Infection with the human immunodeficiency virus (HIV) constitutes a global pandemic, and South Africa forms part of the region known to house over two-thirds of HIV infected individuals worldwide. In the early stages of infection, the C-C chemokine receptor type five (CCR5) is the major HIV-1 co-receptor. The importance of this receptor in HIV infection and disease progression was recognised with the discovery of the CCR5 delta 32 (Δ32) allele. Individuals homozygous for this mutation lack functional CCR5 receptors. Consequently, they are almost completely resistant to HIV infection, while the absence of CCR5 has minimal effects on health. Heterozygous individuals display decreased cell surface CCR5 which slows disease progression. Phenotypic expression of CCR5 is heterogeneous and its relation to genetic mutations in the CCR5 gene is not currently known for the South African population. This together with the effect of CCR5 expression on HIV infection provided the rationale for investigating both the phenotypic and genotypic distribution of CCR5. The aim of this study was therefore 1) to investigate CCR5 phenotypic expression on cluster differentiation four (CD4) T-lymphocytes in a group of South African individuals and 2) to analyse the genetic variation in a South African cohort. Flow cytometric methods were used to measure the phenotypic distribution of CCR5 in 245 individuals by assessing both the percentage of CD4+CCR5+ T-cells and CCR5 density. Sixty five individuals, mostly found within the lower CCR5 receptor density range were selected for DNA sequencing. The study found considerable variability in CCR5 expression with South African individuals expressing relatively high CD4+CCR5+ T-cell percentages. Ethnicity was established as a significant variable affecting CCR5 expression with Black African individuals displaying higher (p <0.05) CD4+CCR5+ T-cell percentages and densities than Caucasians. Genotypic data revealed 70 single nucleotide polymorphisms (SNPs), four insertions and the ∆32 deletion. Results showed that Black African individuals have greater genetic diversity with 39 mutations exclusive to this group. The ∆32 mutation was not detected in the Black African group but was identified in the Caucasian group at a frequency of 18.6 %. Twelve novel mutations were identified in this study with two in the open reading frame (ORF). It is evident from the data that the variability in CCR5 phenotypic expression is difficult to correlate with specific mutations in the gene. This thesis provides information on CCR5 distribution and diversity in the South African population which will be of value to patients, clinicians and health policy officials. / Dissertation (MSc)--University of Pretoria, 2011. / Immunology / Unrestricted

The human leukocyte antigen (HLA) is the most polymorphic region in the human genome and accounts for more than 10% of human diversity. This region plays an important role in matching donors and recipients for transplantation. The South African Bone Marrow Registry (SABMR) does not reflect the demographics of the South African population. The large number of polymorphisms resulting from HLA diversity in the Black South African population and their limited representation in the SABMR reduce the chances of finding adequate matches between donors and recipients in this group. Umbilical cord blood is an alternative to bone marrow for the treatment of fatal diseases. Less strict HLA matching is required due to the naive nature of the T cells in cord blood. A public umbilical cord blood bank is a necessity in trying to cater for the diverse population in South Africa. However, the ethnic diversity of the South African population poses a great challenge in constituting a public umbilical cord blood bank that is representative of the entire population. The Roche designed next generation sequencing (NGS) high resolution (HR) HLA typing kit enables sequencing of additional HLA exons and could improve the degree of matching between individuals to ultimately decrease adverse reactions. An extensive study of the literature was performed to establish the demographics, linguistics, and HLA diversity of the South African population to determine how a public cord blood bank should be constituted. In addition, HLA genotyping was performed by 454 NGS on 20 samples that had previously been HLA typed by conventional methods. The 454 NGS technique made use of a Roche designed medium and high resolution HLA typing kit to genotype the samples. It was possible to assign accurate genotypes to 95.5% of the loci of interest for the total number of 20 samples using the MR kit, compared with 98.5% using the HR kit. In conclusion, the present study indicates the extreme HLA diversity in the South African population, and therefore, recommends constituting the first public umbilical cord blood bank in Gauteng on the basis of race or major ethnic groupings. A minimum number of 10 000 cord blood units is needed to initiate the bank. Furthermore, the 454 NGS platform together with the HR HLA typing kit display potential as an alternative method to be used in a public cord blood bank, as well as routine clinical and diagnostic laboratories, to ultimately improve HLA matching between donors and recipients. / Dissertation (MSc)--University of Pretoria, 2013. / gm2014 / Immunology / unrestricted

