Miriam K. Laufer

Background: Gametocyte carriage varies across Plasmodium falciparum infections. The extent to which this variation reflects differences in the initiation of gametocytogenesis and/or gametocyte maturation remains unclear. We investigated determinants of initiation of gametocytogenesis and gametocyte carriage in P. falciparum infections from Malawi. Methods: A total of 1108 P. falciparum infections from a cohort study conducted in Malawi between 2019 and 2020 underwent ccp4 and pfmget qRT-PCR to measure gametocyte carriage, ap2-g and sbp1 RT-PCR to quantify gametocytogenesis initiation in ring-stage parasites, and pfs230 DNA amplicon sequencing to estimate infection complexity. Mixed-effects logistic regression was used to evaluate predictors of gametocytogenesis initiation and gametocyte carriage, and mixed-effects linear regression was used to evaluate predictors of ap2-g gene transcript abundance and gametocyte density. Results: Gametocyte marker transcripts were detected in 54% (594/1108) of infections and ap2-g transcripts were detected in 41% (429/1047) of infections with detectable ring-stage parasites. Parasite density was positively associated with ap2-g expression, and ap2-g expression was more likely in polyclonal compared to monoclonal infections. Children <6 years old had lower ap2-g transcript abundance than adults but were more likely to carry gametocytes and have higher gametocyte density. Asymptomatic infections showed higher ap2-g transcript abundance and greater gametocyte prevalence and density than symptomatic infections. Conclusions: Our findings suggest that infection complexity and parasite density may play a role in the initiation of gametocytogenesis, whereas host immunity and duration of infection may play a larger role in gametocyte carriage in P. falciparum infections.

Background Persistent human-to-mosquito parasite transmission hinders malaria control in high burden settings. Understanding the human transmission reservoir can support the design of targeted interventions to reduce transmission. Methods In a year-long cohort study in rural Malawi, we used molecular methods to detect all Plasmodium falciparum (Pf) infections and those containing gametocytes, the parasite stage required for transmission, longitudinally at routine surveillance and sick visits. Using population-level analyses, we determined the demographic, temporal, and spatial clustering of infections containing gametocytes and gametocyte density, which predicts transmission. Results Here we show that gametocytes are not randomly distributed among the population or among individuals with Pf infections; gametocytes are detected in only 23% of the population. Among all participants, school-age children have significantly higher incidence of gametocyte-containing infections and high-density gametocyte infections compared to other groups. The presence of school-age children is a key driver of gametocyte frequencies and densities within households, even after adjusting for Pf infection levels. Based on the total gametocyte abundance in the population, we estimate that clearing infections from asymptomatic school-age children in the rainy season would decrease gametocyte abundance by 67% in the population. Conclusions School-age children are the primary driver of ongoing Pf transmission in Malawi and interventions targeting school-age children are needed to effectively reduce Pf infection risk at a population level.

As populations on antiretroviral therapy (ART) in sub-Saharan Africa are aging, hypertension has become an increasingly important comorbidity among people

Background: We evaluated the relationship between maternal HIV and birth outcomes in pregnant women. Setting: Primary health care facilities in Malawi. Methods: In this prospective cohort study, pregnant women attending their first antenatal care visit between 20 and 36 weeks gestation were categorized by HIV status. Women living with HIV were grouped by HIV viral load at antenatal care and delivery (detectable >400 copies/mL), CD4⁺ count at delivery (low 3), and antiretroviral therapy (ART) regimen (tenofovir- and efavirenz-based ART). We evaluated low birth weight (LBW, + count at delivery was associated with LBW. HIV was not significantly associated with small for gestational age. Adverse birth outcomes did not differ by ART regimen. Conclusions: Maternal HIV infection is a risk factor for adverse birth outcomes, and the effect is partially mitigated by viral suppression.

While sulfadoxine-pyrimethamine has been the primary drug in intermittent preventive treatment in pregnancy, dihydroartemisinin-piperaquine (DP) is being considered as an alternative. DP may lead to lower antimalarial antibodies in the mother, resulting in higher risk of malaria in infancy. We probed cord blood sera collected from women enrolled in a clinical trial of sulfadoxine-pyrimethamine vs DP on a protein microarray containing diverse Plasmodium falciparum erythrocyte membrane proteins 1 to measure the impact of intermittent preventive treatment in pregnancy on proteins associated with malaria disease susceptibility. These results suggest that effective maternal malaria prevention may not alter the susceptibility of infants to malaria.

Anopheles gambiae , Anopheles coluzzii , and Anopheles arabiensis are three of the most widespread vectors of malaria parasites, with geographical ranges stretching across wide swaths of Africa. Understanding the population structure of these closely related species, including the extent to which populations are connected by gene flow, is essential for understanding how vector control implemented in one location might indirectly affect vector populations in other locations. Here, we assessed the population structure of each species based on a combined data set of publicly available and newly processed whole-genome sequences. The data set included single nucleotide polymorphisms from whole genomes of 2,410 individual mosquitoes sampled from 128 locations across 19 African countries. We found that A. gambiae sampled from several countries in West and Central Africa showed low genetic differentiation from each other according to principal components analysis (PCA) and ADMIXTURE modeling. Using Estimated Effective Migration Surfaces (EEMS), we showed that this low genetic differentiation indicates high effective migration rates for A. gambiae across this region. Outside of this region, we found eight groups of sampling locations from Central, East, and Southern Africa for which A. gambiae showed higher genetic differentiation, and lower effective migration rates, between each other and the West/Central Africa group. These results indicate that the barriers to and corridors for migration between populations of A. gambiae differ across the geographical range of this malaria vector species. Using the same methods, we found higher genetic differentiation and lower migration rates between populations of A. coluzzii in West and Central Africa than for A. gambiae in the same region. In contrast, we found lower genetic differentiation and higher migration rates between populations of A. arabiensis in Tanzania, compared to A. gambiae in the same region. These differences between A. gambiae , A. coluzzii , and A. arabiensis indicate that migration barriers and corridors may vary, even between very closely related species. Overall, our results demonstrate that migration rates vary both within and between species of Anopheles mosquitoes, presumably based on species-specific responses to the ecological or environmental conditions that may impede or facilitate migration, and the geographical patterns of these conditions across the landscape. Together with previous findings, this study provides robust evidence that migration rates between populations of malaria vectors depend on the ecological context, which should be considered when planning surveillance of vector populations, monitoring for insecticide resistance, and evaluating interventions.

