Drosophila Subobscura Populations Research Articles

Evolution and Plasticity of Gene Expression Under Progressive Warming in Drosophila subobscura.

Understanding the molecular mechanisms of thermal adaptation is crucial to predict the impacts of global warming. However, there is still a lack of research on the effects of rising temperatures over time and of studies involving different populations from the same species. The present study focuses on these two aspects, which are of great importance in understanding how organisms cope and adapt to ongoing changes in their environment. This study investigates the impact of global warming on the gene expression patterns of Drosophila subobscura populations from two different latitudinal locations after 23 generations of evolution. Our results indicate that evolutionary changes depend on the genetic background of the populations, with different starting points for thermal evolution, and that high-latitude populations show more pronounced evolutionary changes, with some evidence of convergence towards low-latitude populations. We found an interplay between plasticity and selection, with the high-latitude population showing fewer initial plastic genes and lower levels of adaptive plasticity, but a greater magnitude of change in both plastic and selective responses during evolution under warming conditions compared with its low-latitude counterpart. A substantial proportion of the transcriptome was observed to be evolving, despite the lack of observable response at higher-order phenotypic traits. The interplay between plasticity and selection may prove to be an essential component in shaping species' evolutionary responses to climate change. Furthermore, the value of conducting studies on multiple populations of the same species is emphasised, given the identification of differences between populations with different backgrounds in several contexts.

Heat-induced female biased sex ratio during development is not mitigated after prolonged thermal selection

BackgroundThe negative impacts of climate change on biodiversity are consistently increasing. Developmental stages are particularly sensitive in many ectotherms. Moreover, sex-specific differences in how organisms cope with thermal stress can produce biased sex ratios upon emergence, with potentially major impacts on population persistence. This is an issue that needs investigation, particularly testing whether thermal selection can alleviate sex ratio distortions in the long-term is a critical but neglected issue. Here, we report an experiment analyzing the sex ratio patterns at different developmental temperatures in Drosophila subobscura populations subjected to long-term experimental evolution (~ 30 generations) under a warming environment.ResultsWe show that exposure to high developmental temperatures consistently promotes sex ratio imbalance upon emergence, with a higher number of female than male offspring. Furthermore, we found that thermal selection resulting from evolution in a warming environment did not alleviate such sex ratio distortions generated by heat stress.ConclusionsWe demonstrate that heat stress during development can lead to clear sex ratio deviations upon emergence likely because of differential survival between sexes. In face of these findings, it is likely that sex ratio deviations of this sort occur in natural populations when facing environmental perturbation. The inability of many insects to avoid thermal shifts during their (more) sessile developmental stages makes this finding particularly troublesome for population subsistence in face of climate warming events.

Detrimental impact of a heatwave on male reproductive behaviour and fertility

Understanding how heatwaves impact on different aspects of mating behaviour and fertility is getting increasingly important. In this context, laboratory fertility and mating experiments involving manipulation and exposure of insects to different thermal conditions are common procedures. To conduct such experiments practical methods such as dyes are needed for an easy, non-invasive discrimination of individuals. We report here a study measuring the effect of an extended heat stress applied to males on several parameters of mating behaviour and fertility of laboratory populations of Drosophila subobscura derived from two distinct European locations. We found highly detrimental effects of heatwave on mating behaviour—with longer (courtship and copulation) latencies and lower mating occurrence but no changes in mating duration—and fertility, with reduced fecundity and reproductive success. Furthermore, we also tested the efficacy of food dye as a marker for individual discrimination and mating occurrence. While food dye did not allow to infer the occurrence of a mating based on a transfer of coloration from male to female, it did not affect mating and fertility, attesting its utility has a method for discriminating individuals within mating experiments in the context of thermal studies. Importantly, despite the fact that the heatwave was only applied in males, we observed an impact on behaviour of females that mated with stressed males, by often refusing their nuptial feeding. This opens possibilities for further integrated research on the changes of female and male mating behaviour and fertility under different thermal scenarios.