Introduction: Cardiovascular disease (CVD) is the leading cause of global mortality and its prevalence is increasing among black South Africans in spite of their favourable lipid profile. Apolipoprotein E (ApoE) is a well-described risk factor for CVD and certain polymorphisms within this gene alter the lipid profile. The author hypothesised that there are population-specific effects within the ApoE gene that are responsible for the favourable lipid profile observed in black South Africans whose effects are being altered by environmental factors. Objectives: The main aim of this study was to investigate the associations between specific ApoE single nucleotide polymorphisms (SNPs) and the lipid profile of a black South African population, taking into account certain environmental and phenotypic factors in order to explore the interaction effects between these variables. Methods: Genotyping within this cross-sectional study (n=1 588), nested within the Prospective Urban and Rural Epidemiology (PURE) study, was achieved using Illumina‘s® GoldenGate Genotyping Assay with VeraCode® technology on the BeadXpress® platform (proprietary multiplex fluorescent hybridisation assays on a bead array substrate) or the Bio-Rad CFX Manager© (version 2.0). The Konelab20i™ auto analyser was used for quantitative determination of serum total cholesterol; high-density lipoprotein cholesterol (HDL-C) and triglyceride (TG) concentrations. Low-density lipoprotein cholesterol concentrations were estimated by the Friedewald equation. Results: All SNPs adhered to the assumptions of the Hardy-Weinberg equilibrium, yet the frequency of the SNPs often differed from that reported in other ethnic groups. The well-reported rs429358 and rs7412 SNPs (as the constituent SNPs of the haplotype-genotypes) presented with the strongest associations with various components of the blood lipid profile in the black South African cohort under investigation. Two gene-environment (rs405509 and rs7412) interaction effects on TG remained significant after conducting post hoc tests. Two genotype-phenotype interaction effects between the rs7412 SNP and body mass index and gamma-glutamyl transferase on the HDL-C concentrations remained significant after conducting post hoc tests. Conclusions: The variety of associations between these particular SNPs and the blood lipid profile determined in the present cohort strongly indicates that it is integral to any public health investigation into CVD development that these SNPs be investigated. This study further produced greater insight into the biological mechanisms underlying serum lipid and cholesterol concentrations in a black South African population. Therefore, from these results it is evident that the lipid profile of black South Africans is most definitely influenced by not only genetic variations in the ApoE gene and certain environmental factors, but by the interaction between these factors as well. The present study is the largest study to date to investigate the effect of polymorphisms in the ApoE gene on the lipid profile of black South Africans. / MSc (Nutrition), North-West University, Potchefstroom Campus, 2014

Introduction: Cardiovascular disease (CVD) is the leading cause of global mortality and its prevalence is increasing among black South Africans in spite of their favourable lipid profile. Apolipoprotein E (ApoE) is a well-described risk factor for CVD and certain polymorphisms within this gene alter the lipid profile. The author hypothesised that there are population-specific effects within the ApoE gene that are responsible for the favourable lipid profile observed in black South Africans whose effects are being altered by environmental factors. Objectives: The main aim of this study was to investigate the associations between specific ApoE single nucleotide polymorphisms (SNPs) and the lipid profile of a black South African population, taking into account certain environmental and phenotypic factors in order to explore the interaction effects between these variables. Methods: Genotyping within this cross-sectional study (n=1 588), nested within the Prospective Urban and Rural Epidemiology (PURE) study, was achieved using Illumina‘s® GoldenGate Genotyping Assay with VeraCode® technology on the BeadXpress® platform (proprietary multiplex fluorescent hybridisation assays on a bead array substrate) or the Bio-Rad CFX Manager© (version 2.0). The Konelab20i™ auto analyser was used for quantitative determination of serum total cholesterol; high-density lipoprotein cholesterol (HDL-C) and triglyceride (TG) concentrations. Low-density lipoprotein cholesterol concentrations were estimated by the Friedewald equation. Results: All SNPs adhered to the assumptions of the Hardy-Weinberg equilibrium, yet the frequency of