Genetic structure of phenotypic robustness in the collaborative cross mouse diallel panel

Developmental stability and canalization describe the ability of developmental systems to minimize phenotypic variation in the face of stochastic micro‐environmental effects, genetic variation and environmental influences. Canalization is the ability to minimize the effects of genetic or environmental effects, whereas developmental stability is the ability to minimize the effects of micro‐environmental effects within individuals. Despite much attention, the mechanisms that underlie these two components of phenotypic robustness remain unknown. We investigated the genetic structure of phenotypic robustness in the collaborative cross (CC) mouse reference population. We analysed the magnitude of fluctuating asymmetry (FA) and among‐individual variation of cranial shape in reciprocal crosses among the eight parental strains, using geometric morphometrics and a diallel analysis based on a Bayesian approach. Significant differences among genotypes were found for both measures, although they were poorly correlated at the level of individuals. An overall positive effect of inbreeding was found for both components of variation. The strain CAST/EiJ exerted a positive additive effect on FA and, to a lesser extent, among‐individual variance. Sex‐ and other strain‐specific effects were not significant. Neither FA nor among‐individual variation was associated with phenotypic extremeness. Our results support the existence of genetic variation for both developmental stability and canalization. This finding is important because robustness is a key feature of developmental systems. Our finding that robustness is not related to phenotypic extremeness is consistent with theoretical work that suggests that its relationship to stabilizing selection is not straightforward.     

Ontogenetic Patterning of Skeletal Fluctuating Asymmetry in Rhesus Macaques and Humans: Evolutionary and Developmental Implications

I address the question of how fluctuating asymmetry (FA)—the distribution of random deviations from bilateral symmetry—varies ontogenetically in the mammalian skeleton. This question is significant because of the light that such patterns can shed on the causes of variation in developmental stability in bone as well as other structures. Based on large ontogenetic skeletal series of Macaca mulatta and Homo sapiens, I report that the FA variances of skeletal metric traits increase ontogenetically. Coupled with the finding that FA variances also accumulate to greater magnitudes in slower growing mammals, this result is consistent with the hypotheses that FA in bone is primarily caused by (a) cumulative effects of asymmetrical mechanical factors, (b) accumulation of variation in the (local) regulation of growth, or (c) a tendency for bone morphology to drift through undirected remodeling. I discuss the implications of these optional explanations for primate evolution and bone development.     

Phenotypic Buffering in a Monogenean: Canalization and Developmental Stability in Shape and Size of the Haptoral Anchors of Ligophorus cephali (Monogenea: Dactylogyridae)

Phenotypic variation results from the balance between sources of variation and counteracting regulatory mechanisms. Canalization and developmental stability are two such mechanisms, acting at two different levels of regulation. The issue of whether or not they act concurrently as a common developmental buffering capacity has been subject to debate. We used geometric morphometrics to quantify the mechanisms that guarantee phenotypic constancy in the haptoral anchors of Ligophorus cephali. Canalization and developmental stability were appraised by estimating inter- and intra-individual variation, respectively, in size and shape of dorsal and ventral anchors. The latter variation was estimated as fluctuating asymmetry (FA) between anchor pairs. The general-buffering-capacity hypothesis was tested by two different methods based on correlations and Principal Components Analyses of the different components of size and shape variation. Evidence for FA in the dorsal and ventral anchors in both shape and size was found. Our analyses supported the hypothesis of a general developmental buffering capacity. The evidence was more compelling for shape than for size and, particularly, for the ventral anchors than for the dorsal ones. These results are in line with previous studies of dactylogyrids suggesting that ventral anchors secure a firmer, more permanent attachment, whereas dorsal anchors are more mobile. Because fixation to the host is crucial for survival in ectoparasites, we suggest that homeostatic development of the ventral anchors has been promoted to ensure the morphological constancy required for efficient attachment. Geometric morphometrics can be readily applied to other host-monogenean models, affording not only to disentangle the effects of canalization and developmental stability, as shown herein, but to further partition the environmental and genetic components of the former.     

