value of T_m corresponds well with mean summer temperatures experienced by the population and suggests that the population has evolved under stabilizing selection in temperatures that fluctuate around this mean temperature. These results suggest local adaptation to temperature in the studied population and allow predicting evolutionary trajectories of thermal reaction norms under changing thermal regimes.     

Size‐induced phenotypic reaction norms in a parasitoid wasp: an examination of life‐history and behavioural traits

The amount of resources available during development often affects body size, causing phenotypic variation in life‐history traits and reproductive behaviours. However, past studies have seldom examined the reaction norms of both life‐history and behavioural traits versus body size. We measured the phenotypic plasticity of several life‐history (age‐specific egg load, egg size, longevity) and behavioural (oviposition rate, host marking rate, walking speed) traits of the egg parasitoid Telenomus podisi Ashmead (Hymenoptera: Scelionidae) in response to body size variation. We predicted that life‐history traits would show more evidence of size compensation than behavioural traits, resulting in fewer positively‐sloped size versus trait reaction norms among the former. As predicted by life‐history models, smaller wasps appear to shift resource allocation towards early‐life reproduction, having a similar egg load to large individuals 9 days after emergence. Surprisingly, longevity was unaffected by body size. However, egg size, the number of offspring produced during oviposition bouts, and the rate of subsequent egg synthesis were greater for larger individuals. In addition, as predicted, the reaction norms of behavioural traits versus body size were all positively sloped. Thus, despite possible adaptive compensatory plasticity of life‐history traits by small individuals, behavioural constraints directly related to body size would contribute to maintaining a positive size–fitness relationship.     

CONTRASTING PATTERNS OF PHENOTYPIC PLASTICITY IN REPRODUCTIVE TRAITS IN TWO GREAT TIT (PARUS MAJOR) POPULATIONS

Phenotypic plasticity is an important mechanism via which populations can respond to changing environmental conditions, but we know very little about how natural populations vary with respect to plasticity. Here we use random‐regression animal models to understand the multivariate phenotypic and genetic patterns of plasticity variation in two key life‐history traits, laying date and clutch size, using data from long‐term studies of great tits in The Netherlands (Hoge Veluwe [HV]) and UK (Wytham Woods [WW]). We show that, while population‐level responses of laying date and clutch size to temperature were similar in the two populations, between‐individual variation in plasticity differed markedly. Both populations showed significant variation in phenotypic plasticity (IxE) for laying date, but IxE was significantly higher in HV than in WW. There were no significant genotype‐by‐environment interactions (GxE) for laying date, yet differences in GxE were marginally nonsignificant between HV and WW. For clutch size, we only found significant IxE and GxE in WW but no significant difference between populations. From a multivariate perspective, plasticity in laying date was not correlated with plasticity in clutch size in either population. Our results suggest that generalizations about the form and cause of any response to changing environmental conditions across populations may be difficult.     

Adaptation to marginal habitats by evolution of increased phenotypic plasticity

In an island population receiving immigrants from a larger continental population, gene flow causes maladaptation, decreasing mean fitness and producing continued directional selection to restore the local mean phenotype to its optimum. We show that this causes higher plasticity to evolve on the island than on the continent at migration-selection equilibrium, assuming genetic variation of reaction norms is such that phenotypic variance is higher on the island, where phenotypes are not canalized. For a species distributed continuously in space along an environmental gradient, higher plasticity evolves at the edges of the geographic range, and in environments where phenotypes are not canalized. Constant or evolving partially adaptive plasticity also alleviates maladaptation owing to gene flow in a heterogeneous environment and produces higher mean fitness and larger population size in marginal populations, preventing them from becoming sinks and facilitating invasion of new habitats. Our results shed light on the widely observed involvement of partially adaptive plasticity in phenotypic clines, and on the mechanisms causing geographic variation in plasticity.     

