Five-year tracking of Plasmodium falciparum allele frequencies in a holoendemic area with indistinct seasonal transitions

Abstract

Background

The renewed malaria eradication efforts require an understanding of the seasonal patterns of frequency of polymorphic variants in order to focus limited funds productively. Although cross-sectional studies in holoendemic areas spanning a single year could be useful in describing parasite genotype status at a given point, such information is inadequate in describing temporal trends in genotype polymorphisms. For Plasmodium falciparum isolates from Kisumu District Hospital, Plasmodium falciparum chloroquine-resistance transporter gene (Pfcrt-K76T) and P. falciparum multidrug resistance gene 1 (PfMDR1-N86Y), were analyzed for polymorphisms and parasitemia changes in the 53 months from March 2008 to August 2012. Observations were compared with prevailing climatic factors, including humidity, rainfall, and temperature.

Methods

Parasitemia (the percentage of infected red blood cells per total red blood cells) was established by microscopy for P. falciparum malaria-positive samples. P. falciparum DNA was extracted from whole blood using a Qiagen DNA Blood Mini Kit. Single nucleotide polymorphism identification at positions Pfcrt-K76T and PfMDR1-N86Y was performed using real-time polymerase chain reaction and/or sequencing. Data on climatic variables were obtained from http://www.tutiempo.net/en/.

Results

A total of 895 field isolates from 2008 (n=169), 2009 (n=161), 2010 (n=216), 2011 (n=223), and 2012 (n=126) showed large variations in monthly frequency of PfMDR1-N86Y and Pfcrt-K76T as the mutant genotypes decreased from 68.4%±15% and 38.1%±13% to 29.8%±18% and 13.3%±9%, respectively. The mean percentage of parasitemia was 2.61%±1.01% (coefficient of variation 115.86%; n=895). There was no correlation between genotype or parasitemia and climatic factors.

Conclusion

This study shows variability in the frequency of Pfcrt-K76T and PfMDR1-N86Y polymorphisms during the study period, bringing into focus the role of cross-sectional studies in describing temporal genotype trends. The lack of correlation between genotypes and climatic changes, especially precipitation, emphasizes the cost of investment in genotype change.

Keywords:

• PfMDR1-N86Y
• Pfcrt-K76T
• polymorphism
• parasitemia

Acknowledgments

We thank Dr R Rashid, KEMRI Center for Clinical Research, Dr Tom Logan, USAMRU-K, and John M Vulule, KEMRI Center for Global Health Research, for supporting this study and giving their permission to publish these data. We also thank all clinical staff at Kisumu East District Hospitals for their assistance. This work was supported by the US Department of Defense, Global Emerging Infections Surveillance and Response System, Silver Spring, MD, USA.

Author contributions

HMA participated in study design, laboratory oversight, manuscript writing, and data analysis and interpretation.

AOA carried out molecular assays, drafting the manuscript or revising it critically for important intellectual content, and final manuscript approval. FLE participated in laboratory oversight, revising manuscript critically for important intellectual content, and final manuscript approval.

DWJ and LI participated in data analysis and molecular assays, manuscript revision, and final approval of the version to be published. ACC performed data analysis, revising the manuscript critically for important intellectual content, and final approval of the version to be published. DO undertook sample processing, manuscript revising, study design, and final approval of manuscript. EAO and CM undertook manuscript revision, acquisition of data, manuscript revising, and final approval of the version to be published. CO performed molecular assays, acquisition of data, manuscript revising, and final approval of version to be published. RY undertook data analysis and molecular assays, manuscript revising, and final approval of version to be published. BA undertook protocol oversight, manuscript writing, conception and design, data analysis and interpretation, and final approval of the version to be published. JDJ and EK participated in protocol oversight, manuscript writing and revision, final approval of version to be published, conception and design of study, and data analysis and interpretation. All authors are in agreement to be accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved.

Disclosure

The authors report no competing interests in this work. The opinions and assertions contained in this work are the private views of the authors and are not to be construed as official or as reflecting the views of the US Department of the Army or the Department of Defense.