Fitness, developmental instability, and the ontogeny of fluctuating asymmetry in Daphnia magna

Fluctuating asymmetry (FA) is the most commonly used measure of developmental instability. The relation between FA and individual fitness remains controversial, partly due to limited knowledge on the mechanisms behind variation in FA. To address this, we investigated the associations between FA, growth and reproduction as well as the ontogeny of FA in a clonal population of Daphnia magna . FA was not correlated with growth and reproduction, either at the between- or the within-individual level, in a high ( N  = 48 individuals) or in a low ( N  = 52 individuals) food-quantity regime. There were therefore no indications of functional effects of FA or of phenotypic trade-offs between developmental stability, growth and reproduction. Individual asymmetries varied randomly in sign and magnitude between subsequent molts ( N  = 19 individuals, 9–11 instars), but the levels of FA were generally lowest at intermediate ages. No feedback between right and left sides was detected. This suggests that FA only reflects the most recent growth history, that developmental instability may increase in old age, and that FA depends on processes operating on each side of the body independently. The results also suggest that FA differences within and among individual Daphnia are largely random, with limited biological significance.  © 2006 The Linnean Society of London, Biological Journal of the Linnean Society , 2006, 88 , 179–192.     

Lack of response to selection for lower fluctuating asymmetry of mutant eyespots in the butterfly Bicyclus anynana

Fluctuating asymmetry (FA) is claimed both to provide a means of evaluating developmental stability, and to reflect an individual's quality or the stress experienced during development. FA refers to the nondirectional variation between left and right sides, whereas directional asymmetry (DA) refers to a significant directional variation between the sides. We studied four eyespots on the dorsal forewing of the tropical butterfly, Bicyclus anynana. Two of the eyespots were specified by a mutant allele, Spotty, that was fixed in the stock. These eyespots showed higher FA than the two flanking, wild-type eyespots, although they are all formed by the same developmental pathway. We applied artificial selection for lower FA of the novel eyespots in an attempt to increase their developmental stability. There was significant variation present in individual FA in our study. However, this did not change as a result of the artificial selection. Most of the variation in FA can be accounted for by individual differences in developmental stability rather than by the applied selection or by environmental variation. Thus, it was not possible to produce any increased developmental stability of the novel eyespots by selecting for low FA. The estimates of realized heritability for both FA and DA of each eyespot were not significantly different from zero. The results suggest that FA provides little, if any, potential for exploring the mechanistic basis of developmental stability.     

Developmental motoneuron cell death and neurotrophic factors

During the development of higher vertebrates, motoneurons are generated in excess. In the lumbar spinal cord of the developing rat, about 6000 motoneurons are present at embryonic day 14. These neurons grow out axons which make contact with their target tissue, the skeletal muscle, and about 50% of the motoneurons are lost during a critical period from embryonic day 14 until postnatal day 3. This process, which is called physiological motoneuron cell death, has been the focus of research aiming to identify neurotrophic factors which regulate motoneuron survival during this developmental period. Motoneuron cell death can also be observed in vitro when the motoneurons are isolated from the embryonic avian or rodent spinal cord. These isolated motoneurons and other types of primary neurons have been a useful tool for studying basic mechanisms underlying neuronal degeneration during development and under pathophysiological conditions in neurodegenerative disorders. Accumulating evidence from such studies suggests that some specific requirements of motoneurons for survival and proper function may change during development. The focus of this review is a synopsis of recent data on such specific mechanisms.     

Phenotypic variation and fluctuating asymmetry in sexually dimorphic feather ornaments in relation to sex and mating system

Secondary sexual characters have been hypothesized to demonstrate increased phenotypic variation between and within individuals as compared to ordinary morphological traits. We tested whether this was the case by studying phenotypic variation, expressed as the coefficient of variation (CV), and developmental instability, measured as fluctuating asymmetry (FA), in ornamental and non-ornamental traits of 70 bird species with feather ornamentation while controlling for similarity among species due to common descent. Secondary sexual characters differed from ordinary morphological traits by showing large phenotypic CV and FA. This difference can be explained by the different mode of selection operating on each kind of trait: a history of intense directional (ornaments) and stabilizing selection (non-ornaments). Phenotypic variation is reduced in the sex with more intense sexual selection (males), but does not differ among species with different mating systems. The strength of stabilizing selection arising from natural selection is associated with decreased CV (wing CV is smaller than tarsus or tail CVs). We found evidence of FA being reduced in ornamental feathers strongly affected by aerodynamics (tail feathers) compared to other ornaments, but only in females. In conclusion, CV and FA were not related, suggesting mat phenotypic plasticity and developmental instability are independent components of phenotypic variation.     

