Bioinformatics-driven development of a queryable cardiometabolic database and its application in a biological setting

A thesis submitted to the Faculty of Science, University of the Witwatersrand, Johannesburg in fulfilment of the requirements for the degree of Doctor of Philosophy. June 2017, JohannesburgAs sequencing and genotyping technologies are advancing, larger and more complex sets of biological data are being produced. Databases can be used to efficiently store and manage the data. Typically, publicly available datasets are accessed through web browsers that offer a user-friendly interface to a database, making complex queries simple to execute. However, research projectspecific data are not commonly stored in this way. In this research, a database (designed in MySQL) and accompanying interface (developed using PHP, HTML and CSS) has been designed for the storage and querying of the quality controlled data from the current project using Metabochip-genotyped Birth to Twenty (Bt20) cohort participants and their female caregivers. Users can easily access the data to generate summary statistics on the phenotype data and download phenotype, single nucleotide polymorphism (SNP) annotation and association analysis data that match user-supplied criteria. Some of the data from the database was used to investigate the genetics of blood pressure (BP) in black South African individuals. Hypertension is a major risk factor for cardiovascular diseases (CVDs). BP variation is known to have a genetic component, but genetic studies in indigenous Africans have been limited. Association analysis, carried out in a merged sample of caregivers and participants, pointed to novel regions of interest in the NOS1AP (DBP and SBP), MYRF (SBP) and POC1B (SBP) genes and two intergenic regions (DACH1|LOC440145 (DBP and SBP) and INTS10|LPL (SBP)). Two SNPs in the MYRF gene met the calculated “array-wide” significance threshold (p<6.7x10-7 for the merged dataset) for multiple testing. Genotype imputation is a useful addition to association studies to increase the SNP panel for association testing. An investigation into the efficiency of imputation in this dataset using a mixed population reference panel was carried out. Imputation was achieved with high confidence in all genes, but a more detailed view of the region was only seen in NOS1AP (DBP and SBP in both the merged and female caregiver datasets) and POC1B (Bt20 participant dataset only). Overall, the research contributed a useful tool for the efficient management of project-specific biological data. The analysis and genotype imputation, which is a promising tool in future studies in this or other African datasets, also provided some insight into the genetics of blood pressure in black South Africans with further functional and replication studies in larger samples required to confirm and explain the findings.MT 201

Genetic risk score for adult body mass index associations with childhood and adolescent weight gain in an African population

Abstract Background Ninety-seven independent single nucleotide polymorphisms (SNPs) are robustly associated with adult body mass index (BMI kg/m2) in Caucasian populations. The relevance of such variants in African populations at different stages of the life course (such as childhood) is unclear. We tested whether a genetic risk score composed of the aforementioned SNPs was associated with BMI from infancy to early adulthood. We further tested whether this genetic effect was mediated by conditional weight gain at different growth periods. We used data from the Birth to Twenty Plus Cohort (Bt20+), for 971 urban South African black children from birth to 18 years. DNA was collected at 13 years old and was genotyped using the Metabochip (Illumina) array. The weighted genetic risk score (wGRS) for BMI was constructed based on 71 of the 97 previously reported SNPs. Results The cross-sectional association between the wGRS and BMI strengthened with age from 5 to 18 years. The significant associations were observed from 11 to 18 years, and peak effect sizes were observed at 13 and 14 years of age. Results from the linear mixed effects models showed significant interactions between the wGRS and age on longitudinal BMI but no such interactions were observed in sex and the wGRS. A higher wGRS was associated with an increased relative risk of belonging to the early onset obese longitudinal BMI trajectory (relative risk = 1.88; 95%CI 1.28 to 2.76) compared to belonging to a normal longitudinal BMI trajectory. Adolescent conditional relative weight gain had a suggestive mediation effect of 56% on the association between wGRS and obesity risk at 18 years. Conclusions The results suggest that genetic susceptibility to higher adult BMI can be tracked from childhood in this African population. This supports the notion that prevention of adult obesity should begin early in life. The genetic risk score combined with other non-genetic risk factors, such as BMI trajectory membership in our case, has the potential to be used to screen for early identification of individuals at increased risk of obesity and other related NCD risk factors in order to reduce the adverse health risk outcomes later

