Mark Janko

Pfhrp2 -Deleted Plasmodium falciparum Parasites in the Democratic Republic of the Congo: A National Cross-sectional Survey

Background Rapid diagnostic tests (RDTs) account for more than two-thirds of malaria diagnoses in Africa. Deletions of the Plasmodium falciparum hrp2 (pfhrp2) gene cause false-negative RDT results and have never been investigated on a national level. Spread of pfhrp2-deleted P. falciparum mutants, resistant to detection by HRP2-based RDTs, would represent a serious threat to malaria elimination efforts. Methods Using a nationally representative cross-sectional study of 7,137 children under five years of age from the Democratic Republic of Congo (DRC), we tested 783 subjects with RDT-/PCR+ results using PCR assays to detect and confirm deletions of the pfhrp2 gene. Spatial and population genetic analyses were employed to examine the distribution and evolution of these parasites. Results We identified 149 pfhrp2-deleted parasites, representing 6.4% of all P. falciparum infections country-wide (95% confidence interval 5.1-8.0%). Bayesian spatial analyses identified statistically significant clustering of pfhrp2 deletions near Kinshasa and Kivu. Population genetic analysis revealed significant genetic differentiation between wild-type and pfhrp2-deleted parasite populations (GST = .046, p ≤ .00001). Conclusions Pfhrp2-deleted P. falciparum is a common cause of RDT-/PCR+ malaria among asymptomatic children in the DRC and appears to be clustered within select communities. Surveillance for these deletions is needed, and alternatives to HRP2-specific RDTs may be necessary.

Genetic Evidence of Importation of Drug-Resistant Plasmodium falciparum to Guatemala from the Democratic Republic of the Congo

Molecular markers and population genetics were effective tracking tools.

Plasmodium falciparum sulfadoxine resistance is geographically and genetically clustered within the DR Congo

Understanding the spatial clustering of Plasmodium falciparum populations can assist efforts to contain drug-resistant parasites and maintain the efficacy of future drugs. We sequenced single nucleotide polymorphisms (SNPs) in the dihydropteroate synthase gene (dhps) associated with sulfadoxine resistance and 5 microsatellite loci flanking dhps in order to investigate the genetic backgrounds, genetic relatedness, and geographic clustering of falciparum parasites in the Democratic Republic of the Congo (DRC). Resistant haplotypes were clustered into subpopulations: one in the northeast DRC, and the other in the balance of the DRC. Network and clonal lineage analyses of the flanking microsatellites indicate that geographically-distinct mutant dhps haplotypes derive from separate lineages. The DRC is therefore a watershed for haplotypes associated with sulfadoxine resistance. Given the importance of central Africa as a corridor for the spread of antimalarial resistance, the identification of the mechanisms of this transit can inform future policies to contain drug-resistant parasite strains.

Malaria surveillance in the Democratic Republic of the Congo: comparison of microscopy, PCR, and rapid diagnostic test.

Malaria surveillance is critical for control efforts, but diagnostic methods frequently disagree. Here, we compare microscopy, PCR, and a rapid diagnostic test in 7137 samples from children in the Democratic Republic of the Congo using latent class analysis. PCR had the highest sensitivity (94.6%) and microscopy had the lowest (76.7%).

A cross-sectional survey of Plasmodium falciparum pfcrt mutant haplotypes in the Democratic Republic of Congo.

