| 1. |
- Aydin-Schmidt, Berit, et al.
(author)
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Field Evaluation of a High Throughput Loop Mediated Isothermal Amplification Test for the Detection of Asymptomatic Plasmodium Infections in Zanzibar
- 2017
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In: PLOS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 12:1
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Journal article (peer-reviewed)abstract
- Background New field applicable diagnostic tools are needed for highly sensitive detection of residual malaria infections in pre-elimination settings. Field performance of a high throughput DNA extraction system for loop mediated isothermal amplification (HTP-LAMP) was therefore evaluated for detecting malaria parasites among asymptomatic individuals in Zanzibar. Methods HTP-LAMP performance was evaluated against real-time PCR on 3008 paired blood samples collected on filter papers in a community-based survey in 2015. Results The PCR and HTP-LAMP determined malaria prevalences were 1.6% (95% CI 1.3-2.4) and 0.7% (95% CI 0.4-1.1), respectively. The sensitivity of HTP-LAMP compared to PCR was 40.8% (CI95% 27.0-55.8) and the specificity was 99.9% (CI95% 99.8-100). For the PCR positive samples, there was no statistically significant difference between the geometric mean parasite densities among the HTP-LAMP positive (2.5 p/mu L, range 0.2-770) and HTP-LAMP negative (1.4 p/mu L, range 0.1-7) samples (p = 0.088). Two lab technicians analysed up to 282 samples per day and the HTP-LAMP method was experienced as user friendly. Conclusions Although field applicable, this high throughput format of LAMP as used here was not sensitive enough to be recommended for detection of asymptomatic low-density infections in areas like Zanzibar, approaching malaria elimination.
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| 2. |
- Jovel, Irina Tatiana, et al.
(author)
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Temporal and Seasonal Changes of Genetic Polymorphisms Associated with Altered Drug Susceptibility to Chloroquine, Lumefantrine, and Quinine in Guinea-Bissau between 2003 and 2012.
- 2015
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In: Antimicrobial Agents and Chemotherapy. - 0066-4804 .- 1098-6596. ; 59:2, s. 872-9
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Journal article (peer-reviewed)abstract
- In 2008, artemether-lumefantrine was introduced in Guinea-Bissau, West Africa, but quinine has also been commonly prescribed for the treatment of uncomplicated Plasmodium falciparum malaria. An efficacious high-dose chloroquine treatment regimen was used previously. Temporal and seasonal changes of genetic polymorphisms associated with altered drug susceptibility to chloroquine, lumefantrine, and quinine have been described. P. falciparum chloroquine resistance transporter (pfcrt) K76T, pfmdr1 gene copy numbers, pfmdr1 polymorphisms N86Y and Y184F, and pfmdr1 sequences 1034 to 1246 were determined using PCR-based methods. Blood samples came from virtually all (n = 1,806) children <15 years of age who had uncomplicated P. falciparum monoinfection and presented at a health center in suburban Bissau (from 2003 to 2012). The pfcrt K76T and pfmdr1 N86Y frequencies were stable, and seasonal changes were not seen from 2003 to 2007. Since 2007, the mean annual frequencies increased (P < 0.001) for pfcrt 76T (24% to 57%), pfmdr1 N86 (72% to 83%), and pfcrt 76 + pfmdr1 86 TN (10% to 27%), and pfcrt 76T accumulated during the high transmission season (P = 0.001). The pfmdr1 86 + 184 NF frequency increased from 39% to 66% (from 2003 to 2011; P = 0.004). One sample had two pfmdr1 gene copies. pfcrt 76T was associated with a lower parasite density (P < 0.001). Following the discontinuation of an effective chloroquine regimen, probably highly artemether-lumefantrine-susceptible P. falciparum (with pfcrt 76T) accumulated, possibly due to suboptimal use of quinine and despite a fitness cost linked to pfcrt 76T. (The studies reported here were registered at ClinicalTrials.gov under registration no. NCT00137514 [PSB-2001-chl-amo], NCT00137566 [PSB-2004-paracetamol], NCT00426439 [PSB-2006-coartem], NCT01157689 [AL-eff 2010], and NCT01704508 [Eurartesim 2012].).
