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Browsing Theses and Dissertations by Author "Bessong, P. O."
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Item Open Access Detection of Cryptosporidium species in stools of HIV/AIDS patients in Bela-Bela, South Africa(2010-06) Makuwa, Stenly Modupi; Bessong, P. O.; Samie, A.; Potgieter, N.See the attached abstract belowItem Open Access Drug Resistance Mutations in Naive HIV-1 South African Patients, and Construction of Molecular Clones to Phenotype Putative Resistance Mutations(2009-03) Mavhandu, Lufuno Grace; Bessong, P. O.; Rekosh, David; Hammarskjold, Marie-LouiseIn countries such as South Africa where access to therapy is progressing data is required on patterns of resistance and evolution of resistance. Thirty protease (PR) and 31 reverse transcriptase (RT) amino acid sequences of HIV primary isolates from drug naNe patients from rural settings in South Africa were examined for resistance mutations. Samples were collected between May and August 2007. Phylogenetic analysis showed that all the sequences were HIV-1 subtype C in both the protease and reverse transcriptase genes. The mean genetic distances among the sequences were 0.0170-0.0786 for the protease, and 0.0045-0.0890 for the reverse transcriptase. However, it was noted that 3 pairs of samples 07VGNF5ZA and 07VGNF6ZA, 07VGNF7ZA and 07VGNF8ZA, 07VGNF10ZA and 07VGNF13ZA did not show any genetic variability among their protease sequences. No major resistance mutation was observed among the protease sequences. However, the following minor resistance mutations were noted: L101N (3/30), A71T (1/30), and T74S (2/30). Examination of the reverse transcriptase gene for resistance mutations reveal the presence of V118I (1/30), V179D (1/30), K103N (2/30). Most of the RT sequences were wild-type, although V118I (3.3%) and k103N (6.7%) associated with resistance to lamivudine and nevirapine, respectively, were observed. In summary, this study has shown that most of the viruses in Limpopo Province, representing the northeastern part of South Africa are HIV-1 subtype C, and that the prevalence of resistant mutations among the drug na"fve patients is still low. Although combination antiretroviral therapy has resulted in a considerable improvement in the treatment of human immunodeficiency virus type 1 (HIV-1) infection, the emergence of resistant virus is a significant obstacle to the effective management of HIV infection and AIDS. Systems to be used in the testing of phenotypic drug resistance and susceptibility are being developed. These may intimately be used in guiding therapy to improve long term suppression of HIV replication. Two proviral chimeric clones making use of pMJ4 and pNL4-3, and two vector plasmids which deletions of sequences encoding HIV-1 protease or reverse transcriptase were constructed for cloning of HIV-1 PCR products. Growth of constructs was monitored by p24 antigen production. Susceptibility to protease and reverse transcriptase inhibitors was measured by using resistance test vectors that contain a Luciferase indicator gene. Cells were co-transfected with packaging plasmids, pluc, and pEnv, resulting in the production of virus particles that were used to infect target cells. Luciferase activity was measured following a single round of replication. The chimeric constructs MJ4 carrying the NL4-3 Apal-Hpal cassette (MJ4/NL4-3) and NL4-3 carrying the MJ4 Apal-Hpal cassette (NL4-3/MJ4) were successfully developed as shown by restriction digestion analysis. Considering growth of the constructed chimeras NL4-3/MJ4 was better than MJ4/NL4-3 although not robustly. Good p24 production was obtained from all four gap-pol plasmids. MJ4/NL4-3 worked better in delivering luciferase to the target cells while NL4-3/ML4 appeared totally devoid of any infectivity. The vectors pCMVGagPol(MJ4)-RREr and pCMVGagPol(NL4.3)-RREr were created and both expressed the viral gag-pol protein. Viral inhibition test showed that the vectors can be inhibited by NRTI, NNRTI and Pl. Inhibition was seen in all drugs in different concentrations indicating that the system works. The results showed that vector systems constructed can be used to evaluate putative drug resistant mutations, coding for resistance to protease and reverse transcriptase inhibitors, detecte� in patient viruses. In addition, the system can also be used to evaluate candidate drugs and assist in the development of new drugs that are active against resistant HIV-1 virus.Item Open Access HIV co-infections with cytomegalovirus, hepatitis c virus and human papillomavirus in northern South Africa(2014-11-03) Rikhotso, Mikateko; Bessong, P. O.; Tebit, Denis.Item Open Access Immunodulation of inflammation in a murine pnemococcal sepsis model(2013-10-01) Musie, Mbulaheni Edgar; Bessong, P. O.; Scheld, M. W.Item Embargo Molecular characterization of entamoeba species and impact of host genetic polymorphisms on amoebiasis(2022-11-10) Davhana, Ndivhudzannyi Caroline; Samie, A.; Bessong, P. O.; ElBakri, A. M. K.Background: Enteric infections constitute