A Critical Enquiry into Variability of Insecticidal Net Use in Cambodia: Implications for Assessing Appropriateness of Malaria Elimination Interventions.

Distributing long-lasting insecticidal nets (LLINs) to individuals living in malaria-endemic regions is a cornerstone of global malaria control. National malaria control programs aim to achieve "universal coverage" of at-risk populations to reach LLINs' full potential to reduce malaria, progress of which is then measured by indicators constructed from standardized questionnaires. Through an exploration of variability in LLIN use in Cambodia, we argue that indicators of universal coverage of LLINs are not sufficiently commensurate with the realities they are intended to measure, limiting the suitability of the data to serve program and policy purposes in a malaria elimination era. Reflecting on the various sources of variability in LLIN use, we apply and extend the concept of "appropriateness" as a third prong to the widely used "efficacy" and "effectiveness" criteria for evaluating LLINs as a tool for malaria prevention. Describing first the different dimensions of the intervention and the sociocultural context separately, we will further show how the variability underlying both is affected and induced by inappropriate aspects of the intervention and the measurements of its impact. We consider the gap between "net use" and the numerical representations of such local net use justifies further exploration of potential strategies to improve LLIN use in subgroups where persisting malaria transmission clusters.

Enteric Fever in Cambodia: Community Perceptions and Practices Concerning Disease Transmission and Treatment.

Enteric fever is a systemic bacterial infection in humans that is endemic in Cambodia and for which antibiotic resistance is increasingly reported. To guide public health programs, this qualitative study sought to explore community perceptions on transmission and treatment. Participant observation was carried out in hospital settings, pharmacies, and at a community level in Phnom Penh. In-depth interviews 39 and one focus group discussion were carried out with blood culture-confirmed enteric fever patients and purposively selected key informants. Informants were theoretically sampled based on initial themes identified using abductive analysis. Nvivo 11 was used for thematic coding. An urgent need to address health literacy concerning the transmission of enteric fever was identified, as lay informants did not link the disease and its symptoms to bacterial contamination of foods and drinks but rather to foods considered "bad" following humoral illness interpretations. As a result, lay informants considered recurrence of enteric fever preventable with appropriate dietary restrictions and Khmer traditional medicines. This study also reveals pluralistic health-care-seeking behavior. For initial and mild symptoms, patients preferred home treatment or traditional healing practices; limited household finances delayed treatment seeking. When symptoms persisted, patients first visited drug outlets or private practitioners, where they received a mix of nonessential medicines and one or more antibiotics often without prescription or confirmation of diagnosis. Inappropriate use of antibiotics was common and was related to diagnostic uncertainty and limited finances, factors which should be addressed during future efforts to improve the uptake of appropriate diagnostics and treatment of enteric fever.

Efficacy of topical mosquito repellent (picaridin) plus long-lasting insecticidal nets versus long-lasting insecticidal nets alone for control of malaria: a cluster randomised controlled trial.

