With malaria elimination back on the international agenda, programs face the challenge of targeting all Plasmodium infections, not only symptomatic cases. As asymptomatic individuals are unlikely to ...seek treatment, they are missed by passive surveillance while remaining infectious to mosquitoes, thus acting as silent reservoirs of transmission. To estimate the risk of asymptomatic infections in various phases of malaria elimination, we need a deeper understanding of the underlying mechanisms favoring carriage over disease, which may involve both pathogen and host factors. Here we review our current knowledge on the determinants leading to Plasmodium falciparum symptomless infections. Understanding the host–pathogen interactions that are most likely to affect transitions between malaria disease states could guide the development of tools to tackle asymptomatic carriers in elimination settings.
Over the past 10 years, knowledge of the burden, economic costs, and consequences of malaria in pregnancy has improved, and the prevalence of malaria caused by Plasmodium falciparum has declined ...substantially in some geographical areas. In particular, studies outside of Africa have increased the evidence base of Plasmodium vivax in pregnancy. Rapid diagnostic tests have been poor at detecting malaria in pregnant women, while PCR has shown a high prevalence of low density infection, the clinical importance of which is unknown. Erythrocytes infected with P falciparum that express the surface protein VAR2CSA accumulate in the placenta, and VAR2CSA is an important target of protective immunity. Clinical trials for a VAR2CSA vaccine are ongoing, but sequence variation needs to be carefully studied. Health system and household costs still limit access to prevention and treatment services. Within the context of malaria elimination, pregnant women could be used to monitor malaria transmission. This Series paper summarises recent progress and highlights unresolved issues related to the burden of malaria in pregnancy.
One of the fundamental steps toward malaria control is the use of antimalarial drugs. The success of antimalarial treatment can be affected by the presence of drug-resistant populations of Plasmodium ...falciparum. To assess resistance, we used molecular methods to examine 351 P. falciparum isolates collected from 4 sentinel sites in Mozambique for K13, pfmdr1, pfcrt, and pfdhps polymorphisms and for plasmepsin2 (pfpm2) and pfmdr1 copy numbers. We found multiple copies of pfpm2 in 1.1% of isolates. All isolates carried K13 wild-type alleles (3D7-like), except 4 novel polymorphisms (Leu619Leu, Phe656Ile, Val666Val, Gly690Gly). Prevalence of isolates with pfcrt mutant (K76T) allele was low (2.3%). Prevalence of isolates with pfdhps mutant alleles (A437G and K540E) was >80%, indicating persistence of sulfadoxine/pyrimethamine resistance; however, markers of artemisinin were absent, and markers of piperaquine resistance were low. Piperaquine resistance isolates may spread in Mozambique as dihydroartemisinin/piperaquine drug pressure increases.
Rapid diagnostic tests (RDTs) are today the most widely used method for malaria diagnosis and are recommended, alongside microscopy, for the confirmation of suspected cases before the administration ...of anti-malarial treatment. The diagnostic performance of RDTs, as compared to microscopy or PCR is well described but the actual analytical sensitivity of current best-in-class tests is poorly documented. This value is however a key performance indicator and a benchmark value needed to developed new RDTs of improved sensitivity.
Thirteen RDTs detecting either the Plasmodium falciparum histidine rich protein 2 (HRP2) or the plasmodial lactate dehydrogenase (pLDH) antigens were selected from the best performing RDTs according to the WHO-FIND product testing programme. The analytical sensitivity of these products was evaluated using a range of reference materials including P. falciparum and Plasmodium vivax whole parasite samples as well as recombinant proteins.
The best performing HRP2-based RDTs could detect all P. falciparum cultured samples at concentrations as low as 0.8 ng/mL of HRP2. The limit of detection of the best performing pLDH-based RDT specifically detecting P. vivax was 25 ng/mL of pLDH.
The analytical sensitivity of P. vivax and Pan pLDH-based RDTs appears to vary considerably from product to product, and improvement of the limit-of-detection for P. vivax detecting RDTs is needed to match the performance of HRP2 and Pf pLDH-based RDTs for P. falciparum. Different assays using different reference materials produce different values for antigen concentration in a given specimen, highlighting the need to establish universal reference assays.