a serious public health problem globally, especially in low and middle-income countries, particularly in areas of poor sanitation, low socio-economic conditions, inadequate water supply and poor hygiene practices. South Africa is one of the developing countries that has been significantly impacted by diarrheal infections, many of which are due to Entamoeba species. It is still not clear why some individuals once infected with E. histolytica develop clinical amoebiasis while others do not show any symptoms. It has been suggested that both the parasite and host factors play a significant role in the outcome of E. histolytica infection. However, this does not explain why only few infected individuals develop symptomatic diseases. This suggests that there are other factors to explain this transition. Tumor necrosis factor-α (TNF-α) is a multifunctional pro-inflammation cytokine which has been considered as one of the pathogenic factors for various diseases. Several studies have reported an association between TNF-α polymorphisms and inflammatory diseases. However, no study has been carried out on the association between polymorphisms in the promoter of the TNF-α gene and high risk of E. histolytica infections. Aim and objective of the study: The overall aim of the study was to determine the molecular characteristics of Entamoeba species in relation to the occurrence of diarrhea among children and determine the impact of host genetic polymorphism on the occurrence of amoebiasis. This aim was addressed by the following primary objectives: a. To investigate the prevalence, distribution and genetic diversity of Entamoeba species and other parasites circulating among children in the Northern part of South Africa. b. To investigate the genetic polymorphism of TNF-α gene promoter region in a cohort of children in Northern South Africa. c. To identify any association between TNF-α promoter gene polymorphism with diarrhea, vomiting, fever, acute lower respiratory infection, gender and malnutrition. d. To identify any potential association that may exist between TNF-α promoter gene polymorphism and parasitic infections. Brief methodology and results: This study was nested within the Madi project and Malnutrition and Enteric Diseases project (MAL-ED) South Africa and received approval from the Health, Safety and Research Ethics Committee of the University of Venda. The stool samples were from Madi project and the blood samples were from the MAL-ED project. A total of 394 stool samples were collected in selected household with children under the age of 5 years who were randomized to receive a silver-impregnated ceramic water, a silver-impregnated ceramic disk, a safe-storage water container, or no intervention, and were followed quarterly for two years from Dzimauli rural communities of South Africa in Limpopo province. All the stool samples were observed under a light microscope for the presence of intestinal parasites. In order to accurately detect Entamoeba species in all faecal samples, polymerase chain reaction (PCR) protocols were performed using genus-specific PCR primers based on small-subunit rRNA gene sequences for E. polecki, E. chattoni, E. dispar, E. histolytica, E. hartmanni and E. coli. Real-time PCR protocol was also used to detect and identify the specific Entamoeba species such as E. histolytica, E. dispar, E. bangladeshi and E. moshkovskii in order to gain information on their accurate prevalence, distribution and genetic diversity in the community. The present study reported an overall prevalence of intestinal parasites including Entamoeba species, Strongyloides stercoralis, Cystoisospora belli and Ascaris lumbricoides as determined by microscopy to be 140 (35.5%), 138 (35.0%), 114 (28.9%) and 78 (19.8%), respectively. The genus-specific PCR was positive for Entamoeba spp. in 140 (35.5%) samples. Real time PCR detected E. histolytica in about 4% of the samples while E. moshkovskii occurred in about 9% of the samples. Results identified by qPCR showed that children in silver-impregnated ceramic water filter group at month 12 had higher E. moshkovskii infection 6 (13.0%), while those in intervention group had lower E. moshkovskii infection 3 (6.8%). None of the participants were infected with Entamoeba bangladeshi. Sequence analysis showed a wide variety of Entamoeba species with E. coli and E. polecki appearing to be the most common organisms followed by E. moshkovskii and E. dispar. Further studies using next generation sequencing technologies are needed to understand the real importance of each of these organisms in the community in terms of the pathogenesis of amoebiasis (Chapter 3). TNF-α is a multi-functional pro-inflammatory cytokine and a primary mediator involved in the early phase of the cytokine cascade and plays an important role in the initiation and regulation of the immune response. The present study aimed to investigate the genetic polymorphism of tumor necrosis factor-α (TNF-α) gene promoter region in a cohort of children in Northern South Africa. A total of 199 blood samples were