The Adaptive Value of Chromosomal Inversions and Climatic Change-Studies on the Natural Populations of Drosophila subobscura from the Balkans.

The adaptive value of the Drosophila subobscura chromosomal inversion polymorphism with regard to environmental effects is well-known. However, the specific details of the inversion adaptations to the global warming scenario deserve to be analyzed. Toward this aim, polymorphism and karyotypes were studied in 574 individuals from Petnica (Serbia) in annual samples taken in June for the period 2019-2022. Comparing the results of Petnica (Cfa: humid subtropical climate) with those from Avala (Serbia: Cfb, temperate oceanic climate) and Font Groga (Barcelona, Spain; Csa: hot-summer Mediterranean climate), significant differences were observed for their chromosomal polymorphism. In Petnica, inversions from U and E chromosomes mainly reacted significantly with regard to temperature, humidity, and rainfall. Moreover, the inversion polymorphism from Petnica (2019-2022) was compared with that from 1995. In this period, a significant increase in mean and maximum temperature was observed. However, to properly explain the observed variations of inversions over time, it was necessary to carefully analyze annual seasonal changes and particular heat wave episodes. Interestingly, yearly fluctuations of U chromosome 'warm'-adapted inversions corresponded with opposite changes in 'non-thermal' inversions. Perhaps these types of inversions were not correctly defined with regard to thermal adaptation, or these fluctuations were also due to adaptations to other physical and/or biological variables. Finally, a joint study of chromosomal inversion polymorphism from many Balkan populations of D. subobscura indicated that different climatic regions presented distinct composition, including thermal-adapted inversions.

Slow and population specific evolutionary response to a warming environment

Adaptation to increasingly warmer environments may be critical to avoid extinction. Whether and how these adaptive responses can arise is under debate. Though several studies have tackled evolutionary responses under different thermal selective regimes, very few have specifically addressed the underlying patterns of thermal adaptation under scenarios of progressive warming conditions. Also, considering how much past history affects such evolutionary response is critical. Here, we report a long-term experimental evolution study addressing the adaptive response of Drosophila subobscura populations with distinct biogeographical history to two thermal regimes. Our results showed clear differences between the historically differentiated populations, with adaptation to the warming conditions only evident in the low latitude populations. Furthermore, this adaptation was only detected after more than 30 generations of thermal evolution. Our findings show some evolutionary potential of Drosophila populations to respond to a warming environment, but the response was slow and population specific, emphasizing limitations to the ability of ectotherms to adapt to rapid thermal shifts.

Sex and population differences underlie variation in reproductive success in a warming environment.

Current rising temperatures are threatening biodiversity. It is therefore crucial to understand how climate change impacts male and female fertility and whether evolutionary responses can help in coping with heat stress. We use experimental evolution to study male and female fertility during the real-time evolution of two historically differentiated populations of Drosophila subobscura under different thermal selection regimes for 23 generations. We aim to (a) tease apart sex-specific differences in fertility after exposure to warming conditions during development, (b) test whether thermal selection can enhance fertility under thermal stress, and (c) address the role of historically distinct genetic backgrounds. Contrary to expectations, heat stress during development had a higher negative impact on female fertility than on male fertility. We did not find clear evidence for enhanced fertility in males or females evolving under warming conditions. Population history had a clear impact on fertility response under thermal stress, particularly in males with those from lower latitude presenting better performance than their higher latitude counterparts. We show that the impact of thermal stress on fertility varies between traits, sexes, and genetic backgrounds. Incorporating these several levels of variation is crucial for a deeper understanding of how fertility evolves under climate change.