Circulating small RNAs, including miRNAs but also isomiRs and other RNA species, have the potential to be used as non-invasive biomarkers for communicable and non-communicable diseases. This study ...aims to characterize and compare small RNA profiles in human biofluids. For this purpose, RNA was extracted from plasma and breast milk samples from 15 healthy postpartum mothers. Small RNA libraries were prepared with the NEBNext® small RNA library preparation kit and sequenced in an Illumina HiSeq2000 platform. miRNAs, isomiRs and clusters of small RNAs were annotated using seqBuster/seqCluster framework in 5 plasma and 10 milk samples that passed the initial quality control. The RNA yield was 81 ng/mL standard deviation (SD): 41 and 3985 ng/mL (SD: 3767) for plasma and breast milk, respectively. Mean number of good quality reads was 4.04 million (M) (40.01% of the reads) in plasma and 12.5M (89.6%) in breast milk. One thousand one hundred eighty two miRNAs, 12,084 isomiRs and 1,053 small RNA clusters that included piwi-interfering RNAs (piRNAs), tRNAs, small nucleolar RNAs (snoRNA) and small nuclear RNAs (snRNAs) were detected. Samples grouped by biofluid, with 308 miRNAs, 1,790 isomiRs and 778 small RNA clusters differentially detected. In summary, plasma and milk showed a different small RNA profile. In both, miRNAs, piRNAs, tRNAs, snRNAs, and snoRNAs were identified, confirming the presence of non-miRNA species in plasma, and describing them for the first time in milk.
Reliable serological tests are required to determine the prevalence of antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and to characterize immunity to the disease in ...order to address key knowledge gaps in the coronavirus disease 2019 (COVID-19) pandemic. Quantitative suspension array technology (qSAT) assays based on the xMAP Luminex platform overcome the limitations of rapid diagnostic tests and enzyme-linked immunosorbent assays (ELISAs) with their higher precision, dynamic range, throughput, miniaturization, cost-efficiency, and multiplexing capacity. We developed three qSAT assays for IgM, IgA, and IgG against a panel of eight SARS-CoV-2 antigens, including spike protein (S), nucleocapsid protein (N), and membrane protein (M) constructs. The assays were optimized to minimize the processing time and maximize the signal-to-noise ratio. We evaluated their performances using 128 prepandemic plasma samples (negative controls) and 104 plasma samples from individuals with SARS-CoV-2 diagnosis (positive controls), of whom 5 were asymptomatic, 51 had mild symptoms, and 48 were hospitalized. Preexisting IgG antibodies recognizing N, M, and S proteins were detected in negative controls, which is suggestive of cross-reactivity to common-cold coronaviruses. The best-performing antibody/antigen signatures had specificities of 100% and sensitivities of 95.78% at ≥14 days and 95.65% at ≥21 days since the onset of symptoms, with areas under the curve (AUCs) of 0.977 and 0.999, respectively. Combining multiple markers as assessed by qSAT assays has the highest efficiency, breadth, and versatility to accurately detect low-level antibody responses for obtaining reliable data on the prevalence of exposure to novel pathogens in a population. Our assays will allow gaining insights into antibody correlates of immunity and their kinetics, required for vaccine development to combat the COVID-19 pandemic.
COVID-19 patients elicit strong responses to the nucleocapsid (N) protein of SARS-CoV-2 but binding antibodies are also detected in prepandemic individuals, indicating potential crossreactivity with ...common cold human coronaviruses (HCoV) and questioning its utility in seroprevalence studies. We investigated the immunogenicity of the full-length and shorter fragments of the SARS-CoV-2 N protein, and the crossreactivity of antibodies with HCoV. We identified a C-terminus region in SARS-CoV2 N of minimal sequence homology with HCoV that was more specific for SARS-CoV-2 and highly immunogenic. IgGs to the full-length SARS-CoV-2 N also recognized N229E N, and IgGs to HKU1 N recognized SARS-CoV-2 N. Crossreactivity with SARS-CoV-2 was stronger for alpha- rather than beta-HCoV despite having less sequence identity, revealing the importance of conformational recognition. Higher preexisting IgG to OC43 N correlated with lower IgG to SARS-CoV-2 N in rRT-PCR negative individuals, reflecting less exposure and indicating a potential protective association. Antibodies to SARS-CoV-2 N were higher in patients with more severe and longer duration of symptoms and in females. IgGs remained stable for at least 3 months, while IgAs and IgMs declined faster. In conclusion, N protein is a primary target of SARS-CoV-2-specific and HCoV crossreactive antibodies, both of which may affect the acquisition of immunity to COVID-19.
