False-negative malaria rapid diagnostic tests in Rwanda: impact of Plasmodium falciparum isolates lacking hrp2 and declining malaria transmission.
Rapid diagnostic tests (RDTs) for histidine rich protein 2 (HRP2) are often used to determine whether persons with fever should be treated with anti-malarials. However, Plasmodium falciparum parasites with a deletion of the hrp2 gene yield false-negative RDTs and there are concerns the sensitivity of HRP2-based RDTs may fall when the intensity of transmission decreases.



This observational study enrolled 9226 patients at three health centres in Rwanda from April 2014 to April 2015. It then compared the sensitivity of RDTs based on HRP2 and the Plasmodium lactate dehydrogenase (pLDH) to microscopy (thick smears) for the diagnosis of malaria. PCR was used to determine whether deletions of the histidine-rich central repeat region of the hrp2 gene (exon 2) were associated with false-negative HRP2-based RDTs.



In comparison to microscopy, the sensitivity and specificity of HRP2- and pLDH-based RDTs were 89.5 and 86.2% and 80.2 and 94.3%, respectively. When the results for both RDTs were combined, sensitivity rose to 91.8% and specificity was 85.7%. Additionally, when smear positivity fell from 46 to 3%, the sensitivity of the HRP2-based RDT fell from 88 to 67%. Of 370 samples with false-negative HRP2 RDT results for which PCR was performed, 140 (38%) were identified as P. falciparum by PCR. Of the isolates identified as P. falciparum by PCR, 32 (23%) were negative for the hrp2 gene based on PCR. Of the 32 P. falciparum isolates negative for hrp2 by PCR, 17 (53%) were positive based on the pLDH RDT.



This prospective study of RDT performance coincided with a decline in the intensity of malaria transmission in Kibirizi (fall in slide positivity from 46 to 3%). This decline was associated with a decrease in HRP2 RDT sensitivity (from 88 to 67%). While P. falciparum isolates without the hrp2 gene were an important cause of false-negative HRP2-based RDTs, most were identified by the pLDH-based RDT. Although WHO does not recommend the use of combined HRP2/pLDH testing in sub-Saharan Africa, these results suggest that combination HRP2/pLDH-based RDTs could reduce the impact of false-negative HRP2-based RDTs for detection of symptomatic P. falciparum malaria.
Tulane University Health Science Center, New Orleans, LA, USA. christina.kozycki@gmail.com.
Christina T
0000-0001-5405-4694
CT
Kozycki
Malaria and Other Parasitic Diseases Division, Rwanda Biomedical Centre, Ministry of Health, Kigali, Rwanda.|Maternal and Child Survival Programme, Jhpiego, Kigali, Rwanda.
Noella
N
Umulisa
School of Medicine and Pharmacy, University of Rwanda, Kigali, Rwanda.
Stephen
S
Rulisa
National Reference Laboratory, Rwanda Biomedical Centre, Ministry of Health, Kigali, Rwanda.
Emil I
EI
Mwikarago
National Reference Laboratory, Rwanda Biomedical Centre, Ministry of Health, Kigali, Rwanda.
Jean Pierre
JP
Musabyimana
Malaria and Other Parasitic Diseases Division, Rwanda Biomedical Centre, Ministry of Health, Kigali, Rwanda.
Jean Pierre
JP
Habimana
Malaria and Other Parasitic Diseases Division, Rwanda Biomedical Centre, Ministry of Health, Kigali, Rwanda.|Swiss Tropical and Public Health Institute, Basel, Switzerland.|University of Basel, Basel, Switzerland.|Quality and Equity Healthcare, Kigali, Rwanda.
Corine
C
Karema
Tulane University Health Science Center, New Orleans, LA, USA.
Donald J
DJ
Krogstad
D000293:Adolescent; D000328:Adult; D000953:Antigens, Protozoan / Q000235:genetics*; D002648:Child; D002675:Child, Preschool; D003430:Cross-Sectional Studies; D003955:Diagnostic Tests, Routine / Q000706:statistics & numerical data*; D005188:False Negative Reactions; D006801:Humans; D007223:Infant; D016778:Malaria, Falciparum / Q000175:diagnosis* / Q000635:transmission; D008875:Middle Aged; D010963:Plasmodium falciparum / Q000235:genetics* / Q000302:isolation & purification; D016133:Polymerase Chain Reaction; D011446:Prospective Studies; D015800:Protozoan Proteins / Q000235:genetics*; D012432:Rwanda; D012680:Sensitivity and Specificity; D055815:Young Adult
D000953:Antigens, Protozoan; C514457:HRP-2 antigen, Plasmodium falciparum; D015800:Protozoan Proteins
Histidine rich protein 2 (HRP2); Malaria; Plasmodium falciparum; Plasmodium lactate dehydrogenase (pLDH); Polymerase chain reaction (PCR); Rapid diagnostic test (RDT); Rwanda; Sensitivity