Philosophiae Doctor - PhD / Semen analysis is the cornerstone for the investigation of male infertility. Semen quality can be influenced by geographical location, age, ejaculatory abstinence, and season. In 2010, the WHO published criteria for human semen characteristics that were markedly lower than those previously reported. Many reports have discussed the methodology used by the WHO to set the 2010 reference values. Some of the limitations of the WHO (2010) study included an undefined ejaculatory abstinence period, the limited representation of different age groups, and a limitation in geographical representation as the study did not include any data from Africa and Middle East. Therefore, the current cohort study was designed to provide retrospective data on semen quality (Africa and Middle East) and reproductive hormones (Middle East) in patients who underwent semen analysis and endocrine investigation at Andrology Laboratories in South Africa and Qatar.

Compared to natural selection, the use of artificial insemination (AI) and other reproductive technologies rapidly increase the rate of genetic change in any population. In order to achieve success with AI, the semen used to inseminate cows must be of the highest possible quality. When semen is frozen, generally only about 50% of the spermatozoa survive the cryopreservation process. Thus, any factors possibly affecting the survival of spermatozoa through the numerous freezing-thawing steps should be studied, in order to identify the optimal conditions for the survival of spermatozoa. The discovery of protective agents within egg yolk and glycerol was a major milestone in sperm cryopreservation. These agents protect bovine spermatozoa during cooling and freezing procedures and result in increased survival rates. Cryopreservation of spermatozoa has become the most common technique for the preservation of male fertility of genetically superior sires even after their death. Using cryopreserved sperm to artificially inseminate females has become standard practice in commercial dairy cattle herds and the application of this reproductive management tool is also expanding to beef herds worldwide. The use of glycerol as a cryoprotectant for bovine spermatozoa is credited as the reason for the success in bovine semen cryopreservation. The purpose of this research was to quantify the effects of different cooling periods, as well as different glycerol equilibration periods on the post-thaw motility percentages and recovery fractions of semen collected from Boran and Nguni bulls. The research was subdivided into two experiments. In each experiment different cooling and glycerol equilibration times were researched. The first experiment involved shorter cooling times (30, 60, 120 and 240 minutes) with each cooling time followed by several longer equilibration times (4, 5, 6, 7 and 8 h). In the second experiment the cooling and equilibration times from the first experiment were reversed. This resulted in longer cooling times (4, 5, 6, 7 and 8 h) with each cooling time having shorter glycerol equilibration times (30, 60, 120 and 240 minutes). An egg yolk-Tris two-step extender was used in both the experiments. The general trend for the glycerol equilibration periods studied in Experiment 1 was that the resulting overall average post-thaw motility percentage and average recovery fraction increased with longer periods. There was a breed difference when comparing the average post-thaw motility percentages after 4, 5, 6 and 8 h (p<0.05), while the average post-thaw motility percentages also tended to differ after 7 h of equilibration. The general trend observed for equilibration periods used in Experiment 2 was that the average post-thaw motility percentage increased as glycerol equilibration period increased up to 120 minutes, but after 240 minutes of glycerol equilibration, there was a slight decline. The differences in average post-thaw motility percentage after the respective glycerol equilibration periods were not statistically significant. The results of each experiment were used to create a matrix that can be used in practice. The matrix using results from Experiment 1 demonstrated that a cooling period glycerol equilibration period combination of 240 minutes and 7 h resulted in the highest (not significantly different from most other combinations) average post-thaw motility rates. The matrix formed from the results of Experiment 2 demonstrated that an 8 h cooling period combined with a 60 minute glycerol equilibration period yielded the highest (not significantly different from most other combinations), average post-thaw motility percentage. / Dissertation (MSc(Agric))--University of Pretoria, 2010. / Animal and Wildlife Sciences / unrestricted