The biology hidden inside residual within‐individual phenotypic variation

Phenotypes vary hierarchically among taxa and populations, among genotypes within populations, among individuals within genotypes, and also within individuals for repeatedly expressed, labile phenotypic traits. This hierarchy produces some fundamental challenges to clearly defining biological phenomena and constructing a consistent explanatory framework. We use a heuristic statistical model to explore two consequences of this hierarchy. First, although the variation existing among individuals within populations has long been of interest to evolutionary biologists, within‐individual variation has been much less emphasized. Within‐individual variance occurs when labile phenotypes (behaviour, physiology, and sometimes morphology) exhibit phenotypic plasticity or deviate from a norm‐of‐reaction within the same individual. A statistical partitioning of phenotypic variance leads us to explore an array of ideas about residual within‐individual variation. We use this approach to draw attention to additional processes that may influence within‐individual phenotypic variance, including interactions among environmental factors, ecological effects on the fitness consequences of plasticity, and various types of adaptive variance. Second, our framework for investigating variation in phenotypic variance reveals that interactions between levels of the hierarchy form the preconditions for the evolution of all types of plasticity, and we extend this idea to the residual level within individuals, where both adaptive plasticity in residuals and canalization‐like processes (stability) can evolve. With the statistical tools now available to examine heterogeneous residual variance, an array of novel questions linking phenotype to environment can be usefully addressed.     

Genetic basis of between‐individual and within‐individual variance of docility

Between‐individual variation in phenotypes within a population is the basis of evolution. However, evolutionary and behavioural ecologists have mainly focused on estimating between‐individual variance in mean trait and neglected variation in within‐individual variance, or predictability of a trait. In fact, an important assumption of mixed‐effects models used to estimate between‐individual variance in mean traits is that within‐individual residual variance (predictability) is identical across individuals. Individual heterogeneity in the predictability of behaviours is a potentially important effect but rarely estimated and accounted for. We used 11 389 measures of docility behaviour from 1576 yellow‐bellied marmots (Marmota flaviventris) to estimate between‐individual variation in both mean docility and its predictability. We then implemented a double hierarchical animal model to decompose the variances of both mean trait and predictability into their environmental and genetic components. We found that individuals differed both in their docility and in their predictability of docility with a negative phenotypic covariance. We also found significant genetic variance for both mean docility and its predictability but no genetic covariance between the two. This analysis is one of the first to estimate the genetic basis of both mean trait and within‐individual variance in a wild population. Our results indicate that equal within‐individual variance should not be assumed. We demonstrate the evolutionary importance of the variation in the predictability of docility and illustrate potential bias in models ignoring variation in predictability. We conclude that the variability in the predictability of a trait should not be ignored, and present a coherent approach for its quantification.     

Nonlinear effects of temperature on body form and developmental canalization in the threespine stickleback

Theoretical models predict that nonlinear environmental effects on the phenotype also affect developmental canalization, which in turn can influence the tempo and course of organismal evolution. Here, we used an oceanic population of threespine stickleback (Gasterosteus aculeatus) to investigate temperature‐induced phenotypic plasticity of body size and shape using a paternal half‐sibling, split‐clutch experimental design and rearing offspring under three different temperature regimes (13, 17 and 21 °C). Body size and shape of 466 stickleback individuals were assessed by a set of 53 landmarks and analysed using geometric morphometric methods. At approximately 100 days, individuals differed significantly in both size and shape across the temperature groups. However, the temperature‐induced differences between 13 and 17 °C (mainly comprising relative head and eye size) deviated considerably from those between 17 and 21 °C (involving the relative size of the ectocoracoid, the operculum and the ventral process of the pelvic girdle). Body size was largest at 17 °C. For both size and shape, phenotypic variance was significantly smaller at 17 °C than at 13 and 21 °C, indicating that development is most stable at the intermediate temperature matching the conditions encountered in the wild. Higher additive genetic variance at 13 and 21 °C indicates that the plastic response to temperature had a heritable basis. Understanding nonlinear effects of temperature on development and the underlying genetics are important for modelling evolution and for predicting outcomes of global warming, which can lead not only to shifts in average morphology but also to destabilization of development.     

