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Background    Plasmodium knowlesi  is the most common cause of malaria in Malaysia. However, microscopic diagnosis is inaccurate and rapid diagnostic tests (RDTs) are insufficiently sensitive. PCR is sensitive and specific but not feasible at a district level. Loop-mediated isothermal amplification (LAMP) shows potential with only basic requirements. A commercially available LAMP assay, the Eiken Loopamp™ MALARIA Pan Detection kit, is sensitive for  Plasmodium falciparum  and  Plasmodium vivax , but has not previously been evaluated for  P. knowlesi . This study aims to determine the sensitivity of this LAMP assay for detecting  P. knowlesi  infection.    Methods  Study participants included 73 uncomplicated malaria patients with PCR species confirmation: 50  P. knowlesi , 20  P. falciparum  and 3  P. vivax . Nineteen malaria-negative, non-endemic area controls were also included. The sensitivity of the Eiken Loopamp™ MALARIA Pan Detection kit (Pan LAMP) for detecting each  Plasmodium  species was evaluated. Sensitivity and specificity of the Eiken Loopamp™ MALARIA  Pf  Detection kit ( Pf  LAMP) for  P. falciparum  were also determined. The limit of detection for each LAMP assay was evaluated, with results compared to PCR. All  P. knowlesi  patients were also tested by CareStart™ (Pf/VOM) and OptiMAL-IT™ (Pan/Pf) RDTs.    Results  The sensitivity of the Pan LAMP assay was 100% for  P. knowlesi  (95% CI 92.9–100),  P. falciparum  (95% CI 83.2–100), and  P. vivax  (95% CI 29.2–100). The  Pf  LAMP was 100% sensitive and specific for  P. falciparum  detection, with all  P. knowlesi  samples having a negative reaction. LAMP sensitivity was superior to both RDTs, with only 10 and 28% of  P. knowlesi  samples testing positive to CareStart™ and OptiMAL-IT™, respectively. Limit of detection using the Pan LAMP for both  P. knowlesi  and  P. vivax  was 2 parasites/μL, comparable to PCR. For  P. falciparum  both the Pan LAMP and  Pf  LAMP demonstrated a limit of detection of 20 parasites/μL.    Conclusions  The Eiken Loopamp™ MALARIA Pan Detection kit is sensitive for detection of  P. knowlesi  in low parasitaemia clinical infections, as well as  P. falciparum  and  P. vivax.  However, a  P. knowlesi -specific field assay in a simpler format would assist correct species identification and initiation of optimal treatment for all malaria patients.

malaria journal

Detection of Plasmodium knowlesi, Plasmodium falciparum and Plasmodium vivax using loop-mediated isothermal amplification (LAMP) in a co-endemic area in Malaysia