21st International Congress for Tropical Medicine and Malaria (ICTMM) – 2024: Day 3

MESA Correspondents bring you cutting-edge coverage from the ICTMM 2024 Congress “Global Responses and Interdisciplinary Research Towards Eliminating Tropical Diseases”.

Malaria – Molecular Biology

Sarah A. Cassidy-Seyoum (Menzies School of Health Research, Australia) presented the challenges and potential solutions for the countrywide implementation of glucose-6-phosphate dehydrogenase (G6PD) testing for Plasmodium vivax malaria treatment in Cambodia. Cambodia aims to eliminate malaria by 2025 despite a high prevalence of P. vivax infection. The drug used to kill hypnozoites of P. vivax can cause severe hemolysis in G6PD deficient individuals. To address the issue, biosensor G6PD test was implemented and its role in P. vivax case management was assessed in three high-burden provinces. The surveillance data was analysed, and 67 interviews and 29 focus group discussions were conducted in 2022 and 2023. From the data, most patients were diagnosed by village malaria workers (VMWs). Only 49% reached the health center because of issues related to cost, patient perception, previous treatment experience, and rumors. 94% accepted the G6PD test. She concluded by pointing out that bringing treatment closer to patients through VMWs would improve access to malaria treatment.

Muhammad Fikri Heikal (Khon Kaen University, Thailand) explored the role of Opisthorchis viverrini helminth defense molecules (HDM) in shaping host immunity. HDM is a secretory product of helminth with structural characteristics that mimic the mammalian antimicrobial peptide LL-37. These molecules are conserved across liver fluke species. Fasciola hepatica (FhHDM) neutralized lipopolysaccharide (LPS)-induced inflammation while Clonorchis sinensis HDM (CsHDM) does not affect LPS-induced inflammation. Distinct migration routes influence immunomodulation strategies among liver flukes. F. hepatica suppresses M1 macrophage activation, while O. viverrini and C. sinensis activate M1 macrophages. Heikal investigated the role of the O. viverrini HDM (OvHDM) C-terminal synthetic peptide in LPS-induced inflammation using the in vitro ELISA technique. The results showed that OvHDM increased the level of LPS-induced inflammation. HDM from different parasites showed a distinct effect on LPS-induced inflammation. HDM is utilized by the liver flukes to modulate the immune response to facilitate their survival.

Brendan Crabb (Burnet Institute, Australia) explained how inhibiting lipid transfer and ring development can stop malaria parasites. When the merozoite makes contact with a red blood cell (RBC), it begins to invade the RBC and eventually develops into the ring stage. An aryl-acetamide compound called MMV006833 (M-833) prevents Plasmodium falciparum from growing inside the RBC after invasion. It targets the P. falciparum steroidogenic acute regulatory lipid transfer protein (PfSTART-1). In the presence of START inhibitors, merozoites are unable to transform into amoeboid rings. Crabb suggested that START is involved in the transfer of lipids necessary to establish the parasite’s parasitophorous vacuole membrane and for the internal reorganization of membranes needed for the transformation of merozoites into rings.

Badriah Alkathiri (Chungbuk National University, Democratic People’s Republic of Korea) discussed the identification of tick-borne pathogens and endosymbionts in ticks using metagenomic next-generation sequencing (NGS). Ticks are considered the second most significant biological vector, with a high capacity to harbor and transmit various pathogens. Alkathiri assessed tick-borne bacteria in ticks distributed in Korea using 16S rRNA metabarcoding. A total of 6,907 ticks were pooled into 534 pools. DNA was extracted from each pool and subjected to metagenomic sequencing. Verification of Bartonella, Rickettsia, Wolbachia, and Ehrlichia was performed using PCR. The results showed that the prevalence of Rickettsiella was high at 67.28%, while Coxiella was at 22.12%. Rickettsia, Wolbachia, and Ehrlichia were also detected in the samples. Further molecular characterization and phylogenetic analysis revealed that 70.45% of Rickettsia and 2.60% of Wolbachia were found in Haemaphysalis longicornis, while 16.81% of Wolbachia were found in Haemaphysalis flava. The presence of Wolbachia in Haemaphysalis longicornis and Haemaphysalis flava represents novel findings.

