Browsing by Author "Van Heerden, Carel J."

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    Distinct host-immune response toward species related intracellular mycobacterial killing : a transcriptomic study
    (Taylor & Francis, 2020) Madhvi, Abhilasha; Mishra, Hridesh; Chegoua, Novel N.; Tromp, Gerard; Van Heerden, Carel J.; Pietersen, R. D.; Leisching, Gina; Baker, Bienyameen
    The comparison of the host immune response when challenged with pathogenic and nonpatho- genic species of mycobacteria can provide answers to the unresolved question of how pathogens subvert or inhibit an effective response. We infected human monocyte derived macrophages (hMDMs) with different species of mycobacteria, in increasing order of pathogenicity, i.e. M. smegmatis, M. bovis BCG, and M. tuberculosis R179 that had been cultured in the absence of detergents. RNA was isolated post-infection and transcriptomic analysis using amplicons (Ampliseq) revealed 274 differentially expressed genes (DEGs) across three species, out of which we selected 19 DEGs for further validation. We used qRT-PCR to confirm the differential expression of 19 DEGs. We studied biological network through Ingenuity Pathway Analysis® (IPA) which revealed up-regulated pathways of the interferon and interleukin family related to the killing of M. smegmatis. Apart from interferon and interleukin family, we found one up-regulated (EIF2AK2) and two down-regulated (MT1A and TRIB3) genes as unique potential targets found by Ampliseq and qRT-PCR which may be involved in the intracellular mycobacterial killing. The roles of these genes have not previously been described in tuberculosis. Multiplex ELISA of culture supernatants showed increased host immune response toward M. smegmatis as compared to M. bovis BCG and M.tb R179. These results enhance our understanding of host immune response against M.tb infection.
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    Identification of an iron-responsive subtype in two children diagnosed with relapsing-remitting multiple sclerosis using whole exome sequencing
    (Elsevier, 2019-06) Van Rensburg, Susan J.; Peeters, Armand V.; Van Toorn, Ronald; Schoeman, Johan; Moremi, Kelebogile E.; Van Heerden, Carel J.; Kotze, Maritha J.
    Background: Multiple sclerosis is a disorder related to demyelination of axons. Iron is an essential cofactor in myelin synthesis. Previously, we described two children (males of mixed ancestry) with relapsing-remitting multiple sclerosis (RRMS) where long-term remission was achieved by regular iron supplementation. A genetic defect in iron metabolism was postulated, suggesting that more advanced genetic studies could shed new light on disease pathophysiology related to iron. Methods: Whole exome sequencing (WES) was performed to identify causal pathways. Blood tests were performed over a 10 year period to monitor the long-term effect of a supplementation regimen. Clinical wellbeing was assessed quarterly by a pediatric neurologist and regular feedback was obtained from the schoolteachers. Results: WES revealed gene variants involved in iron absorption and transport, in the transmembrane protease, serine 6 (TMPRSS6) and transferrin (TF) genes; multiple genetic variants in CUBN, which encodes cubilin (a receptor involved in the absorption of vitamin B12 as well as the reabsorption of transferrin-bound iron and vitamin D in the kidneys); SLC25A37 (involved in iron transport into mitochondria) and CD163 (a scavenger receptor involved in hemorrhage resolution). Variants were also found in COQ3, involved with synthesis of Coenzyme Q10 in mitochondria. Neither of the children had the HLA-DRB1*1501 allele associated with increased genetic risk for MS, suggesting that the genetic contribution of iron-related genetic variants may be instrumental in childhood MS. In both children the RRMS has remained stable without activity over the last 10 years since initiation of nutritional supplementation and maintenance of normal iron levels, confirming the role of iron deficiency in disease pathogenesis in these patients. Conclusion: Our findings highlight the potential value of WES to identify heritable risk factors that could affect the reabsorption of transferrin-bound iron in the kidneys causing sustained iron loss, together with inhibition of vitamin B12 absorption and vitamin D reabsorption (CUBN) and iron transport into mitochondria (SLC25A37) as the sole site of heme synthesis. This supports a model for RRMS in children with an apparent iron-deficient biochemical subtype of MS, with oligodendrocyte cell death and impaired myelination possibly caused by deficits of energy- and antioxidant capacity in mitochondria.
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    Optimization of automated ribosomal intergenic spacer analysis for the estimation of microbial diversity in fynbos soil
    (Academy of Science of South Africa -- ASSAF, 2010-08) Slabbert, Etienne; Van Heerden, Carel J.; Jacobs, Karin
    Automated ribosomal intergenic spacer analysis (ARISA) has become a commonly used molecular technique for the study of microbial populations in environmental samples. The reproducibility and accuracy of ARISA, with and without the polymerase chain reaction (PCR) are important aspects that influence the results and effectiveness of these techniques. We used the primer set ITS4/ITS5 for ARISA to assess the fungal community composition of two sites situated in the Sand Fynbos. The primer set proved to deliver reproducible ARISA profiles of the fungal community composition with little variation observed between ARISA-PCRs. Variation that occurred in a sample due to repeated DNA extraction is expected for ecological studies. This reproducibility made ARISA a useful tool for the assessment and comparison of diversity in ecological samples. In this paper, we also offered particular suggestions concerning the binning strategy for the analysis of ARISA profiles.
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    Targeted next-generation sequencing identifies novel variants in candidate genes for Parkinson’s disease in Black South African and Nigerian patients
    (BioMed Central, 2020-02-04) Oluwole, Oluwafemi G.; Kuivaniemi, Helena; Abrahams, Shameemah; Haylett, William L.; Vorster, Alvera A.; Van Heerden, Carel J.; Kenyon, Colin P.; Tabb, David L.; Fawale, Michael B.; Sunmonu, Taofiki A.; Ajose, Abiodun; Olaogun, Matthew O.; Rossouw, Anastasia C.; Van Hillegondsberg, Ludo S.; Carr, Jonathan; Ross, Owen A.; Komolafe, Morenikeji A.; Tromp, Gerard; Bardien, Soraya
    Background: The prevalence of Parkinson’s disease (PD) is increasing in sub-Saharan Africa, but little is known about the genetics of PD in these populations. Due to their unique ancestry and diversity, sub-Saharan African populations have the potential to reveal novel insights into the pathobiology of PD. In this study, we aimed to characterise the genetic variation in known and novel PD genes in a group of Black South African and Nigerian patients. Methods: We recruited 33 Black South African and 14 Nigerian PD patients, and screened them for sequence variants in 751 genes using an Ion AmpliSeq™ Neurological Research panel. We used bcftools to filter variants and annovar software for the annotation. Rare variants were prioritised using MetaLR and MetaSVM prediction scores. The effect of a variant on ATP13A2’s protein structure was investigated by molecular modelling. Results: We identified 14,655 rare variants with a minor allele frequency ≤ 0.01, which included 2448 missense variants. Notably, no common pathogenic mutations were identified in these patients. Also, none of the known PD-associated mutations were found highlighting the need for more studies in African populations. Altogether, 54 rare variants in 42 genes were considered deleterious and were prioritized, based on MetaLR and MetaSVM scores, for follow-up studies. Protein modelling showed that the S1004R variant in ATP13A2 possibly alters the conformation of the protein. Conclusions: We identified several rare variants predicted to be deleterious in sub-Saharan Africa PD patients; however, further studies are required to determine the biological effects of these variants and their possible role in PD. Studies such as these are important to elucidate the genetic aetiology of this disorder in patients of African ancestry.