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New strategies and tools for Plasmodium falciparum case management and surveillance in the era of imminent resistance to artemisinin-based combination therapy in Tanzania.
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Women's and Children's Health. Muhimbili University of Health and Allied Sciences. (Andreas Mårtensson)
2020 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Artemether-lumefantrine has been an efficacious first line treatment for uncomplicated Plasmodium falciparum malaria in Tanzania since its introduction in 2006. Interest has developed in understanding the observation of high residual PCR determined positivity rates on day 3 after supervised artemether-lumefantrine treatment in the magnitude of almost 30% in previous assessments from 2015 in Bagamoyo district, Tanzania. Deep sequencing has recently been used to study these Bagamoyo parasites with delayed clearance, and the clearance times by PCR of some P. falciparum sub-populations were similar to artemisinin resistant parasites in Myanmar as assessed by microscopy, albeit lacking the described mutations in the Kelch13 propeller gene associated with artemisinin resistance. Moreover, molecular epidemiological studies from Bagamoyo, have shown temporal selection of lumefantrine associated genetic tolerance/resistance markers (pfmdr1 - N86, 184F, D1246 and pfcrt - K76) in the parasite population following wide scale use of artemether-lumefantrine but without signs of compromised treatment efficacy. On the other hand, traditional epidemiological studies have reported that imported malaria cases in Zanzibar from Tanzania mainland contribute to regressing the malaria elimination efforts in this pre-elimination part of the country.

This PhD project explored efficacy and safety of extending the artemether-lumefantrine regimen from standard 3 days to 6 days and adding single low dose primaquine (0.25mg/kg) as a new strategy that can be used in order to protect the therapeutic lifespan of artemether-lumefantrine. Also, whole-genome sequencing was used to study genomic epidemiology of P. falciparum population between Tanzania mainland and Zanzibar.

The results revealed that extended artemether-lumefantrine treatment did not have superior efficacy in the current context of artemether-lumefantrine sensitive P. falciparum parasites. However, the safety profile was excellent and similar to standard 3 days treatment. Parasite detection by molecular methods was 84% on day 3 after artemether-lumefantrine treatment. Meanwhile, significant decreases in the effective population sizes were inferred in both Tanzania mainland and Zanzibar parasite populations, that coincide with a period of decreasing malaria transmission in Tanzania. The parasite population from Tanzania mainland and Zanzibar were found to be connected, implying importation of cases from high transmission mainland to pre elimination regions of Zanzibar.

Utility of these results is during exploring options of alternative artemisinin-based combination therapy regimens to protect their therapeutic efficacy in an era of imminent artemisinin resistance in sub Saharan Africa. Moreover, the genomic epidemiological findings in this project may be of interest for malaria elimination programs, in the incorporation of molecular tools in future malaria elimination strategies and resistance surveillance, in the context of understanding importation of malaria from high to low transmission regions.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2020. , p. 100
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 1661
Keywords [en]
Malaria, Plasmodium falciparum, artemether-lumefantrine, drug resistance, Tanzania
National Category
Infectious Medicine
Research subject
Medical Science
Identifiers
URN: urn:nbn:se:uu:diva-407806ISBN: 978-91-513-0929-3 (print)OAI: oai:DiVA.org:uu-407806DiVA, id: diva2:1420848
Public defence
2020-05-25, Rudbecksalen, Rudbeck entreplan, C11, Uppsala, 10:00 (English)
Opponent
Supervisors
Funder
Sida - Swedish International Development Cooperation Agency, 16/9875007059Swedish Research Council, 2016-0577Available from: 2020-04-29 Created: 2020-04-01 Last updated: 2020-05-19
List of papers
1. Detection of Plasmodium falciparum by Light Microscopy, Loop-Mediated Isothermal Amplification, and Polymerase Chain Reaction on Day 3 after Initiation of Artemether-Lumefantrine Treatment for Uncomplicated Malaria in Bagamoyo District, Tanzania: A Comparative Trial
Open this publication in new window or tab >>Detection of Plasmodium falciparum by Light Microscopy, Loop-Mediated Isothermal Amplification, and Polymerase Chain Reaction on Day 3 after Initiation of Artemether-Lumefantrine Treatment for Uncomplicated Malaria in Bagamoyo District, Tanzania: A Comparative Trial
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2019 (English)In: American Journal of Tropical Medicine and Hygiene, ISSN 0002-9637, E-ISSN 1476-1645, Vol. 101, no 5, p. 1144-1147Article in journal (Refereed) Published
Abstract [en]

Microscopy-determined Plasmodium falciparum positivity rates exceeding 10% on day 3 after initiation of artemisinin-based combination therapy (ACT) is an important indicator of artemisinin resistance. However, microscopy does not detect low-density parasitemia, contrary to molecular tools such as loop-mediated isothermal amplification (LAMP) and polymerase chain reaction (PCR). We compared microscopy, LAMP, and PCR for detection of P. falciparum on day 3 after ACT in 256 patients with uncomplicated malaria in Bagamoyo District, Tanzania. Day 3 positivity rates were 0%, 84.8%, and 84.4% for each method, respectively. The sensitivity and specificity of LAMP against PCR was 100% (95% CI, 96.1-100) and 77.4% (95% CI, 58.9-90.4) when quantitative PCR-determined parasite densities were ≥ two parasites/µL. Loop-mediated isothermal amplification had comparable diagnostic accuracy to PCR and could potentially represent a field-friendly tool for determining day 3 positivity rates. However, what day 3 P. falciparum positivity determined using molecular methods represents needs to be further elucidated.

