Exploring community participation in vectorborne disease control in Southeast Asia: a scoping review protocol.

INTRODUCTION: Vector borne diseases (VBDs) present significant public health challenges in Southeast Asia (SEA), and the increasing number of cases threatens vulnerable communities. Inadequate vector control and management have been linked to the spread of VBDs. To address these issues, community participation has been proposed as a promising approach to enhance health programmes and control of VBDs. This article outlines a protocol for a scoping review of the published literature on community-participation approaches to control VBDs in the SEA region. The primary research question is 'How does community participation complement the control of VBDs in SEA?' This review aims to provide an overview of various approaches and identify barriers and facilitators to effective implementation. METHODS AND ANALYSIS: The research questions will guide the scoping review. In stage 1, peer-reviewed publications from PubMed, Web of Science and Scopus will be searched using predefined search terms related to community-based approaches and VBDs in the SEA region, English, Indonesian and Malay published between 2012 and 2022. In stage 2, the references from relevant articles will be screened for eligibility. In stage 3, eligible articles will be charted in Microsoft Excel to facilitate the review process, and studies will be characterised based on the investigated diseases; this review will also highlight the methodological context of these studies. In stage 4, a thematic analysis will be conducted to derive meaningful findings from the dataset relevant to the research inquiry, followed by writing the results in stage 5. This scoping review aims to be the first to explore community participation in VBD control in the SEA population, providing valuable insights for future research and stakeholders involved in disease control. ETHICS AND DISSEMINATION: This scoping review does not require ethical approval because the methodology synthesises information from available articles. This review is planned for dissemination in academic journals, conference presentations and shared with stakeholders as part of knowledge sharing among those involved in VBD control.

Origin of the human malaria parasite Plasmodium vivax.

The geographic origin of Plasmodium vivax, a leading cause of human malaria, has been the subject of much speculation. Here we review the evolutionary history of P. vivax and P. vivax-like parasites in humans and non-human primates on three continents, providing overwhelming evidence for an African origin. This conclusion is consistent with recent reports showing that Duffy-negative humans in Africa are, in fact, susceptible to P. vivax, with parasites invading Duffy-antigen-expressing erythroid precursors. Thus, the African origin of P. vivax not only explains the distribution of the Duffy-negative genotype but also provides new insight into the history and status of P. vivax malaria in Africa and efforts geared toward its eradication.

A smartphone-based crowd-sourced real-time surveillance platform (apple snail inspector) for the invasive snails: a design and development study.

BACKGROUND: The large amphibious freshwater apple snail is an important invasive species in China, but there is currently no method available for their surveillance. The development and popularization of smartphones provide a new platform for research on surveillance technologies for the early detection and effective control of invasive species. METHODS: The ASI surveillance system was developed based on the infrastructure of the WeChat platform and Amap. The user can directly enter the game interface through the WeChat port on their mobile phone, and the system automatically obtains their location. The user can then report the location of apple snails. The administrator can audit the reported information, and all information can be exported to Microsoft Excel version 2016 for analysis. The map was generated by ArcGIS 10.2 and was used to characterize the spatial and temporal distribution of apple snails in Jiangsu Province. RESULTS: The architecture of ASI consists of three parts: a mobile terminal, a server terminal and a desktop terminal. We published more than 10 tweets on the official WeChat account of the system to announce it to the public, and a total of 207 users in 2020 and 2021 correctly reported sightings of apple snails. We identified 550 apple snails breeding sites in 2020 and 2021, featuring ponds (81%), parks (17%) and farmland (2%). In addition, most of the locations contained snail eggs, and the reporting times mainly occurred between May and September. CONCLUSIONS: The ASI is an effective surveillance system that can be used to identify the breeding locations of apple snails and provides the basis of prevention and control for its dispersal. Its successful development and operation provide new potential avenues for surveillance of other public health issues.

Knowledge, compliance, and challenges in anti-malarial products usage: a systematic review of at-risk communities for zoonotic malaria.