the SNPs often differed from that reported in other ethnic groups. The well-reported rs429358 and rs7412 SNPs (as the constituent SNPs of the haplotype-genotypes) presented with the strongest associations with various components of the blood lipid profile in the black South African cohort under investigation. Two gene-environment (rs405509 and rs7412) interaction effects on TG remained significant after conducting post hoc tests. Two genotype-phenotype interaction effects between the rs7412 SNP and body mass index and gamma-glutamyl transferase on the HDL-C concentrations remained significant after conducting post hoc tests. Conclusions: The variety of associations between these particular SNPs and the blood lipid profile determined in the present cohort strongly indicates that it is integral to any public health investigation into CVD development that these SNPs be investigated. This study further produced greater insight into the biological mechanisms underlying serum lipid and cholesterol concentrations in a black South African population. Therefore, from these results it is evident that the lipid profile of black South Africans is most definitely influenced by not only genetic variations in the ApoE gene and certain environmental factors, but by the interaction between these factors as well. The present study is the largest study to date to investigate the effect of polymorphisms in the ApoE gene on the lipid profile of black South Africans. / MSc (Nutrition), North-West University, Potchefstroom Campus, 2014

Positive identification can be problematic if fingerprinting, DNA, dental history, etc. are no longer available. This may be possible through techniques such as facial approximation, but any form of craniofacial identification requires intimate knowledge of human craniofacial anatomy. Where children are involved, craniofacial changes due to facial growth further complicate matters and require knowledge of tissue thickness and variation in facial shapes. These have hardly been studied in children of African descent. The aims of this study were to provide data on tissue thickness and craniofacial proportions of South African Black and Coloured children and to document the lateral profile shape changes between the ages of 6 and 13 years. Tissue thickness was measured using cephalograms of South African children (n = 388). After digitizing the images, tissue thickness measurements were taken at 11 mid-facial landmarks from each image using the iTEM measuring program. Craniofacial proportions were assessed through assessing standardized anterior and lateral facial photographs of 1749 children. Measurements of facial features were taken using iTEM, from which 28 standard facial indices were calculated. For both tissue thickness and craniofacial indices comparisons between groups per age, sex and ancestry were statistically analyzed. In addition, geometric morphometrics were used to describe lateral facial shape changes and differences age, sex and ancestry (n = 800). The results showed that tissue thickness differences at lower face landmarks are more pronounced in age groups per ancestry as opposed to differences per age and sex. Facial profile per facial shape, class and ancestry showed differences at all landmarks. Craniofacial indices indicated that Coloured children have wider heads, foreheads and faces compared to Black children. The height of the nose and lower lip is longer in Coloured children compared to Black children. In Coloured children, mandibular height and lower face height is shorter in relation to total face height. Males have wider heads, foreheads, mandibles and faces compared to females. The degree of prognathism is dictated by ancestry and to a lesser extent by age and sex as findings showed that maxillary prognathism was more prominent in Black children, while mandibular prognathism were more pronounced in male children. South Africans have a relative concave lateral facial profile due to the maxilla and mandible being more prognathic than in North American children. Differences in lateral face shape between children of various ages, sexes and ancestral groups were visualized through the relative displacement of landmarks related to the forehead and lower face. The resultant differences in lateral facial profile can assist in more accurate estimation of age and ancestry of unknown children. This research created reference datasets for tissue thickness and craniofacial indices of South African children of Black and Coloured ancestry per age and sex that will be useful in the diagnosis of facial dysmorphology and for facial reconstruction / approximation of juvenile remains. It also shed more light on facial growth patterns in the various groups. / Thesis (PhD)--University of Pretoria, 2015. / tm2015 / Anatomy / PhD / Unrestricted