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| 3. |
- Jovel, Irina T., et al.
(author)
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Unexpected selections of Plasmodium falciparum polymorphisms in previously treatment-naive areas after monthly presumptive administration of three different anti-malarial drugs in Liberia 1976-78
- 2017
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In: Malaria Journal. - : Springer Science and Business Media LLC. - 1475-2875. ; 16
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Journal article (peer-reviewed)abstract
- Background: To assess the effect on malaria prevalence, village specific monthly administrations of pyrimethamine, chlorproguanil, chloroquine or placebo were given to children in four previously treatment-naive Liberian villages, 1976-78. Plasmodium falciparum in vivo resistance developed to pyrimethamine only. Selection of molecular markers of P.falciparum resistance after 2 years of treatment are reported. Methods: Blood samples were collected from 191 study children in a survey in 1978. Polymorphisms in pfcrt, pfmdr1, pfdhfr, pfdhps, pfmrp1 and pfnhe1 genes were determined using PCR-based methods. Results: Pfcrt 72-76 CVIET was found in one chloroquine village sample, all remaining samples had pfcrt CVMNK. Pfmdr1 N86 prevalence was 100%. A pfmdr1 T1069(ACT -> ACG) synonymous polymorphism was found in 30% of chloroquine village samples and 3% of other samples (P = 0.008). Variations in pfnhe1 block I were found in all except the chloroquine treated village (P < 0.001). Resistance associated pfdhfr 108N prevalence was 2% in the pyrimethamine village compared to 45-65% elsewhere, including the placebo village (P = 0.001). Conclusions: Chloroquine treatment possibly resulted in the development of pfcrt 72-76 CVIET. Selection of pfmdr1 T1069(ACG) and a pfnhe1 block 1 genotypes indicates that chloroquine treatment exerted a selective pressure on P. falciparum. Pyrimethamine resistance associated pfdhfr 108N was present prior to the introduction of any drug. Decreased pfdhfr 108N frequency concurrent with development of pyrimethamine resistance suggests a non-pfdhfr polymorphisms mediated resistance mechanism.
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| 4. |
- Jovel Quiñonez Dalmau, Irina Tatiana
(author)
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Aspects of molecular markers in drug resistant malaria
- 2014
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Doctoral thesis (other academic/artistic)abstract
- BACKGROUND: There were an estimated 207 million cases of malaria in 2012 of which 91% were due to Plasmodium falciparum. Antimalarial drug resistance constitutes a major problem in the efforts to control malaria. Drug resistance typically arises by the gradual accumulation of genetic changes that enable parasites to tolerate gradually increasing drug concentrations until fully resistant parasites develop. The aim of this thesis was to determine temporal and geographic frequencies of genetic polymorphisms linked to P. falciparum and P. vivax resistance after exposure to various antimalarial drugs. METHODS: Polymorphisms in P. falciparum and P. vivax multidrug resistance 1 (pfmdr1 and pvmdr1), dihydrofolate reductase (pfdhfr and pvdhfr) genes, P. falciparum chloroquine resistance transporter (pfcrt), dihydropteroate synthase (pfdhps), V-type H+ pyrophosphatase (pfvp2), Na+/H+ exchanger-1 (pfnhe1) and multidrug-resistant protein 1 (pfmrp1) genes were determined in field samples. P. falciparum and P. vivax samples from Honduras, where drug resistant malaria has not been detected, were analysed for paper I. Samples from Honduras, Colombia, Liberia, Guinea-Bissau, Tanzania, Iran, Thailand and Vanuatu that represented varying origins and frequency of chloroquine (CQ) resistant P. falciparum malaria were analysed for paper II. Samples from 4 villages in Liberia in 1978 after children were administered village specific treatments of CQ, pyrimethamine (PYR), chlorproguanil or placebo monthly for 2 years were used for paper III. Samples (n=1806) from virtually all children aged <15 years with uncomplicated malaria between 2003 and 2012 in Guinea- Bissau were analysed for paper IV. Artemether-lumefantrine replaced an effective high dose CQ treatment in Guinea-Bissau in 2008 but quinine was prescribed to 43% of the children between 2010 and 2012. RESULTS and CONCLUSIONS: Paper I. No genetic polymorphisms associated with CQ or sulphadoxine-pyrimethamine (SP) resistance were found indicating that P. falciparum remains CQ sensitive in Honduras. CQ resistance associated pvmdr1 976F (7/37) and SP resistance associated pvdhfr 57L+58R (2/57) were detected suggesting a degree of tolerance to CQ and SP in P. vivax. Paper II. Pfvp2 SNPs were found in only 20/367 patient samples. Nevertheless, the V405 + K582 + P711 haplotype was associated with pfcrt 76T (P=0.007) in line with previous in vitro data. However, the limited variation suggests that the results should be interpreted with caution. Paper III. Pfcrt 72-76 CVIET without other downstream resistance associated SNPs occurred in 1/178 samples from the village using CQ in Liberia. CVIET might have developed in Liberia in 1978 but the haplotype was not detected in 1981 suggesting that it probably did not provide a selective advantage in this area. Pfdhfr 108N was significantly lower (1/46, P=0.001) in the Liberian village using PYR compared with other villages (64/123). Pfdhfr 108N probably occurred as a wild type allele prior to the introduction of antifolates in Liberia. Decreased pfdhfr 108N frequency concurrently with the development of in vivo PYR resistance suggests a resistant mechanism other than pfdhfr point mutations. Paper IV. Between 2003 and 2007 pfcrt K76T and pfmdr1 N86Y frequencies were stable in Guinea-Bissau. After 2007, mean annual pfcrt 76T (24->57%) and pfmdr1 86N (72->83%) frequencies increased (p<0.001). Increased pfcrt 76T was probably driven by quinine that was as commonly used artemether-lumefantrine. Pfmdr1 86+184 NF frequency increased (39->66%) between 2003 and 2011 (p=0.004), possibly driven by artemether-lumefantrine.
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| 5. |
- Jovel Quiñonez Dalmau, Irina Tatiana
(author)
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Spread and distribution of drug resistance and compensatory mutations in Plasmodium falciparum
- 2012
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Licentiate thesis (other academic/artistic)abstract
- In 2010 there were an estimated 216 million cases of malaria worldwide. In Honduras there were ~9000 cases of which 88% were due to Plasmodium vivax mono-infection. Chloroquine (CQ) resistant Plasmodium falciparum have spread throughout the world curtailing its use. The only exception appears to be north of Panama where CQ reportedly remains efficacious and the drug of choice for treating both P. falciparum and P. vivax. Resistance to antimalarials is associated with specific genetic polymorphisms and recently a putative H+ pump (pfvp2) has been suggested to be linked to CQ resistant P. falciparum. The aim of this thesis was to identify resistance associated genetic polymorphisms in P. falciparum and P.vivax from Honduras and to describe the worldwide distribution of pfvp2 polymorphisms and their correlation to CQ resistance. Resistance associated genetic polymorphisms in P. falciparum and P. vivax multidrug resistance gene (pfmdr1 and pvmdr1), dihydrofolate reductase (pfdhfr and pvdhfr), P. falciparum chloroquine resistance transporter (pfcrt), dihydropteroate synthase (pfdhps) and V-type H+ pyrophosphatase (pfvp2) were identified in field samples using PCR based methods. From Honduras, 37 P. falciparum and 64 P. vivax samples, collected from symptomatic patients were used. In addition, 50 samples from each of Colombia, Liberia, Guinea-Bissau, Tanzania, Iran, Thailand and Vanuatu were used. The samples represented a time period from 1978 to 2009 and areas with different prevalence of CQ resistant P. falciparum. In samples from Honduras no genetic polymorphisms associated with CQ or sulphadoxinepyrimethamine (SP) resistance were found in P. falciparum. In P. vivax, the CQ resistance associated pvmdr1 976F allele was found in 7/37 samples and the SP resistance associated pvdhfr 57L+58R alleles were found in 2/57 samples. When analysing the worldwide collection of samples, the pfvp2 405V, 582K and 711P haplotype was associated with the for CQ resistance essential allele, pfcrt 76T (P=0.007). Samples with pfvp2 405I and/or 582R and/or 711S were significantly more common in Liberia in 1978-1980 (P=0.01), all African countries (P=0.004) and all African countries + Honduras (P=0.01) compared to the rest of the world. Our results suggest that P. falciparum and P. vivax in Honduras are sensitive to CQ and SP. However, small numbers of P. vivax had genetic polymorphisms suggesting a degree of tolerance to CQ and SP. The association between pfcrt 76T and the pfvp2 405V, 582K and 711P haplotype suggest that this haplotype is associated with CQ resistance. This is in line with previous research that has described increased expression of pfvp2 during CQ exposure. The higher frequency of pfvp2 405I and/or 582R and/or 711S in CQ sensitive settings in Africa and Honduras suggests a larger variation in the pfvp2 genome prior to the spread of CQ resistance further supporting the association between pfvp2 and CQ resistance.
