Decentralisation of the multidrug resistant tuberculosis (MDR TB) programme to primary health care (PHC) facilities in the Nelson Mandela Metropolitan was implemented in order to improve the effectiveness of MDR TB services. This study explored the knowledge gaps of nurses at PHC facilities as regards MDR TB. A quantitative, cross-sectional descriptive study was conducted; data was collected using a structured questionnaire. Non-probability sampling was applied in this study. A convenient sampling technique was used and 25 of the 42 facilities were selected. Thirty-two respondents completed the questionnaire with a response rate of 64%. Descriptive statistics were used to describe the data. Only 38% of the nurses had been trained on MDR TB. Overall scores were high with a mean knowledge score of 61%. However there were knowledge gaps regarding side effects of MDR TB medication. This study revealed gaps in knowledge of certain areas of MDR TB management / Health Studies / M.A. (Public Health)

Magister Pharmaceuticae - MPharm / Many studies have reported low cure rates for multidrug-resistant tuberculosis (MDRTB) patients and MDR-TB patients co-infected with human immunodeficiency virus (HIV). However, little is known about the effect of HIV infection and antiretroviral therapy on the treatment outcomes of MDR-TB in South Africa. Therefore, the objectives of the study are: to find out whether HIV infection and interactions between ARVs and second line anti-TB drugs have an impact on the following MDR-TB treatment outcomes: cure rate and treatment failure at Brewelskloof Hospital. MDR-TB patients were treated for 18-24 months. The study was designed as a case-control retrospective study comparing MDR-TB treatment outcomes between HIV positive (cases) and HIV negative patients (controls). Patients were included in the study only if they complied with the following criteria: sensitivity to second line anti-TB drugs, MDR-TB infection, co-infection with HIV (for some of them), male and female patients, completion of treatment between 1 January 2006 and 31 December 2008. Any patients that presented with extreme drug-resistant tuberculosis (XDR-TB) were excluded from the study. Data were retrospectively collected from each patient’s medical records. There were a total of 336 patients of which 242 (72%) were MDR-TB patients and 94 (27.9%) MDRTB co-infected with HIV patients. Out of the 242 MDR-TB patients, 167 (69.2%) were males and 75 (30.7%) were females. Of the 94 patients with MDR-TB co-infected with HIV, 51 (54.2%) males and 43 (45.7%) females. Patients with multidrug-resistant tuberculosis co-infected with HIV who qualify for antiretroviral therapy were treated with stavudine, lamivudine and efavirenz while all MDR-TB patients were given kanamycin, ethionamide, ofloxacin, cycloserine and pyrazinamide. The cure rate of MDR-TB in HIV (+) patients and in HIV (-) patients is 34.5% and 30 % respectively. There is no significant difference between both artes (pvalue = 0.80). The MDR-TB cure rate in HIV (+) patients taking antiretroviral drugs and in HIV (+) patients without antiretroviral therapy is 35% and 33% respectively. The difference between both rates is not statistically significant. The study shows that 65 (28.0%) patients completed MDR-TB treatment but could not be classified as cured or failure, 29 (12.5%) patients failed, 76 (32.7%) defaulted, 18 (7.7%) were transferred out and 44 (18.9%) died. As far as treatment completed and defaulted is concerned, there is no significant statistical difference between HIV (+) and HIV (-) The number of patients who failed the MDR-TB treatment and who were transferred out is significantly higher in the HIV (-) group than in the HIV (+) group. Finally the number of MDR-TB patients who died is significantly higher in the HIV (+) group). The median (range) duration of antiretroviral therapy before starting anti-tuberculosis drugs is 10.5 (1-60) months. According to this study results, the MDR-TB treatment cure rate at Brewelkloof hospital is similar to the cure rate at the national level. The study also hows that HIV infection and antiretroviral drugs do not influence any influence on MDR-TB treatment outcomes. / South Africa

Magister Pharmaceuticae - MPharm / The aim of the study was to determine firstly, kanamycin plasma concentrations using liquid chromatography coupled with mass spectrometry (LC/MS); secondly, to investigate the PK parameters of kanamycin in patients infected with MDR-TB and in patients co-infected with MDR-TB and HIV; thirdly, to assess the influence of HIV infection and renal impairment on the PK of kanamycin and fourthly, to find out whether there is any interaction between antiretroviral drugs and kanamycin.

