A comparative study of competitive ability between two cactophilic species in their natural hosts

Abstract Competition is a major aspect of the ecology of insect communities exploiting ephemeral and fragmented resources. We analysed the effect of intraspecific (single species culture) and interspecific (mixed species culture) competition on larval viability, developmental time and wing length in the cactophilic Drosophila buzzatii and Drosophila koepferae (Diptera: Drosophilidae) reared in cultured media prepared with fermenting tissues of three common natural cactus hosts in nature at different densities. Our results show that all traits measured were affected by both intra‐ and interspecifc competition, although the effect of competition depended on the Drosophila species and the rearing cactus. In fact, flies tended to have a lower viability, shorter wing size and longer developmental time as a function of increasing density in single species culture in both D. buzzatii and D. koepferae (intraespecific competition). Besides, the performance of both species was seriously affected (shorter body size, slower developmental times, lower viability) by the presence of heterospecific competitors except in the case of D. koepferae reared in its primary host plant, Trichocereus terschekii. We also show that D. koepferae successfully utilized Opuntia quimilo, which is absent in most parts of its distribution range. We discuss the roles of intra‐ and interspecific competition as determinants of the relative abundance of these two species in the arid zones of Southern South America.     

Mating success depends on rearing substrate in cactophilic <Emphasis Type="Italic">Drosophila</Emphasis>

Drosophila buzzatii and D. koepferae coexist in the arid lands of southern South America and exploit different types of cactus as breeding hosts. The former prefers to lay eggs on the rotting pads of prickly pears (genus Opuntia) whereas D. koepferae exhibits greater acceptance for columnar cacti (e.g., Echinopsis terschekii). Here, we demonstrate that the rearing cacti affect male mating success, flies reared in each species’ preferred host exhibited enhanced mating success than those raised in secondary hosts. Opuntia sulphurea medium endows D. buzzatii males with greater mating ability while D. koepferae males perform better when flies develop in Echinopsis terschekii. These effects are not mediated through body size, even in D. buzzatii whose body size happens to be affected by the rearing cacti. This scenario, which is consistent with the evolution of host specialization and speciation through sensory drive, emphasizes the importance of habitat isolation in the coexistence of these cactophilic Drosophila.     

The evolutionary history of Drosophila buzzatii XXVII

The correlation between body size and longevity was tested in an Argentinian natural population of Drosophila buzzatii. Mean thorax length of flies newly emerging from rotting cladodes of Opuntia vulgaris was significantly smaller than that of two samples of flies caught at baits. The present results which might be interpreted as directional selection for longevity favoring larger flies are in agreement with previous results achieved in a Spanish natural population of D. buzzatii. Flies emerging from different substrates showed significant differences in thorax length, suggesting that an important fraction of phenotypic variance can be attributed to environmental variability. However, laboratory and field work in different populations of D. buzzatii showed a significant genetic component for thorax length variation.     

Phenotypic plasticity in Drosophila cactophilic species: the effect of competition,density, and breeding sites

Changes in the environmental conditions experienced by naturally occurring populations are frequently accompanied by changes in adaptive traits allowing the organism to cope with environmental unpredictability. Phenotypic plasticity is a major aspect of adaptation and it has been involved in population dynamics of interacting species. In this study, phenotypic plasticity (i.e., environmental sensitivity) of morphological adaptive traits were analyzed in the cactophilic species Drosophila buzzatii and Drosophila koepferae (Diptera: Drosophilidae) considering the effect of crowding conditions (low and high density), type of competition (intraspecific and interspecific competition) and cacti hosts (Opuntia and Columnar cacti). All traits (wing length, wing width, thorax length, wing loading and wing aspect) showed significant variation for each environmental factor considered in both Drosophila species. The phenotypic plasticity pattern observed for each trait was different within and between these cactophilic Drosophila species depending on the environmental factor analyzed suggesting that body size‐related traits respond almost independently to environmental heterogeneity. The effects of ecological factors analyzed in this study are discussed in order to elucidate the causal factors investigated (type of competition, crowding conditions and alternative host) affecting the election of the breeding site and/or the range of distribution of these cactophilic species.     

