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.     

Fluctuating asymmetry of sexual and nonsexual traits in stalk‐eyed flies: a poor indicator of developmental stress and genetic quality

It has been proposed that females use fluctuating asymmetry (FA) in sexual ornaments to assess male quality. FA of sexual traits is predicted to show greater sensitivity to stress than FA of nonsexual traits, and to be heritable. We used a half‐sib mating design and manipulation of larval food environment to test these predictions on stalk‐eyed flies, Cyrtodiopsis dalmanni, in which females prefer males with larger eyespans. We measured size and FA of eyestalks and of two nonsexually selected characters, wing length and width. We found no evidence of an increase in FA under larval food stress in any of the individual traits, although trait size decreased under stress. We combined FA across traits into a single composite index, and found that males reared in the most benign larval environment had significantly higher composite FA than males reared on other media. There was no such effect in females. Heritability of FA was not significantly different from zero in any of the traits, in any of the environments, although trait sizes showed high heritability. We conclude that FA in sexual and nonsexual traits is a poor indicator of developmental stress and genetic quality.     

Developmental stability in yellow dung flies (Scathophaga stercoraria): fluctuating asymmetry,heterozygosity and environmental stress

The genetic basis for developmental stability, the ability of an organism to withstand genetic and environmental disturbance of development, is poorly understood. Fluctuating asymmetry (FA: small random deviations from symmetry in paired, bilateral traits) is the most widely used measure of developmental stability, and evidence suggests FA is weakly and negatively associated with genome‐wide heterozygosity. We investigated the genetic basis of developmental stability in the yellow dung fly. Fly lines were inbred for 16 generations at which time they were homozygous at the phosphoglucomutase (PGM) loci and PGM appears to influence FA in at least one other taxon. After 16 generations of inbreeding, lines homozygous for different PGM alleles were crossed and levels of FA for four metric traits were compared in the inbred and crossed flies. We also compared FA levels in these flies with previously gathered data on wild‐type (second generation outcrossed) flies, and additionally looked at the effects of two environmental stresses (larval food limitation and increased temperature) on FA. There were no significant differences in any measure of FA, nor in mean FA, in any trait when inbred and crossed flies were compared. Comparison of FA in these and wild flies also revealed no significant differences. Food limitation had no influence on FA, whereas heat stress increased FA of naturally, but not sexually, selected traits. Our results do not show a negative relationship between heterozygosity and FA, but support the notion that FA levels are stress, trait and taxon specific.     

Condition dependence of sexual ornament size and variation in the stalk-eyed fly Cyrtodiopsis dalmanni (Diptera: Diopsidae)

We used the stalk-eyed fly Cyrtodiopsis dalmanni to examine predictions made by condition-dependent handicap models of sexual selection. Condition was experimentally varied by manipulation of larval food availability. Cyrtodiopsis dalmanni is a highly dimorphic species exhibiting strong sexual selection, and the male sexual ornament (exaggerated eyespan) showed strong condition-dependent expression relative to the homologous trait in females and nonsexual traits. Male eyespan also showed a great increase in standardized variance under stress, unlike nonsexual traits. The inflated variance of the male ornament was primarily attributable to condition-dependent (but body-size-independent) increase in variance. Thus, evaluation of male eyespan allows females to gain additional information about male condition over and above that given by body size. These findings accord well with condition-dependent handicap models of sexual selection.     

Fluctuating asymmetry in certain morphological traits in laboratory populations of Drosophila ananassae.