The emergence and spread of multi-drug resistant (MDR-TB) and extensively drugresistant tuberculosis (XDR-TB) are a major medical and public problem threatening the global health. The objectives of this study were to (i) determine the prevalence of MDR-TB and XDR-TB in the Eastern Cape; (ii) analyze patterns of gene mutations in MDR-TB and (iii) identify gene mutations associated with resistance to second line injectable drugs in XDR-TB isolates. A total of 1520 routine sputum specimens sequentially received within a period of 12 months i.e. February 2012 to February 2013 from all MDR-TB and XDR-TB patients treated by Hospitals and clinics in the Eastern Cape were included in this study, of which 1004 had interpretable results. Samples were analyzed with the Genotype MTBDRplus VER 2.0 assay kit (Hain Lifescience) for detection of resistance to Rifampicin and Isoniazid while solid and liquid culture drug susceptibility tests were used for ethambutol, streptomycin, ethionamide, ofloxacin, capreomycin and amikacin. PCR and sequence analysis of short regions of target genes gyrA, (encode subunit of DNA topoisomerase gyrase), rrs (16S rRNA) and tlyA (encodes a 2’-O-methyltransferase) were performed on 20 XDR-TB isolates. MTBDRplus kit results and drug susceptibility tests identified 462 MDR-TB, 284 pre-XDR and 258 XDR-TB isolates from 267 clinics and 25 hospitals in the Eastern Cape. There was a high frequency of resistance to streptomycin, ethionamide, amikacin, ofloxacin and capreomycin. Mutation patterns indicated differences between the health districts as well as differences between the facilities within the health districts. The most common mutation patterns observed were: (i) ΔWT3, ΔWT4, MUT1 [D516V+del515] (rpoB), ΔWT, MUT1 [S315T1] (katG), ΔWT1 [C15T] (inhA) [39 MDR, 204 XDR-TB and 214 pre XDR-TB isolates], (ii) ΔWT8, MUT3 [L533P+S531L] (rpoB), ΔWT, MUT1 [S315T1] [145 MDR, 18 pre-XDR and 3 XDR-TB solates] and (iii) ΔWT3, WT4 [D516Y+del515] (rpoB), ΔWT, MUT1 [S315T1] (katG) [75 MDR, 1 pre-XDR and 7 XDR-TB isolates]. Mutations in inhA promoter regions were strongly associated with XDR-TB isolates. Two thirds (66.6 percent (669/1004) of the isolates had inhA mutations present with 25.4 percent (170/669) found among the MDR isolates, 39.2 percent (262/669) among the pre-XDR isolates and 35.4 percent (237/669) among the XDR-TB isolates, which implies that these resistant isolates are being spread by transmission within the community and circulating in the province. There was good correlation between XDR-TB drug susceptibility test results and sequence analyses of the gyrA and rrs genes. The majority of XDR-TB isolates contained mutations at positions C269T (6/20) and 1401G (18/20) in gyrA and rrs genes respectively. Sequence analysis of short regions of gyrA and rrs genes may be useful for detection of fluoroquinolone and amikacin/ kanamycin resistance in XDR-TB isolates but the tlyA gene is not a sensitive genetic marker for capreomycin resistance. This study highlighted the urgent need for the development of rapid diagnostics for XDR-TB and raised serious concerns regarding ineffective patientmanagement resulting in ongoing transmission of extremely resistant strains of XDRTB in the Eastern Cape suggesting that the Eastern Cape could be fast becoming the epicenter for the development of Totally Drug-resistant Tuberculosis (TDR-TB) in South Africa.

