Thermal phenotypic plasticity of pre- and post-copulatory male harm buffers sexual conflict in wild Drosophila melanogaster

  1. Claudia Londoño-Nieto
  2. Roberto García-Roa
  3. Clara Garcia-Co
  4. Paula González
  5. Pau Carazo

  • Curated by eLife

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    This study has important implications for the impact of sexual conflict on population viability under different temperatures. The authors propose that male harm to females in sexual conflict can be reduced as a function of temperature within the optimal reproductive range of a species. The evidence for this proposal is currently incomplete because there is methodological detail that needs to be further clarified. The results could have implications for the likelihood of the evolutionary rescue of species facing the climate crisis.

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Abstract

Strong sexual selection frequently leads to sexual conflict and ensuing male harm, whereby males increase their reproductive success at the expense of harming females. Male harm is a widespread evolutionary phenomenon with a strong bearing on population viability. Thus, understanding how it unfolds in the wild is a current priority. Here, we sampled a wild Drosophila melanogaster population and studied male harm across the normal range of temperatures under which it reproduces optimally in nature by comparing female lifetime reproductive success and underlying male harm mechanisms under monogamy (i.e. low male competition/harm) vs. polyandry (i.e. high male competition/harm). While females had equal lifetime reproductive success across temperatures under monogamy, polyandry resulted in a maximum decrease of female fitness at 24°C (35%), reducing its impact at both 20°C (22%), and 28°C (10%). Furthermore, female fitness components and pre- (i.e. harassment) and post-copulatory (i.e. ejaculate toxicity) mechanisms of male harm were asymmetrically affected by temperature. At 20°C, male harassment of females was reduced, and polyandry accelerated female actuarial aging. In contrast, the effect of mating on female receptivity (a component of ejaculate toxicity) was affected at 28°C, where the mating costs for females decreased and polyandry mostly resulted in accelerated reproductive aging. We thus show that, across a natural thermal range, sexual conflict processes and their effects on female fitness components are plastic and complex. As a result, the net effect of male harm on overall population viability is likely to be lower than previously surmised. We discuss how such plasticity may affect selection, adaptation and, ultimately, evolutionary rescue under a warming climate.

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  1. Version published to 10.7554/elife.84759 on eLife
  2. eLife

    Author Response

    Reviewer #1 (Public Review):

    The study tackles the topic of male harm (sexual selection favoring male reproductive strategies that incur a reduction of female fitness) from an interesting angle. The authors put emphasis on using wild-collected populations and studying them within their normal thermal range of reproductive conditions. Where previous studies have used temperature variation as a proxy for stressful environmental change, this approach should instead clarify what can be the role of male harm on female fitness in natural conditions. A minor caveat regarding this point is the fact the polygamy treatment also has a heavily male-biased sex ratio (3:1). The authors argue that this sex ratio is within the range of normal variation in that species, but it is likely that the average is still (1:1) in natural populations and using a male-biased sex ratio could magnify the intensity of male harm. This does not undermine the conclusions regarding the temperature sensitivity of sexual conflict but should be acknowledged.

    The authors find that varying temperature within a range found in natural conditions affects the reproductive interactions between males and females, particularly through male-harm mechanisms. Male harm, measured as a reduction in lifetime reproductive success (LRS) from monogamy to polygamy settings is present at 20C, stronger at 24, and absent or undetectable at 28C. Female senescence is always faster in the polygamy mating systems as compared to monogamy, but the effect appears strongest at 20C. Mating behaviors of males and females in these different settings are used to attempt to uncover underlying mechanisms of the sensitivity of male harm to temperature.

    A weakness of the manuscript in its current form is the lack of clarity about the experimental design, which makes understanding the results a long and involved procedure, even for someone who is familiar with the field. If the authors consider revising the manuscript, I suggest giving a better overview of the experimental design(s) earlier in the manuscript, perhaps supported by a diagram or flowchart. I also suggest structuring the results better to aid the reader (e.g., make clearer distinctions between results that come from the different experiments). Finally, some additional figures and statistical tests corrected for multiple testing would help get a better feel of some aspects of the dataset.