Assessing computational genomics skills: Our experience in the H3ABioNet African bioinformatics network

The H3ABioNet pan-African bioinformatics network, which is funded to support the Human Heredity and Health in Africa (H3Africa) program, has developed node-assessment exercises to gauge the ability of its participating research and service groups to analyze typical genome-wide datasets being generated by H3Africa research groups. We describe a framework for the assessment of computational genomics analysis skills, which includes standard operating procedures, training and test datasets, and a process for administering the exercise. We present the experiences of 3 research groups that have taken the exercise and the impact on their ability to manage complex projects. Finally, we discuss the reasons why many H3ABioNet nodes have declined so far to participate and potential strategies to encourage them to do so

Genetics of HIV-associated sensory neuropathy in black Southern Africans

HIV-associated sensory neuropathy (HIV-SN) is a common complication associated with human immunodeficiency virus (HIV)-infection. A common symptom of HIV-SN is pain. Variation at specific loci within certain candidate genes has been suggested to alter susceptibility to developing HIV-SN as well as the susceptibility to developing pain and the intensity of the pain experienced. Few studies, however, have been conducted in individuals of black African ancestry. The aim of the current research was to conduct an in-depth study, in a black Southern African population, of genes previously associated with susceptibility to developing HIV-SN (TNFA and surrounding genes and IL12B) and variations in pain susceptibility and intensity (GCH1, KCNS1, IL1B, IL6, CCL2 and CCR2) in other neuropathic pain states. Single nucleotide polymorphisms (SNPs) identified in the literature were supplemented with population appropriate tagSNPs to improve assessment of the genes in an African population. Genotyping of previously collected deoxyribonucleic acid (DNA) samples was carried out using a GoldenGate assay with VeraCode microbeads and data were read on an Illumina BeadXpress Reader. Data were statistically analysed to assess the association of the genetic variants with susceptibility to developing HIV-SN and pain and variability of pain intensity in those patients with painful HIV-SN. Although some SNPs and haplotypes in the genes investigated associated with HIV-SN susceptibility (TNFA region), pain susceptibility (TNFA region, IL12B, KCNS1, IL6 and CCR2) or pain intensity (TNFA region, KCNS1, IL1B and IL6), none of the results were consistent with that which has been found in previous studies in non-African populations. Reasons for this may be that associations are population-specific or model-specific. Limitations of the study included the use of a relatively small sample, the method of sampling (convenience sampling), genotyping failure and tagging inefficiency in some instances, and the fact that there is no Southern African population dataset to use for tagSNP selection. My findings emphasise the importance of conducting genetic association studies in separate ethnic groups

Genetic risk score for adult body mass index associations with childhood and adolescent weight gain in an African population

Abstract Background Ninety-seven independent single nucleotide polymorphisms (SNPs) are robustly associated with adult body mass index (BMI kg/m2) in Caucasian populations. The relevance of such variants in African populations at different stages of the life course (such as childhood) is unclear. We tested whether a genetic risk score composed of the aforementioned SNPs was associated with BMI from infancy to early adulthood. We further tested whether this genetic effect was mediated by conditional weight gain at different growth periods. We used data from the Birth to Twenty Plus Cohort (Bt20+), for 971 urban South African black children from birth to 18 years. DNA was collected at 13 years old and was genotyped using the Metabochip (Illumina) array. The weighted genetic risk score (wGRS) for BMI was constructed based on 71 of the 97 previously reported SNPs. Results The cross-sectional association between the wGRS and BMI strengthened with age from 5 to 18 years. The significant associations were observed from 11 to 18 years, and peak effect sizes were observed at 13 and 14 years of age. Results from the linear mixed effects models showed significant interactions between the wGRS and age on longitudinal BMI but no such interactions were observed in sex and the wGRS. A higher wGRS was associated with an increased relative risk of belonging to the early onset obese longitudinal BMI trajectory (relative risk = 1.88; 95%CI 1.28 to 2.76) compared to belonging to a normal longitudinal BMI trajectory. Adolescent conditional relative weight gain had a suggestive mediation effect of 56% on the association between wGRS and obesity risk at 18 years. Conclusions The results suggest that genetic susceptibility to higher adult BMI can be tracked from childhood in this African population. This supports the notion that prevention of adult obesity should begin early in life. The genetic risk score combined with other non-genetic risk factors, such as BMI trajectory membership in our case, has the potential to be used to screen for early identification of individuals at increased risk of obesity and other related NCD risk factors in order to reduce the adverse health risk outcomes later

Genetic risk score for adult body mass index associations with childhood and adolescent weight gain in an African population.

