Wing morphology and fluctuating asymmetry depend on the host plant in cactophilic Drosophila

As in most insect groups, host plant shifts in cactophilic Drosophila represent environmental challenges as flies must adjust their developmental programme to the presence of different chemical compounds and/or to a microflora that may differ in the diversity and abundance of yeasts and bacteria. In this context, wing morphology provides an excellent opportunity to investigate the factors that may induce changes during development. In this work, we investigated phenotypic plasticity and developmental instability of wing morphology in flies on the cactophilic Drosophila buzzatii and Drosophila koepferae raised on alternative breeding substrates. We detected significant differences in wing size between and within species, and between flies reared on different cactus hosts. However, differences in wing shape between flies emerged from different cactus hosts were not significant either in D. buzzatii or in D. koepferae. Our results also showed that morphological responses involved the entire organ, as variation in size and shape correlated between different portions of the wing. Finally, we studied the effect of the rearing cactus host on developmental instability as measured by the degree of fluctuating asymmetry (FA). Levels of FA in wing size were significantly greater in flies of both species reared in non-preferred when compared with those reared in preferred host cacti. Our results are discussed in the framework of an integrative view aimed at investigating the relevance of host plant shifts in the evolution of the guild of cactophilic Drosophila species that diversified in South America.     

Courtship song recognition in the Drosophila melanogaster complex: heterospecific songs make females receptive in D. melanogaster, but not in D. sechellia

Abstract. The courtship song emitted by male wing vibration has been regarded as one of the most important signals in sexual isolation in the species of the Drosophila melanogaster complex. Inter- and intraspecific crosses were observed using males whose wings were removed (mute) or females whose aristae were removed (deaf). Females of D. melanogaster, D. simulans , and D. mauritiana mated with heterospecific males in the song-present condition (cross between normal females and winged males) more often than in the no-song condition (cross between normal females and wingless males or between aristaless females and winged males) or they showed no preference between the two conditions. It is possible that in these females heterospecific courtship songs play a role as if they were conspecific. In contrast, the females of D. sechellia mated with D. melanogaster or D. simulans males in the no-song condition more often than in the song-present condition, suggesting that they reject males with heterospecific song. Female mate recognition depending on the courtship song in D. melanogaster, D. simulans , and D. mauritiana is considered to be relatively broader and that in D. sechellia narrower.     

Transposable Element Dynamics in Two Sibling Species: Drosophila Melanogaster and Drosophila Simulans

Transposable elements (TEs) in the two sibling species, Drosophila melanogaster and D. simulans, differ considerably in amount and dynamics, with D. simulans having a smaller amount of TEs than D. melanogaster. Several hypotheses have been proposed to explain these differences, based on the evolutionary history of the two species, and claim differences either in the effective size of the population or in genome characteristics. Recent data suggest, however, that the higher amount of TEs in D. melanogaster could be associated with the worldwide invasion of D. melanogaster a long time ago while D. simulans is still under the process of such geographical spread. Stresses due to new environmental conditions and crosses between migrating populations could explain the mobilization of TEs while the flies colonize. Colonization and TE mobilization may be strong evolutionary forces that have shaped and are still shaping the eukaryote genomes.     

The effect of developmental temperature on the genetic architecture underlying size and thermal clines in Drosophila melanogaster and D. simulans from the east coast of Australia

Body size and thermal tolerance clines in Drosophila melanogaster occur along the east coast of Australia. However the extent to which temperature affects the genetic architecture underlying the observed clinal divergence remains unknown. Clinal variation in these traits is associated with cosmopolitan chromosome inversions that cline in D. melanogaster. Whether this association influences the genetic architecture for these traits in D. melanogaster is unclear. Drosophila simulans shows linear clines in body size, but nonlinear clines in cold resistance. Clinally varying inversions are absent in D. simulans. Line-cross and clinal analyses were performed between tropical and temperate populations of D. melanogaster and D. simulans from the east coast of Australia to investigate whether clinal patterns and genetic effects contributing to clinal divergence in wing centroid size, thorax length, wing-to-thorax ratio, cold and heat resistance differed under different developmental temperatures (18 °C, 25 °C, and 29 °C). Developmental temperature influenced the genetic architecture in both species. Similarities between D. melanogaster and D. simulans suggest clinally varying inversion polymorphisms have little influence on the genetic architecture underlying clinal divergence in size in D. melanogaster. Differing genetic architectures across different temperatures highlight the need to consider different environments in future evolutionary and molecular studies of phenotypic divergence.     

