Sperm collection in Black-legged Kittiwakes and characterization of sperm velocity and morphology

Background:Collecting and studying live sperm is central to many important fields of biology.Yet,a simple method to collect live sperm is lacking in wild seabird species.Here,we describe a non?invasive method to collect viable sperm samples based on a simple massage technique applied to male Black?legged Kittiwakes(Rissa tridactyla).Methods:We studied a colony breeding at Kongsfjorden,Svalbard and successfully obtained sperm samples from 32 males.With a subset of samples(n = 12 males),we compared the suitability of several extenders(0.9% NaCl,PBS,Earle's balance salt solution,Dulbecco's modified Eagle medium) in maintaining sperm alive long enough for analyses.With another 18 ejaculates,we conducted computer assisted sperm analyses using the CASA plugin for ImageJ.We provide details about the settings to be used for such analyses.Lastly,droplets from 20 ejaculates were smeared on glass slides and preserved with formalin to characterize sperm morphology in terms of total sperm length,sperm head length,midpiece length and flagellum length,and percentage of abnormal sperm.Results:With this method and under field conditions,we were able to obtain sufficient amounts of live sperm to assess traits related to sperm quality(e.g.sperm morphology,percentage of motile sperm,sperm velocity).We found that two extenders,Earle's balanced salt solution and Dulbecco modified Eagle's medium,yielded similarly good results.Additionally,we investigated whether specific behaviours were associated with successful sperm collection and whether sperm collection success depended on how long before laying sperm collection was attempted.Finally,we provide mean values for sperm morphology,sperm swimming ability and percentage of motile sperm,which may prove useful for future comparative analyses,and we report high levels of sperm abnormality and within?ejaculate variation in sperm morphology.Conclusions:We discuss the high percentage of abnormal sperm and high within?ejaculate variation in sperm morphology in light of sperm competition theory and conclude that these figures are likely due to relaxed post?cop?ulatory sexual selection,kittiwakes being strictly monogamous.Finally,we suggest that this method could be applied to other seabird species sharing similar ecology.

Experimental manipulation of reproductive tactics in Seba’s short-tailed bats:consequences on sperm quality and oxidative status

To reproduce,males have to fertilize the female’s eggs,sometimes in competition with ejaculates of other males.In species where males display alternative reproductive tactics,whereby territorial males secure mating and non-territorial males have to sneak copulations,the latter might be expected to invest relatively more resources towards sperm quality compared with the territorial males.Sperm cells are especially vulnerable to oxidative stress,which reduces male fertility.Therefore,antioxidant resources are expected to modulate sperm quality,and might be allocated differently between reproductive tactics.To test the link between reproductive tactics,redox profile and sperm quality,we experimentally induced changes in the reproductive tactics of 39 captive males Seba's short-tailed bats Carollia perspicillata.We monitored the blood and ejaculate oxidative balance,and the sperm quality before,7 days and 21 days after the manipulation of reproductive tactic.Although ejaculates'oxidative damage was negatively related to sperm velocity,males exhibited similar blood and ejaculates redox profiles and similar sperm quality,regardless of their reproductive tactic.Possibly,these results arise as a consequence of some constraints having been lifted during the experiment.Our results also suggest that,in Seba's short-tailed bats,the expression of alternative reproductive tactics is not subjected to strong oxidative constraints.Furthermore,our results could reflect an absence of trade-off between pre-and post-copulatory traits in harem males,as they could be selected to invest both in female attraction and sperm quality,as a consequence of their inability to fully monopolize females.