D. P. Froman

Sperm mobility determines the outcome of sperm competition in the domestic fowl

The aim of this study was to establish whether the mobility of sperm of the domestic fowl, as measured by an in vitro assay, predicted the outcome of sperm competition. Thirteen pairs of New Hampshire roosters, comprising one male categorized as having high–mobility sperm and the other as having average–mobility sperm, were used. Each male provided 25 times 106 sperm, which were mixed and artificially inseminated into between four and seven New Hampshire hens, each of which produced 2 to 11 offspring. The experiment was conducted twice, such that the same pair of males inseminated the same females. Paternity was assigned by using microsatellite markers. There was a clear effect of sperm–mobility phenotype on the outcome of sperm competition: in all 13 pairs the high–mobility male fathered the majority of offspring (73.3% overall; p < 0.0001). The proportion of offspring fathered by the high–mobility male within pairs varied significantly between male pairs (p < 0.0005). This effect was associated with the difference in sperm–mobility scores between males within pairs: there was a significant positive relationship between the proportion of offspring fathered by the high–mobility male and the ratio of mobility scores between males (p < 0.05). In addition, compared with their success predicted from the non–competitive situation, in the competitive situation high–mobility males were disproportionately successful in fertilizing eggs compared with average–mobility males. This may occur because female sperm storage is limited in some way and a greater proportion of high–mobility sperm gain access to the females sperm storage tubules. There was no evidence that female effects accounted for any of the variation in paternity.

Sperm mobility: mechanisms of fertilizing efficiency, genetic variation and phenotypic relationship with male status in the domestic fowl, Gallus gallus domesticus

When females are sexually promiscuous, sexual selection continues after insemination through sperm competition and cryptic female choice, and male traits conveying an advantage in competitive fertilization are selected for. Although individual male and ejaculate traits are known to influence paternity in a competitive scenario, multiple mechanisms co‐occur and interact to determine paternity. The way in which different traits interact with each other and the mechanisms through which their heritability is maintained despite selection remain unresolved. In the promiscuous fowl, paternity is determined by the number of sperm inseminated into a female, which is mediated by male social dominance, and by the quality of the sperm inseminated, measured as sperm mobility. Here we show that: (i) the number of sperm inseminated determines how many sperm reach the female sperm‐storage sites, and that sperm mobility mediates the fertilizing efficiency of inseminated sperm, mainly by determining the rate at which sperm are released from the female storage sites, (ii) like social status, sperm mobility is heritable, and (iii) subdominant males are significantly more likely to have higher sperm mobility than dominant males. This study indicates that although the functions of social status and sperm mobility are highly interdependent, the lack of phenotypic integration of these traits may maintain the variability of male fitness and heritability of fertilizing efficiency.

Social competitiveness associated with rapid fluctuations in sperm quality in male fowl.

When females copulate with multiple males, paternity is determined by the competitive ability of a male to access females and by the ability of its ejaculates to out-compete those of other males over fertilization. The relationship between the social competitiveness of a male and the fertilizing quality of its sperm has therefore crucial implications for the evolution of male reproductive strategies in response to sexual selection. Here, we present a longitudinal experimental study of the relationship between social status and sperm quality. We monitored sperm quality in socially naive male domestic fowl, Gallus gallus domesticus, before and after exposure to a social challenge which comprised two stages. In the first stage, social dominance was established in male pairs divergent in sperm quality, and in the second, social status was experimentally manipulated by re-shuffling males across pairs. We show that sperm quality fluctuates within males both before and after a social challenge. Importantly, such fluctuations followed consistently different patterns in males that displayed different levels of social competitiveness in the social challenge. In particular, following the social challenge, sperm quality dropped in males that won both contests while the sperm quality of males that lost both contests remained constant. Together, these results indicate that males of different social competitiveness are predisposed to specific patterns of fluctuations in sperm quality. These rapid within-male fluctuations may help explain the recent findings of trade-offs between male social and gametic competitive abilities and may help maintain phenotypic variability in these traits.

Deduction of a Model for Sperm Storage in the Oviduct of the Domestic Fowl (Gallus domesticus)