Zinc and cadmium accumulation in cabbage aphid (Brevicoryne brassicae) host plants and developmental instability

Developmental instability in morphological characters can occur during individual development due to various environmental stresses. Fluctuating asymmetry (FA) is often used as a measurement of developmental instability, but within-environment variation (CV_e) is also considered an indicator of developmental instability. Cabbage aphid ( Brevicoryne brassicae ) populations were reared on zinc- (Zn) or cadmium- (Cd) contaminated cabbage and radish plants. Developmental instability indicators were measured and their relations with fitness were explored. Results revealed that cabbage aphids exposed to Cd and Zn displayed considerable developmental instability, particularly fluctuating asymmetry. Differences in developmental instability between the two metals were also detected, as well as differences between the two developmental instability measurements. For almost all measured traits, FA was greater on Cd- and Zn-contaminated compared to non-contaminated host plants. In contrast, CV_e of some traits was greater on non-contaminated host plants, yet for other traits CV_e was greater on contaminated host plants. There were also non-significant inverse relationships between FA and fitness of cabbage aphid populations. Due to weak correlations between FA and different patterns of two developmental instability measurements, this study does not support the hypothesis that developmental instability is a useful bioindicator of environmental quality.     

High variation in developmental instability under non-normal developmental error: a Bayesian perspective

The developmental mechanisms behind developmental instability (DI) are only poorly understood. Nevertheless, fluctuating asymmetry (FA) is often used a surrogate for DI. Based on statistical arguments it is often assumed that individual levels of FA are only weakly associated with the underlying DI. Patterns in FA therefore need to be interpreted with caution, and should ideally be transformed into patterns in DI. In order to be able to achieve that, assumptions about the distribution of developmental errors must be made. Current models assume that errors during development are additive and independent such that they yield a normal distribution. The observation that the distribution of FA is often leptokurtic has been interpreted as evidence for between-individual variation in DI. This approach has led to unrealistically high estimates of between-individual variation in DI, and potentially incorrect interpretations of patterns in FA, especially at the individual level. Recently, it has been suggested that the high estimates of variation in DI may be biased upward because either developmental errors are log-normal or gamma distributed and/or low measurement resolution of FA. A proper estimation of the amount (and shape) of heterogeneity in DI is crucial for the interpretation of patterns in FA and their transformation into patterns in DI. Yet, incorrect model assumptions may render misleading inferences. We therefore develop a statistical model to evaluate the sensitivity of results under the normal error model against the two alternative distributions as well as to investigate the importance of low measurement resolution. An analysis of simulated and empirical data sets indicated that bias due to misspecification of the developmental error distribution can be substantial, yet, did not appear to reduce estimates of variation in DI in empirical data sets to a large extent. Effects of low measurement resolution were neglectable. The importance of these results are discussed in the context of the interpretation of patterns in FA.     

FLUCTUATING ASYMMETRY DOES NOT CONSISTENTLY REFLECT SEVERE DEVELOPMENTAL DISORDERS IN HUMAN FETUSES

Developmental instability (DI), as measured by fluctuating asymmetry (FA), may reflect fitness and facilitate the expression of morphological variation. Insights in the underlying mechanisms and magnitude of DI during early development would increase our understanding of its role in evolutionary biology. We studied associations between FA and congenital abnormalities of different origins and functional systems in deceased human fetuses. Major congenital abnormalities corresponded to severe, often-lethal developmental disorders disrupting normal development from early organogenesis onward, but only moderately increased FA. Lower FA with age also supported the hypothesis that more severe abnormalities, leading to an earlier death, increased DI. Although FA related significantly to measures of fitness or health, we anticipated stronger associations because fetal health problems were detrimental. Furthermore, elevated FA occurred in only 4 of 17 disorders (left–right patterning, limb defects, and problems of bronchopulmonary and urogenital system). Fetuses experiencing major abnormalities other than these four types did not show increased FA. This suggests that the functional importance of symmetry in limbs has resulted in strong selection for symmetry and reduced its sensitivity to stress. Finally, the observed patterns suggest that specific developmental pathways have a stronger effect on DI than others do.     

Fluctuating asymmetry in farmed Atlantic salmon (Salmo salar) juveniles: also a maternal matter?

Developmental stability reflects the degree to which phenotypic expression is unaffected by random accidents or developmental noise. Developmental stability may be measured by phenodeviance or fluctuating asymmetry (FA), and estimation of developmental stability has attracted substantial interest because it appears to represent a relatively simple method to identify sub lethal stress exposure and to assess animal welfare. As a part of a long-term study, the work presented here primarily aimed to investigate impacts on developmental instability in farmed salmon offspring ten months post hatch attributable to maternal cortisol administration prior to spawning and mild hyperthermia exerted during incubation. Main results show that maternal cortisol enhancement increased the level of FA in pectoral and pelvic fins, but did not affect the frequency of malformations in offspring. Mild hyperthermia during incubation increased weight and fork length and also increased pelvic fin FA. Malformed fish were heavier and longer than the normal ones, and pelvic fin asymmetry was positively related to condition factor. These results illustrate plausible lasting impacts on offspring development due to the maternal endocrinological state at spawning and indicate that developmental instability in farmed salmon juveniles may mirror aspects of the broodstock’s housing conditions.     

