Plasmodium falciparum phenotypic and genotypic resistance profile during the emergence of Piperaquine resistance in Northeastern Thailand

Abstract

Malaria remains a public health problem in Thailand, especially along its borders where highly mobile populations can contribute to persistent transmission. This study aimed to determine resistant genotypes and phenotypes of 112 Plasmodium falciparum isolates from patients along the Thai-Cambodia border during 2013-2015. The majority of parasites harbored a pfmdr1-Y184F mutation. A single pfmdr1 copy number had CVIET haplotype of amino acids 72-76 of pfcrt and no pfcytb mutations. All isolates had a single pfk13 point mutation (R539T, R539I, or C580Y), and increased % survival in the ring-stage survival assay (except for R539I). Multiple copies of pfpm2 and pfcrt-F145I were detected in 2014 (12.8%) and increased to 30.4% in 2015. Parasites containing either multiple pfpm2 copies with and without pfcrt-F145I or a single pfpm2 copy with pfcrt-F145I exhibited elevated IC₉₀ values of piperaquine. Collectively, the emergence of these resistance patterns in Thailand near Cambodia border mirrored the reports of dihydroartemisinin-piperaquine treatment failures in the adjacent province of Cambodia, Oddar Meanchey, suggesting a migration of parasites across the border. As malaria elimination efforts ramp up in Southeast Asia, host nations militaries and other groups in border regions need to coordinate the proposed interventions.

Figure 1

Prevalence of antimalarial drug resistance mutations in Thailand from 2013–2015. Bars indicate the prevalence of parasites with single and amplified pfpm2 copy number. The line graphs indicate the prevalence of parasites harboring the pfcrt-F145I mutation, amplified pfmdr1 copy number and pfk13-C580Y and pfk13-R539T mutations.

Figure 2

In vitro characterization. (A) RSA and pfk13 mutations. RSA_0-3 h survival rate for standard laboratory-adapted clones (W2 for ART-sensitive control, IPC-4884 and IPC-5202 for ART-resistance control) and culture-adapted clinical isolates. The dashed line represents the 1% survival rate cut-off that differentiates ART-resistance (≥ 1% red-dashed line) from ART-sensitive (< 1%) parasites in RSAs. Median and interquartile ranges are shown. (B) IC₉₀ to PIP in parasites with pfcrt-F145I mutation, with and without pfpm2 amplification. Copy number variations of pfpm2 are shown on the x-axis, and PIP-IC₉₀ is shown on the y-axis (log₁₀ scale).