Ocean acidification alters sperm responses to egg-derived chemicals in a broadcast spawning mussel

The continued emissions of anthropogenic carbon dioxide are causing progressive ocean acidification (OA). While deleterious effects of OA on biological systems are well documented in the growth of calcifying organisms, lesser studied impacts of OA include potential effects on gamete interactions that determine fertilization, which are likely to influence the many marine species that spawn gametes externally. Here, we explore the effects of OA on the signalling mechanisms that enable sperm to track egg-derived chemicals (sperm chemotaxis). We focus on the mussel Mytilus galloprovincialis, where sperm chemotaxis enables eggs to bias fertilization in favour of genetically compatible males. Using an experimental design based on the North Carolina II factorial breeding design, we test whether the experimental manipulation of seawater pH (comparing ambient conditions to predicted end-of-century scenarios) alters patterns of differential sperm chemotaxis. While we find no evidence that male–female gametic compatibility is impacted by OA, we do find that individual males exhibit consistent variation in how their sperm perform in lowered pH levels. This finding of individual variability in the capacity of ejaculates to respond to chemoattractants under acidified conditions suggests that climate change will exert considerable pressure on male genotypes that can withstand an increasingly hostile fertilization environment.     

Female‐induced remote regulation of sperm physiology may provide opportunities for gamete‐level mate choice

In sedentary externally fertilizing species, direct interactions between mating partners are limited and prefertilization communication between sexes occurs largely at the gamete level. Certain combinations of eggs and sperm often have higher fertilization success than others, which may be contingent on egg‐derived chemical factors that preferentially attract sperm from compatible males. Here, we examine the mechanisms underlying such effects in the marine mussel Mytilus galloprovincialis, where differential sperm attraction has recently been shown to be associated with variation in offspring viability. Specifically, we focus on the sperm surface glycans, an individually unique layer of carbohydrates that moderate self‐recognition and other cellular‐level interactions. In many species egg‐derived factors trigger remarkable changes in the sperm's glycan layer, physiology, and swimming behavior, and thus potentially moderate mate choice at the gamete level. Here, we show that sperm glycan modifications and the strength of acrosome reaction are both dependent on specific male–female interactions (male–female combination). We also find associations between female‐induced sperm glycan changes and the Ca²⁺ influx into sperm–‐a key regulator of fertilization processes from sperm capacitation to gamete fusion. Together, our results suggest that female‐induced remote regulation of sperm physiology may constitute a novel mechanism of gamete‐level mate choice.     

Chemically moderated gamete preferences predict offspring fitness in a broadcast spawning invertebrate

Sperm chemoattraction, where sperm locate unfertilized eggs by following a concentration gradient of egg-derived chemoattractants, has been widely documented across numerous taxa. While marine invertebrates are favoured models for understanding the underlying mechanisms of sperm chemoattraction, the evolutionary forces underpinning the process remain enigmatic. Here, we show that in mussels (Mytilus galloprovincialis), chemically moderated gamete preferences promote assortative fertilizations between genetically compatible gametes. When offered the choice of egg clutches from two females, sperm exhibited consistent but differential ‘preferences’ for chemical cues secreted from conspecific eggs. Critically, our data reveal that the preferences shown by sperm during the egg-choice trials are highly predictive of early embryonic viability when eggs and sperm from the same individuals are mixed during standard (no-choice) fertilization assays. Moreover, we demonstrate that by experimentally separating chemoattractants from eggs, sperm swimming behaviour is differentially regulated by egg-derived chemoattractants, and that these changes in sperm behaviour are highly consistent with observed patterns of gamete preferences, fertilization and larval survival. Together, this integrated series of experiments reveals that the behaviour of sperm is fine-tuned to respond differentially to the chemical signals emitted from different conspecific eggs, and that these choices have measurable fitness benefits.     

Egg‐induced changes to sperm phenotypes shape patterns of multivariate selection on ejaculates

Sperm cells exhibit extraordinary phenotypic diversity and rapid rates of evolution, yet the adaptive value of most sperm traits remains equivocal. Recent findings suggest that to understand how selection targets ejaculates, we must recognize that female‐imposed physiological conditions often alter sperm phenotypes. These phenotypic changes may influence the relationships among sperm traits and their association with fitness. Here, we show that chemical substances released by eggs (known to modify sperm physiology and behaviour) alter patterns of selection on a suite of sperm traits in the mussel Mytilus galloprovincialis. We use multivariate selection analyses to characterize linear and nonlinear selection acting on sperm traits in (a) seawater alone and (b) seawater containing egg‐derived chemicals (egg water). Our analyses revealed that nonlinear selection on canonical axes of multiple traits (notably sperm velocity, sperm linearity and percentage of motile sperm) was the most important form of selection overall, but importantly these patterns were only evident when sperm phenotypes were measured in egg water. These findings reveal the subtle way that females can alter patterns of selection, with the implication that overlooking environmentally moderated changes to sperm, may result in erroneous interpretations of how selection targets phenotypic (co)variation in sperm traits.     

