Vibrio cholerae, the causative agent of the diarrheal disease cholera, must acquire iron to survive. Although iron is relatively abundant, it forms insoluble ferric complexes in the presence of oxygen. The more soluble ferrous iron is limited to anaerobic or reducing environments. To meet the nutritional needs of the cell, V. cholerae encodes many different ferric iron transport systems but only one characterized ferrous iron transporter, Feo. Feo is widely distributed in bacteria and archaea, but the mechanism for transport is not known. In this study, basic characterization of the V. cholerae feoABC operon was performed to gain further understanding about a critical iron transport system. Each gene in the operon, feoA, feoB, and feoC, was found to be required for ferrous iron uptake. FeoB, an inner membrane protein, is considered to be the ferrous permease but functions for FeoA and FeoC are not known. These studies show that neither FeoA nor FeoC is required for expression of feoB, suggesting that these proteins are required for Feo function. Analysis of the composition of the Feo transporter using a bacterial adenylate cyclase two-hybrid system indicated interactions between Feo proteins, specifically, between FeoC and the cytoplasmic portion of FeoB. This result indicates that feoC encodes a protein that interacts with FeoB and is necessary for ferrous iron transport. / text

Various concentrations of ferrous and ferric iron solutions were held at room temperature for 60 min before they were assayed for ferrous iron, which may be unstable due to oxidation. The ferrous and ferric solutions (in pH 2 HCl) were maintained as such for 60 min without the use of chelators. There was no significant oxidation of ferrous iron. Also, four different levels of each ferrous and ferric iron were injected into proximal duodenal loops of rat intestine and uptake was determined at four different time intervals. Two iron-replete rats were assigned to each of the treatments. The in situ experiments showed that iron was taken up rapidly from pH 2.0 solutions of ferrous and ferric iron. Maximum amount of iron was taken up in the first 10 min. Uptake of ferrous iron was significantly greater (p < 0.05) than uptake of ferric iron, and there were significant differences in total uptake among the four iron levels used. Uptake, absorption, and adsorption kinetics of both ferrous and ferric iron were determined in situ for both iron-replete and iron-deficient rats. Deficiency caused greater uptake and absorption, confirming a biological adaptation of these processes. Both uptake and absorption were greater for ferrous than for ferric iron and were possibly taken up by different pathways or by a ferrous-ferric pathway with preference for ferrous. Uptake and absorption kinetics were biphasic for both ferrous and ferric iron. The first phase demonstrated saturation kinetics and was followed by a nonsaturable phase at higher concentrations of luminal iron. Iron deficiency altered the uptake and absorption kinetics of ferrous and ferric iron, but not always in a similar manner, suggesting that ferrous and ferric iron were each taken up by a separate pathway. Indications were that enhanced absorption during deficiency was largely due to adaptation of ferric uptake. Iron adsorption was directly proportional to luminal iron concentration, but it was greater for ferric than for ferrous, possibly due to charge interactions. Iron deficiency caused increased adsorption of both ferrous and ferric iron, supporting the notion that adsorption acts to maintain iron in a form available for uptake.