Past history shapes evolution of reproductive success in a global warming scenario

Adaptive evolution is critical for animal populations to thrive in the fast-changing natural environments. Ectotherms are particularly vulnerable to global warming and, although their limited coping ability has been suggested, few real-time evolution experiments have directly accessed their evolutionary potential. Here, we report a long-term experimental evolution study addressing the evolution of Drosophila thermal reaction norms, after ∼30 generations under different dynamic thermal regimes: fluctuating (daily variation between 15 and 21 °C) or warming (daily fluctuation with increases in both thermal mean and variance across generations). We analyzed the evolutionary dynamics of Drosophila subobscura populations as a function of the thermally variable environments in which they evolved and their distinct background. Our results showed clear differences between the historically differentiated populations: high latitude D. subobscura populations responded to selection, improving their reproductive success at higher temperatures whereas their low latitude counterparts did not. This suggests population variation in the amount of genetic variation available for thermal adaptation, an aspect that needs to be considered to allow for better predictions of future climate change responses. Our results highlight the complex nature of thermal responses in face of environmental heterogeneity and emphasize the importance of considering inter-population variation in thermal evolution studies.

High developmental temperature leads to low reproduction despite adult temperature.

Phenotypic plasticity can help organisms cope with changing thermal conditions and it may depend on which life-stage the thermal stress is imposed: for instance, exposure to stressful temperatures during development can trigger a positive plastic response in adults. Here, we analyze the thermal plastic response of laboratory populations of Drosophila subobscura, derived from two contrasting latitudes of the European cline. We measured reproductive performance through fecundity characters, after the experimental populations were exposed to five thermal treatments, with different combinations of developmental and adult temperatures (14°C, 18°C, or 26°C). Our questions were whether (1) adult performance changes with exposure to higher (or lower) temperatures during development; (2) flies raised at lower temperatures outperform those developed at higher ones, supporting the "colder is better" hypothesis; (3) there is a cumulative effect on adult performance of exposing both juveniles and adults to higher (or lower) temperatures; (4) there is evidence for biogeographical effects on adult performance. Our main findings were that (1) higher developmental temperatures led to low reproductive performance regardless of adult temperature, while at lower temperatures reduced performance only occurred when colder conditions were persistent across juvenile and adult stages; (2) flies raised at lower temperatures did not always outperform those developed at other temperatures; (3) there were no harmful cumulative effects after exposing both juveniles and adults to higher temperatures; (4) both latitudinal populations showed similar thermal plasticity patterns. The negative effect of high developmental temperature on reproductive performance, regardless of adult temperature, highlights the developmental stage as very critical and most vulnerable to climate change and associated heat waves.

Local adaptation at fine spatial scale through chromosomal inversions and mito-nuclear epistasis: Findings in Drosophila subobscura (Diptera: Drosophilidae)

To explore local adaptation in wild populations at a fine spatial scale we characterized the genetic variability of eight closely located populations of Drosophila subobscura and its associations w ...

How phenotypic convergence arises in experimental evolution.

Evolutionary convergence is a core issue in the study of adaptive evolution, as well as a highly debated topic at present. Few studies have analyzed this issue using a "real-time" or evolutionary trajectory approach. Do populations that are initially differentiated converge to a similar adaptive state when experiencing a common novel environment? Drosophila subobscura populations founded from different locations and years showed initial differences and variation in evolutionary rates in several traits during short-term (∼20 generations) laboratory adaptation. Here, we extend that analysis to 40 more generations to analyze (1) how differences in evolutionary dynamics among populations change between shorter and longer time spans, and (2) whether evolutionary convergence occurs after 60 generations of evolution in a common environment. We found substantial variation in longer term evolutionary trajectories and differences between short- and longer term evolutionary dynamics. Although we observed pervasive patterns of convergence toward the character values of long-established populations, populations still remain differentiated for several traits at the final generations analyzed. This pattern might involve transient divergence, as we report in some cases, indicating that more generations should lead to final convergence. These findings highlight the importance of longer term studies for understanding convergent evolution.

Effects of photoperiod on life-history and thermal stress resistance traits across populations of Drosophila subobscura.