Evolution of a desaturase involved in female pheromonal cuticular hydrocarbon biosynthesis and courtship behavior in Drosophila

Drosophila species exhibit polymorphism in female pheromonal cuticular hydrocarbons, with 7-monoenes produced in Drosophila simulans and 7,11-dienes in most populations of Drosophila melanogaster (5,9-dienes in several African populations). A female-biased desaturase, desatF, expressed only in D. melanogaster is involved in the synthesis of 7,11-dienes. We investigated the role of desatF in 5,9-diene flies. We constructed a 5,9-diene strain knock-down for desatF and showed that desatF is involved in 5,9-diene formation. We also studied D. melanogaster/D. simulans hybrids. These hybrid females produced dienes and received normal courtship from D. melanogaster males, but copulation success was reduced. With D. simulans males, courtship was decreased and no copulation occurred. Hybrids with a chromosomal deletion of the D. melanogaster desatF gene had no dienes and received normal courtship from D. simulans males; depending on the D. simulans parental strain, 7-19% of them succeeded in mating. D. simulans desatF promoter region shows 21-23% gaps and 86-89% identity with D. melanogaster promoter region, the coding region 93-94% identity, depending on the strain. These differences could explain the functional polymorphism of desatF observed between both species, contributing to different cuticular hydrocarbon profiles, that constitute an effective barrier between species.     

Conceptual translation of timeless reveals alternative initiating methionines in Drosophila.

We have sequenced genomic fragments which encode the N-terminus of the TIMELESS (TIM) clock protein in Drosophila simulans and D. yakuba. We observe that in these two species, the initiating methionine appears to lie downstream of the one proposed to encode the translational start inD.melanogaster, thereby truncating the N-terminus by 23 amino acids. We then sequenced the corresponding 5'fragment in a number of D. melanogaster individuals from different strains. We observed a polymorphism which strongly suggests that the originally proposed start site cannot be utilised in some individuals, and that these flies will initiate translation of TIM at the downstream ATG. Given the current interest in TIM regulation in D. melanogaster, it is important to correctly define the N-terminus in this species.     

Effects of aggregation pheromone on individual behaviour and food web interactions: a field study on Drosophila

Abstract.  1. The effects of an aggregation pheromone on individual behaviour and food web interactions were investigated in two ecological communities, using Drosophila melanogaster and D. simulans as focal species.
2. Fruit substrates with aggregation pheromone were significantly more attractive to adult D. melanogaster and D. simulans than control fruit substrates, and the response was positively dose dependent. Competing species and natural enemies were also significantly attracted to substrates with the aggregation pheromone of D. melanogaster and D. simulans .
3. Significantly more eggs were deposited on pheromone-treated fruits than on control fruits, and the microdistribution of eggs within fruits was correlated to the microdistribution of the pheromone. The aggregation pheromone induced more females to share the breeding site.
4. The extremely high densities of fruit flies in the large aggregations appeared to reduce the oviposition rate of females. Physical interactions with conspecific and heterospecifics were frequently observed in the aggregations, and often led to patch leaving of the fruit flies.
5. Competition for food among larvae occurred at high densities and parasitism was density dependent. Aggregation pheromones can be directly responsible for these patterns through their effects on the con- and heterospecific behaviour.
6. The combined results show that aggregation pheromones affect a multitude of aspects in the ecology of interacting animals. The importance of incorporating the communication signals in ecological theory of aggregations is discussed.     