Although bacterial leaching of sulphidic minerals is a well-known phenomenon, it is only in the last ten years that full-scale bacterial leaching plants have been commissioned for gold processing. In order for bacterial leaching to compete successfully with other pretreatment processes for refractory ores, particularly with established technologies such as roasting and pressure leaching, it needs to be efficient. This requires the optimization of the parameters affecting the leaching reaction and the growth of bacteria. The entire biotreatment process is agitation leaching, carried out in stirred reactors or Pachuca type reactors. The bacterial oxidation is a complex reaction involving gaseous, liquid and solid phases. The interactions are highly complex, and analysis is complicated by the presence of solids in the leaching medium. Inspite of the amount of research that has been performed, kinetic and process models are underdeveloped. Since kinetic data varies widely with the type and source of concentrate, experimental data should be generated before doing the full-scale reactor design. In sizing reactors for a commercial scale process, it would be useful to have a mathematical model that one could use to predict the amount and rate of release of metal, as a function of the various operating parameters of the system. G.R.Halli arsenical gold sulphide concentrate obtained from Hutti Gold Mines Ltd., Karnataka, was chosen for our study, because of its high refractoriness. An indegenous strain of Thiobacillus ferrooxidans was used for biooxidation. The experiments were conducted in a well-agitated stirred tank reactor under controlled conditions. Sparged air was supplemented with carbon-dioxide for optimized growth. In this work, more than 90% gold and 95% silver could be recovered from the sulphidic gold concentrate when bioleaching was used ahead of cyanidation, compared to 40% and 50% by direct cyanidation. A generalized model, which accounts for both direct bacterial attack and indirect chemical leaching, has been proposed for the biooxidation of refractory gold concentrates. The bacterial balance, therefore, accounts for its growth both on solid substrate and in solution, and for the attachment to and detachment from the surface. The overall process is considered to consist of several sub-processes, each of which can be described in terms of a mechanism and related rate expressions. These sub-processes were studied seperately under kinetically controlled conditions. The key parameters appearing in the rate equations were evaluated using the experimental data. Since the refractory concentrate contains pyrite and arsenopyrite as the major leachable entities, leaching studies have been done on pure pyrite and arsenopyrite as test minerals and the key parameters in the rate equations are evaluated using this data. The model so developed is tested with the leaching kinetics of the concentrate. The growth of bacteria is dependent on the availability of the substrate, ferrous iron, and the dependence is modelled by the widely accepted Monod equation. The effect of carbon dioxide supplementation on the bacterial activity was studied and the optimal concentration for growth was found to be l%(v/v). Studies on indirect chemical leaching showed that the rate is sensitive to surface area of concentrate. Indirect rate constant of arsenopyrite was found to be greater than that of pyrite, since pyrite is more nobler than arsenopyrite. Conditions of direct leaching alone was obtained at high pulp density and using substrate adapted bacteria. The rate constant of arsenopyrite was found to be greater than that of pyrite. The parameters obtained were tested with the overall batch leaching data of the concentrate and favourable comparision was obtained. Thus, it has been possible to isolate the various simultaneous sub-processes occurring during the leaching and propose useful models to describe these processes in some detail. The model has been extended successfully to predict the continuous leaching behaviour using the parameters obtained from the batch data. Studies on the effect of residence time and pulp density on steady state behaviour showed that there is a critical residence time and pulp density below which washout conditions occur. The critical residence time at 10% pulp density was found to be 11 hrs. Operation at pulp densities lower than 5% and residence times lower than 72 hrs is not favourable for efficient leaching. Studies on the effect of initial ferric iron concentration showed that there exists an optimum concentration of ferric iron at which the time required to reach steady state is minimum.