In vitro fertilization experiments using sockeye salmon reveal that bigger eggs are more fertilizable under sperm limitation

Although theory and widespread evidence show that the evolution of egg size is driven primarily by offspring and maternal fitness demands, an additional explanation invokes sperm limitation as a selective force that could also influence egg size optima. Levitan proposed that constraints from gamete encounter in external fertilization environments could select for enlargement of ova to increase the physical size of the fertilization target. We test this theory using in vitro fertilization experiments in an externally fertilizing fish. Sockeye salmon (Onchorhyncus nerka) females show considerable between-individual variation in ovum size, and we explored the consequences of this natural variation for the fertilization success of individual eggs under conditions of sperm limitation. By engineering consistent conditions where in vitro fertilization rate was always intermediate, we were able to compare the sizes of fertilized and unfertilized eggs across 20 fertilization replicates. After controlling for any changes in volume through incubation, results showed that successfully fertilized eggs were significantly larger than the eggs that failed to achieve fertilization. Under conditions without sperm limitation, fertility was unaffected by egg size. Our findings therefore support Levitan''s theory, demonstrating empirically that some element of egg size variation could be selected by fertilization demands under sperm limitation. However, further research on sperm limitation in natural spawnings is required to assess the selective importance of these results.     

Ejaculate evolution in external fertilizers: Influenced by sperm competition or sperm limitation?

The evolution of sperm quality and quantity is shaped by various selective processes, with sperm competition generally considered the primary selective agent. Particularly in external fertilizers, however, sperm limitation through gamete dispersal can also influence gamete investments, but empirical data examining this effect are limited. Here, we studied the relative importance of sperm competition and the spawning conditions in explaining the macroevolutionary patterns of sperm size and number within two taxa with external fertilization but differences in their reproductive biology. In frogs, sperm swim slowly but for up to hours as they penetrate the gelatinous egg coating, whereas fish sperm typically swim fast, are very short‐lived (seconds to minutes), and often face a relatively higher risk of being moved away from the ova by currents. Our phylogenetic models and path analyses revealed different trajectories of ejaculate evolution in these two taxa. Sperm size and number responded primarily to variation in sperm competition in the anurans, but more strongly to egg number and water turbulence in the fishes. Whereas the results across anurans align with the general expectation that sexual selection is the main driver of ejaculate evolution, our findings across the fishes suggest that sperm limitation has been underappreciated.     

Length rather than year‐round spawning,affects reproductive performance of RAS‐reared F‐generation pikeperch,Sander lucioperca (Linnaeus, 1758) – Insights from practice

The continuous production of large numbers of high quality gametes is essential for aquaculture, particularly in candidate species, such as pikeperch, Sander lucioperca (L.). The common practice of year‐round reproduction is under suspicion of inflicting adverse effects on the quality of the gametes through the disturbance of endogenous rhythms. We hypothesized that such perturbation does not affect RAS‐reared F‐generation broodstock. Reproductive performance (number of eggs) and gamete quality (fertilization and hatching rate) were assessed over the course of 3 years covering six independent, photothermal shifted spawning seasons in a commercial pikeperch hatchery (n = 31 egg batches of F‐generation fish in total). No substantial differences in fertilization or hatching rates could be detected between the individual spawning seasons. Fecundity varied, but there are indications for a size effect on female fecundity with intermediate sized females producing higher number of eggs (~65–70 cm). Low egg quality could be detected in batches of very large fish. In conclusion, size‐specific broodstock composition, but not year‐round reproduction of F‐generation pikeperch spawners affects the reproductive performance.     

