In this project we are building a mathematical model to track the movement of spermatozoa during the process of chemotaxis. Our model is built on an off-lattice spherical biased random walk in 3-dimensional space, an extension of previous conventional deterministic 2-dimensional models. The sperm's type of movement is decided based on a comparison of the current and previous chemoattractant concentration which can be used to see whether it is approaching the egg. From the statistical analysis of the simulation results, we find that chemotaxis is an effective mechanism to increase the number of sperm reaching the egg.

A Hodgkin-Huxley style system of ODEs was developed to model the ion channel activity of A. punctulata sperm flagellum during exposure to resact during chemotaxis. Empirical data was used in conjunction with parameter estimation methods in an attempt for the model to reproduce realistic Voltage potentials and ion concentrations. The change in calcium concentration is of particular interest, as it is essential in the waveform of the flagellum during chemotaxis.

While the central role of spermatozoa in sexual reproduction and fertilization is well understood, many functional attributes of sperm have yet to be elucidated at the molecular level. One key to ultimately understand the molecular basis of sperm function is to comprehensively characterize its biochemical composition. This crucial information has been lacking, as molecular characterization of the sperm cell cannot be assessed by classic gene expression assays since mature spermatozoa are transcriptionaly inert. Whole-cell shotgun proteomic approaches have revolutionized the molecular analysis of sperm form and function. We have utilized improved methodologies to re-analyze the D. melanogaster sperm proteome and characterize five additional Drosophila species sperm proteomes. This methodology, which included a 1D SDS-PAGE prefractionation step, resulted in good reproducibility between biological replicates and high quality sperm proteomes. An interspecific analysis of the sperm proteomes revealed that despite variation in protein composition, Drosophila sperm proteomes have a consistent functional profile and 519 proteins were identified a being conserved across the melanogaster subgroup within a phylogenetic framework. Evolution of the sperm proteome was explored in Mus musculus through the utilization of targeted proteomic datasets completed, which provided subcellular localizations for sperm components. This study resulted in several novel findings, including evidence for accelerated evolution as well as an enrichment of positive selection on genes found in the cell membrane and acrosome. This may be a result of the selective pressures encountered by these membrane proteins during sperm development, maturation and transit through the female reproductive tract where the sperm cell membrane, and eventually the acrosome, are exposed to the extracellular milieu and are available for direct cell-cell interactions. These findings not only reveal the varying evolutionary pressures acting on a single cell type but also highlights the utility of the proteomics technique in clarifying protein interaction and evolutionary history.

Forensic analysis of deoxyribonucleic acid (DNA) collected from sexual assault evidence is a multi-step process that requires a great amount of time and resources. A large percentage of samples are mixtures containing DNA from a major female contributor and at least one minor male contributor. The amount of male DNA present is often much less than that of the female, making it difficult to achieve a full short-tandem repeat (STR) profile for identification purposes. The current method employed by many forensic laboratories to separate sperm DNA from non-sperm DNA is the differential extraction. Although a robust and reliable method when applied to liquid samples, the procedure has failed to evolve significantly since first developed.1,2 Between the time it has been collected and tested, sexual assault evidence becomes dried and aged, contributing to the potential loss and degradation of already low amounts of DNA and increasing the likelihood of an incomplete profile.2 This study tests the effectiveness of a combination of enzymes to release DNA from sperm using a variety of substrates. Although this method extracted greater amounts of male DNA than the traditional Qiagen® extraction, further research is necessary to determine if the application of this new method can improve or eventually replace the current procedures. / 2018-06-16T00:00:00Z

by So Siu Cheung, Eddie. / "September 2000." / Thesis (Ph.D.)--Chinese University of Hong Kong, 2000. / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Mode of access: World Wide Web. / Abstracts in English and Chinese.