BACKGROUND: Although effective topical repellents provide personal protection against malaria, whether mass use of topical repellents in addition to long-lasting insecticidal nets can contribute to a further decline of malaria is not known, particularly in areas where outdoor transmission occurs. We aimed to assess the epidemiological efficacy of a highly effective topical repellent in addition to long-lasting insecticidal nets in reducing malaria prevalence in this setting. METHODS: A cluster randomised controlled trial was done in the 117 most endemic villages in Ratanakiri province, Cambodia, to assess the efficacy of topical repellents in addition to long-lasting insecticidal nets in controlling malaria in a low-endemic setting. We did a pre-trial assessment of village accessibility and excluded four villages because of their inaccessibility during the rainy season. Another 25 villages were grouped because of their proximity to each other, resulting in 98 study clusters (comprising either a single village or multiple neighbouring villages). Clusters were randomly assigned (1:1) to either a control (long-lasting insecticidal nets) or intervention (long-lasting insecticidal nets plus topical repellent) study group after a restricted randomisation. All clusters received one long-lasting insecticidal net per individual, whereas those in the intervention group also received safe and effective topical repellents (picaridin KBR3023, SC Johnson, Racine, WI, USA), along with instruction and promotion of its daily use. Cross-sectional surveys of 65 randomly selected individuals per cluster were done at the beginning and end of the malaria transmission season in 2012 and 2013. The primary outcome was Plasmodium species-specific prevalence in participants obtained by real-time PCR, assessed in the intention-to-treat population. Complete safety analysis data will be published seperately; any ad-hoc adverse events are reported here. This trial is registered with ClinicalTrials.gov, number NCT01663831. FINDINGS: Of the 98 clusters that villages were split into, 49 were assigned to the control group and 49 were assigned to the intervention group. Despite having a successful distribution system, the daily use of repellents was suboptimum. No post-intervention differences in PCR plasmodium prevalence were observed between study groups in 2012 (4·91% in the control group vs 4·86% in the intervention group; adjusted odds ratio [aOR] 1·01 [95% CI 0·60-1·70]; p=0·975) or in 2013 (2·96% in the control group vs 3·85% in the intervention group; aOR 1·31 [0·81-2·11]; p=0·266). Similar results were obtained according to Plasmodium species (1·33% of participants in the intervention group vs 1·10% in the intervention group were infected with Plasmodium falciparum; aOR 0·83 [0·44-1·56]; p=0·561; and 1·85% in the control group vs 2·67% in the intervention group were infected with Plasmodium vivax; aOR 1·51 [0·88-2·57]; p=0·133). 41 adverse event notifications from nine villages were received, of which 33 were classified as adverse reactions (11 of these 33 were cases of repellent abuse through oral ingestion, either accidental or not). All participants with adverse reactions fully recovered and 17 were advised to permanently stop using the repellent. INTERPRETATION: Mass distribution of highly effective topical repellents in resource-sufficient conditions did not contribute to a further decline in malaria endemicity in a pre-elimination setting in the Greater Mekong subregion. Daily compliance and appropriate use of the repellents remains the main obstacle. FUNDING: Bill & Melinda Gates Foundation.

Characterizing Types of Human Mobility to Inform Differential and Targeted Malaria Elimination Strategies in Northeast Cambodia.

Human population movements currently challenge malaria elimination in low transmission foci in the Greater Mekong Subregion. Using a mixed-methods design, combining ethnography (n = 410 interviews), malariometric data (n = 4996) and population surveys (n = 824 indigenous populations; n = 704 Khmer migrants) malaria vulnerability among different types of mobile populations was researched in the remote province of Ratanakiri, Cambodia. Different structural types of human mobility were identified, showing differential risk and vulnerability. Among local indigenous populations, access to malaria testing and treatment through the VMW-system and LLIN coverage was high but control strategies failed to account for forest farmers' prolonged stays at forest farms/fields (61% during rainy season), increasing their exposure (p = 0.002). The Khmer migrants, with low acquired immunity, active on plantations and mines, represented a fundamentally different group not reached by LLIN-distribution campaigns since they were largely unregistered (79%) and unaware of the local VMW-system (95%) due to poor social integration. Khmer migrants therefore require control strategies including active detection, registration and immediate access to malaria prevention and control tools from which they are currently excluded. In conclusion, different types of mobility require different malaria elimination strategies. Targeting mobility without an in-depth understanding of malaria risk in each group challenges further progress towards elimination.

Assuring access to topical mosquito repellents within an intensive distribution scheme: a case study in a remote province of Cambodia.