BACKGROUND: Zoonotic malaria is a growing public health threat in the WHO Southeast Asia (SEA) and Western Pacific (WP) regions. Despite vector-control measures, the distribution of Macaque fascicularis and M. nemestrina, and Anopheles mosquitoes carrying non-human simian malaria parasites poses challenges to malaria elimination. The systematic review assesses the literature on knowledge and malaria-preventive practices in zoonotic malaria-affected areas across the WHO SEA and WP, aiming to identify challenges for malaria control. METHODS: Peer-reviewed articles published in English, Malay and Indonesian between January 2010 and December 2022 were searched in OVID Medline, Scopus, Web of Science, and Google Scholar. Studies of any design-excluding reviews, conference proceedings, and reports from all WHO SEA and WP countries vulnerable to zoonotic malaria-were included. Backwards-reference screening and thematic analysis were conducted. RESULTS: Among 4,174 initially searched articles, 22 peer-reviewed articles met the inclusion criteria. An additional seven articles were identified through backwards-reference screening, resulting in a total of 29 articles for this review. Half of these studies were conducted in Cambodia, Myanmar, Malaysia, and Thailand, mainly in forests and remote communities. The review highlighted inconsistencies in the operationalization of knowledge, and five major themes were identified related to knowledge: causation and transmission, symptoms, treatment, severity and complications, and malaria prevention. While participants generally had some understanding of malaria causation/transmission, minority and indigenous ethnic groups demonstrated limited knowledge and held misconceptions, such as attributing malaria to drinking dirty water. Preventive practices included traditional and non-traditional or modern methods-with a preference for traditional approaches to avoid mosquito bites. Challenges to malaria control included feasibility, cost, and access to healthcare services. CONCLUSION: This review provides insights into knowledge, local understandings, and preventive practices related to malaria in the WHO SEA and WP regions. The findings highlight the need for future research to explore the knowledge of at-risk communities regarding zoonotic malaria, their perceive threat of the disease and factors exposing them to zoonotic malaria. New strategies must be developed for zoonotic malaria programs tailored to local contexts, emphasizing the significance of community participation, health education, and socio-behavioural change initiatives. It is important to consider the interconnectedness of human health, environmental and non-human primates conservation. Socio-cultural nuances should also be carefully considered in the design and implementation of these programs to ensure their effect tailored to local contexts.

The two parasite species formerly known as Plasmodium ovale.

Plasmodium ovale was the last of the exclusively human malaria parasites to be described, in 1922, and has remained the least well studied. Beginning in 1995, two divergent forms of the parasite, later termed 'classic' and 'variant', were described. By 2010, it was realised that these forms are two closely related, but genetically distinct and non-recombining species; they were given the names Plasmodium ovale curtisi and Plasmodium ovale wallikeri. Since then, substantial additional data have confirmed that the two parasites are indeed separate species, but the trinomial nomenclature has often led to confusion about their status, with many authors describing them as subspecies. We hereby formally name them Plasmodium ovalecurtisi and Plasmodium ovalewallikeri.

Role of pre-existing immunity in driving the dengue virus serotype 2 genotype shift in the Philippines: A retrospective analysis of serological data.

OBJECTIVE: The invasion of dengue virus (DENV)-2 Cosmopolitan genotype into the Philippines, where the Asian II genotype previously circulated challenges the principle of dengue serotype-specific immunity. Assessment of antibodies in this population may provide a mechanistic basis for how new genotypes emerge in dengue-endemic areas. METHODS: We evaluated the neutralizing antibody (nAb) and antibody-dependent enhancement (ADE) responses against the two genotypes using archived serum samples collected from 333 patients with confirmed dengue in Metro Manila, Philippines, before, during, and after the introduction of the Cosmopolitan genotype. We quantified nAb titers in baby hamster kidney (BHK-21) cells with or without the Fcγ receptor IIA (FcγRIIA) to detect the capacity of virus-antibody complexes to neutralize or enhance DENV. RESULTS: The nAb potency of the archived serum samples against the two genotypes was greatly affected by the presence of FcγRIIA. We found significant differences in nAb titers between the two genotypes in BHK-21 cells with FcγRIIA (P <0.0001). The archived serum samples were incapable of fully neutralizing the Cosmopolitan genotype, but instead strongly promoted its ADE compared to the Asian II genotype (P <0.0001). CONCLUSION: These results reinforce the role of pre-existing immunity in driving genotype shifts. Our finding that specific genotypes exhibit differing susceptibilities to ADE by cross-reactive antibodies may have implications for dengue vaccine development.

