The independent effect of living in malaria hotspots on future malaria infection: an observational study from Misungwi, Tanzania.

BACKGROUND: As malaria transmission declines, continued improvements of prevention and control interventions will increasingly rely on accurate knowledge of risk factors and an ability to define high-risk areas and populations at risk for focal targeting of interventions. This paper explores the independent association between living in a hotspot and prospective risk of malaria infection. METHODS: Malaria infection status defined by nPCR and AMA-1 status in year 1 were used to define geographic hotspots using two geospatial statistical methods (SaTScan and Kernel density smoothing). Other malaria risk factors for malaria infection were explored by fitting a multivariable model. RESULTS: This study demonstrated that residing in infection hotspot of malaria transmission is an independent predictor of malaria infection in the future. CONCLUSION: It is likely that targeting such hotspots with better coverage and improved malaria control strategies will result in more cost-efficient uses of resources to move towards malaria elimination

Population pharmacokinetics of primaquine and its metabolites in African males

Background: Primaquine (PQ) is the prototype 8-aminoquinoline drug, a class which targets gametocytes and hypnozoites. The World Health Organization (WHO) recommends adding a single low dose of primaquine to the standard artemisinin-based combination therapy (ACT) in order to block malaria transmission in regions with low malaria transmission. However, the haemolytic toxicity is a major adverse outcome of primaquine in glucose-6-phosphate dehydrogenase (G6PD)-deficient subjects. This study aimed to characterize the pharmacokinetic properties of primaquine and its major metabolites in G6PD-deficient subjects. Methods: A single low-dose of primaquine (0.4–0.5 mg/kg) was administered in twenty-eight African males. Venous and capillary plasma were sampled up to 24 h after the drug administration. Haemoglobin levels were observed up to 28 days after drug administration. Only PQ, carboxy-primaquine (CPQ), and primaquine carbamoyl-glucuronide (PQCG) were present in plasma samples and measured using liquid chromatography mass spectrometry. Drug and metabolites’ pharmacokinetic properties were investigated using nonlinear mixed-effects modelling. Results: Population pharmacokinetic properties of PQ, CPQ, and PQCG can be described by one-compartment disposition kinetics with a transit-absorption model. Body weight was implemented as an allometric function on the clearance and volume parameters for all compounds. None of the covariates significantly affected the pharmacokinetic parameters. No significant correlations were detected between the exposures of the measured compounds and the change in haemoglobin or methaemoglobin levels. There was no significant haemoglobin drop in the G6PD-deficient patients after administration of a single low dose of PQ. Conclusions: A single low-dose of PQ was haematologically safe in this population of G6PD-normal and G6PD-deficient African males without malaria. Trial registration NCT0253576

Primaquine to reduce transmission of Plasmodium falciparum malaria in Mali : a single-blind, dose-ranging, adaptive randomised phase 2 trial

Background Single low doses of primaquine, when added to artemisinin-based combination therapy, might prevent transmission of Plasmodium falciparum malaria to mosquitoes. We aimed to establish the activity and safety of four low doses of primaquine combined with dihydroartemisinin-piperaquine in male patients in Mali. Methods In this phase 2, single-blind, dose-ranging, adaptive randomised trial, we enrolled boys and men with uncomplicated P falciparum malaria at the Malaria Research and Training Centre (MRTC) field site in Ouelessebougou, Mali. All participants were confirmed positive carriers of gametocytes through microscopy and had normal function of glucose-6-phosphate dehydrogenase (G6PD) on colorimetric quantification In the first phase, participants were randomly assigned (1:1:1) to one of three primaquine doses: 0 mg/kg (control), 0.125 mg/kg, and 0.5 mg/kg. Randomisation was done with a computer-generated randomisation list (in block sizes of six) and concealed with sealed, opaque envelopes. In the second phase, different participants were sequentially assigned (1:1) to 0.25 mg/kg primaquine or 0.0625 mg/kg primaquine. Primaquine tablets were dissolved into a solution and administered orally in a single dose. Participants were also given a 3 day course of dihydroartemisinin-piperaquine, administered by weight (320 mg dihydroartemisinin and 40 mg piperaquine per tablet). Outcome assessors were masked to treatment allocation, but participants were permitted to find out group assignment. Infectivity was assessed through membrane feeding assays, which were optimised through the beginning part of phase one. The primary efficacy endpoint was the mean within-person percentage change in mosquito infectivity 2 days after primaquine treatment in participants who completed the study after optimisation of the infectivity assay, had both a pre-treatment infectivity measurement and at least one follow-up infectivity measurement, and who were given the correct primaquine dose. The safety endpoint was the mean within-person change in haemoglobin concentration during 28 days of study follow-up in participants with at least one follow-up visit. This study is registered with ClinicalTrials.gov, number NCT01743820. Findings Between Jan 2,2013, and Nov 27,2014, we enrolled 81 participants. In the primary analysis sample (n=71), participants in the 0.25 mg/kg primaquine dose group (n=15) and 0.5 mg/kg primaquine dose group (n=14) had significantly lower mean within-person reductions in infectivity at day 2-92.6% (95% CI 78.3-100; p=0.0014) for the 0.25 mg/kg group; and 75.0% (45.7-100; p=0.014) for the 0.5 mg/kg primaquine group compared with those in the control group (n=14; 11.3% [-27.4 to 50.0]). Reductions were not significantly different from control for participants assigned to the 0.0625 mg/kg dose group (n=16; 41.9% [1.4-82.5]; p=0.16) and the 0.125 mg/kg dose group (n=12; 54.9% [13.4-96.3]; p=0.096). No clinically meaningful or statistically significant drops in haemoglobin were recorded in any individual in the haemoglobin analysis (n=70) during follow-up. No serious adverse events were reported and adverse events did not differ between treatment groups. Interpretation A single dose of 0.25 mg/kg primaquine, given alongside dihydroartemisinin-piperaquine, was safe and efficacious for the prevention of P falciparum malaria transmission in boys and men who are not deficient in G6PD. Future studies should assess the safety of single-dose primaquine in G6PD-deficient individuals to define the therapeutic range of primaquine to enable the safe roll-out of community interventions with primaquine.Peer reviewe