Orientador: João Carlos Pinheiro Ferreira / Banca: José Roberto Sartori / Banca: Nadja Susana Mogyca Leandro / Resumo: No presente estudo foram correlacionados os resultados das contagem de perfurações espermáticas na membrana perivitelínica externa de ovos incubáveis com as taxas de eclosão e fertilidade, objetivando-se analisar a possibilidade do uso dessa técnica como alternativa aos métodos tradicionais de avaliação da fertilidade, executados no incubatório. As avaliações foram realizadas a cada três semanas durante todo o período reprodutivo de um lote comercial de matrizes de corte da linhagem Avian Cobb 48, submetidas à monta natural a partir da 25a semana de idade. Foram realizadas 10 avaliações, sendo utilizados 30 ovos em cada avaliação. Foi realizada a extração, fixação e coloração da membrana perivitelínica externa para possibilitar a visualização das perfurações ao microscópio óptico. A idade da ave influenciou negativamente o número de perfurações espermáticas na membrana perivitelínica externa, bem como a taxa de fertilidade e de eclosão. Houve correlação positiva entre o número de perfurações espermáticas na membrana perivitelínica e a taxa de fertilidade (r= 0,885; P<0,05), assim como com a taxa de eclosão (r= 0,800; P<0,05). É possível inferir que a contagem de perfurações espermáticas pode ser utilizada como ferramenta de avaliação da fertilidade de ovos incubáveis / Abstract: In this study we correlated the results of counting sperm penetration in the outer perivitelline layer of hatching eggs to the fertility and hatching rates, aiming to examine the possibility of using this technique as an alternative to traditional methods of fertility evaluation, performed in the hatchery. The evaluations were performed every three weeks throughout the reproductive period of a flock of commercial broiler strain Cobb Avian 48, submitted to natural mating from the 25th week of age. Were performed 10 evaluations, 30 eggs were used in each assessment. Was extracted, fixed and stained outer perivitelline layer to enable viewing of holes in an optical microscope. The bird's age negatively influenced the number of sperm holes in the outer perivitelline layer and the fertility rate and hatching. A positive correlation between the number of sperm holes in the perivitelline layer and the fertility rate (r = 0.885, P <0.05), as well as the hatching rate (r = 0.800, P <0.05). It can infer that the sperm count of holes can be used as a tool for assessing the fertility of eggs hatching / Mestre

Post-copulatory sexual selection is a strong evolutionary force, affecting morphological and behavioural traits in males and females in species with polyandrous mating systems. Many insects are subject to sperm competition; sperm from rival males compete to fertilise ova. Since sperm are finite, males should allocate them economically, tailoring ejaculate allocation to suit the reproductive potential of individual matings. Theory suggests when sperm competition risk is high, males should increase sperm numbers to achieve greater reproductive success than their rivals, but evidence of this expected fitness consequence of ejaculate allocation is largely lacking. In this thesis, I use Callosobruchus maculatus beetles to investigate the causes of ejaculate allocation patterns, and to examine whether ejaculate allocation does affect male reproductive success. In Chapter 3, I investigate the effect of rival male presence on ejaculate size and find that, while males grouped with rivals as adults produce bigger ejaculates, their increased effort unexpectedly does not lead to increased reproductive success. In Chapter 4, I examine whether larval conditions also affect ejaculate size, and find that, contrary to sperm competition theory, males reared under dense conditions produce smaller ejaculates than those reared solitarily, and that male reproductive success is consequently elevated in males reared at low larval densities compared to those reared at high densities. In Chapter 5, I then demonstrate that ejaculates produced by low density males contain more sperm than ejaculates produced by high density males, suggesting males do not respond to sperm competition level represented by larval density, but instead suffer resource limitation when reared at high density. In Chapter 6, I investigate the effects of water provision on ejaculate size, and find that males given water produce larger ejaculates, and females given water receive smaller ejaculates. Finally, I link my findings with those of other studies, and suggest my most important result is that plasticity of ejaculate allocation cannot be assumed to be an adaptive behaviour; studies directly measuring the fitness effects of male ejaculate allocation are needed, even when observed patterns conform to theory.

