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In Vitro Growth Phenotypes of Single Parasite Lineages Cloned from Multiclonal Malaria Isolates

Poster
Closeup of the head of a brown mosquito with a blurred background.

ASTMH 2021 Annual Meeting

Virtual Event

November 18, 2021

Abstract

Measurement of malaria parasite proliferation in cultured erythrocytes is critical for elucidating key determinants of phenotypes, including drug susceptibility, virulence, and fitness. Multiple parasite lineages with different proliferation rates or fitness may coexist within a clinical isolate, resulting in complex growth interactions and variations in phenotype. We measured proliferation rates of three Plasmodium falciparum Cambodian isolates, including IPC_3445 (MRA-1236), IPC_5202 (MRA-1240), IPC_6403 (MRA-1285), and parasite lineages previously cloned from each of these isolates by limiting dilution. Following synchronization, in vitro cultures were maintained over four consecutive asexual parasite life cycles, with parasite sampling at the end of every cycle to estimate parasitemia and growth rate. In parallel with clinical isolates and component parasite lineages, growth rates and relative changes in parasitemia were measured for laboratory parasite lines 3D7 (MRA-102) and DD2 (MRA-150) as controls. We observed significant differences in fold-change in parasitemia (FC), and parasite growth rate (GR) between parasite isolates and clonal lineages that make up each isolate. For example, while isolate MRA-1240 exhibits similar a proliferation rate to one of its constituent lineages, MRA1240-hap1 (GR: 1.03 ± 0.02 vs. 1.02 ± 0.05; FC: 67 ± 6 vs. 62 ± 2; p > 0.05), the other two component lineages (MRA1240-hap2 and MRA1240-hap3) exhibit markedly different growth profiles. We observed that the most abundant parasite haplotype often dominates the growth phenotype, masking the effect of minority haplotypes akin to recent observations from drug susceptibility testing. Our results also show diminished proliferation of isolate MRA-1236 (GR: 0.90 ± 0.02; FC: 39 ± 4) relative to the component lineages MRA1236-hap1 (GR: 1.11 ± 0.02; FC: 101 ± 3) and MRA1236-hap2 (GR: 1.05 ± 0.04; FC: 88 ± 6) suggestive of competitive suppression. All parasite lines are available through BEI Resources and have well-defined in vitro growth phenotypes useful for both research and development of interventions against malaria

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Presenter

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Standwell C. Nkhoma, PhD

Scientist, BEI Resources

Standwell Nkhoma, PhD, is a malaria scientist within the MR4-BEI Resources Program at ATCC. His work at ATCC has largely focused on designing new genome-wide assays for genotyping malaria parasites and developing new approaches to enhance authentication of MR4-BEI Resources reagents. Recent outputs from this work include a recently published manuscript demonstrating the need to clone multiclonal isolates to yield single parasite lineages with well-defined phenotypes and genotypes and a poster demonstrating how complex interactions between parasite lineages within a single isolate affect parasite proliferation rates. Dr. Nkhoma earned his PhD in Molecular Biology and Biochemistry from the University of Liverpool, UK.

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