Impact of Sulfadoxine-Pyrimethamine Resistance on Effectiveness of Intermittent Preventive Therapy for Malaria in Pregnancy at Clearing Infections and Preventing Low Birth Weight.

BACKGROUND: Owing to increasing sulfadoxine-pyrimethamine (SP) resistance in sub-Saharan Africa, monitoring the effectiveness of intermittent preventive therapy in pregnancy (IPTp) with SP is crucial. METHODS: Between 2009 and 2013, both the efficacy of IPTp-SP at clearing existing peripheral malaria infections and the effectiveness of IPTp-SP at reducing low birth weight (LBW) were assessed among human immunodeficiency virus-uninfected participants in 8 sites in 6 countries. Sites were classified as high, medium, or low resistance after measuring parasite mutations conferring SP resistance. An individual-level prospective pooled analysis was conducted. RESULTS: Among 1222 parasitemic pregnant women, overall polymerase chain reaction-uncorrected and -corrected failure rates by day 42 were 21.3% and 10.0%, respectively (39.7% and 21.1% in high-resistance areas; 4.9% and 1.1% in low-resistance areas). Median time to recurrence decreased with increasing prevalence of Pfdhps-K540E. Among 6099 women at delivery, IPTp-SP was associated with a 22% reduction in the risk of LBW (prevalence ratio [PR], 0.78; 95% confidence interval [CI], .69-.88; P < .001). This association was not modified by insecticide-treated net use or gravidity, and remained significant in areas with high SP resistance (PR, 0.81; 95% CI, .67-.97; P = .02). CONCLUSIONS: The efficacy of SP to clear peripheral parasites and prevent new infections during pregnancy is compromised in areas with >90% prevalence of Pfdhps-K540E. Nevertheless, in these high-resistance areas, IPTp-SP use remains associated with increases in birth weight and maternal hemoglobin. The effectiveness of IPTp in eastern and southern Africa is threatened by further increases in SP resistance and reinforces the need to evaluate alternative drugs and strategies for the control of malaria in pregnancy.

The A581G Mutation in the Gene Encoding Plasmodium falciparum Dihydropteroate Synthetase Reduces the Effectiveness of Sulfadoxine-Pyrimethamine Preventive Therapy in Malawian Pregnant Women.

BACKGROUND: The A581 G: mutation in the gene encoding Plasmodium falciparum dihydropteroate synthase (dhps), in combination with the quintuple mutant involving mutations in both dhps and the gene encoding dihydrofolate reductase (dhfr), the so-called sextuple mutant, has been associated with increased placental inflammation and decreased infant birth weight among women receiving intermittent preventive treatment with sulfadoxine-pyrimethamine (IPTp-SP) during pregnancy. METHODS: Between 2009 and 2011, delivering women without human immunodeficiency virus infection were enrolled in an observational study of IPTp-SP effectiveness in Malawi. Parasites were detected by polymerase chain reaction (PCR); positive samples were sequenced to genotype the dhfr and dhps loci. The presence of K540 E: in dhps was used as a marker for the quintuple mutant. RESULTS: Samples from 1809 women were analyzed by PCR; 220 (12%) were positive for P. falciparum. A total of 202 specimens were genotyped at codon 581 of dhps; 17 (8.4%) harbored the sextuple mutant. The sextuple mutant was associated with higher risks of patent infection in peripheral blood (adjusted prevalence ratio [aPR], 2.76; 95% confidence interval [CI], 1.82-4.18) and placental blood (aPR 3.28; 95% CI, 1.88-5.78) and higher parasite densities. Recent SP use was not associated with increased parasite densities or placental pathology overall and among women with parasites carrying dhps A581 G: . CONCLUSIONS: IPTp-SP failed to inhibit parasite growth but did not exacerbate pathology among women infected with sextuple-mutant parasites. New interventions to prevent malaria during pregnancy are needed urgently.