The Cytochrome P450 family of enzymes is responsible for the majority of Phase I metabolism, and has been identified as an important source of pharmacokinetic variation in therapeutic responses. CYP2C19 and CYP2D6, metabolising >35% of commonly prescribed medications, are two of the most important pharmacogenetic markers that have been studied with the aim of improving treatment response and reducing adverse drug reactions. The Food and Drug Administration (FDA) approved AmpliChip CYP450 Test (AmpliChip) was compared to a previously developed PCR-RFLP platform and a newly developed XLPCR+ Sequencing platform for the ability to identifying genotype and predicting phenotype for CYP2C19 and CYP2D6 respectively. The AmpliChip was found not to be genotypically comprehensive enough for evaluating CYP2C19 genotype, not robust enough for determining CYP2D6 genotype and inaccurate in predicting phenotype for both. The XLPCR+ Sequencing method identified three novel alleles and one sub-variant. Advances in online column-switching solid phase extraction generated a rapid and robust LCMS/ MS method for simultaneously quantifying the probe drugs omeprazole (CYP2C19 substrate), dextromethorphan (CYP2D6 substrate) and their metabolites. Antimodes were identified for phenotypic cut-offs which offered measured phenotype for comparison to predicted phenotype. Omeprazole metabolism by CYP2C19 correlated well with predicted phenotype in a demographically representative South African cohort. There are concerns regarding the use of omeprazole as a probe drug as participants predicted to be ultrarapid metabolisers for CYP2C19 had similar rates to extensive metabolisers. Regardless of this concern, decreased metabolism was assigned to the CYP2C19*15 for the first time. CYP2D6 predicted phenotype correlated very well with measured phenotype, validating the suitability of dextromethorphan use for measuring CYP2D6 metabolism. Substrate modified activity score using 0.5 to predict intermediate metabolisers fine-tuned the XLPCR+ Sequencing platform for phenotype prediction. This finding, along with observations in CYP2C19 metabolism of omeprazole, highlights the importance of substrate specific phenotype prediction strategies. Controversially, attempts to associate CYP2D6 phenotype prediction with risperidone-related adverse drug reactions has yielded conflicting results. The XL-PCR+Sequencing platform was able to discount this association by predicting a variety of metabolisers in a pilot cohort selected to be experiencing risperidone-related adverse drug reactions. The comprehensive capability of the XL-PCR+Sequencing allowed for the identification of an additional novel allele in this cohort. The data presented in thisthesis has provided insight into the relationship between predicted and measured phenotype for CYP2C19 and CYP2D6 in the South African population. The XL-PCR+Sequencing platform can be used for future research or can be applied to improve treatment outcome. The LC-MS/MS method developed could be used for future evaluations of predicted and measured phenotype with the ability to be adjusted for therapeutic drug monitoring. This thesis advances pharmacogenetics of CYP2C19 and CYP2D6 for use in the South African population. / Thesis (PhD)--University of Pretoria, 2013. / gm2014 / Pharmacology / unrestricted

Thesis (M.Sc. (Physiology)) -- University of Limpopo, 2019 / Background Type 2 diabetes mellitus (T2DM) is an increasing challenge globally, and is estimated to affect 439 million adults by 2030. This estimate is linked to an unhealthy lifestyle with characteristics such as low physical activity (PA) and high plasma glucose levels (PGLs). Studies associating PA with insulin resistance and diabetes among adults and adolescents have been conducted widely in developed countries. Such studies are scanty among rural populations, especially in Africa. Assessment of the burden of diabetes and associated lifestyle risk factors in developing countries is essential in order to encourage appropriate intervention strategies to counter the increasing prevalence. Aim and objectives The aim of this study was to investigate the relationship between PA and T2DM among rural young adults aged 22 to 30 years in Ellisras area in Limpopo Province, South Africa Methods A total of 713 young adults (349 males and 364 females) who have been part of the Ellisras Longitudinal Study participated in the current study. Physical activity data was collected using a validated questionnaire. After an overnight fast, participants provided fasting venous blood samples for determination of plasma glucose and insulin. Insulin resistance was estimated using the homeostasis model assessment of insulin resistance. Anthropometric measurements (waist circumference and height) were performed using standard procedures. Linear and logistic regressions were used to assess the relationship between PA, pre-diabetes, insulin resistance and T2DM; and the odds of having T2DM with low PA levels. Results The prevalence of physical inactivity was 67.3 and 71.0% for males and females, respectively. That of pre-diabetes was between 45.7% and 50.2%. The prevalence of diabetes was 9.6% for males and 10.1% for females while for insulin resistance was 22.9% for males and 29.3% for females. Linear regression found a significant relationship (p<0.05) between physical activity and blood glucose (ß =5.715; 95% CI 4.545; 6.885), waist circumference (ß = 37.572; 95% CI 25.970; 49.174) and waist-toheight ratio (ß = 0.192; 95% CI 0.087; 0.296). Logistic regression found a significant (p<0.05) relationship between low physical activity and T2DM (Odds ratio = 2.890; 95% CI 1.715; 4.870) and insulin resistance (Odds ratio = 1.819; 95% CI 1.266; 2.614). Conclusion Physical activity is low in this population, and is independently associated with T2DM and insulin resistance. KEY WORDS Type 2 diabetes mellitus; pre-diabetes; insulin resistance; physical activity; young adults; rural South African population. / Vrije University, Amsterdam, The Netherlands, and the University of Limpopo