Multiple paternity and sperm competition in the sibling species Drosophila buzzatii and Drosophila koepferae

Sperm competition (SC) is a major component of sexual selection that enhances intra‐ and intersexual conflicts and may trigger rapid adaptive evolution of sexual characters. The actual role of SC on rapid evolution, however, is poorly understood. Besides, the relative contribution of distinctive features of the mating system to among species variation in the strength of SC remains unclear. Here, we assessed the strength of SC and mating system factors that may account for it in the closely related species Drosophila buzzatii and Drosophila koepferae. Our analyses reveal higher incidence of multiple paternity and SC risk in D. buzzatii wild‐inseminated females. The estimated number of fathers per brood was 3.57 in D. buzzatii and 1.95 in D. koepferae. In turn, the expected proportion of females inseminated by more than one male was 0.89 in D. buzzatii and 0.58 in D. koepferae. Laboratory experiments show that this pattern may be accounted for by the faster rate of stored sperm usage observed in D. koepferae and by the greater female remating rate exhibited by D. buzzatii. We also found that the male reproductive cost of SC is also higher in D. buzzatii. After a female mated with a second male, first‐mating male fertility was reduced by 71.4% in D. buzzatii and only 33.3% in D. koepferae. Therefore, we may conclude that postmating sexual selection via SC is a stronger evolutionary force in D. buzzatii than in its sibling.     

Experimental Evolution of Alkaloid Tolerance in Sibling <Emphasis Type="Italic">Drosophila</Emphasis> Species with Different Degrees of Specialization

Drosophila buzzatii and Drosophila koepferae are sibling species with marked ecological differences related to their patterns of host exploitation. D. buzzatii is a polyphagous species with a sub-cosmopolitan distribution, while D. koepferae is endemic to the mountain plateaus of the Andes, where it exploits alkaloidiferous columnar cacti as primary hosts. We use experimental evolution to study the phenotypic response of these cactophilic Drosophila when confronting directional selection to cactus chemical defenses for 20 generations. Flies adapted to cactus diets also experienced higher viability on alkaloid-enriched media, suggesting the selection of adaptive genetic variation for chemical-stress tolerance. The more generalist species D. buzzatii showed a rapid adaptive response to moderate levels of secondary metabolites, whereas the columnar cacti specialist D. koepferae tended to maximize fitness under harder conditions. The evolutionary dynamic of fitness-related traits suggested the implication of metabolic efficiency as a key mediator in the adaptive response to chemical stress. Although we found no evidence of adaptation costs accompanying specialization, our results suggest the involvement of compensatory evolution. Overall, our study proposes that differential adaptation to secondary metabolites may contribute to varying degrees of host specialization, favoring niche partitioning among these closely related species.     

A three-locus system of interspecific incompatibility underlies male inviability in hybrids between Drosophila buzzatii and D. koepferae

In hybrids between the sibling species D. buzzatii and D. koepferae, both sexes are more or less equally viable in the F₁: However, backcross males to D. buzzatii are frequently inviable, apparently because of interspecific genetic incompatibilities that are cryptic in the F₁. We have performed a genetic dissection of the effects of the X chromosome from D. koepferae. We found only two cytological regions, termed hmi-1 and hmi-2, altogether representing 9% of the whole chromosome, which when introgressed into D. buzzatii cause inviability of hybrid males. Observation of the pattern of asynapsis of polytene chromosomes (incomplete pairing, marking introgressed material) in females and segregation analyses were the technique used to infer the X chromosome regions responsible for this hybrid male inviability. The comparison of these results with those previously obtained with the same technique for hybrid male sterility in this same species pair indicate that in the X chromosome of D. koepferae there are at least seven times more regions that produce hybrid male sterility than hybrid male inviability. We have also found that the inviability brought about by the introgression of hmi-1 is suppressed by the cointrogression of two autosomal sections from D. koepferae. Apparently, these three regions conform to a system of species-specific complementary factors involved in an X-autosome interaction that, when disrupted in backcross hybrids by recombination with the genome of its sibling D. buzzatii, brings about hybrid male inviability.     

Aedeagal Divergence in Sympatric Populations of Two Sibling Species of Cactophilic Drosophila (Diptera: Drosophilidae): Evidence of Character Displacement?

