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Abstract Background Malaria diagnosis in many malaria-endemic countries relies mainly on the use of rapid diagnostic tests (RDTs). The majority of commercial RDTs used in Africa detect the Plasmodium falciparum histidine-rich protein 2 (PfHRP2). pfhrp2/3 gene deletions can therefore lead to false-negative RDT results. This study aimed to evaluate the frequency of PCR-confirmed, false-negative P. falciparum RDT results in Monrovia, Liberia. Methods PfHRP2-based RDT (Paracheck Pf®) and microscopy results from 1038 individuals with fever or history of fever (n = 951) and pregnant women at first antenatal care (ANC) visit (n = 87) enrolled in the Saint Joseph’s Catholic Hospital (Monrovia) from March to July 2019 were used to assess the frequency of false-negative RDT results. True–false negatives were confirmed by detecting the presence of P. falciparum DNA by quantitative PCR in samples from individuals with discrepant RDT and microscopy results. Samples that were positive by 18S rRNA qPCR but negative by PfHRP2-RDT were subjected to multiplex qPCR assay for detection of pfhrp2 and pfhrp3. Results One-hundred and eighty-six (19.6%) and 200 (21.0%) of the 951 febrile participants had a P. falciparum-positive result by RDT and microscopy, respectively. Positivity rate increased with age and the reporting of joint pain, chills and shivers, vomiting and weakness, and decreased with the presence of coughs and nausea. The positivity rate at first ANC visit was 5.7% (n = 5) and 8% (n = 7) by RDT and microscopy, respectively. Out of 207 Plasmodium infections detected by microscopy, 22 (11%) were negative by RDT. qPCR confirmed absence of P. falciparum DNA in the 16 RDT-negative but microscopy-positive samples which were available for molecular testing. Among the 14 samples that were positive by qPCR but negative by RDT and microscopy, 3 only amplified pfldh, and among these 3 all were positive for pfhrp2 and pfhrp3. Conclusion There is no qPCR-confirmed evidence of false-negative RDT results due to pfhrp2/pfhrp3 deletions in this study conducted in Monrovia (Liberia). This indicates that these deletions are not expected to affect the performance of PfHRP2-based RDTs for the diagnosis of malaria in Liberia. Nevertheless, active surveillance for the emergence of PfHRP2 deletions is required.

Abstract Background Disruption of malaria control strategies during the West African 2014–2016 Ebola epidemic led to an increase in malaria-attributable mortality. However, recent data on malaria infection in vulnerable groups, such as pregnant women, are lacking in this post-Ebola scenario. This cross-sectional study aimed to assess the prevalence of Plasmodium falciparum infection and of molecular markers of drug resistance among pregnant women attending antenatal care in Monrovia, capital of Liberia. Methods From October 2016 to June 2017, all pregnant women attending their first antenatal care visit at the Saint Joseph’s Catholic Hospital, Monrovia, were invited to participate in the study. In addition to their routine antenatal care tests, capillary blood spotted onto filter papers were collected from all consenting participants to determine presence of P. falciparum by real-time quantitative PCR. Molecular markers of anti-malarial drug resistance were assessed through Sanger sequencing and quantitative PCR in specimens positive for P. falciparum analysis. Results Of the 195 women participants, 24 (12.3%) were P. falciparum-positive by qPCR. Infected women tended to be more commonly primigravidae and younger than uninfected ones. Parasite densities were higher in primigravidae. Fever was more frequently detected among the infected women. No statistically significant association between P. falciparum infection and haemoglobin levels or insecticide-treated net use was found. While high prevalence of genetic polymorphisms associated with chloroquine and amodiaquine resistance were detected, no molecular markers of artemisinin resistance were observed. Conclusion Plasmodium falciparum infections are expected to occur in at least one in every eight women attending first ANC at private clinics in Monrovia and outside the peak of the rainy season. Young primigravidae are at increased risk of P. falciparum infection. Molecular analyses did not provide evidence of resistance to artemisinins among the P. falciparum isolates tested. Further epidemiological studies involving pregnant women are necessary to describe the risk of malaria in this highly susceptible group outside Monrovia, as well as to closely monitor the emergence of resistance to anti-malarials, as recommended by the Liberian National Malaria Control Programme.