National Category
Public Health, Global Health, Social Medicine and Epidemiology Infectious Medicine
Identifiers
urn:nbn:se:uu:diva-397526 (URN)10.4269/ajtmh.19-0298 (DOI)000497187300034 ()31549618 (PubMedID)
Funder
Sida - Swedish International Development Cooperation Agency, Bil-Tz 16/9875007059Swedish Research Council, 2016-0577
Available from: 2019-11-21 Created: 2019-11-21 Last updated: 2020-04-01Bibliographically approved
2. Parasite clearance, cure rate, post-treatment prophylaxis and safety of standard 3-day versus an extended 6-day treatment of artemether-lumefantrine and a single low-dose primaquine for uncomplicated Plasmodium falciparum malaria in Bagamoyo district, Tanzania– a randomized controlled trial.
Open this publication in new window or tab >>Parasite clearance, cure rate, post-treatment prophylaxis and safety of standard 3-day versus an extended 6-day treatment of artemether-lumefantrine and a single low-dose primaquine for uncomplicated Plasmodium falciparum malaria in Bagamoyo district, Tanzania– a randomized controlled trial.
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

Background:

Artemisinin-based combination therapy (ACT) resistant Plasmodium falciparum represents an increasing threat to Africa. Extended ACT regimens from standard 3 to 6 days may represent a means to prevent its development and potential spread in Africa.

Methods: 

Standard 3-day treatment with artemether-lumefantrine (control) was compared to extended 6-day treatment and single low-dose primaquine (intervention); in a randomized controlled, parallel group, superiority clinical trial of patients aged 1-65 years with microscopy confirmed uncomplicated P. falciparum malaria, enrolled in Bagamoyo district, Tanzania. The study evaluated parasite clearance, including proportion of PCR detectable P. falciparum on days 5 and 7 (primary endpoint), cure rate, post-treatment prophylaxis, safety and tolerability. Clinical, and laboratory assessments, including ECG were conducted during 42 days of follow-up. Blood samples were collected for parasite detection (by microscopy and PCR), molecular genotyping and pharmacokinetic analyses. Kaplan-Meier survival analyses were done for both parasite clearance and recurrence. Results. A total of 280 patients were enrolled, 141 and 139 in the control and intervention arm, respectively, of whom 121 completed 42 days follow-up in each arm. There was no difference in proportion of PCR positivity across the arms at day 5 (80/130 (61.5%) vs 89/134 (66.4%), p=0.44), or day 7 (71/129 (55.0%) vs 70/134 (52.2%), p=0.71). Day 42 microscopy determined cure rates (PCR adjusted) were 97.4% (100/103) and 98.3% (110/112), p=0.65, in the control and intervention arm, respectively. Microscopy determined crude recurrent parasitemia during follow-up was 21/121 (17.4%) in the control and 14/121 (11.6%) in the intervention arm, p=0.20, and it took 34 days and 42 days in the respective arms for 90% of the patients to remain without recurrent parasitemia. Lumefantrine exposure was significantly higher in intervention arm from D3 to D42, but cardiac, biochemical and hematological safety was high and similar in both arms.

Conclusion:

Extended 6-day artemether-lumefantrine treatment and a single low-dose of primaquine was not superior to standard 3-day treatment for ACT sensitive P. falciparum infections, but importantly equally efficacious and safe. Thus, extended artemether-lumefantrine treatment may be considered as a future treatment regimen for ACT resistant P. falciparum, to prolong the therapeutic lifespan of ACT in Africa.

Keywords
Malaria, Plasmodium falciparum, artemether-lumefantrine, drug resistance, Tanzania
National Category
Medical and Health Sciences
Research subject
Infectious Diseases
Identifiers
urn:nbn:se:uu:diva-407921 (URN)
Funder
Sida - Swedish International Development Cooperation Agency, Bil-Tz 16/9875007059
Available from: 2020-03-31 Created: 2020-03-31 Last updated: 2020-04-01
3. Electrocardiographic safety evaluation of a prolonged artemether-lumefantrine treatment in patients with uncomplicated Plasmodium falciparum malaria in Bagamoyo District, Tanzania.
Open this publication in new window or tab >>Electrocardiographic safety evaluation of a prolonged artemether-lumefantrine treatment in patients with uncomplicated Plasmodium falciparum malaria in Bagamoyo District, Tanzania.
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(English)Manuscript (preprint) (Other academic)
Abstract [en]

 Background:

Extended artemisinin-based combination therapy (ACT) for treatment of uncomplicated Plasmodium falciparum malaria with already existing drug regimens such as artemether-lumefantrine, might be effective in tackling the emerging ACT resistance. However, given the history of cardiotoxicity among antimalarial drugs structurally similar to lumefantrine, the potential effect of extended artemether-lumefantrine treatment on the electrocardiographic (ECG) QTc interval is of high concern.

