Prevalence of malaria resistance-associated mutations in Plasmodium falciparum circulating in 2017-2018, Bo, Sierra Leone.

Introduction: In spite of promising medical, sociological, and engineering strategies and interventions to reduce the burden of disease, malaria remains a source of significant morbidity and mortality, especially among children in sub-Saharan Africa. In particular, progress in the development and administration of chemotherapeutic agents is threatened by evolved resistance to most of the antimalarials currently in use, including artemisinins. Methods: This study analyzed the prevalence of mutations associated with antimalarial resistance in Plasmodium falciparum from 95 clinical samples collected from individuals with clinically confirmed malaria at a hospital in Bo, Sierra Leone between May 2017 and December 2018. The combination of polymerase chain reaction amplification and subsequent high throughput DNA sequencing was used to determine the presence of resistance-associated mutations in five P. falciparum genes - pfcrt, pfmdr1, pfdhfr, pfdhps and pfkelch13. The geographic origin of parasites was assigned using mitochondrial sequences. Results: Relevant mutations were detected in the pfcrt (22%), pfmdr1 (>58%), pfdhfr (100%) and pfdhps (>80%) genes while no resistance-associated mutations were found in the pfkelch13 gene. The mitochondrial barcodes were consistent with a West African parasite origin with one exception indicating an isolate imported from East Africa. Discussion: Detection of the pfmdr1 NFSND haplotype in 50% of the samples indicated the increasing prevalence of strains with elevated tolerance to artemeter + lumefantrine (AL) threatening the combination currently used to treat uncomplicated malaria in Sierra Leone. The frequency of mutations linked to resistance to antifolates suggests widespread resistance to the drug combination used for intermittent preventive treatment during pregnancy.

Are malaria transmission-blocking vaccines acceptable to high burden communities? Results from a mixed methods study in Bo, Sierra Leone.

BACKGROUND: Malaria transmission-blocking vaccines (TBVs) could help break the cycle of malaria transmission by conferring community rather than individual protection. When introducing new intervention strategies, uptake is dependent on acceptability, not just efficacy. In this exploratory study on acceptability of TBVs in Sierra Leone, it was hypothesized that TBVs would be largely acceptable to adults and health workers in areas with relatively few ongoing malaria interventions, and that (i) knowledge of malaria and vaccines, (ii) health behaviours associated with malaria and vaccines, and (iii) attitudes towards different vaccines types could lead to greater TBV acceptability. METHODS: This study used a mixed methods approach in Bo, Sierra Leone, to understand community knowledge, attitudes, and practices related to malaria and vaccination in general. This included: (i) a population-based cross-sectional survey (n=615 adults), (ii) 6 focus group discussions with parents, and (iii) 20 key informant interviews. The concept of a TBV was explained to participants before they were asked about their willingness to accept this vaccine modality as part of an integrated malaria elimination programme. RESULTS: This study found that most adults would be willing to receive a TBV vaccine. Respondents noted mostly positive past experiences with adult and childhood vaccinations for other infectious diseases and high levels of engagement in other malaria prevention behaviors such as bed nets. Perceived barriers to TBV acceptance were largely focused on general community-level distribution of a vaccine, including personal fears of vaccination and possible costs. After an explanation of the TBV mechanism, nearly all focus group and interview participants believed that community members would accept the vaccine as part of an integrated malaria control approach. Both parents and health workers offered insight on how to successfully roll-out a future TBV vaccination programme. CONCLUSIONS: The willingness of community members in Bo, Sierra Leone to accept a TBV as part of an integrated anti-malarial strategy suggests that the atypical mechanism of TBV action might not be an obstacle to future clinical trials. This study's findings suggests that perceived general barriers to vaccination implementation, such as perceived personal fears and vaccine cost, must be addressed in future clinical and implementation research studies.

Comparison of capillary and venous blood for malaria detection using two PCR-based assays in febrile patients in Sierra Leone.