BACKGROUND: The public health value of a vector control tool depends on its epidemiological efficacy, but also on its ease of implementation. This study describes an intensive distribution scheme of a topical repellent implemented in 2012 and 2013 for the purpose of a cluster-randomized trial using the existing public health system. The trial aimed to assess the effectiveness of repellents in addition to long-lasting insecticidal nets (LLIN) and occurred in a province of Cambodia. Determinants for accessibility and consumption of this tool were explored. METHODS: 135 individuals were appointed to be repellent distributors in 57 villages. A 2-weekly bottle exchange programme was organized. Distributors recorded information regarding the amount of bottles exchanged, repellent leftover, and reasons for not complying in household data sheets. Distributor-household contact rates and average 2-weekly consumption of repellent were calculated. Household and distributors characteristics were obtained using questionnaires, surveying 50 households per cluster and all distributors. Regression models were used to explore associations between contact and consumption rates and determinants such as socio-economic status. Operational costs for repellent and net distribution were obtained from the MalaResT project and the provincial health department. RESULTS: A fourfold increase in distributor-household contact rates was observed in 2013 compared to 2012 (median2012 = 20 %, median2013 = 88.9 %). Consumption rate tripled over the 2-year study period (median2012 = 20 %, median2013 = 57.89 %). Contact rates were found to associate with district, commune and knowing the distributor, while consumption was associated with district and household head occupation. The annual operational cost per capita for repellent distribution was 31 times more expensive than LLIN distribution (USD 4.33 versus USD 0.14). DISCUSSION: After the existing public health system was reinforced with programmatic and logistic support, an intense 2-weekly distribution scheme of a vector control tool over a 2-year period was operated successfully in the field. Lack of associations with socio-economic status suggested that the free distribution strategy resulted in equitable access to repellents. The operational costs for the repellent distribution and exchange programme were much higher than LLIN distribution. Such effort could only be justified in the context of malaria elimination where these interventions are expected to be limited in time.

Factors influencing the use of topical repellents: implications for the effectiveness of malaria elimination strategies.

In Cambodia, despite an impressive decline in prevalence over the last 10 years, malaria is still a public health problem in some parts of the country. This is partly due to vectors that bite early and outdoors reducing the effectiveness of measures such as Long-Lasting Insecticidal Nets. Repellents have been suggested as an additional control measure in such settings. As part of a cluster-randomized trial on the effectiveness of topical repellents in controlling malaria infections at community level, a mixed-methods study assessed user rates and determinants of use. Repellents were made widely available and Picaridin repellent reduced 97% of mosquito bites. However, despite high acceptability, daily use was observed to be low (8%) and did not correspond to the reported use in surveys (around 70%). The levels of use aimed for by the trial were never reached as the population used it variably across place (forest, farms and villages) and time (seasons), or in alternative applications (spraying on insects, on bed nets, etc.). These findings show the key role of human behavior in the effectiveness of malaria preventive measures, questioning whether malaria in low endemic settings can be reduced substantially by introducing measures without researching and optimizing community involvement strategies.

High Mobility and Low Use of Malaria Preventive Measures Among the Jarai Male Youth Along the Cambodia-Vietnam Border.

Malaria control along the Vietnam-Cambodia border presents a challenge for both countries' malaria elimination targets as the region is forested, inhabited by ethnic minority populations, and potentially characterized by early and outdoor malaria transmission. A mixed methods study assessed the vulnerability to malaria among the Jarai population living on both sides of the border in the provinces of Ratanakiri (Cambodia) and Gia Lai (Vietnam). A qualitative study generated preliminary hypotheses that were quantified in two surveys, one targeting youth (N = 498) and the other household leaders (N = 449). Jarai male youth, especially in Cambodia, had lower uptake of preventive measures (57.4%) and more often stayed overnight in the deep forest (35.8%) compared with the female youth and the adult population. Among male youth, a high-risk subgroup was identified that regularly slept at friends' homes or outdoors, who had fewer bed nets (32.5%) that were torn more often (77.8%). The vulnerability of Jarai youth to malaria could be attributed to the transitional character of youth itself, implying less fixed sleeping arrangements in nonpermanent spaces or non-bed sites. Additional tools such as long-lasting hammock nets could be suitable as they are in line with current practices.

Re-imagining malaria: heterogeneity of human and mosquito behaviour in relation to residual malaria transmission in Cambodia.