Time to drop the term 'simian malaria parasites'.

The term 'simian malaria parasites' has crept into the modern malaria literature as a synonym for 'non-human primate malaria parasites', most commonly referring to species of Plasmodium infective to Old World monkeys. As humans are also simians, we contend that this usage is erroneous, and should not be used.

Disentangling the intersection of inequities with health and malaria exposure: key lessons from rural communities in Northern Borneo.

BACKGROUND: The increasing incidence of Plasmodium knowlesi malaria poses a significant challenge to efforts to eliminate malaria from Malaysia. Macaque reservoirs, outdoors-biting mosquitoes, human activities, and agricultural work are key factors associated with the transmission of this zoonotic pathogen. However, gaps in knowledge regarding reasons that drive malaria persistence in rural Kudat, Sabah, Northern Borneo remain. This study was conducted to address this knowledge gap, to better understand the complexities of these entangled problems, and to initiate discussion regarding new countermeasures to address them. This study aims to highlight rural community members' perspectives regarding inequities to health relating to P. knowlesi malaria exposure. METHODS: From January to October 2022, a study using qualitative methods was conducted in four rural villages in Kudat district of Sabah, Malaysia. A total of nine in-depth interviews were conducted with community and faith leaders, after the completion of twelve focus group discussions with 26 photovoice participants. The interviews were conducted using the Sabah Malay dialect, audio-recorded, transcribed, and translated into English. The research team led the discussion and analysis, which was approved by participants through member checking at the community level. RESULTS: Participants identified disparity in health as a key issue affecting their health and livelihoods. Injustice in the social environment was also identified as a significant challenge, including the importance of listening to the voices of affected communities in disentangling the social and economic phenomena that can impact malaria control. Specific concerns included inadequate access to health-related resources and degradation of the environment. Participants recommended improving access to water and other necessities, increasing the availability of malaria control commodities in healthcare facilities, and developing sustainable programs to reduce socioeconomic disparities. CONCLUSION: Inequities to health emerged as a key concern for malaria control in rural Kudat, Sabah. A locally targeted malaria programme cantered on improving the social and economic disparities associated with health outcomes, could be a potential strategy for malaria prevention in such areas. Community-level perspectives gathered from this study can be used as a foundation for future discussions and dialogues among policymakers and community members for achieving greater transparency, improving social equity, and interoperability in addressing P. knowlesi malaria control.

The public health response to a Plasmodium malariae outbreak in Penampang district, Sabah during a COVID-19 movement control order.

BACKGROUND: Since 2018, no indigenous human malaria cases has been reported in Malaysia. However, during the recent COVID-19 pandemic the World Health Organization is concerned that the pandemic might erode the success of malaria control as there are reports of increase malaria cases in resource limited countries. Little is known how the COVID-19 pandemic has impacted malaria in middle-income countries like Malaysia. Here the public health response to a Plasmodium malariae outbreak occurred in a village in Sabah state, Malaysia, during a COVID-19 movement control order is reported. METHODS: An outbreak was declared following the detection of P. malariae in July 2020 and active case detection for malaria was performed by collecting blood samples from residents residing within 2 km radius of Moyog village. Vector prevalence and the efficacy of residual insecticides were determined. Health awareness programmes were implemented to prevent future outbreaks. A survey was conducted among villagers to understand risk behaviour and beliefs concerning malaria. RESULTS: A total of 5254 blood samples collected from 19 villages. Among them, 19 P. malariae cases were identified, including the index case, which originated from a man who returned from Indonesia. His return from Indonesia and healthcare facilities visit coincided with the movement control order during COVID-19 pandemic when the healthcare facilities stretched its capacity and only serious cases were given priority. Despite the index case being a returnee from a malaria endemic area presenting with mild fever, no malaria test was performed at local healthcare facilities. All cases were symptomatic and uncomplicated except for a pregnant woman with severe malaria. There were no deaths; all patients recovered following treatment with artemether-lumefantrine combination therapy. Anopheles balabacensis and Anopheles barbirostris were detected in ponds, puddles and riverbeds. The survey revealed that fishing and hunting during night, and self-treatment for mild symptoms contributed to the outbreak. Despite the index case being a returnee from a malaria-endemic area presenting with mild fever, no malaria test was performed at local healthcare facilities. CONCLUSION: The outbreak occurred during a COVID-19 movement control order, which strained healthcare facilities, prioritizing only serious cases. Healthcare workers need to be more aware of the risk of malaria from individuals who return from malaria endemic areas. To achieve malaria elimination and prevention of disease reintroduction, new strategies that include multisectoral agencies and active community participation are essential for a more sustainable malaria control programme.