In der vorliegenden Arbeit wurde die Methode der magnetischen Kernspinresonanz (NMR) eingesetzt, um beim Schadstoffabbau stattfindende Prozesse und geochemische Reaktionen zerstörungs- und beprobungsfrei zu untersuchen. Dies ist möglich, da die gelösten Elektronenakzeptoren Sauerstoff und Eisen paramagnetisch sind und somit einen Ein uss auf die NMRRelaxationszeiten ausüben. Der lineare Zusammenhang zwischen der gelösten Sauerstoff- beziehungsweise Eisen-Konzentration und den NMR-Relaxationsraten 1/T1 und 1/T2 wurde quantifiziert. Weiterhin wurde der bereits bekannte Einfluss der Matrixoberflächen von porösen Medien auf die Relaxation von Wasser nachgewiesen. Die paramagnetischen Zentren auf Sandoberflächen führen ebenfalls zu einer Verkürzung der Relaxationszeiten. Es wurde gezeigt, dass die kleinsten Korngrößen der verwendeten Sande den größten Einfluss auf die Oberflächenrelaxation haben. Wird die Oberflächenrelaxation berücksichtigt, ist auch in porösen Medien die ermittelte lineare Abhängigkeit der Relaxationszeiten von der Ionenkonzentration anwendbar, um den Gehalt an gelösten paramagnetischen Ionen aus Relaxationsmessungen zu ermitteln. Beispielhaft wurde der Anstieg der Eisen(III)-Konzentration in der Porenlösung von natürlichen Sanden infolge der Auflösung eisenhaltiger Mineralien von den Oberflächen zeitlich und räumlich detailliert betrachtet. Eine durchgeführte Modellierung zeigte, dass das Reaktionssystem zu Beginn der Reaktion von der Diffusion dominiert wird, am Ende dann die Reaktionsgeschwindigkeit der bestimmende Parameter ist. Die beim biologischen Schadstoffabbau auftretenden Redoxprozesse des Eisens wurden durch rein chemische Reaktionen unter Verwendung von Oxidations- und Reduktionsmitteln simuliert. Die zeitlich und räumlich detaillierte Beobachtung des Anstiegs beziehungsweise des Abfalls der gelösten Eisen(III)-Konzentration in der (Poren-)Lösung war mit NMR-Relaxometrie trotz der Schnelligkeit der Reaktionen möglich. Mit Hilfe der anschliessenden Modellierung wurde der wichtige Einfluss des pH-Wertes auf den genauen Ablauf der Reaktionen deutlich gemacht. Nur in sehr sauren pH-Bereichen (pH < 3) liegen die Eisen(III)-Ionen in Lösung vor. Weiterhin wurde der Einfluss der Mikroorganismen selbst auf die NMR-Relaxations- und Diffusionsmessungen untersucht. Im Rahmen dieser Arbeit wurde an Medien mit Lactobacillus und Penicillium eine Verschiebungen in den Relaxationszeitverteilungen hin zu kleineren Relaxationszeiten gemessen. Dies basiert auf der bereits bekannten Verringerung der Mobilität der Spins innerhalb der Biomasse. Für Bakterien von Geobacter metallireducens konnte erstmals der Verbrauch von Eisen(III)-Ionen durch Reduktion während des Wachstum anhand der ansteigenden T2-Relaxationszeit gezeigt werden.

La dénitrification autotrophe utilisant le fer Ferreux est un bioprocédé innovant pour l'élimination des nitrates, en même temps que l'oxydation du fer dans les eaux usées. Les dénitrifiants chimio-autotrophes convertissent le nitrate en azote gazeux et l'oxydation du Fe(II) conduit à la production de précipités de fer ferrique qui peuvent ensuite être enlevés et récupérés. La possibilité de maintenir une dénitrification autotrophe avec le fer ferreux en utilisant une culture mixte de Thiobacillus, un inoculum de boue activée et des cultures pures de la souche Pseudogulbenkiania 2002 et de T. denitrificans dans différentes conditions de pH et d'EDTA:Fe(II) a été initialement étudiée dans des essais biologiques par lots. Des ratios plus faibles d’EDTA:Fe(II) se traduisent par une efficacité et des taux d'élimination des nitrates plus élevés. La culture mixte de Thiobacillus présente le taux d'élimination de nitrate le plus élevé, égal à 1.18 mM•(g VSS•d)-1.Par la suite, la culture mixte de Thiobacillus a été ensemencée dans deux réacteurs à lit tassé à flux montant identiques. Les deux réacteurs (réacteur 1 et 2) ont reçu respectivement 120 et 60 mg / L de nitrate et une alimentation différente de Fe (II) afin de respecter un rapport molaire Fe(II):NO3- de 5:1. L’EDTA a été supplémenté à un rapport molaire EDTA:Fe(II) de 0,5:1. Le pH, le TRH et la température étaient de 6,5-7,0, 31 h et 22 ± 2 ° C. Dans le réacteur 1, le TRH a été raccourci de 31 à 24 h et la concentration de NO3- a été maintenue stable à 250 mg / L. Inversement, le réacteur 2 a été mis en fonctionnement avec un TRH décroissant et une concentration de NO3- en alimentation, maintenant ainsi un taux de charge de NO3- stable. Après environ 80 jours d'incubation, l'élimination des nitrates était de 88% dans le réacteur 1 pour un THR de 31 h. L'élimination de nitrates la plus élevée obtenue dans le réacteur 2 était de 80%. Une diminution du TRH de 31 à 24 h n'a pas affecté l'élimination du nitrate dans le réacteur 1, alors que dans le réacteur 2 l'élimination du nitrate a diminué à 64%.De plus, l'influence des métaux lourds (Ni, Cu, Zn) sur la dénitrification autotrophe utilisant du fer ferreux a été évaluée dans des essais biologiques discontinus, en utilisant les mêmes quatre cultures microbiennes différentes. L'efficacité et les taux d'élimination des nitrates les plus élevés ont été obtenus avec la culture mixte dominante de Thiobacillus, alors que la souche Pseudogulbenkiania de 2002 était la moins efficace. Cu s'est avéré être le métal le plus inhibiteur pour les cultures mixtes. Un impact plus faible a été observé lorsque le Zn a été ajouté. Le Ni présentait l'effet inhibiteur le plus faible. Une sensibilité plus élevée à la toxicité des métaux a été observée pour les cultures pures. Enfin, la caractérisation minérale des précipités obtenus pour les expériences avec du Cu, Ni et Zn a été étudiée. Chez les témoins abiotiques, l'oxydation chimique du Fe (II) a entraîné la formation d'hématite. Un mélange de différents (hydro)oxides de Fe(III) a été observé pour toutes les cultures microbiennes, et en particulier : i) un mélange d'hématite, d'akaganéite et / ou