Piperaquine resistant Cambodian Plasmodium falciparum clinical isolates: in vitro genotypic and phenotypic characterization

• Published in: Malaria Journal
• Main Authors: Nonlawat Boonyalai, Brian A. Vesely, Chatchadaporn Thamnurak, Chantida Praditpol, Watcharintorn Fagnark, Kirakarn Kirativanich, Piyaporn Saingam, Chaiyaporn Chaisatit, Paphavee Lertsethtakarn, Panita Gosi, Worachet Kuntawunginn, Pattaraporn Vanachayangkul, Michele D. Spring, Mark M. Fukuda, Chanthap Lon, Philip L. Smith, Norman C. Waters, David L. Saunders, Mariusz Wojnarski
• Format: Article in Journal/Newspaper
• Language: English
• Published: BMC 2020
• Subjects: Drug combination; Exonuclease; Malaria; PfCRT; Piperaquine resistance; Plasmepsin; Arctic medicine. Tropical medicine; RC955-962; Infectious and parasitic diseases; RC109-216; Arctic
• Online Access: https://doi.org/10.1186/s12936-020-03339-w; https://doaj.org/article/d710747a8c344a03b5f36b788ad69daa

Abstract Background High rates of dihydroartemisinin–piperaquine (DHA–PPQ) treatment failures have been documented for uncomplicated Plasmodium falciparum in Cambodia. The genetic markers plasmepsin 2 (pfpm2), exonuclease (pfexo) and chloroquine resistance transporter (pfcrt) genes are associated with PPQ resistance and are used for monitoring the prevalence of drug resistance and guiding malaria drug treatment policy. Methods To examine the relative contribution of each marker to PPQ resistance, in vitro culture and the PPQ survival assay were performed on seventeen P. falciparum isolates from northern Cambodia, and the presence of E415G-Exo and pfcrt mutations (T93S, H97Y, F145I, I218F, M343L, C350R, and G353V) as well as pfpm2 copy number polymorphisms were determined. Parasites were then cloned by limiting dilution and the cloned parasites were tested for drug susceptibility. Isobolographic analysis of several drug combinations for standard clones and newly cloned P. falciparum Cambodian isolates was also determined. Results The characterization of culture-adapted isolates revealed that the presence of novel pfcrt mutations (T93S, H97Y, F145I, and I218F) with E415G-Exo mutation can confer PPQ-resistance, in the absence of pfpm2 amplification. In vitro testing of PPQ resistant parasites demonstrated a bimodal dose–response, the existence of a swollen digestive vacuole phenotype, and an increased susceptibility to quinine, chloroquine, mefloquine and lumefantrine. To further characterize drug sensitivity, parental parasites were cloned in which a clonal line, 14-B5, was identified as sensitive to artemisinin and piperaquine, but resistant to chloroquine. Assessment of the clone against a panel of drug combinations revealed antagonistic activity for six different drug combinations. However, mefloquine-proguanil and atovaquone–proguanil combinations revealed synergistic antimalarial activity. Conclusions Surveillance for PPQ resistance in regions relying on DHA–PPQ as the first-line treatment is dependent on the ...