Molecular Detection and Identification of Piroplasm in Cattle from Kathmandu Valley, Nepal.

BACKGROUND: Tick-borne protozoan parasites (TBPPs) cause significant problems for domestic animals' health in Nepal. TBPPs are routinely diagnosed by labor-intensive blood smear microscopy. In Nepal, there are some reports of Babesia and Theileria in cattle, although species identification is rarely performed. Therefore, we performed conventional nested PCR (nPCR) followed by sequence analysis to identify TBPP species infecting cattle in Nepal. METHODS: One hundred and six blood samples were collected from cattle in the Kathmandu Valley. Thin blood smears were prepared for microscopic examination. Parasite DNA was extracted from the blood, and nPCR and sequencing were performed to identify the TBPPs present. RESULTS: Among the 106 samples, 45 (42.5%) were positive for piroplasm (Babesia spp. and Theileria spp.) via microscope observation and 56 (52.8%) samples were positive via nPCR. The obtained PCR products were used for direct sequencing, and we identified the species as B. bigemina, B. bovis, T. annulate and T. orientalis. Phylogenetic analyses showed that the B. bovis, B. bigemina and T. orientalis sequences from this study belonged to each species clade. On the other hand, T. annulate was divided into two clades in the analysis, and our T. annulate sequences were also divided in these two clades. The piroplasm-positive cattle showed lower hemoglobin and red blood cells than healthy cattle. CONCLUSIONS: To the best of our knowledge, this study is the first to apply molecular detection and species determination of TBPPs in cattle in Nepal. The results of this study may be used as a starting point for the development of successful TBPP surveillance and prevention programs in Nepal.

Exploring barriers to and facilitators of malaria prevention practices: a photovoice study with rural communities at risk to Plasmodium knowlesi malaria in Sabah, Malaysia.

BACKGROUND: The control of Plasmodium knowlesi malaria remains challenging due to the presence of macaque monkeys and predominantly outdoor-biting Anopheles mosquitoes around human settlements. This study aims to explore the barriers and facilitators related to prevention of mosquito bites among rural communities living in Sabah, Malaysia using the participatory visual method, photovoice. METHODS: From January through June 2022, 26 participants were recruited from four villages in Kudat, Sabah, using purposive sampling. Participants were male and female villagers, aged > 18 years old. After photovoice training in the villages, participants documented facilitators of and barriers related to avoiding mosquito bites using their own smartphone cameras, and provided narratives for their photos. Twelve Focus Group Discussions (FGDs) sessions in three rounds were held to share and discuss the photos, and to address challenges to the avoidance of mosquito bites. All discussions were conducted in the Sabah Malay dialect, and were video and audio recorded, transcribed, and analyzed using reflexive thematic analysis. The Ideation Model, a meta-theoretical model of behaviour change, underpinned this study. RESULTS: The most common types of barriers identified by participants included (I) intrapersonal factors such as low perceived threat of malaria, (II) livelihood and lifestyle activities consisting of the local economy and socio-cultural activities, and (III) physical and social environment. The facilitators were categorized into (I) intrapersonal reasons, including having the opportunity to stay indoors, especially women who are housewives, (II) social support by the households, neaighbours and healthcare workers, and (III) support from healthcare services and malaria awareness program. Participants emphasized the importance of stakeholder's support in implementing feasible and affordable approaches to P. knowlesi malaria control. CONCLUSION: Results provided insights regarding the challenges to preventing P. knowlesi malaria in rural Kudat, Sabah. The participation of communities in research was valuable in expanding knowledge of local challenges and highlighting possible ways to overcome barriers. These findings may be used to improve strategies for zoonotic malaria control, which is critical for advancing social change and minimizing health disparities in malaria prevention.