Student Number : 9807768F - MSc (Med) research report - Faculty of Health Sciences / Fanconi anaemia (FA) is an autosomal recessive disorder characterised by aplastic anaemia (AA) and a high risk of developing acute myeloid leukaemia (AML). It is unknown whether heterozygote carriers are also predisposed to developing these disorders. The black South African population group is ideal for FA mutation screening because the presence of a founder mutation, FANCG 637-643, makes screening relatively straight forward. Three individuals with AML (115 screened) and one with AA (78 screened) were found to be heterozygous for the black South African founder mutation. From our data it seems unlikely that this mutation places heterozygous carriers of the mutation at high risk of developing AML or AA. Three children with AA out of 26 screened, were homozygous for the mutation. This finding reiterates the importance of screening all children with AA for FA. The frequency of certain congenital abnormalities in black South African FA patients was compared to patients described by other research groups. The frequencies of the abnormalities were similar to other FANCG cohorts described but significant differences to a group of FA patients from unspecified complementation groups were found. This difference could be because different complementation groups are associated more or less strongly with specific abnormalities. It was found previously that particular congenital abnormalities in FA patients are associated with a poor haematological outcome. We concluded that black South African FANCG patients have a high risk of early development of AA even though they do not have a high frequency of congenital abnormalities.

INTRODUCTION AND AIM - Cardiovascular disease is globally a major risk factor for morbidity and mortality. It is caused by various factors, one of which is an abnormal haemostatic process. Fibrinogen is a haemostatic factor that is considered to be an independent risk factor for cardiovascular disease. Elevated fibrinogen can be caused by environmental and genetic factors which increase the risk of the occurrence of thrombosis. The fibrinogen y' chain, which is one of the three chains of fibrinogen, has two different variants, the yA and y’. The presence of the fibrinogen y’ chain has been associated with thrombotic disorders. Many studies have investigated the fibrinogen variables in Caucasian individuals, but only a few such studies have been conducted on non-Caucasian individuals. The genetic diversity of ethnic groups differs and could cause differences in the fibrinogen variables between these groups. Fibrinogen is known to increase with age; therefore to explain changes over time in fibrinogen concentrations it was also important to investigate whether genetic determinants and possible gene–environment interactions influenced fibrinogen over time. In this study the main aim was to determine the change in the fibrinogen variables over a five-year period within a black South African cohort subdivided according to genotypes associated with fibrinogen variables, and to determine whether the observed changes were modulated by environmental factors. PARTICIPANTS AND METHODS - Data [baseline (n=2010) and follow-up (n=1288)] were collected in the Prospective Urban and Rural Epidemiology (PURE) study during 2005 and 2010 from apparently healthy black men and women aged between 35 and 65 years and residing in rural or urban settlements. Experimental methods included analysis of fibrinogen and fibrinogen y’ concentrations, single nucleotide polymorphisms (SNPs) and determination of environmental factors associated with the fibrinogen variables. RESULTS - The fibrinogen variables increased significantly from 2005 to 2010 in both the rural and urban participants, as well as in both men and women. The major environmental factors that affected the fibrinogen variables were C-reactive protein (CRP), interleukin-6 (IL-6), body mass index (BMI), glycated haemoglobin (HbA1c), age, blood lipids, human immunodeficiency virus (HIV) and tobacco use. Fibrinogen increased consistently from 2005 to 2010 in the respective genotypes of all SNPs analysed, except in the FGG 9340 T>C homozygous mutant carriers. Fibrinogen y’ also increased in general in most genotypes from 2005 to 