Magister Public Health - MPH / In Zambia, there had been a large scaling up of new interventions to control malaria since 2003, which included the distribution of rapid diagnostic tests (RDTs), used to immediately determine if someone with symptoms suggestive of malaria actually has malaria; training of health workers in the use of the RDTs; and the prescription of artemisinin-based combination therapy (ACT) to which the malaria parasite is sensitive, rather than the old treatment regime of chloroquine to which the malaria parasite had become resistant. The use of RDTs to confirm the presence of malaria before treating for it with ACT became known as the „test and treat‟ policy. Previously, since the 1960s, in malaria endemic areas such as Zambia, children presenting with fever (the commonest symptom of malaria) without any obvious other cause for the fever, were assumed to have malaria and were hence treated for it with chloroquine. This was known as "presumptive treatment" of malaria. The combination of "presumptive treatment" and the use of a single medication led to the development of high levels of resistance to chloroquine, to the extent that it is now no longer an effective treatment for malaria. Years after the introduction of the "test and treat" policy, it was still unclear to what extent it was being implemented, as there was initial reluctance by health workers to test all children presenting with fever for malaria and if they did test they may not have followed the management guidelines of treating those who test positive with ACT and further investigating those who test negative for the cause of the fever. It seemed that staff had gotten used to the "presumptive treatment" approach to malaria over almost 4 decades and hence were quite reluctant to abandon it. The conflicting guidelines for malaria treatment in children between IMCI and "test and treat‟ has promoted a paradox between presumptive treatment for malaria and "test and treat" approach as IMCI teaches health workers to treat febrile children presumptively for malaria whereas the "test and treat" approach requires them to first make a definitive diagnosis before treating. Hence although the "test and treat" approach was instituted to overcome the problems with presumptive treatment approach it now had to contend with the competing and contradictory influence of the IMCI approach. This study therefore aimed to assess what proportion of children aged five years and younger who presented with fever were managed via the "test and treat" guidelines and which factors were associated with this, in Lusaka District, Zambia. Methodology: A cross sectional analytical study design was used based on a review of medical records. A sample size of 800 medical records of children presenting with fever was selected from 10 out of the 23 health care facilities in Lusaka, using a multistage stratified random sampling technique. Four hundred records were sampled from 2008 records (five years after commencement of the "test and treat" policy) and 400 from 2011 records (eight years after commencement of the "test and treat" policy). Trained data collectors used a data extraction tool to transcribe demographic and clinical data from the medical records in a standardized manner. Data Analysis: Univariate descriptive statistics analysis was performed using measures of central tendency and measures of dispersion to analyze numerical (continuous) variables such as age, weight and body temperature; and using frequencies for categorical variables such as gender, area of residence, RDTs/microscopy malaria tests conducted, received ACT if RDT positive, presence of an ACT treatment chart on the health centre wall and availability of a weighing scale. To determine the relationship between variables, bivariate analysis via the prevalence ratio was conducted. Results: Just over half (55%) of all children with fever were tested for malaria in 2008 and this gratifyingly increased to (73%) in 2011. Overall, the proportion of children correctly and appropriately treated with ACT, which means that those who tested positive for malaria were given ACT, was 85% in 2008 but regrettably dropped to 72% in 2011. Although "presumptive treatment" decreased from 24% in 2008 to 11% in 2011, the proportion of children with fever not tested for malaria, and although not treated for malaria, but left without a definitive diagnosis of their fever being made, remained high but dropping (22% in 2008 and 16% in 2011). Similarly the proportion of children who tested negative for malaria but then did not undergo any further investigation also unfortunately remained very high and rising (57% in 2008 and 89% in 2011). A combination of the above poor clinical management practises resulted in only 38% of