Research Report on Use of Polymerase Chain Reaction (PCR) for the diagnosis of Malaria in Nepal

Abstract

Background: Polymerase chain reaction method has been found to be more sensitive for Plamodium vivax (91%) and Plasmodium falcifarum (89%). Level of parasite detection has been reported to be as low as 1 to 10 parasites per micro litre of blood in patient sample. Diagnosis by polymerase chain reaction has also been reported to be species specific for all four species of human Plasmodium. Hence there is a need to evaluate a non-microscopic diagnostic technique which should be sensitive, specific and may be available in central facilities of the country for evaluation of drug efficacy, drug resistance to study genetic diversity of parasite found in Nepal. Methods: During the study period (January-September 2003) blood samples from 283 suspected malaria patients from Dhanusha and Kanchanpur districts located in central and far western Nepal were collected and further analyzed utilizing conventional Giemsa stained malaria microscopy and Polymerase Chain Reaction based on 18sssr DNA gene. Results: Out of total 283 samples only 66 (23.32%) samples were positive for malarial parasite (Plasmodium falcifarum-26, Plasmodium vivax-40). The prevalence of malaria in Kanchanpur district during the outbreak season 2003 was 23.32%. Plasmodium falcifarum malaria cases were 9.18% and Plasmodium vivax malaria cases were 14.13% of the total collected cases whereas 39.39% and 60.60% of the total positive cases respectively. This rate of prevalence is much higher than the expected national prevalence rate. Comparative evaluation of polymerase chain reaction and Giemsa stained blood smear microscopy method showed that polymerase chain reaction is almost as sensitive as microscopy. Polymerase chain reaction could detect 2 cases of falcifarum malaria which could not be detected by microscopy. Conclusions: Polymerase chain reaction is a useful diagnostic technique for malaria diagnostic research especially in drug resistant and low parasitaemic cases. Keywords: malaria; microscopy; plasmodium falcifarum; plasmodium vivax; polymerase chain reaction.