Fluctuating asymmetry (FA, subtle random deviations from perfect bilateral symmetry) is often used as a measure of developmental instability (DI), which results from perturbations in developmental pathways caused by genetic or environmental stressors. During the present study, we estimated FA in 5 morphological traits, viz. wing length (WL), wing to thorax ratio (W:T), sternopleural bristle number (SBN), sex-comb tooth number (SCTN), and ovariole number (ON) in 18 laboratory populations of Drosophila ananassae. FA levels of measured traits differed significantly among populations except for SBN (in males and females) and W:T ratio (in females). Positional fluctuating asymmetry (PFA), a sensitive measure of DI, also varied significantly among the populations for SBN in females and SCTN in males. Interestingly, both males and females were similar for nonsexual traits. However, when FA across all traits (sexual and nonsexual) was combined into a single composite index (CFA), significant differences were found for both populations and sexes. Males showed higher CFA values than females, suggesting that males are more prone to developmental perturbations. The magnitude of FA differed significantly among traits, being lowest for nonsexual traits (SBN, WL, W:T ratio) and highest for sexual traits (SCTN and ON). The trait size of sexual traits (SCTN and ON) was positively correlated with their asymmetry. The possible reasons for variation in FA both among traits and among populations, and the usefulness of FA as an indicator of developmental stress and phenotypic quality in D. ananassae are discussed.     

Does fluctuating asymmetry of wing traits capture relative environmental stress in a lepidopteran?

1. Fluctuating asymmetry (FA) is hypothesized to be a useful predictor of population canalization, especially for organisms at risk from environmental change.
2. Identification of traits that meet statistical criteria as FA measures remains a challenge.
3. Here, a laboratory experiment subjected immature butterflies (Vanessa cardui) to diet and temperature conditions of varying stress levels. Variation in dietary macronutrient ratio (protein: carbohydrate) and rearing temperature (optimal: 25°C; elevated: 32°C) was introduced as stressors. Temperature and nutrition are key variables influencing ectotherm growth and fitness and so are likely to be important stressors that influence FA.
4. Individuals subjected to stressful conditions were predicted to show elevated FA of three wing size traits, as well as increased mortality and decreased adult body size.
5. Trait FA did not vary across treatments. Instead, treatment levels impacted viability: The combined incidence of pupal death and expression of significant wing malformations increased in treatment levels designated as stressful. Variation in adult dry mass also reflected predicted stress levels. Results suggest that individuals predicted to display increased FA either died or displayed gross developmental aberrations.
6. This experiment illustrates important constraints on the investigation of FA, including selection of appropriate traits and identification of appropriate levels of stressors to avoid elevated mortality. The latter concern brings into question the utility of FA as an indicator of stress in vulnerable, natural populations, where stress levels cannot be controlled, and mortality and fitness effects are often not quantifiable.

     

Fluctuating asymmetry,fecundity and development time in Drosophila: is there an association under optimal and stress conditions?

A number of studies have reported a significant negative association between fluctuating asymmetry (FA) of bilateral morphological traits and individual fitness traits, but almost all of these are unreplicated and based on small sample sizes using single trait estimates of FA. We therefore tested if there was a relationship between the FA of five bilateral traits and fecundity and development time in Drosophila in a multiple replicated experimental design. Stressed treatments were included to increase the variability of traits and to test whether associations among traits were affected by changes in the environment. Significant positive relationships were found between the size of wing characters and mean fecundity for the 5‐day period and this relationship tended to be stronger in the stress treatments. No association was found between FA and mean fecundity for any of the traits measured. Similarly, a significant positive relationship was detected between wing trait size and development time but no association was detected between trait FA and development time. There were no differences between mean fecundity or development time of extreme asymmetry phenotypes compared with modal phenotypes. These results are discussed with reference to suggestions in the literature that FA can be used to estimate individual fitness.     

Stability in development and variability in morphological signs in a natural population of Drosophila melanogaster: seasonal dynamics in 1999

Seasonal dynamics of developmental stability and variability of morphological traits was examined in a natural population of Drosophila melanogaster in order to compare these two parameters as indicators of temperature stress. Morphometric (thorax length and wing length) and meristic (number of sternopelural and orbital bristles) were studied. Variability was measured as the coefficient of variation. Stability of development was estimated as fluctuating asymmetry (FA) of bilateral traits. Thorax length and wing length did not exhibit consistent seasonal trends whereas wing loading significantly decreased. Significant seasonal changes in FA were not detected in any trait examined. Two traits showed reduced variation in autumn. The use of FA as an indicator of ecological stresses in insects is discussed on the basis of these results and the literature evidence.     

