We are looking for a highly motivated student to apply to the E2M2 doctoral school (University of Lyon 1) on the following topic: Ageing in ejaculate quality in a wild mammal
•Context and objectives of the PhD project:
Decades of studies in the wild have revealed that reproductive ageing (i.e. the decline in reproductive performance with increasing age) is the rule rather than the exception in mammalian females (Lemaître et al. 2020). However, the occurrence of reproductive ageing in males was for a long time largely dismissed by evolutionary ecologists. This bias can be explained by both methodological (e.g. the difficulty of assigning paternity in absence of molecular tools) and biological (e.g. the long-lasting belief of a germline shielded from the accumulation of damages, the « Weismann Barrier”) causes. As a consequence, evidence that males also suffer from a decline in reproductive performance with increasing age in the wild has only emerged recently. Overall, male reproductive ageing is in fact also pervasive in mammals as revealed by a meta-analysis performed by our team (i.e. reproductive ageing detected in at least 60% of the species studied, Cambreling et al., in review). The next step is now to decipher the biological and physiological determinants of male reproductive ageing in the wild.
In most mammals, males are successively confronted with two types of reproductive competition (Simmons et al. 2017). First, they compete for gaining mating opportunities (i.e. precopulatory competition) and their success in the ability to secure mating is therefore determined by the allocation to pre-copulatory sexual traits (e.g. weapon size, secretion of olfactory compounds). Second, they compete for gaining fertilization opportunities (i.e. post-copulatory competition). Indeed, when females mate with different males during a given period of sexual receptivity, sperm from these different males compete to fertilize one or several oocytes (Simmons et al. 2017). Male fertilization success under post-copulatory competition is mostly determined by post-copulatory sexual traits (e.g. sperm quantity and quality, Pizzari & Parker 2009). Any decline in male allocation to pre- and/or post-copulatory sexual traits could thus contribute to the overall decline in reproductive success (Lemaître & Gaillard 2017). So far, ageing in pre-copulatory traits has been relatively well documented. For instance, a recent compilation of published studies has notably revealed that senescence in antler length is pervasive (and modulated by environmental conditions) across cervids (Cambreling et al. 2023). However, while ageing in pre-copulatory traits appears to be widespread in the wild, our knowledge regarding the occurrence of ageing in post-copulatory traits in wild mammals, as well as on the ecological and biological factors that could modulate its magnitude, remains scarce.
The aim of this PhD project is to quantify the decline in ejaculate quality (sperm quantity and quality) with increasing male age, as well as to identify some of its biological and ecological drivers using a population of roe deer (Capreolus capreolus) intensively monitored at the level of individual as a biological model. This project will be divided in three axes.
Axis 1: Quantifying ageing in ejaculate quality in roe deer
The PhD student will test the hypothesis that both sperm quality and quantity decline with increasing age in male roe deer. Indeed, recent advances from studies performed in men as well as in laboratory and domestic animals have revealed that male gametes are far from being ageing-proof (Monaghan & Metcalfe 2019) and that the decline in sperm-related traits can even be exacerbated by some aspects of somatic ageing (e.g. degradation of the testicular tissue). However, if evidence of ageing in sperm traits (e.g. concentration and motility) has been documented in humans and laboratory rodents (Johnson et al. 2015, Sanghvi et al. 2024), the picture appears much less clear in mammals in the wild (Sanghvi et al. 2024). The difficulty of studying ejaculate ageing in the wild can largely be attributed to various methodological challenges including the use of non-lethal techniques to collect ejaculates, a sampling strategy encompassing individuals spanning the full age range of the species, and a collection of repeated samples per individual to control for selective processes known to bias the detection of ageing. The methodological approach we recently developed (see Methods below) will allow the PhD student to overcome these methodological challenges and to quantify accurately ageing patterns in ejaculate quality in roe deer.
Axis 2: Assessing the pattern of covariation between secondary sexual traits and sperm-related traits throughout the male life course.
The PhD student will test the hypothesis that the allocation to conspicuous antlers (e.g. long antlers) is a honest signal of individual fertility, and thus a reliable predictor of ejaculate quality, at least during early adulthood. Indeed, antler is a condition-dependent sexual trait, with males of highest quality being able to grow every year particularly long antlers (e.g. Vanpé et al. 2007 in roe deer), and data from hunted red deer (Cervus elaphus) have revealed antler size is positively associated with measures of sperm velocity (Malo et al. 2005). However, the relationship between these pre- and post-copulatory sexual traits has never been studied from an ageing perspective. Yet, we can predict that the decline in body mass with increasing age, which occurs at around 6.5 years of age in the population of Trois-Fontaines (Douhard et al. 2017) will reinforce the resource-based allocation trade-off expected to occur between pre- and post-copulatory traits (Simmons et al. 2017). Old males could thus be expected to cheat in order to maintain their phenotypic quality and thereby copulate with females, leading them to favour allocation to antler growth at the expense of sperm-related traits. These analyses will include various measures of phenotypic quality ranging from body condition to locomotory performance.
