Prevalence of Pfhrp2/3 gene deletions among false negative rapid antigen test results in central India

Background &objectives: The diagnosis of Plasmodium falciparum malaria is widely dependent on the P. falciparum histidine rich protein 2 (PfHRP2) antigens based rapid diagnostic tests. There are few possible factors like Pfhrp2 polymorphism, Pfhrp2 deletion and density of malaria parasite which can affect the sensitivity of the Pf-HRP2-based RDT. The primary objective of the investigation was to check whether the Pfhrp2 gene deletion is the primary cause of RDT false negative cases.

Febrile patients from three districts of Chhattisgarh, India were screened for malaria during 2016-2017 by microscopy and RDT. All microscopy P. falciparum positive samples were validated by PCR. Microscopy positive and RDT negative samples were analyzed for the presence of Exon 2, across Exon 1-2, upstream and downstream of both the Pfhrp2 and Pfhrp3 genes fragment by PCR.

Out of 203 screened samples, 85 were detected positive for P. falciparum malaria based on microscopy and PCR. Among these 85 P. falciparum positive samples, 4 samples were observed Pf-HRP2 RDT negative. Although, it signified that the RDTs used were reliable with sensitivity of 95.3% (81/85). 3/4 PfHRP2-RDT negative samples of the P. falciparum isolates exhibited complete deletion of Pfhrp2 and Pfhrp3 genes and one sample was found RDT false negative due to high parasite density.

Pfhrp2 and Pfhrp3 deletions that result in false negative RDTs were uncommon in our setting. The continued monitoring of RDTS which results in false negative tests due to Pfhrp2/3 gene deletion is the need of the hour for an effective malaria elimination strategy.