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| 6. |
- Malmberg, Maja, et al.
(author)
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Temporal trends of molecular markers associated with artemether-lumefantrine tolerance/resistance in Bagamoyo district, Tanzania.
- 2013
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In: Malaria journal. - : Springer Science and Business Media LLC. - 1475-2875. ; 12
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Journal article (peer-reviewed)abstract
- Development and spread of Plasmodium falciparum resistance to artemisinin-based combination therapy (ACT) constitutes a major threat to recent global malaria control achievements. Surveillance of molecular markers could act as an early warning system of ACT-resistance before clinical treatment failures are apparent. The aim of this study was to analyse temporal trends of established genotypes associated with artemether-lumefantrine tolerance/resistance before and after its deployment as first-line treatment for uncomplicated malaria in Tanzania 2006.
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| 7. |
- Morris, Ulrika, et al.
(author)
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A cluster randomised controlled trial of two rounds of mass drug administration in Zanzibar, a malaria pre-elimination setting-high coverage and safety, but no significant impact on transmission.
- 2018
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In: BMC Medicine. - : Springer Science and Business Media LLC. - 1741-7015. ; 16:1
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Journal article (peer-reviewed)abstract
- BACKGROUND: Mass drug administration (MDA) has the potential to interrupt malaria transmission and has been suggested as a tool for malaria elimination in low-endemic settings. This study aimed to determine the effectiveness and safety of two rounds of MDA in Zanzibar, a pre-elimination setting.METHODS: A cluster randomised controlled trial was conducted in 16 areas considered as malaria hotspots, with an annual parasite index of > 0.8%. The areas were randomised to eight intervention and eight control clusters. The intervention included two rounds of MDA with dihydroartemisinin-piperaquine and single low-dose primaquine 4 weeks apart in May-June 2016. Primary and secondary outcomes were cumulative confirmed malaria case incidences 6 months post-MDA and parasite prevalences determined by PCR 3 months post-MDA. Additional outcomes included intervention coverage, treatment adherence, occurrence of adverse events, and cumulative incidences 3, 12, and 16 months post-MDA.RESULTS: Intervention coverage was 91.0% (9959/10944) and 87.7% (9355/10666) in the first and second rounds, respectively; self-reported adherence was 82.0% (881/1136) and 93.7% (985/1196). Adverse events were reported in 11.6% (147/1268) and 3.2% (37/1143) of post-MDA survey respondents after both rounds respectively. No serious adverse event was reported. No difference in cumulative malaria case incidence was observed between the control and intervention arms 6 months post-MDA (4.2 and 3.9 per 1000 population; p = 0.94). Neither was there a difference in PCR-determined parasite prevalences 3 months post-MDA (1.4% and 1.7%; OR = 1.0, p = 0.94), although having received at least the first MDA was associated with reduced odds of malaria infection (aOR = 0.35; p = 0.02). Among confirmed malaria cases at health facilities, 26.0% and 26.3% reported recent travel outside Zanzibar in the intervention and control shehias (aOR ≥ 85; p ≤ 0.001).CONCLUSIONS: MDA was implemented with high coverage, adherence, and tolerability. Despite this, no significant impact on transmission was observed. The findings suggest that two rounds of MDA in a single year may not be sufficient for a sustained impact on transmission in a pre-elimination setting, especially when the MDA impact is restricted by imported malaria. Importantly, this study adds to the limited evidence for the use of MDA in low transmission settings in sub-Saharan Africa.TRIAL REGISTRATION: ClinicalTrials.gov, NCT02721186 (registration date: March 29, 2016).