Grain of environment explains variation in the strength of genotype × environment interaction

Theory predicts that genetic variation in phenotypic plasticity (genotype × environment interaction or G × E) should be eroded by selection acting across environments. However, it appears that G × E is often maintained under selection, although not universally. This variation in the presence and strength of G × E requires explanation. Here I ask whether the explanation may lie in the grain of the environment at which G × E is expressed. The grain (or grain size) of the environment refers to the scale of environmental heterogeneity relative to generation time – that is, relative to the window of operation of selection – with higher rates of heterogeneity occurring in finer‐grained environments. The hypothesis that the grain of the environment explains variation in the expression of G × E encapsulates variation in the power of selection to shape reaction norms: selection should be able to erode G × E in fine‐grained environments but lose its power as the grain becomes coarser. I survey studies of G × E in sexual traits and demonstrate that the strength of G × E varies with the grain of the environment across which it is expressed, with G × E being stronger in coarser‐grained environments. This result elucidates when G × E is most likely to be sustained in the reaction norms of fitness‐related traits and when its evolutionary consequences will be most pronounced.     

Population differences in response to hypoxic stress in Atlantic salmon

Understanding whether populations can adapt to new environmental conditions is a major issue in conservation and evolutionary biology. Aquatic organisms are increasingly exposed to environmental changes linked with human activities in river catchments. For instance, the clogging of bottom substratum by fine sediments is observed in many rivers and usually leads to a decrease in dissolved oxygen concentrations in gravel beds. Such hypoxic stress can alter the development and even be lethal for Atlantic salmon (Salmo salar) embryos that spend their early life into gravel beds. In this study, we used a common garden experiment to compare the responses to hypoxic stress of four genetically differentiated and environmentally contrasted populations. We used factorial crossing designs to measure additive genetic variation of early life‐history traits in each population. Embryos were reared under normoxic and hypoxic conditions, and we measured their survival, incubation time and length at the end of embryonic development. Under hypoxic conditions, embryos had a lower survival and hatched later than in normoxic conditions. We found different hypoxia reaction norms among populations, but almost no population effect in both treatments. We also detected significant sire × treatment interactions in most populations and a tendency for heritability values to be lower under stressful conditions. Overall, these results reveal a high degree of phenotypic plasticity in salmon populations that nevertheless differ in their adaptive potential to hypoxia given the distinct reaction norms observed between and within populations.     

Heterozygosity predicts clutch and egg size but not plasticity in a house sparrow population with no evidence of inbreeding

We investigated the link between heterozygosity and the reaction norm attributes of reproductive performance in female house sparrows (Passer domesticus). We collected data on clutch size, egg size, hatching success and nestling survival in 2816 nesting attempts made by 791 marked individuals over a 16-year period. Pedigree analysis revealed no evidence of inbreeding. Neither parent-offspring regression nor an animal model revealed significant heritability in clutch or egg size. We selected 42 females that laid at least seven clutches at our study site and used a survey of 21 autosomal microsatellite loci to estimate heterozygosity for each female. We controlled for phenotypic plasticity and found that both clutch and egg size showed significant positive correlations with heterozygosity. We found no evidence that heterozygosity influenced the slope of individual reaction norms. Further analysis suggested that clutch size was affected by heterozygosity across the genome, but egg size had more complex relationships, with evidence favouring the influence of multiple loci. Given the apparent lack of inbreeding and large population size, our results suggest associative overdominance as the likely mechanism for the impact of heterozygosity, but also created a puzzle about the process producing associations between neutral markers and the genes affecting clutch size or egg size. One possible explanation is a long-term residual effect of the historical bottleneck that occurred when house sparrows were introduced into North America. The existence of heterozygosity-fitness correlations in a population with considerable phenotypic plasticity and little inbreeding implies that the effects of heterozygosity may be more significant than previously thought.     