Jahnvi Jakhan (ICMR National Institute of Malaria Research, India) addressed that glucose-6-phosphate dehydrogenase deficiency (G6PD-d) is an erythroenzymopathy leading to hemolytic anemia under oxidative stress. Although in other parts of the world, the correlation between G6PD and malaria has been extensively researched and implemented, it is still poorly understood in India. The gap being the failure to elucidate the spectrum of mutation causing G6PdD-d with context to P. vivax malaria in India. To address this gap, Jakhan aimed to genotypically correlate G6PD variants found in Delhi and Goa regions of India where such data are missing in P. vivax infected individuals. From a total of 46 retrospective samples, molecular analysis revealed the presence of two most common polymorphic markers, 1311 C>T and IVSXI 93T>C mutations. Some studies suggested that the presence of these two mutations in an individual could lower G6PD enzyme activity. Besides, two novel mutations, 1388 G>T and 1398 C>T were also found. The bioinformatic studies showed a higher SIFT and higher MutPred2 score suggesting deleterious nature to the encoded protein leading to a greater propensity to be pathogenic. The limitations of the study include the inability to perform G6PD phenotypic profiling for determination of G6PD-d status as the majority of samples were dried blood spots and also due to small sample size in this study.

Youseuf Suleiman (University of Liverpool, UK) addressed the need for accurate and reliable markers for tick identification through his talk. Challenges included scarcity of genomic data, suitable markers, and the presence of cryptic species with similar morphological features that might lead to misidentification. While a complete nuclear genome is a gold standard, their large genomic sizes (1-7 Gb) have posed difficulties with assembly and annotation. Suleiman then introduced tick mitochondrial genomes (mitogenomes) with their considerably smaller sizes (14-17  Kb), high copy number, and maternal inheritance which was ideal for accurate and reliable taxonomic differentiation. With this, complete mitochondrial sequences of nine tick cell lines derived from Amblyomma, Rhipicephalus, Hyalomma, and Ixodes spp., sourced from the Tick Cell Biobank were produced. Additionally, the first mitogenome of Amblyomma variegatum was produced. From the data, Suleiman could reliably place Amblyomma variegatum as sister to Amblyomma hebraeumand corroborate the taxonomic placement of the tick cell lines. The study highlighted the significance of mitochondrial genomics in advancing the understanding of tick taxonomy and evolutionary relationships.

Pedro Ferreira (Life and Health Sciences Research Institute, Portugal) discussed the severity of cerebral malaria (CM) and the lack of a comprehensive research approach for studies. Thereafter, Ferreira proposed an in vitro system by using cellular approaches to identify molecular alterations in human brain vasculature cells that resemble the blood-brain barrier in CM. Induced pluripotent stem cells (iPSCs) culture was used to generate the brain organoids, thereafter, embryoid bodies formed after 6 days of spontaneous differentiation. After 11 days, neurospheres were formed using a neural induction medium. Finally, the brain organoids were ready after 30 days of post-differentiation. Through transcriptomic analysis, the specific gene expression profiles in human brain microvascular endothelial cells (HBMEC) activated by P. falciparum were characterized. From there, the impact of HBMEC-P. falciparum-activated secretomes were evaluated in human brain organoids which showed induced specific and significant alterations in transcriptome, and the slowing down of normal development in brain organoids. This model is novel in investigating factors influencing CM contributing to understanding pathogenesis, brain injury, and dysfunction.

Ase Berg (Stavanger University Hospital, Norway) started off by raising awareness about the onset of malaria before proceeding to introduce circulating markers of extracellular matrix (ECM). Berg explained that the ECM has roles in wound healing, and it contributes to constructive regrowth, without which, scarring and fibrosis occur. A clinical study was done with 205 participants with P. falciparum infection from 2017 to 2020. The circulating levels of six ECM remodeling mediators (KL 40, CysB, ENRAGE, CXCL 16, GDF 15, and GAL-3) were quantified by enzyme immunoassay. Results showed that all six components had significantly elevated readings with levels increasing with malaria severity in severe and complicated malaria compared to uncomplicated malaria. Berg added that the increasing levels could even predict death. From further exploration of different ECM parameters versus severity criteria, hyperparasitemia triggered all of the ECM remodeling mediators. Cerebral malaria and renal failure showed elevated levels for all except YKL 40, liver failure with heightened ENRAGE and GAL-3, severe anemia with GDF 15, and hemolysis with GDF 15 and GAL-3.