Developmental stability covaries with genome-wide and single-locus heterozygosity in house sparrows

Fluctuating asymmetry (FA), a measure of developmental instability, has been hypothesized to increase with genetic stress. Despite numerous studies providing empirical evidence for associations between FA and genome-wide properties such as multi-locus heterozygosity, support for single-locus effects remains scant. Here we test if, and to what extent, FA co-varies with single- and multilocus markers of genetic diversity in house sparrow (Passer domesticus) populations along an urban gradient. In line with theoretical expectations, FA was inversely correlated with genetic diversity estimated at genome level. However, this relationship was largely driven by variation at a single key locus. Contrary to our expectations, relationships between FA and genetic diversity were not stronger in individuals from urban populations that experience higher nutritional stress. We conclude that loss of genetic diversity adversely affects developmental stability in P. domesticus, and more generally, that the molecular basis of developmental stability may involve complex interactions between local and genome-wide effects. Further study on the relative effects of single-locus and genome-wide effects on the developmental stability of populations with different genetic properties is therefore needed.     

FLUCTUATING ASYMMETRY IN A SALIX HYBRID SYSTEM: THE IMPORTANCE OF GENETIC VERSUS ENVIRONMENTAL CAUSES

To examine the effects of hybridization and environmental stress on developmental instability, we examined fluctuating asymmetry (FA), the variance in random deviations from perfect symmetry in bilaterally symmetrical traits, for leaf symmetry in a Salix hybrid system. An abiotic environmental stress (water stress), an interspecific biotic stress (pathogen attack), and an intraspecific biotic stress (competition) were examined to determine which factors increase developmental instability. None of these three environmental stressors significantly increased FA. However, genetic stress through hybridization was detected; hybrid plants showed significantly higher levels of FA than parental species. In contrast to hybridization providing greater developmental stability through heterozygosity, these results suggest that complex, nonadditive interactions provided developmental stability and that developmental instability increased when coadapted gene complexes were disrupted through hybridization. In addition, plant biomass was significantly, negatively correlated with FA, suggesting that those individuals that were more able to buffer themselves against the disruptive effects of environmental stress may have a selective advantage over those that are less able to buffer themselves against these disruptive effects.     

Lopsided Fish in the Snake River Basin — Fluctuating Asymmetry as a way of Assessing Impact of Hatchery Supplementation in Chinook Salmon, Oncorhynchus tshawytscha

The use of developmental instability (an individual's failure to produce a consistent phenotype in a given environment) was evaluated to detect the effects of outplanting hatchery fish on wild salmon. Juvenile chinook salmon were collected in 1989, 1990, and 1991 from five drainages in the Snake River Basin. In each drainage we attempted to collect fish from streams with no hatchery supplementation (wild), naturally spawning fish from streams with hatchery supplementation (natural), and fish collected at a hatchery. Forty fish were collected per site and the number of elements in bilateral characters were counted on each side of the fish. Indices of fluctuating asymmetry (FA), a measure of minor, random deviations in perfect symmetry of bilateral counts, were calculated as an estimator of developmental instability. Analysis of character counts from seven paired characters revealed normal distributions. Only one of the characters displayed counts that were statistically larger on one side than the other, indicating that directional asymmetry (DA) or antisymmetry was not a major bias of FA. However, the means of all individual characters revealed a non-statistically significant left side bias. We analyzed our data using two indices of FA (FA1 and FA5) with different levels of sensitivity to DA. Differences in both FA indices were found among years, with collection sites in 1989 having significantly larger FA values than in 1991 (FA p < 0.01). Levels of FA among wild, natural, and hatchery fish were comparatively small (FA1 p = 0.17). This suggests developmental conditions were different in the first year of the study than in the last. The cause of these differences may be linked to either genetic or environmental variation or to gene—environment interactions, but the general population declines of salmon that occurred during this time obscures more specific conclusions.     

