Lead Institution: University of California, San Francisco
Research Areas
Uganda is emblematic of the challenges faced by high burden countries, where routine surveillance systems are inadequate to assess trends in the burden of malaria or to monitor the impact of control interventions. Through PRISM, researchers have implemented a comprehensive malaria surveillance program including enhanced health facility-based surveillance and detailed longitudinal studies. Complementary laboratory-based studies include surveillance for markers of antimalarial drug and insecticide resistance and serologic measures of malaria exposure. These studies have greatly improved the understanding of the epidemiology of malaria in Uganda and of the impact of control interventions. Researchers plan to strategically focus on quantifying the impact of malaria control interventions, working in close collaboration with partners at the Uganda National Malaria Control Division. The research team will utilize specimens collected from the surveillance system to generate genomic and serologic data to address key questions about malaria epidemiology, transmission, diagnostics, and antimalarial drug and insecticide resistance. A central goal of the ICEMR is to improve malaria surveillance to better assess the impact of malaria control interventions and guide evidence-based utilization of existing and novel interventions to reduce the malaria burden using an adaptive approach.
The PRISM program consists of three research projects linked together in an integrated manner to maximize scientific discovery. Research Project 1 (Surveillance and Impact Evaluation Project) will utilize health facility and community-based malaria surveillance data from sites with varied transmission intensity and control interventions to monitor trends, estimate the impact of interventions, and provide clinical data and specimens for the program’s other research projects. Research Project 2 (Resistance Project) will characterize the evolution of genotypic markers of drug and insecticide resistance and assess the impacts of resistance on malaria transmission. Research Project 3 (Molecular Epidemiology Project) will characterize key parasite and vector genomic traits and human serological responses.
These highly interrelated projects are being conducted in settings with varied malaria epidemiology and differing population level control intervention to provide critical information needed to optimize strategies for the control and ultimate elimination of malaria in Uganda.
Key Achievements
The establishment of an enhanced health-facility-based malaria surveillance network to monitor temporal and geographic trends in malaria burden and assess the impact of population level control interventions
The basic principle of the PRISM health facility-based malaria surveillance system has been to improve the quality and utility of primary data generated at selected health facilities. This has been accomplished by collecting individual level data in an electronic format and providing training and support to improve malaria case management. Researchers currently work at 38 health centers, referred to as Malaria Reference Centers (MRCs). Since 2018 an emphasis has been placed on collecting data on village of residence for all patients presenting to the MRCs. These data along with the identification of target areas around the MRCs and enumeration surveys to estimate the populations of these target areas have allowed researchers to generate estimates of malaria incidence within the target areas. In addition, these target areas have provided a sampling frame for conducting cross-sectional community surveys to estimate the coverage of malaria control interventions and generate data on parasite prevalence. These surveillance data have been used to measure the impact of population level vector control interventions including indoor residual spaying of insecticide (IRS) and the universal distribution of different types of long-lasting insecticidal bednets (LLINs). These assessments have been instrumental in helping to shape policy and respond to changes over time.
The use of comprehensive cohort studies and entomological surveillance to improve our understanding of the epidemiology of malaria in different settings
PRISM has been at the forefront of conducting comprehensive cohort studies. These studies enroll residents of households randomly selected from enumerated catchment areas and follow them over an extended period including detailed entomological surveillance within the cohort households. Studies have provided a wealth of high-quality epidemiological data on measures of disease, infection, and transmission in a variety of settings in addition to generating a large specimen biobank that has been used for a variety of complementary laboratory studies. Research showed a dramatic resurgence of malaria in eastern Uganda was temporally associated with a change to clothianidin-based IRS formulations and emergence of Anopheles funestus as the predominant vector. Malaria burden subsequently decreased after a shift back to IRS with Actellic. This work highlights the ability of malaria vectors to rapidly circumvent control efforts and the importance of high-quality surveillance systems to assess the impact of malaria control interventions and generate timely, actionable data.
The use of molecular tools to better understand the epidemiology of blood stage infections
In highly endemic countries like Uganda, the malaria parasite reservoir is dynamic, and includes symptomatic and asymptomatic infections. Asymptomatic infections comprise a large proportion of the parasite reservoir and are important because studies have shown that even submicroscopic (low-density infections that are below the level of detection by microscopy) asymptomatic infections can transmit parasites to mosquitoes. In cohort studies, ICEMR researchers use a highly sensitive quantitative PCR assay, in addition to microscopy, to longitudinally assess parasite density in each participant. Use of highly sensitive molecular diagnostics shows these techniques are critical to characterize the parasite reservoir more fully, given that microscopic infections represent only the “tip of the iceberg.” Furthermore, these techniques can be used to identify asymptomatic individuals who provide persistent reservoirs of transmission in lower transmission settings; to identify infected populations who, although asymptomatic, may benefit clinically from treatment; and to evaluate the results of interventions. In addition to characterizing infections by qPCR, researchers also use amplicon deep-sequencing to genotype parasite DNA. This allows the team to follow individual clones (strains) in cohort participants and study the dynamics of these infections over time. Because polyclonal infections can occur due to co-infection (one mosquito bite transmitting multiple clones) or superinfection (multiple bites), genotyping to distinguish the clones present in an infection is necessary to assess the molecular force of infection (mFOI), or the incidence of genetically distinct parasite clones acquired over time, and to estimate the rate of clearance of infections. Researchers used amplicon sequencing to estimate the clearance of asymptomatic Plasmodium falciparum infections and found that females consistently cleared their infections at a faster rate than males, even after adjusting for age, baseline infection status, and parasite density. They also found no evidence for a sex-based difference in exposure to infection through behaviors or as measured by mFOI. The team has used amplicon deep-sequencing to better understand transmission dynamics. By carefully characterizing infections in human participants, they can track which parasite clones are transmitted to mosquitoes during membrane feeding experiments.