2010, except in the FGG 10034 C>T mutant allele carriers, where a decrease was observed. It was determined that CRP was the only environmental factor that influenced the change in fibrinogen over time and that FGG 10034 C>T was the only SNP that influenced the change in fibrinogen y’ over the five years. Four gene–environment interactions also influenced fibrinogen on a cross-sectional level, i.e. FGA 2224 G>A with age, FGB Arg448Lys with HIV status, FGB 1643 C>T with urbanisation and FGB 1038 G>A with HbA1c. Only the FGG 9340 T>C with HbA1c interaction was found to predict change in fibrinogen concentrations over the five years. CONCLUSION - Both environmental and genetic factors significantly influenced the fibrinogen variables cross-sectionally as well as prospectively. It was clear that the influence of the environmental factors was mediated by genetic polymorphisms and vice versa, as can be seen by the gene–environment interactions found in this study. An important finding of this study was that the interaction of HbA1c with two SNPs on fibrinogen variables may explain the known inconsistent relationship found between fibrinogen concentrations and diabetes. / MSc (Dietetics), North-West University, Potchefstroom Campus, 2014

INTRODUCTION AND AIM - Cardiovascular disease is globally a major risk factor for morbidity and mortality. It is caused by various factors, one of which is an abnormal haemostatic process. Fibrinogen is a haemostatic factor that is considered to be an independent risk factor for cardiovascular disease. Elevated fibrinogen can be caused by environmental and genetic factors which increase the risk of the occurrence of thrombosis. The fibrinogen y' chain, which is one of the three chains of fibrinogen, has two different variants, the yA and y’. The presence of the fibrinogen y’ chain has been associated with thrombotic disorders. Many studies have investigated the fibrinogen variables in Caucasian individuals, but only a few such studies have been conducted on non-Caucasian individuals. The genetic diversity of ethnic groups differs and could cause differences in the fibrinogen variables between these groups. Fibrinogen is known to increase with age; therefore to explain changes over time in fibrinogen concentrations it was also important to investigate whether genetic determinants and possible gene–environment interactions influenced fibrinogen over time. In this study the main aim was to determine the change in the fibrinogen variables over a five-year period within a black South African cohort subdivided according to genotypes associated with fibrinogen variables, and to determine whether the observed changes were modulated by environmental factors. PARTICIPANTS AND METHODS - Data [baseline (n=2010) and follow-up (n=1288)] were collected in the Prospective Urban and Rural Epidemiology (PURE) study during 2005 and 2010 from apparently healthy black men and women aged between 35 and 65 years and residing in rural or urban settlements. Experimental methods included analysis of fibrinogen and fibrinogen y’ concentrations, single nucleotide polymorphisms (SNPs) and determination of environmental factors associated with the fibrinogen variables. RESULTS - The fibrinogen variables increased significantly from 2005 to 2010 in both the rural and urban participants, as well as in both men and women. The major environmental factors that affected the fibrinogen variables were C-reactive protein (CRP), interleukin-6 (IL-6), body mass index (BMI), glycated haemoglobin (HbA1c), age, blood lipids, human immunodeficiency virus (HIV) and tobacco use. Fibrinogen increased consistently from 2005 to 2010 in the respective genotypes of all SNPs analysed, except in the FGG 9340 T>C homozygous mutant carriers. Fibrinogen y’ also increased in general in most genotypes from 2005 to 2010, except in the FGG 10034 C>T mutant allele carriers, where a decrease was observed. It was determined that CRP was the only environmental factor that influenced the change in fibrinogen over time and that FGG 10034 C>T was the only SNP that influenced the change in fibrinogen y’ over the five years. Four gene–environment interactions also influenced fibrinogen on a cross-sectional level, i.e. FGA 2224 G>A with age, FGB Arg448Lys with HIV status, FGB 1643 C>T with urbanisation and FGB 1038 G>A with HbA1c. Only the FGG 9340 T>C with HbA1c interaction was found to predict change in fibrinogen concentrations over the five years. CONCLUSION - Both environmental and genetic factors significantly influenced the fibrinogen variables cross-sectionally as well as prospectively. It was clear that the influence of the environmental factors was mediated by genetic polymorphisms and vice versa, as can be seen by the gene–environment interactions found in this study. An important finding of this study was that the interaction of HbA1c with two SNPs on fibrinogen variables may explain the known inconsistent relationship found between fibrinogen concentrations and diabetes. / MSc (Dietetics), North-West University, Potchefstroom Campus, 2014