Aedeagal morphology of two sibling cactophilic species, Drosophila buzzatii Patterson & Wheeler and Drosophila koepferae Fontdevila & Wasserman, was analyzed in nine allopatric and three sympatric locations throughout South America. Morphological differences were detected for both aedeagus size and shape between sympatric and allopatric populations of D. buzzatii, despite the significant variability within both groups. Populations of D. buzzatii sympatric with D. koepferae displayed smaller aedeagus than the allopatric ones as well as more differentiated aedeagus shape. The shape differences were non-allometric and mainly consisted in a change of curvature of the dorsal margin of the aedeagus being more pronounced in males from populations sympatric with D. koepferae. It is concluded that aedeagal morphology presented some degree of character displacement in both size and shape in populations of D. buzzatii in sympatry with D. koepferae. These results might suggest the existence of mechanisms of interspecific recognition and hybridization prevention between these species that include the morphology of the male genitalia.     

Reaction norms across and genetic parameters at different temperatures for thorax and wing size traits in Drosophila aldrichi and D. buzzatii

Reaction norms across seven constant and one fluctuating temperature of development were measured for thorax length and several wing size traits for up to 10 isofemale lines of each of the cactophilic Drosophila species, D. aldrichi and D. buzzatii, originating from the same locality. Maximum thorax length was reached at different low to intermediate temperatures for the two species, whereas wing length was highest at the lowest temperature in both species. Various ratio parameters showed pronounced species differences. The reaction norm for the wing loading index (wing length/thorax length) decreased monotonically with temperature in both species, but was much steeper and spanned a wider range in D. aldrichi than in D. buzzatii, suggesting either that wing loading is not a good characterization of flight capacity or, more likely, that flight optimization does not occur in the same manner in both species. The vein ratio (distal length/proximal length of the third vein) increased with temperature in D. buzzatii but decreased in D. aldrichi. Wing development in the two species thus is very different, with the proximal part of the wing in D. buzzatii more closely allied to the thorax than to the distal part. Among line variation was significant for all traits in both species, and most pronounced for thorax length and the ratio parameters. Coefficients of variation were significantly different between the species for all traits, with those in D. aldrichi higher than in D. buzzatii. Genetic variance in plasticity was significant for all traits in D. buzzatii, but only for seven out of 12 in D. aldrichi. Additive genetic variances for all traits in both species were significantly larger than zero. Genetic correlations between thorax length and several wing length parameters, and between these and wing area, were positive and generally significant in both species. The genetic correlation between the distal and the proximal length of the third vein was not significantly different from zero in D. aldrichi, but negative and significant in D. buzzatii. Heritabilites varied significantly among temperatures for almost all traits in both species. Phenotypic variances were generally higher in D. aldrichi than in D. buzzatii, and commonly highest at the extreme temperatures in the former species. At the high temperature the genetic variances also were usually highest in D. aldrichi. The data clearly suggest that the process of thermal adaptation is species specific and caution against generalizations based on the study of single species.     

Coexistence of ecologically similar colonising species III. Drosophila aldrichi and D. buzzatii: larval performance on,and adult preference for,three Opuntia cactus species

Two Drosophila species, D. buzzatti and D. aldrichi, coexist on several species of Opuntia cacti in Australia, primarily on O. tomentosa and O. streptacantha in the northern part of the cactus distribution, and on O. stricta in the south. Thorax length of field-collected adults was less, and the variance in length greater, than that for flies reared on simulated rots in the laboratory, indicating that these species are affected by crowding in nature. A larval performance index, measured on simulated cactus rots at low, moderate and high densities in single-species cultures, and at moderate and high densities in mixed-species cultures, was used to compare the relative intensity of intra- and interspecific competition at the same total larval density per 5 g necrotic cactus. Larval performance of both fly species was greatest on O. streptacantha, intermediate on O. tomentosa, and least on O. stricta in both single-species and mixed-species cultures. On O. stricta, the performances of D. aldrichi and D. buzzatii were not different when in single-species cultures, but that of D. aldrichi decreased significantly in mixed-species cultures. On the other two cactus species, the performances of D. aldrichi and D. buzzattii were not different in mixed-species cultures. The order of preferences by adult females for the cacti differed from that for larval performance, with females of both species prefering O. stricta. Analysis of microbial numbers growing on the cacti showed little difference among cacti at the rot age used for testing adult preference, but later growth was greater on O. tomentosa and O. streptacantha, the cacti that best supported larvae. Differential larval performance on O. stricta may contribute to the rare presence of D. aldrichi in the southern part of the cactus distribution, while the superior quality of O. tomentosa and O. streptacantha (larger rot size and higher microbial concentration) may reduce competition and facilitate cocxistence of the fly species in the north.     