Comparison of molecular quantification of Plasmodium falciparum gametocytes by Pfs25 qRT-PCR and QT-NASBA in relation to mosquito infectivity.

BACKGROUND: Quantifying gametocyte densities in natural malaria infections is important to estimate malaria transmission potential. Two molecular methods (Pfs25 mRNA quantitative reverse transcriptase PCR (qRT-PCR) and Pfs25 mRNA quantitative nucleic acid sequence based amplification (QT-NASBA)) are commonly used to determine gametocyte densities in clinical and epidemiological studies and allow gametocyte detection at densities below the microscopic threshold for detection. Here, reproducibility of these measurements and the association between estimated gametocyte densities and mosquito infection rates were compared. METHODS: To quantify intra- and inter-assay variation of QT-NASBA and qRT-PCR, a series of experiments was performed using culture-derived mature Plasmodium falciparum gametocytes from three different parasite isolates (NF54, NF135, NF166). Pfs25 mRNA levels were also determined in samples from clinical trials in Mali and Burkina Faso using both methods. Agreement between the two methods and association with mosquito infection rates in membrane feeding assays were assessed. RESULTS: Intra- and inter-assay variability was larger in QT-NASBA compared to qRT-PCR, particularly at low gametocyte densities (100 gametocyte per ?L). Samples collected in one of the two transmission studies had extremely low gametocyte densities by both molecular methods, which is suggestive of RNA degradation due to an unknown number of freeze-thaw cycles and illustrates the reliance of molecular gametocyte diagnostics on a reliable cold-chain. CONCLUSIONS: The experiments indicate that both qRT-PCR and QT-NASBA are of value for quantifying mature gametocytes in samples collected in field studies. For both assays, estimated gametocyte densities correlated well with mosquito infection rates. QT-NASBA is less reproducible than qRT-PCR, particularly for low gametocyte densities

Efficacy and safety of primaquine and methylene blue for prevention of Plasmodium falciparum transmission in Mali: a phase 2, single-blind, randomised controlled trial.

