The objective of this study was to uncover molecular and cellular dynamics in spermatozoa from Holstein bulls with Good (11 bulls) and Poor (5 bulls) cryotolerance. Post-thaw sperm kinetics, membrane integrity, and DNA fragmentation were assessed. Data was analyzed using principal component analysis. The spermatozoa from Good bulls had a higher number of cells with intact membranes (P = 0.029), nonragmented DNA (P = 0.018), post-thaw viability (P < 0.001) in comparison to those of the Poor bulls. The Good bulls also had a faster average path velocity (VAP; P = 0.017), straight-line velocity (VSL; P = 0.036), a greater distance average path (DAP; P = 0.006) and distance straight line (DSL; P = 0.011). No differences were found in total antioxidant capacity (TAC), number of live cells or other kinetic parameters between spermatozoa from Good and Poor groups, showing that no one characteristic can be used to determine cryotolerance.

A series of recent studies using Drosophila melanogaster suggest that while males may benefit from having access to many partners, female fitness is reduced by extended cohabitation and sexual interaction with males. Yet, even if repeated sexual interactions are harmful to females, limited male exposure will ultimately be detrimental due to sperm-depletion and infertility. Females are therefore expected to balance mating opportunities and sperm storage capacity to maximize lifetime reproductive success. I introduced extended mating deprivation as a selective pressure to experimentally evolve lines of D. melanogaster for characters related to mating and postcopulatory sexual selection. Evolution of the mate-deprived lines over several dozen generations demonstrated that restricted sexual access was indeed a potent selective pressure. I consistently found that when males were removed for an extended time period, female fitness declined substantially, suggesting that mate-deprivation over nine days was harmful. Under these conditions, selected-line males responded to mate-reduced conditions and demonstrated a 13% increase in reproductive success compared to controls. Experimental females had a 15% increase in fertility compared to controls. I investigated a series of developmental characteristics that may have been altered by the selection regime, and while there was some evidence of evolved change, these results were not consistent. Although the data at hand do not substantiate a detailed characterization, both sexes in the experimental populations demonstrated increased fitness after extended mate-deprivation, thus evolutionary change appears to have occurred via selection on one or both relevant male ejaculate characteristics: sperm number and survival, and factors affecting female late-life fertility. / Thesis (Master, Biology) -- Queen's University, 2008-04-28 23:05:42.835

Magister Scientiae (Medical Bioscience) - MSc(MBS) / A functional sperm is critical for successful fertilization in order to deliver an intact genome to the site of fertilization. It is often characterized by high motility and normal morphology. Moreover, sperm hyperactivated motility is imperative for both detachment from the oviductal wall and for penetration into the zona pellucida, subsequently resulting in fertilization. Several semen parameters such as volume, colour, sperm morphology and sperm concentration are used to clinically discriminate between fertile and sub-fertile males. Additionally, several sperm functional tests assess sperm function and a male’s fertility potential. A sperm feature that is not currently assessed clinically, but could possibly discriminate between fertility and infertility, is hyperactivation. The aim of this project was to investigate motility degrees (good, medium and poor) of sperm subpopulations and induce hyperactivation in each subpopulation, as well as to sperm in semen, by addition of caffeine and procaine. This was achieved by separating three sperm subpopulations from a semen sample using the Puresperm density gradient separating technique. Sperm subpopulations were exposed to 5mM caffeine and 2 mM procaine respectively for 15, 30, 60, 90 and 120 minutes. Sperm in semen was exposed to caffeine and procaine using a flush technique and analysed at 0, 5, 15, 30, 45 and 60 minutes. Sperm displaying hyperactivation was determined using cut-offs for curvilinear velocity, linearity and amplitude of lateral head displacement. The results indicate significant differences in overall percentage motility, sperm kinematic parameters and hyperactivation among the three subpopulations (p<0.05). Procaine and caffeine both induced hyperactivation in subpopulations, although the most pronounced effect of procaine was evident after 15-30 minutes compared to caffeine (60-90 minutes) in subpopulations. Maximum hyperactivation of sperm in semen was seen after 15- 30 minutes in both procaine and caffeine. Moreover, caffeine had significantly higher stimulating effect than procaine. The results suggest that the existence and distinct motility characteristics of subpopulations should be considered in future during clinical assessment of male fertility, especially when assessing hyperactivation. The immediate and higher stimulation response of sperm with the flush technique indicates that the technique may be an ideal sperm functional test compared to the separation technique. The separation technique may be used to categorize sperm subpopulation of a patient in terms of motility (high motile or low motile) and to stimulate such subpopulations with chemicals for use in assisted reproduction technologies.