A study of wing morphology and fluctuating asymmetry in interspecific hybrids between Drosophila buzzatii and D. koepferae

In this work we investigate the effect of interspecific hybridization on wing morphology using geometric morphometrics in the cactophilic sibling species D. buzzatii and D. koepferae. Wing morphology in F₁ hybrids exhibited an important degree of phenotypic plasticity and differs significantly from both parental species. However, the pattern of morphological variation between hybrids and the parental strains varied between wing size and wing shape, across rearing media, sexes, and crosses, suggesting a complex genetic architecture underlying divergence in wing morphology. Even though there was significant fluctuating asymmetry for both, wing size and shape in F₁ hybrids and both parental species, there was no evidence of an increased degree of fluctuating asymmetry in hybrids as compared to parental species. These results are interpreted in terms of developmental stability as a function of a balance between levels of heterozygosity and the disruption of coadaptation as an indirect consequence of genomic divergence.     

Molecular Population Genetics of the α<Emphasis Type="Italic">-Esterase5</Emphasis> Gene Locus in Original and Colonized Populations of <Emphasis Type="Italic">Drosophila buzzatii</Emphasis> and Its Sibling <Emphasis Type="Italic">Drosophila koepferae</Emphasis>

Several studies have suggested that esterase-2 (EST-2) may be the target of natural selection in the cactophilic fly Drosophila buzzatii. In this work, we analyzed nucleotide variation in a fragment of α-esterase5 (αE5), the gene encoding EST-2, in original (Argentinian) and colonized (Australian) populations of D. buzzatii and in its sibling D. koepferae. Estimates of nucleotide heterozygosity in D. buzzatii were similar in Australia and Argentina, although we detected a loss of singletons in colonized populations, suggesting a moderate founder effect. Interspecific comparisons revealed that D. buzzatii was more polymorphic for nonsynonymous variation, whereas D. koepferae was more variable for synonymous and noncoding sites. The two major chromosomal arrangements (2st and 2j) in D. buzzatii displayed similar levels of nucleotide variation, whereas 2jz ³ was monomorphic. The sequenced region allowed the discrimination of a greater number of EST-2 protein variants in the Australian sample than in the Argentinean sample. In D. koepferae, nucleotide variation in αE5 does not depart from neutral expectations, although tests of population structure were significant for silent variation. In contrast, D. buzzatii has probably undergone a recent population expansion in its South American range. In addition, the McDonald and Kreitman test revealed an excess of nonsynonymous polymorphism in both original and colonized populations of this species. Electronic Supplementary Material Electronic Supplementary material is available for this article at and accessible for authorised users. [Reviewing Editor: Dr. Richard Kliman]     

Differential Rates of Male Genital Evolution in Sibling Species of <Emphasis Type="Italic">Drosophila</Emphasis>

Genital morphology in animals with internal fertilization is considered to be among the fastest evolving traits. Sexual selection is often proposed as the main driver of genital diversification but the exact selection mechanisms involved are usually unclear. In addition, the mechanisms operating may differ even between pairs of sibling species. We investigated patterns of male genital variation within and between natural populations of the cactophilic fly Drosophila koepferae ranging its entire geographic distribution and compared them with those previously observed in its sibling species, D. buzzatii. Using both mtDNA and nDNA markers we found that genital shape variation in D. koepferae is more restricted than expected for neutral evolution, suggesting the predominance of stabilizing selection. We also detected dissimilar patterns of divergence between populations of D. koepferae that were allopatric and sympatric with D. buzzatii. The constrained evolution inferred for D. koepferae’s genitalia clearly contrasts with the rapid divergence and higher morphological disparity observed in the populations of D. buzzatii. Finally, different possible scenarios of male genital evolution in each species and within the radiation of D. buzzatii cluster are discussed.     

