Nonradioactive heteroduplex tracking assay for the detection of minority-variant chloroquine-resistant Plasmodium falciparum in Madagascar Public Deposited

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  • Juliano, Jonathan
    • Affiliation: School of Medicine, Division of Infectious Diseases, Department of Medicine
  • Meshnick, Steven R.
    • Affiliation: Gillings School of Global Public Health, Department of Epidemiology
  • Ramarosandratana, Benjamin
    • Other Affiliation: Service de Lutte contre le Paludisme, Antananarivo, Madagascar
  • Mwapasa, Victor
    • Other Affiliation: Department of Community Health, College of Medicine, University of Malawi, Blantyre, Malawi
  • Ariey, Frédéric
    • Other Affiliation: Institut Pasteur du Cambodge, Phnom Penh, Cambodia
  • Randrianarivelojosia, Milijaona
    • Other Affiliation: Unité de Recherche sur le Paludisme, Institut Pasteur de Madagascar, Antananarivo, Madagascar
  • Abstract Background Strains of Plasmodium falciparum genetically resistant to chloroquine (CQ) due to the presence of pfcrt 76T appear to have been recently introduced to the island of Madagascar. The prevalence of such resistant genotypes is reported to be low (< 3%) when evaluated by conventional PCR. However, these methods are insensitive to low levels of mutant parasites present in patients with polyclonal infections. Thus, the current estimates may be an under representation of the prevalence of the CQ-resistant P. falciparum isolates on the island. Previously, minority variant chloroquine resistant parasites were described in Malawian patients using an isotopic heteroduplex tracking assay (HTA), which can detect pfcrt 76T-bearing P. falciparum minority variants in individual patients that were undetectable by conventional PCR. However, as this assay required a radiolabeled probe, it could not be used in many resource-limited settings. Methods This study describes a digoxigenin (DIG)-labeled chemiluminescent heteroduplex tracking assay (DIG-HTA) to detect pfcrt 76T-bearing minority variant P. falciparum. This assay was compared to restriction fragment length polymorphism (RFLP) analysis and to the isotopic HTA for detection of genetically CQ-resistant parasites in clinical samples. Results Thirty one clinical P. falciparum isolates (15 primary isolates and 16 recurrent isolates) from 17 Malagasy children treated with CQ for uncomplicated malaria were genotyped for the pfcrt K76T mutation. Two (11.7%) of 17 patients harboured genetically CQ-resistant P. falciparum strains after therapy as detected by HTA. RFLP analysis failed to detect any pfcrt K76T-bearing isolates. Conclusion These findings indicate that genetically CQ-resistant P. falciparum are more common than previously thought in Madagascar even though the fitness of the minority variant pfcrt 76T parasites remains unclear. In addition, HTAs for malaria drug resistance alleles are promising tools for the surveillance of anti-malarial resistance. The use of a non-radioactive label allows for the use of HTAs in malaria endemic countries.
Date of publication
  • doi:10.1186/1475-2875-8-47
  • 19291288
Resource type
  • Article
Rights statement
  • In Copyright
Rights holder
  • Jonathan J Juliano et al.; licensee BioMed Central Ltd.
Journal title
  • Malaria Journal
Journal volume
  • 8
Journal issue
  • 1
Page start
  • 47
  • English
Is the article or chapter peer-reviewed?
  • Yes
  • 1475-2875
Bibliographic citation
  • Malaria Journal. 2009 Mar 16;8(1):47
  • Open Access
  • BioMed Central Ltd

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