CHARACTERIZING SELECTION ON PHENOTYPIC PLASTICITY IN RESPONSE TO NATURAL ENVIRONMENTAL HETEROGENEITY

Adaptive genetic differentiation and adaptive phenotypic plasticity can increase the fitness of plant lineages in heterogeneous environments. We examine the relative importance of genetic differentiation and plasticity in determining the fitness of the annual plant, Erodium cicutarium, in a serpentine grassland in California. Previous work demonstrated that the serpentine sites within this mosaic display stronger dispersal‐scale heterogeneity than nonserpentine sites. We conducted a reciprocal transplant experiment among six sites to characterize selection on plasticity expressed by 180 full‐sibling families in response to natural environmental heterogeneity across these sites. Multivariate axes of environmental variation were constructed using a principal components analysis of soil chemistry data collected at every experimental block. Simple linear regressions were used to characterize the intercept, and slope (linear and curvilinear) of reaction norms for each full‐sibling family in response to each axis of environmental variation. Multiple linear regression analyses revealed significant selection on trait means and slopes of reaction norms. Multivariate analyses of variance demonstrated genetic differentiation between serpentine and nonserpentine lineages in the expression of plasticity in response to three of the five axes of environmental variation considered. In all but one case, serpentine genotypes expressed a stronger adaptive plastic response than nonserpentine genotypes.     

Adaptive divergence in body size overrides the effects of plasticity across natural habitats in the brown trout

The evolution of life-history traits is characterized by trade-offs between different selection pressures, as well as plasticity across environmental conditions. Yet, studies on local adaptation are often performed under artificial conditions, leaving two issues unexplored: (i) how consistent are laboratory inferred local adaptations under natural conditions and (ii) how much phenotypic variation is attributed to phenotypic plasticity and to adaptive evolution, respectively, across environmental conditions? We reared fish from six locally adapted (domesticated and wild) populations of anadromous brown trout (Salmo trutta) in one semi-natural and three natural streams and recorded a key life-history trait (body size at the end of first growth season). We found that population-specific reaction norms were close to parallel across different streams and Q_ST was similar – and larger than F_ST – within all streams, indicating a consistency of local adaptation in body size across natural environments. The amount of variation explained by population origin exceeded the variation across stream environments, indicating that genetic effects derived from adaptive processes have a stronger effect on phenotypic variation than plasticity induced by environmental conditions. These results suggest that plasticity does not “swamp” the phenotypic variation, and that selection may thus be efficient in generating genetic change.     

Genetic and environmental components of growth in nestling blue tits (Parus caeruleus)

We investigated the effect of brood‐size mediated food availability on the genetic and environmental components of nestling growth in the blue tit (Parus caeruleus), using a cross‐fostering technique. We found genetic variation for body size at most nestling ages, and for duration of mass increase, but not of tarsus growth. Hence, nestling growth in our study population seems to have the potential to evolve further. Furthermore, significant genotype–environment interactions indicated heritable variation in reaction norms of growth rates and growth periods, i.e. that our study population had a heritable plasticity in the growth response to environmental conditions. The decreasing phenotypic variance with nestling age indicated compensatory growth in all body traits. Furthermore, the period of weight increase was longer for nestlings growing up in enlarged broods, while there was no difference to reduced broods in the period of tarsus growth. At fledging, birds in enlarged broods had shorter tarsi and lower weights than birds in reduced broods, but there was no difference in wing length or body condition between the two experimental groups. The observed flexibility in nestling growth suggests that growing nestlings are able to respond adaptively to food constraint by protecting the growth of ecologically important traits.     

Variation in Caenorhabditis elegans dauer larva formation

Dauer larvae of Caenorhabditis elegans are formed when young larvae experience conditions of low food availability and high conspecific population density; non-dauer, third stage larvae are formed in conditions of plenty. This developmental response to environmental conditions is an example of phenotypic plasticity; that is, an environmentally induced change in phenotype and, as such, a manifestation of a genotype-environment interaction. Extensive variation was found in reaction norms of phenotypic plasticity of dauer formation among wild lines of C. elegans. Recombinant-inbred lines were constructed from parental lines with very different reaction norms of dauer formation. These recombinant-inbred lines had a wide range of reaction norms, of a range greater than that set by the parental lines. The natural variation in reaction norms of dauer formation in C. elegans is, presumably, an adaptation to enhance fitness under the lines' different natural prevailing conditions. The genetic basis of this variation, as well as its phenotypic consequences, are now ripe for further investigation.