Antigen persistence of rapid diagnostic tests in pregnant women in Nanoro, Burkina Faso, and the implications for the diagnosis of malaria in pregnancy

Abstract

Objectives To evaluate persistence of several Plasmodium antigens in pregnant women after treatment and compare diagnostics during treatment follow‐up. Methods Thirty‐two pregnant women (N = 32) with confirmed malaria infection by a histidine‐rich protein 2 (HRP2)‐based rapid diagnostic test (RDT) and microscopy were followed for 28 days after artemisinin‐based combination therapy (ACT). A Plasmodium lactate dehydrogenase (pLDH)‐based RDT and two ELISAs based on the detection of dihydrofolate reductase–thymidylate synthase (DHFR‐TS) and haeme detoxification protein (HDP) were compared with each other and to RT‐PCR at each visit. Results The mean visit number (95% confidence interval) on which the HRP2‐based RDT was still positive after treatment was 3.4 (2.7–4.1) visits with some patients still positive at day 28. This is significantly later than the pLDH‐based RDT [0.84 (0.55–1.1)], microscopy (median 1, range 1–3), DHFR‐TS‐ELISA [1.7 (1.1–2.3)] and RT‐PCR (median 2, range 1–5) (P < 0.05), but not significantly later than HDP‐ELISA [2.1 (1.6–2.7)]. Lower gravidity and higher parasite density at day 0 resulted in significantly longer positive results with most tests (P < 0.05). Conclusions HRP2 can persist up to 28 days after ACT treatment; therefore, this test is not suitable for treatment follow‐up in pregnant women and can generate problems when using this test during intermittent preventive treatment (IPTp). DHFR‐TS is less persistent than HRP2, making it a potentially interesting target for diagnosis.

Evaluation of Antigen Detection Tests, Microscopy, and Polymerase Chain Reaction for Diagnosis of Malaria in Peripheral Blood in Asymptomatic Pregnant Women in Nanoro, Burkina Faso

“…A P. falciparum-specific 18S ribosomal DNA RT-PCR was performed on a CFX96™ real-time PCR detection system (Bio-Rad Laboratories, Hercules, CA) and a FAM-labeled Taqman probe as described. 16,19,20 If there were discordant results in duplicate samples (i.e., one positive result and one negative result), the RT-PCR for that sample was repeated. Results were not considered positive until they gave a signal in at least two of the duplicate samples.…”

“…The ELISA was then performed with α-HDP H16 (5 μg/mL) or α-DHFR-TS D20 (5 μg/mL) as described. 16 Data collection and statistical analysis. Sample size was calculated to be able to determine with 95% confidence if the sensitivity and specificity was 90% ± 5%, resulting in at least 150 positive women and 150 negative women to be recruited.…”

“…15,16 Briefly, thick and thin blood smears were stained with Giemsa and parasites were counted against 200 leukocytes, with parasite negative results based on screening of 100 microscopic fields at 1,000 + magnification. In case of lower parasitemia ( 10 parasites/200 leukocytes), parasites were counted against 500 leukocytes.…”

“…For HRP2 RDT, n = 418; for RT-PCR, n = 417; for microscopy, n = 380; for Plasmodium lactate dehydrogenase (pLDH) RDT, n = 412; and for enzyme-linked immunosorbent assays, n = 347. in sealed bags with desiccant until transport and further processing in the Netherlands. DNA was isolated from the protein saver cards according to the method of Boom and others 18 and as described 16 and kept at −20 C until use. A P. falciparum-specific 18S ribosomal DNA RT-PCR was performed on a CFX96™ real-time PCR detection system (Bio-Rad Laboratories, Hercules, CA) and a FAM-labeled Taqman probe as described.…”

Malaria Diagnostics in Clinical Trials

“…45 Thus, HRP2-based RDTs should not be used for patient follow-up for at least one month after treatment. 46 Lactate dehydrogenase is a less sensitive target, and LDH-specific assays can be designed to detect P. falciparum LDH or non-P. falciparum LDH. Lactate dehydrogenase circulates in the peripheral blood for a shorter time than HRP2.…”