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| 8. |
- Mwaiswelo, Richard, et al.
(author)
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Adding a single low-dose of primaquine (0.25 mg/kg) to artemether-lumefantrine did not compromise treatment outcome of uncomplicated Plasmodium falciparum malaria in Tanzania : a randomized, single-blinded clinical trial
- 2016
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In: Malaria Journal. - : Springer Science and Business Media LLC. - 1475-2875. ; 15
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Journal article (peer-reviewed)abstract
- Background: The World Health Organization (WHO) recently recommended the addition of a single low-dose of the gametocytocidal drug primaquine (PQ) to artemisinin-based combination therapy (ACT) in low transmission settings as a component of pre-elimination or elimination programmes. However, it is unclear whether that influences the ACT cure rate. The study assessed treatment outcome of artemether-lumefantrine (AL) plus a single PQ dose (0.25 mg/kg) versus standard AL regimen for treatment of acute uncomplicated Plasmodium falciparum malaria in Tanzania. Methods: A randomized, single-blinded, clinical trial was conducted in Yombo, Bagamoyo district, Tanzania. Acute uncomplicated P. falciparum malaria patients aged >= 1 year, with the exception of pregnant and lactating women, were enrolled and treated with AL plus a single PQ dose (0.25 mg/kg) or AL alone under supervision. PQ was administered together with the first AL dose. Clinical and laboratory assessments were performed at 0, 8, 24, 36, 48, 60, and 72 h and on days 7, 14, 21, and 28. The primary end-point was a polymerase chain reaction (PCR)-adjusted adequate clinical and parasitological response (ACPR) on day 28. Secondary outcomes included: fever and asexual parasitaemia clearance, proportion of patients with PCR-determined parasitaemia on day 3, and proportion of patients with Pfmdr1 N86Y and Pfcrt K76T on days 0, 3 and day of recurrent infection. Results: Overall 220 patients were enrolled, 110 were allocated AL + PQ and AL, respectively. Parasite clearance by microscopy was fast, but PCR detectable parasitaemia on day 3 was 31/109 (28.4 %) and 29/108 (26.9 %) in patients treated with AL + PQ and AL, respectively (p = 0.79). Day 28 PCR-adjusted ACPR and re-infection rate was 105/105 (100 %) and 101/102 (99 %) (p = 0.31), and 5/107 (4.7 %) and 5/8 (4.8 %) (p = 0.95), in AL + PQ and AL arm, respectively. There was neither any statistically significant difference in the proportion of Pfmdr1 N86Y or Pfcrt K76T between treatment arms on days 0, 3 and day of recurrent infection, nor within treatment arms between days 0 and 3 or day 0 and day of recurrent infection. Conclusion: The new WHO recommendation of adding a single low-dose of PQ to AL did not compromise treatment outcome of uncomplicated P. falciparum malaria in Tanzania.
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| 9. |
- Mwaiswelo, Richard, et al.