The aim of this study was to assess community members’ knowledge levels regarding Drug Resistant TB and HIV and AIDS. The study was conducted at ward 40 in Green bushes area in Nelson Mandela Metropolitan Municipality (NMMM). A quantitative research method was used in this study. Random sampling is the type of probability sampling method that was used in this study. The sample consisted of 100 respondents above 18 years who were randomly selected from the beneficiary list of for the RDP houses in Green bushes area in the Nelson Mandela Metropolitan Municipality. Data for this study were collected using close ended questions which were administered by the researcher to the selected participants. Data was analysed using bivariate and descriptive statistics according to the identified themes. The study revealed that community members had high knowledge levels regarding Drug Resistant TB and HIV and AIDS prevention, transmission modes and diagnosis and treatment methods. The findings revealed that community members were highly knowledgeable and aware of the fact that abstaining and practising safe sex were means of preventing the spread of HIV and AIDS as it was spread through unprotected sex, while opening of windows and minimisation of close contact with HIV positive people and children with people infected with Drug Resistant TB are infection control measures or methods of preventing the spread of the disease. Additionally, the study indicated that female respondents were more aware and knowledgeable about prevention, transmission modes and diagnosis and treatment of Drug Resistant TB and HIV and AIDS than male respondents. Furthermore, the findings revealed that the respondents were highly knowledgeable and aware about transmission of Drug Resistant TB and HIV and AIDS; knowledgeable about prevention and less knowledgeable about diagnosis and treatment. A high percentage of female respondents knew that there was no vaccine to neither prevent nor cure HIV and AIDS and that antiretroviral drug were used to manage it. The study also showed that female respondents knew that all people irrespective of race and economic class can be infected with Drug Resistant TB and HIV and AIDS. It is important to note that the respondents between 41-60 years possessed more knowledge regarding Drug Resistant TB and HIV and AIDS than the respondents who were between 18-40 years. Lastly, the study showed that there were significant differences in gender and knowledge and no significant differences in age and knowledge of the respondents regarding Drug Resistant TB and HIV and AIDS. It is recommended that in future, research regarding knowledge levels about Drug Resistant TB and HIV and AIDS be extended to other wards in the Nelson Mandela Metropolitan Municipality (NMMM). Accurate knowledge should be provided by ensuring that educational materials that are developed, are appropriate for the various levels of literacy, and that more appropriate and relevant information regarding these diseases is made more accessible to community members in their home languages. The researcher further recommends that during training interventions and educational campaigns more emphasis should be put on prevention, diagnosis and treatment of Drug Resistant TB and HIV and AIDS.

Thesis (MSc. (Medical Sciences)) -- University of Limpopo, 2015 / Introduction: Multidrug-resistant tuberculosis (MDR-TB) caused by resistance to at least rifampicin (RIF) and isoniazid (INH) drugs is a growing public health concern in South Africa. The detection of MDR-TB still relies on culture despite advancement in molecular diagnostic technology. Currently MTBDRplus and GeneXpert are the only available assays used in rapid diagnosis of MDR-TB using chromosomal mutations in drug target regions. Some strains are missed by these assays due to their limitation in mutational detection profile. Novel Seegene Anyplex assays simultaneously detect TB and resistance to RIF and INH using fifteen and six mutational probes, respectively within 3 hours. Limpopo Province has limited information on the circulating strains of TB. Aim: To determine drug-resistant Mycobacterium tuberculosis (M. tuberculosis) mutations using Anyplex™ MTB/NTM/MDR-TB real time assay and characterise the drug-resistant strains. Methods: We prospectively collected 204 clinical samples at Modimolle MDR-TB unit and retrospectively used 104 culture isolates from MRC laboratory in Pretoria. The MTBDRplus assay was used to screen for M. tuberculosis and drug resistant mutations to RIF and INH drugs. Anyplex™ MTB/NTM/MDR-TB assay was used for rapid detection of M. tuberculosis and drug resistance to RIF and INH within 3 hours. The discordance between phenotypic and genotypic assays was resolved by sequencing and the Anyplex™ resistant profiles were spoligotyped. Diagnostic data was collected from NHLS and MRC databases and analysed using the Microsoft excel and Epi Info version 3.5. Descriptive statistics (percentages and frequencies) were used to explain proportions. Results: The Anyplex™ MTB/NTM assay detected M. tuberculosis in 69/111(62%) and 100/104 (96%) of clinical and culture samples respectively. The sensitivities, specificity, PPV and NPV obtained for both RIF and INH resistance by Anyplex™ MDR-TB assay were 67%, 59%, 67%, 55% and 15%, 100%, 100% and 17%, respectively. Anyplex™ MTB/NTM/MDR-TB resolved 23/45 (51%) of discordant vi samples. Sequencing of remaining discordant isolates revealed L511P, L533P and D516Y mutations within rpoB gene. A novel R385W mutation within katG was also detected. Spoligotyping of Anyplex™ MDR-TB resistant clinical isolates revealed Euro American clade with 20% followed by 15% Manu2, 5% East African Indian, 5% H37Rv, 5% atypical and 50% were orphans. Conclusion: The novel Anyplex™ MTB/NTM/MDR-TB assay is a rapid and valid technique for detecting M. tuberculosis and most common mutations conferring resistance to RIF and INH. However further investigations are required, as the assay has a lower sensitivity as compared to already endorsed techniques. / National Research Foundation (NRF) and University of Limpopo TB Grant