Desiccation resistance and mating behaviour in laboratory populations of Drosophila simulans originating from the opposing slopes of Lower Nahal Oren (Israel)

Lower Nahal Oren in Northern Israel, often referred to as 'Evolution Canyon', has been proposed as a microscale model site for ecological evolution. However, conflicting stress resistance and mating assay results contribute to controversy over the Nahal Oren model. In this study, we further tested the Nahal Oren model, while extending its focus from Drosophila melanogaster to its sister species, Drosophila simulans. Using fly populations derived from the opposing canyon slopes and acclimated to laboratory conditions for 11-22 generations, we did not find a significant slope effect on desiccation resistance (P = 0.96) or body metabolic fuel content (P > 0.43), which would indicate a genetic basis for adaptation to local resource limitation. Multiple-choice mating assays (47-48% homotypic couples in two replicate populations) did not indicate divergence from a random mating pattern between north- and south-facing slope flies. In conclusion, our findings do not support divergence of D. simulans populations across Lower Nahal Oren.     

Comparative Analysis of Morphological Traits Among Drosophila Melanogaster and D. Simulans: Genetic Variability, Clines and Phenotypic Plasticity

The two sibling cosmopolitan species, Drosophila melanogaster and D. simulans, are able to proliferate under very different climatic conditions. This has resulted in local adaptations, which are often arranged in latitudinal clines. Such clines are documented for body weight, wing and thorax length, sternopleural and abdominal bristle number, ovariole number and thoracic pigmentation. The overall magnitude of geographical differentiation is, however, much less in D. simulans than in D. melanogaster, and latitudinal clines are less pronounced. The fact that natural populations live under different climates raises the problem of interaction between temperature and phenotype. The reaction norms of morphometrical traits have been investigated as a function of growth temperature. The shapes of the response curves vary according to the investigated trait. They are generally curvilinear and can be described by calculating characteristic values after polynomial adjustments. For a given trait, the reaction norms of the two species are similar in their shape, although some significant differences may be observed. Within each species, significant differences are also observed between geographic populations: reaction norms are not parallel and the divergence is better marked when more distant populations (e.g., temperate and tropical) are compared. It thus appears that besides mean trait value, phenotypic plasticity is also a target of natural selection. A specific analysis of wing shape variation according to growth temperature was also undertaken. Reaction norms with different shapes may be observed in various parts of the wing: the major effect is found between the basis and the tip of the wing, but in a similar way in the two species. By contrast, some ratios, called wing indices by taxonomists, may exhibit completely different reaction norms in the two species. For a single developmental temperature (25 degrees C) the phenotypic variability of morphometrical traits is generally similar in the two species, and also the genetic variability, estimated by the intraclass correlation. A difference exists, however, for the ovariole number which is less variable in D. simulans. Variance parameters may vary according to growth temperature, and a detailed analysis was made on wing dimensions. An increase of environmental variability at extreme, heat or cold temperatures, has been found in both species. Opposite trends were, however, observed for the genetic variability: a maximum heritability in D. simulans at middle temperatures, corresponding to a minimum heritability in D. melanogaster. Whether such a difference exists for other traits and in other populations deserves further investigations. In conclusion, morphometrical analyses reveal a large amount of significant differences which may be related to speciation and to the divergence of ecological niches. Within each species, numerous geographic variations are also observed which, in most cases, reflect some kinds of climatic adaptation.     

Interspecific competition in Drosophila II. Competitive outcome in some 2–resource environments

Populations of Drosophila metanogaster and D. simulans were made to compete in a number of 2– resource environments. The resources used were all based on a standard Drosophila medium and differed from each other only in their concentrations of ethanol. Each experiment comprised three cages started at a species–frequency of 0.8 D. melanogaster and three at 0.2 D. melanogaster. This enabled screening for a wide range of equilibria.
Reversal of competitive superiority was achieved (unintentionally) by raising the alcohol level of one of the resources. Thus while D. simulans won in an environment with media containing 0 and 896 ethanol, D. melanogaster won if the ethanol concentrations were 0 and 10%, or 0 and 12%. This parallels the reversal of dominance reported earlier for single–resource systems. However, in none of the 2–resource systems studied was there a stable equilibrium in species–frequency–in fact there was no significant frequency–dependence at all.
An artificial selection experiment produced a stock of D. melanogaster ovipositing a higher proportion of eggs on the alcoholic resource. Thus, this stock was more divergent from D. simulans , in at least one aspect of resource use, than was the original stock of D. melanogaster. However, competition between stock D. simulans and selected D. melanogaster showed no frequency–dependence. Again, reversal of competitive dominance was unintentionally achieved.
The implications of these results are discussed, and the need for a comprehensive study of resource–utilization in a pair of competing species is stressed.     