Insight into the pathogenesis of fetal growth restriction in placental malaria: decreased placental glucose transporter isoform 1 expression.

Placental malaria, especially when complicated with intervillositis, can cause fetal growth restriction. Transplacental glucose transport by glucose transporter isoform 1 (GLUT-1) on the syncytiotrophoblast microvillous and basal plasma membranes regulates fetal growth. We found that GLUT-1 expression in the microvillous plasma membrane of Plasmodium falciparum-negative placenta biopsy specimens was comparable to that in P. falciparum-positive placenta biopsy specimens with or without intervillositis, whereas GLUT-1 expression in the basal plasma membrane was lowest in P. falciparum-positive placenta biopsy specimens with intervillositis, compared with the other 2 specimen types (P ≤ .0016). GLUT-1 expression in the basal plasma membrane also correlated negatively with monocyte infiltrate density (r = -0.43; P = .003) and positively with birth weight (r = 0.28; P = .06). These findings suggest that intervillositis, more than placental malaria per se, might cause fetal growth restriction, through impaired transplacental glucose transport.

Independent lineages of highly sulfadoxine-resistant Plasmodium falciparum haplotypes, eastern Africa.

Sulfadoxine-resistant Plasmodium falciparum undermines malaria prevention with sulfadoxine/pyrimethamine. Parasites with a highly resistant mutant dihydropteroate synthase (dhps) haplotype have recently emerged in eastern Africa; they negated preventive benefits of sulfadoxine/pyrimethamine, and might exacerbate placental malaria. We explored emerging lineages of dhps mutant haplotypes in Malawi, the Democratic Republic of the Congo, and Tanzania by using analyses of genetic microsatellites flanking the dhps locus. In Malawi, a triple-mutant dhps SGEG (mutant amino acids are underlined) haplotype emerged in 2010 that was closely related to pre-existing double-mutant SGEA haplotypes, suggesting local origination in Malawi. When we compared mutant strains with parasites from the Democratic Republic of the Congo and Tanzania by multiple independent analyses, we found that SGEG parasites were partitioned into separate lineages by country. These findings support a model of local origination of SGEG dhps haplotypes, rather than geographic diffusion, and have implications for investigations of emergence and effects of parasite drug resistance.

The effects of malaria and intermittent preventive treatment during pregnancy on fetal anemia in Malawi.

BACKGROUND: Fetal anemia is common in malarious areas and is a risk factor for infant morbidity and mortality. Malaria during pregnancy may cause decreased cord hemoglobin (Hb) and fetal anemia among newborns. Intermittent preventive treatment during pregnancy (IPTp) with sulfadoxine-pyrimethamine (SP) is protective against malaria but may also affect hematopoiesis and contribute to fetal anemia. METHODS: Peripheral, placental, and cord blood were examined for malaria parasitemia and Hb concentration in a cross-section of 3848 mothers and infants delivered at Queen Elizabeth Central Hospital in Blantyre, Malawi between 1997 and 2006. Unconditional linear and logistic regressions were performed with multiple imputation for missing covariates to assess the associations between malaria, IPTp with SP, and fetal anemia. RESULTS: The overall prevalence of fetal anemia was 7.9% (n = 304). Malaria parasitemia at delivery was associated with an adjusted decrease in cord Hb of -0.24 g/dL (95% confidence interval [CI], -.42 to -.05). The adjusted prevalence odds ratio for the effect of malaria on fetal anemia was 1.41 (95% CI, 1.05-1.90). Primigravidae who did not take IPTp had infants at highest risk for fetal anemia, and density of parasitemia was correlated with the decrease in cord Hb. There was no significant association between SP use and cord Hb or fetal anemia. CONCLUSIONS: Malaria during pregnancy, but not IPTp, decreases cord Hb and is a risk factor for fetal anemia in Malawi. Intermittent preventive treatment during pregnancy with SP may continue to be safe and effective in preventing malaria during pregnancy and fetal anemia despite development of SP resistance.