(author)
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Prevalence of and Risk Factors Associated with Polymerase Chain Reaction-Determined Plasmodium falciparum Positivity on Day 3 after Initiation of Artemether-Lumefantrine Treatment for Uncomplicated Malaria in Bagamoyo District, Tanzania
- 2019
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In: American Journal of Tropical Medicine and Hygiene. - : American Society of Tropical Medicine and Hygiene. - 0002-9637 .- 1476-1645. ; 100:5, s. 1179-1186
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Journal article (peer-reviewed)abstract
- Prevalence of and risk factors associated with polymerase chain reaction (PCR)-determined Plasmodium falciparum positivity were assessed on day 3 after initiation of treatment, pre-implementation and up to 8 years post-deployment of artemether-lumefantrine as first-line treatment for uncomplicated malaria in Bagamoyo district, Tanzania. Samples originated from previously reported trials conducted between 2006 and 2014. Cytochrome b-nested PCR was used to detect malaria parasites from blood samples collected on a filter paper on day 3. Chi-square and McNemar chi-squared tests, logistic regression models, and analysis of variance were used as appropriate. Primary outcome was based on the proportion of patients with day 3 PCR-determined P. falciparum positivity. Overall, 256/584 (43.8%) of screened patients had day 3 PCR-determined positivity, whereas only 2/584 (0.3%) had microscopy-determined asexual parasitemia. Day 3 PCR-determined positivity increased from 28.0% (14/50) in 2006 to 74.2% (132/178) in 2007-2008 and declined, thereafter, to 36.0% (50/139) in 2012-2013 and 27.6% (60/217) in 2014. When data were pooled, pretreatment microscopy-determined asexual parasitemia ≥ 100,000/µL, hemoglobin < 10 g/dL, age < 5 years, temperature ≥ 37.5°C, and year of study 2007-2008 and 2012-2013 were significantly associated with PCR-determined positivity on day 3. Significant increases in P. falciparum multidrug resistance gene 1 N86 and P. falciparum chloroquine resistant transporter K76 across years were not associated with PCR-determined positivity on day 3. No statistically significant association was observed between day 3 PCR-determined positivity and PCR-adjusted recrudescence. Day 3 PCR-determined P. falciparum positivity remained common in patients treated before and after implementation of artemether-lumefantrine in Bagamoyo district, Tanzania. However, its presence was associated with pretreatment characteristics. Trials registration numbers: NCT00336375, ISRCTN69189899, NCT01998295, and NCT02090036.
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| 10. |
- Mwaiswelo, Richard, et al.
(author)
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Safety of a single low-dose of primaquine in addition to standard artemether-lumefantrine regimen for treatment of acute uncomplicated Plasmodium falciparum malaria in Tanzania
- 2016
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In: Malaria Journal. - : Springer Science and Business Media LLC. - 1475-2875. ; 15
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Journal article (peer-reviewed)abstract
- Background: This study assessed the safety of the new World Health Organization (WHO) recommendation of adding a single low-dose of primaquine (PQ) to standard artemisinin-based combination therapy (ACT), regardless of individual glucose-6-phosphate dehydrogenase (G6PD) status, for treatment of acute uncomplicated Plasmodium falciparum malaria in Tanzania. Methods: Men and non-pregnant, non-lactating women aged >= 1 year with uncomplicated P. falciparum malaria were enrolled and randomized to either standard artemether-lumefantrine (AL) regimen alone or with a 0.25 mg/kg single-dose of PQ. PQ was administered concomitantly with the first AL dose. All drug doses were supervised. Safety was evaluated between days 0 and 28. G6PD status was assessed using rapid test (CareStart (TM)) and molecular genotyping. The primary endpoint was mean percentage relative reduction in haemoglobin (Hb) concentration (g/dL) between days 0 and 7 by genotypic G6PD status and treatment arm. Results: Overall, 220 patients, 110 per treatment arm, were enrolled, of whom 33/217 (15.2 %) were phenotypically G6PD deficient, whereas 15/110 (13.6 %) were genotypically hemizygous males, 5/110 (4.5 %) homozygous females and 22/110 (20 %) heterozygous females. Compared to genotypically G6PD wild-type/normal [ 6.8, 95 % confidence interval (CI) 4.67-8.96], only heterozygous patients in AL arm had significant reduction in day-7 mean relative Hb concentration (14.3, 95 % CI 7.02-21.55, p=0.045), however, none fulfilled the pre-defined haemolytic threshold value of >= 25 % Hb reduction. After adjustment for baseline parasitaemia, Hb, age and sex the mean relative Hb reduction was not statistically significant in both heterozygous and hemizygous/homozygous patients in both arms. A majority of the adverse events (AEs) were mild and unrelated to the study drugs. However, six (4.4 %) episodes, three per treatment arm, of acute haemolytic anaemia occurred between days 0 and 7. Three occurred in phenotypically G6PD deficient patients, two in AL and one in AL + PQ arm, but none in genotypically hemizygous/homozygous patients. All patients with acute haemolytic anaemia recovered without medical intervention. Conclusion: The findings support that the WHO recommendation of adding a single low-dose of PQ to standard AL regimen is safe for the treatment of acute uncomplicated P. falciparum malaria regardless of G6PD status in Tanzania.
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