A Novel Nested Multiplex Polymerase Chain Reaction Assay for Malaria Diagnosis Using the Hydroxymethyl Dihydropterin Pyrophosphokinase-Dihydropteroate Synthase (hppk-dhps) Gene

Am J Trop Med Hyg. 2023 Sep 11:tpmd230130. doi: 10.4269/ajtmh.23-0130. Online ahead of print.ABSTRACTThere are many techniques for malaria diagnosis. Currently, the nested polymerase chain reaction (PCR) method based on a small subunit ribosomal RNA gene (18S rRNA) has been used as a confirmatory method. However, this method is time-consuming, laborious, and costly. Therefore, the objective of this study was to develop nested multiplex PCR for Plasmodium species identification using the dihydropterin pyrophosphokinase-dihydropteroate synthase (hppk-dhps) gene. Genus- and species-specific primers for the hppk-dhps gene were designed. The performance of the novel nested multiplex PCR was compared with 18S rRNA nested PCR. A total of 115 blood samples were used in this study, including 84 infected samples and 31 uninfected samples. Analysis of the blood samples by nested multiplex PCR targeting the hppk-dhps gene identified 81 infected cases. The level of agreement between this novel method and 18S rRNA nested PCR was 97.4%. Further, the novel method successfully detected all human malaria parasites except Plasmodium ovale and detected mixed Plasmodium falciparum/Plasmodium vivax infections. The sensitivity and specificity obtained from this novel method were 96.4% and 100%, respectively. The limit of detection of the hppk-dhps nested multiplex PCR for P. falciparum and P. vivax was 500 parasites/µL and 4 parasites/µL, respectively. The lowest parasite gDNA detected by this me...
Source: The American Journal of Tropical Medicine and Hygiene - Category: Tropical Medicine Authors: Source Type: research