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PhD (Dietetics), North-West University, Potchefstroom Campus Background: Non-resolving inflammation is characteristic of tuberculosis (TB). This leads to lung tissue damage and anaemia of infection, which is associated with poor clinical outcomes. Iron supplementation may have limited efficacy and may favour bacterial growth. Therefore, anti-inflammatory and pro-resolving host-directed therapy (HDT) have been suggested. Omega-3 long-chain polyunsaturated fatty acids (n-3 LCPUFAs) including eicosapentaenoic and docosahexaenoic acid (EPA/DHA) may provide such a nutritional approach for HDT. Aim: The aim of this thesis was to determine the effects of EPA/DHA and iron supplementation, alone and in combination, on inflammatory and clinical outcomes in Mycobacterium tuberculosis (Mtb)-infected mice and whether these effects were dependent on n-3 polyunsaturated (n-3 PUFA) status prior to infection. Methods: Male C3HeB/FeJ mice were conditioned on n-3 PUFA sufficient or deficient diets for six weeks prior to infection. One week post Mtb infection, n-3 PUFA sufficient mice were randomised to 1) continue on the n-3 PUFA sufficient diet, or were switched to an 2) EPA/DHA-supplemented, 3) iron-supplemented, or 4) EPA/DHA and iron-supplemented diet. Mice conditioned on an n-3 PUFA deficient diet were randomised to 1) continue on the n-3 PUFA-deficient diet, or switched to the 2) n-3 PUFA sufficient, or 3) EPA/DHA-supplemented diets. The mice received these diets for three weeks until euthanasia. Results: The phospholipid fatty acid composition of cell membranes of mice reflected the dietary fatty acid content. Pro-resolving lung lipid mediator profiles were found in the EPA/DHA-supplemented groups in both n-3 PUFA sufficient and low-status arms, and also when combined with iron. Additionally, EPA/DHA supplementation resulted in lower bacterial loads and lung pathology in the sufficient, but not in the low-status group. Iron and EPA/DHA supplementation both individually lowered systemic and lung cytokines, together with improved anaemia of infection markers. However, whilst EPA/DHA supplementation lowered lung T cells, iron resulted in higher lung immune cell counts. Iron also had no effect on lung pathology or bacterial load but lowered body weight gain. There were iron x EPA/DHA interactions to attenuate the lowering effects of iron or EPA/DHA on anaemia of infection, and of iron for higher immune cell counts. Furthermore, compared with the n-3 PUFA sufficient diet, EPA/DHA supplementation provided superior benefits in body weight gain, bacterial load, and lung inflammation in low-status mice. Conclusions: The findings of this thesis showed that EPA/DHA supplementation, after the initial inflammatory response, has antibacterial and inflammation-resolving benefits and improves markers of anaemia of infection in TB, depending on n-3 PUFA status. On the other hand, iron promotes anti-inflammatory effects and improves markers of anaemia, but enhances immune cell recruitment and lowers body weight gain. Providing combination iron and EPA/DHA treatment attenuates their individual beneficial effects. Lastly, in low-status mice, EPA/DHA provides superior effects compared with an n-3 PUFA sufficient diet. Considering this, iron may not be detrimental concerning worsening the bacterial burden in TB, but moreover, n-3 LCPUFA therapy may be a promising approach as HDT in TB. Doctoral