    I believe that the conclusions are generally justified and the results overall convincing. Overall, this is an impressive study with a lot of dimensions to it. Its complexity is a challenge and may require additional effort from the authors to make it easier to access. The core of the question is answered by LRS measures, but the authors have also provided a wealth of behavioral data as well as other fitness components. The manuscript could be greatly improved by putting more effort into linking the different metrics together to track down potential mechanisms for the observed variation in male-harm-induced reduction in female LRS. The discussion would also benefit from considering the female side of the sexual conflict coevolution arms race.

    We are thankful for the nice words and constructive appraisal of our work. As stated above, reviews like this are extraordinarily helpful. The reviewer mentions four main points that we have addressed:

    1. We now expand a bit on the justification to use a (3:1) male-biased sex ratio in the methods section (lines 150-155). We also acknowledge potential limitations of this design in the discussion (lines 563-571).
    2. To clarify the methods, we have placed this section before the results. This, in itself, has significantly improved the clarity of the manuscript. We have also substantially re-written the methods and results (including adding some tables) to streamline the text while providing all the necessary details, and have also included several diagrams to illustrate all our experiments (in the SM, see Figs. S1.1 to S1.5) along with a general schematic figure of the general design that we present early on in the main text (in the introduction, see Fig. 1).
    3. As suggested, we have re-run all analyses using the Benjamini-Hochberg procedure in order to correct for inflation of type I error rate due to multiple testing. We have also included in the SM a complementary set of models that also test for this via post hoc Tukey contrasts. Both these approached corroborate our initial findings, and thus contribute to strengthen our results.
    4. We now explicitly discuss the female side of things in the discussion (lines 636-647).

    Reviewer #2 (Public Review):

    Londoño-Nieto et al. investigated the influence of temperature on the form and intensity of sexual conflict in Drosophila melanogaster. They aimed to test the effect of naturally occurring temperature fluctuations on a wild population of Drosophila while disentangling pre- and postcopulatory episodes of sexual conflict. To this end, they exposed females to males under monogamy or polyandry, hence manipulating the degree of male harm experienced by females. The effect of temperature was explored by exposing these groups to 20, 24, or 28{degree sign}C. They found that female fitness suffered from male harm most at 24{degree sign}C and less at the other two temperatures. Interestingly, pre- and postcopulatory episodes of sexual conflict were affected differently by temperature. Overall, these data suggest that the relationship between sexual conflict and temperature can be strong and complex. Hence, these results can have important implications for the impact of sexual conflict on population viability, especially in light of the climate crisis.

    We want to thank the reviewer for the time invested in reading and reviewing our work. We are glad to read that the reviewer found our results interesting and considered our study to be of importance to the field.

    This paper tackles a highly relevant question using an established model organism for sexual conflict and contains a rich dataset obtained using a series of carefully planned experiments and analysed in an appropriate way. Importantly, the authors used biologically meaningful temperatures and mating treatments, which increases the relevance of the data. The main conclusions are well supported by the data. Nevertheless, the devil is in the detail, and given the way the authors frame their study (i.e. testing a natural population under naturally occurring temperature fluctuations) and their results (i.e. sexual conflict is buffered by temperature effects in the wild) there are some limitations to be considered:

    We appreciate the positive feedback! The reviewer identified potential limitations and made good suggestions that have only served to improve our manuscript considerably, for which we are very grateful. Details follow on how we have dealt with each specific comment.

    1. The authors frame their study as addressing the question of how sexual conflict reacts to naturally occurring temperature fluctuations in the wild. Nevertheless, the population used in this experiment had been kept for nearly 3 years in the laboratory prior to the experiment. Importantly, the authors ensured that the laboratory population maintained genetic diversity, by regularly crossing wild lines into it. Nevertheless, this population remained for some time in the laboratory under standardized conditions. The applied temperature fluctuations are in a biologically meaningful range (though only during the reproductive season), but it remains unclear if the applied fluctuations were in a standardized way (i.e. pre-programmed) or included random fluctuations (i.e. a more natural setting). This laboratory setup has certainly clear advantages, for example, it enables the exclusion of any effects other than the temperature on sexual conflict. Nevertheless, how these will then ultimately play out in the wild could be a different story.