I Made Susila Utama (Prof Ngoerah Hospital, Indonesia) discussed the recent malaria characteristics at Prof Ngoerah Hospital, Bali. Utama raised the awareness that although Bali was malaria-free, it was still at risk of contracting malaria from travelers. As such, a descriptive study was done in the hospital from 2019 to 2023 regarding patient characteristics, type of Plasmodium, severity of infection and treatment outcome. From this, 37 cases were treated, 78.4% (29) being male patients from which up to 24.32% (9) had severe malaria but none died. P. vivax infections were the highest, 51.35%, followed closely by P. falciparum infections, 45.94%, and the lowest being P. malariae (2.71%). From severe malaria manifestations, jaundice was the highest occurrence at 77.77%, followed by renal dysfunction at 44.4%. The current treatment for uncomplicated malaria included treatment with 3 days dehydroartemycine (DHP) and primaquine (PQ) for 1 or 14 days. For severe malaria and P. vivax infections, artesunate IV was added into the treatment regime. Finally, for the treatment of P. malariae, artesunate IV and 3 days DHP were administered without the inclusion of PQ.

Symposium P08 – Introducing and Scaling up Emerging Diagnostic Solutions for Malaria Case Detection and Management

Win Han Oo (Burnet Institute, Australia) gave an account of the challenges in malaria case detection, pre-existing limitations, and emerging threats in the Asia-Pacific region. Current malaria diagnostic tools include clinical diagnosis, microscopy, rapid diagnostic tests (RDTs), and molecular methods. However, there are specific challenges in the Asia-Pacific region, such as subclinical malaria, which can represent a significant burden due to parasite reservoirs that often go undetected and untreated. Also, drug resistance and the need to detect and treat all malaria cases is another hurdle. Additionally, hrp2/3 gene deletions cause false negatives in malaria RDTs. Detecting hypnozoites in P. vivax is also challenging due to its dormant asymptomatic liver-stage form, to address this, Han Oo introduced the G6PD test, which has proven to be an effective screening tool. All these challenges required the need for a robust quality assurance framework and the need for additional data for comparative performance. He drew attention to how critical case detection is with effective diagnostics, as timely and accurate treatment is necessary to detect and eliminate residual malaria transmission. He concluded by alluding to the limitations of current diagnostic tools at the programmatic level which warrants investment in research for continuous development.

Allison Golden (PATH, USA) discussed new diagnostic tools for case detection and management of malaria touching on the ongoing challenges in diagnosing malaria infections using microscopy and rapid diagnostic tests (RDTs). In P. falciparum detection, the emergence of hrp2/3 gene deletions compromises RDT performance, requiring improvements in sensitivity. Analytical testing of malaria using next-generation RDTs can provide comparative performance data to evaluate RDT results by benchmarking and running tests with panels, collecting data, and analyzing results for evidence and input. Golden simulated the clinical sensitivity of RDTs using laboratory analytical data compared to WHO-prequalified RDTs and demonstrated that improved RDTs will address hrp2/3 gene deletions and provide enhanced sensitivity for PfLDH and PvLDH. Quantitative and semi-quantitative point-of-care G6PD tests are also currently available. Golden concluded that high-sensitivity P. vivax diagnostics alongside  G6PD diagnostic tools can provide access to radical cures.

Yu Nandar Aung (Independent Researcher, Myanmar) analyzed the health economic considerations and value proposition for new diagnostic products to support malaria control and elimination. Malaria is one of hundreds of diseases contributing to the global burden of disease. Deciding its importance when there are competing priorities is based on value for money and affordability. Aung conducted a cost-effectiveness analysis to assess the budget impact of malaria burdens. She also used a case study from Lao PDR to understand the cost-effectiveness of G6PD testing for Plasmodium vivax radical cure and its budgetary implications. A decision tree model was used to evaluate the effectiveness of radical cure using primaquine (PMQ) under unsupervised and supervised conditions. The results showed that the supervised intervention was the costliest but also the most effective method. The G6PD quantitative test was found to be cost-effective in the setting of Laos. However, rolling out the G6PD test to all health facilities would be expensive and have a limited impact, while a selective rollout based on P. falciparum caseload would have the highest impact on the budget.

