Multiplex real-time PCR detection of P. falciparum, P. vivax and P. malariae in human blood samples.
Two duplex real-time PCR assays were developed to diagnose three human parasites: Plasmodium falciparum, Plasmodium vivax and Plasmodium malariae. TaqMan duplex real-time PCR was evaluated in 263 blood samples of suspected malaria patients by comparing results against those obtained with microscopy and nested PCR. Compared with nested PCR, duplex real-time PCR assays showed 100% sensitivity and specificity. Duplex real-time PCR detected all mixtures of P. falciparum and P. vivax DNA, except at threshold detection limits for both parasites in which P. vivax was not amplified. Threshold detection limits of real-time PCR were 3.1, 0.3 and 0.8 parasites per microlitre of blood for P. falciparum, P. vivax and P. malariae, respectively. Duplex real-time PCR allows the detection of malarial cases, including mixed species infection, it simplifies analysis and reduces cost. Thus, this protocol may prove invaluable for use in the diagnosis of human infection, trial treatments and epidemiologic studies in which high-throughput analyses are often required.
Veron
Vincent
V
Laboratoire Hospitalo-universitaire de Parasitologie et Mycologie Médicale, Equipe EA3593, UFR de Médecine, Université des Antilles et de la Guyane, Cayenne, French Guiana. vincent.veron@guyane.univ-ag.fr
Simon
Stephane
S
Carme
Bernard
B
D000818:Animals; D016054:DNA, Protozoan / Q000737:chemistry / Q000302:isolation & purification; D003937:Diagnosis, Differential; D006801:Humans; D008288:Malaria / Q000175:diagnosis* / Q000469:parasitology; D018512:Parasitemia / Q000175:diagnosis* / Q000469:parasitology; D010963:Plasmodium falciparum / Q000235:genetics / Q000302:isolation & purification*; D010965:Plasmodium malariae / Q000235:genetics / Q000302:isolation & purification*; D010966:Plasmodium vivax / Q000235:genetics / Q000302:isolation & purification*; D016133:Polymerase Chain Reaction / Q000379:methods* / Q000592:standards; D012680:Sensitivity and Specificity
D016054:DNA, Protozoan