An Introduction to Chonghao Alan Yin Malaria is a disease which has plagued humanity for millennia. In 2016, analysis of ancient DNA from two individuals who lived in first century Italy, at the height of the Roman Empire’s power, showed fragments of mitochondrial DNA from parasites, (1) one of four species of Plasmodium spp. known to infect humans, and arguably the deadliest. In 2018, it is estimated that there were 228 million cases of malaria globally leading to 405000 deaths, of which roughly two-thirds came from children under the age of five. (2) The CDC estimates that direct costs due to malaria amount to at least 12 billion US dollars per year, (3) and this is only looking at direct costs – costs associated with medical services and intervention. Malaria as a disease is most prevalent and most intense in the tropical and subtropical parts of the world, disproportionately so in Africa, which claimed 94% of deaths in 2018. (2) It is well established that malaria is a vector-borne disease in which mosquitoes pertaining to the genus Anopheles transmit Plasmodium spp. to humans. Of these, the Anopheles gambiae is the most efficient vector, and is found only in sub-Saharan Africa. (4) I remember hearing from Prof Brendan Crabb, Director of the , describing malaria as a “disease of poverty,” and a “key driver in poverty” in one of my lectures in 2019. That statement has stuck with me to this day. It is well known that poorer countries, especially those near the equator, geographically correlate with an increased risk of malaria. Yet, as Gallup and Sachs (4) note, it is uncertain whether poverty alone significantly affects the degree of success of malaria control. From 1969 to 1976, the WHO sponsored a non-profit study investigating the impact of malaria control measures on transmission rates. The location of interest was Garki, Nigeria, where it was estimated that a person would receive 174 bites per night by Anopheles gambiae and 94 bites per night by Anopheles funestus vectors during the wet season. (5) More than $6 million would be spent over 7 years, and although rates of mosquito bites decreased by 90%, this did not meaningfully change the rate of malaria transmission. Strategies included residual insecticide spraying, reaching an average 99% coverage of buildings, in addition to mass administration of the sulfalene-pyrimethamine drug as prophylaxis. Perhaps more despairingly, the strategies employed here were noted to be very intense, expensive, and ultimately financially unsustainable for long-term use in the region. Even so, these strategies were of little benefit. The topic of malaria control is highly complex. To understand the pathology of malaria, and thus the rationale of evidence-based strategies in combatting malaria, great efforts have been placed in studying the genetics and complicated molecular involved. P. falciparum parasites in particular have three stages of development: a mosquito stage, where sexual reproduction occurs, resulting in the formation of haploid organisms called sporozoites, which then infect people. At first, liver cells are infected and rupture, releasing millions of parasites into the bloodstream and leading to invasion of red blood cells (RBCs), where the magnitude of parasites increases by factors of up to 106. It is here in the blood stage where viral load is at its peak and where disease occurs, which may happen up to a month after the initial mosquito bite. Once inside a RBC, the parasite generates an enclosure around itself known as a parasitophorous vacuole (PV) membrane, and begins producing proteins which are exported across this membrane and into RBC cytosol through special PTEX protein translocons located at the surface of the PV membrane. (6) Infected RBCs take on a characteristic rigid, bumpy appearance due to the presence of knob- associated histidine-rich protein (KAHRP), which helps the infected cell bind to blood vessel walls by providing stability against large shear forces due to blood flow. Actual binding is due to the presence of Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1). This process of binding, referred to as cytoadherence, is what allows the infected RBC to in effect avoid the body’s splenic clearance methods, which would normally detect changes to cell shape and deformability. (7) Therefore, with a failure to clear infected RBCs, sequestration of those infected cells occurs, and this is essential to the survival of the parasite. Astonishingly, there have been as many as 60 var genes identified (there very likely being many more undiscovered at present), each leading to a slightly different PfEMP1 protein, and thus binding to different receptors in different locations within the body. Only one var gene is expressed at a time; this is under epigenetic control and selective pressure. When the body develops immunity to any particular PfEMP1, ‘switching’ of var genes will occur. As a result, waves of parasitaemia occur with a cycle of clearance, switching and replication. (8) Affected individuals who survive to adulthood, even to the age of 5, will likely have immunity to malaria due to having encountered so many PfEMP1 variants. In the case of P. falciparum, new challenges are emerging, including resistance to first line drug treatments in southeast Asia. Artemisinin, and its derivatives, are cornerstone drugs used in first line treatment of P. falciparum