    Agree. We clarify now that we meant pre-programmed fluctuations and acknowledge this limitation in the methods (lines 124-131).

    1. The authors highlight clearly that temperature fluctuations in the wild might play an important part in how sexual conflict plays out in natural populations. This very interesting and highly relevant point might lead the reader to assume that this is what was actually tested in the experiment. Nevertheless, in the experiments, different constant temperatures were applied to the flies, while only the stock population was kept at a fluctuating temperature regime. Hence, the influence of fluctuations during episodes of sexual conflict remains untested. While the present data show that sexual conflict can be modulated by temperature, the effect of naturally occurring fluctuations on the net cost of sexual conflict to a population remains unclear.

    Again, a fair point that we acknowledge in the current version (lines 571-575). “Second, our treatment temperatures were stable, designed to study how coarse-grain changes in temperature across the adult lifespan of flies may influence how sexual conflict unfolds in nature. Thus, future studies will need to encompass how fine-grained fluctuation (i.e., repeated variation of temperature across an adult’s lifespan) may affect male harm for a more comprehensive picture of temperature effects on sexual conflict in the wild”.

    1. The authors conclude that the effect of sexual conflict can be buffered by temperature in the wild. In general, I agree with this, although a more conservative way of framing this would be to say that temperature modulates or moderates sexual conflict instead of buffers it. If there really is a buffering effect of temperature in the wild remains to be tested, I believe. This will depend on how actual changes in temperature affect this dynamic (see point 2). In addition, I think another interesting open question is what the mechanism behind the observed differences might be. Are male and female interests really more aligned at different temperatures (i.e. males plastically reduce harm)? This would really buffer the harm of sexual conflict at those temperatures. Nevertheless, alternatively, males might not be perfectly adapted to manipulate the female optimally at lower or higher temperatures. This would mean that if the temperatures change, males might evolve to increase the manipulation of females, and hence the scope for sexual conflict might not change in the end under this scenario. Nevertheless, as the authors themselves state: 'An intriguing possibility is thus that SFPs are more effective at lowering female re-mating rates at warm temperatures, thereby buffering these costs.' Therefore, a temperature-dependent increase in the effectiveness of male manipulation might counterintuitively reduce sexual conflict in this species.

    We echo both points in the current version of the paper (see lines 633-655).

    1. In the end the authors argue that the climate crisis might have 'unexpected positive consequences via its effect on male harm'. Sexual conflict is indeed widespread, but it takes many different forms (as has been nicely described in the introduction of this paper). Because the studied system seems to be quite a specific example, it is questionable how far spread this phenomenon is in nature. In addition, it remains unclear how male harm will evolve in response to the climate crisis (see point 3). Finally, the relative fitness of females increased in the present experiment, as the tested range was within the reproductive optimum of the species. Nevertheless, the relative importance of the positive effect of sexual conflict on fitness outside of optimal temperatures seems questionable.

    Agree. Altogether, we have tried to tone down our conclusions regarding the implication of our results for a climate change scenario, and acknowledge all the points highlighted by the reviewer in the current version of the manuscript (see lines 563-575).

    Nonetheless, I believe these results to be of exceeding interest to the scientific community and of importance to the field. It opens up many potential research directions and adds further data to the fascinating field of sexual conflict, SFPs, and male harm in Drosophila.

    We are thrilled to read that the reviewer found our study of exceeding interest.