Spatial Associations, Size–Distance Relationships and Population Structure of Two Dominant Life Forms in a Semiarid Enclave of the Venezuelan Andes

The role of three thorny legume species as nurse plants and competitive relationships with columnar cacti were evaluated in a semiarid enclave of the Venezuelan Andes. Abundance and size of three columnar cacti species (Stenocereus griseus, Cereus repandus and Pilosocereus tillianus) under isolated shrubs of three thorny legumes species (Prosopis juliflora, Acacia farnesiana and A. macracantha) were recorded and compared with open areas. Using size–distance data we inferred the intensity of intra- and interspecific competition between both life forms in a “cardonal” (xeric zone) and “espinar” (mesic zone) of the enclave. Sixty-one columnar cacti were found beneath the three thorny legume species (92%), while only 5 cactus individuals were found in open areas (8%). Comparison of observed and expected number of cactus individuals shows that S. griseus and C. repandus are significantly associated with isolated shrubs of A. farnesiana and A. macracantha. Although P. juliflora contributed more than 40% of the total legume plant cover, the number of columnar cacti under its isolated shrubs was significantly lower than would be expected by chance. In all, 19 of 21 possible plant–plant combinations between and within columnar cacti and legume shrubs were recorded (espinar: 18, cardonal: 13, common combinations: 12). In both zones, intra- and interspecific combinations among columnar cacti species were relatively high (positive correlation between the sum of neighbour plant sizes and the distances separating them). Our results strongly suggest, at least for case of S. griseus and C. repandus, that these columnar cacti species require nurse plants for their establishment. The results on interference need still support from further research. We discuss the effects of plant–plant positive interactions on natural regeneration of these columnar cacti.     

Genetic variation and plasticity of thorax length and wing length in Drosophila aldrichi and D. buzzatii

Reaction norms across three temperatures of development were measured for thorax length, wing length and wing length/thorax length ratio for ten isofemale lines from each of two populations of Drosophila aldrichi and D. buzzatii. Means for thorax and wing length in both species were larger at 24 °C than at either 18 °C or 31 °C, with the reduction in size at 18 °C most likely due to a nutritional constraint. Although females were larger than males, the sexes were not different for wing length/thorax length ratio. The plasticity of the traits differed between species and between populations of each species, with genetic variation in plasticity similar for the two species from one locality, but much higher for D. aldrichi from the other. Estimates of heritabilities for D. aldrichi generally were higher at 18 °C and 24 °C than at 31 °C, but for D. buzzatii they were highest at 31 °C, although heritabilities were not significantly different between species at any temperature. Additive genetic variances for D. aldrichi showed trends similar to that for heritability, being highest at 18 °C and decreasing as temperature increased. For D. buzzatii, however, additive genetic variances were lowest at 24 °C. These results are suggestive that genetic variation for body size characters is increased in more stressful environments. Thorax and wing lengths showed significant genetic correlations that were not different between the species, but the genetic correlations between each of these traits and their ratio were significantly different. For D. aldrichi, genetic variation in the wing length/thorax length ratio was due primarily to variation in thorax length, while for D. buzzatii, it was due primarily to variation in wing length. The wing length/thorax length ratio, which is the inverse of wing loading, decreased linearly as temperature increased, and it is suggested that this ratio may be of greater adaptive significance than either of its components.     

High transposition rates of Osvaldo, a new Drosophila buzzatii retrotransposon

Transposition of a new Drosophila retrotransposon was investigated. Total genomic Southern analysis and polytene in situ hybridizations in D. buzzatii strains and other related species using a 6 kb D. buzzatii clone (cDb314) showed a dispersed, repetitive DNA pattern, suggesting that this clone contains a transposable element (TE). We have sequenced the cDb314 clone and demonstrated that it contains all the conserved protein sequences and motifs typical of retrovirus-related sequences. Although cDb314 does not include the complete TE, the protein sequence alignment demonstrates that it includes a defective copy of a new long terminal repeat (LTR) retrotransposon, related to the gypsy family, which we have named Osvaldo. Using a D. buzzatii inbred line in which all insertion sites are known, we have measured Osvaldo transposition rates in hybrids between this D. buzzatii line and its sibling species D. koepferae. The results show that Osvaldo transposes in bursts at high rate, both in the D. buzzatii inbred line and in species hybrids.This paper is dedicated posthumously to Osvaldo A. Reig in recognition of his contributions to evolutionary biology and his early appreciation of the role of transposable elements in evolution     

Variation of life-history and morphometrical traits in Drosophila buzzatii and Drosophila simulans collected along an altitudinal gradient from a Canary island