IntroductionOrganisms use environmental cues to match their phenotype with the future availability of resources and environmental conditions. Changes in the magnitude and frequency of environmental cues such as photoperiod and temperature along latitudes can be used by organisms to predict seasonal changes. While the role of temperature variation on the induction of plastic and seasonal responses is well established, the importance of photoperiod for predicting seasonal changes is less explored.Materials and methodsHere we studied changes in life‐history and thermal stress resistance traits in Drosophila subobscura in response to variation in photoperiod (6:18, 12:12 and 18:6 light:dark cycles) mimicking seasonal variations in day length. The populations of D. subobscura were collected from five locations along a latitudinal gradient (from North Africa and Europe). These populations were exposed to different photoperiods for two generations, whereafter egg‐to‐adult viability, productivity, dry body weight, thermal tolerance, and starvation resistance were assessed.ResultsWe found strong effects of photoperiod, origin of populations, and their interactions on life‐history and stress resistance traits. Thermal resistance varied between the populations and the effect of photoperiod depended on the trait and the method applied for the assessment of thermal resistance.PerspectivesOur results show a strong effect of the origin of population and photoperiod on a range of fitness‐related traits and provide evidence for local adaptation to environmental cues (photoperiod by population interaction). The findings emphasize an important and often neglected role of photoperiod in studies on thermal resistance and suggest that cues induced by photoperiod may provide some buffer enabling populations to cope with a more variable and unpredictable future climate.

Predictable phenotypic, but not karyotypic, evolution of populations with contrasting initial history

The relative impact of selection, chance and history will determine the predictability of evolution. There is a lack of empirical research on this subject, particularly in sexual organisms. Here we use experimental evolution to test the predictability of evolution. We analyse the real-time evolution of Drosophila subobscura populations derived from contrasting European latitudes placed in a novel laboratory environment. Each natural population was sampled twice within a three-year interval. We study evolutionary responses at both phenotypic (life-history, morphological and physiological traits) and karyotypic levels for around 30 generations of laboratory culture. Our results show (1) repeatable historical effects between years in the initial state, at both phenotypic and karyotypic levels; (2) predictable phenotypic evolution with general convergence except for body size; and (3) unpredictable karyotypic evolution. We conclude that the predictability of evolution is contingent on the trait and level of organization, highlighting the importance of studying multiple biological levels with respect to evolutionary patterns.

Local adaptation of reproductive performance during thermal stress.

Considerable evidence exists for local adaptation of critical thermal limits in ectotherms following adult temperature stress, but fewer studies have tested for local adaptation of sublethal heat stress effects across life-history stages. In organisms with complex life cycles, such as holometabolous insects, heat stress during juvenile stages may severely impact gametogenesis, having downstream consequences on reproductive performance that may be mediated by local adaptation, although this is rarely studied. Here, we tested how exposure to either benign or heat stress temperature during juvenile and adult stages, either independently or combined, influences egg-to-adult viability, adult sperm motility and fertility in high- and low-latitude populations of Drosophila subobscura. We found both population- and temperature-specific effects on survival and sperm motility; juvenile heat stress decreased survival and subsequent sperm motility and each trait was lower in the northern population. We found an interaction between population and temperature on fertility following application of juvenile heat stress; although fertility was negatively impacted in both populations, the southern population was less affected. When the adult stage was also subject to heat stress, the southern population exhibited positive carry-over effects whereas the northern population's fertility remained low. Thus, the northern population is more susceptible to sublethal reproductive consequences following exposure to juvenile heat stress. This may be common in other organisms with complex life cycles and current models predicting population responses to climate change, which do not take into account the impact of juvenile heat stress on reproductive performance, may be too conservative.

Tracking changes in chromosomal arrangements and their genetic content during adaptation.