Background: A growing body of evidence has found links between endocrine disruptor phthalates and male reproductive disorders, but the mechanisms underlying these relationships are poorly known. Seminal plasma metabolomes may mediate associations of phthalate exposure with impaired semen quality. Objective: To identify seminal plasma metabolomes associated with poor semen quality and evaluate their associations with urinary phthalate metabolites among 660 Chinese adult men. Method: The seminal plasma metabolic profiles were acquired using an untargeted approach based on liquid chromatography-high resolution mass spectrometry. We explored the differences in seminal plasma metabolites between participants with poor and good semen quality and evaluated cross-sectional associations between discriminatory metabolic biomarkers and urinary phthalate metabolites. Results: Differences between poor and good semen quality groups were observed in relation to 25 seminal plasma metabolites, mostly related to the metabolism of polyunsaturated fatty acids (PUFA) and acylcarnitine (all p < 0.05). After adjusting for various confounders and multiple tests, metabolites were all significantly associated with one or more individual sperm quality parameters (motility, concentration, total count, and morphology) (all p < 0.05). Among identified metabolic biomarkers, seminal plasma L-palmitoylcarnitine, linoelaidyl carnitine, and oleic acid were inversely associated with urinary mono-(2-ethylhexyl) phthalate (MEHP), and seminal plasma L-acetylcarnitine was inversely associated with the proportion of di-(2-ethylhexyl)-phthalate metabolites (DEHP) excreted as MEHP in urine (%MEHP) (all p < 0.05). Mediation analysis revealed that oleic acid and L-acetylcarnitine mediated significant proportions (6.7% and 17%, respectively) of the positive associations between urinary DEHP metabolites and the percentage of spermatozoa with an abnormal head. Conclusions: Elevated urinary phthalate metabolites may impact semen quality by causing metabolic disorders of seminal plasma PUFAs and acylcarnitine. These pathways warrant further investigation.

No description available.

Thesis (M.Sc. (Medical Sciences)) -- University of Limpopo, 2021 / Infertility, defined as the inability to conceive following one year of unprotected sexual intercourse, respectively affects 25% of couples globally. Oxidative stress (OS) has been greatly related to the idiopathic cause of infertility and Camellia sinensis contains antioxidants that may enhance reproductive functions. This study focussed on the effects of Camellia sinensis (green and black tea) on human sperm functions in both normal and abnormal samples. Semen samples (n= 59) collected from donors were liquefied, analysed, and classified as normal (n=40) and abnormal (n= 19) using the WHO criteria. Samples were washed and exposed to aqueous leaf extracts of green and black tea (0, 0.4, 4, 40, 405 μg/ml) for 1 hour. Human Tubular Fluid (HTF) served as the control. The respective sperm parameters were analysed (sperm motility, vitality, DNA fragmentation, mitochondrial membrane potential (MMP), capacitation and acrosome reaction (CTC) and reactive oxygen species (ROS). Green and black tea significantly increased vitality, and intact MMP, while it significantly reduced, CTC, and intracellular ROS as well as DNA fragmented spermatozoa in both normal and abnormal samples compared to the control (p<0.05). A significant increase in sperm CTC, ROS, with a decrease in sperm vitality, and intact MMP was observed in the abnormal compared to the normal samples (p<0.05). No significant change in motility was observed between normal and abnormal samples compared to their respective controls, in both green and black tea (p>0.05). Camellia sinensis improved human sperm function in vitro and may be attributed to its antioxidant activity. / National Research Foundation (NRF)

Thirty five million people are currently living with HIV-1 today with women accounting for half of infected individuals globally. Sexual transmission is the main route of HIV transmission with approximately 40% of HIV infections occurring when the mucosal lining of the female genital tract (FGT) is exposed to HIV in semen from an infected male partner. Seminal plasma (SP), the fluid portion of semen, is a complex fluid which plays an immunomodulatory role in the FGT for successful conception, largely due to its high concentrations of TGF-β1. Several factors in SP from HIV-uninfected men have been shown to either inhibit or enhance HIV infection in target cells, however it is not clear how SP from HIV infected men would modulate genital epithelial cells (GECs), the first cells that encounter HIV in the FGT. The overall goals of this thesis were to compare inflammatory and regulatory cytokine concentrations in SP from HIV-uninfected and infected men, and subsequently compare GEC cytokine responses following exposure to SP from HIV-uninfected and HIV-infected men. I also investigated how SP and TGF-β regulated cytokine production and barrier function in GECs in the presence of HIV. The results summarized in this thesis demonstrated that HIV infection leads to different cytokine profiles in SP, based on stage of HIV-1 infection. HIV-infected men in acute stage contained higher levels of proinflammatory cytokines in their SP compared to HIV-uninfected and chronically infected men (CI men) which subsequently lead to higher levels of proinflammatory cytokines from GECs compared to CI men. In the follow up to this study we found that active TGF-β, which was found in higher concentrations in SP from CI men and led to decreased inflammatory response from GECs, was compartmentalized between blood plasma and seminal plasma. Higher levels of active TGF-β in SP correlated with decreased semen viral load and the immune activation marker sCD14 leading us to believe that ART-naive CI men in our cohort were naturally controlling their immune activation status, as active TGF-β levels were lower in ART-treated men. Short-term exposure of GECs to SP from CI men or TGF-β at comparable concentrations to SP protected the GEC barrier against HIV by decreasing inflammatory cytokines and preventing tight junction breakage. However, long-term exposure to TGF-β in the presence of HIV further increased inflammation in GECs suggesting a biphasic role for TGF-β in the FGT. This body of work summarized in this thesis demonstrates for the first time how semen from HIV-infected men modulates FGT epithelial cell cytokine responses and barrier function, an important consideration in the design of local therapeutic strategies to protect the FGT against HIV infection. / Thesis / Doctor of Philosophy (Medical Science)