infections, especially as part of Artemisinin-based combination therapies (ACTs). Although Artemisinin was first discovered in the 1970s, the precise mechanism of action of dihydroartemisinin (DHA), the active metabolite of clinically relevant Artemisinin derivatives, was not clear until recently. (9) These efforts have been motivated by cases of artemisinin resistance which have been linked to Pfkelch13 gene mutations, first described in Cambodia in 2008 and now spread across the Greater Mekong region, (10). Posing a serious threat to malaria control, recent evidence suggests that the Pfkelch13 C580Y mutant allele has become the dominant lineage in the Greater Mekong region, overtaking other Pfkelch13 mutations. (11) More worrying is that resistance to piperaquine, the partner drug to DHA in dihydroartemisinin-piperaquine (DHA- P) ACT, one of five first line ACTs recommended by the WHO, has also become increasingly common, often present in conjunction to Pfkelch13 mutations. (11) Together, these have led to substantial decreases in DHA-P treatment efficacy in Cambodia since 2008. (11) Thus, in addition to managing further spread of multi-drug resistant parasites across the southeast Asia region towards India, novel drug development is sorely needed, and represents a new hope. One potential target for drug development lies in the aforementioned PTEX translocon. Once inside an infected cell, because the parasite essentially isolates itself from the rest of the RBC cytoplasm, it is then reliant on utilising the PTEX translocon in order to pass necessary proteins such as KAHRP, which constitutes a key survival mechanism, through the PV membrane to the RBC’s surface. Therefore, disrupting or blocking the function of PTEX shows promise. Great strides have been made in studying the structure of the PTEX complex, as demonstrated recently by Ho et al, (6) who have managed to capture the 3D structure of PTEX. Figure 1. The PTEX Translocon. (6) In addition to navigating drug resistance and refining our understanding of the underlying molecular biology in the discovery and design of new drug treatments, other barriers have also been identified in malaria control, ranging from social, cultural, financial and institutional issues. Systematic reviews in recent years have identified various shared themes in the barriers to malaria control. One factor recognised by Paul et al. (12), specifically in East Africa, was that community education levels often led to poor adherence to treatments. This was consistent with the findings of Maslove et al. (13) across sub-Saharan Africa, who identified that due to cultural norms and traditional beliefs of the disease, traditional therapies were preferred to more evidence-based medicines and healthcare clinics. Furthermore, the disease itself was often considered to be caused by factors inconsistent with the literature, such as environmental factors and even supernatural causes. Other factors identified include financial costs, such as the cost of insecticide-treated bed nets, or level of access to healthcare systems, with distance and transport costs also cited. (13) Another factor consistent across both reviews was the quality of healthcare present in communities; health workers were often patronising, or lacked sufficient training, thus negatively influencing community perceptions of the healthcare system. (13) On a broader level, operational and logistical issues have been identified in the policymaking process. Given the complexity of malaria control, multiple organisations are often implicated across local, national, and international levels. At each level, different time frames, priorities and strategies may confound the approach and scope of antimalaria measures. (12) Therefore, some considerations in addressing these issues may lie in implementing better educational programs targeted towards promoting health literacy amongst local communities, simultaneously raising awareness, and understanding of the disease, while also fostering greater engagement with healthcare systems. Overall, a greater shift towards more evidence- based practices is needed while also respecting cultural traditions to avoid further straining relationships between the community and healthcare systems. Finally, a greater level of coordination is needed amongst the organisations involved in malaria control. Indeed, the topic of malaria control is truly multidisciplinary. Throughout this article, we have attempted to explore the burden of malaria on the underdeveloped parts of the world, particularly in Africa, the fundamental science underpinning the pathology of malaria, a brief update on antimalarial drug resistance in southeast Asia, and ultimately the need for novel drug development. Yet, even if these problems are solved, relevant issues in the management of global malaria control efforts must still be addressed, which is no trivial task. The topic of malaria and issue of malaria control thus represents a blending of many disciplines and fields which I enjoy individually, yet when combined, is greater than the sum of its parts.

Acknowledgements: To Prof Brendan Crabb for stimulating my initial interest in malaria and to Assoc Prof Jack Richards, my mentor for GHMP 2020 in providing support, personal insight on malaria and great conversations regarding a clinician- career pathway.

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