Philosophiae Doctor - PhD / In the marine environment, penguins have been described as curators and serve a critical role in ecological balance. The African penguin (Spheniscus demersus) has undergone a rapid population decline, mainly due to disturbances in their natural habitat. The African penguin was up-listed from vulnerable to endangered on the IUCN Red List for Threatened Species in 2010 and thus urgent conservation action is required. Integral to long-term conservation action of any species is a basic knowledge of its reproductive biology, which is currently lacking for African penguins. The main aim of this investigation was to evaluate techniques for the collection of semen in African penguin and to determine sperm quality in order to cryopreserve sperm for in vitro fertilization (IVF) purposes of captive and wild populations. Semen was collected once a week during two breeding seasons from two captive African penguins. Ejaculates (n=51) were obtained over two breeding seasons (Jan-Feb and Jun-Oct) and evaluated for semen volume, sperm concentration, sperm vitality, sperm motility and sperm morphology. In addition twelve (six females and six males, n=4 were breeding pairs) captive African penguins were monitored for hormone (estradiol, testosterone, progesterone) levels prior to and after the egg-laying period.

PhD / The aim of this thesis was to investigate the possibility of integrating frozen-thawed boar semen into reproductive technologies and into commercial production of pigs in Australia. This was to be achieved by establishing a semen freezing and AI regime that was of a standard acceptable to industry, and integrating the resultant frozen-thawed sperm into other reproductive technologies, such as flow cytometric sperm sorting and IVF. Initially, a protocol for freezing and thawing boar semen was established, based on the method described by Westendorf et al. (1975) and attempts were made to modify this protocol to improve the post-thaw sperm quality, as determined by in vitro assessment of motility, acrosome integrity and longevity. First, the egg yolk used in the freezing extenders was investigated, and the chicken yolk was replaced with either duck or quail yolk. It was shown that there was no benefit in substituting yolk from duck or quail for the chicken yolk traditionally used in freezing extender. Second, the effect of seminal plasma addition to the freezing extender, or seminal plasma addition to resuspension medium post-thaw was tested. Incorporating whole seminal plasma into the freezing extender at levels above 50% was found to be detrimental to post-thaw sperm quality. Reducing levels to 20% of the final volume improved acrosome integrity, but adversely affected motility of sperm. However, adding 20% seminal plasma to the resuspension medium used after thawing of boar semen had no significant influence on sperm quality compared with resuspension in medium without seminal plasma. The antioxidant catalase, and the iron chelator desferal added to the freezing extender, did not improve post-thaw sperm quality, nor was any benefit seen with addition of these substrates to the resuspension medium post-thaw. However, the bioactive phospholipid PAF and its regulating enzyme PAF:AH appeared to enhance post-thaw motility and acrosome integrity of sperm, respectively, when added to the semen pre-freezing. Unfortunately, due to the restrictions imposed on rPAF:AH as a research drug, it was not possible to test the in vivo effects at this time. After the in vitro experiments were completed, the in vivo fertility of frozen-thawed sperm was tested using the optimal freezing protocol and a novel technology, enabling non-surgical deep intrauterine insemination of sows. The aim was to establish the lowest possible dose of frozen-thawed sperm that could be used, without compromising fertility. Successful pregnancies were achieved with doses as low as 62.5 x 106 frozen-thawed sperm but the farrowing rates were too low to be practicable on a commercial scale. This is the first report of litters born after insemination of such a low dose of frozen-thawed sperm and using the novel DIU insemination technique. However, it was concluded that a double dose of 250 x 106 frozen-thawed sperm was the minimum dose required for maintaining acceptable fertility. Reduction in sperm numbers to such an extent made it possible to consider non-surgical insemination of sex-sorted, frozen-thawed semen. Previously, pregnancies had been achieved only after surgical insemination of sex-sorted boar sperm, or with DIU insemination of unfrozen sperm, immediately after sex-sorting. The low numbers of sex-sorted sperm available restricted the inseminate dose used here to 50 x106 motile sperm. A litter of 5 piglets was born after a low-dose, DIU insemination of sex-sorted, frozen-thawed sperm. This is the first report of piglets born after insemination with sex-sorted frozen-thawed sperm and non-surgical insemination. The low farrowing rate achieved in this experiment prompted the investigation of integrating sex-sorted, frozen-thawed boar sperm into IVF. Morulae were produced after IVF with sex-sorted, frozen-thawed sperm and successfully transferred using non-surgical techniques. This is the first report of pregnancy achieved with non-surgical transfer of embryos produced after IVF and IVC of IVM oocytes with sex-sorted, frozen-thawed boar sperm. Unfortunately, the pregnancy did not hold, and the embryos were lost prior to Day 32, but PCR of non-transferred embryos confirmed successful pre-selection of sex. Overall, this thesis demonstrated that it is still not economically feasible to incorporate frozen-thawed boar semen into the commercial production of pigs although it has considerable application in breeding programmes. However, the development of novel techniques enabling reduction in sperm dose, and for non-surgical transfer of embryos into recipient sows and incorporation of frozen-thawed semen into these technologies means that progress is being made with the integration of reproductive technologies and frozen-thawed semen into the pig industry.