In the Democratic Republic of the Congo (DRC), artesunate-amodiaquine is first-line therapy for falciparum malaria; little is known about the prevalence of molecular markers of parasite drug resistance. Across the DRC, we genotyped 166 parasites in Plasmodium falciparum chloroquine resistance transporter (pfcrt) using polymerase chain reaction (PCR) and sequencing. Of these parasites, 73 (44%) parasites were pure wild-type CVMNK, 55 (31%) parasites were chloroquine-resistant CVIET: , 35 (21.1%) parasites were mixed CVMNK and CVIET: , and 3 parasites were other genotypes. Ninety-two infections (55.4%) harbored the pfcrt K76T: substitution that is highly correlated with chloroquine failure. The amodiaquine-resistant S: VMNT: haplotype was absent. Geographically, pfcrt haplotypes were not clearly clustered. Chloroquine accounted for 19.4% of antimalarial use, and amodiaquine accounted for 15.3% of antimalarial use; there were no associations between drug use and mutant haplotype prevalence. In the DRC, our molecular survey indicates that resistance to chloroquine is substantial but that resistance to amodiaquine is absent. These contrasting findings highlight the need for molecular surveillance of drug resistance to inform malaria control policies.

Changing spatial patterns and increasing rurality of HIV prevalence in the Democratic Republic of the Congo between 2007 and 2013.

The Democratic Republic of the Congo (DRC) has one of the lowest HIV prevalence in sub-Saharan Africa, estimated at 1.1% [0.9-1.3] of adults aged 15-49 in 2013 (UNAIDS). Within the 2 million km(2) country, however, there exists spatial variation in HIV prevalence, with the highest HIV prevalence observed in the large cities of Kinshasa and Lubumbashi. Globally, HIV is an increasingly rural disease, diffusing outwards from urban centers of high HIV prevalence to places where HIV was previously absent or present at very low levels. Utilizing data collected during Demographic and Health Surveillance (DHS) in 2007 and 2013 in the DRC, we sought to update the map of HIV prevalence in the DRC as well as to explore whether HIV in the DRC is an increasingly rural disease or remains confined to urban areas. Bayesian kriging and regression indicate that HIV prevalence in rural areas of the DRC is higher in 2013 than in 2007 and that increased distance to an urban area is no longer protective against HIV as it was in 2007. These findings suggest that HIV education, testing and prevention efforts need to diffuse from urban to rural areas just as HIV is doing.

The links between agriculture, Anopheles mosquitoes, and malaria risk in children younger than 5 years in the Democratic Republic of the Congo: a population-based, cross-sectional, spatial study

Summary Background The relationship between agriculture, Anopheles mosquitoes, and malaria in Africa is not fully understood, but it is important for malaria control as countries consider expanding agricultural projects to address population growth and food demand. Therefore, we aimed to assess the effect of agriculture on Anopheles biting behaviour and malaria risk in children in rural areas of the Democratic Republic of the Congo (DR Congo). Methods We did a population-based, cross-sectional, spatial study of rural children (<5 years) in the DR Congo. We used information about the presence of malaria parasites in each child, as determined by PCR analysis of dried-blood spots from the 2013–14 DR Congo Demographic and Health Survey (DHS). We also used data from the DHS, a longitudinal entomological study, and available land cover and climate data to evaluate the relationships between agriculture, Anopheles biting behaviour, and malaria prevalence. Satellite imagery was used to measure the percentage of agricultural land cover around DHS villages and Anopheles sites. Anopheles biting behaviour was assessed by Human Landing Catch. We used probit regression to assess the relationship between agriculture and the probability of malaria infection, as well as the relationship between agriculture and the probability that a mosquito was caught biting indoors. Findings Between Aug 13, 2013, and Feb 13, 2014, a total of 9790 dried-blood spots were obtained from the DHS, of which 4612 participants were included in this study. Falciparum malaria infection prevalence in rural children was 38·7% (95% uncertainty interval [UI] 37·3–40·0). Increasing exposure to agriculture was associated with increasing malaria risk with a high posterior probability (estimate 0·07, 95% UI −0·04 to 0·17; posterior probability [estimate >0]=0·89), with the probability of malaria infection increased between 0·2% (95% UI −0·1 to 3·4) and 2·6% (–1·5 to 6·6) given a 15% increase in agricultural cover, depending on other risk factors. The models predicted that large increases in agricultural cover (from 0% to 75%) increase the probability of infection by as much as 13·1% (95% UI −7·3 to 28·9). Increased risk might be due to Anopheles gambiae sensu lato, whose probability of biting indoors increased between 11·3% (95% UI −15·3 to 25·6) and 19·7% (–12·1 to 35·9) with a 15% increase in agriculture. Interpretation Malaria control programmes must consider the possibility of increased risk due to expanding agriculture. Governments considering initiating large-scale agricultural projects should therefore also consider accompanying additional malaria control measures. Funding National Institutes of Health, National Science Foundation, Bill & Melinda Gates Foundation, Presidents Malaria Initiative, and Royster Society of Fellows at the University of North Carolina at Chapel Hill.

