Estimating malaria antigen dynamics and the time to negativity of next-generation malaria rapid diagnostic tests

Abstract

Background: Rapid diagnostic tests (RDTs) used to diagnose Plasmodium falciparum predominantly target the antigen Histidine Rich Protein 2 (HRP2) exclusively. With the emergence of hrp2/hrp3 gene deletions, RDTs targeting other antigens such as the essential enzyme Lactate Dehydrogenase (LDH) are needed. The dynamics of LDH relative to HRP2 are currently not well described but are needed to inform the use of next-generation (NG-) LDH and HRP2 RDTs that are designed to address hrp2/hrp3 gene deletions.

Methods: A longitudinal cohort study conducted in a low transmission setting in Namibia was leveraged to compare HRP2 and LDH decay rates. Passive and active case detection were used to recruit individuals with positive HRP2-RDT results. Study participants were treated and subsequently followed weekly until they received two consecutive HRP2-RDT negative results. Blood specimens were characterized for antigen concentration and parasite density. Antigen decay rates were calculated and used to estimate time to negativity (TTN) of NG-RDTs: two HRP2 and LDH-based RDTs (Rapigen Pf and a WHO prequalified Pf/Pv RDT) and an LDH-only RDT (Rapigen Pf/Pv).

Results: In 135 participants, the starting geometric mean concentrations for HRP2 and LDH were 899 ng/mL and 344 ng/mL respectively. Both antigens followed a biphasic decay rate, with a faster decay rate in the first phase. For current RDTs with an analytical sensitivity of 1 ng/mL for HRP2 and 5 ng/mL for LDH, TTN was 44 and 4 days, respectively. With a NG-RDT with LDH analytical sensitivity of 0.37 ng/mL, average TTN was 9 days. Multiple levels of analytical sensitivity were also modeled.

Conclusions: In the detection of P. falciparum malaria, LDH versus HRP2-based RDTs had a faster TTN due to a combination of lower accumulated antigen concentrations and faster decay rates, even for more sensitive LDH-based RDTs. Detection of LDH versus HRP2 by RDT is more likely to reflect a new or very recent infection. For NG-RDTs that target both antigens, HRP2 is likely to contribute more to the test signal than LDH in recently treated infections unless the infection has hrp2/hrp3 gene deletions. Antigen decay data combined with analytical sensitivity contributes to understanding RDT performance and interpretation.

Fig. 1

HRP2 and pLDH antigen dynamics posttreatment. HRP2 data reported first in Ntuku et al. [4]. Red points indicate the data from each individual over time, the pink lines show the fitted biphasic decay for each participant, and the solid black lines show the overall fit to all the data. Dashed black lines indicate the uRDT LOD value of 80 pg/mL. Panels A and C present the decay on a log10 scale, and panels B and D show the untransformed data. The y-axis for panels B and D is restricted to amplify the biphasic inflection

Fig. 2

Combined survival curve for different hypothetical limits of detection (pg/mL) for each antigen. pLDH measurement lines are in shades of red, and HRP2 measurement lines are in shades of blue. Dashed lines represent the median TTN for each test line, and 95% Confidence intervals are represented by matching-coloured ribbons

Fig. 3

Survival curves for NxTek, CareStart, WHOPQ-RDT, and Rapigen Malaria Ag RDTs among participants who cleared infection. Dotted lines represent the median TTN for each RDT. The solid blue NxTek and CareStart lines were calculated using primary data from this study reported first in Ntuku et al. [4]. The dashed red, orange, and yellow lines were estimated using RDT benchmarking data published in Golden et al. [16]. 95% confidence intervals are represented by matching-coloured ribbons

Fig. 4

Survival curves for WHOPQ-RDT and Rapigen Malaria Ag Pf RDT among participants who cleared infection. The WHOPQ-RDT is shown in panel A, while the Rapigen Malaria Ag Pf RDT is shown in panel B. Blue lines indicate the combined HRP2/PfLDH trend, while yellow and red lines represent the HRP2 and PfLDH-only lines, respectively. Dashed lines represent the median TTN for each RDT/test line, and 95% Confidence intervals are represented by matching-coloured ribbons