evaluated from children who were part of the MAL-ED study cohort. The TNF-α gene at positions ‑1031T/C and ‑308G/A were genotyped by polymerase chain reaction–restriction fragment length polymorphism (PCR–RFLP) assay and confirmed by DNA sequencing. Out of these 199 participants, 94(47.2%) were males and 105(52.8%) were females. Of all the children, 23(11.6%) had low birth weight. A strong association was noted between the CC homozygous genotype at position -1031 and children with diarrhea (P=0.043, OR=4.167, 95% CT=0.942-18.43); whereas TC heterozygous genotype was significantly common in healthy children with no diarrhea (P=0.019, OR=0.446, 95% CT=0.226- 0.882). The T-allele was significantly common in children with diarrhea (P=0.043, OR=0.240, 95% CT=0.054-1.062). A strong association was also noted between the TT homozygous genotype at position -1031 and children with dehydration (P=0.014, 95% CI= 1.224-12.443). A strong association was noted between the GA heterozygous genotype at position -308 and children with diarrhea (P=0.040, OR=2.579, 95% CT=1.019-6.528); while AA homozygous genotype was significantly common in healthy children with no diarrhea (P=0.012, OR=0.3420, 95% CT=143-0.815). Heterozygous GA genotype was more common among healthy children with no dehydration and the result was statistically insignificant (P = 0.894, X2 = 0.018, OR = 1.095, 95% CT = 0.288-4.168); while a strong association was also noted between the heterozygous GA genotype at position -308 and children with vomiting (P=0.019, OR=2.694, 95% CT=1.160-6.256). The G allele was significantly common in children with vomiting (P=0.010, OR=2.816, 95% CT=1.263-6.279). Our study has for the first time revealed that the -1031(T/C) polymorphism of TNF-α promotor gene is associated with diarrhea, dehydration and acute lower respiratory infection among children at five years of age, while the -308(G/A) polymorphism was associated with diarrhea and vomiting among these children (Chapter 4). Intestinal parasitic diseases are common in developing countries including South Africa and have been documented to be the most common in children under the age of five. The present study aimed to identify any potential association that may exist between TNF-α promoter gene polymorphism and parasitic infections. A total of 199 blood samples was evaluated from children who were part of the MAL-ED study cohort. The DNA was used to investigate polymorphism in the promoter region of the TNF-α gene at position 1031T/C. The polymorphisms were detected by polymerase chain reaction–restriction fragment length polymorphism (PCR–RFLP) assay. The TC genotype at position -1031 was significantly higher in healthy controls children than in children who were infected with Entamoeba species (59.9% vs 29.4%, P = 0.015) and Entamoeba coli (59.1% vs 30.8%, P = 0.046), indicating that TC genotype may be protective against Entamoeba infections and Entamoeba coli infections. The CC genotype at position -1031 was more common among children with parasite and diarrhea and the results was statistically significant (P = 0.04). This study has revealed that the CC genotype may be is a risk factor for symptomatic parasitic infections, while the TC genotype might be protective of Entamoeba infections among children in Dzimauli community (Chapter 5). Overall conclusion: The present study demonstrated new data on the prevalence, distribution and genetic diversity of Entamoeba species and other parasites in Northern South Africa. Several Entamoeba species are circulating in the region and the importance of E. moshkovskii in the pathogenesis of amoebiasis seems to be underestimated. This study also revealed for the first time that the -1031(T/C) polymorphism of TNF-α promotor gene is associated with diarrhea and acute lower respiratory infection, Entamoeba species and Cyclospora infections among the children in Dzimauli community, while - 308(G/A) was associated with vomiting and overall illness. Information generated from this study will be useful for understanding the transmission, source of infection and clinical outcome of infection with parasitic infections. However, larger studies need to be conducted in order to confirm these findings.Item Open Access Molecular detection of norovirus GI ang GII genotypes in children less than two years of age and impact on child growth(2014-11-03) Moloro, Glenton Thabo; Samie, A.; Bessong, P. O.Item Open Access Molecular diagnosis and characterization of clinical isolates of entamoeba histolytica, giadia lamblia and cyptosporidium species from the United Arab Emirates and South Africa(2014-11-03) ElBakri, Ali Mohammed Kamal; Bessong, P. O.; Potgieter, N.; AbuOdeh, R. OItem Open Access Screening of herbal preparation (Pheko) for anti HIV-1 replication properties(2015-01-14) Matume, Nontokozo Daphney; Bessong, P. O.; Sekhoacha, M.Item Open Access Serologic markers and molecular pidemiology of HBV in an HIV infected cohort from Cameroon(2016-05) Magoro, Tshifhiwa; Bessong, P. O.; Mavhandu, Lufuno G.; Masebe, Tracy M.See the attached abstract below