title: High levels of sulphadoxine-pyrimethamine resistance Pfdhfr-Pfdhps quintuple mutations: a cross sectional survey of six regions in Tanzania.
abstract: BACKGROUND
  In 2006, the first-line anti-malarial drug treatment in Tanzania was changed from sulphadoxine-pyrimethamine (SP) to artemether-lumefantrine (ALu), an artemisinin-based combination (ACT), since when the use of SP has been restricted for intermittent preventive treatment in pregnancy (IPTp). A number of Plasmodium falciparum mutations are known to be associated with resistance to SP, but it is not known if the prevalence of these mutations is increasing or decreasing under the conditions of reduced levels of SP use. This study reports on the current SP resistant quintuple Pfdhfr-Pfdhps mutations in six regions of Tanzania.
  
  
  METHODS
  Finger-prick blood on filter paper and rapid diagnostic test strips from P. falciparum-positive individuals of all age groups attending health facilities in six regions of Tanzania between June 2010 and August 2011 were obtained. Using chelex-100 extracted DNA, genotyping was done for mutations on codons 51, 59 and 108 of Pfdhfr and 437 and 540 of Pfdhps genes using PCR-RFLP technique.
  
  
  RESULTS
  A total of 802 malaria-positive samples were screened and genotyped. The prevalence of Pfdhfr 51I, Pfdhps 437G and 540E varied between the regions (p < 0.001) whereas Pfdhfr 59R (FE 10.79, p = 0.225) and 108 N (FE 10.61, p = 0.239) did not vary between the regions. The Pfdhfr triple mutant was above 84% and close to fixation levels in all regions, whereas the Pfdhps double mutation ranged from 43.8 to 97% between the regions. The quintuple mutant (IRNGE) was the most prevalent in all regions and it varied significantly from 37.5 to 90.2% (χ(2) = 1.11, p <0.001).
  
  
  CONCLUSIONS
  There is evidence of persistent high levels of SP resistance markers in Tanzania with evidence of quintuple mutations that are likely to become fixed in the population. This threatens the future of SP not only in IPTp programmes, but as a combination drug for ACT. Continuous monitoring of SP-IPTp efficacy should be encouraged subsequent to searching for alternative drugs for IPTp in East Africa.
authors.forename: Sungwa I
authors.initials: SI
authors.lastname: Matondo
authors.forename: Godfrey S
authors.initials: GS
authors.lastname: Temba
authors.forename: Adelaida A
authors.initials: AA
authors.lastname: Kavishe
authors.forename: Julius S
authors.initials: JS
authors.lastname: Kauki
authors.forename: Akili
authors.initials: A
authors.lastname: Kalinga
authors.forename: Marco
authors.initials: M
authors.lastname: van Zwetselaar
authors.forename: Hugh
authors.initials: H
authors.lastname: Reyburn
authors.affiliation: Kilimanjaro Christian Medical University College and Kilimanjaro Clinical Research Institute, Moshi, Tanzania. rekavishe@yahoo.com.
authors.forename: Reginald A
authors.initials: RA
authors.lastname: Kavishe
mesh_terms: D000328:Adult; D000962:Antimalarials / Q000494:pharmacology*; D002648:Child; D003430:Cross-Sectional Studies; D004094:Dihydropteroate Synthase / Q000235:genetics / Q000378:metabolism; D004338:Drug Combinations; D004351:Drug Resistance; D005260:Female; D006801:Humans; D016778:Malaria, Falciparum / Q000188:drug therapy / Q000453:epidemiology / Q000469:parasitology*; D008297:Male; D009154:Mutation; D010963:Plasmodium falciparum / Q000187:drug effects* / Q000235:genetics*; D016133:Polymerase Chain Reaction; D012150:Polymorphism, Restriction Fragment Length; D011247:Pregnancy; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism; D011739:Pyrimethamine / Q000494:pharmacology*; D013413:Sulfadoxine / Q000494:pharmacology*; D013636:Tanzania; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics / Q000378:metabolism
chemical_list: D000962:Antimalarials; D004338:Drug Combinations; D015800:Protozoan Proteins; C001205:fanasil, pyrimethamine drug combination; D013413:Sulfadoxine; D013762:Tetrahydrofolate Dehydrogenase; D004094:Dihydropteroate Synthase; D011739:Pyrimethamine
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