Stability of Development and Variability in Morphological Traits in a Natural Population of Drosophila melanogaster: Seasonal Dynamics in 1999

Seasonal dynamics of developmental stability and variability of morphological traits was examined in a natural population of Drosophila melanogaster in order to compare these two parameters as indicators of temperature stress. Morphometric (thorax length and wing length) and meristic (number of sternopelural and orbital bristles) were studied. Variability was measured as the coefficient of variation. Stability of development was estimated as fluctuating asymmetry (FA) of bilateral traits. Thorax length and wing length did not exhibit consistent seasonal trends whereas wing loading significantly decreased. Significant seasonal changes in FA were not detected in any trait examined. Two traits showed reduced variation in autumn. The use of FA as an indicator of ecological stresses in insects is discussed on the basis of these results and the literature evidence.     

Parasites increase fluctuating asymmetry of maleDrosophila nigrospiracula: Implications for sexual selection

Fluctuating asymmetry (minor deviations from perfect bilateral symmetry) is manifested by individuals less able to buffer environmental stress during development. I utilized a system of two naturally-occurring parasites ofDrosophila nigrospiracula to test whether parasitic infection during host development yields elevated degrees of fluctuating asymmetry in two morphological traits of males. This hypothesis has important implications for sexual selection, as it may explain why asymmetric males are often found to be sexually disadvantaged. In my system, nematodes infect larvae and therefore are more likely to disrupt development than mites which only parasitize adult flies. As predicted, nematode-infected maleD. nigrospiracula had a higher degree of bristle asymmetry than did mite-infested and control (carrying neither parasite) males. There was also a significant relation between nematode number and degree of asymmetry. There was a significant negative relation between nematode load and size of adult males, implicating a causal link between nutritional stress during host development and fluctuating asymmetry. Patterns of wing length asymmetry were inconsistent with those of bristle asymmetry. Nematode-infected males did not differ in wing length asymmetry relative to mite-infested and control males, nor was there a significant relation between nematode number and wing asymmetry. This inconsistency in expression of asymmetry may reflect different intensities of selection operating on each morphological trait.     

Effect of developmental temperature stress on fluctuating asymmetry in certain morphological traits in Drosophila ananassae

In the present study, the effect of thermal stress on the variability and fluctuating asymmetry (FA) in different morphological traits, viz., thorax length (TL), sternopleural bristle number (SBN), wing length (WL), wing-to-thorax (W/T) ratio, sex comb tooth number (SCTN) and ovariole number (ON), was investigated in 10 isofemale lines of Drosophila ananassae. The phenotypic and genetic variability is higher in the flies reared at low (20 °C) and at high (30 °C) temperatures as compared to that of standard (25 °C) temperature. Further, the levels of FA of measured traits differed significantly among the three temperature regimes except SBN and SCTN in males and SBN and W/T ratio in females. Moreover, the magnitude of positional fluctuating asymmetry is similar in males reared at three different developmental temperatures for SBN and SCTN but it varies significantly for SBN in females. However, when FA across all the traits was combined into a composite index (CFA), significant differences were found for both temperature regimes and sexes. Males showed higher CFA at 30 °C whereas in females it was higher at 20 °C. The results suggest that temperature increases the levels of variability and FA but the effect seems to be trait and sex specific in D. ananassae.     

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.     

Temperature and genotypic effects on life history and fluctuating asymmetry in a field strain of Culex pipiens

Fluctuating asymmetry (FA) has been proposed as a tool to measure levels of stress experienced by populations of organisms during development. To be of value as a bio-marker to highlight conditions at particular sites, it is important that variation in FA is due to environmental (eg pollution) variation and not genetic variation among populations and families, in other words heritability for FA should be very close to zero. A full-sib design was set up in which families of Culex pipiens mosquitoes collected from the field were reared at three different developmental temperatures. The effects of temperature and family on developmental rate, egg to adult survival and four wing morphological measures were assessed. There was both a temperature and a family effect on development rate and survival. Temperature affected all four wing traits, but an influence of family was only evident in two of the wing traits. Two separate measures of FA for each of the wing traits were obtained. The mean estimates of FA were mainly around 1% of the value of the character measured. There was evidence of an increase in FA with increase in temperature stress. Heritability was estimated for the wing traits and wing trait FA's using restricted estimation maximum likelihood. The estimates of heritability for the wing traits were small and, individually, did not differ significantly from zero. There was also no evidence of heritable genetic variation for any of the wing trait FA's. The results are discussed in relation to other studies where FA heritabilities have been estimated and in relation to the use of FA as an indicator of environmental stress.     