    Reviewer #3 (Public Review):

    In this paper, the authors explore the effects of the environment, specifically temperature, on male harm to females. Male harm is the phenomenon where males reduce female fitness in polyandrous systems, where a single female may mate with multiple males. The selection of males to increase their reproductive success in male-male competition can lead to genetic conflict that increases male fitness at the expense of female fitness. Typically, male harm has been studied in single environments under optimal conditions. However, there is an increasing focus on the effect of the environment on fitness costs of male harm to females, as a way to better understand the effect of male harm on population fitness in more realistic ecological contexts. In this paper, the authors add to these studies by exploring the effect of temperature on male harm and female fitness, using the fruit fly Drosophila melanogaster, as a model system. They find that temperature affects the impact of male harm on female fitness, with male harm having the greatest effect at 24˚C relative to 20˚C and 28˚C. The authors then go on to disentangle how temperature affects the various components of male harm that impact female fitness (e.g. harassment, ejaculate toxicity). The paper demonstrates that male harm depends on ecological context, which has implications for understanding its impact on population fitness under realistic ecological scenarios, particularly with respect to climate change.

    The strength of the paper is that it demonstrates that male harm (presented as differences in female life reproductive success between monogamous and polyandrous matings) changes with temperature. The authors dissect this general observation by showing that different aspects of precopulatory reproductive behavior, for example, male-male aggression, copulation rate, and female rejection rate, also change with temperature. Further, they demonstrate that correlates for male ejaculate quality also change with temperature, suggesting that temperature also affects postcopulatory mechanisms of male harm.

    The weakness of the paper is that the method and results section are difficult to follow, which negatively impacts the interpretation of the data. The experiments are complex and need to be for what the authors are studying. Nevertheless, the paper is written in a way that makes it challenging for the reader to fully understand how precisely the experiments were conducted. Further, the authors do not explain clearly how some of the experiments relate to the phenomenon ostensibly being assayed. For example, a more detailed explanation of why mating duration and remating latency are assays for ejaculate quality in the context of sperm competition would be very helpful in interpreting the data. Further, a clearer explanation of the statistical analyses conducted

    Thank you for the positive, detailed and constructive review. We agree with all the weaknesses laid out and we have strived to address all of them in the current version. This includes a mayor rearrangement, structuring and re-write of the methods and results section and extra statistical analyses. Please find the details below.

  3. eLife

    eLife assessment

    This study has important implications for the impact of sexual conflict on population viability under different temperatures. The authors propose that male harm to females in sexual conflict can be reduced as a function of temperature within the optimal reproductive range of a species. The evidence for this proposal is currently incomplete because there is methodological detail that needs to be further clarified. The results could have implications for the likelihood of the evolutionary rescue of species facing the climate crisis.

  4. eLife

    Reviewer #1 (Public Review):

    The study tackles the topic of male harm (sexual selection favoring male reproductive strategies that incur a reduction of female fitness) from an interesting angle. The authors put emphasis on using wild-collected populations and studying them within their normal thermal range of reproductive conditions. Where previous studies have used temperature variation as a proxy for stressful environmental change, this approach should instead clarify what can be the role of male harm on female fitness in natural conditions. A minor caveat regarding this point is the fact the polygamy treatment also has a heavily male-biased sex ratio (3:1). The authors argue that this sex ratio is within the range of normal variation in that species, but it is likely that the average is still (1:1) in natural populations and using a male-biased sex ratio could magnify the intensity of male harm. This does not undermine the conclusions regarding the temperature sensitivity of sexual conflict but should be acknowledged.

    The authors find that varying temperature within a range found in natural conditions affects the reproductive interactions between males and females, particularly through male-harm mechanisms. Male harm, measured as a reduction in lifetime reproductive success (LRS) from monogamy to polygamy settings is present at 20C, stronger at 24, and absent or undetectable at 28C. Female senescence is always faster in the polygamy mating systems as compared to monogamy, but the effect appears strongest at 20C. Mating behaviors of males and females in these different settings are used to attempt to uncover underlying mechanisms of the sensitivity of male harm to temperature.
    A weakness of the manuscript in its current form is the lack of clarity about the experimental design, which makes understanding the results a long and involved procedure, even for someone who is familiar with the field. If the authors consider revising the manuscript, I suggest giving a better overview of the experimental design(s) earlier in the manuscript, perhaps supported by a diagram or flowchart. I also suggest structuring the results better to aid the reader (e.g., make clearer distinctions between results that come from the different experiments). Finally, some additional figures and statistical tests corrected for multiple testing would help get a better feel of some aspects of the dataset.