de ferrihydrite a été observé dans les précipités des expériences réalisées avec la culture mixte dominée par la présence de Thiobacillus; ii) en plus d'hématite, de l'akaganeite et / ou de la ferrihydrite, la maghémite a été identifiée lorsque la culture pure de T. denitrificans a été utilisée; iii) l'utilisation de la culture pure de la souche Pseudogulbenkiania 2002 a entraîné la formation d'hématite et de maghémite; enfin, l'enrichissement en boues activées a permis la production d'hématite et de magnétite en plus de la maghémite. Aucune différence concernant la minéralogie des précipités n'a été observée avec l'addition de Cu, alors que l'addition de Ni et de Zn a probablement stimulé la formation de maghémite. Une caractérisation minérale supplémentaire est cependant nécessaire / Ferrous iron mediated autotrophic denitrification is an innovative bioprocess for nitrate removal, simultaneously with iron oxidation in wastewaters. Chemoautotrophic denitrifiers convert nitrate to nitrogen gas and Fe(II) oxidation results in the production of ferric iron precipitates that can be subsequently removed and recovered. The feasibility of maintaining Fe(II)-mediated autotrophic denitrification with a Thiobacillus mixed culture, an activated sludge inoculum and pure cultures of Pseudogulbenkiania strain 2002 and T. denitrificans under different pH and EDTA:Fe(II) conditions was initially investigated in batch bioassays. Lower EDTA: Fe(II) ratios resulted in higher nitrate removal efficiency and rates. The Thiobacillus mixed culture resulted in the highest specific nitrate removal rate, equal to 1.18 mM•(g VSS•d)-1.Subsequently, the Thiobacillus mixed culture was seeded in two identical up-flow packed bed reactors. The two reactors (reactor 1 and 2) were fed with 120 and 60 mg/L of nitrate, respectively, and a different Fe(II) feed in order to respect a molar ratio Fe(II):NO3- 5:1. EDTA was supplemented at a EDTA:Fe(II) molar ratio 0.5:1. The pH, HRT and temperature were 6.5-7.0, 31 h and 22±2°C. In reactor 1, HRT was shortened from 31 to 24 h and NO3- concentration was maintained stable at 250 mg/L. Conversely, reactor 2 was operated with decreasing HRT and feed NO3- concentration, thus maintaining a stable NO3- loading rate. After approximately 80 d of incubation, nitrate removal was 88% in reactor 1 at HRT of 31 h. The highest nitrate removal achieved in reactor 2 was 80%. A HRT decrease from 31 to 24 h did not affect nitrate removal in reactor 1, whereas nitrate removal decreased to 64% in reactor 2.Moreover, the influence of heavy metals (Ni, Cu, Zn) on Fe(II)-mediated autotrophic denitrification was assessed in batch bioassays. The highest nitrate removal efficiency and rates were achieved with the Thiobacillus-dominated mixed culture, whereas Pseudogulbenkiania strain 2002 was the least effective. Cu showed to be the most inhibitory metal for mixed cultures. A lower impact was observed when Zn was supplemented. Ni showed the lowest inhibitory effect. A higher sensitivity to metal toxicity was observed for the pure cultures. Finally, the mineral characterization of the precipitates obtained in the experiments with Cu, Ni and Zn was investigated. In abiotic controls, the chemical Fe(II) oxidation resulted in hematite formation. A mixture of different Fe(III) (hydr)oxides was observed with all microbial cultures, and in particular: i) a mixture of hematite, akaganeite and/or ferrihydrite was observed in the precipitates of the experiments carried out with the Thiobacillus-dominated mixed culture; ii) on top of hematite, akaganeite and/or ferrihydrite, maghemite was identified when the T.denitrificans pure culture was used; iii) the use of the pure culture of Pseudogulbenkiania strain 2002 resulted in hematite and maghemite formation; finally, the activated sludge enrichment allowed the production of hematite and magnetite besides maghemite. No difference in the mineralogy of the precipitates was observed with the addition of Cu, whereas the addition of Ni and Zn likely stimulated the formation of maghemite. Further mineral characterization is however required

A new class of treatment processes called Advanced Reduction Processes (ARP) has been proposed. The ARPs combine activation methods and reducing agents to form highly reactive reducing radicals that degrade oxidized contaminants. Batch screening experiments were conducted to identify effective ARP by applying several combinations of activation methods (ultraviolet light, ultrasound, electron beam, microwaves) and reducing agents (dithionite, sulfite, ferrous iron, sulfide) to degradation of five target contaminants (perchlorate, nitrate, perfluorooctanoic acid, 2,4 dichlorophenol, 1,2 dichloroethane) at 3 pH levels (2.4, 7.0, 11.2). These experiments identified the combination of sulfite activated by ultraviolet light produced by a low pressure mercury vapor lamp as an effective ARP. More detailed kinetic experiments were conducted with nitrate and perchlorate as target compounds and nitrate was found to degrade more rapidly than perchlorate. The effects of pH, sulfite concentration, and light intensity on perchlorate and nitrate degradation were investigated. The effectiveness of the sulfite/UV-L treatment process improved with increasing pH for both perchlorate and nitrate.