children with fever in 2008 and unfortunately dropping to only 33% in 2011 being correctly managed (tested for malaria via RDT or microscopy and treated with ACT if positive, while further investigated for the cause of fever if negative). On preparedness of the health facility to implement the "test and treat" policy, it was noted that only 4 out of 10 health facilities were at least minimally prepared to do so, but paradoxically on bivariate analysis those minimally prepared were less likely (PR 0.62; 95% CI 0.41-0.94) to correctly manage the patients in 2011 than those who were unprepared. A similar paradox occurred for those correctly treated with ACT after testing positive, with facilities which were minimally prepared being less likely to do so (PR 0.28; 95% CI 0.14-0.58) in 2011 than those facilities which were unprepared to implement the "test and treat" policy. However these associations were inconsistent over time, as the associations were not present in 2008. Similarly all other factors such as staff category (doctor, nurse, clinical officer) and type of presenting symptoms besides fever (anorexia, lethargy, pallor) assessed, were not consistently associated with testing for malaria in both 2008 and 2011. The same applied for the other two main outcome variables of 'treated with ACT after test positive for malaria' and 'correctly managed child with fever', in that there were no factors that showed a consistent association with them in both 2008 and 2011. Conclusion: Testing of children with fever for malaria is at a low level but rose between 2008 and 2011. Paradoxically the proportion of those diagnosed with malaria who were correctly treated with ACT dropped between 2008 and 2011, as did the proportion of children with fever who were correctly managed. No factors assessed in this study were found to be consistently associated in both 2008 and 2011 with either testing for malaria, or treating confirmed malaria cases with ACT, or managing patients with fever correctly. Recommendations: In order for health workers to correctly implement the "test and treat" policy, which involves a series of complex steps, they ought to be formally trained to do so, mentored and constructively supervised. Additionally health facilities should be adequately equipped to enable health workers to fully implement the policy. Further studies to assess factors associated with the correct management of malaria via the "test and treat" policy are warranted.

Malaria accounts for 198 million cases worldwide; with a high mortality rate. 584000 deaths were reported in 2013. Malaria is a re-emerging disease globally due to drug resistance, parasite recrudescence and non-availability of a vaccine. Chloroquine, quinine and antifolates served as frontline antimalarial drugs for decades. Development of resistance to chloroquine and antifolates, and the decreased efficacy of mefloquine, and even quinine, in malaria-endemic regions, has led to artemisinin derivatives evolving as frontline drugs. Artemisinin is a potent antimalarial compound and clears around 104 parasites per cycle. Despite being a potent antimalarial, artemisinin derivatives suffer from poor pharmacokinetic properties and short half lives. This has led to the development of artemisinin-based combination therapies (ACTs) using a partner drug with a longer half-life. However, resistance to ACTs has been reported in the last few years, perhaps due to lack of adherence to prescribed regimens or suboptimal treatment and the use of counterfeit drugs. Therefore there is an urgent need to develop an alternative ACT which overcomes these limitations. This thesis entitled “Adjunct therapy with curcumin for the treatment of malaria: studies in a murine model” describes the antimalarial activity of curcumin and artemisinin and the adjunct role of curcumin in the prevention of parasite recrudescence and cerebral malaria. The thesis is divided into three chapters: The first chapter entitled “Introduction: Malaria and anti-malarial drugs” consists of a brief introduction of malaria, the parasite life cycle and currently known antimalarial drugs. During the course of infection, the Plasmodium undergoes sporogony in the mosquito, and merogony and schizogony in the human host. All these life cycle stages are briefly described with depictions. A major part of this chapter is dedicated to describe antimalarial compounds under the following headings 1. Quinoline derivatives 2. 