SEXUAL TRAITS ARE SENSITIVE TO GENETIC STRESS AND PREDICT EXTINCTION RISK IN THE STALK‐EYED FLY,DIASEMOPSIS MEIGENII

The handicap principle predicts that sexual traits are more susceptible to inbreeding depression than nonsexual traits. However, this hypothesis has received little testing and results are inconsistent. We used 11 generations of full‐sibling mating to test the effect of inbreeding on sexual and nonsexual traits in the stalk‐eyed fly Diasemopsis meigenii. Consistent with the theoretical predictions, the male sexual trait (eyespan) decreased more than nonsexual traits (female eyespan and male wing length), even after controlling for body size variation. In addition, male eyespan was a reliable predictor of line extinction, unlike other nonsexual traits. After 11 generations, inbred lines were crossed to generate inbred and outbred families. All morphological traits were larger in outbred individuals than inbred individuals. This heterosis was greater in male eyespan than in male wing length, but not female eyespan. The elevated response in male eyespan to genetic stress mirrored the result found using environmental stress during larval development and suggests that common mechanisms underlie the patterns observed. Overall, these results support the hypothesis that male sexual traits suffer more from inbreeding depression than nonsexual traits and are in line with predictions based on the handicap principle.     

Variation in morphological traits and trait asymmetry in field Drosophila serrata from marginal populations

Drosophila serrata occurs along the eastern coast of Australia with a southern range boundary near Sydney. To compare levels of phenotypic variation in marginal and central populations, we examined morphological variation in populations of this species from the southern range boundary and two more northerly populations. The populations differed for wing traits and there was an increase in wing size in the marginal locations which persisted under laboratory culture. The means of wing and bristle traits increased under laboratory culture, whereas wing trait coefficients of variation and variances decreased. Heritability estimates for wing size traits tended to be lower in the field compared with the laboratory, whereas bristle and crossvein length heritabilities were similar across environments. There was evidence for heritable variation in wing and bristle traits in both the marginal and more northern populations, suggesting that genetic variation was not limiting in marginal populations. Fluctuating asymmetry (FA) was also assessed as a measure of genomic and environmental stress. There were no consistent differences among populations for the FA of individual traits, or for a total FA score summed across traits. FA levels in field parents and laboratory‐reared progeny were similar. Overall, the results do not support the conjecture that levels of phenotypic and genetic variability differ between central and marginal D. serrata populations.     

Can evolution of sexual dimorphism be triggered by developmental temperatures?

Genetic prerequisites for the evolution of sexual dimorphism, sex-specific heritabilities and low or negative genetic correlations between homologous traits in males and females are rarely found. However, sexual dimorphism is evolving rapidly following environmental change, suggesting that sexual dimorphism and its genetic background could be environmentally sensitive. Yet few studies have explored the sensitivity of the genetic background of sexual dimorphism on environmental variation. In this study, on Drosophila melanogaster, we used a large nested full-sib-half-sib breeding design where families were split into four different developmental temperatures: two constant temperature treatments of 25 and 30 °C and two cycling temperatures with means of 25 and 30 °C, respectively. After emergence, we tested heat shock tolerance of adult flies. We found that sexual dimorphism was strongly affected by temperature during development. Moreover, we found that female heritability was significantly lower in flies developing at hot temperature and more so under hot and cycling temperatures. Interestingly, most of the genetic variation for heat shock tolerance was orthogonal (i.e. noncorrelated) between sexes, allowing independent evolution of heat shock tolerance in males and females. These findings give support to the hypothesis that the evolution of sexual dimorphism can be influenced by the environments experienced during development.     