Background: Ninety-seven independent single nucleotide polymorphisms (SNPs) are robustly associated with adult body mass index (BMI kg/m2) in Caucasian populations. The relevance of such variants in African populations at different stages of the life course (such as childhood) is unclear. We tested whether a genetic risk score composed of the aforementioned SNPs was associated with BMI from infancy to early adulthood. We further tested whether this genetic effect was mediated by conditional weight gain at different growth periods. We used data from the Birth to Twenty Plus Cohort (Bt20+), for 971 urban South African black children from birth to 18 years. DNA was collected at 13 years old and was genotyped using the Metabochip (Illumina) array. The weighted genetic risk score (wGRS) for BMI was constructed based on 71 of the 97 previously reported SNPs. Results: The cross-sectional association between the wGRS and BMI strengthened with age from 5 to 18 years. The significant associations were observed from 11 to 18 years, and peak effect sizes were observed at 13 and 14 years of age. Results from the linear mixed effects models showed significant interactions between the wGRS and age on longitudinal BMI but no such interactions were observed in sex and the wGRS. A higher wGRS was associated with an increased relative risk of belonging to the early onset obese longitudinal BMI trajectory (relative risk = 1.88; 95%CI 1.28 to 2.76) compared to belonging to a normal longitudinal BMI trajectory. Adolescent conditional relative weight gain had a suggestive mediation effect of 56% on the association between wGRS and obesity risk at 18 years. Conclusions: The results suggest that genetic susceptibility to higher adult BMI can be tracked from childhood in this African population. This supports the notion that prevention of adult obesity should begin early in life. The genetic risk score combined with other non-genetic risk factors, such as BMI trajectory membership in our case, has the potential to be used to screen for early identification of individuals at increased risk of obesity and other related NCD risk factors in order to reduce the adverse health risk outcomes later

Genetic risk score for adult body mass index associations with childhood and adolescent weight gain in an African population

Abstract Background Ninety-seven independent single nucleotide polymorphisms (SNPs) are robustly associated with adult body mass index (BMI kg/m2) in Caucasian populations. The relevance of such variants in African populations at different stages of the life course (such as childhood) is unclear. We tested whether a genetic risk score composed of the aforementioned SNPs was associated with BMI from infancy to early adulthood. We further tested whether this genetic effect was mediated by conditional weight gain at different growth periods. We used data from the Birth to Twenty Plus Cohort (Bt20+), for 971 urban South African black children from birth to 18 years. DNA was collected at 13 years old and was genotyped using the Metabochip (Illumina) array. The weighted genetic risk score (wGRS) for BMI was constructed based on 71 of the 97 previously reported SNPs. Results The cross-sectional association between the wGRS and BMI strengthened with age from 5 to 18 years. The significant associations were observed from 11 to 18 years, and peak effect sizes were observed at 13 and 14 years of age. Results from the linear mixed effects models showed significant interactions between the wGRS and age on longitudinal BMI but no such interactions were observed in sex and the wGRS. A higher wGRS was associated with an increased relative risk of belonging to the early onset obese longitudinal BMI trajectory (relative risk = 1.88; 95%CI 1.28 to 2.76) compared to belonging to a normal longitudinal BMI trajectory. Adolescent conditional relative weight gain had a suggestive mediation effect of 56% on the association between wGRS and obesity risk at 18 years. Conclusions The results suggest that genetic susceptibility to higher adult BMI can be tracked from childhood in this African population. This supports the notion that prevention of adult obesity should begin early in life. The genetic risk score combined with other non-genetic risk factors, such as BMI trajectory membership in our case, has the potential to be used to screen for early identification of individuals at increased risk of obesity and other related NCD risk factors in order to reduce the adverse health risk outcomes later

Additional file 1: Table S1. of Insights into the genetics of blood pressure in black South African individuals: the Birth to Twenty cohort