Drug-Resistance and Population Structure of Plasmodium falciparum Across the Democratic Republic of Congo Using High-Throughput Molecular Inversion Probes

Probing all known drug-resistance loci using molecular inversion probes across the Democratic Republic of the Congo reveals prevalence, frequency, and geographical distribution of drug-resistant parasites, as well as spread of sulfadoxine-resistance mutations despite minimal official sulfadoxine use.

Effect of individual and community-level bed net usage on malaria prevalence among under-fives in the Democratic Republic of Congo

BackgroundUnderstanding the contribution of community-level long-lasting, insecticidal net (LLIN) coverage to malaria control is critical to planning and assessing intervention campaigns. The Democratic Republic of Congo (DRC), which has one of the highest burdens of malaria cases and deaths and has dramatically scaled up LLIN ownership in recent years thus it is an ideal setting to evaluate the effect of individual versus community-level use to prevent malaria among children under the age of 5.ResultsData were derived from the 2013–2014 DRC Demographic and Health Survey. Community-level LLIN usage was significantly associated with protection against malaria, even when individual-level LLIN usage was included in the model. In stratified analysis, higher levels of community LLIN coverage enhanced the protective effect of individual LLIN usage, resulting in lower malaria prevalence among individuals who used a LLIN. A sub-analysis of individual LLIN usage by insecticide type revealed deltamethrin-treated nets were more protective than permethrin-treated nets, suggesting that mosquitoes in the DRC are more susceptible to deltamethrin.ConclusionsThis study examines the effects of individual and community-level LLIN usage in young children in an area of high ITN usage. Individual and community LLIN usage were significantly associated with protection against malaria in children under 5 in the DRC. Importantly, the protective effect of individual LLIN usage against malaria is enhanced when community LLIN coverage is higher, demonstrating the importance of increasing community-level LLIN usage. LLINs treated with deltamethrin were shown to be more protective against malaria than LLINs treated with permethrin. Demographic and Health Surveys are thus a novel and important means of surveillance for insecticide resistance.

Individual and household characteristics of persons with Plasmodium falciparum malaria in sites with varying endemicities in Kinshasa Province, Democratic Republic of the Congo

BackgroundThe Democratic Republic of the Congo (DRC) bears a large share of global malaria burden despite efforts to control and eliminate the disease. More detailed understanding of individual and household level characteristics associated with malaria are needed, as is an understanding of how these characteristics vary spatiotemporally and across different community-level malaria endemicities. An ongoing study in Kinshasa Province is designed to address gaps in prior malaria surveillance in the DRC by monitoring malaria across seasons, age groups and in high and low malaria sites. Across seven sites, 242 households and 1591 individuals are participating in the study. Results of the enrollment questionnaire, rapid diagnostic tests and PCR testing of dried blood spots are presented.ResultsOverall malaria prevalence in the study cohort is high, 27% by rapid diagnostic test and 31% by polymerase chain reaction, and malaria prevalence is highly varied across very small geographic distances. Malaria prevalence is highest in children aged 6–15. While the majority of households own bed nets, bed net usage is less than 50%.ConclusionsThe study cohort will provide an understanding of how malaria persists in populations that have varying environmental exposures, varying community-level malaria, and varying access to malaria control efforts.