Indicators of physiological stress and the elaboration of sexual traits in the collared flycatcher

Stress may have consequences for the evolution of condition-dependentsexual traits. For example, stress may be related to sexualtraits through immune function, and sexual traits can reflecthow individuals bear the costs of stress-mediated immunosuppression.However, male traits may be directly associated with stress,and such traits would then indicate stress tolerance. Here,we present initial results for the relationship between physiologicalstress estimated by the levels of heat shock proteins (HSP60and HSP70) and heterophil/lymphocyte ratio and the elaborationof sexual traits, such as forehead and wing patch size and songfeatures in the collared flycatcher Ficedula albicollis. Malesproducing longer and more versatile songs had significantlyhigher levels of HSP70, but other traits were unrelated to stress.In general, effect sizes for the relationship between stressand sexual traits had broad confidence intervals and variedbetween being small and medium effects. Immunoglobulin levels,leukocyte abundance, haemoparasite prevalence, male age, anddate and time effects did not affect the relationship betweenstress and sexual traits. These preliminary results, servinga basis for further experimental studies indicate that the relationshipbetween sexual traits and stress does not seem to be strong,but stress may partially constrain the expression of some sexualtraits.     

Intraspecific mating system evolution and its effect on complex male secondary sexual traits: Does male–male competition increase selection on size or shape?

Sexual selection is generally held responsible for the exceptional diversity in secondary sexual traits in animals. Mating system evolution is therefore expected to profoundly affect the covariation between secondary sexual traits and mating success. Whereas there is such evidence at the interspecific level, data within species remain scarce. We here investigate sexual selection acting on the exaggerated male fore femur and the male wing in the common and widespread dung flies Sepsis punctum and S. neocynipsea (Diptera: Sepsidae). Both species exhibit intraspecific differences in mating systems and variation in sexual size dimorphism (SSD) across continents that correlates with the extent of male–male competition. We predicted that populations subject to increased male–male competition will experience stronger directional selection on the sexually dimorphic male foreleg. Our results suggest that fore femur size, width and shape were indeed positively associated with mating success in populations with male‐biased SSD in both species, which was not evident in conspecific populations with female‐biased SSD. However, this was also the case for wing size and shape, a trait often assumed to be primarily under natural selection. After correcting for selection on overall body size by accounting for allometric scaling, we found little evidence for independent selection on any of these size or shape traits in legs or wings, irrespective of the mating system. Sexual dimorphism and (foreleg) trait exaggeration is therefore unlikely to be driven by direct precopulatory sexual selection, but more so by selection on overall size or possibly selection on allometric scaling.     

A time series of evolution in action: a latitudinal cline in wing size in South American Drosophila subobscura

Drosophila subobscura is geographically widespread in the Old World. Around the late 1970s, it was accidentally introduced into both South and North America, where it spread rapidly over broad latitudinal ranges. This invading species offers opportunities to study the speed and predictability of trait evolution on a geographic scale. One trait of special interest is body size, which shows a strong and positive latitudinal cline in many Drosophila species, including Old World D. subobscura. Surveys made about a decade after the invasion found no evidence of a size cline in either North or South America. However, a survey made in North America about two decades after the invasion showed that a conspicuous size cline had evolved and (for females) was coincident with that for Old World flies. We have now conducted parallel studies on 10 populations (13 degrees of latitude) of flies, collected in Chile in spring 1999. After rearing flies in the laboratory for several generations, we measured wing sizes and compared geographic patterns (versus latitude or temperature) for flies on all three continents. South American females have now evolved a significant latitudinal size cline that is similar in slope to that of Old World and of North American flies. Rates of evolution (haldanes) for females are among the highest ever measured for quantitative traits. In contrast, the size cline is positive but not significant for South or North American males. At any given latitude, South American flies of both sexes are relatively large; this in part reflects the relatively cool climate of coastal Chile. Interestingly, the sections of the wing that generate the size cline for females differ among all three continents. Thus, although the evolution of overall wing size is predictable on a geographic scale (at least for females), the evolution of size of particular wing components is decidedly not.     

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.