Thesis (M. Sc. (Chemistry)) -- University of Limpopo, 2021 / Tuberculosis is one of the top 10 leading causes of death in the world. The development of drug resistant strains of Mycobacterium tuberculosis such as Multidrug resistant (MDR) and Extensively drug resistant (XDR) strains further complicate the TB control. Medicinal plants present a possible source for new potential antitubercular drugs. They have played an important role in drug discovery, with many pharmaceutical products originating from them. Isolation and characterisation of new antitubercular compounds from plant extracts is relevant today because of the development of resistant strains. The aim of the study was to evaluate the antimycobacterial activity of the leave extracts of Croton gratissimus and Leonotis leonorus. The first step was to extract fine powder leaves of the two plant species using four (dichloromethane, acetone, hexane and ethanol/water) different solvent systems. Isolation of the fractions was done using column chromatography and preparative thin layer chromatography. Minimum inhibitory concentrations were determined using the broth dilution method and the values were recorded in μg/mL. All the isolated fractions from both plant species were evaluated for preliminary in-vitro antimycobacterial activity. Some of the isolated fractions showed an increased activity against the pathogen as compared to the crude extracts. All the crude extracts of the two plants had activity with MIC90 values greater than 125 μg/mL. Seven fractions obtained from Croton gratissimus showed potential activity against the pathogen with MIC90 values ranging from 30.61 to 64.88 μg/mL. Leonotis leonurus had three fractions with promising activity with MICs ranging from 1.963 to 62.51 μg/mL. The crude extracts of the two plant species showed that the two plant species have antioxidant properties. The qualitative antioxidant assay showed that DCM crude extracts had more antioxidants than all other extracts because of more clear zones against the purple background colour on the TLC plates. These was confirmed by the qualitative antioxidant assay where DCM crude extracts was able to inhibit the highest percentage of DPPH at different concentrations than all other solvent extracts. The DCM crude extracts of L. leonurus and Croton gratissimus inhibited 87 and 93 % of DPPH respectively at 250 μg/mL. The structures of the compounds within the isolated fractions were elucidated using NMR and confirmed by MS and FTIR spectroscopies. The NMR data showed that the isolated fractions were not pure compounds but mixtures of closely related compounds. The compounds whose structures were elucidated included two labdane diterpenoids (Croton A and Croton B) and a Cembranolide ((5E,10E,13R)-4-isopropyl-7,11-dimethyl-15-oxo-14-oxa-bicyclo [11.2.1] hexadeca-5,10-dien-7-yl acetate) from Croton gratissimus and a phenol (4-(3,3,4,4-tetramethylheptyl) benzene-1,2-diol)) from Leonotis leonurus. / National Research Foundation (NRF) and Sasol Foundation

Tuberculosis (TB) is a leading public health problem across the world. For various reasons, TB and multidrug-resistant tuberculosis (MDR-TB) have increased. Clarification on TB/HIV co-infection and homelessness as risk factors for TB and MDR-TB is required to inform policy interventions to reduce TB-related morbidity, mortality, and healthcare costs. In this quantitative study, data from the Houston Health Department (N = 341) were analyzed to explore the relationship between TB and MDR-TB outcomes and TB/HIV co-infection and type of housing/homelessness. Foreign-born persons are disproportionately affected in the United States. The socio-ecological model provided a theoretical framework for the investigation. Multiple and logistic regression analyses were conducted to investigate the relationships between variables, controlling for age and gender. Results indicate that HIV infected persons were more likely than non-infected persons to contract TB, and homeless persons were more likely than non-homeless persons to contract TB/MDR-TB, suggesting that high TB/HIV co-infection rates increase prevalence of TB and MDR-TB while improvements in housing reduce prevalence of TB and MDR-TB. However, no significant associations between variables were found. The odds ratio, Exp(B) = 0.000, p -?¥ 0.90, 95% Cl [0.000, with no upper bound values] was observed for both independent variables. Regular screening for TB/HIV co-infection among persons with high TB and MDR-TB risk profiles is recommended. Further investigation is required. Inclusion of more covariates could further elucidate more evidence of an association between variables. Study findings may support interventions to reduce TB-related morbidity, leading to positive social change.

