The Comparison of Real-time-PCR-HRM and Microscopy Methods for Detection of Mixed Plasmodium spp. Infections in Laghman Province, Afghanistan

Background: Laghman province, in the east of Afghanistan, is one of the most malaria endemic regions with prominence of Plasmodium vivax and Plasmodium falciparum. So far, no study has been conducted to investigate the extent of mixed infections in this area. Objective: In this study, we aimed to evaluate the prevalence of mixed infections of malaria in Laghman province by using a new and highly sensitive molecular method (real-time polymerase chain reaction high resolution melting analysis) and compare its results with microscopically confirmed cases of malaria. Methods: In general, 347 infected individuals have been referred to Khalwati laboratory that is located in the center of Laghman province from May to November of 2018. Microscopic analysis was performed on prepared thick and thin blood films under 00 lens with oil immersion. The real- time-PCR-HRM assay was performed using an ABI 7500 Fast Real-time PCR system. Results: In a microscopic examination, out of 347 patients referred to the Center, 267 (76.94%) cases were detected to be Plasmodium vivax, 79 (22.76%) Plasmodium falciparum and 1 (0.28%) case of mixed of two species. However, by using Real-time PCR-HRM technique, 249 (71.75%) were detected Plasmodium vivax, 79 (22.76%) were Plasmodium falciparum and 19 (5.47%) were mixed of two species. Conclusion: Our result indicated that the Real-time PCR-HRM method is more accurate and more reliable than microscopic method in the diagnosis of malaria mixed infections.