Seeing malaria through the eyes of affected communities: using photovoice to document local knowledge on zoonotic malaria causation and prevention practices among rural communities exposed to Plasmodium knowlesi malaria in Northern Borneo Island.

BACKGROUND: Many rural communities in Malaysian Borneo and Southeast Asia are at risk of Plasmodium knowlesi malaria. Multiple factors contribute to infection, however, a deep understanding of illness causation and prevention practices among at-risk communities remains limited. This study aims to document local knowledge on malaria causation and preventive practices of rural communities in Sabah, Malaysia, using photovoice-a participatory research method. METHODS: From January to June 2022, a photovoice study was conducted with rural communities in Matunggong subdistrict, Malaysia, to explore their experiences with and local knowledge of non-human primate malaria and prevention practices. The study included (1) an introductory phase in which participants were introduced to the photovoice method; (2) a documentation phase in which participants captured and narrated photos from their communities; (3) a discussion phase in which participants discussed photos and relevant topics through a series of three focus group discussions (FGDs) per village; and (4) a dissemination phase where selected photos were shared with key stakeholders through a photo exhibition. A purposively selected sample of 26 participants (adults > 18 years old, male, and female) from four villages participated in all phases of the study. The study activities were conducted in Sabah Malay dialect. Participants and the research team contributed to data review and analyses. RESULTS: Rural communities in Sabah, Malaysia possess local knowledge that attributes non-human primate malaria to natural factors related to the presence of mosquitoes that bite humans and which carry "kuman-malaria" or malaria parasite. Participants revealed various preventive practises ranging from traditional practises, including burning dried leaves and using plants that produce foul odours, to non-traditional approaches such as aerosols and mosquito repellents. By engaging with researchers and policymakers, the participants or termed as co-researchers in this study, showcased their ability to learn and appreciate new knowledge and perspectives and valued the opportunity to share their voices with policymakers. The study successfully fostered a balance of power dynamics between the co-researchers, research team members and policymakers. CONCLUSION: There were no misconceptions about malaria causation among study participants. The insights from study participants are relevant because of their living experience with the non-human malaria. It is critical to incorporate rural community perspectives in designing locally effective and feasible malaria interventions in rural Sabah, Malaysia. Future research can consider adapting the photovoice methodology for further research with the community toward building locally tailored-malaria strategies.

The diversity of Plasmodium falciparum isolates from asymptomatic and symptomatic school-age children in Kinshasa Province, Democratic Republic of Congo.