Background: Human life without sperm is not possible. Therefore, it is alarming that the fertilizing ability of human spermatozoa is continuously decreasing. The reasons for that are widely unknown, but there is hope that metabolomics-based investigations may be able to contribute to overcoming this problem. This review summarizes the attempts made so far. Methods: We will discuss liquid chromatography–mass spectrometry (LC-MS), gas chromatography (GC), infrared (IR) and Raman as well as nuclear magnetic resonance (NMR) spectroscopy. Almost all available studies apply one of these methods. Results: Depending on the methodology used, different compounds can be detected, which is (in combination with sophisticated methods of bioinformatics) helpful to estimate the state of the sperm. Often, but not in all cases, there is a correlation with clinical parameters such as the sperm mobility. Conclusions: LC-MS detects the highest number of metabolites and can be considered as the method of choice. Unfortunately, the reproducibility of some studies is poor, and, thus, further improvements of the study designs are needed to overcome this problem. Additionally, a stronger focus on the biochemical consequences of the altered metabolite concentrations is also required

Exploring the genetics of the efficiency of fertile AI dose production in rabbits The general aim of this thesis has been to analyse sources of variation for some of the most important components of fertile artificial insemination (AI) dose production in order to explore the interest and limitations of different strategies for their genetic improvement in a paternal line of rabbits selected for growth rate. These components refer to seminal production and quality traits, being considered the male reproductive performance (fertility and prolificacy) as the final expression of the effect of the seminal characteristics and the effect of the interaction among them and with the female. Genetic analyses of the seminal traits involved in AI dose production and growth rate were modelled using threshold and linear multiple-trait mixed models. Relationship between fertility and pH of the semen was analysed either using mixed or recursive mixed models. Male and female genetic contributions to fertility were estimated using additive or product threshold models and both models were compared by its ability of predicting fertility data. Existence of genotype x artificial insemination conditions for male effect on fertility and prolificacy was estimated under a Character state model. Finally, the product threshold model was used for estimating separately the effect of the environmental temperature on male and on female contributions to fertility. All inferences of this thesis have been done under a Bayesian approach. Male libido and variables related to the quality of the ejaculate such as presence of urine and calcium carbonates in the ejaculate, individual sperm motility, semen pH and suitability for AI of the ejaculate (which involves the subjective combination of several semen quality traits) were found to be lowly heritable, but repeatable. / Tusell Palomero, L. (2011). Exploring the genetics of the efficiency of fertile AI dose production in rabbits [Tesis doctoral]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/11842