BACKGROUND: In certain regions in Southeast Asia, where malaria is reduced to forested regions populated by ethnic minorities dependent on slash-and-burn agriculture, malaria vector populations have developed a propensity to feed early and outdoors, limiting the effectiveness of long-lasting insecticide-treated nets (LLIN) and indoor residual spraying (IRS). The interplay between heterogeneous human, as well as mosquito behaviour, radically challenges malaria control in such residual transmission contexts. This study examines human behavioural patterns in relation to the vector behaviour. METHODS: The anthropological research used a sequential mixed-methods study design in which quantitative survey research methods were used to complement findings from qualitative ethnographic research. The qualitative research existed of in-depth interviews and participant observation. For the entomological research, indoor and outdoor human landing collections were performed. All research was conducted in selected villages in Ratanakiri province, Cambodia. RESULTS: Variability in human behaviour resulted in variable exposure to outdoor and early biting vectors: (i) indigenous people were found to commute between farms in the forest, where malaria exposure is higher, and village homes; (ii) the indoor/outdoor biting distinction was less clear in forest housing often completely or partly open to the outside; (iii) reported sleeping times varied according to the context of economic activities, impacting on the proportion of infections that could be accounted for by early or nighttime biting; (iv) protection by LLINs may not be as high as self-reported survey data indicate, as observations showed around 40% (non-treated) market net use while (v) unprotected evening resting and deep forest activities impacted further on the suboptimal use of LLINs. CONCLUSIONS: The heterogeneity of human behaviour and the variation of vector densities and biting behaviours may lead to a considerable proportion of exposure occurring during times that people are assumed to be protected by the distributed LLINs. Additional efforts in improving LLIN use during times when people are resting in the evening and during the night might still have an impact on further reducing malaria transmission in Cambodia.

Spatial clustering and risk factors of malaria infections in Ratanakiri Province, Cambodia.

BACKGROUND: Malaria incidence worldwide has steadily declined over the past decades. Consequently, increasingly more countries will proceed from control to elimination. The malaria distribution in low incidence settings appears patchy, and local transmission hotspots are a continuous source of infection. In this study, species-specific clusters and associated risk factors were identified based on malaria prevalence data collected in the north-east of Cambodia. In addition, Plasmodium falciparum genetic diversity, population structure and gene flows were studied. METHOD: In 2012, blood samples from 5793 randomly selected individuals living in 117 villages were collected from Ratanakiri province, Cambodia. Malariometric data of each participant were simultaneously accumulated using a standard questionnaire. A two-step PCR allowed for species-specific detection of malaria parasites, and SNP-genotyping of P. falciparum was performed. SaTScan was used to determine species-specific areas of elevated risk to infection, and univariate and multivariate risk analyses were carried out. RESULT: PCR diagnosis found 368 positive individuals (6.4%) for malaria parasites, of which 22% contained mixed species infections. The occurrence of these co-infections was more frequent than expected. Specific areas with elevated risk of infection were detected for all Plasmodium species. The clusters for Falciparum, Vivax and Ovale malaria appeared in the north of the province along the main river, while the cluster for Malariae malaria was situated elsewhere. The relative risk to be a malaria parasite carrier within clusters along the river was twice that outside the area. The main risk factor associated with three out of four malaria species was overnight stay in the plot hut, a human behaviour associated with indigenous farming. Haplotypes did not show clear geographical population structure, but pairwise Fst value comparison indicated higher parasite flow along the river. DISCUSSION: Spatial aggregation of malaria parasite carriers, and the identification of malaria species-specific risk factors provide key insights in malaria epidemiology in low transmission settings, which can guide targeted supplementary interventions. Consequently, future malaria programmes in the province should implement additional specific policies targeting households staying overnight at their farms outside the village, in addition to migrants and forest workers.

An innovative tool for moving malaria PCR detection of parasite reservoir into the field.