Antenatal receipt of sulfadoxine-pyrimethamine does not exacerbate pregnancy-associated malaria despite the expansion of drug-resistant Plasmodium falciparum: clinical outcomes from the QuEERPAM study.

BACKGROUND: Antenatal intermittent preventive therapy with 2 doses of sulfadoxine-pyrimethamine (IPTp-SP) is the mainstay of efforts in sub-Saharan Africa to prevent pregnancy-associated malaria (PAM). Recent studies report that drug resistance may cause IPTp-SP to exacerbate PAM morbidity, raising fears that current policies will cause harm as resistance spreads. METHODS: We conducted a serial, cross-sectional analysis of the relationships between IPTp-SP receipt, SP-resistant Plasmodium falciparum, and PAM morbidity in delivering women during a period of 9 years at a single site in Malawi. PAM morbidity was assessed by parasite densities, placental histology, and birth outcomes. RESULTS: The prevalence of parasites with highly SP-resistant haplotypes increased from 17% to 100% (P < .001), and the proportion of women receiving full IPTp (≥2 doses) increased from 25% to 82% (P < .001). Women who received full IPTp with SP had lower peripheral (P = .018) and placental (P < .001) parasite densities than women who received suboptimal IPTp (<2 doses). This effect was not significantly modified by the presence of highly SP-resistant haplotypes. After adjustment for covariates, the receipt of SP in the presence of SP-resistant P. falciparum did not exacerbate any parasitologic, histologic, or clinical measures of PAM morbidity. CONCLUSIONS: In this longitudinal study of malaria at delivery, the receipt of SP as IPTp did not potentiate PAM morbidity despite the increasing prevalence and fixation of SP-resistant P. falciparum haplotypes. Even when there is substantial resistance, SP may be used in modified IPTp regimens as a component of comprehensive antenatal care.

Maternal syphilis infection is associated with increased risk of mother-to-child transmission of HIV in Malawi.

OBJECTIVE: To determine the association between maternal syphilis and HIV mother-to-child transmission (MTCT). DESIGN: Prospective cohort study. METHODS: Pregnant women admitted at Queen Elizabeth Central Hospital (Malawi) in late third trimester were screened for HIV (by HIV rapid tests) and syphilis (by rapid plasma regain test and Treponema pallidum hemagglutination assay). HIV-infected women and their infants received nevirapine, according to the HIVNET 012 protocol. They were followed up at 6 and 12 weeks postpartum. Infant HIV infection was diagnosed by DNA PCR. FINDINGS: Of the 1155 HIV-infected women enrolled, 1147 had syphilis test results, of whom 92 (8.0%) were infected. Only 751 HIV-positive women delivered live singleton infants who were tested for HIV at birth. Of these, 65 (8.7%) were HIV-infected, suggesting in utero (IU) HIV MTCT. Of the 686 infants who were HIV-negative at birth, 507 were successfully followed up. Of these, 89 (17.6%) became HIV-infected, suggesting intrapartum/postpartum (IP/PP) HIV MTCT. Maternal syphilis was associated with IU HIV MTCT, after adjusting for maternal log10 HIV-1 viral load and low birth weight (LBW) [adjusted relative risk (ARR), 2.77; 95% CI, 1.40-5.46]. Furthermore, maternal syphilis was associated with IP/PP HIV MTCT (ARR, 2.74; 95% CI, 1.58-4.74), after adjusting for recent fever, breast infection, LBW and maternal log10 HIV-1 viral load. CONCLUSION: Maternal syphilis is associated with IU and IP/PP HIV MTCT. Screening and early treatment of maternal syphilis during pregnancy may reduce pediatric HIV infections.

The effect of Plasmodium falciparum malaria on peripheral and placental HIV-1 RNA concentrations in pregnant Malawian women.