Van Ahn Ngo Thi (Health Poverty Action, Vietnam) spoke about the challenges and opportunities for the introduction of new products for malaria elimination in Vietnam, a country with low malaria burden, targeting Plasmodium vivax that contributes to most of the malaria cases in the country. It was stated that the number of malaria cases in Vietnam in the last decade had reduced significantly with only 17 out of 46 provinces yet to achieve elimination. To help Vietnam achieve its elimination target, Thi and his team launched a project that covered eight provinces, targeting endemic and border populations near these areas. She stated that the project conducted training for health staff to detect active cases and conduct G6PD testing which enables safe and effective P. vivax treatment with primaquines. Within one year they successfully achieved over 70% coverage of their targeted population with the G6PD testing. She mentioned the limited opportunity for application due to low endemicity as one of the challenges for implementing new products such as G6PD in non-endemic regions. She recommended routine training on the operating procedure of new products to mitigate the challenge. 

Soy Ty Kheang (Health and Social Development, Cambodia) introduced new diagnostic tools for the radical cure of P. vivax in G6PD-d patients and shared lessons learned from Cambodia. The case study in Cambodia focused on equal access to safe radical cures. P. vivax, the predominant malaria species in Cambodia accounting for about 90% of cases, and a relatively high prevalence of G6PD deficiency. Key implementers are health centers (HCs) and village malaria workers (VMWs), who perform malaria diagnostic using diagnostic tests (RDTs) and provide first-line malaria treatment with artesunate-mefloquine (ASMQ). Patients detected by VMWs were referred to HCs. The survey enrolled over 1,000 P. vivax cases in target HCs from 2019 to 2020, with over 600 participants taking the G6PD test. The study reported over 100 cases of G6PD deficiency among the participants. Treatment with ASMQ from the survey led to a significant drop in hemoglobin levels by more than 25% after one week of the regimen, compared to patients on a two-weeks regimen. Therefore, the study proposed a follow-up one week into an eight-week ASMQ treatment. The health facility-community care model for P. vivax radical cure, including patients with G6PD deficiency, proved to be feasible since the uptake improved quickly after implementation.

Plenary 6 – Genomic Epidemiology and Phenotypic Diversity of Malaria Parasites 

David Conway (London School of Hygiene and Tropical Medicine – LSHTM, UK) presented the genomic epidemiology and phenotypic diversity of malaria parasites, noting the significant rise in malaria genomics publications over the past 47 years. He explained that advances in genome sequencing have greatly improved the understanding of molecular infections and cellular processes, contributing to a deeper understanding of the biology and genomics of malaria, in addition to the expansion of population genomic studies. Conway highlighted the variation in guanine-cytosine (GC) content and codon usage among malaria species, emphasizing their influence on genomic traits. He pointed out that chemogenomics is now being used to identify potential targets for antimalarial compounds. He also described how single-cell transcriptomics has provided detailed insights into parasite development, particularly through the expression of Gametocyte Development 1 (GDV1), which promotes sexual commitment and increases the expression of the antigen MSPDBL2. Conway mentioned that the discovery of genomic sequences with multiple copies has enhanced DNA detection which improves diagnostic accuracy. However, he raised concerns over the emergence of hrp2/3 gene deletions, which compromises the effectiveness of rapid diagnostic tests (RDTs), alongside the rise of partial artemisinin resistance. He further explained that genomic studies have uncovered multiple sources of Plasmodium falciparum resistance to artemisinin, stressing the need for population genomic analysis to distinguish subpopulations. Additionally, Conway reported that P. vivax has a distinct genomic structure, with SNPs increasing the resolution of sub-populations. Finally, he pointed out that ancient parasite genomes provide insights into historical malaria in Europe, and that Plasmodium knowlesi has been divided into three genetic populations. He stressed on the necessity of studying both phenotype and genotype together, which could reveal critical information about parasite evolution and resistance development.

Symposium P23 – Innovative biocontrol approaches for mosquito-borne diseases elimination: Role and factors for success.