Differential responses to artificial selection on oviposition site preferences in Drosophila melanogaster and D. simulans

The preference–performance relationship in plant–insect interactions is a central theme in evolutionary ecology. Among many insects, eggs are vulnerable and larvae have limited mobility, making the choice of an appropriate oviposition site one of the most important decisions for a female. We investigated the evolution of oviposition preferences in Drosophila melanogaster Meigen and Drosophila simulans Sturtevant by artificially selecting for the preference for 2 natural resources, grape and quince. The main finding of our study is the differential responses of D. melanogaster and D. simulans. Although preferences evolved in the experimental populations of D. melanogaster, responses were not consistent with the selection regimes applied. In contrast, responses in D. simulans were consistent with expectations, demonstrating that this species has selectable genetic variation for the trait. Furthermore, crosses between D. simulans divergent lines showed that the genetic factors involved in grape preference appear to be largely recessive. In summary, our artificial selection study suggests that D. melanogaster and D. simulans possess different genetic architectures for this trait.     

Ancestral polymorphism and recent invasion of transposable elements in Drosophila species

ABSTRACT: BACKGROUND: During the evolutionary history of transposable elements, some processes, such as ancestral polymorphisms and horizontal transfer of sequences between species, can produce incongruences in phylogenies. We investigated the evolutionary history of the transposable elements Bari and 412 in the sequenced genomes of the Drosophila melanogaster group and in the sibling species D. melanogaster and D. simulans using traditional phylogenetic and network approaches. RESULTS: The maximum likelihood (ML) phylogenetic analyses revealed incongruences and unresolved relationships for both the Bari and 412 elements. The DNA transposon Bari within the D. ananassae genome is more closely related to the element of the melanogaster complex than to the sequence in D. erecta, which is inconsistent with the species phylogeny. Divergence analysis and the comparison of the rate of synonymous substitutions per synonymous site of the Bari and host gene sequences explain the incongruence as an ancestral polymorphism inherited stochastically by the derived species. Unresolved relationships were observed in the ML phylogeny of both elements involving D. melanogaster, D. simulans and D. sechellia. A network approach was used to attempt to resolve these relationships. The resulting tree suggests recent transfers of both elements between D. melanogaster and D. simulans. The divergence values of the elements between these species support this conclusion. CONCLUSIONS: We showed that an ancestral polymorphism and recent invasion of genomes due to introgression or horizontal transfer between species occurred during the evolutionary history of the Bari and 412 elements in the melanogaster group. These invasions likely occurred in Africa during the Pleistocene, before the worldwide expansion of D. melanogaster and D. simulans.     

DNA variability and divergence at the notch locus in Drosophila melanogaster and D. simulans: a case of accelerated synonymous site divergence

DNA diversity in two segments of the Notch locus was surveyed in four populations of Drosophila melanogaster and two of D. simulans. In both species we observed evidence of non-steady-state evolution. In D. simulans we observed a significant excess of intermediate frequency variants in a non-African population. In D. melanogaster we observed a disparity between levels of sequence polymorphism and divergence between one of the Notch regions sequenced and other neutral X chromosome loci. The striking feature of the data is the high level of synonymous site divergence at Notch, which is the highest reported to date. To more thoroughly investigate the pattern of synonymous site evolution between these species, we developed a method for calibrating preferred, unpreferred, and equal synonymous substitutions by the effective (potential) number of such changes. In D. simulans, we find that preferred changes per "site" are evolving significantly faster than unpreferred changes at Notch. In contrast we observe a significantly faster per site substitution rate of unpreferred changes in D. melanogaster at this locus. These results suggest that positive selection, and not simply relaxation of constraint on codon bias, has contributed to the higher levels of unpreferred divergence along the D. melanogaster lineage at Notch.     