    I believe that the conclusions are generally justified and the results overall convincing. Overall, this is an impressive study with a lot of dimensions to it. Its complexity is a challenge and may require additional effort from the authors to make it easier to access. The core of the question is answered by LRS measures, but the authors have also provided a wealth of behavioral data as well as other fitness components. The manuscript could be greatly improved by putting more effort into linking the different metrics together to track down potential mechanisms for the observed variation in male-harm-induced reduction in female LRS. The discussion would also benefit from considering the female side of the sexual conflict coevolution arms race.

  5. eLife

    Reviewer #2 (Public Review):

    Londoño-Nieto et al. investigated the influence of temperature on the form and intensity of sexual conflict in Drosophila melanogaster. They aimed to test the effect of naturally occurring temperature fluctuations on a wild population of Drosophila while disentangling pre- and post-copulatory episodes of sexual conflict. To this end, they exposed females to males under monogamy or polyandry, hence manipulating the degree of male harm experienced by females. The effect of temperature was explored by exposing these groups to 20, 24, or 28{degree sign}C. They found that female fitness suffered from male harm most at 24{degree sign}C and less at the other two temperatures. Interestingly, pre- and postcopulatory episodes of sexual conflict were affected differently by temperature. Overall, these data suggest that the relationship between sexual conflict and temperature can be strong and complex. Hence, these results can have important implications for the impact of sexual conflict on population viability, especially in light of the climate crisis.

    This paper tackles a highly relevant question using an established model organism for sexual conflict and contains a rich dataset obtained using a series of carefully planned experiments and analysed in an appropriate way. Importantly, the authors used biologically meaningful temperatures and mating treatments, which increases the relevance of the data. The main conclusions are well supported by the data. Nevertheless, the devil is in the detail, and given the way the authors frame their study (i.e. testing a natural population under naturally occurring temperature fluctuations) and their results (i.e. sexual conflict is buffered by temperature effects in the wild) there are some limitations to be considered:

    1. The authors frame their study as addressing the question of how sexual conflict reacts to naturally occurring temperature fluctuations in the wild. Nevertheless, the population used in this experiment had been kept for nearly 3 years in the laboratory prior to the experiment. Importantly, the authors ensured that the laboratory population maintained genetic diversity, by regularly crossing wild lines into it. Nevertheless, this population remained for some time in the laboratory under standardized conditions. The applied temperature fluctuations are in a biologically meaningful range (though only during the reproductive season), but it remains unclear if the applied fluctuations were in a standardized way (i.e. pre-programmed) or included random fluctuations (i.e. a more natural setting). This laboratory setup has certainly clear advantages, for example, it enables the exclusion of any effects other than the temperature on sexual conflict. Nevertheless, how these will then ultimately play out in the wild could be a different story.

    2. The authors highlight clearly that temperature fluctuations in the wild might play an important part in how sexual conflict plays out in natural populations. This very interesting and highly relevant point might lead the reader to assume that this is what was actually tested in the experiment. Nevertheless, in the experiments, different constant temperatures were applied to the flies, while only the stock population was kept at a fluctuating temperature regime. Hence, the influence of fluctuations during episodes of sexual conflict remains untested. While the present data show that sexual conflict can be modulated by temperature, the effect of naturally occurring fluctuations on the net cost of sexual conflict to a population remains unclear.