4-aminoquinolines 3. Antifolates 4. Artemisinin derivatives 5. Antibiotics and 6. Curcumin. The second chapter is aimed at examining the ability of curcumin-arteether (a synthetic derivative of artemisinin) combination therapy in preventing parasite recrudescence in a murine model through immunomodulation employing various immunological, molecular biological, and biochemical techniques. The use of suboptimal doses of antimalarial drugs leads to recrudescence or relapse of malaria (reappearance of the parasite in blood after antimalarial regimen). In the present study we have addressed this issue by the use of curcumin as an adjunct molecule with α,β arteether (a synthetic derivative of artemisinin). We have studied recrudescence in a Swiss mice model. A suboptimal dose was standardized by the use of different doses of α,β arteether (AE) ranging from 250µg to 1500 µg. We found 750 µg to be a suboptimal dose and studied the adjunct nature of curcumin when animals were treated with AE suboptimal dose or AE+curcumin (AC) combination treatment and monitored the survival of animals. Our results clearly demonstrate that ~95% of animals treated with the suboptimal AE dose died of recrudescent malaria but there was almost 100% survival of AC-treated animals; these animals were under observation for at least 3 months. We have studied the effect of curcumin in a recrudescence model at the molecular level. Curcumin by itself has antimalarial activity, but only in combination with α,β arteether prevented recrudescence. Our results indicate that curcumin has immunomodulatory activity. Serum cytokine analysis and spleen mRNA analysis for proinflammatory and anti-inflammatory mediators indicate that AC treatment effectively reduced both mRNA and serum cytokine levels of IFNγ, TNFα, IL-12 and effectively increased both mRNA and serum levels IL-10 and antibodies of the IgG subclass. Using TLR2 and IL-10 knockout animals, we have conclusively demonstrated that TLR2 is involved in the production of IL-10, and IL-10 is required for the AC-mediated protection of animals during the recrudescence period. We conclude that curcumin is able to prevent parasite recrudescence essentially by switching the Th1 response to a Th2 response. The third chapter deals with the study the effect of areether-curcumin (AC) combination therapy in the prevention of Experimental Cerebral Malaria. Although malaria mortality rates have decreased by an impressive 47% between 2000 and 2013, it is still a major affliction of mankind (WHO 2014). Plasmodium falciparum infection causes human cerebral malaria (HCM). The mortality rate in HCM is unacceptably high (15–20%), despite the availability of artemisinin-based therapy. HCM is characterized by a rapid progression from headache, general malaise, and prostration to hemiparesis, ataxia, unrousable coma, and death. Paediatric HCM deaths are mostly due to respiratory arrest. Alternatively, death may be due to parasite-mediated injury to a sensitive location; a small lesion due to parasite in brain stem can cause sudden respiratory arrest. In HCM, cytoadherence of pRBCs in brain microvasculature has been implicated as a major contributing factor for CM pathology. The failure of a large number of adjunct therapies in HCM demands the development of new intervention strategies. An effective adjunct therapy is urgently needed. Experimental Cerebral Malaria (ECM) in mice manifests many of the neurological features of HCM. In this study, we have demonstrated the efficacy of curcumin and PLGA nanocurcumin in the treatment of Experimental Cerebral Malaria (ECM), using the Plasmodium berghei ANKA-infected mouse model (C57BL/6). Curcumin/PLGA nanocurcumin alone can prevent the onset of ECM. We have shown that curcumin/PLGA nanocurcumin can prevent CD8+ T cell, CXCR3+ CD8 T cell and parasite-infected RBC (pRBC) sequestration in the brain. These are also the essential parameters underlying HCM. We have also demonstrated that curcumin effectively inhibits T cell proliferation in spleen. We have explained the anti-inflammatory effects of curcumin by showing the inhibition of NF-B in both brain and spleen, which is a plausible explanation. But, curcumin/PLGA nanocurcumin treated animals died later due to build up of parasitemia in blood and subsequent anemia. Moreover, a combination therapy with arteether and curcumin given even after the onset of neurological symptoms can completely cure and protect the animals against mortality. We have tested AC-combination after the onset of symptoms to mimic patient conditions in HCM, since the murine regimens reported were not successful in the treatment of HCM. Our results clearly demonstrate that AC treatment even after the onset of symptoms ensures 100% survival. Since the bioavailability of curcumin is reported to be poor, we have also tested the efficacy of PLGA nanocurcumin and find that it is superior to native curcumin in terms of therapeutic effects. It is concluded that curcumin would be an ideal adjunct drug to be used with the artemisinin derivatives to treat malaria, including cerebral malaria.