Four experiments were conducted to evaluate the viability and fertility of bovine spermatozoa encapsulated in microcapsules and microgels. In Experiment I, one of two morphologically distinct sperm types i.e. marker or unmarked bull spermatozoa (100 x 10⁶ sperm/bull) were encapsulated in protamine sulfate microcapsules and simultaneously inseminated with the reciprocal sperm type unencapsulated. Insemination of both sperm types unencapsulated served as a control. Accessory sperm embedded in the zona pellucida were counted and morphologically classified 6 to 7 d post insemination. From microencapsulated inseminates, accessory sperm populations did not increase over the unencapsulated controls, but contributed 25.7% of the accessory sperm population. ln Experiment 2, an in vitro study was performed to evaluate the maintenance of viability for bovine spermatozoa encapsulated in PIPES, HEPES, or saline microgels. Neat semen was pooled from five bulls (50 x 10° sperm/bull), encapsulated in alginate microgels, and incubated at 37 C for 8 h. The unencapsulated control displayed greater maintenance of viability for percent intact acrosomes and motility when compared to all treatments. By 8 h incubation, PlPES and HEPES were not significantly different, but demonstrated greater maintenance of viability when compared to saline microgel treatments. In Experiment 3, PIPES microgels were heterospermically inseminated with equal numbers (20 x l0⁶ sperm/bull) of frozen-thawed marker bull and normal bull spermatozoa as explained in Experiment 1. Microencapsulated treatments contributed significantly lower numbers of accessory sperm when compared to unencapsulated controls. In Experiment 4, one of the two morphologically distinct sperm types (20 x l0⁶ frozen-thawed sperm/bull) were encapsulated in protamine sulfate microcapsules and the reciprocal sperm type was encapsulated in PIPES microgels. A total of 21 accessory sperm were recovered from 30 embryos which demonstrates the ability of microencapsulated spermatozoa to fertilize an oocyte. / Master of Science

Introduction International guidelines endorse the screening of patients for human immunodeficiency virus (HIV), hepatitis B virus (HBV), hepatitis C virus (HCV) and Chlamydia trachomatis before assisted reproductive techniques (ART). At present no such guidelines exists in South Africa. At the Reproductive and Endocrine Unit (referred to as “the Unit”) of Steve Biko Academic Hospital, all patients with unknown HIV status are counselled and a blood sample is collected during the initial visit for automated laboratory based HIV screening. These HIV results are not available before semen samples are processed. Furthermore, patients are not screened for HBV, HCV and Chlamydia trachomatis. Couples attending the Unit are of a low to middle socio-economic status and experience financial constraints. Moreover, automated laboratory based assays are expensive to perform. Rapid testing is a cost effective and practical method from screening patients, with a 20–30 minute result turnover time. Until screening at the Unit is improved, the possible identification of semen characteristics that could indicate HIV infection would be a useful tool. Materials and Methods The following rapid point-of-care assays were evaluated: Determine® HIV-1/2 combo test (n=100), Determine® HBsAg test (n=100), DIAQUICK HCV kit (n=74), and the DIAQUICK Chlamydia trachomatis kit (n=30). For profiling, parameters from a basic semen analysis of HIV-positive males (n=60) were compared with HIV-negative males (n=60). Information pertaining to CD4 count, antiretroviral treatment and plasma viral load of HIV-positive males were analysed. Results From all patients included in the study, 8% tested positive for HIV. The risk of a female being HIV-positive was 3.73 times higher than for males. In the pilot study to explore rapid testing for HBV and HCV, 1% and 1.4% of patients tested positive respectively. When testing for Chlamydia trachomatis 31.3% of females, but no males tested positive. Comparing semen profiles, no significant differences were found between samples from HIV positive and negative males or between HIV positive males categorised by CD4 cell count (p>0.05). For the HIV-positive group with a detectable plasma HIV viral load (>40 copies/ml), a significant difference was observed in the semen viscosity (p=0.0460). Significant differences were noted in the sperm motility (immotile sperm p=0.0456, progressive sperm p=0.0192) of patients receiving antiretroviral (ARV) therapy. Discussion and Conclusion The use of rapid testing is an acceptable and feasible option for improving current screening protocols at the Unit. The absence of definite alterations in the semen characteristics of HIV-positive men further motivates the need for a simpler, point-of-care screening protocol. The prevalence of HBV was lower than that reported in the general population of South Africa and further investigation is needed. Although the sample size was small, HCV prevalence was similar to that of the general population. One third of females tested positive for Chlamydia trachomatis. The methodology used was possibly not appropriate for males. This study highlighted the need for guidelines that address the specialised needs of ART clinics in resource-limited and developing countries with a high HIV prevalence. / Dissertation (MSc)--University of Pretoria, 2013. / gm2014 / Obstetrics and Gynaecology / unrestricted