Pertussis (whooping cough) is a reemergent, highly contagious respiratory infection of public health concern. Infants prior to initiation of their primary vaccination series are the most vulnerable to severe infection, and even death. Vaccination during pregnancy is an efficacious means of reducing infection in infants. This approach relies on boosting maternal immunity and passive transfer of antibodies to the infant via placenta and breast milk. Similarly, maternal vaccination post-partum can enhance maternal-infant immunity. To support the analysis of pertussis immunity in the context of maternal-infant immunization, we developed a high throughput multiplex assay for simultaneous quantification of serum IgG antibodies against pertussis vaccine antigens: pertussis toxin (PT), filamentous hemagglutinin (FHA), pertactin (PRN), and fimbriae (FIM2/3), and against tetanus (TT) and diphtheria toxoids (DT), using the Meso Scale Discovery (MSD) platform. The assay was qualified, and specificity, sensitivity, accuracy, precision, linearity, and robustness were demonstrated. The assay was subsequently adapted for quantification of IgG and IgA in breast milk. Applied to a serological survey of pregnant women living in the United States and sub-Saharan Africa, this method revealed differences in magnitude and breadth of antibody profile, consistent with history of vaccination. A longitudinal analysis of Tdap responses in women vaccinated post-partum demonstrated a rapid increase in serum IgG that remained elevated for up to 24 months. Likewise, high levels of vaccine-specific IgA and IgG antibodies were present in breast milk, although they exhibited faster decay. This multiplex MSD assay is a reliable and practical tool for quantification of pertussis, tetanus, and diphtheria antibodies in serum and breast milk in serosurveys or vaccine studies. IMPORTANCE: Pertussis (whooping cough) has reemerged in recent years. Vaccination during pregnancy is an effective approach to prevent illness during the first months of life. We developed a multiplex assay for quantification of pertussis, tetanus, and diphtheria serum antibodies using the Meso Scale Discovery (MSD) platform; the method was qualified, and specificity, precision, accuracy, linearity, and limits of quantification were defined. It was also adapted for quantification of antibodies in breast milk. We successfully determined serostatus in women from different regions and with different vaccination histories, as well as responses to Tdap in blood and breast milk post-partum. This is the first description of a multiplex assay for the quantification of pertussis, tetanus, and diphtheria antibodies in breast milk.

We evaluated detectable viral load (VL) in pregnant women established on antiretroviral therapy (ART) for at least 6 months before conception and those self-reported as ART naive at first antenatal care (ANC) at two government clinics in Southern Malawi. We used logistic regression to identify the predictors of detectable viral load (VL), defined as any measure greater than 400 copies/ml. Of 816 women, 67.9% were established on ART and 32.1% self-reported as ART naive. Among women established on ART, 10.8% had detectable VL and 9.9% had VL >1000 copies/ml (WHO criteria for virological failure). In adjusted analysis, among women established on ART, virological failure was associated with younger age (p = .02), "being single/widowed" (p = 0.001) and no previous deliveries (p = .05). One fifth of women who reported to be ART-naive were found to have an undetectable VL at first ANC. None of the demographic factors could significantly differentiate those with high versus low VL in the ART-naive sub-sample. In this cohort, approximately 90% of women who had initiated ART prior to conception had an undetectable VL at first ANC. This demonstrates good success of the ART program but identifies high risk populations that require additional support.

Background: Infants under 6 months of age are often excluded from malaria surveillance and observational studies. The impact of malaria during early infancy on health later in childhood remains unknown. Methods: Infants from two birth cohorts in Malawi were monitored at quarterly intervals and whenever they were ill from birth through 24 months for Plasmodium falciparum infections and clinical malaria. Poisson regression and linear mixed effects models measured the effect of exposure to malaria in infancy on subsequent malaria incidence, weight-for-age z-scores (WAZ), and haemoglobin concentrations after 6 months. Results: Infants with at least one P. falciparum infection during their first 6 months had increased incidence ratio (IRR) of P. falciparum infection (IRR = 1.27, 95% CI, 1.06–1.52) and clinical malaria (IRR = 2.37, 95% CI, 2.02–2.80) compared to infants without infection. Infants with clinical malaria had increased risk of P. falciparum infection incidence between 6 and 24 months (IRR = 1.64, 95% CI, 1.38–1.94) and clinical malaria (IRR = 1.85, 95% CI, 1.48–2.32). Exposure to malaria was associated with lower WAZ over time (p = 0.02) and lower haemoglobin levels than unexposed infants at every time interval (p = 0.02). Conclusions: Infants experiencing malaria infection or clinical malaria are at increased risk of subsequent infection and disease, have poorer growth, and lower haemoglobin concentrations.