Abstract The mechanism of sperm storage in the fowl oviduct has remained a mystery since the 1960s, when sperm storage tubules (SST) were discovered between the shell gland and vagina. Previously, it was known that only motile sperm could ascend the vagina and enter these tubules. However, the means by which sperm resided therein was not clear. Research with computer-assisted sperm motion analysis has demonstrated that 1) seminal plasma glutamate acts as a motility agonist via N-methyl-d-aspartate receptors; 2) motility depends on extracellular Ca2+ and Na+; 3) straight-line velocity is a variable with a skewed distribution; 4) sperm cell trajectory is a function of straight-line velocity; and 5) specific inhibition of phospholipase A2 renders sperm immotile. An additional experiment demonstrated that Ca2+ acts as a second messenger and thereby modulates the content of long-chain acylcarnitine within sperm. Therefore, it is proposed that 1) the release of endogenous fatty acids fuels sperm as they ascend the vagina; (2) on entering the SST, motile sperm maintain position against a fluid current generated by SST epithelial cells; 3) resident sperm metabolize exogenous fatty acids released from lipid-laden epithelial cells; (4) motile sperm emerge from the SST when their velocity declines to a threshold at which retrograde movement begins; and 5) the skewed distribution of straight-line velocity accounts for the exponential pattern of sperm emergence from the SST. In summary, sperm residence within and emergence from the SST are phenomena most likely explicable in terms of sperm cell motility.

Sperm Mobility: Phenotype in Roosters (Gallus domesticus) Determined by Mitochondrial Function

Abstract Previously, inheritance of sperm mobility entailed a maternal additive genetic effect, and sperm ATP content was correlated (r = 0.80) with phenotype. The present study was conducted to determine if mitochondrial function was critical to phenotypic expression. Whereas phenotype was independent of mitochondrial helix length, phenotype was correlated with sperm oxygen consumption (r = 0.83) using random-bred roosters. Aberrant mitochondria characterized immobile sperm, as evidenced by transmission-electron microscopy. Such mitochondria were swollen and contained disorganized cristae. Additional experiments were performed with roosters from lines selected for low or high sperm mobility. A threefold difference in sperm oxygen consumption was observed between lines. Single nucleotide polymorphisms were observed in mitochondrial DNA by sequencing replicate mitochondrial genomes from each line. An A-to-G substitution in the gene encoding tRNAArg was inherited consistently, as evidenced by restriction fragment length polymorphism analysis using two male and two female progeny per family group and 14 family groups per line. Motile concentration in semen from low-line males was half that observed in semen from high-line males, as evidenced by computer-assisted sperm motion analysis. Likewise, 47% of sperm from low-line males contained aberrant mitochondria, compared to 4% for high-line males. In summary, sperm mobility phenotype was dependent on mitochondrial function, which in turn was altered by genetic selection. Fowl deferent duct fluid contains a high concentration of glutamate. We propose that variation in sperm mobility phenotype stems from the extent to which glutamate induces excessive mitochondrial Ca2+ uptake before ejaculation.

Sperm competition dynamics: ejaculate fertilising efficiency changes differentially with time

BackgroundA fundamental challenge in evolutionary biology is to resolve the mechanisms that maintain paternity a hypervariable fitness component. Because females are often sexually promiscuous, this challenge hinges on establishing the mechanisms through which the ejaculates of different males compete for fertilisation (sperm competition). The competitive quality of an ejaculate is mediated by the relative number of live sperm and their motile performance. The differential rate at which rival ejaculates lose their fertilising efficiency over time is therefore expected to influence the outcome of sperm competition.ResultsHere, we artificially inseminated into sets of replicate domestic hens, Gallus gallus domesticus, experimentally engineered heterospermic ejaculates containing a large number of low-quality sperm from one male, and a lower number of high-quality sperm from another male. Large, low-quality ejaculates fertilised the first eggs produced after insemination, but small, high-quality ejaculates prevailed in the long run despite their numerical disadvantage.ConclusionTogether, these results provide the first experimental demonstration that the relative competitive value of an ejaculate changes drastically over the time during which competing ejaculates are stored within the reproductive tract of a female, resulting in a marked temporal pattern of variation in paternity. A high level of replication makes these results robust. However, our study was restricted to few males of a well characterised study population, and future work should explore the generality of these results.

Sperm Mobility: Phenotype in Roosters (Gallus domesticus) Determinedby Concentration of Motile Sperm and Straight Line Velocity

Abstract Previous research demonstrated that sperm mobility, i.e., the net movement of a sperm population, is a quantitative trait of the domestic fowl. However, the cellular basis for this trait was unknown. In the present work, individual motile sperm were evaluated with a Hobson SpermTracker in order to identify one or more properties of motile sperm that could account for variation in sperm mobility observed among males. A method was validated for assessing sperm motion over an erythrocyte monolayer at body temperature. A small-scale experiment with roosters from the tails and center of a normal distribution of sperm mobility phenotypes (n = 33 roosters) demonstrated that straight line velocity (VSL) and motile concentration were critical to expression of phenotype. The importance of these variables was confirmed with a large-scale experiment using a representative subpopulation (n = 100 roosters). VSL of individual sperm at 41°C ranged between 5 and 100 μm/sec. VSL averaged 32, 39, and 40 μm/sec for low, average, and high sperm mobility phenotypes. Sperm were diluted to 1.2 × 106/ml for motion analysis. Mean motile concentrations were 0.52, 0.84, and 0.95 × 106/ml for low, average, and high sperm mobility phenotypes. Motile concentration was correlated with sperm mobility (r = 0.71). VSL appeared to have an additive effect as it was correlated with straightness of sperm cell trajectory (r = 0.79).

