Introduction. Malaria is currently one of the most prevalent parasite-transmitted diseases caused by parasites of the genus Plasmodium. Misidentification of human malaria parasites especially P. knowlesi based on microscopic examination is very common. The objectives of this paper were to accurately identify the incidence of human malaria parasites in the interior division of Sabah, Malaysian Borneo, based on small subunit ribosomal RNA (ssrRNA) and to determine the misidentification rate in human malaria parasites. Methods. Nested PCR was used to detect the presence of human malaria parasites. A total of 243 blood spot samples from patients who had requested for blood film for malaria parasite (BFMP) analyses were used in this study. Results. Nested PCR findings showed that there was no P. malariae infection while the highest prevalent malaria parasite was P. knowlesi, followed by P. vivax, P. falciparum, and mixed infection. Only 69.5% of the 243 samples giving consistent nested PCR and microscopic results. Conclusion. The preliminary findings from molecular detection of malaria showed that P. knowlesi was the most prevalent Plasmodium species in the interior division of Sabah. The findings from this paper may provide a clearer picture on the actual transmission of different Plasmodium species in this region. 1. Introduction Malaria is a tropical disease caused by parasites of the genus Plasmodium. Around 250 million of malaria cases and one million deaths caused by malaria are reported around the world, especially in underdeveloped and remote regions [1]. In most South East Asia countries, malaria remains a serious threat to public health [2]. Furthermore, this disease is the most common vector-borne parasitic disease in remote areas of Malaysia [3]. Four Plasmodium parasites, namely, Plasmodium falciparum, P. vivax, P. malaria, and P. ovale, are well recognized to cause malaria worldwide. Recently, P. knowlesi has been acknowledged as the “fifth human malaria species” following its discovery in humans in Malaysian Borneo [4]. Since then, naturally acquired P. knowlesi infection was found to be widespread in Malaysian Borneo and Pahang in West Malaysia [5]. Routine microscopic examination has been considered as the “gold standard” for the diagnosis of Plasmodium parasite infection due to its simple, rapid, and cost-effective features [6]. Nevertheless, this method is prone to misdiagnosis in the cases of mixed infections and low-level parasitemia [7]. The aims of this study were to accurately identify the malaria parasites in humans and to determine
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