Background Nutritional treatment in the intensive care unit (ICU) has evolved from meeting nutritional requirements to manipulating patient outcome. Pharmaconutrition, referring to nutrients that are applied for their pharmacological properties, forms part of the standard nutritional care plan. The most abundant amino acid in the body, glutamine, is also the most-researched pharmaconutrient. It is an independent predictor of mortality in ICU patients, at both deficient and very high levels. Glutamine supplementation is recommended in the ICU setting for its proven outcome benefits. However, recent data showed that glutamine supplementation increases mortality risk in certain patient groups. Moreover, it suggested that not all ICU patients are glutamine deficient. Therefore, the main aim of this study was to investigate the plasma glutamine levels of adult ICU patients, on admission to the ICU. In addition, to elucidate the profile of ICU patients that can be expected to present with a glutamine deficiency or excess, with regards to gender, diagnosis and inflammatory markers. Methods In this observational, cross-sectional study, 60 mixed ICU adult patients admitted to two hospitals in the North West province were included in the study group. Blood sampling was conducted within 24 hours following ICU admission, to determine plasma glutamine, interleukin (IL)-6 and C-reactive protein (CRP) levels. Plasma glutamine levels were compared with those of a control group of healthy individuals, matched by age, race, and gender. Gender-related differences in plasma glutamine levels were investigated, as well as differences between patients with various medical conditions. The relationship between plasma glutamine levels and IL-6 or CRP was examined. Additionally, a CRP concentration cut-off point at which glutamine becomes deficient was determined by means of a receiver operating characteristic (ROC) curve. Results and discussion Intensive care unit patients had significantly lower plasma glutamine levels than healthy individuals on day one of ICU admission (p < 0.0001). However, only 38.3% (n = 23) had deficient plasma glutamine levels (< 420 μmol/L), while 6.7% (n = 4) presented with supra-normal levels (> 930 μmol/L). No significant difference could be detected between the plasma glutamine levels of male and female ICU patients (p = 0.116). Likewise, levels between diagnosis categories were also not significantly different (p = 0.325). There was a significant inverse association between plasma glutamine levels and CRP concentrations (r = -0.44, p < 0.05), and a trend towards an inverse association with IL-6 (r = - 0.23, p = 0.08). A CRP cut-off value of 95.5 mg/L was determined, above which plasma glutamine values became deficient; however, more research is needed to confirm this result. Conclusion and recommendations This research therefore showed that ICU patients, when compared with healthy individuals, had lower plasma glutamine levels on day one of admission to the ICU. However, not all were glutamine deficient, as the majority had normal and some presented with supra-normal plasma glutamine levels. An individualised approach should therefore be followed in identifying candidates for glutamine supplementation. The patients‟ condition alone may not be sufficient to predict glutamine status, but an association between plasma glutamine levels and CRP was firmly established, as well as a cut- off CRP-value above which glutamine can be expected to become deficient, which could be of use in this regard. MSc (Dietetics), North-West University, Potchefstroom Campus, 2015

MSc (Nutrition), North-West University, Potchefstroom Campus Introduction - Globally, hypertensive disorders affect 10% of pregnancies and are the most common risk factors for maternal and perinatal morbidity and mortality. Between 2011 and 2013, 14.8 % of maternal mortalities were associated with hypertensive disorders in South Africa. Adequate intake and status of n-3 long chain polyunsaturated fatty acids (LCPUFA) have been linked to improved endothelial function and reduced blood pressure. However, the South African longitudinal Prospective Urban Rural Epidemiology (PURE) study has shown a positive association between plasma n-3 LCPUFA and hypertension and a negative association between n-6 LCPUFA and hypertension. In addition, in South Africa, pregnant women are supplemented routinely with 60 mg/d of elemental iron to prevent iron deficiency. This supplementation programme may lead to oxidative stress and increased inflammation, especially in women with sufficient iron status, increasing their risk to develop gestational hypertension. This study assessed the relationship of n-3 and n-6 LCPUFA status with blood pressure in pregnant African women receiving routine iron supplementation. Methods - This was a longitudinal analysis where 250 pregnant women were recruited at

MSc (Dietetics), North-West University, Potchefstroom Campus Background: Tuberculosis (TB) bacilli successfully survive within the host. It causes an immune response that is defensive but also harmful to the host’s lung tissue, because of perpetuating inflammation. Non-resolving inflammation in TB persists and can occur in both latent and active TB. Omega-3 polyunsaturated fatty acids (n-3 PUFA) have anti-inflammatory and inflammation-resolving activity due to their conversion to lipid mediators and influence on inflammatory cytokines. Aim: This project aimed to determine the effect of n-3 PUFA status prior to infection on lung inflammation and pathology in Mycobacterium tuberculosis (Mtb)-infected C3HeB/FeJ mice. Methods: Uninfected mice (n=20) were conditioned on either n-3 PUFA sufficient (n-3FAS) or deficient (n-3FAD) diets for six weeks and continued these diets post Mtb infection for an additional four weeks until euthanasia. Lung bacterial load, lung- and spleen-weight indexes and lung histology (free alveolar space), cytokines and lipid mediators were assessed. Results: The low n-3 PUFA status group (n-3FAD) had a trend to present with lower bacterial loads (p=0.095) and presented with lower spleen-weight indexes (p=0.041), more free alveolar space (p