Condition dependence of developmental stability in the sexually dimorphic fly Telostylinus angusticollis (Diptera: Neriidae)

Developmental stability is widely regarded as a condition‐dependent trait, but its relation to genotype and environment, and extent of developmental integration, remain contentious. In Telostylinus angusticollis, the dorsocentral bristles exhibit striking variation in developmental stability, manifested as fluctuating asymmetry (FA) in bristle position (‘positional FA’) and failure to develop some bristles (‘bristle loss’), in natural and laboratory populations. To determine whether this variation reflects condition, I tested for effects of genotype and environment (larval diet quality), and examined covariation with condition‐dependent traits. Positional FA was not affected by genotype or environment. However, positional FA covaried negatively with secondary sexual trait expression in males, and with sexual dimorphism in body shape, but covaried positively with body size in females. Bristle loss reflected both genotype and larval diet. Flies reared on poor‐quality diet exhibited a similar rate of bristle loss as wild flies. Both positional FA and bristle loss were greater in males. These results suggest that the relation between developmental stability and condition is complex and sex dependent.     

Fluctuating asymmetry--indicator of what?

In a population the optimal phenotype is promoted by buffering mechanisms that keep inter- and intra-individual variation low. A link exists between canalization, that controls phenotypic variation, and developmental stability, mostly measured as fluctuating asymmetry of bilateral traits (FA). Both types of variation are associated with the functional importance of a trait, and both are increased by stress of various kinds. But there are also several instances of non-congruence. The concept of developmental stability has been found elusive, and low FA is not the unambiguous measure of well being and good genes that has been claimed. It can be concluded that developmental stability is partly governed by specific, as yet unknown, molecular processes.     

Conserved and novel gene expression between regeneration and asexual fission in <Emphasis Type="Italic">Nematostella vectensis</Emphasis>

Due to work in model systems (e.g., flies and mice), the molecular mechanisms of embryogenesis are known in exquisite detail. However, these organisms are incapable of asexual reproduction and possess limited regenerative abilities. Thus, the mechanisms of alternate developmental trajectories and their relation to embryonic mechanisms remain understudied. Because these developmental trajectories are present in a diverse group of animal phyla spanning the metazoan phylogeny, including cnidarians, annelids, and echinoderms, they are likely to have played a major role in animal evolution. The starlet sea anemone Nematostella vectensis, an emerging model system, undergoes larval development, asexual fission, and complete bi-directional regeneration in the field and laboratory. In order to investigate to what extent embryonic patterning mechanisms are utilized during alternate developmental trajectories, we examined expression of developmental regulatory genes during regeneration and fission. When compared to previously reported embryonic expression patterns, we found that all genes displayed some level of expression consistent with embryogenesis. However, five of seven genes investigated also displayed striking differences in gene expression between one or more developmental trajectory. These results demonstrate that alternate developmental trajectories utilize distinct molecular mechanisms upstream of major developmental regulatory genes such as fox, otx, and Hox-like.     

Artificial selection reveals heritable variation for developmental instability

Fluctuating (nondirectional) asymmetry (FA) of bilaterally paired structures on a symmetrical organism is commonly used to assay the developmental instability (DI) caused by environmental or genetic factors. Although evidence for natural selection to reduce FA has been reported, evidence that FA (and by extension DI) is heritable is weak. We report the use of artificial selection to demonstrate heritable variation in the fluctuating asymmetry of interlandmark distances within the wing in an outbred population of Drosophila melanogaster. Our estimates for the heritability of FA range from 0% to 1% and result in estimates for the heritability of DI as large as 20%, comparable to values typical for life-history traits. These values indicate the existence of evolutionarily relevant genetic variation for DI and the effectiveness of selection for reduced FA suggests that natural selection has not fixed all the genetic variants that would improve developmental stability in these populations.     

Heat shock in the developmentally sensitive period of butterfly eyespots fails to increase fluctuating asymmetry

Fluctuating asymmetry (FA) is considered to provide a means of evaluating developmental stability and to reflect an individual's quality or the stress experienced during development. Stress is predicted to increase the phenotypic variation of both FA and trait size. In this study we examined the effect of a particular heat shock on both FA and size of eyespots in the butterfly, Bicyclus anynana. We also examined whether those eyespots thought to be involved in partner choice and sexual selection were particularly sensitive to stress. We applied a heat shock of 39.5 degrees C for 3 h before, during, and after a sensitive period in eyespot development. We examined the FA, variation in FA, size, and variation in size of five eyespots, two on the dorsal forewing (sexually selected traits), two on the ventral forewing, and one on the ventral hindwing (nonsexually selected traits). For each sex and treatment, the heat shock did not result in significant changes in mean trait size and FA nor in the variation of size and FA. There were no differences in the response to the heat shock between sexually and nonsexually selected traits. We discuss how the increased production of heat shock proteins, including HSP60, may have stabilized development and how this might explain the results.