Quantification of multiple infections of <it>Plasmodium falciparum in vitro</it>

• Main Authors: Wacker Mark A, Turnbull Lindsey B, Walker Leah A, Mount Michael C, Ferdig Michael T
• Format: Article
• Language: English
• Published: BMC 2012-05-01
• Series: Malaria Journal
• Online Access: http://www.malariajournal.com/content/11/1/180

<p>Abstract</p> <p>Background</p> <p>Human malaria infections caused by the parasite <it>Plasmodium falciparum</it> often contain more than one genetically distinct parasite. Despite this fact, nearly all studies of multiple strain <it>P. falciparum</it> infections have been limited to determining relative densities of each parasite within an infection. In light of this, new methods are needed that can quantify the absolute number of parasites within a single infection.</p> <p>Methods</p> <p>A quantitative PCR (qPCR) method was developed to track the dynamic interaction of <it>P. falciparum</it> infections containing genetically distinct parasite clones in cultured red blood cells. Allele-specific primers were used to generate a standard curve and to quantify the absolute concentration of parasite DNA within multi-clonal infections. Effects on dynamic growth relationships between parasites under drug pressure were examined by treating mixed cultures of drug sensitive and drug resistant parasites with the anti-malarial drug chloroquine at different dosing schedules.</p> <p>Results</p> <p>An absolute quantification method was developed to monitor the dynamics of <it>P. falciparum</it> cultures <it>in vitro.</it> This method allowed for the observation of competitive suppression, the reduction of parasites numbers due to the presence of another parasite, and competitive release, the improved performance of a parasite after the removal of a competitor. These studies demonstrated that the presence of two parasites led to the reduction in density of at least one parasite. The introduction of drug to a mixed culture containing both a drug resistant and drug sensitive parasites resulted in an increased proportion of the drug resistant parasite. Moreover, following drug treatment, the resistant parasite experienced competitive release by exhibiting a fitness benefit greater than simply surviving drug treatment, due to the removal of competitive suppression by the sensitive parasite.</p> <p>Conclusions</p> <p>The newly developed assay allowed for the examination of the dynamics of two distinct clones <it>in vitro</it>; both competitive suppression and release were observed. A deeper understanding of the dynamic growth responses of multiple strain <it>P. falciparum</it> infections, with and without drug pressure, can improve the understanding of the role of parasite interactions in the spread of drug resistant parasites, perhaps suggesting different treatment strategies.</p>