 

Method:

Male and non-pregnant females aged 1–65 years, diagnosed with uncomplicated P. falciparum malaria in Bagamoyo district, Tanzania, were randomized into two arms. The intervention arm received an extended, i.e. 6-day, course of artemether-lumefantrine and an additional single low-dose primaquine (0.25 mg/kg) administered together with the last artemether-lumefantrine dose. The control arm received the standard weight-based 3-day course. ECGs were performed at day 0 and 4–5 hours after the last dose at day 5. QT intervals were read manually using the tangent method and automatically. Bazett’s (QTcB) and Fridericia’s (QTcF) formulae were used for correction for heart rate. Descriptive statistics were used to calculate baseline characteristics and the number of supra-thresholds QTc intervals (QTc prolongation >500, change in QTc interval (DQTc) >60 ms). The mean change in QTc interval in and between the two arms was compared using the paired t-test and independent samples t-test, respectively.

 

Results:

A total of 195 patients were enrolled, 103 and 92 in the intervention and control arm, respectively. No patient experienced QTc intervals >500 ms on day 5 by both formulae. Patients with DQTc >60 ms, for QTcF were 6/103 (5.8%) vs 2/92 (2.2%) in the intervention and control arms, respectively. For QTcB was 2/103 (1.9%) vs 1/92 (1.1%) in the intervention and control arms, respectively. The mean difference in DQTc interval was statistically significant between the two arms with both correction formulae, 11.4 ms 50 (p= 0.010) and 13.4 ms (p= 0.001), for QTcB and QTcF respectively.

 

Conclusion:

The extended 6-day course of artemether-lumefantrine did not reveal clinically relevant QTc prolonging effects. However, significant QTcF prolongation and presence of patients with supra-threshold QTc values observed in the intervention arm underscore the importance of further monitoring of QTc parameters in extended artemether-lumefantrine treatment.

Keywords
Malaria, Plasmodium falciparum, cardiotoxicity, artemether–lumefantrine, Tanzania, prolonged treatment, artemisinin resistance, ECG
National Category
Clinical Medicine
Research subject
Infectious Diseases
Identifiers
urn:nbn:se:uu:diva-407932 (URN)
Funder
Swedish Research Council, 2016-0577
Available from: 2020-03-31 Created: 2020-03-31 Last updated: 2020-04-01
4. Falciparum malaria from coastal Tanzania and Zanzibar remains highly connected despite effective control efforts on the archipelago
Open this publication in new window or tab >>Falciparum malaria from coastal Tanzania and Zanzibar remains highly connected despite effective control efforts on the archipelago
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2020 (English)In: Malaria Journal, ISSN 1475-2875, E-ISSN 1475-2875, Vol. 19, article id 47Article in journal (Refereed) Published
Abstract [en]

Background: Tanzania's Zanzibar archipelago has made significant gains in malaria control over the last decade and is a target for malaria elimination. Despite consistent implementation of effective tools since 2002, elimination has not been achieved. Importation of parasites from outside of the archipelago is thought to be an important cause of malaria's persistence, but this paradigm has not been studied using modern genetic tools.

Methods: Whole-genome sequencing (WGS) was used to investigate the impact of importation, employing population genetic analyses of Plasmodium falciparum isolates from both the archipelago and mainland Tanzania. Ancestry, levels of genetic diversity and differentiation, patterns of relatedness, and patterns of selection between these two populations were assessed by leveraging recent advances in deconvolution of genomes from polyclonal malaria infections.

Results: Significant decreases in the effective population sizes were inferred in both populations that coincide with a period of decreasing malaria transmission in Tanzania. Identity by descent analysis showed that parasites in the two populations shared long segments of their genomes, on the order of 5 cM, suggesting shared ancestry within the last 10 generations. Even with limited sampling, two of isolates between the mainland and Zanzibar were identified that are related at the expected level of half-siblings, consistent with recent importation.

Conclusions: These findings suggest that importation plays an important role for malaria incidence on Zanzibar and demonstrate the value of genomic approaches for identifying corridors of parasite movement to the island.

Place, publisher, year, edition, pages
BMC, 2020
Keywords
Plasmodium, Malaria, Population genetics
National Category
Infectious Medicine
Identifiers
urn:nbn:se:uu:diva-407643 (URN)10.1186/s12936-020-3137-8 (DOI)000513890400003 ()31992305 (PubMedID)
Funder
Swedish Research Council
Available from: 2020-03-31 Created: 2020-03-31 Last updated: 2020-04-01Bibliographically approved

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