BACKGROUND: Primaquine and methylene blue are gametocytocidal compounds that could prevent Plasmodium falciparum transmission to mosquitoes. We aimed to assess the efficacy and safety of primaquine and methylene blue in preventing human to mosquito transmission of P falciparum among glucose-6-phosphate dehydrogenase (G6PD)-normal, gametocytaemic male participants. METHODS: This was a phase 2, single-blind, randomised controlled trial done at the Clinical Research Centre of the Malaria Research and Training Centre (MRTC) of the University of Bamako (Bamako, Mali). We enrolled male participants aged 5-50 years with asymptomatic P falciparum malaria. G6PD-normal participants with gametocytes detected by blood smear were randomised 1:1:1:1 in block sizes of eight, using a sealed-envelope design, to receive either sulfadoxine-pyrimethamine and amodiaquine, sulfadoxine-pyrimethamine and amodiaquine plus a single dose of 0·25 mg/kg primaquine, dihydroartemisinin-piperaquine, or dihydroartemisinin-piperaquine plus 15 mg/kg per day methylene blue for 3 days. Laboratory staff, investigators, and insectary technicians were masked to the treatment group and gametocyte density of study participants. The study pharmacist and treating physician were not masked. Participants could request unmasking. The primary efficacy endpoint, analysed in all infected patients with at least one infectivity measure before and after treatment, was median within-person percentage change in mosquito infectivity 2 and 7 days after treatment, assessed by membrane feeding. This study is registered with ClinicalTrials.gov, number NCT02831023. FINDINGS: Between June 27, 2016, and Nov 1, 2016, 80 participants were enrolled and assigned to the sulfadoxine-pyrimethamine and amodiaquine (n=20), sulfadoxine-pyrimethamine and amodiaquine plus primaquine (n=20), dihydroartemisinin-piperaquine (n=20), or dihydroartemisinin-piperaquine plus methylene blue (n=20) groups. Among participants infectious at baseline (54 [68%] of 80), those in the sulfadoxine-pyrimethamine and amodiaquine plus primaquine group (n=19) had a median 100% (IQR 100 to 100) within-person reduction in mosquito infectivity on day 2, a larger reduction than was noted with sulfadoxine-pyrimethamine and amodiaquine alone (n=12; -10·2%, IQR -143·9 to 56·6; p<0·0001). The dihydroartemisinin-piperaquine plus methylene blue (n=11) group had a median 100% (IQR 100 to 100) within-person reduction in mosquito infectivity on day 2, a larger reduction than was noted with dihydroartemisinin-piperaquine alone (n=12; -6·0%, IQR -126·1 to 86·9; p<0·0001). Haemoglobin changes were similar between gametocytocidal arms and their respective controls. After exclusion of blue urine, adverse events were similar across all groups (59 [74%] of 80 participants had 162 adverse events overall, 145 [90%] of which were mild). INTERPRETATION: Adding a single dose of 0·25 mg/kg primaquine to sulfadoxine-pyrimethamine and amodiaquine or 3 days of 15 mg/kg per day methylene blue to dihydroartemisinin-piperaquine was highly efficacious for preventing P falciparum transmission. Both primaquine and methylene blue were well tolerated. FUNDING: Bill & Melinda Gates Foundation, European Research Council

Performance of a Limiting-Antigen Avidity Enzyme Immunoassay for Cross-Sectional Estimation of HIV Incidence in the United States

Background: A limiting antigen avidity enzyme immunoassay (HIV-1 LAg-Avidity assay) was recently developed for cross-sectional HIV incidence estimation. We evaluated the performance of the LAg-Avidity assay alone and in multi-assay algorithms (MAAs) that included other biomarkers. Methods and Findings: Performance of testing algorithms was evaluated using 2,282 samples from individuals in the United States collected 1 month to >8 years after HIV seroconversion. The capacity of selected testing algorithms to accurately estimate incidence was evaluated in three longitudinal cohorts. When used in a single-assay format, the LAg-Avidity assay classified some individuals infected >5 years as assay positive and failed to provide reliable incidence estimates in cohorts that included individuals with long-term infections. We evaluated >500,000 testing algorithms, that included the LAg-Avidity assay alone and MAAs with other biomarkers (BED capture immunoassay [BED-CEIA], BioRad-Avidity assay, HIV viral load, CD4 cell count), varying the assays and assay cutoffs. We identified an optimized 2-assay MAA that included the LAg-Avidity and BioRad-Avidity assays, and an optimized 4-assay MAA that included those assays, as well as HIV viral load and CD4 cell count. The two optimized MAAs classified all 845 samples from individuals infected >5 years as MAA negative and estimated incidence within a year of sample collection. These two MAAs produced incidence estimates that were consistent with those from longitudinal follow-up of cohorts. A comparison of the laboratory assay costs of the MAAs was also performed, and we found that the costs associated with the optimal two assay MAA were substantially less than with the four assay MAA. Conclusions: The LAg-Avidity assay did not perform well in a single-assay format, regardless of the assay cutoff. MAAs that include the LAg-Avidity and BioRad-Avidity assays, with or without viral load and CD4 cell count, provide accurate incidence estimates

A Comparison of Two Measures of HIV Diversity in Multi-Assay Algorithms for HIV Incidence Estimation

