Abstract
Plasmodium ovale curtisi (Poc) and Plasmodium ovale wallikeri (Pow) have been described as two distinct species, only distinguishable by molecular methods such as PCR. Because of no well-defined endemic area and a variable clinical presentation as higher thrombocytopenia and nausea associated with Pow infection and asymptomatic forms of the pathology with Poc infection, rapid and specific identification of Plasmodium ovale curtisi and Plasmodium ovale wallikeri are needed. The aim of the study was to evaluate a new quantitative real-time PCR coupled with high resolution melting revelation (qPCR-HRM) for identification of both species. Results were compared with a nested-PCR, considered as a gold standard for Pow and Poc distinction. 356 samples including all human Plasmodium species at various parasitaemia were tested. The qPCR-HRM assay allowed Poc and Pow discrimination in 66 samples tested with a limit of detection evaluated at 1 parasite/µL. All these results were concordant with nested-PCR. Cross-reaction was absent with others blood parasites. The qPCR-HRM is a rapid and convenient technique to Poc and Pow distinction.
Highlights
Plasmodium ovale curtisi (Poc) and Plasmodium ovale wallikeri (Pow) have been described as two distinct species, only distinguishable by molecular methods such as PCR
We developed a new qPCR technique with High Resolution melting (HRM) revelation with the aim to reliably differentiate Pow and Poc, using samples from patients with imported malaria received at the French Malaria Reference Center (FNMRC)
Of the 356 samples were included in the study period, 25 were negative for Plasmodium infection and 331 were positive according to qPCR-Taqman results (Supplementary Table S1): 219 were positive for Plasmodium falciparum (Pf), 69 Po, 20 for Plasmodium malariae (Pm), 17 for Plasmodium vivax (Pv). 6 samples present mixed infection: 4P. ovale spp + Pf, 1P. ovale spp + Pm and 1P. ovale spp + Pm
Summary
Plasmodium ovale curtisi (Poc) and Plasmodium ovale wallikeri (Pow) have been described as two distinct species, only distinguishable by molecular methods such as PCR. The qPCR-HRM assay allowed Poc and Pow discrimination in 66 samples tested with a limit of detection evaluated at 1 parasite/μL. All these results were concordant with nested-PCR. Seven cases of Plasmodium ovale spp relapse (defined by malaria disease more than one year after the return from an endemic area) occurred in these patients (46% of the 15 observed relapses)[3]. Malaria symptoms are nonspecific and the observation of thick and thin stained blood smears remained the reference method of diagnosis[4,5] This technique is rapid and unexpensive, but required an experienced microscopist to ensure the quality of the result. (for Pf, Pv, Pm and P. ovale spp) and nested-PCR (for Poc and Pow) as the reference for species determination
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