Dickson Wilson Lwetoijera (Ifakara Health Institute, Tanzania) raised awareness about worldwide malaria cases. He stated that despite the death rates dropping by approximately 20%, the number of cases in the world still remained more or less unchanged for more than two decades. The progress for malaria eradication had started to reverse because of the impact of climate change, insecticide resistance, urban malaria vectors, humanitarian crisis, insufficient compliance, and others, so there is a need for novel tools for malaria eradication. This is where engineered gene drives come into play for population modification to propagate antimalarial effects. However, its development and implementation require sustained capacity building with regard to African stakeholders. Lwetoijera then elaborated on transmission zero for population modification via mosquito gene drives the first transgenic mosquito made in Africa by Transmission Zero, the global malaria research programme developing genetic vector control strategies. The modification gene drive technology comprises separate strains for effector and drive functions where the effector trait was responsible for blocking Plasmodium transmission and the driver for propagating itself and effector in mosquito populations.

Leo Barack (Asia Pacific Malaria Elimination Network – APMEN, Singapore) emphasized the role of regional leadership as being essential to provide long-lasting, affordable, and sustainable solutions like gene drives. These novel approaches in regional malaria elimination should include adaptability, scalability, application, cultural sensitivity, and compatibility with context in countries. He highlighted the role of regional leadership being pivotal in making this happen via the promotion of research and development through defined research priorities, resource, and funding allocation. Besides, regional leadership is also crucial in advocacy for policies in terms of its development and implementation as well as establishing regulatory frameworks. Lastly, he stressed the role of coordination and collaboration in determining multisectoral efforts with integration along with joint initiatives and shared knowledge to create platforms for further advancement of research.

Mouhamed Drabo (Imperial College London, United Kingdom) discussed strategies for gaining legitimacy with local communities in research projects. Target Malaria is a non-profit research association consisting of seven institutions from Africa, Europe, and North America. Their goal is to develop and share new, cost-effective, and sustainable genetic technologies to modify mosquitoes and reduce malaria transmission. The principles for engaging with Target Malaria prioritize reaching out to the most relevant groups, involving project representatives in the engagement process, and starting the engagement early while maintaining it consistently. In order to gain community trust, the project is committed to being open and accountable, undergoing external review, sharing relevant study results with stakeholders, and building community knowledge about malaria. Additionally, the project aims to conduct power analyses, establish active and sustainable community participation, and ensure transparency and effective communication. At the same time, it intends to maintain permanent dialogue, create partnerships with local leaders and community organizations, and establish feedback processes. 

Cheong Huat Tan (National Environment Agency Singapore, Singapore) shared his experience with the Wolbachia project implemented in Singapore. The project aimed to reduce the mosquito population to a level that inhibits disease transmission. It was carried out in several phases in areas with a high risk of dengue and a high population of Aedes aegypti mosquitoes. A guidance framework for the sterile insect technique was established to assist in decision-making, testing, and implementation. He mentioned that the assessment of using Wolbachia to suppress Aedes aegypti should consider effectiveness, safety, and engagement with leaders, experts, and the public collectively. Tan further explained that strong collaborative networks between government agencies, academia, research partners, and industry are essential. To support the implementation of the project, it is important to leverage established infrastructure and networks. The release of male Aedes mosquitoes should be tailored to the local environment. Implementing data-driven strategies enables informed decision-making, optimizes strategies, measures effectiveness, and enhances both cost-effectiveness and scalability.

Symposium P20 – Accelerating towards zero vivax malaria – The Vivax Serology Partnership (VISPA)

Rhea Longley (Walter and Eliza Hall Institute of Medical Research – WEHI, Australia) explained the use of Plasmodium vivax serological exposure markers to uncover hidden malaria infections. Longley highlighted that resistance to traditional malaria control measures in P. vivax facilitated by hidden liver-stage hypnozoites remain a challenge for malaria elimination. Longley and her team explored the ability of serological markers to detect past and present infections in addition to detecting the possibility of recurrent infection. She stated that the prospects in serological markers led to the establishment of vivax serology partnership (VISPA), which aims to provide evidence to introduce serology-based identification that will help in integrating drug treatment interventions into global national policy and develop novel diagnostic tools based on their serological tests.