The HIV epidemic is concentrated in sub-Saharan Africa. However, limited information exists on its impact on women and infant's health since the introduction of antiretroviral drugs in this region, ...where health resources are often scarce.
The effect of HIV infection on maternal health, birth outcomes and infant health was analysed in two contemporary cohorts of HIV-uninfected and HIV-infected pregnant women from southern Mozambique. Pregnant women attending the first antenatal care visit were followed until one month after delivery. Antiretroviral therapy was administered based on CD4+T cell count and clinical stage. Maternal and neonatal morbidity and mortality, as well as pregnancy outcomes were assessed by mother's HIV status.
A total of 1183 HIV-uninfected and 561 HIV-infected pregnant women were enrolled. HIV-infected women were more likely to have anaemia both at the first antenatal care visit and at delivery than HIV-uninfected women (71.5% versus 54.8% and 49.4% versus 40.6%, respectively, p<0.001). Incidence of hospital admissions during pregnancy was increased among HIV-infected women (RR, 2.04, 95%CI, 1.45; 2.86; p<0.001). At delivery, 21% of HIV-infected women reported being on antiretroviral therapy, and 70% having received antiretroviral drugs for prevention of mother to child transmission of HIV. The risk of stillbirths was doubled in HIV-infected women (RR, 2.16 95%CI 1.17; 3.96, p = 0.013). Foetal anaemia was also increased among infants born to HIV-infected women (10.6% versus 7.3%, p = 0.022). No differences were found in mean birth weight, malaria, prematurity and maternal and neonatal deaths between groups.
HIV infection continues to be associated with significant maternal morbidity and poor neonatal health outcomes. Efforts should urgently be made to identify the barriers that impede improvements on the devastating effects of HIV in African women and their infants.
ClinicalTrials.gov NCT 00811421.
Mozambique has historically been one of the countries with the highest malaria burden in the world. Starting in the 1960s, malaria control efforts were intensified in the southern region of the ...country, especially in Maputo city and Maputo province, to aid regional initiatives aimed to eliminate malaria in South Africa and eSwatini. Despite significant reductions in malaria prevalence, elimination was never achieved. Following the World Health Organization's renewed vision of a malaria-free-world, and considering the achievements from the past, the Mozambican National Malaria Control Programme (NMCP) embarked on the development and implementation of a strategic plan to accelerate from malaria control to malaria elimination in southern Mozambique. An initial partnership, supported by the Bill and Melinda Gates Foundation and the La Caixa Foundation, led to the creation of the Mozambican Alliance Towards the Elimination of Malaria (MALTEM) and the Malaria Technical and Advisory Committee (MTAC) to promote national ownership and partner coordination to work towards the goal of malaria elimination in local and cross-border initiatives. Surveillance systems to generate epidemiological and entomological intelligence to inform the malaria control strategies were strengthened, and an impact and feasibility assessment of various interventions aimed to interrupt malaria transmission were conducted in Magude district (Maputo Province) through the "Magude Project". The primary aim of this project was to generate evidence to inform malaria elimination strategies for southern Mozambique. The goal of malaria elimination in areas of low transmission intensity is now included in the national malaria strategic plan for 2017-22 and the NMCP and its partners have started to work towards this goal while evidence continues to be generated to move the national elimination agenda forward.
Malaria programmes use Plasmodium falciparum histidine-rich protein-2 (PfHRP2) based rapid diagnostic tests (RDTs) for malaria diagnosis. The deletion of this target antigen could potentially lead to ...misdiagnosis, delayed treatment and continuation of active transmission.
Plasmodium falciparum isolates (n = 1162) collected in Southern Mozambique were assessed by RDTs, microscopy and/or 18SrRNA qPCR. pfhrp2 and pfhrp3 deletions were investigated in isolates from individuals who were negative by RDT but positive by microscopy and/or qPCR (n = 69) using gene-specific PCRs, with kelch13 PCR as the parasite DNA control.
Lack of pfhrp2 PCR amplification was observed in one of the 69 isolates subjected to molecular analysis 1.45% (95% CI 0.3-7.8%).
The low prevalence of pfhrp2 deletions suggests that RDTs will detect the vast majority of the P. falciparum infections. Nevertheless, active surveillance for changing deletion frequencies is required.
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