Abstract
BackgroundPlasmodium vivax is the most prevalent human malaria parasite and is likely to increase proportionally as malaria control efforts more rapidly impact the prevalence of Plasmodium falciparum. Despite the prominence of P. vivax as a major human pathogen, vivax malaria qualifies as a neglected and under-studied tropical disease. Significant challenges bringing P. vivax into the laboratory, particularly the capacity for long-term propagation of well-characterized strains, have limited the study of this parasite’s red blood cell (RBC) invasion mechanism, blood-stage development, gene expression, and genetic manipulation.Methods and resultsPatient isolates of P. vivax have been collected and cryopreserved in the rural community of Ampasimpotsy, located in the Tsiroanomandidy Health District of Madagascar. Periodic, monthly overland transport of these cryopreserved isolates to the country’s National Malaria Control Programme laboratory in Antananarivo preceded onward sample transfer to laboratories at Case Western Reserve University, USA. There, the P. vivax isolates have been cultured through propagation in the RBCs of Saimiri boliviensis. For the four patient isolates studied to-date, the median time interval between sample collection and in vitro culture has been 454 days (range 166–961 days). The median time in culture, continually documented by light microscopy, has been 159 days; isolate AMP2014.01 was continuously propagated for 233 days. Further studies show that the P. vivax parasites propagated in Saimiri RBCs retain their ability to invade human RBCs, and can be cryopreserved, thawed and successfully returned to productive in vitro culture.Conclusions/significanceLong-term culture of P. vivax is possible in the RBCs of Saimiri boliviensis. These studies provide an alternative to propagation of P. vivax in live animals that are becoming more restricted. In vitro culture of P. vivax in Saimiri RBCs provides an opening to stabilize patient isolates, which would serve as precious resources to apply new strategies for investigating the molecular and cellular biology of this important malaria parasite.
Highlights
Plasmodium vivax is the most prevalent human malaria parasite and is likely to increase proportion‐ ally as malaria control efforts more rapidly impact the prevalence of Plasmodium falciparum
Patient Ext2838 (AMP2016.01), a 3-year old male presented with a temperature of 36.7 °C, was malaria rapid diagnostic test (RDT) positive and slide-positive for P. vivax (5509 parasites/ μL, 0.11%)
Amino acid sequence comparisons are presented in Additional file 2: Figures S1 and S2, respectively. These results show that the P. vivax Duffy binding protein (PvDBP) and PvAMA-1 haplotype sequences from the cultures on days 86 and 202 (2014.01b and 2014.01c) were identical to those obtained from the patient blood sample (2014.01a)
Summary
Plasmodium vivax is the most prevalent human malaria parasite and is likely to increase proportion‐ ally as malaria control efforts more rapidly impact the prevalence of Plasmodium falciparum. Significant challenges bringing P. vivax into the laboratory, the capacity for long-term propagation of wellcharacterized strains, have limited the study of this parasite’s red blood cell (RBC) invasion mechanism, blood-stage development, gene expression, and genetic manipulation. Rounding out the achievements arising from reliable culture of P. falciparum is the ability to perform genetic manipulation of the parasite to verify genetic underpinnings of the parasite’s complex biology [22, 37,38,39]. Much of this has required development of methods for long-term culture, storage, and rejuvenation of well-characterized parasite strains
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