被子植物受精机制的研究进展

被子植物的受精是一个复杂而精巧的过程。花粉管到达子房,通过退化助细胞进入胚囊,释放出两个精细胞。原来在花粉管中相互联结的两个精细胞在退化助细胞中分开,一个与卵细胞融合,另一个与中央细胞融合,完成双受精。目前对双受精过程中有关雌、雄配子识别的机制还知之甚少。本文介绍了目前被子植物精、卵细胞融合前后的细胞周期变化、退化助细胞的功能、精细胞在退化助细胞中迁移的研究动态、精细胞的倾向受精和卵细胞的激活等被子植物受精生物学领域中的一些新的研究成果和发展趋势。     

Sperm mixing in the polyandrous leaf‐cutting ant Acromyrmex echinatior

The insemination of queens by sperm from multiple males (polyandry) has evolved in a number of eusocial insect lineages despite the likely costs of the behavior. The selective advantages in terms of colony fitness must therefore also be significant and there is now good evidence that polyandry increases genetic variation among workers, thereby improving the efficiency of division of labor, resistance against disease, and diluting the impact of genetically incompatible matings. However, these advantages will only be maximized if the sperm of initially discrete ejaculates are mixed when stored in queen spermathecae and used for egg fertilization in a “fair raffle.” Remarkably, however, very few studies have addressed the level of sperm mixing in social insects. Here we analyzed sperm use over time in the highly polyandrous leaf‐cutting ant Acromyrmex echinatior. We genotyped cohorts of workers produced either 2 months apart or up to over a year apart, and batches of eggs laid up to over 2 years apart, and tested whether fluctuations in patriline distributions deviated from random. We show that the representation of father males in both egg and worker cohorts does not change over time, consistent with obligatorily polyandrous queens maximizing their fitness when workers are as genetically diverse as possible.     

Assessing the potential for post‐ejaculatory female choice in a polyandrous beach‐spawning fish

In species with limited opportunities for pre‐ejaculatory sexual selection (behavioural components), post‐ejaculatory mechanisms may provide opportunities for mate choice after gametes have been released. Recent evidence from a range of taxa has revealed that cryptic female choice (i.e., female‐mediated differential fertilization bias), through chemical cues released with or from eggs, can differentially regulate the swimming characteristics of sperm from various males and ultimately determine male fertilization success under sperm competition. We assessed the potential role that such female‐modulated chemical cues play in influencing sperm swimming characteristics in beach‐spawning capelin (Mallotus villosus), an externally fertilizing fish that mates as couples (one male and one female) or threesomes (two males and one female) with presumably limited opportunities for pre‐ejaculatory sexual selection. We assayed sperm swimming characteristics under varying doses and donor origins of egg cues and also examined the possibility of assortative mating based on body size. We found mating groups were not associated by size, larger males did not produce better quality ejaculates, and egg cues (regardless of dosage or donor identity) did not influence sperm swimming characteristics. Our findings suggest that intersexual pre‐ejaculatory sexual selection and cryptic female choice mediated by female chemical cues are poorly developed in capelin, possibly due to unique natural selection constraints on reproduction.     

Sperm use economy of honeybee (Apis mellifera) queens

The queens of eusocial ants, bees, and wasps only mate during a very brief period early in life to acquire and store a lifetime supply of sperm. As sperm cannot be replenished, queens have to be highly economic when using stored sperm to fertilize eggs, especially in species with large and long‐lived colonies. However, queen fertility has not been studied in detail, so that we have little understanding of how economic sperm use is in different species, and whether queens are able to influence their sperm use. This is surprising given that sperm use is a key factor of eusocial life, as it determines the fecundity and longevity of queens and therefore colony fitness. We quantified the number of sperm that honeybee (Apis mellifera) queens use to fertilize eggs. We examined sperm use in naturally mated queens of different ages and in queens artificially inseminated with different volumes of semen. We found that queens are remarkably efficient and only use a median of 2 sperm per egg fertilization, with decreasing sperm use in older queens. The number of sperm in storage was always a significant predictor for the number of sperm used per fertilization, indicating that queens use a constant ratio of spermathecal fluid relative to total spermathecal volume of 2.364 × 10^?6 to fertilize eggs. This allowed us to calculate a lifetime fecundity for honeybee queens of around 1,500,000 fertilized eggs. Our data provide the first empirical evidence that honeybee queens do not manipulate sperm use, and fertilization failures in worker‐destined eggs are therefore honest signals that workers can use to time queen replacement, which is crucial for colony performance and fitness.     

Cross-species fertilization: the hamster egg receptor,Juno, binds the human sperm ligand,Izumo1

Fertilization is the culminating event in sexual reproduction and requires the recognition and fusion of the haploid sperm and egg to form a new diploid organism. Specificity in these recognition events is one reason why sperm and eggs from different species are not normally compatible. One notable exception is the unusual ability of zona-free eggs from the Syrian golden hamster (Mesocricetus auratus) to recognize and fuse with human sperm, a phenomenon that has been exploited to assess sperm quality in assisted fertility treatments. Following our recent finding that the interaction between the sperm and egg recognition receptors Izumo1 and Juno is essential for fertilization, we now demonstrate concordance between the ability of Izumo1 and Juno from different species to interact, and the ability of their isolated gametes to cross-fertilize each other in vitro. In particular, we show that Juno from the golden hamster can directly interact with human Izumo1. These data suggest that the interaction between Izumo1 and Juno plays an important role in cross-species gamete recognition, and may inform the development of improved prognostic tests that do not require the use of animals to guide the most appropriate fertility treatment for infertile couples.     