Correlation of findings in our study with previous studies. (XLSX 13 kb

Publisher's PDF: Role of TNF block genetic variants in HIV-associated sensory neuropathy in black Southern Africans. DOI: 10.1038/ejhg.2014.104

<b>Post-print copy of: </b>Wadley AL, Hendry LM, Kamerman PR, Chew CSN, Price P, Cherry CL, Lombard Z. Role of TNF block genetic variants in HIV-associated sensory neuropathy in black Southern Africans. European Journal of Human Genetics 23: 363-368, 2015 DOI: 10.1038/ejhg.2014.104, PMID: 24896147<div><br></div><div><b>Abstract: </b>HIV-associated sensory neuropathy (HIV-SN) is a common neurological complication of HIV infection. The TNF block is a region within the central MHC that contains many immunoregulatory genes. Polymorphisms and haplotypes of the TNF block have been associated with increased risk of HIV-SN in Asians and whites. Here we investigated genetic associations with HIV-SN in 342 black Southern Africans (190 cases and 152 neuropathy-free controls) using single nucleotide polymorphisms (SNPs) spanning the TNF block and a set of haplotypes defined by 31 SNPs in Asian and white populations (denoted FVa). We included population-appropriate tagSNPs derived from an African population (Yoruban, YRI, HapMap) and derived extended haplotypes comprising 61 SNPs (denoted FVa_ext b). We found no association between HIV-SN and carriage of two SNPs (TNF-1031/rs1799964*C and BAT1 (intron10)/rs9281523*C) associated with HIV-SN in whites and Asians. Additionally, a haplotype containing TNF-1031/rs1799964*C associated with increased risk of HIV-SN in Asians, but was not present in this African population. However, alleles of seven SNPs associated with reduced risk of HIV-SN (corrected for age, height and multiple comparisons). These were rs11796*A, rs3130059*G, rs2071594*C, NFKBIL1-62/rs2071592*A, rs2071591*A, LTA+252/rs909253*G, rs1041981*C. One haplotype (FV18_ext1), not containing these alleles, was associated with increased risk of HIV-SN after correction for age, height and multiple comparisons. Our results confirm the involvement of genes in the TNF block in altering risk for HIV-SN, but genotypes critical in this African population differed from those affecting HIV-SN in whites and Asians. These differences support the need for genetic association studies in diverse populations.</div

Insights into the genetics of blood pressure in black South African individuals: the Birth to Twenty cohort

Abstract Background Cardiovascular diseases (CVDs) are the leading cause of non-communicable disease deaths globally, with hypertension being a major risk factor contributing to CVDs. Blood pressure is a heritable trait, with relatively few genetic studies having been performed in Africans. This study aimed to identify genetic variants associated with variance in systolic (SBP) and diastolic (DBP) blood pressure in black South Africans. Methods Genotyping was performed using the Metabochip in a subset of participants (mixed sex; median age 17.9) and their adult female caregivers (median age 41.0) from the Birth to Twenty cohort (n = 1947). Data were analysed as a merged dataset (all participants and caregivers together) in GEMMA (v0.94.1) using univariate linear mixed models, incorporating a centered relatedness matrix to account for the relatedness between individuals and with adjustments for age, sex, BMI and principal components of the genotype information. Results Association analysis identified regions of interest in the NOS1AP (DBP: rs112468105 - p = 7.18 × 10−5 and SBP: rs4657181 - p = 4.04 × 10−5), MYRF (SBP: rs11230796 - p = 2.16 × 10−7, rs400075 - p = 2.88 × 10−7) and POC1B (SBP: rs770373 - p = 7.05 × 10−5, rs770374 - p = 9.05 × 10−5) genes and some intergenic regions (DACH1|LOC440145 (DBP: rs17240498 - p = 4.91 × 10−6 and SBP: rs17240498 - p = 2.10 × 10−5) and INTS10|LPL (SBP: rs55830938 - p = 1.30 × 10−5, rs73599609 - p = 5.78 × 10−5, rs73667448 - p = 6.86 × 10−5)). Conclusions The study provided further insight into the contribution of genetic variants to blood pressure in black South Africans. Future functional and replication studies in larger samples are required to confirm the role of the identified loci in blood pressure regulation and whether or not these variants are African-specific