Axis 3: Investigating the role of heavy metal exposure on ejaculate quality ageing
The PhD student will test the hypothesis that exposure to various toxic metals (e.g. lead, cadmium, mercury, arsenic, aluminium, chromium) will have detrimental impact on sperm-related traits, notably through an acceleration of the age-specific decline in both sperm quantity and quality in male roe deer. The growing diversity of chemical substances produced and released into the environment is indeed a major concern for wildlife. It is now recognized that the majority of organisms are exposed to various contaminants of anthropogenic origin, and that this exposure can compromise their reproduction (as fully recognized in men, Crocetto et al. 2023). However, while the short-term effects of acute exposure to various contaminants, usually considered individually, have been widely studied, the long-term effects of chronic exposure to low doses of multiple chemicals are much less documented, especially in the wild. Although a recent study conducted by our team suggests that cadmium exposure is associated to smaller testes in male roe deer of unknown age (Vuarin et al. unpublished data), whether such exposure could modulate reproductive ageing remains to be tested as it could have lasting unappreciated consequences on ejaculate quality ageing.
• Methods:
This project will be based on the monitoring of roe deer (Capreolus capreolus) in the Territoire d’Etude et d’Expérimentation (TEE) of the Trois-Fontaines forest (northeast France), a long-lasting project (50 years) conducted via a collaboration between the Office Français de le Biodiversité (OFB) and the LBBE. This population is the subject of a capture-mark-recapture program for 50 years (Gaillard et al., 1993). Every winter, captures are organized and various phenotypic measurements (e.g. body mass, antler length) and biological samples (e.g. blood, hair) are collected from known age individuals. For the past 2 years, we have started to perform additional captures in summer (from mid-June to end of July) just before the rutting season, where we specifically target males. We have successfully implemented a non-lethal protocol for ejaculate collection. Sperm quality and quantity data are already available for more than 50 collected samples in 2023-2024, encompassing 44 individuals ranging from 1 to 10 years old, and thus already covering a large range of the roe deer lifespan. We expect to collect a minimum of 30-35 samples in the summer of 2025. In addition, the PhD student will pursue data collection on a similar number of males, in close collaboration with the field team, during the first two summers of the PhD project (2026/2027), leading to a sample size of at least 150 samples for the analyses. The PhD student will thus estimate a various range of sperm quality parameters following golden-standard methods in Sperm Computer Analysis (SCA). All procedures have been approved by the Ethical Committee of Lyon 1 University.
Complementary data such as antler size, body mass, and body size (hind foot length) are (and will be) also collected during the summer captures, as well as biological samples such as hair that will be further used for trace metal analysis. These analyses will enable the quantification of long-term effects of chronic exposure to low doses of 22 essential and non-essential metals in all sampling males (see Herrada et al., 2024). These analyses will be carried out by the Chrono-environnement laboratory (UMR 6249 CNRS-Université de Besançon), with which a collaboration has already been initiated (‘IMPULSION’ project of Pauline Vuarin). Trace metals will also be quantified in seminal fluid in order to test whether contaminants accumulate in this matrix and if so, whether it correlates with ejaculate quality. Finally, thanks to a new collaboration between the LBBE and the Laboratoire Interdisciplinaire de Biologie de la Motricité (LIBM) of the University Savoie Mont Blanc, males will be equipped with biologgers enabling the quantification of fine scale movement patterns and their energetic costs.
Statistical methods that will be used to test the hypotheses from the 3 different axes will mainly involve principal component analyses (PCA) and generalized linear mixed effect models (GLMER) to deal with repeated measurements over individuals’ life.
• Expected results:
This PhD project should lead to an in-depth understanding of ageing in ejaculate quality by notably providing new insight regarding the occurrence and the drivers of ageing in ejaculate quality. Moreover, we expect that our analyses performed on roe deer (a species known to be a relevant indicator of environmental contamination, see Draghi et al., 2023) will generate robust results regarding the effects of pollutants exposure on sperm quantity and quality, a topic that constitutes a major public health issue for wildlife and human populations. We expect to publish a minimum of one paper per axis of this PhD project.
• Profil and skills required:
We aim to recruit a PhD student with a solid background in at least one of the following areas: evolutionary biology, biology of ageing, physiology of reproduction or biostatistics. We are looking for a student with excellent organizational skills and who is also extremely rigorous, both in terms of data handling and statistical processing. A good knowledge of R programming is required. The student will participate in the roe deer captures and in the collection of biological tissues (e.g. sperm samples) from anesthetized animals during field sessions. The student must therefore be comfortable with the idea of handling animals in the field. Overall, she/he will spend a few weeks in the TEE of Trois-Fontaines for data collection (basically a few days in January/February for the winter captures and 4/5 weeks from mid-June to late-July for the summer captures). It is important to note that data collection in the field involves fairly long time slots (with experimentations that can finish at 10:00 pm / 11:00 pm). Occasionally, field activities may also take place during weekends. Importantly, the PhD student will have access to a regulatory training in animal experimentation that will take place at Lyon, and which is mandatory when using animals for scientific purposes. A driving licence is not mandatory but would constitute a valuable additional asset.