Chromium Picolinate Reduces Laying Hen Serum and Egg Yolk Cholesterol

Abstract One hundred 55-wk-old Single Comb White Leghorn hens were divided randomly into four experimental groups. Hens were caged individually. Each treatment group was fed one of four experimental diets for 35 consecutive d. The control diet was a standard layer ration. The other diets were supplemented with chromium picolinate at 200, 400, or 800 μg/kg. The objective of the experiment was to determine the effect of chromium picolinate on egg production, egg quality, and serum traits. Dietary supplementation with chromium picolinate had no effect on egg production and a negligible effect on egg weight. In contrast, chromium picolinate lowered (P

Sperm Mobility: Deduction of a Model Explaining Phenotypic Variation in Roosters (Gallus domesticus)

Abstract In previous work, variation in sperm mobility phenotype was attributed to the proportion of ejaculated fowl sperm containing dysfunctional mitochondria. In the present work, latent mitochondrial dysfunction was inferred from patterns of sperm egress from the oviducts sperm-storage tubules. In addition, experiments were performed to help explain how mitochondrial function could be compromised in viable sperm cells. Confocal microscopy demonstrated that sperm Ca2+ content differed between low and high sperm-mobility phenotypes when sperm were stained with rhod-2 AM, a Ca2+-specific dye. Fluorescence was associated with the nuclear envelope, a variant of the endoplasmic reticulum, and greater fluorescence was observed in sperm from low sperm-mobility males. Fluorescence was reduced by 50% when motile sperm were rendered immotile by incubation with a Ca2+ chelator. Thus, a relationship was established between a dynamic intracellular Ca2+ pool and sperm motility. Sperm N-methy-d-aspartic acid (NMDA) receptors were inferred by the action of d-homocysteinesulfinic acid, a potent NMDA receptor agonist. Seminal plasma from low sperm mobility males was characterized by an elevated glutamate concentration. Thapsigargin, which inhibits the smooth endoplasmic reticulum Ca2+ pump and thereby promotes Ca2+ efflux, rendered sperm immotile. This effect was blocked by cyclosporin A, which prevents the formation of the mitochondrial permeability transition pore (PTP) in response to elevated mitochondrial Ca2+ content. In summary, we propose that 1) glutamate enables Ca2+ uptake into sperm before ejaculation, 2) excessive Ca2+ uptake triggers formation of the PTP in a subpopulation of sperm, and 3) sperm mobility is decreased in proportion.

PHYSIOLOGY AND ENDOCRINOLOGY SYMPOSIUM: A proteome-based model for sperm mobility phenotype

Sperm mobility is defined as sperm movement against resistance at body temperature. Although all mobile sperm are motile, not all motile sperm are mobile. Sperm mobility is a primary determinant of male fertility in the chicken. Previous work explained phenotypic variation at the level of the sperm cell and the mitochondrion. The present work was conducted to determine if phenotypic variation could be explained at the level of the proteome using semen donors from lines of chickens selected for low or high sperm mobility. We began by testing the hypothesis that premature mitochondrial failure, and hence sperm immobility, arose from Ca(2+) overloading. The hypothesis was rejected because staining with a cell permeant Ca(2+)-specific dye was not enhanced in the case of low mobility sperm. The likelihood that sperm require little energy before ejaculation and the realization that the mitochondrial permeability transition can be induced by oxidative stress arising from inadequate NADH led to the hypothesis that glycolytic enzymes might differ between lines. This possibility was confirmed by 2-dimensional electrophoresis for aldolase and phosphoglycerate kinase 1. This outcome warranted evaluation of the whole cell proteome by differential detergent fractionation and mass spectrometry. Bioinformatics evaluation of proteins with different expression levels confirmed the likelihood that ATP metabolism and glycolysis differ between lines. This experimental outcome corroborated differences observed between lines in previous work, which include mitochondrial ultrastructure, sperm cell oxygen consumption, and straight line velocity. Although glycolytic proteins were more abundant within highly mobile sperm, quantitative PCR of representative testis RNA, which included mRNA for phosphoglycerate kinase 1, found no difference between lines. In summary, we propose a proteome-based model for sperm mobility phenotype in which a genetic predisposition puts sperm cells at risk of premature mitochondrial failure as they pass through the excurrent ducts of the testis. In other words, we attribute mitochondrial failure to sperm cell and reproductive tract attributes that interact to affect sperm in a stochastic manner before ejaculation. In conclusion, our work provides a starting point for understanding chicken semen quality in terms of gene networks.