An interspecific QTL study of Drosophila wing size and shape variation to investigate the genetic basis of morphological differences

The Drosophila wing has been used as a model in studies of morphogenesis and evolution; the use of such models can contribute to our understanding of mechanisms that promote morphological divergence among populations and species. We mapped quantitative trait loci (QTL) affecting wing size and shape traits using highly inbred introgression lines between D. simulans and D. sechellia, two sibling species of the melanogaster subgroup. Eighteen QTL peaks that are associated with 12 wing traits were identified, including two principal components. The wings of D. simulans and D. sechellia significantly diverged in size; two of the QTL peaks could account for part of this interspecific divergence. Both of these putative QTLs were mapped at the same cytological regions as other QTLs for intraspecific wing size variation identified in D. melanogaster studies. In these regions, one or more loci could account for intra- and interspecific variation in the size of Drosophila wings. Three other QTL peaks were related to a pattern of interspecific variation in wing size and shape traits that is summarized by one principal component. In addition, we observed that female wings are significantly larger and longer than male wings and the second, fourth and fifth longitudinal veins are closer together at the distal wing area. This pattern was summarized by another principal component, for which one QTL was mapped.     

Molecular variation at the In(2L)t proximal breakpoint site in natural populations of Drosophila melanogaster and D. simulans

A previous study of nucleotide polymorphism in a Costa Rican population of Drosophila melanogaster found evidence for a nonneutral deficiency in the number of haplotypes near the proximal breakpoint of In(2L)t, a common inversion polymorphism in this species. Another striking feature of the data was a window of unusually high nucleotide diversity spanning the breakpoint site. To distinguish between selective and neutral demographic explanations for the observed patterns in the data, we sample alleles from three additional populations of D. melanogaster and one population of D. simulans. We find that the strength of associations among sites found at the breakpoint varies between populations of D. melanogaster. In D. simulans, analysis of the homologous region reveals unusually elevated levels of nucleotide polymorphism spanning the breakpoint site. As with American populations of D. melanogaster, our D. simulans sample shows a marked reduction in the number of haplotypes but not in nucleotide diversity. Haplotype tests reveal a significant deficiency in the number of haplotypes relative to the neutral expectation in the D. simulans sample and some populations of D. melanogaster. At the breakpoint site, the level of divergence between haplotype classes is comparable to interspecific divergence. The observation of interspecific polymorphisms that differentiate major haplotype classes in both species suggests that haplotype classes at this locus are considerably old. When considered in the context of other studies on patterns of variation within and between populations of D. melanogaster and D. simulans, our data appear more consistent with the operation of selection than with simple demographic explanations.     

Comparison of alcohol dehydrogenase expression in Drosophila melanogaster and D. simulans

Alcohol dehydrogenase (ADH) gene expression was analyzed in Drosophila melanogaster and its sibling species D. simulans. The levels of ADH activity, ADH-cross-reacting material (CRM), and ADH-mRNA were analyzed for several strains of each species, which derive from diverse geographic locations around the world. There is considerable quantitative variation in ADH activity, CRM level, and RNA level among strains within species at all developmental stages. However, the only consistent differences between the two species are in pupal RNA level and in late-adult activity and CRM level. Late-adult melanogaster flies that are homozygous for the Slow allozyme have approximately twice the level of ADH activity and CRM as do simulans flies. The regression of activity on CRM over strains is highly significant and essentially the same for each species, which means that most, if not all, of the activity difference between the species is due to a difference in concentration of the ADH protein. In contrast, there is no significant regression of CRM level on mRNA level in adults of either species; nor is there a significant difference in RNA level between species. Therefore, the difference in ADH protein concentration is not due to RNA template availability. Thus, the interspecific difference in ADH level in adults must be due either to a difference in the rate of translation of the two RNAs or to a difference in protein stability.     

Intraspecific diversity of nucleotide sequences within the adenine + thymine-rich region of mitochondrial DNA molecules of Drosophila mauritiana, Drosophila melanogaster and Drosophila simulans.

Mitochondrial DNA (mtDNA) molecules from Drosophila mauritiana, D. melanogaster, and D. simulans contain a single adenine + thymine (A+T)-rich region, which is similarly located in all molecules, but varies in size among these species. Using agarose gel electrophoresis and electron microscopy, a difference in occurrence of one EcoRI site, and a difference in size (approximately 0.7 kb) of the A+T-rich regions was found between mtDNA molecules of flies of two female lines of D. mauritiana. In heteroduplexes constructed between these two kinds of mtDNA molecules, two or three regions of strand separation, each comprising single strands of unequal length, were apparent near the center of the A+T-rich region. Using the structural differences between D. mauritiana mtDNA molecules it was demonstrated the mtDNA of this species is maternally inherited. Differences in length of A+T-rich regions were also found between mtDNA molecules of two geographically separated strains of D. melanogaster, and between mtDNA molecules of two geographically separated strains of D. simulans. However, in both cases, in heteroduplexes constructed between mtDNA molecules of different strains of one species, the A+T-rich regions appeared completely paired.     