    3. The authors conclude that the effect of sexual conflict can be buffered by temperature in the wild. In general, I agree with this, although a more conservative way of framing this would be to say that temperature modulates or moderates sexual conflict instead of buffers it. If there really is a buffering effect of temperature in the wild remains to be tested, I believe. This will depend on how actual changes in temperature affect this dynamic (see point 2). In addition, I think another interesting open question is what the mechanism behind the observed differences might be. Are male and female interests really more aligned at different temperatures (i.e. males plastically reduce harm)? This would really buffer the harm of sexual conflict at those temperatures. Nevertheless, alternatively, males might not be perfectly adapted to manipulate the female optimally at lower or higher temperatures. This would mean that if the temperatures change, males might evolve to increase the manipulation of females, and hence the scope for sexual conflict might not change in the end under this scenario. Nevertheless, as the authors themselves state: 'An intriguing possibility is thus that SFPs are more effective at lowering female re-mating rates at warm temperatures, thereby buffering these costs.' Therefore, a temperature-dependent increase in the effectiveness of male manipulation might counterintuitively reduce sexual conflict in this species.

    4. In the end the authors argue that the climate crisis might have 'unexpected positive consequences via its effect on male harm'. Sexual conflict is indeed widespread, but it takes many different forms (as has been nicely described in the introduction of this paper). Because the studied system seems to be quite a specific example, it is questionable how far spread this phenomenon is in nature. In addition, it remains unclear how male harm will evolve in response to the climate crisis (see point 3). Finally, the relative fitness of females increased in the present experiment, as the tested range was within the reproductive optimum of the species. Nevertheless, the relative importance of the positive effect of sexual conflict on fitness outside of optimal temperatures seems questionable.

    Nonetheless, I believe these results to be of exceeding interest to the scientific community and of importance to the field. It opens up many potential research directions and adds further data to the fascinating field of sexual conflict, SFPs, and male harm in Drosophila.

  6. eLife

    Reviewer #3 (Public Review):

    In this paper, the authors explore the effects of the environment, specifically temperature, on male harm to females. Male harm is the phenomenon where males reduce female fitness in polyandrous systems, where a single female may mate with multiple males. The selection of males to increase their reproductive success in male-male competition can lead to genetic conflict that increases male fitness at the expense of female fitness. Typically, male harm has been studied in single environments under optimal conditions. However, there is an increasing focus on the effect of the environment on fitness costs of male harm to females, as a way to better understand the effect of male harm on population fitness in more realistic ecological contexts. In this paper, the authors add to these studies by exploring the effect of temperature on male harm and female fitness, using the fruit fly Drosophila melanogaster, as a model system. They find that temperature affects the impact of male harm on female fitness, with male harm having the greatest effect at 24˚C relative to 20˚C and 28˚C. The authors then go on to disentangle how temperature affects the various components of male harm that impact female fitness (e.g. harassment, ejaculate toxicity). The paper demonstrates that male harm depends on ecological context, which has implications for understanding its impact on population fitness under realistic ecological scenarios, particularly with respect to climate change.

    The strength of the paper is that it demonstrates that male harm (presented as differences in female life reproductive success between monogamous and polyandrous matings) changes with temperature. The authors dissect this general observation by showing that different aspects of pre-copulatory reproductive behavior, for example, male-male aggression, copulation rate, and female rejection rate, also change with temperature. Further, they demonstrate that correlates for male ejaculate quality also change with temperature, suggesting that temperature also affects post-copulatory mechanisms of male harm.

    The weakness of the paper is that the method and results section are difficult to follow, which negatively impacts the interpretation of the data. The experiments are complex and need to be for what the authors are studying. Nevertheless, the paper is written in a way that makes it challenging for the reader to fully understand how precisely the experiments were conducted. Further, the authors do not explain clearly how some of the experiments relate to the phenomenon ostensibly being assayed. For example, a more detailed explanation of why mating duration and remating latency are assays for ejaculate quality in the context of sperm competition would be very helpful in interpreting the data. Further, a clearer explanation of the statistical analyses conducted

  7. Version published to 10.1101/2022.11.10.515924v2 on bioRxiv
  8. Version published to 10.1101/2022.11.10.515924v1 on bioRxiv