Selection for high gamete encounter rates explains the success of male and female mating types

Sexual reproduction occurs in many small eukaryotes by fusion of similar gametes (isogamy). In the absence of distinguishable sperm and eggs, male and female mating types are missing. However, species with distinct males and females have so prospered that almost all familiar plants and animals have these mating types. Why has sexual reproduction involving sperm and eggs been so successful? An answer is obtained by considering physical limitations on encounter rates between gametes. A biophysical model based on well-established relationships produces fitness landscapes for the evolution of gamete size and energy allocation between motility and pheromone production. These landscapes demonstrate that selection for high gamete encounter rates favors large, pheromone-producing eggs and small, motile sperm. Thus, broadcast-spawning populations with males and females can reproduce at lower population densities and survive under conditions where populations lacking males and females go extinct. It appears that physical constraints on gamete encounter rates are sufficient to explain the first two steps in the isogamy-->anisogamy-->oogamy-->internal fertilization evolutionary sequence observed in several lineages of the eukaryotes. Unlike previous models, assumptions concerning zygote fitness or decreasing speed of swimming with increasing gamete size are not required.     

Releasing small ejaculates slowly increases per‐gamete fertilization success in an external fertilizer: Galeolaria caespitosa (Polychaeta: Serpulidae)

The idea that male reproductive strategies evolve primarily in response to sperm competition is almost axiomatic in evolutionary biology. However, externally fertilizing species, especially broadcast spawners, represent a large and taxonomically diverse group that have long challenged predictions from sperm competition theory—broadcast spawning males often release sperm slowly, with weak resource‐dependent allocation to ejaculates despite massive investment in gonads. One possible explanation for these counter‐intuitive patterns is that male broadcast spawners experience strong natural selection from the external environment during sperm dispersal. Using a manipulative experiment, we examine how male reproductive success in the absence of sperm competition varies with ejaculate size and rate of sperm release, in the broadcast spawning marine invertebrate Galeolaria caespitosa (Polychaeta: Serpulidae). We find that the benefits of Fast or Slow sperm release depend strongly on ejaculate size, but also that the per‐gamete fertilization rate decreases precipitously with ejaculate size. Overall, these results suggest that, if males can facultatively adjust ejaculate size, they should slowly release small amounts of sperm. Recent theory for broadcast spawners predicts that sperm competition can also select for Slow release rates. Taken together, our results and theory suggest that selection often favours Slow ejaculate release rates whether males experience sperm competition or not.     

CRYPTIC CHOICE OF CONSPECIFIC SPERM CONTROLLED BY THE IMPACT OF OVARIAN FLUID ON SPERM SWIMMING BEHAVIOR

Despite evidence that variation in male–female reproductive compatibility exists in many fertilization systems, identifying mechanisms of cryptic female choice at the gamete level has been a challenge. Here, under risks of genetic incompatibility through hybridization, we show how salmon and trout eggs promote fertilization by conspecific sperm. Using in vitro fertilization experiments that replicate the gametic microenvironment, we find complete interfertility between both species. However, if either species’ ova were presented with equivalent numbers of both sperm types, conspecific sperm gained fertilization precedence. Surprisingly, the species’ identity of the eggs did not explain this cryptic female choice, which instead was primarily controlled by conspecific ovarian fluid, a semiviscous, protein‐rich solution that bathes the eggs and is released at spawning. Video analyses revealed that ovarian fluid doubled sperm motile life span and straightened swimming trajectory, behaviors allowing chemoattraction up a concentration gradient. To confirm chemoattraction, cell migration tests through membranes containing pores that approximated to the egg micropyle showed that conspecific ovarian fluid attracted many more spermatozoa through the membrane, compared with heterospecific fluid or water. These combined findings together identify how cryptic female choice can evolve at the gamete level and promote reproductive isolation, mediated by a specific chemoattractive influence of ovarian fluid on sperm swimming behavior.     