In der vorliegenden Arbeit wurde die Methode der magnetischen Kernspinresonanz (NMR) eingesetzt, um beim Schadstoffabbau stattfindende Prozesse und geochemische Reaktionen zerstörungs- und beprobungsfrei zu untersuchen. Dies ist möglich, da die gelösten Elektronenakzeptoren Sauerstoff und Eisen paramagnetisch sind und somit einen Ein uss auf die NMRRelaxationszeiten ausüben. Der lineare Zusammenhang zwischen der gelösten Sauerstoff- beziehungsweise Eisen-Konzentration und den NMR-Relaxationsraten 1/T1 und 1/T2 wurde quantifiziert. Weiterhin wurde der bereits bekannte Einfluss der Matrixoberflächen von porösen Medien auf die Relaxation von Wasser nachgewiesen. Die paramagnetischen Zentren auf Sandoberflächen führen ebenfalls zu einer Verkürzung der Relaxationszeiten. Es wurde gezeigt, dass die kleinsten Korngrößen der verwendeten Sande den größten Einfluss auf die Oberflächenrelaxation haben. Wird die Oberflächenrelaxation berücksichtigt, ist auch in porösen Medien die ermittelte lineare Abhängigkeit der Relaxationszeiten von der Ionenkonzentration anwendbar, um den Gehalt an gelösten paramagnetischen Ionen aus Relaxationsmessungen zu ermitteln. Beispielhaft wurde der Anstieg der Eisen(III)-Konzentration in der Porenlösung von natürlichen Sanden infolge der Auflösung eisenhaltiger Mineralien von den Oberflächen zeitlich und räumlich detailliert betrachtet. Eine durchgeführte Modellierung zeigte, dass das Reaktionssystem zu Beginn der Reaktion von der Diffusion dominiert wird, am Ende dann die Reaktionsgeschwindigkeit der bestimmende Parameter ist. Die beim biologischen Schadstoffabbau auftretenden Redoxprozesse des Eisens wurden durch rein chemische Reaktionen unter Verwendung von Oxidations- und Reduktionsmitteln simuliert. Die zeitlich und räumlich detaillierte Beobachtung des Anstiegs beziehungsweise des Abfalls der gelösten Eisen(III)-Konzentration in der (Poren-)Lösung war mit NMR-Relaxometrie trotz der Schnelligkeit der Reaktionen möglich. Mit Hilfe der anschliessenden Modellierung wurde der wichtige Einfluss des pH-Wertes auf den genauen Ablauf der Reaktionen deutlich gemacht. Nur in sehr sauren pH-Bereichen (pH < 3) liegen die Eisen(III)-Ionen in Lösung vor. Weiterhin wurde der Einfluss der Mikroorganismen selbst auf die NMR-Relaxations- und Diffusionsmessungen untersucht. Im Rahmen dieser Arbeit wurde an Medien mit Lactobacillus und Penicillium eine Verschiebungen in den Relaxationszeitverteilungen hin zu kleineren Relaxationszeiten gemessen. Dies basiert auf der bereits bekannten Verringerung der Mobilität der Spins innerhalb der Biomasse. Für Bakterien von Geobacter metallireducens konnte erstmals der Verbrauch von Eisen(III)-Ionen durch Reduktion während des Wachstum anhand der ansteigenden T2-Relaxationszeit gezeigt werden.