Yes / Mutations in the gyrase genes cause fluoroquinolone resistance in Mycobacterium tuberculosis. However, the predictive value of these markers for clinical outcomes in patients with MDR-TB is unknown to date. The objective of this study was to determine molecular markers and breakpoints predicting second-line treatment outcomes in M. tuberculosis patients treated with fourth-generation fluoroquinolones. We analysed treatment outcome data in relation to the gyrA and gyrB sequences and MICs of ofloxacin, gatifloxacin and moxifloxacin for pretreatment M. tuberculosis isolates from 181 MDR-TB patients in Bangladesh whose isolates were susceptible to injectable drugs. The gyrA 90Val, 94Gly and 94Ala mutations were most frequent, with the highest resistance levels for 94Gly mutants. Increased pretreatment resistance levels (>2 mg/L), related to specific mutations, were associated with lower cure percentages, with no cure in patients whose isolates were resistant to gatifloxacin at 4 mg/L. Any gyrA 94 mutation, except 94Ala, predicted a significantly lower proportion of cure compared with all other gyrA mutations taken together (all non-94 mutants + 94Ala) [OR = 4.3 (95% CI 1.4-13.0)]. The difference in treatment outcome was not explained by resistance to the other drugs. Our study suggests that gyrA mutations at position 94, other than Ala, predict high-level resistance to gatifloxacin and moxifloxacin, as well as poor treatment outcome, in MDR-TB patients in whom an injectable agent is still effective.

Tuberculosis (TB) is the world’s most deadly infectious disease and causes enormous public health problems. The comorbidity with HIV and the rise of multidrug-resistant TB strains impede successful therapy through drug-drug interactions and the lack of efficient second-line treatments. The aim of this thesis was to support the improvement of anti-TB therapy through development of pharmacometric models, specifically focusing on the novel drug bedaquiline, pharmacokinetic interactions and methods for pooled population analyses. A population pharmacokinetic model of bedaquiline and its metabolite M2, linked to semi-mechanistic models of body weight and albumin concentrations, was developed and used for exposure-response analysis. Treatment response was quantified by measurements of mycobacterial load and early bedaquiline exposure was found to significantly impact the half-life of bacterial clearance. The analysis represents the first successful characterization of a concentration-effect relationship for bedaquiline. Single-dose Phase I studies investigating potential interactions between bedaquiline and efavirenz, nevirapine, ritonavir-boosted lopinavir, rifampicin and rifapentine were analyzed with a model-based approach. Substantial effects were detected in several cases and dose-adjustments mitigating the impact were suggested after simulations. The interaction effects of nevirapine and ritonavir-boosted lopinavir were also confirmed in patients with multidrug-resistant TB on long-term treatment combining the antiretrovirals and bedaquiline. Furthermore, the outcomes from model-based analysis were compared to results from conventional non-compartmental analysis in a simulation study. Non-compartmental analysis was found to consistently underpredict the interaction effect when most of the concentration-time profile was not observed, as commonly is the case for compounds with very long terminal half-life such as bedaquiline. To facilitate pooled analyses of individual patient data from multiple sources a structured development procedure was outlined and a fast diagnostic tool for extensions of the stochastic model components was developed. Pooled analyses of nevirapine and rifabutin pharmacokinetics were performed; the latter generating comprehensive dosing recommendations for combined administration of rifabutin and antiretroviral protease inhibitors. The work presented in this thesis demonstrates the usefulness of pharmacometric techniques to improve treatment of TB and especially contributes evidence to inform optimized dosing regimens of new and old anti-TB drugs in various clinical contexts.