BACKGROUND: Rapid and sensitive diagnostics are critical tools for clinical case management and public health control efforts. Both capillary and venous blood are currently used for malaria detection and while diagnostic technologies may not be equally sensitive with both materials, the published data on this subject are scarce and not conclusive. METHODS: Paired clinical samples of venous and capillary blood from 141 febrile individuals in Bo, Sierra Leone, were obtained between January and May 2019 and tested for the presence of Plasmodium parasites using two multiplexed PCR assays: the FilmArray-based Global Fever Panel (GFP) and the TaqMan-based Malaria Multiplex Sample Ready (MMSR) assay. RESULTS: No significant differences in Plasmodium parasite detection between capillary and venous blood for both assays were observed. The GFP assay was more sensitive than MMSR for all markers that could be compared (Plasmodium spp. and Plasmodium falciparum) in both venous and capillary blood. CONCLUSIONS: No difference was found in malaria detection between venous and capillary blood using two different PCR-based detection assays. This data gives support for use of capillary blood, a material which can be obtained easier by less invasive methods, for PCR-based malaria diagnostics, independent of the platform.

Use of real-time multiplex PCR, malaria rapid diagnostic test and microscopy to investigate the prevalence of Plasmodium species among febrile hospital patients in Sierra Leone.

BACKGROUND: Malaria continues to affect over 200 million individuals every year, especially children in Africa. Rapid and sensitive detection and identification of Plasmodium parasites is crucial for treating patients and monitoring of control efforts. Compared to traditional diagnostic methods such as microscopy and rapid diagnostic tests (RDTs), DNA based methods, such as polymerase chain reaction (PCR) offer significantly higher sensitivity, definitive discrimination of Plasmodium species, and detection of mixed infections. While PCR is not currently optimized for routine diagnostics, its role in epidemiological studies is increasing as the world moves closer toward regional and eventually global malaria elimination. This study demonstrates the field use of a novel, ambient temperature-stabilized, multiplexed PCR assay in a small hospital setting in Sierra Leone. METHODS: Blood samples from 534 febrile individuals reporting to a hospital in Bo, Sierra Leone, were tested using three methods: a commercial RDT, microscopy, and a Multiplex Malaria Sample Ready (MMSR) PCR designed to detect a universal malaria marker and species-specific markers for Plasmodium falciparum and Plasmodium vivax. A separate PCR assay was used to identify species of Plasmodium in samples in which MMSR detected malaria, but was unable to identify the species. RESULTS: MMSR detected the presence of any malaria marker in 50.2% of all tested samples with P. falciparum identified in 48.7% of the samples. Plasmodium vivax was not detected. Testing of MMSR P. falciparum-negative/universal malaria-positive specimens with a panel of species-specific PCRs revealed the presence of Plasmodium malariae (n = 2) and Plasmodium ovale (n = 2). The commercial RDT detected P. falciparum in 24.6% of all samples while microscopy was able to detect malaria in 12.8% of tested specimens. CONCLUSIONS: Wider application of PCR for detection of malaria parasites may help to fill gaps existing as a result of use of microscopy and RDTs. Due to its high sensitivity and specificity, species coverage, room temperature stability and relative low complexity, the MMSR assay may be useful for detection of malaria and epidemiological studies especially in low-resource settings.

Estimating the size of urban populations using Landsat images: a case study of Bo, Sierra Leone, West Africa.

BACKGROUND: This is the third paper in a 3-paper series evaluating alternative models for rapidly estimating neighborhood populations using limited survey data, augmented with aerial imagery. METHODS: Bayesian methods were used to sample the large solution space of candidate regression models for estimating population density. RESULTS: We accurately estimated the population densities and counts of 20 neighborhoods in the city of Bo, Sierra Leone, using statistical measures derived from Landsat multi-band satellite imagery. The best regression model proposed estimated the latter with an absolute median proportional error of 8.0%, while the total population of the 20 neighborhoods was estimated with an error of less than 1.0%. We also compare our results with those obtained using an empirical Bayes approach. CONCLUSIONS: Our approach provides a rapid and effective method for constructing predictive models for population densities and counts utilizing remote sensing imagery. Our results, including cross-validation analysis, suggest that masking non-urban areas in the Landsat section images prior to computing the candidate covariate regressors should further improve model generality.

Seroprevalence of hepatitis B surface antigen (HBsAg) in Bo, Sierra Leone, 2012-2013.

