Browsing by Author "Jooste, Tracey"

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    The detection of Citrus tristeza virus genetic variants using pathogen specific electronic probes
    (Stellenbosch : Stellenbosch University, 2017-03) Jooste, Tracey; Maree, H. J.; Burger, J. T.; Visser, M.; Stellenbosch University. Faculty of AgriSciences. Dept. of Genetics.
    ENGLISH ABSTRACT: Citrus tristeza virus (CTV), a complex pathogen of citrus spp., is endemic to South Africa and has been responsible for great losses locally and internationally. CTV causes severe stem pitting in grapefruit, which forms an important sector of South Africa's citrus production and export market. The limited understanding of CTV’s ability to cause severe disease in one host while no symptoms in another restricts the implementation of effective management strategies. The conservation of plant biosecurity relies on the rapid identification of pathogenic organisms including viruses. While there are many molecular assays available for the detection of plant viruses, these are often limited in their ability to test for multiple viruses simultaneously. However, with next-generation sequencing (NGS) based metagenomic analysis it is possible to detect multiple viruses within a sample, including low-titre and novel viruses, at the same time. Conventional NGS data analysis has computational limitations during contig assembly and similarity searches in sequence databases, which prolongs the time required for a diagnostic result. In this study, an alternative targeted method was explored for the simultaneous detection of 11 recognised citrus viruses in NGS data using electronic probes (e-probes). E-probes were designed, optimised and screened against raw, unassembled NGS data in order to minimise the bioinformatic processing time required. The e-probes were able to accurately detect their cognate viruses in simulated datasets, without any false negatives or positives. The efficiency of the e-probe based approach was validated with NGS datasets generated from different RNA preparations: dsRNA from ‘Mexican’ lime infected with different CTV genotypes, dsRNA from field samples, as well as small RNA and total RNA from grapefruit infected with the CTV T3 genotype. A set of probes were made publically available that is able to accurately detect CTV in NGS data irrespective of which genotype the plants are infected with. The results were confirmed by performing de novo assemblies of the high quality read datasets and subsequent BLAST analyses. This sequence based detection method eliminates the need for NGS data assembly, ultimately reducing the virus-detection turnaround time.
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    Human whole genome sequencing in South Africa
    (Nature, 2021-01) Glanzmann, Brigitte; Jooste, Tracey; Ghoor, Samira; Gordon, Richard; Mia, Rizwana; Mao, Jun; Li, Hao; Charls, Patrick; Douman, Craig; Kotze, Maritha J.; Peeters, Armand V.; Loots, Glaudina; Esser, Monika; Tiemessen, Caroline T.; Wilkinson, Robert J.; Louw, Johan; Gray, Glenda; Warren, Robin M.; Moller, Marlo; Kinnear, Craig
    The advent and evolution of next generation sequencing has considerably impacted genomic research. Until recently, South African researchers were unable to access affordable platforms capable of human whole genome sequencing locally and DNA samples had to be exported. Here we report the whole genome sequences of the first six human DNA samples sequenced and analysed at the South African Medical Research Council’s Genomics Centre. We demonstrate that the data obtained is of high quality, with an average sequencing depth of 36.41, and that the output is comparable to data generated internationally on a similar platform. The Genomics Centre creates an environment where African researchers are able to access world class facilities, increasing local capacity to sequence whole genomes as well as store and analyse the data.
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    Identification of novel DNA methylation signatures in the development of cardiovascular disease
    (2021) Jooste, Tracey; Johnson, Rabia; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Biomedical Sciences: Medical Physiology.
    ENGLISH ABSTRACT: Cardiovascular diseases (CVDs) remain the leading cause of death worldwide, claiming approximately 17.9 million lives annually. The last few decades have seen an exponential increase in the prevalence of major CVD risk factors, such as obesity, insulin resistance (IR), and type 2 diabetes mellitus (T2DM) in underdeveloped countries, including South Africa. This increase is concomitant to escalating CVD incidence and in part due to increased consumption of diets high in fat and sugar, and sedentary lifestyles. Several mechanisms have been implicated in the pathogenesis of diabetic-induced CVD and recently, increasing evidence suggests that dysregulation of the epigenome may play an important role in the development of T2DM and related cardiac complications. More specifically, aberrant DNA methylation has been extensively investigated and implicated in the development of chronic diseases such as obesity, T2DM and CVD. Nonetheless, it has been suggested that some active dietary compounds may reverse this epigenetic phenomenon which allows for the modification of the transcription of critical genes associated with the progression of CVD. Several plant-polyphenols have been reported to influence gene transcription by altering DNA methylation status. Thus, targeting DNA methylation could provide a promising approach for alternative prevention strategies. This study explored the gene expression networks activated during diet-induced CVD and the ability of a green rooibos extract, Afriplex GRT, to alter this consequence. In addition, the study aimed to evaluate aberrant DNA methylation associated with diet-induced CVD to further elucidate pathophysiology. To this end, cardiomyocytes exposed to high glucose and palmitate (HG + Pal) displayed a diminished antioxidant defence system, mitochondrial dysfunction and increased apoptosis, indicative of cardiac stress. Additionally, the combinatory treatment with HG + Pal induced transcriptional changes associated with inflammation, oxidative stress, altered lipid metabolism and increased contractile dysfunction, ultimately promoting the development of atherosclerosis and hypertrophic cardiomyopathy. Interestingly, post treatment with Afriplex GRT or Aspalathin had no significant effect on the metabolic and molecular derangements induced under HG + Pal stress. Similarly, RNA sequencing conducted on cardiac tissue of Wistar rats that received a high fat, high sugar (HFHS) diet, revealed the downregulation of differentially expressed genes (DEGs) involved in host antioxidant activity and inflammatory response, accompanied with an increase in hypertrophic gene expression possibly affecting cardiac muscle functionality. Supplementation with Afriplex GRT™ yielded no high confidence results for the amelioration of the transcriptomic signatures resulting from HFHS diet feeding. To profile DNA methylation throughout disease progression, cardiac tissue of male Wistar rats maintained on a HFHS diet were subjected to whole genome bisulfite sequencing (WGBS). The latter revealed aberrant DNA methylation of genes linked to the phagosome, platelet activation, toll-like receptor signalling and diabetic cardiomyopathy. Furthermore, hypomethylation within the intergenic and gene body regions of several differentially methylated genes (DMGs) overlapped with DEGs identified in the RNA sequencing analysis. Collectively these results demonstrate the ability of the HFHS diet to act as a pathological stimulus capable of inducing altered gene expression and DNA methylation associated with a heightened proinflammatory and lipid metabolism response that increased the risk of CVD development.