Detection of malaria infection with light microscopy and Nested- PCR methods and phylogenetic Analysis in peripheral blood expansion obtained from Sistan- Baluchestan Province

Introduction and Objectives: Malaria is still one of the most important life-threatening infection disease. According to the WHO reports, there were 215 million cases of malaria in 2015. Malaria cause by the genus Plasmodium include P. falciparum, P. vivax, P. malariae, and P. ovale. The aim of this study was to detect of malaria infection with light microscopy and Nested- PCR methods and phylogenetic Analysis in peripheral blood expansion obtained from Sistan- Baluchestan Province. Materials and Methods: 55 blood smears were collected from patients with suspected to malaria in a 6-year period in Sistan- Baluchestan Province. Diagnosis of plasmodium species was performed by Light microscopy and Nested- PCR methods. In addition, seven Plasmodium-positive samples were used to genetic diversity determination. Results: In total, 55 of 55 (100%) cases were positive by LM and included 3 P. falciparum (3.1%), 48 p. vivax (49.48%) and 4 mixed infection with P. falciparum and P. vivax (4.12%). 94.54% (n=52) were positive with Nested PCR and involved 45 P. vivax (46.39%) and 7 (7.21%) mixed infection with P. falciparum and P. vivax. The results of phylogenetic analysis showed that S25, S26, S61 plu 3, S62, S72 plu 3, S74, and S75 isolates belong to Sistan- Baluchestan province. These isolates located three different subclades. These subclades were named I to III. Isolates S26, S75, S62, were in sub clade I, S25 and S74 isolates located in subclade II. All of subclade I and II were P. vivax. Subclade III involved S61 plu3 and S72 plu3 isolates and were P. falciparum. Conclusion: Nested PCR had high sensitivity and specificity than microscopy method and was considered as a good approach for malaria detection.

Keywords:

Malaria , Light microscopy , Nested PCR , Plasmodium falciparum , Plasmodium vivax ,