There is considerable evidence for an adaptive role of inversions, but how their genetic content evolves and affects the subsequent evolution of chromosomal polymorphism remains controversial. Here, we track how life-history traits, chromosomal arrangements and 22 microsatellites, within and outside inversions, change in three replicated populations of Drosophila subobscura for 30 generations of laboratory evolution since founding from the wild. The dynamics of fitness-related traits indicated adaptation to the new environment concomitant with directional evolution of chromosomal polymorphism. Evidence of selective changes in frequency of inversions was obtained for seven of 23 chromosomal arrangements, corroborating a role for inversions in adaptation. The evolution of linkage disequilibrium between some microsatellites and chromosomes suggested that adaptive changes in arrangements involved changes in their genetic content. Several microsatellite alleles increased in frequency more than expected by drift in targeted inversions in all replicate populations. In particular, there were signs of selection in the O3+4 arrangement favouring a combination of alleles in two loci linked to the inversion and changing along with it, although the lack of linkage disequilibrium between these loci precludes epistatic selection. Seven other alleles increased in frequency within inversions more than expected by drift, but were not in linkage disequilibrium with them. Possibly these alleles were hitchhiking along with alleles under selection that were not specific to those inversions. Overall, the selection detected on the genetic content of inversions, despite limited coverage of the genome, suggests that genetic changes within inversions play an important role in adaptation.

Inversions and adaptation to the plant toxin ouabain shape DNA sequence variation within and between chromosomal inversions of Drosophila subobscura.

Adaptation is defined as an evolutionary process allowing organisms to succeed in certain habitats or conditions. Chromosomal inversions have the potential to be key in the adaptation processes, since they can contribute to the maintenance of favoured combinations of adaptive alleles through reduced recombination between individuals carrying different inversions. We have analysed six genes (Pif1A, Abi, Sqd, Yrt, Atpα and Fmr1), located inside and outside three inversions of the O chromosome in European populations of Drosophila subobscura. Genetic differentiation was significant between inversions despite extensive recombination inside inverted regions, irrespective of gene distance to the inversion breakpoints. Surprisingly, the highest level of genetic differentiation between arrangements was found for the Atpα gene, which is located outside the O1 and O7 inversions. Two derived unrelated arrangements (O3+4+1 and O3+4+7) are nearly fixed for several amino acid substitutions at the Atpα gene that have been described to confer resistance in other species to the cardenolide ouabain, a plant toxin capable of blocking ATPases. Similarities in the Atpα variants, conferring ouabain resistance in both arrangements, may be the result of convergent substitution and be favoured in response to selective pressures presumably related to the presence of plants containing ouabain in the geographic locations where both inversions are present.

Keeping your options open: Maintenance of thermal plasticity during adaptation to a stable environment.

Phenotypic plasticity may allow species to cope with environmental variation. The study of thermal plasticity and its evolution helps understanding how populations respond to variation in temperature. In the context of climate change, it is essential to realize the impact of historical differences in the ability of populations to exhibit a plastic response to thermal variation and how it evolves during colonization of new environments. We have analyzed the real-time evolution of thermal reaction norms of adult and juvenile traits in Drosophila subobscura populations from three locations of Europe in the laboratory. These populations were kept at a constant temperature of 18ºC, and were periodically assayed at three experimental temperatures (13ºC, 18ºC, and 23ºC). We found initial differentiation between populations in thermal plasticity as well as evolutionary convergence in the shape of reaction norms for some adult traits, but not for any of the juvenile traits. Contrary to theoretical expectations, an overall better performance of high latitude populations across temperatures in early generations was observed. Our study shows that the evolution of thermal plasticity is trait specific, and that a new stable environment did not limit the ability of populations to cope with environmental challenges.

Effect of lead pollution on fitness and its dependence on heterozygosity in Drosophila subobscura.

Lead is one of the most present contaminants in the environment, and different species respond differently to this type of polution. If combined with genomic stress, lead may act synergistically, causing significant decrease of fitness components. We used two genetically diverse Drosophila subobscura populations (regarding both putatively adaptive inversion and microsatellite loci polymorphisms) originating from two ecologically distinct habitats. To establish different levels of genome heterozygosity, series of intraline, intrapopulation and interpopulation crosses were made. The progeny were reared on a standard medium and a medium with 200 μg/mL of lead acetate. Development time was significantly extended to all groups reared on lead. The progeny of intraline crosses showed significantly extended development time compared to all other groups. The obtained results suggest that genome heterozygosity reduces the effect of lead pollution.

