Population density effects on gamete traits and fertilisation dynamics under varying sperm environments in mussels
Craig D.H Sherman1*, Vincent Careau2, Clelia Gasparini3, Kim J. Weston1, Jonathan P. Evans4
1School of Life and Environmental Sciences, Deakin University, Victoria 3216, Australia.
2Department of Biology, University of Ottawa, Ottawa, ON, Canada.
3Department of Biology, University of Padova, Padova, Italy
4Centre for Evolutionary Biology, School of Biological Sciences, University of Western Australia, Crawley 6009, WA, Australia
*Corresponding author
E-mail address: craig.sherman@deakin.edu.au
Running title: Gamete traits and fertilisation dynamics
Gamete traits can vary widely among species, populations and individuals, influencing fertilisation dynamics and overall reproductive fitness. Sexual selection can play an important role in determining the evolution of gamete traits with local environmental conditions determining the strength and direction of sexual selection. Here we test for signatures of post-mating selection on gamete traits in relation to population density, and possible interactive effects of population density and sperm concentration on sperm motility and fertilisation rates among natural populations of mussels. Our study shows that males from high density populations produce smaller sperm compared with males from low density populations, but we detected no effect of population origin on egg size. Our results also reveal that females from low density populations tended to exhibit lower fertilisation rates across a range of sperm concentrations, although this became less important as sperm concentration increased. Variances in fertilisation success were higher for females than males and the effect of gamete compatibility between males and females increases as sperm concentrations increase. These results suggest that local population density can influence gamete traits and fertilisation dynamics but also highlight the importance of phenotypic plasticity in governing sperm-egg interactions in a highly dynamic selective environment.
KEY WORDS : sexual selection, sperm competition, polyspermy, maternal effect, genotype by environment, cryptic female choice.
It is now widely recognised that in most sexually reproducing taxa sexual selection can continue after mating in the form of sperm competition, where ejaculates from rival males compete for fertilisation (Parker 1970), and cryptic female choice, where females influence the outcome of these contests (Eberhard 1996; Birkhead and Moller 1998; Parker 2020). This has resulted in a myriad of adaptations in both sexes; for example, selection can favour specific gamete traits that make ejaculates more competitive in the race to fertilise eggs (Pizzari and Parker 2009; Simmons and Fitzpatrick 2012; Lüpold et al. 2020), gamete plasticity in response to local environmental conditions (Crean and Marshall 2008), sperm chemoattractants that function to attract sperm from specific (e.g. compatible) males (Evans et al. 2012; Oliver and Evans 2014; Kekäläinen and Evans 2017; Lymbery et al. 2017), or gamete recognition proteins that ultimately determine whether sperm can fuse with an egg (Palumbi 1999; Swanson and Vacquier 2002; Levitan and Ferrell 2006; Evans and Sherman 2013).
Post-mating sexual selection has been argued to be particularly important in broadcast spawning organisms that release their gametes into the water for external fertilisation (reviewed in Evans and Sherman 2013; Kekäläinen and Evans 2018; Evans and Lymbery 2020). Indeed, one of the only opportunities for mate choice and mating competition in broadcast spawning organisms occurs through gamete-level interactions. However, broadcast spawning marine invertebrates face unique challenges that are linked to the spawning environment. In particular, variation in adult population density, and therefore the density of gametes in the water, is predicted to influence the intensity and direction of post-mating sexual selection (Parker et al. 1997; Franke et al. 2002; Levitan 2004; Levitan and Ferrell 2006; Marshall and Bolton 2007; Sherman et al. 2015; Hadlow et al. 2022). In low density populations, the rapid dilution of gametes can result in low fertilisation rates (Vogel et al. 1982; Levitan and Petersen 1995; Styan and Butler 2000; Levitan 2004), thereby generating selection on gamete traits in both sexes to increase the probablity of egg-sperm encounters and fusion. For example, in females this may include the production of fewer, larger eggs to increase egg-sperm collision rates, greater investment in sperm-attracting chemoattractants, greater egg longevity, and less selective membrane blocks to sperm penetration (Yund 2000; Riffell et al. 2004; Marshall and Evans 2005; Levitan 2006; Levitan and Ferrell 2006; Kosman and Levitan 2014; Evans and Lymbery 2020). Under these sperm-limiting conditions males will maximixe their fitness by increasing gamete encounter rates, for example by producing a smaller number of larger and/or longer lived sperm or altering swimming patterns (Parker 1993; Benzie and Dixon 1994; Parker 1998; Yund 2000; Snook 2005) (but see Crean and Marshall 2008).