<p>Sperm competition, the contest between sperm from rival males for fertilizations, is an important evolutionary force shaping sperm characteristics. Theory predicts that males experiencing elevated levels of sperm competition will invest more in sperm number, size and speed. While broad support exists for the idea that elevations in sperm competition lead to increased investment in sperm production, there is mixed support for the role of sperm competition in shaping sperm size and swimming speed. In this thesis, using a combination of within-species and comparative studies, I describe how sperm competition has influenced sperm traits in fishes and critically test a number of predictions from sperm competition theory. In the marine plainfin midshipman fish (Porichthys notatus) and the fresh-water shell brooding cichlid Telmatochromis vittatus, I show that the males who experience the highest level of sperm competition had faster but not longer sperm. Instead, selection appears to have acted on sperm energetics, increasing energy production to drive sperm movement in males who experience more intense levels of sperm competition. In a comparative study using Tanganyikan cichlids, I show that males in species experiencing high levels of sperm competition (i.e. promiscuous species) had both longer and faster sperm than males of closely related species unlikely to experience sperm competition (i.e. monogamous species). I also uncovered a predicted but previously inadequately tested relationship between sperm size and speed. This relationship holds across, but not within, species and I discuss possible explanations for differences between and within species. Finally, I used directional tests of trait evolution to assess how selection acts to increase sperm swimming speed and provide evidence that the evolution of fast swimming sperm preceded the evolution of long sperm across cichlid fishes. Together, the results of this thesis show that spenn competition promotes the evolution of faster swimming spenn in fishes and highlights the importance of sperm energetics in detennining the competitive success of ejaculates.</p> / Thesis / Doctor of Philosophy (PhD)

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Chicken spermatozoa undergo a post-testicular maturation process similar to that reported for mammals. Association of secreted epididymal proteins and glycoproteins with the sperm surface appears to be complete at the anterior ductus deferens. However, antigenic change to, and glycosylation and deglycosylation of seminal plasma and sperm surface-associated proteins, selective loss of proteins from the sperm surface, as well as secretion of some seminal plasma proteins and selective absorption of others by epithelial cells lining the excurrent ducts, continues in the ductus deferens and ampulla. Hypertonic treatment or neuraminidase treatment of spermatozoa without apparent loss of sperm integrity in-vitro, reduced the ability of spermatozoa to gain access to the uterovaginal insemination. This indicates a clear role for sperm surface-associated proteains and sperm surfarce sialic acid in vaginal sperm transport. Sperm surface-associated proteins were also extracted by glycerol in a temperature-dependent and concentration-dependent manner. This was accompanied at room temperature by a glycerol concentration-dependent reduction in sperm motility. These effects may be implicated in the contraceptive action of glycerol in the chicken vagina. Spermatozoa from several avian and mammalian species entered quail uterovaginal junction SSTs in-vitro, and turkey spermatozoa were found in chicken uterovaginal junction SSTs following uterovaginal junction but not intravaginal insemination, thus showing that the SSTs are not selective, and identifying the vagina as a major site of oviducal sperm selection. Spermatozoa washed from the vagina following intravaginal insemination had immunoglobulin bound to their surface, which was shown to be associated with cell death. Spermatozoa recovered from the anterior oviduct however, were generally devoid of bound immunoglobulin. Furthermore, sperm access to the newly ovulated egg in-vivo following incubation with vaginal mucosa in-vitro did not differ significantly from that of control spermatozoa despite fewer viable spermatozoa inseminated, suggesting a true 'selection' of spermatozoa in the vagina. Ovarian pocket fluid, postulated to be the milieu in which fertilisation takes place in-vivo, altered sperm surface antigenicity and reduced sperm motility in-vitro, suggesting that spermatozoa may naturally undergo oviduct-induced changes prior to fertilisation. Sperm-egg interaction appears specific and receptor-mediated, as in mammals. Spermatozoa showed no preference for the animal pole of the egg, and heterologous gamete combinations indicated limited, order-dependent rather than species-dependent specificity. Presenting carbohydrate residues on the sperm surface and sperm surface antigenicity showed a lack of change proportional to species divergence, although the vagina clearly presents the main barrier to interspecies fertilisation within the order Galliformes.