No trade-off between high and low temperature tolerance in a winter acclimatized Danish Drosophila subobscura population

Coping with cold winter conditions is a major challenge for many insects.In early spring we observed newly emerged Drosophila subobscura, which had overwintered as larvae and pupae. As temperatures increase during spring these flies are faced with higher minimum and maximum temperatures in their natural microhabitat. Thus, there is a potential costly mismatch between winter and early spring acclimatization and the increased ambient temperatures later in adult life.We obtained individuals from a natural Danish population of D. subobscura and acclimated them in the laboratory to 20°C for one generation, and compared critical thermal maximum (CTmax) and minimum (CTmin) to that of individuals collected directly from their natural microhabitat. The two populations (laboratory and field) were subsequently both held in the laboratory at 20°C and tested for their CTmax and CTmin every third day for 28days.At the first day of testing, field acclimatized D. subobscura had both higher heat and cold resistance compared to laboratory flies, and thereby a considerable larger thermal scope. Following transfer to the laboratory, cold and heat resistance of the field flies decreased over time relative to the laboratory flies. Despite the substantial decrease in thermal tolerances the thermal scope remained larger for field acclimatized individuals for the duration of the experiment.We conclude that flies acclimatized to their natural microhabitat had increased cold resistance, without a loss in heat tolerance. Thus while a negative correlation between cold and heat tolerance is typically observed in laboratory studies in Drosophila sp., this was not observed for field acclimatized D. subobscura in this study. We suggest that this is an adaptation to juvenile overwintering in temperate cold environments, where developmental (winter) temperatures can be much lower than temperatures experienced by reproducing adults after emergence (spring). The ability to gain cold tolerance through acclimatization without a parallel loss of heat tolerance affects thermal scope and suggests that high and low thermal tolerance act through mechanisms with different dynamics and reversibility.

Evolution of mating behavior between two populations adapting to common environmental conditions.

Populations from the same species may be differentiated across contrasting environments, potentially affecting reproductive isolation among them. When such populations meet in a novel common environment, this isolation may be modified by biotic or abiotic factors. Curiously, the latter have been overlooked. We filled this gap by performing experimental evolution of three replicates of two populations of Drosophila subobscura adapting to a common laboratorial environment, and simulated encounters at three time points during this process. Previous studies showed that these populations were highly differentiated for several life-history traits and chromosomal inversions. First, we show initial differentiation for some mating traits, such as assortative mating and male mating rate, but not others (e.g., female mating latency). Mating frequency increased during experimental evolution in both sets of populations. The assortative mating found in one population remained constant throughout the adaptation process, while disassortative mating of the other population diminished across generations. Additionally, differences in male mating rate were sustained across generations. This study shows that mating behavior evolves rapidly in response to adaptation to a common abiotic environment, although with a complex pattern that does not correspond to the quick convergence seen for life-history traits.

How much can history constrain adaptive evolution? A real-time evolutionary approach of inversion polymorphisms in Drosophila subobscura.

Chromosomal inversions are present in a wide range of animals and plants, having an important role in adaptation and speciation. Although empirical evidence of their adaptive value is abundant, the role of different processes underlying evolution of chromosomal polymorphisms is not fully understood. History and selection are likely to shape inversion polymorphism variation to an extent yet largely unknown. Here, we perform a real-time evolution study addressing the role of historical constraints and selection in the evolution of these polymorphisms. We founded laboratory populations of Drosophila subobscura derived from three locations along the European cline and followed the evolutionary dynamics of inversion polymorphisms throughout the first 40 generations. At the beginning, populations were highly differentiated and remained so throughout generations. We report evidence of positive selection for some inversions, variable between foundations. Signs of negative selection were more frequent, in particular for most cold-climate standard inversions across the three foundations. We found that previously observed convergence at the phenotypic level in these populations was not associated with convergence in inversion frequencies. In conclusion, our study shows that selection has shaped the evolutionary dynamics of inversion frequencies, but doing so within the constraints imposed by previous history. Both history and selection are therefore fundamental to predict the evolutionary potential of different populations to respond to global environmental changes.