At the other end of the population density spectrum, males from high-density populations face increased sperm competition, while females face an increased risk of polyspermy - where eggs are fertilized by two or more sperm (usually resulting in cell or zygote death). As a consequence, selection may favour female reproductive strategies that minimise the risk of polyspermy. Specifically, when sperm are abundant and the risks of polyspermy are high, females are predicted to produce smaller eggs, invest less in sperm-attracting chemoattractants, or increase the selectivity of membrane gamete recognition systems to reduce the risk of lethal polyspermy (Levitan and Ferrell 2006; Levitan et al. 2007; Kamel et al. 2010; Kosman and Levitan 2014). Males, on the other hand should produce higher numbers of smaller sperm (Parker 1998; Snook 2005; Parker 2006) or release smaller quantities of sperm over a longer period of time (Bode and Marshall 2007; Marshall and Bolton 2007). Consequently, as population density (and sperm abundance) increases, selection should favour reproductive strategies in males that further exacerbate the reproductive costs incurred by females (polyspermy) (Evans and Lymbery 2020).This complex interplay between population density and the differential selection pressures faced by males and females can lead to sexual conflict, where the optimal mating strategy of one sex reduces the reproductive fitness of the other (Parker 2006; Kamel et al. 2010). Despite this prediction, however, there are only a limited number of studies that have explored selection on gamete traits and fertilization dynamics from natural populations with varying adult densities (however, see Levitan 2002, 2004; Levitan and Ferrell 2006; Levitan 2012).
Blue mussels of the genus Mytilus are external spawners and provide an ideal experimental system for understanding the combined effects of population density, gamete traits and sperm abundance on fertilisation dynamics (Evans et al. 2012; Sherman et al. 2015; Evans and Lymbery 2020). Blue mussels are found on hard substrates of nearshore and intertidal habitats in temperate and subarctic regions of the northern and sourthern hemispheres (Hilbish et al. 2000) and are predominant species found in temperate Australian waters (Westfall and Gardner 2013; Ab Rahim et al. 2016; Popovic et al. 2020; Zbawicka et al. 2021). Populations vary naturally in population abundance (Cockrell et al. 2015) with likely concomitant variation in sperm concentrations among natural spawning events. Moreover, recent work on AustralianMytilus has revealed that ecologically relevant variation in sperm concentrations can have dramatic effects on fertilisation kinetics (Sherman et al. 2015). Furthermore, there is evidence fromMytilus for strong patterns of gamete selectivitiy, such that specific combinations of gametes from both sexes generate predictable differences in fertilisation rates and offspring viability (Evans et al. 2012; Oliver and Evans 2014; Sherman et al. 2015; Lymbery et al. 2017). However, the extent to which such patterns depend on the fertilisation environment has yet to be determined.
In this study we explore the effect of adult population density on variation in gamete traits and fertilisation dynamics across a range of sperm concentration environments in mussels from natural populations. We expected that selection will have acted on gamete traits in both sexes to optimise fertilization success under different egg-sperm ratios. Specifically, we explore whether population origin influences gamete traits in both sexes, and whether experimental adjustments to sperm-egg ratios during in vitro fertilisation trials influence fertilisation kinetics and pairwise patterns of fertilisation rates (i.e. gamete compatibility). One expectation from the latter trials is that eggs will become more selective as the risk of polyspermy (high sperm concentrations) increase, resulting stronger gametic (male-by-female) interactions under high sperm densities.