Competition between Plasmodium falciparum strains in clinical infections during in vitro culture adaptation

Abstract

We evaluated the dynamics of parasite populations during in vitro culture adaptation in 15 mixed Plasmodium falciparum infections, which were collected from a hypoendemic area near the China-Myanmar border. Allele types at the msp1 block 2 in the initial clinical samples and during subsequent culture were quantified weekly using a quantitative PCR method. All mixed infections carried two allele types based on the msp1 genotyping result. We also genotyped several polymorphic sites in the dhfr, dhps and mdr1 genes on day 0 and day 28, which showed that most of the common sites analyzed were monomorphic. Two of the three clinical samples mixed at dhps 581 remained stable while one changed to wild-type during the culture. During in vitro culture, we observed a gradual loss of parasite populations with 10 of the 15 mixed infections becoming monoclonal by day 28 based on the msp1 allele type. In most cases, the more abundant msp1 allele types in the clinical blood samples at the beginning of culture became the sole or predominant allele types on day 28. These results suggest that some parasites may have growth advantages and the loss of parasite populations during culture adaptation of mixed infections may lead to biased results when comparing the phenotypes such as drug sensitivity of the culture-adapted parasites.

Attenuation of Plasmodium falciparum in vitro drug resistance phenotype following culture adaptation compared to fresh clinical isolates in Cambodia

“…Differences in pfmdr 1 genotype pre- and post-culture have been described in polyclonal infections [ 24 ]. Loss of parasite sub-populations during culture adaptation were observed in 60–70 % of polyclonal samples collected from both Uganda and the China-Myanmar border [ 5 , 6 ]. Fitness costs to resistant parasite sub-populations during in vitro culture have been postulated to alter parasite genotype [ 5 – 7 ].…”

“…Loss of parasite sub-populations during culture adaptation were observed in 60–70 % of polyclonal samples collected from both Uganda and the China-Myanmar border [ 5 , 6 ]. Fitness costs to resistant parasite sub-populations during in vitro culture have been postulated to alter parasite genotype [ 5 – 7 ]. These alterations may confound in vitro drug susceptibility results for culture-adapted parasites, obscuring potential correlations with the ex vivo assay, particularly in high transmission areas where multiplicity of infection is high [ 2 , 20 , 32 , 33 ].…”

“…This is particularly true for those genes which the parasite may have had to modify to survive in the presence of drug, and which may carry a fitness cost for the parasite. Several studies on parasite population dynamics demonstrated clonal fluctuations and selection of resistant genotypes during malaria treatment [ 2 – 4 ] with evidence of loss of parasite population characteristics during culture adaptation [ 5 – 7 ].…”

From Circulation to Cultivation: Plasmodium In Vivo versus In Vitro

“…(F) In vitro adaptation leads to reduced genetic heterogeneity. For example, two-thirds of polyclonal samples became monoclonal during the initial 4 weeks of adaptation [40].…”

Susceptibility Testing of Medically Important Parasites

“…The former approach requires a long-term culture system to be set up and for the isolate to adapt to the artificial system. The risk here is that culture may select for a certain genotype when an infection is often polygenic (so-called multiplicity of infection), especially in regions of hyperendemicity such as Africa (18). Selection may occur for the fitter clone or genotype, which may not reflect the situation in the patient.…”