Multiplication rate variation in the human malaria parasite Plasmodium falciparum

Lee Murray,Mahamadou Diakite,Sarah J Tarr,Lindsay B Stewart,David J Conway,Alfred Amambua-Ngwa,Ambroise D Ahouidi

doi:10.1038/s41598-017-06295-9

Abstract

It is important to understand intrinsic variation in asexual blood stage multiplication rates of the most virulent human malaria parasite, Plasmodium falciparum. Here, multiplication rates of long-term laboratory adapted parasite clones and new clinical isolates were measured, using a newly standardised assay of growth from low starting density in replicate parallel cultures with erythrocytes from multiple different donors, across multiple cycles. Multiplication rates of long-term established clones were between 7.6 and 10.5 fold per 48 hours, with clone Dd2 having a higher rate than others (clones 3D7, HB3 and D10). Parasite clone-specific growth was then analysed in co-culture assays with all possible heterologous pairwise combinations. This showed that co-culture of different parasites did not affect their replication rates, indicating that there were no suppressive interactions operating between parasites. Multiplication rates of eleven new clinical isolates were measured after a few weeks of culture, and showed a spectrum of replication rates between 2.3 and 6.0 fold per 48 hours, the entire range being lower than for the long-term laboratory adapted clones. Multiplication rate estimates remained stable over time for several isolates tested repeatedly up to three months after culture initiation, indicating considerable persistence of this important trait variation.

Highlights

The malaria parasite Plasmodium falciparum is responsible for up to half a million human deaths each year[1], but there are basic features of its biology that remain poorly understood
Parasite multiplication rates have been assessed for only a few laboratory adapted clones of P. falciparum[13], and from genetic cross progeny of two clones[14], but these may not be representative of natural parasite populations
Substantial intrinsic variation in P. falciparum multiplication rates is revealed, with long-term laboratory adapted lines multiplying between 7.6 and 10.5 fold every 48 hours and clinical isolates that have been cultured for only a few weeks multiplying between 2.3 and 6.0 fold every 48 hours

Summary

Introduction

The malaria parasite Plasmodium falciparum is responsible for up to half a million human deaths each year[1], but there are basic features of its biology that remain poorly understood. Studies on rodent malaria models have given contrasting conclusions as to whether competitive suppression occurs between different co-infecting genotypes[19, 20], and co-culture studies on P. falciparum have not yielded consistent findings on interactions between different clones[21,22,23] It remains unknown how malaria parasites would be able to detect the presence of other clones within a multi-clone infection, or within heterologous co-cultures, potential intercellular communication mechanisms are suggested[24, 25]. A reliable assay is here described for quantifying P. falciparum asexual blood stage multiplication rates in a standardised manner over multiple cycles, using biological triplicate measurements in different erythrocytes This is applied to measure the multiplication rate variation among unrelated laboratory clones cultured separately, and in mixed co-cultured combinations, as well as the multiplication rates of a panel of clinical isolates from three endemic countries

Methods

Results

Conclusion