Rahmat Sagara (Oxford University Clinical Research Unit – OUCRU, Indonesia) discussed his efforts to quantify the impact of novel intervention strategies aimed at accelerating Plasmodium vivax elimination in the high-malaria-endemic region of Papua, Indonesia. He reported that Papua has the highest number of malaria cases in Indonesia. Sagara outlined Indonesia’s malaria program strategy, which includes universal access to healthcare, surveillance, policy development, and empowering communities. He explained that multiple interventions are being used to eliminate the malaria burden, as part of an acceleration plan. Mathematical modeling was employed to estimate the impact of expanding malaria control efforts which included simulations of a range of intervention scenarios using serological diagnostic tools that quantified their impact, identified the optimal deployment strategies, and determined the required levels of G6PD testing. Based on his analysis, Sagara estimated the death rate, maximum age, and the entomological inoculation rate (EIR) at equilibrium. He concluded that, according to Indonesia’s elimination plan, the country is expected to be malaria-free by 2030.

Rintis Noviyanti (National Research and Innovation Agency – BRIN, Indonesia) reported on her study using serological markers to predict Plasmodium vivax relapses in a cohort of returning Indonesian soldiers. She explained that malaria elimination efforts are hindered by P. vivax due to its ability to form asymptomatic, dormant hypnozoites in the liver, which can reactivate and cause relapses. Noviyanti’s primary objective was to validate a sero-diagnostic test to identify soldiers who were carrying these dormant forms of the parasite. She mentioned two battalions of soldiers from Surabaya and Malang, deployed to the malaria-endemic region of Papua, who were involved in the study. According to her findings, some soldiers, particularly from the Malang battalion, experienced up to three malaria relapses within a six-month period. Plasma samples from these soldiers were analyzed using Luminex assays to detect antibody responses to P. vivax proteins. Noviyanti reported that a total of 592 samples were initially collected, with 553 remaining viable for analysis. She highlighted that median antibody titers for eight P. vivax biomarkers were measured over time, comparing those who relapsed to those who did not. Noviyanti concluded that these serological markers have the potential to serve as a novel public health intervention, aiding malaria elimination efforts by identifying individuals at risk of relapse.

Ayleen Alicia Kosasih (Oxford University Clinical Research Unit – OUCRU, Indonesia) presented the efficacy of serological screening and treatment to prevent Plasmodium vivax incidence among school children in North Sumatra, Indonesia in a randomized-controlled trial. Detection and treatment remain pivotal for the control and elimination of malaria in both endemic and non-endemic regions. This led to their study that evaluated the effectiveness of serological test and treatment (PvSeroTAT) intervention over a period of a year, with the aim of reducing malaria incidence among school children in the country. The study enrolled over 1,000 participants and the method was found to effectively treat P. vivax seropositive subjects and interestingly reduced the rate of recurrent parasitemia by over 50%. In addition, Kosasih pointed out that the method predicted children who were more likely to have recurrent P. vivax infection. She concluded that the intervention can help reduce recurrent P. vivax infection and reiterated that scaling up the intervention will help in halting malaria transmission in the region.

Rachael Farquhar (Burnet Institute, Australia) described the efforts to develop evidence-based policy and guidelines for the introduction of Plasmodium vivax serological testing and treatment strategies in the Asia Pacific region. She highlighted P. vivax as a major obstacle to reaching the 2030 malaria elimination goals in the region. Farquhar explained that the vivax serology partnership (VISPA) is collaborating with stakeholders to support National Malaria Control Programs (NMCPs) in implementing P. vivax serological testing and treatment (PvSeroTAT). Interviews and focus group discussions are being conducted to assess the feasibility and readiness of health systems for PvSeroTAT in two scenarios: diagnosing relapse risk to guide radical cure and population-based screening for recent infections. Farquhar shared initial insights from surveys conducted after the 2023 APMEN vivax working group meeting, which aimed to identify barriers to serologically guided treatment at both the health system and community levels. VISPA has been implemented in several countries, including Laos, Cambodia, the Philippines, and Latin American nations. She underlined that the findings will help inform the communication toolkit for NMCPs to enhance engagement with PvSeroTAT strategies.