• Research environment:
The PhD student will be based in the Laboratoire de Biométrie et Biologie Evolutive (LBBE, UMR CNRS 5558, University of Lyon 1) and more specifically in the Evolutionary Biodemography team. The student will have regular exchange with the researchers involved in the roe deer project, the field technicians and engineers from the LBBE’s biotechnology unit, and staff from the Office Français de la Biodiversité (OFB). If she/he wishes, the PhD student will have the opportunity to join the LBBE doctoral students’ association: Les Pinsons migRateurs. The aim of this friendly association is to maintain and promote scientific animation, social mediation and cohesion activities within the non-permanent staff of the LBBE, as well as to help the professional integration of young researchers.
• Contact:
Students interested in this research project can send an email to Jean-François Lemaître (jean-francois.lemaitre@univ-lyon1.fr) with cc to Pauline Vuarin (Pauline.vuarin@univ-lyon1.fr) and Jean-Michel Gaillard (jean-michel.gaillard@univ-lyon1.fr).
• References:
– Cambreling, S., Gaillard, J. M., Pellerin, M., Vanpé, C., Débias, F., Delorme, D., … & Lemaître, J. F. (2023). Natal environmental conditions modulate senescence of antler length in roe deer. Frontiers in Ecology and Evolution, 11, 1139235.
– Cambreling, S. Ronget, V., Remot, F., Gaillard, J-M. & Lemaître, J-F. (Under review) Male reproductive senescence in mammals is pervasive and aligned with the slow-fast continuum.
– Crocetto, F., Risolo, R., Colapietro, R., Bellavita, R., Barone, B., Ballini, A., … & Brancorsini, S. (2023). Heavy metal pollution and male fertility: an overview on adverse biological effects and socio-economic implications. Endocrine, Metabolic & Immune Disorders-Drug Targets, 23(2), 129-146.
– Douhard, F., Gaillard, J. M., Pellerin, M., Jacob, L., & Lemaître, J. F. (2017). The cost of growing large: Costs of post‐weaning growth on body mass senescence in a wild mammal. Oikos, 126(9), 1329-1338.
– Draghi, S., Agradi, S., Riva, F., Tarhan, D., Bilgiç, B., Dokuzeylül, B., … & Curone, G. (2023). Roe Deer (Capreolus capreolus) hair as a bioindicator for the environmental presence of toxic and trace elements. Toxics, 11(1), 49.
– Gaillard, J. M., Delorme, D., Boutin, J. M., Van Laere, G., Boisaubert, B., & Pradel, R. (1993). Roe deer survival patterns: a comparative analysis of contrasting populations. Journal of Animal Ecology, 778-791.
– Herrada, A., Bariod, L., Saïd, S., Rey, B., Bidault, H., Bollet, Y., … & Vuarin, P. (2024). Minor and trace element concentrations in roe deer hair: A non-invasive method to define reference values in wildlife. Ecological Indicators, 159, 111720.
– Johnson, S. L., Dunleavy, J., Gemmell, N. J., & Nakagawa, S. (2015). Consistent age-dependent declines in human semen quality: a systematic review and meta-analysis. Ageing research reviews, 19, 22-33.
– Lemaître, J. F., & Gaillard, J. M. (2017). Reproductive senescence: new perspectives in the wild. Biological Reviews, 92(4), 2182-2199.
– Lemaître, J. F., Ronget, V., & Gaillard, J. M. (2020). Female reproductive senescence across mammals: a high diversity of patterns modulated by life history and mating traits. Mechanisms of Ageing and Development, 192, 111377.
– Malo, A. F., Roldan, E. R., Garde, J., Soler, A. J., & Gomendio, M. (2005). Antlers honestly advertise sperm production and quality. Proceedings of the Royal Society B: Biological Sciences, 272(1559), 149-157.
– Monaghan, P., & Metcalfe, N. B. (2019). The deteriorating soma and the indispensable germline: gamete senescence and offspring fitness. Proceedings of the Royal Society B, 286(1917), 20192187.
– Pizzari, T., & Parker, G. A. (2009). Sperm competition and sperm phenotype. Sperm biology, 207-245.
– Sanghvi, K., Vega-Trejo, R., Nakagawa, S., Gascoigne, S. J., Johnson, S. L., Salguero-Gómez, R., … & Sepil, I. (2024). Meta-analysis shows no consistent evidence for senescence in ejaculate traits across animals. Nature communications, 15(1), 558.
– Simmons, L. W., Lüpold, S., & Fitzpatrick, J. L. (2017). Evolutionary trade-off between secondary sexual traits and ejaculates. Trends in ecology & evolution, 32(12), 964-976.
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