Background: Multi-assay algorithms (MAAs) can be used to estimate HIV incidence in cross-sectional surveys. We compared the performance of two MAAs that use HIV diversity as one of four biomarkers for analysis of HIV incidence. Methods: Both MAAs included two serologic assays (LAg-Avidity assay and BioRad-Avidity assay), HIV viral load, and an HIV diversity assay. HIV diversity was quantified using either a high resolution melting (HRM) diversity assay that does not require HIV sequencing (HRM score for a 239 base pair env region) or sequence ambiguity (the percentage of ambiguous bases in a 1,302 base pair pol region). Samples were classified as MAA positive (likely from individuals with recent HIV infection) if they met the criteria for all of the assays in the MAA. The following performance characteristics were assessed: (1) the proportion of samples classified as MAA positive as a function of duration of infection, (2) the mean window period, (3) the shadow (the time period before sample collection that is being assessed by the MAA), and (4) the accuracy of cross-sectional incidence estimates for three cohort studies. Results: The proportion of samples classified as MAA positive as a function of duration of infection was nearly identical for the two MAAs. The mean window period was 141 days for the HRM-based MAA and 131 days for the sequence ambiguity-based MAA. The shadows for both MAAs were <1 year. Both MAAs provided cross-sectional HIV incidence estimates that were very similar to longitudinal incidence estimates based on HIV seroconversion. Conclusions: MAAs that include the LAg-Avidity assay, the BioRad-Avidity assay, HIV viral load, and HIV diversity can provide accurate HIV incidence estimates. Sequence ambiguity measures obtained using a commercially-available HIV genotyping system can be used as an alternative to HRM scores in MAAs for cross-sectional HIV incidence estimation

The prevalence, incidence and risk factors of herpes simplex virus type 2 infection among pregnant Zimbabwean women followed up nine months after childbirth

Background Herpes simplex virus type 2 (HSV-2) is the leading cause of genital ulcer disease worldwide. The virus can be transmitted to neonates and there are scarce data regarding incidence of HSV-2 among women in pregnancy and after childbirth. The aim of this study is to measure the incidence and risk factors for HSV-2 infection in women followed for 9 months after childbirth. Methods Pregnant women were consecutively enrolled late in pregnancy and followed at six weeks, four and nine months after childbirth. Stored samples were tested for HSV-2 at baseline and again at nine months after childbirth and HSV-2 seropositive samples at nine months after childbirth (seroconverters) were tested retrospectively to identify the seroconversion point. Results One hundred and seventy-three (50.9%) of the 340 consecutively enrolled pregnant women were HSV-2 seronegative at baseline. HSV-2 incidence rate during the 10 months follow up was 9.7 (95% CI 5.4-14.4)/100 and 18.8 (95% CI 13.9-26.1)/100 person years at risk (PYAR) at four months and nine months after childbirth respectively. Analysis restricted to women reporting sexual activity yielded higher incidence rates. The prevalence of HSV-2 amongst the HIV-1 seropositive was 89.3%. Risk factors associated with HSV-2 seropositivity were having other sexual partners in past 12 months (Prevalence Risk Ratio (PRR) 1.8 (95% CI 1.4-2.4) and presence of Trichomonas vaginalis (PRR 1.7 95% CI 1.4-2.1). Polygamy (Incidence Rate Ratio (IRR) 4.4, 95% CI 1.9-10.6) and young age at sexual debut (IRR 3.6, 95% CI 1.6-8.3) were associated with primary HSV-2 infection during the 10 months follow up. Conclusions Incidence of HSV-2 after childbirth is high and the period between late pregnancy and six weeks after childbirth needs to be targeted for prevention of primary HSV-2 infection to avert possible neonatal infections

Efficacy of Single-Dose Primaquine With Artemisinin Combination Therapy on Plasmodium falciparum Gametocytes and Transmission: An Individual Patient Meta-Analysis.

BACKGROUND: Since the World Health Organization recommended single low-dose (0.25 mg/kg) primaquine (PQ) in combination with artemisinin-based combination therapies (ACTs) in areas of low transmission or artemisinin-resistant Plasmodium falciparum, several single-site studies have been conducted to assess efficacy. METHODS: An individual patient meta-analysis to assess gametocytocidal and transmission-blocking efficacy of PQ in combination with different ACTs was conducted. Random effects logistic regression was used to quantify PQ effect on (1) gametocyte carriage in the first 2 weeks post treatment; and (2) the probability of infecting at least 1 mosquito or of a mosquito becoming infected. RESULTS: In 2574 participants from 14 studies, PQ reduced PCR-determined gametocyte carriage on days 7 and 14, most apparently in patients presenting with gametocytemia on day 0 (odds ratio [OR],?0.22; 95% confidence interval [CI], .17-.28 and OR,?0.12; 95% CI, .08-.16, respectively). Rate of decline in gametocyte carriage was faster when PQ was combined with artemether-lumefantrine (AL) compared to dihydroartemisinin-piperaquine (DP) (P?=?.010 for day 7). Addition of 0.25 mg/kg PQ was associated with near complete prevention of transmission to mosquitoes. CONCLUSIONS: Transmission blocking is achieved with 0.25 mg/kg PQ. Gametocyte persistence and infectivity are lower when PQ is combined with AL compared to DP