Variation in three life‐history traits (developmental time, preadult viability and daily female productivity) and five morphometrical traits (thorax length, wing length, wing width, wing/thorax ratio and wing‐aspect ratio) was studied at three developmental temperatures (20, 25 and 30 °C) in Drosophila buzzatii and Drosophila simulans collected on the island of La Gomera (Canary Archipelago). The flies originated from five closely situated localities, representing different altitudes (from 20 to 886 m above sea level) and a range of climatic conditions. We found statistically significant population effects for all traits in D. buzzatii and for most of the traits in D. simulans. Although no correlations of trait values with altitude were detected, geographical patterns for three life‐history traits and body size in D. buzzatii indicated that short‐range geographical variation in this species could be maintained by local climatic selection. Five of eight traits showed population‐by‐temperature interactions either in D. buzzatii or in D. simulans, but in all cases except wing width in D. buzzatii this could not be interpreted as adaptive responses to thermal conditions in the localities. The range of plastic changes across temperatures for particular traits differed between species, indicating a possibility for different levels of environmental stress experienced by the natural populations. The reaction norm curves and the response of within‐population variability to thermal treatments suggested better adaptations to higher and lower temperatures for D. buzzatii and D. simulans, respectively. The levels of among‐population differentiation depended on developmental temperature, implying environmental effects on the expression of the genetic variance. At 20 and 25 °C, interpopulation variability in D. buzzatii was higher than in D. simulans, while at 30 °C the opposite trend was observed. © 2005 The Linnean Society of London, Biological Journal of the Linnean Society, 2005, 84 , 119–136.     

Evolution of male genitalia: environmental and genetic factors affect genital morphology in two <Emphasis Type="Italic">Drosophila</Emphasis> sibling species and their hybrids

Background  

The rapid evolution of genital morphology is a fascinating feature that accompanies many speciation events. However, the underlying patterns and explanatory processes remain to be settled. In this work we investigate the patterns of intraspecific variation and interspecific divergence in male genitalic morphology (size and shape) in the cactophilic sibling species Drosophila buzzatii and D. koepferae. Genital morphology in interspecific hybrids was examined and compared to the corresponding parental lines.     

Cloning and sequencing of the breakpoint regions of inversion <Emphasis Type="Italic">5g</Emphasis> fixed in <Emphasis Type="Italic">Drosophila buzzatii</Emphasis>

Chromosomal inversions are ubiquitous in Drosophila both as intraspecific polymorphisms and interspecific differences. Many gaps still remain in our understanding of the mechanisms that generate them. Previous work has shown that in Drosophila buzzatii, three polymorphic inversions were generated by ectopic recombination between copies of the transposon Galileo. In this study, we have characterized the breakpoint regions of inversion 5g, fixed in D. buzzatii and absent in Drosophila koepferae and other closely related species. A novel approach comprising four experimental steps was used. First, D. buzzatii BAC clones encompassing the breakpoints were identified and their ends sequenced. Then, breakpoint regions were mapped at high resolution in the Drosophila mojavensis genome sequence. Finally, breakpoint regions were isolated by polymerase chain reaction in D. buzzatii and D. koepferae and sequenced. Our aim was to shed light on the mechanism that generated inversion 5g and specifically to test for an implication of the transposon Galileo. No evidence implicates Galileo or other transposable elements in the origin of inversion 5g that was generated most likely by two independent breaks and non-homologous end-joining repair. Our results show that different inversion-generating mechanisms may coexist within the same lineage and suggest a hypothesis for the evolutionary time and mode of their operation. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. Josefa González and Alfredo Ruiz contributed equally.     

Variation of body size within and between wild populations of Drosophila buzzatii

Four populations of the cactophilous species D. buzzatii have been compared with respect to the phenotypic variation of thorax and wing length of wild versus laboratory reared flies. Three of the strains were intercrossed to provide parent, F₁ and F₂ comparisons as a test of co-adaptation. The genetic contribution to phenotypic variation of laboratory reared flies was estimated from the correlation between sibs derived from random pair mating and reared individually in separate cultures. The average natural temperature during development was estimated from the relations between the wing/thorax ratio and temperature in laboratory tests.The variance of thorax and wing length of wild flies was several times greater than that of laboratory reared flies and the increase was attributed primarily to variation in larval food supply although temperature fluctuation is also important. There was no evidence of heterosis or F₂ break-down in the crosses. For two of the populations the heritability of thorax length was high, 60–70%, and substantially lower for the third. The average temperature estimated from the wing/thorax/temperature relationship differed between sites. The reduction of body size below the potential maximum averaged 30% for two and 20% for the other population, with a wide spread about these values. The evidence is discussed in relation to assessing the nature of ecological variation by comparing the variation of morphological traits in wild and laboratory reared flies.