OBJECTIVE: The aim of this study was to determine the prevalence of hepatitis B surface antigen (HBsAg) among febrile individuals tested at Mercy Hospital Research Laboratory (MHRL) in Bo, Sierra Leone. RESULTS: A total of 860 febrile individuals ages 5 years and older were tested by MHRL between July 2012 and June 2013 with a Standard Diagnostics Bioline HBsAg rapid diagnostic test. The overall HBsAg prevalence rate was 13.7%, including a rate of 15.5% among males and 12.6% among females. The HBsAg rate did not differ by child or adult age group (p > 0.5). The prevalence rate in Bo was similar to the 11-15% HBsAg prevalence rates reported in the past decade from other studies across West Africa. Scaling up the infant hepatitis B vaccination program in Sierra Leone will be important for reducing the future burden of disease and premature death attributable to chronic viral hepatitis B disease.

Prevalence of markers of HIV infection among febrile adults and children in Bo, Sierra Leone, 2012-2013.

OBJECTIVE: The goal of this study was to examine the prevalence of HIV among febrile patients seeking care in Mercy Hospital, Bo, Sierra Leone, in 2012-2013. RESULTS: A total of 1207 febrile persons were tested for HIV with Determine™ and SD Bioline rapid diagnostic tests kits that detect the presence of HIV antibodies and HIV p24 antigens. The overall prevalence of HIV among the tested patients was 8.9%, which is considerably higher than the < 2% prevalence of HIV reported previously in the general population. While these results are not sufficient to prove a causal relationship, the obtained data imply that HIV positive individuals may be more likely to suffer from febrile infectious diseases than individuals without HIV infection. Increasing the availability and use of HIV testing services will allow antiretroviral therapy to be accessed in a timely manner and improve health status among people living with HIV.

Surveillance of Vector-Borne Infections (Chikungunya, Dengue, and Malaria) in Bo, Sierra Leone, 2012-2013.

Malaria remains a significant cause of morbidity and mortality in West Africa, but the contribution of other vector-borne infections (VBIs) to the burden of disease has been understudied. We used rapid diagnostic tests (RDTs) for three VBIs to test blood samples from 1,795 febrile residents of Bo City, Sierra Leone, over a 1-year period in 2012-2013. In total, 24% of the tests were positive for malaria, fewer than 5% were positive for markers of dengue virus infection, and 39% were positive for IgM directed against chikungunya virus (CHIKV) or a related alphavirus. In total, more than half (55%) of these febrile individuals tested positive for at least one of the three VBIs, which highlights the very high burden of vector-borne diseases in this population. The prevalence of positives on the Chikungunya IgM and dengue tests did not vary significantly with age (P > 0.36), but higher rates of malaria were observed in children < 15 years of age (P < 0.001). Positive results on the Chikungunya IgM RDTs were moderately correlated with rainfall (r2 = 0.599). Based on the high prevalence of positive results on the Chikungunya IgM RDTs from individuals Bo and its environs, there is a need to examine whether an ecological shift toward a greater burden from CHIKV or related alphaviruses is occurring in other parts of Sierra Leone or the West African region.

Stratified Sampling of Neighborhood Sections for Population Estimation: A Case Study of Bo City, Sierra Leone.

There is a need for better estimators of population size in places that have undergone rapid growth and where collection of census data is difficult. We explored simulated estimates of urban population based on survey data from Bo, Sierra Leone, using two approaches: (1) stratified sampling from across 20 neighborhoods and (2) stratified single-stage cluster sampling of only four randomly-sampled neighborhoods. The stratification variables evaluated were (a) occupants per individual residence, (b) occupants per neighborhood, and (c) residential structures per neighborhood. For method (1), stratification variable (a) yielded the most accurate re-estimate of the current total population. Stratification variable (c), which can be estimated from aerial photography and zoning type verification, and variable (b), which could be ascertained by surveying a limited number of households, increased the accuracy of method (2). Small household-level surveys with appropriate sampling methods can yield reasonably accurate estimations of urban populations.

Use of the FilmArray System for Detection of Zaire ebolavirus in a Small Hospital in Bo, Sierra Leone.

Laboratories associated with small hospitals often have limited expertise, personnel, and equipment to rapidly identify rare and emerging infectious diseases. We describe the successful use of the FilmArray system for rapid detection of Ebola virus directly from clinical samples in 6 out of 83 tested subjects in a small health care center in Sierra Leone.

