CHINESE JOURNAL OF PARASITOLOGY AND PARASITIC DISEASES >

2020 , Vol. 38 >Issue 4: 423 - 428

DOI: https://doi.org/10.12140/j.issn.1000-7423.2020.04.004

ORIGINAL ARTICLES

Evaluation of the efficacy of loop-mediated isothermal amplification in detecting Plasmodium falciparum and other species of Plasmodium

  • Shui-mao ZHOU ,
  • Zu-wu TU ,
  • Yan YANG ,
  • Fang CHEN ,
  • Xi-shuai JIA
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  • 1 Wuhan Centers for Disease Prevention and Control, Wuhan 430015, China
    2 Hubei Provincial Center for Disease Control and Prevention, Wuhan 430070, China

Received date: 2020-03-07

  Online published: 2020-09-09

Supported by

Wuhan Municipal Health and Family Planning Commission Medical Research Project(WG17B04);Wuhan Municipal Health and Family Planning Commission Medical Research Project(WG18Q05);Seventh Batch of Young and Middle-aged Medical Backbone Talent Training Projects of Wuhan Municipal Health and Health Commission

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Abstract

Objective To detect the DNA of malaria parasites using the loop-mediated isothermal amplification (LAMP) method and evaluate the efficacy of this method.Methods Blood samples on filter papers were collected from confirmed malaria patients returning from endemic areas of other countries during 2013-2018. According to the 18S rRNA sequences of malaria parasites, LAMP primers for Plasmodium falciparum and other species of Plasmodium were designed using the Primer Explorer V5 software. The LAMP method for detecting malaria parasites was established and its sensitivity, specificity, detection limit and amplification efficiency were assessed and compared with the nested-PCR. The effect of calcein indicator on LAMP reaction was tested.Results The LAMP and nested-PCR were performed separately on 96 blood samples, including 47 with P. falciparum and 49 with other species of Plasmodium. Either method found 93 samples as malaria-positive. The LAMP method produced positive results for all the 47 blood samples with P. falciparum, while having a false negative rate of 2.1% among the samples with other Plasmodium spp.. The LAMP method provided negative results for 10 samples with Leishmania, 8 with Schistosomiasis japonicum, and 38 blood samples from healthy participants, highly consistent with the results of nested-PCR (Kappa value, 0.956). The sensitivity of LAMP for P. falciparum, P. vivax, P. ovale and P. malariae was 25, 3, 4, and 2 parasites/μl blood, respectively. Amplification efficiency test showed that the turbity time (Tt) and peak value of rate (Df) of LAMP for the four malaria species were less than 60 min and above 0.14, respectively, of them the amplification efficacy was particularly high for P. falciparum (Tt, 20.4 min; Df, 0.16). In visualized LAMP assay for the four malaria species, using calcein indicator delayed LAMP reaction Tt about 8-22 min, and lower the Df by about 21%-35%.Conclusion The LAMP method is highly sensitive, specific, of low detection limit and has potentials for application on-site in the field and in primary medical settings.

Key words： Plasmodium; LAMP; Nested-PCR; P. falciparum

Cite this article

Shui-mao ZHOU , Zu-wu TU , Yan YANG , Fang CHEN , Xi-shuai JIA . Evaluation of the efficacy of loop-mediated isothermal amplification in detecting Plasmodium falciparum and other species of Plasmodium[J]. CHINESE JOURNAL OF PARASITOLOGY AND PARASITIC DISEASES, 2020 , 38(4) : 423 -428 . DOI: 10.12140/j.issn.1000-7423.2020.04.004

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