BACKGROUND: To achieve the goal of malaria elimination in low transmission areas such as in Cambodia, new, inexpensive, high-throughput diagnostic tools for identifying very low parasite densities in asymptomatic carriers are required. This will enable a switch from passive to active malaria case detection in the field. METHODS: DNA extraction and real-time PCR assays were implemented in an "in-house" designed mobile laboratory allowing implementation of a robust, sensitive and rapid malaria diagnostic strategy in the field. This tool was employed in a survey organized in the context of the MalaResT project (NCT01663831). RESULTS: The real-time PCR screening and species identification assays were performed in the mobile laboratory between October and November 2012, in Rattanakiri Province, to screen approximately 5,000 individuals in less than four weeks and treat parasite carriers within 24-48 hours after sample collection. An average of 240 clinical samples (and 40 quality control samples) was tested every day, six/seven days per week. Some 97.7% of the results were available <24 hours after the collection. A total of 4.9% were positive for malaria. Plasmodium vivax was present in 61.1% of the positive samples, Plasmodium falciparum in 45.9%, Plasmodium malariae in 7.0% and Plasmodium ovale in 2.0%. CONCLUSIONS: The operational success of this diagnostic set-up proved that molecular testing and subsequent treatment is logistically achievable in field settings. This will allow the detection of clusters of asymptomatic carriers and to provide useful epidemiological information. Fast results will be of great help for staff in the field to track and treat asymptomatic parasitaemic cases. The concept of the mobile laboratory could be extended to other countries for the molecular detection of malaria or other pathogens, or to culture vivax parasites, which does not support long-time delay between sample collection and culture.

Injections, cocktails and diviners: therapeutic flexibility in the context of malaria elimination and drug resistance in Northeast Cambodia.

BACKGROUND: Adherence to effective malaria medication is extremely important in the context of Cambodia's elimination targets and drug resistance containment. Although the public sector health facilities are accessible to the local ethnic minorities of Ratanakiri province (Northeast Cambodia), their illness itineraries often lead them to private pharmacies selling "cocktails" and artemether injections, or to local diviners prescribing animal sacrifices to appease the spirits. METHODS: The research design consisted of a mixed methods study, combining qualitative (in-depth interviews and participant observation) and quantitative methods (household and cross-sectional survey). RESULTS: Three broad options for malaria treatment were identified: i) the public sector; ii) the private sector; iii) traditional treatment based on divination and ceremonial sacrifice. Treatment choice was influenced by the availability of treatment and provider, perceived side effects and efficacy of treatments, perceived etiology of symptoms, and patient-health provider encounters. Moreover, treatment paths proved to be highly flexible, changing mostly in relation to the perceived efficacy of a chosen treatment. CONCLUSIONS: Despite good availability of anti-malarial treatment in the public health sector, attendance remained low due to both structural and human behavioral factors. The common use and under-dosage of anti-malaria monotherapy in the private sector (single-dose injections, single-day drug cocktails) represents a threat not only for individual case management, but also for the regional plan of drug resistance containment and malaria elimination.

Using CF11 cellulose columns to inexpensively and effectively remove human DNA from Plasmodium falciparum-infected whole blood samples.

BACKGROUND: Genome and transcriptome studies of Plasmodium nucleic acids obtained from parasitized whole blood are greatly improved by depletion of human DNA or enrichment of parasite DNA prior to next-generation sequencing and microarray hybridization. The most effective method currently used is a two-step procedure to deplete leukocytes: centrifugation using density gradient media followed by filtration through expensive, commercially available columns. This method is not easily implemented in field studies that collect hundreds of samples and simultaneously process samples for multiple laboratory analyses. Inexpensive syringes, hand-packed with CF11 cellulose powder, were recently shown to improve ex vivo cultivation of Plasmodium vivax obtained from parasitized whole blood. This study was undertaken to determine whether CF11 columns could be adapted to isolate Plasmodium falciparum DNA from parasitized whole blood and achieve current quantity and purity requirements for Illumina sequencing. METHODS: The CF11 procedure was compared with the current two-step standard of leukocyte depletion using parasitized red blood cells cultured in vitro and parasitized blood obtained ex vivo from Cambodian patients with malaria. Procedural variations in centrifugation and column size were tested, along with a range of blood volumes and parasite densities. RESULTS: CF11 filtration reliably produces 500 nanograms of DNA with less than 50% human DNA contamination, which is comparable to that obtained by the two-step method and falls within the current quality control requirements for Illumina sequencing. In addition, a centrifuge-free version of the CF11 filtration method to isolate P. falciparum DNA at remote and minimally equipped field sites in malaria-endemic areas was validated. CONCLUSIONS: CF11 filtration is a cost-effective, scalable, one-step approach to remove human DNA from P. falciparum-infected whole blood samples.