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Research article
George Bwire
Muhimbili University of Health and Allied Sciences School of Pharmacy
Corresponding Author
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Background : High levels of Plasmodium falciparum resistance prompted withdrawal of sulfadoxine-pyrimethamine (SP) as the first-line treatment for uncomplicated malaria in Tanzania. However, SP was limited for intermittent preventive treatment during pregnancy (IPTp) especially where there is moderate to high malaria transmission. This study reports the patterns of P. falciparum dihydrofolate reductase ( Pfdhfr ) and dihydropteroate synthetase ( Pfdhps ) mutations.
Methods: Parasite genomic DNA was extracted from dried blood spots prepared by finger prick. Batched samples (384) were sequenced in a single MiSeq lane combining all PCR products. Samples were de-plexed using the multiplexing adapters and individual CRAM files were aligned to a modified amplicon reference genome. Genotyping of Pfdhfr and Pfdhps mutations were done using bcftools as well as custom scripts to filter and translate genotypes into drug resistance haplotypes.
Results: The Pfdhfr was analyzed from 445 samples, the wild type (WT) Pfdhfr haplotype NCSI was detected in only six (1.3%) samples. Triple Pfdhfr IRN I haplotype was dominant, contributing to 84% (n=374) of haplotypes. The total of 446 samples were studied for Pfdhps . WT for Pfdhps was found in 6.7% (n=30) of all samples detected. Double Pfdhps haplotype (S GE AA) accounted for 83% of mutations of the Pfdhps gene. The overall prevalence of K540E was 90.4% (n=396) while A581G was 1.1% (n=5). Additionally, 91.4% (n=447) genotypes where detected from 489 sequenced samples. Of 447 genotypes detected only 0.9% (n=4) of samples were WT (SAKAA-NCSI). Quintuple mutation, S GE AA- IRN I accounted 71.4% of concomitant Pfdhfr/Pfdhps mutations where 0.2% (n=1) had septuple mutation, AG K GS - IRN I.
Conclusions : Despite the high prevalence of mutations in Pfdhfr and Pfdhps gene but the current mutations at Pfdhfr K540E and Pfdhps A581G are within the recommended WHO range, stopping IPTp-SP is recommended in areas where the Pfdhfr K540E prevalence is >95 % and Pfdhps A581G is >10 % as SP is likely to be ineffective). Nevertheless, saturation in Pfdhfr and Pfdhps haplotypes is alarming, therefore screening for alternative antimalarial drug for IPTp-SP is recommended.
Keywords
Pfdhfr, Pfdhps, Sulfadoxine-pyrimethamine, Resistance, Tanzania
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Reviewer #3 agreed
On 10 Jun, 2020
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Received 10 Jun, 2020
Review #2 received
Received 24 May, 2020
Reviewer #2 agreed
On 22 May, 2020
Review #1 received
Received 22 May, 2020
Reviewers invited
Invitations sent on 19 May, 2020
Reviewer #1 agreed
On 19 May, 2020
Editor assigned
On 02 Apr, 2020
Submission checks complete
On 01 Apr, 2020
Editor invited
On 01 Apr, 2020
Version 1
Posted 06 Jan, 2020
No comments provided
Editorial decision: Major revision
On 02 Apr, 2020
Review #2 received
Received 27 Mar, 2020
Reviewer #2 agreed
On 05 Mar, 2020
Review #1 received
Received 19 Jan, 2020
Editor assigned
On 06 Jan, 2020
Reviewers invited
Invitations sent on 06 Jan, 2020
Reviewer #1 agreed
On 06 Jan, 2020
Submission checks complete
On 02 Jan, 2020
Editor invited
On 02 Jan, 2020
First submitted
On 18 Dec, 2019
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See the published version of this preprint at BMC Infectious Diseases.
This is a preprint, a preliminary version of a manuscript that has not completed peer review at a journal. Research Square does not conduct peer review prior to posting preprints. The posting of a preprint on this server should not be interpreted as an endorsement of its validity or suitability for dissemination as established information or for guiding clinical practice.