Background Iron deficiency (ID) is the world‟s most prevalent micronutrient deficiency and predominantly affects developing countries, also South Africa. In areas with low fish consumption and high n-6 PUFA vegetable oil intake, there is a risk for having inadequate n-3 PUFA status. Both iron and n-3 PUFA play important roles in the immune response, and supplementation is a strategy to alleviate deficiencies. However, little is known about potential interactive effects between concurrent iron and n-3 PUFA supplementation on the immune system. This is also important in the context that iron supplementation may be unsafe and may increase morbidity and mortality. Aim The overall aim of this thesis was to assess the effects of iron and docosahexaenoic (DHA)/eicosapentaenoic acid (EPA) supplementation, alone and in combination, on the immune system of ID children. More specifically, these effects were investigated on the occurrence and duration of illness and school-absenteeism due to illness, peripheral blood mononuclear cell (PBMC), red blood cell (RBC) and plasma total phospholipid fatty acid composition, iron status, fatty acid-derived immune modulators and targeted PBMC gene expression. Furthermore, association of PBMC, RBC and plasma total phospholipid fatty acid composition with allergic disease, were also examined. Design In a 2-by-2 factorial, randomised, double-blind, placebo-controlled trial, South African children (n = 321, aged 6–11 y) were randomly assigned to receive oral supplements of either 1) iron (50 mg as ferrous sulphate) plus placebo; 2) DHA/EPA (420/80 mg) plus placebo; 3) iron plus DHA/EPA (420/80 mg); or 4) placebo plus placebo for 8.5 mo, four times per week. Absenteeism and illness symptoms were recorded and biochemical parameters for compliance as well as parameters fundamental to immune function were assessed at baseline and endpoint. Furthermore, in a cross-sectional design, associations of allergic disease with baseline fatty acid composition of PBMC, RBC and plasma were examined. Results The combination of iron and DHA/EPA significantly attenuated respiratory illness caused by iron supplementation. DHA/EPA supplementation alone improved respiratory symptoms at school, but increased headache-related absenteeism. DHA/EPA and iron supplementation individually tended to increase and decrease anti-inflammatory DHA and EPA-derived mediators, respectively. Furthermore the anti-inflammatory DHA-derived immune mediator, 17HDHA was higher in the DHA/EPA plus placebo and iron plus DHA/EPA groups than in the iron plus placebo group. Also, the pro-inflammatory arachidonic acid (AA)-derived modulators (5- and 15-hydroxyeicosapentaenoic acid) were significantly lower in the iron plus DHA/EPA group compared to the placebo plus placebo groups. In the study population, 27.2% of the children had allergic disease and AA in PBMC phospholipids was significantly lower in the allergic children than in the non-allergic children. In RBC phospholipids dihomo-gamma-linolenic acid (DGLA) and the ratio of DGLA: linoleic acid (LA) correlated negatively and the n-6:n-3 PUFA ratio positively with total immunoglobulin E (tIgE). Furthermore, trans-C18:1n-9, tended to be higher in the allergic group. Conclusion DHA/EPA prevented respiratory illness caused by iron supplementation and although DHA/EPA on its own reduced respiratory morbidity when the children were present at school, surprisingly it increased the likelihood of being absent with headache and fever. The biochemical findings compliment the clinical results and support previous observations about DHA/EPA supplementation to reduce inflammation, but add to the current knowledge base that a relatively high oral dose of non-haem iron modulates circulating lipid-derived immune modulators and related gene expression. Furthermore, when supplementing with iron and DHA/EPA combined, in this ID population with low fish intake, the anti-inflammatory effect of DHA/EPA is maintained concurrently with attenuation of respiratory morbidity. This finding support the notion that excess iron (probably as non-transferrin bound iron) becomes available for pathogens and is probably why we found that iron increased respiratory infectious morbidity. The improved clinical outcome with combined supplementation seems to be related to increased lipid-mediator synthesis gene expression and the availability of DHA/EPA, leading to a more pro-resolving profile and enhanced immune competence. Overall these results give better insight into immune function and infectious morbidity in relation to n-3 PUFA and iron status and treatment, as well as the possible association of fatty acid status with allergic disease in young South-African school children. / PhD (Nutrition), North-West University, Potchefstroom Campus, 2015