MSc (Dietetics), North-West University, Potchefstroom Campus Background: Tuberculosis (TB) is currently a major health problem worldwide, and despite improvement, mortality rates are not ideal. Since the immune response, underlying protection against TB is incompletely understood, long durations of treatment time and poor health outcomes remain a problem. Therefore, host directed non-pharmaceutical interventions, supporting TB treatment, may be a promising approach to improve outcomes. Previous studies have found anti-inflammatory treatment to improve TB outcomes and the anti-inflammatory and pro-resolving properties of omega-3 (n-3) polyunsaturated fatty acids (PUFAs) treatment have been proven beneficial in other inflammatory diseases. Due to the fact that TB is known to be a disease of non-resolving inflammation leading to host tissue damage and poor clinical outcomes, the main aim of this study was to investigate whether sufficient and deficient n-3 PUFA status, after TB infection, had an influence on markers of morbidity, disease progression and anaemia of infection (AI). Methods: Eighteen 8 to 12-week-old C3HeB/FeJ mice were infected with TB via the intranasal route (high dose acute infection), and 12 mice via the aerosol route (low dose chronic infection) after conditioning for six weeks on an n-3 PUFA sufficient (FAS) or deficient (FAD) diet. Red blood cell (RBC) fatty acid status, whole blood haemoglobin (Hb), and body weight were determined the day before infection and markers of AI (Hb, plasma ferritin, transferrin receptor (TfR) and hepcidin), morbidity (appearance, respiratory rate, and body weight) and disease progression and clinical outcomes (lung bacillary load, organ indexes, and lung cytokine concentrations) were analyzed and compared between the two different groups 35 days after infection upon euthanization. Results: At euthanasia, intranasal and aerosol groups showed differences in PUFA composition with regards to all n-6 PUFA (all p < 0.001) and n-3 PUFAs (all p < 0.001, except docosahexaenoic acid (DHA) in aerosol subgroup p = 0.003) in RBC. Mice from the aerosol FAD subgroup had lower Hb concentrations prior to infection (p = 0.003), and a lower bacterial load (p = 0.008) and spleen-body-weight-index (p = 0.006) at 35 days post infection. In contrast, in the aerosol infected mice, the decrease in Hb in the n-3 FAS subgroup were more than in the n-3 FAD subgroup during the five weeks of TB infection (p = 0.027). No effects were found post-infection in other markers of AI. Inflammatory marker profiles showed a trend (p=0.061) towards lower pro-inflammatory cytokine IL-12 in the aerosol n-3 FAD subgroup. Conclusion: The lower-dose aerosol infection model was generally more sensitive to effects of n-3 PUFA sufficiency and deficiency. Without infection, n-3 PUFA sufficiency may contribute to improved Hb concentrations compared to n-3 PUFA deficiency. However, n-3 PUFA sufficiency may give rise to a bigger decrease in Hb concentrations after infection. Furthermore, contradicting to what was expected n-3 PUFA deficiency caused slower disease progression compared to an n-3 PUFA sufficient status. Future research should investigate whether an additional n-3 PUFA supplement together with TB medication (rather than only providing sufficient n-3 PUFA intake) may improve markers of AI and whether morbidity may be affected positively or even worsened in this case. Masters

The following case study was discussed during the fourth-year dietetics evaluation process at North-West University, Potchefstroom Campus. It is a reflection of the opinion of the dietitians, students and lecturers involved, describes the actions taken during the nutritional management of the case, and is based on current literature and guidelines relevant to the topic.