The use of molecular tools and membrane feeding assays to better understand human to mosquito transmission
A key objective of the PRISM program has been to quantify gametocyte production during P. falciparum infections and determine human and mosquito factors that influence gametocyte transmission. Using data from cohort studies researchers have demonstrated that many incident infections are short-lived and spontaneously clear before gametocytes were produced, while gametocyte production was near-universal among infections that lasted 12 weeks or longer. Infections in individuals with sickle-cell trait (HbAS) were more likely to produce gametocytes and produced gametocytes at higher densities compared to those in wildtype (HbAA) individuals. Importantly, gametocyte carriage was considerably higher among asymptomatic infections than clinical malaria cases. Gametocyte density was positively associated with total parasite density among asymptomatic infections, in which gametocytes were often produced throughout the duration of infection, but not among clinical malaria cases, in which relative gametocyte densities were considerably lower. The researchers’ mosquito feeding assays have demonstrated a strong positive association between gametocyte density and mosquito infection rates, and evidence for a lower transmission potential of gametocytes that are sampled during clinical malaria attacks compared to asymptomatic infections. When considering both the occurrence and duration of infections over time and their infectivity to mosquitoes, clinical malaria cases were estimated to be responsible for <5% of all infected mosquitoes; rather, the large majority of mosquito infections were from asymptomatic individuals. Intriguingly, a small minority of asymptomatic school-age children were responsible for most mosquito infections.
Assessment of temporal and geographic trends in measures of antimalarial drug and insecticide resistance
ICEMR researchers conducted serial surveillance of the prevalence of P. falciparum drug resistance markers across Uganda since 2010. Prevalence of mutations associated with resistance to chloroquine and other aminoquinolines (in particular PfCRT 76T and PfMDR1 86Y) have decreased markedly. Duplications of pfmdr1 and plasmepsin genes associated with resistance to mefloquine and piperaquine, respectively, in SE Asia are very uncommon across Uganda. Prevalence of 5 mutations associated with moderate resistance to sulfadoxine-pyrimethamine (SP; PfDHFR 51I, 59R, 108N; PfDHPS 437G, 540E) has remained very high with prevalence of two additional mutations associated with high level resistance to SP (PfDHFR 164L; PfDHPS 581G) increased markedly, especially in western Uganda. Until recently, studies from the group showed no convincing evidence for artemisinin partial resistance in Uganda based on parasitological or molecular criteria. However, this situation has changed remarkably in recent years. Researchers have demonstrated the emergence of two validated markers of artemisinin resistance, PfK13 469Y and 675V, in northern Uganda. They have recently demonstrated the emergence of two additional validated resistance markers, PfK13 469F and 561H in southwestern Uganda. Taken together, these results demonstrate combined prevalence of validated markers of artemisinin partial resistance >50% in both northern and southwestern Uganda, with spread of the mutations first seen in northern Uganda to districts around the country in 2021-22. These results suggest stable prevalence of artemisinin partial resistance across Uganda, and concern regarding the antimalarial efficacies of ACTs.
Key objectives of the PRISM program have been to use transcriptomic and genomic approaches to identify novel mediators of insecticide resistance and to monitor anopheline insecticide resistance at diverse sites against a background of varying control interventions and malaria endemicity. These data have been used to identify novel origins of some target-site resistance mutations and strong signatures of selection, unique to the Ugandan data set, around some candidate insecticide resistance gene families. Complementary whole genome microarray and candidate genome wide association studies have been used to identify pyrethroid resistance associated variants in both An. gambiae and An. arabiensis. Characterization of phenotypic and genotypic markers of pyrethroid resistance have been conducted at 11 sites with varied vector control strategies. These studies have documented widespread pyrethroid resistance across Uganda, higher in An. gambiae s.s. than in An. arabiensis. Addition of PBO to pyrethroids increased mortality, supporting deployment of PBO LLINs in Uganda.
Regional Impact
There has recently been a dramatic increase in the scale up of control interventions and reduction in the burden of malaria across Africa. However, progress has not been uniform, and in fact has been slowest in countries with the highest burden, such as Uganda. Malaria covers a wide range of epidemiological settings in the country. This ICEMR is conducting studies in health centers and surrounding communities around Uganda, ranging from areas of relatively low transmission intensity to areas with some of the highest transmission intensities recorded in the world. Researchers hope to use the varied settings to evaluate intervention strategies and assess optimal control methods.
Study Sites
View Associated sites for the Uganda ICEMR in a larger map
Collaborating Institutions
- Infectious Diseases Research Collaboration (IDRC), Kampala, Uganda
- Makerere University College of Health Sciences, Kampala, Uganda
- Liverpool School of Tropical Medicine, Liverpool, UK
- London School of Hygiene and Tropical Medicine, London, UK
- Radboud Institute for Health Sciences, Netherlands
- Stanford University, Stanford, CA
Staff
Co-Principal Investigators: Moses R. Kamya MD, Ph.D.; Grant Dorsey, MD, Ph.D.
Project Leads
- Surveillance and Impact Evaluation: Joaniter I. Nankabirwa, Makerere University and Infectious Diseases Research Collaboration (IDRC), Uganda
- Molecular Epidemiology: Isaac Ssewanyana, Infectious Diseases Research Collaboration (IDRC), Uganda
- Resistance: Samuel Nsobya, Infectious Diseases Research Collaboration (IDRC), Uganda