Attitudes toward home-based malaria testing in rural and urban Sierra Leone.

BACKGROUND: The purpose of this study was to examine malaria testing practices and preferences in Bo, Sierra Leone, and to ascertain interest in and willingness to take a home-based rapid diagnostic test administered by a community health volunteer (CHV) or a trained family member rather than travelling to a clinical facility for laboratory-based testing. METHODS: A population-based, cross-sectional survey of 667 randomly-sampled rural households and 157 urban households was conducted in December 2013 and January 2014. RESULTS: Among rural residents, 69% preferred a self/family- or CHV-conducted home-based malaria test and 20% preferred a laboratory-based test (with others indicating no preference). Among urban residents, these numbers were 38% and 44%, respectively. If offered a home-based test, 28% of rural residents would prefer a self/family-conducted test and 68% would prefer a CHV-assisted test. For urban residents, these numbers were 21% and 77%. In total, 36% of rural and 63% of urban residents reported usually taking a diagnostic test to confirm suspected malaria. The most common reasons for not seeking malaria testing were the cost of testing, waiting to see if the fever resolved on its own, and not wanting to travel to a clinical facility for a test. In total, 32% of rural and 27% of urban participants were very confident they could perform a malaria test on themselves or a family member without assistance, 50% of rural and 62% of urban participants were very confident they could perform a test after training, and 56% of rural and 33% of urban participants said they would pay more for a home-based test than a laboratory-based test. CONCLUSION: Expanding community case management of malaria to include home testing by CHVs and family members may increase the proportion of individuals with febrile illnesses who confirm a positive diagnosis prior to initiating treatment.

Herbs and herbal combinations used to treat suspected malaria in Bo, Sierra Leone.

ETHNOPHARMACOLOGICAL RELEVANCE: Most adults in West Africa treat acute febrile illnesses with local herbs, but the patterns of herbs used for malaria have not been recently described in Sierra Leone. MATERIALS AND METHODS: We used a population-based cross-sectional approach to interview 810 randomly-sampled rural and urban adult residents of Bo, Sierra Leone, in December 2013 and January 2014 about their use of herbal remedies when they suspect they have malaria. RESULTS: In total, 55% of the participants reported taking one or more of seven herbs to treat symptoms of malaria. Among herb users, the most commonly used anti-malarial herbs were Moringa oleifera (moringa, 52%) and Sarcocephalus latifolius (yumbuyambay, 50%). The other herbs used included Senna siamea (shekutoure, 18%), Cassia sieberiana (gbangba, 18%), Uvaria afzelii (gone-botai, 14%), Morinda chrysorhiza (njasui, 14%), and Craterispermum laurinum (nyelleh, 7%). Combination herbal therapy was common, with 37% of herb users taking two or more herbs together when ill with suspected malaria. CONCLUSIONS: Indigenous medical knowledge about herbal remedies and combinations of local herbs remains an integral part of malaria case management in Sierra Leone.

Methods for determining the uncertainty of population estimates derived from satellite imagery and limited survey data: a case study of Bo city, Sierra Leone.

This study demonstrates the use of bootstrap methods to estimate the total population of urban and periurban areas using satellite imagery and limited survey data. We conducted complete household surveys in 20 neighborhoods in the city of Bo, Sierra Leone, which collectively were home to 25,954 persons living in 1,979 residential structures. For five of those twenty sections, we quantized the rooftop areas of structures extracted from satellite images. We used bootstrap statistical methods to estimate the total population of the pooled sections, including the associated uncertainty intervals, as a function of sample size. Evaluations based either on rooftop area per person or on the mean number of occupants per residence both converged on the true population size. We demonstrate with this simulation that demographic surveys of a relatively small proportion of residences can provide a foundation for accurately estimating the total population in conjunction with aerial photographs.

Reemergence of chikungunya virus in Bo, Sierra Leone.

We diagnosed 400 possible IgM-positive cases of chikungunya virus in Bo, Sierra Leone, during July 2012-January 2013 by using lateral flow immunoassays. Cases detected likely represent only a small fraction of total cases. Further laboratory testing is required to confirm this outbreak and characterize the virus.