Background Iron deficiency (ID) is the world‟s most prevalent micronutrient deficiency and predominantly affects developing countries, also South Africa. In areas with low fish consumption and high n-6 PUFA vegetable oil intake, there is a risk for having inadequate n-3 PUFA status. Both iron and n-3 PUFA play important roles in the immune response, and supplementation is a strategy to alleviate deficiencies. However, little is known about potential interactive effects between concurrent iron and n-3 PUFA supplementation on the immune system. This is also important in the context that iron supplementation may be unsafe and may increase morbidity and mortality. Aim The overall aim of this thesis was to assess the effects of iron and docosahexaenoic (DHA)/eicosapentaenoic acid (EPA) supplementation, alone and in combination, on the immune system of ID children. More specifically, these effects were investigated on the occurrence and duration of illness and school-absenteeism due to illness, peripheral blood mononuclear cell (PBMC), red blood cell (RBC) and plasma total phospholipid fatty acid composition, iron status, fatty acid-derived immune modulators and targeted PBMC gene expression. Furthermore, association of PBMC, RBC and plasma total phospholipid fatty acid composition with allergic disease, were also examined. Design In a 2-by-2 factorial, randomised, double-blind, placebo-controlled trial, South African children (n = 321, aged 6–11 y) were randomly assigned to receive oral supplements of either 1) iron (50 mg as ferrous sulphate) plus placebo; 2) DHA/EPA (420/80 mg) plus placebo; 3) iron plus DHA/EPA (420/80 mg); or 4) placebo plus placebo for 8.5 mo, four times per week. Absenteeism and illness symptoms were recorded and biochemical parameters for compliance as well as parameters fundamental to immune function were assessed at baseline and endpoint. Furthermore, in a cross-sectional design, associations of allergic disease with baseline fatty acid composition of PBMC, RBC and plasma were examined. Results The combination of iron and DHA/EPA significantly attenuated respiratory illness caused by iron supplementation. DHA/EPA supplementation alone improved respiratory symptoms at school, but increased headache-related absenteeism. DHA/EPA and iron supplementation individually tended to increase and decrease anti-inflammatory DHA and EPA-derived mediators, respectively. Furthermore the anti-inflammatory DHA-derived immune mediator, 17HDHA was higher in the DHA/EPA plus placebo and iron plus DHA/EPA groups than in the iron plus placebo group. Also, the pro-inflammatory arachidonic acid (AA)-derived modulators (5- and 15-hydroxyeicosapentaenoic acid) were significantly lower in the iron plus DHA/EPA group compared to the placebo plus placebo groups. In the study population, 27.2% of the children had allergic disease and AA in PBMC phospholipids was significantly lower in the allergic children than in the non-allergic children. In RBC phospholipids dihomo-gamma-linolenic acid (DGLA) and the ratio of DGLA: linoleic acid (LA) correlated negatively and the n-6:n-3 PUFA ratio positively with total immunoglobulin E (tIgE). Furthermore, trans-C18:1n-9, tended to be higher in the allergic group. Conclusion DHA/EPA prevented respiratory illness caused by iron supplementation and although DHA/EPA on its own reduced respiratory morbidity when the children were present at school, surprisingly it increased the likelihood of being absent with headache and fever. The biochemical findings compliment the clinical results and support previous observations about DHA/EPA supplementation to reduce inflammation, but add to the current knowledge base that a relatively high oral dose of non-haem iron modulates circulating lipid-derived immune modulators and related gene expression. Furthermore, when supplementing with iron and DHA/EPA combined, in this ID population with low fish intake, the anti-inflammatory effect of DHA/EPA is maintained concurrently with attenuation of respiratory morbidity. This finding support the notion that excess iron (probably as non-transferrin bound iron) becomes available for pathogens and is probably why we found that iron increased respiratory infectious morbidity. The improved clinical outcome with combined supplementation seems to be related to increased lipid-mediator synthesis gene expression and the availability of DHA/EPA, leading to a more pro-resolving profile and enhanced immune competence. Overall these results give better insight into immune function and infectious morbidity in relation to n-3 PUFA and iron status and treatment, as well as the possible association of fatty acid status with allergic disease in young South-African school children. / PhD (Nutrition), North-West University, Potchefstroom Campus, 2015