Analysis of gamete membrane dynamics during double fertilization of Arabidopsis

Angiosperms have a unique sexual reproduction system called “double fertilization.” One sperm cell fertilizes the egg and another sperm cell fertilizes the central cell. To date, plant gamete membrane dynamics during fertilization has been poorly understood. To analyze this unrevealed gamete subcellular behavior, live cell imaging analyses of Arabidopsis double fertilization were performed. We produced female gamete membrane marker lines in which fluorescent proteins conjugated with PIP2a finely visualized egg cell and central cell surfaces. Using those lines together with a sperm cell membrane marker line expressing GCS1-GFP, the double fertilization process was observed. As a result, after gamete fusion, putative sperm plasma membrane GFP signals were occasionally detected on the egg cell surface adjacent to the central cell. In addition, time-lapse imaging revealed that GCS1-GFP signals entered both the egg cell and the central cell in parallel with the sperm cell movement toward the female gametes during double fertilization. These findings suggested that the gamete fusion process based on membrane dynamics was composed of (1) plasma membrane fusion on male and female gamete surfaces, (2) entry of sperm internal membrane components into the female gametes, and (3) plasmogamy.     

Sperm should evolve to make female meiosis fair

Genomic conflicts arise when an allele gains an evolutionary advantage at a cost to organismal fitness. Oögenesis is inherently susceptible to such conflicts because alleles compete for inclusion into the egg. Alleles that distort meiosis in their favor (i.e., meiotic drivers) often decrease organismal fitness, and therefore indirectly favor the evolution of mechanisms to suppress meiotic drive. In this light, many facets of oögenesis and gametogenesis have been interpreted as mechanisms of protection against genomic outlaws. That females of many animal species do not complete meiosis until after fertilization, appears to run counter to this interpretation, because this delay provides an opportunity for sperm‐acting alleles to meddle with the outcome of female meiosis and help like alleles drive in heterozygous females. Contrary to this perceived danger, the population genetic theory presented herein suggests that, in fact, sperm nearly always evolve to increase the fairness of female meiosis in the face of genomic conflicts. These results are consistent with the apparent sperm dependence of the best characterized female meiotic driversin animals. Rather than providing an opportunity for sperm collaboration in female meiotic drive, the “fertilization requirement” indirectly protects females from meiotic drivers by providing sperm an opportunity to suppress drive.     

Recent progress on sperm characterization in flowering plants

The organization, isolation and physiology of the angiosperm male gamete has recently emerged as an area of special interest. Detailed in vivo studies have revealed: (1) that the functional unit of male reproduction is a'male germ unit,'composed of two sperm cells and a physically associated vegetative nucleus; (2) that the two sperm cells are often cytoplasmically dimorphic, and (3) that some sperm cells appear to undergo preferential fertilization, preferentially fusing with either the egg or the proendospermaric central cell. Male gamete isolation has provided enrichments of up to 10⁷ cells ml^–1, with sperm integrity and viability demonstrated by the flunrochromatic reaction and ATP levels. Although results from biochemical and hybridoma antibody characterization are still preliminary, they indicate that sperm cells possess polypeptide expression patterns that are different from those of surrounding cells. These findings suggest the existence of an independent developmental program in the angiosperm male gamete.     

SPERM‐LIMITED FECUNDITY IN NEMATODES: HOW MANY SPERM ARE ENOUGH?

The Bateman principle, which holds that oocytes are the limiting gamete in reproduction, is violated in a variety of species. Self-fertilizing hermaphrodites of the nematode Caenorhabditis elegans provide an example of a system in which sperm number limits lifetime reproductive output, in this species due to the protandrous nature of sperm production that in turn delays the onset of fertilization. This reproductive delay forms the basis of a trade-off between generation time and total fecundity, in which sperm number plays a pivotal role. I use an age-structured population model to describe the number of sperm that maximize fitness, given larval development time and rates of gamete production. The model predicts the evolution of sperm numbers that are consistent with empirical data for C. elegans provided that precocious larval sperm production is taken into account. Several testable hypotheses follow from the model regarding how natural selection and environmental variation may influence patterns of sperm production among populations or species with a similar mode of reproduction.     

Multiple mating,sperm transfer and oviposition pattern in the giant sperm species,Drosophila bifurca

Many diverse traits are involved in gamete systems, and several models have analysed sperm length variation in terms of the intensity of sperm competition. This study investigates mating, sperm transfer and oviposition patterns in Drosophila bifurca, which possesses the longest sperm in the animal kingdom (about 6 cm). The prediction is that sperm gigantism should prevent male–male interaction. In this study, we examine how sperm transfer varies as males mate with a series of females, and how female receptivity changes with time after mating. As predicted, we found an extremely limited overlap of ejaculates owing to (1) reduced sperm transfer to females that had already mated, and (2) female remating depended both on the amount of sperm transferred and the modes of egg laying. The amount of sperm transferred to the female is discussed in relation to the peculiar morphology of the male reproductive tract and to sexual dimorphism and ecological hypotheses.