BACKGROUND: Understanding Plasmodium falciparum population diversity and transmission dynamics provides information on the intensity of malaria transmission, which is needed for assessing malaria control interventions. This study aimed to determine P. falciparum allelic diversity and multiplicity of infection (MOI) among asymptomatic and symptomatic school-age children in Kinshasa Province, Democratic Republic of Congo (DRC). METHODS: A total of 438 DNA samples (248 asymptomatic and 190 symptomatic) were characterized by nested PCR and genotyping the polymorphic regions of pfmsp1 block 2 and pfmsp2 block 3. RESULTS: Nine allele types were observed in pfmsp1 block2. The K1-type allele was predominant with 78% (229/293) prevalence, followed by the MAD20-type allele (52%, 152/293) and RO33-type allele (44%, 129/293). Twelve alleles were detected in pfmsp2, and the 3D7-type allele was the most frequent with 84% (256/304) prevalence, followed by the FC27-type allele (66%, 201/304). Polyclonal infections were detected in 63% (95% CI 56, 69) of the samples, and the MOI (SD) was 1.99 (0.97) in P. falciparum single-species infections. MOIs significantly increased in P. falciparum isolates from symptomatic parasite carriers compared with asymptomatic carriers (2.24 versus 1.69, adjusted b: 0.36, (95% CI 0.01, 0.72), p = 0.046) and parasitaemia > 10,000 parasites/µL compared to parasitaemia < 5000 parasites/µL (2.68 versus 1.63, adjusted b: 0.89, (95% CI 0.46, 1.25), p < 0.001). CONCLUSION: This survey showed low allelic diversity and MOI of P. falciparum, which reflects a moderate intensity of malaria transmission in the study areas. MOIs were more likely to be common in symptomatic infections and increased with the parasitaemia level. Further studies in different transmission zones are needed to understand the epidemiology and parasite complexity in the DRC.

Plasmodium malariae: the persisting mysteries of a persistent parasite.

Plasmodium malariae is a 'neglected malaria parasite' in as much as the amount of research conducted on it pales into insignificance when compared to that pertaining to Plasmodium falciparum and Plasmodium vivax, its more notorious and pathogenic cousins. There has, however, been an increase in interest in this parasite over the past decade. Principally, this is because of the increasing use of sensitive molecular detection techniques that have revealed a wider than previously recorded prevalence in some regions (particularly in Africa), and high numbers of chronic, asymptomatic infections.

Ring-stage growth arrest: Metabolic basis of artemisinin tolerance in Plasmodium falciparum.

The emergence and spread of artemisinin-tolerant malaria parasites threatens malaria control programmes worldwide. Mutations in the propeller domain of the Kelch13 protein confer Plasmodium falciparum artemisinin resistance (ART-R). ART-R is linked to the reduced susceptibility of temporary growth-arrested ring-stage parasites, but the metabolic mechanisms remain elusive. We generated two PfKelch13 mutant lines via CRISPR-Cas9 gene editing which displayed a reduced susceptibility accompanied by an extended ring stage. The metabolome of ART-induced ring-stage growth arrest parasites carrying PfKelch13 mutations showed significant alterations in the tricarboxylic acid (TCA) cycle, glycolysis, and amino acids metabolism, pointing to altered energy and porphyrin metabolism with metabolic plasticity. The critical role of these pathways was further confirmed by altering metabolic flow or through chemical inhibition. Our findings uncover that the growth arrestment associated with ART-R is potentially attributed to the adaptative metabolic plasticity, indicating that the defined metabolic remodeling turns out to be the trigger for ART-R.

A song for the unsung: The relevance of Plasmodium vinckei as a laboratory rodent malaria system.

Rodent malaria parasites (RMPs) allow the study of malaria parasite biology across its entire life cycle through a vertebrate host and a mosquito vector under laboratory conditions. Among the four RMPs originally collected from wild thicket rats in sub-Saharan Central Africa and adapted to laboratory mice, Plasmodium vinckei has the largest geographical range and includes the largest number of sub-species, demonstrating its deep genetic diversity. Despite affording the same advantages as other RMP species and additionally displaying a large degree of phenotypic and genotypic diversity, P. vinckei has seen limited use in the laboratory. Here, we review the contribution of P. vinckei to our understanding of malaria and highlight the areas where it could offer an advantage over other RMP species in future studies.

A systematic review of asymptomatic Plasmodium knowlesi infection: an emerging challenge involving an emerging infectious disease.