Stomach contents of 19 mature sperm whales, 18 males and one female, that stranded on New Zealand beaches between the mid 1990s and 2004 were examined, identified and measured. Three of the stomachs were empty. All other samples consisted almost entirely of cephalopod beaks. A total of 23,223 cephalopod beaks (10,647 upper and 12,576 lower), representing at least 36 species in 17 families were found in the remaining 16 stomachs. Non-cephalopod remains in the stomachs of sperm whales stranded in New Zealand included limited quantities of fish, salps, crustacean exoskeletons, a copepod, some wood and sand.The present investigation represents the most comprehensive study of the diet of sperm whales in New Zealand since the early 1960s. The results show that oceanic squid of the families Histioteuthidae, Cranchiidae, Onychoteuthidae and Octopoteuthidae are the most common remains found in the stomachs of sperm whales stranded on New Zealand beaches, with the families Onychoteuthidae, Histioteuthidae, Octopoteuthidae and Architeuthidae being the most important by estimated weight in whale diet, and the families Cranchiidae, Pholidoteuthidae and Ancistrocheiridae secondarily so.The beaks of three cephalopod species thought to be restricted to Antarctic waters (Kondakovia longimana, Mesonychoteuthis hamiltoni and Psychroteuthis glacialis) were found in 12 of the stomachs, suggesting these whales had recently migrated into New Zealand from more southern feeding grounds. The amount of local cephalopod beaks in the stomachs suggests some of the stranded sperm whales did not feed much within New Zealand waters in the days prior to stranding.The beaks of Taningia danae, Octopoteuthis megaptera, Octopoteuthis sp. 'Giant' and Lepidoteuthis grimaldii are illustrated and described. Oblique and lateral illustrations of the lower beaks are given, as well as sections of the rostrum, jaw angle, shoulder and lateral wall, to show the major identifying features for each of the species.Squid are an important component of food chains in the Southern Ocean and they act as both high-level predators and prey for apex predators. Therefore, seasonal fluctuations in their abundance must have cascading effects on the diets of apex predators. With increasing global fishing effort, and with cephalopods representing over 4% of the global annual catch, there are competing interests between the ocean's top teuthophagous predators and the fishing industry.Uncertainty of the effects fisheries have on the marine ecosystem has stimulated numerous research studies in recent years. However, despite the economic and ecological importance of cephalopods, there are few ecological studies on them or their significance in the trophic systems of the deep-sea and their life cycles and distribution patterns are only now beginning to be understood. Additional dietary studies that investigate the cephalopod composition and size-class structure in the diet of predators are needed to assess their importance in deepsea food webs, and the potential impact that deep-sea fisheries might have on associated and dependant species, namely apex oceanic predators.The results of this study provide the first significant insight into the diet of the sperm whale, one of the most important apex predators in New Zealand waters.