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

Background Rapid diagnostic tests (RDTs) used to diagnose Plasmodium falciparum (Pf) 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.

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).

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.

In the detection of Pf 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 of RDTs contributes to our understanding of their performance and interpretation. Competing Interest Statement The authors have declared no competing interest. Funding Statement The project was funded by the Bill and Melinda Gates Foundation (A128488 and INV1135840). The funders contributed but did not lead the study design, and had no role in data collection and analysis, decision to publish, or preparation of the manuscript. Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: The study protocol was approved by the Research Unit of the MOHSS of Namibia (Ref: NH), the University of Namibia Research Ethics Committee (MRC/382/2018) and the University of California, San Francisco Committee for Human Research (17 24178). I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes Data Availability After publication, data collected from this study are available upon request to the corresponding author. Available data include de-identified individual participant data and a data dictionary defining each field in the set. Requests to conduct analyses outside the scope of this publication will be reviewed by the principal investigators (MSH and DM) to determine whether a proposed use of the data is scientifically and ethically appropriate and does not conflict with constraints or informed consent limitations identified by the institutions that granted ethical approval for the study. Requests to reanalyse the data presented in this Article will not require such review. Abbreviations - AIC - Akaike information criterion - BIC - Bayesian information criterion - HRP2 - histidine-rich protein 2 - KM - Kaplan-Meier - LDH - lactate dehydrogenase - LOD - limit of detection - MOHSS - Ministry of Health and Social Services - MSAT - mass screening and treatment - Pan-LDH - all-malaria lactate dehydrogenase - Pf - Plasmodium falciparum - PfLDH - Plasmodium falciparum lactate dehydrogenase - pLDH - Plasmodium lactate dehydrogenase - PvLDH - Plasmodium vivax lactate dehydrogenase - RDT - rapid diagnostic test - TTN - time to negativity [in the title] - uRDT - ultrasensitive rapid diagnostic test - WHO - World Health Organization