Defence strategies against a parasitoid wasp in Drosophila: fight or flight?

Hosts may defend themselves against parasitism through a wide variety of defence mechanisms, but due to finite resources, investment in one defence mechanism may trade-off with investment in another mechanism. We studied resistance strategies against the parasitoid wasp Leptopilina boulardi in two Drosophila species. We found that D. melanogaster had significantly lower physiological resistance against L. boulardi than D. simulans, and hypothesized that D. melanogaster might instead invest more heavily in other forms of defence, such as behavioural defence. We found that when given a choice between clean oviposition sites and sites infested with wasps, both D. melanogaster and D. simulans detected and avoided infested sites, which presumably limits later exposure of their offspring to infection. Unlike D. simulans, however, D. melanogaster laid significantly fewer eggs than controls in the forced presence of wasps. Our findings suggest that D. melanogaster relies more heavily on behavioural avoidance as defence against wasp parasitism than D. simulans, and that this may compensate for a lack of physiological defence.     

Growth temperature and genetic variability of wing dimensions in Drosophila: opposite trends in two sibling species

Thirteen linear wing dimensions were measured in 10 isofemale lines of Drosophila melanogaster and D. simulans grown at seven constant temperatures from 12 to 31 degrees C. Within-line (environmental) variability, estimated by the within-line coefficient of variation (CVw), exhibited similar variation patterns in the two species, that is higher values at extreme (low or high) temperatures. The magnitude of variation was, however, greater in D. simulans, which appears to be more responsive to thermal change. A clear hyperbolic relationship between trait mean value and CVw was also observed in both species, arising from measurement errors which are relatively more pronounced on shorter traits. Genetic variability was analysed by considering both the genetic CV (CVg, evolvability) and isofemale line heritability (intraclass correlation). Both parameters provided independent information, as shown by a lack of correlation between them. Moreover, CVg was negatively correlated with trait mean value, while heritability showed a positive correlation. With respect to thermal environment, both parameters exhibited similar reaction patterns which contrasted the two species. Genetic variability in D. melanogaster followed a convex reaction norm, with higher values at extreme (high or low) temperatures, and this observation agrees with previous independent investigations. Surprisingly, D. simulans revealed an opposite pattern, with a maximum genetic variability in the middle of the range. Such data point to the danger of drawing general conclusions from the analysis of a single species.     

Different mechanisms underlie phenotypic plasticity and interspecific variation for a reproductive character in drosophilids (Insecta: Diptera)

The insect ovary is a modular structure, the functional unit of which is the ovariole. Ovariole number is positively correlated with potential reproductive output. Among drosophilids (Insecta: Diptera), ovariole number shows both phenotypic plasticity and substantial interspecific and interpopulational variation. Here we examine the mechanistic connection between phenotypic plasticity and genetically fixed variation in ovariole number within the melanogaster species group. When a laboratory population of Drosophila melanogaster was reared under reduced food conditions, differences in ovariole number were entirely due to alterations in cell differentiation during the wandering stage at the very end of larval development. Cell growth and cell death were not affected. When these same flies were reared under a variety of temperatures, ovariole number differences arose during the latter half of the third (final) larval instar. Cell differentiation was affected, although cell number was not, and ovariole number differences were established before metamorphosis. In contrast, genetically fixed, interspecific and interpopulational variability in ovariole number was caused by alterations in the dynamics of cell differentiation and by cell number differences. Furthermore, the stages affected were different in different species and populations in the melanogaster species group, ranging from the first (D. sechellia) through the middle of the third (D. simulans and D. mauritiana) larval stage. Therefore, the mechanistic bases for plasticity-based variability are largely distinct from the mechanistic bases for interspecific and interpopulational variability. Our results suggest that phenotypic plasticity indicates evolutionary flexibility in underlying ontogenetic processes.