Place of PCR methods in malaria diagnosis

Background: Gold-standard method for malaria diagnosis is microscopic examination of Giemsa stained thick and thin blood smears. This method is cheap and simple but fastidious and requires experienced microscopists. In recent years, molecular biology techniques have been applied with success in the microbiology field because of their great sensitivity and specificity. The aim of this study is the evaluation of Polymerase Chain Reaction (PCR) in the detection of low parasitaemia and mixed infections.
Methods: A total of 191 blood samples were included in the study. They were collected from patients admitted to hospital because of suspicion of malaria infection, and distributed as follows: 105 from Liege (Belgium), 42 from Lubumbashi (Democratic Republic of Congo), and 44 from Cayenne (French Guiana). Two PCR techniques targeting the small sub-unit rRNA gene of Plasmodium were tested in comparison with microscopy. The real-time PCR was specific of Plasmodium sp. and the semi-nested multiplex PCR was able to detect each of the four species.
Results: The real-time PCR sensitivity was 97% and 100% for multiplex PCR. The specificity of both techniques was 96%. Multiplex PCR detected 2 mixed infections that were missed by microscopy. In 4 cases, both PCR techniques permit to detect parasitaemia after treatment while microscopy was already negative. In one case, parasite DNA was detected by PCR one day before the microscopy became positive.
Conclusions: Both PCR techniques presented the same detection limit. The PCR methods had a better sensitivity than microscopy. They detected P. falciparum and P. vivax respectively 7 and 6 days after beginning of treatment. Multiplex PCR allowed species identification and mixed infection determination that could confirm and complete the microscopic examination. Real-time PCR was quicker than nested PCR and could be used for screening in addition to the gold-standard method