BACKGROUND: In the last decade Plasmodium knowlesi has been detected in humans throughout South East Asia. The highest risk groups for this infection are males, adults and those performing forest-related work. Furthermore, asymptomatic cases of P. knowlesi malaria have been reported including among women and children. METHODS: Pubmed, Scopus and the Web of Science databases for literature describing asymptomatic P. knowlesi malaria published between 2010 and 2020 were searched. A systematic literature review was conducted to identify studies reporting the prevalence and incidence of laboratory confirmed asymptomatic P. knowlesi cases in humans, their clinical and demographic characteristics, and methods used to diagnose these cases. RESULTS: By analysing over 102 papers, thirteen were eligible for this review. Asymptomatic P. knowlesi infections have been detected in 0.03%-4.0% of the population depending on region, and infections have been described in children as young as 2 years old. Various different diagnostic methods were used to detect P. knowlesi cases and there were differing definitions of asymptomatic cases in these studies. The literature indicates that regionally-differing immune-related mechanisms may play a part on the prevalence of asymptomatic P. knowlesi. CONCLUSION: Differing epidemiological characteristics of asymptomatic P. knowlesi malaria in different regions reinforces the need to further investigate disease transmission mechanics. Effective public health responses to changes in P. knowlesi epidemiology require proactive intervention and multisectoral collaboration.

Generating Trust in Participatory Research on Plasmodium knowlesi Malaria: A Study with Rural Community Gatekeepers during the COVID-19 Pandemic.

BACKGROUND: Plasmodium knowlesi malaria is a zoonotic infection that affects rural communities in South East Asia. Although the epidemiology of the disease has been extensively researched, the voices of individuals within affected communities often go unheard. Here, we describe a study that explores the importance of gatekeepers in conducting research among rural communities, their perspectives on the challenges encountered when attempting to avoid malaria infection, and their views on participatory research. METHODS: Between 1 November 2021 and 28 February 2022, we conducted a study in Kudat district, Sabah, using a multi-method design. All participants consented to the study, which included health care workers (HCWs) (n = 5), community leaders (n = 8), and faith leaders (n = 1). We conducted interviews, transect walks, and observations with gatekeepers to ensure data trustworthiness. All interviews were conducted in the Sabah Malay dialect. The sessions were audio- and video-recorded, transcribed into English and analyzed using thematic analysis. RESULTS: Between 2017 and 2021, the number of cases of P. knowlesi malaria detected in humans ranged from 35 to 87 in villages under the care of the Lotong primary health care clinic. The challenges in controlling malaria include social norms, lifestyles, socioeconomic factors, environmental factors, and limitations of basic resources. Critical discussions regarding participation with the gatekeepers identified that face-to-face interviews were preferable to online discussions, and influenced willingness to participate in future research. CONCLUSION: This study was conducted among village gatekeepers during the COVID-19 pandemic and generated information to drive methodological changes, opening up new ideas by sharing perspectives on challenges in P. knowlesi malaria control among vulnerable communities. The study generated trust in the community and expanded knowledge regarding participation that is critical for future community-based studies.

Disparate selection of mutations in the dihydrofolate reductase gene (dhfr) of Plasmodium ovale curtisi and P. o. wallikeri in Africa.

Plasmodium ovale curtisi and P. ovale wallikeri are both endemic in sub-Saharan Africa, the Middle East and Southeast Asia. Molecular surveillance data for drug resistance in P. ovale spp. is limited at present. We analysed polymorphisms in the podhfr, pocrt and pocytb genes of P. ovale spp. in 147 samples collected from travelers returning to China from Africa. Two podhfr mutations, S58R and S113N/T were detected in P. ovale curtisi with high/moderate frequencies of 52.17% and 17.39%, respectively. Evidence of positive selection (dN/dS = 2.41) was found for podhfr in P. ovale curtisi and decreased diversity (He) of microsatellite markers flanking the mutant alleles suggests that selective sweeps have occurred for both. Mutations E34G (1.50%) and L43V (1.50%) in pocrt of P. ovale curtisi, and E34G (3.70%), I102M (1.80%) and V111F (1.80%) of P. ovale wallikeri were found at low frequencies. Mutations R66K (6.20%), R75K (11.63%) and R95K (3.88%) of pocytb were found in both P. ovale curtisi and P. ovale wallikeri. These results suggest that the podhfr gene of P. ovale curtisi may be subject to drug selection in Africa, warranting further attention. We observed significant differences in the prevalence and distribution of podhfr mutations between the two P. ovale species, suggestive of fundamental biological differences between them.