Learn more about In Review
Research article
George Bwire
Muhimbili University of Health and Allied Sciences School of Pharmacy
Corresponding Author
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
Peer Review Timeline
CURRENT STATUS: Published
View the published article at BMC Infectious Diseases.
No community comments so far
Reviewer #3 agreed
On 10 Jun, 2020
Review #3 received
Received 10 Jun, 2020
Review #2 received
Received 24 May, 2020
Reviewer #2 agreed
On 22 May, 2020
Review #1 received
Received 22 May, 2020
Reviewers invited
Invitations sent on 19 May, 2020
Reviewer #1 agreed
On 19 May, 2020
Editor assigned
On 02 Apr, 2020
Submission checks complete
On 01 Apr, 2020
Editor invited
On 01 Apr, 2020
Version 1
Posted 06 Jan, 2020
No comments provided
Editorial decision: Major revision
On 02 Apr, 2020
Review #2 received
Received 27 Mar, 2020
Reviewer #2 agreed
On 05 Mar, 2020
Review #1 received
Received 19 Jan, 2020
Editor assigned
On 06 Jan, 2020
Reviewers invited
Invitations sent on 06 Jan, 2020
Reviewer #1 agreed
On 06 Jan, 2020
Submission checks complete
On 02 Jan, 2020
Editor invited
On 02 Jan, 2020
First submitted
On 18 Dec, 2019
Metrics
Comments: 0
PDF Downloads: ...
HTML Views: ...
License:
This work is licensed under a CC BY 4.0 License. Read Full License
View the published article at BMC Infectious Diseases.
Background : High levels of Plasmodium falciparum resistance prompted withdrawal of sulfadoxine-pyrimethamine (SP) as the first-line treatment for uncomplicated malaria in Tanzania. However, SP was limited for intermittent preventive treatment during pregnancy (IPTp) especially where there is moderate to high malaria transmission. This study reports the patterns of P. falciparum dihydrofolate reductase ( Pfdhfr ) and dihydropteroate synthetase ( Pfdhps ) mutations.
Methods: Parasite genomic DNA was extracted from dried blood spots prepared by finger prick. Batched samples (384) were sequenced in a single MiSeq lane combining all PCR products. Samples were de-plexed using the multiplexing adapters and individual CRAM files were aligned to a modified amplicon reference genome. Genotyping of Pfdhfr and Pfdhps mutations were done using bcftools as well as custom scripts to filter and translate genotypes into drug resistance haplotypes.
Results: The Pfdhfr was analyzed from 445 samples, the wild type (WT) Pfdhfr haplotype NCSI was detected in only six (1.3%) samples. Triple Pfdhfr IRN I haplotype was dominant, contributing to 84% (n=374) of haplotypes. The total of 446 samples were studied for Pfdhps . WT for Pfdhps was found in 6.7% (n=30) of all samples detected. Double Pfdhps haplotype (S GE AA) accounted for 83% of mutations of the Pfdhps gene. The overall prevalence of K540E was 90.4% (n=396) while A581G was 1.1% (n=5). Additionally, 91.4% (n=447) genotypes where detected from 489 sequenced samples. Of 447 genotypes detected only 0.9% (n=4) of samples were WT (SAKAA-NCSI). Quintuple mutation, S GE AA- IRN I accounted 71.4% of concomitant Pfdhfr/Pfdhps mutations where 0.2% (n=1) had septuple mutation, AG K GS - IRN I.
Conclusions : Despite the high prevalence of mutations in Pfdhfr and Pfdhps gene but the current mutations at Pfdhfr K540E and Pfdhps A581G are within the recommended WHO range, stopping IPTp-SP is recommended in areas where the Pfdhfr K540E prevalence is >95 % and Pfdhps A581G is >10 % as SP is likely to be ineffective). Nevertheless, saturation in Pfdhfr and Pfdhps haplotypes is alarming, therefore screening for alternative antimalarial drug for IPTp-SP is recommended.
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