OBJECTIVE: To investigate the effect of placental Plasmodium falciparum malaria infection on peripheral and/or placental HIV-1 viral load. DESIGN: A cross-sectional study of HIV-infected pregnant women, with and without placental malaria, delivering at Queen Elizabeth Central Hospital in Malawi. METHODS: Peripheral blood samples were collected from consenting women and tested for HIV. HIV-infected women received nevirapine at the onset of labor. At delivery, placental blood and tissue specimens were collected. HIV-1 RNA concentrations were measured in peripheral and placental plasma samples, and malaria infection was determined by placental histopathology. RESULTS: Of the 480 HIV-infected women enrolled, 304 had placental histopathology performed, of whom 74 (24.3%) had placental malaria. Compared with women without placental malaria, those with placental malaria had a 2.5-fold higher geometric mean peripheral HIV-1 RNA concentration (62,359 versus 24 814 copies/ml; P = 0.0007) and a 2.4-fold higher geometric mean placental HIV-1 RNA concentration (11,733 versus 4919 copies/ml; P = 0.008). In multivariate analyses, after adjusting for CD4 cell count and other covariates, placental malaria was associated with a 1.7-fold increase in geometric mean peripheral HIV-1 RNA concentration (47,747 versus 27,317 copies/ml; P = 0.02) and a 2.0-fold increase in geometric mean placental HIV-1 RNA concentration (9670 versus 4874 copies/ml; P = 0.03). CONCLUSION: Placental malaria infection is associated with an increase in peripheral and placental HIV-1 viral load, which might increase the risk of mother-to-child transmission of HIV.

Adaptive evolution and fixation of drug-resistant Plasmodium falciparum genotypes in pregnancy-associated malaria: 9-year results from the QuEERPAM study.

Sulfadoxine-pyrimethamine (SP) has been widely deployed in Africa for malaria control and molecular evidence of parasite drug-resistance is prevalent. However, the temporal effects on the selection of Plasmodium falciparum are not well understood. We conducted a retrospective serial cross-sectional study between 1997 and 2006 to investigate changes in drug-resistant malaria among pregnant women delivering at a single hospital in Blantyre, Malawi. P. falciparum parasites were genotyped for parasite clone multiplicity and drug-resistance mutations, and the strength of selection upon mutant genotypes was quantified. Five mutations in the dihydrofolate reductase and dihydropteroate synthase genes began at moderate frequencies and achieved fixation by 2005; the frequency of the highly-SP-resistant "quintuple mutant" haplotype increased from 19% to 100%. The selective advantage of alleles and haplotypes were quantified with selection coefficients: Selection was positive on all mutant alleles and haplotypes associated with SP resistance, and the relative fitness of the quintuple mutant haplotype was 0.139 (95% C.I. 0.067-0.211), indicating a substantial positive selective advantage. Mutations that confer higher levels of resistance to SP did not emerge. SP-resistant haplotypes were rapidly selected for and fixed in P. falciparum populations infecting pregnant women while SP was widely deployed in Malawi. These results underscore the pressing need for new preventive measures for pregnancy-associated malaria and provide a real-world model of the selection landscape malaria parasites.

Decreasing burden of malaria in pregnancy in Malawian women and its relationship to use of intermittent preventive therapy or bed nets.

BACKGROUND: The World Health Organization recommends insecticidal bednets and intermittent preventive treatment to reduce malaria in pregnancy. Longitudinal data of malaria prevalence and pregnancy outcomes are valuable in gauging the impact of these antimalarial interventions. METHODOLOGY/PRINCIPAL FINDINGS: We recruited 8,131 women delivering in a single Malawian hospital over 9 years. We recorded demographic data, antenatal prescription of intermittent preventive therapy during pregnancy with sulfadoxine-pyrimethamine and bed net use, and examined finger-prick blood for malaria parasites and hemoglobin concentration. In 4,712 women, we examined placental blood for malaria parasites and recorded the infant's birth weight. Peripheral and placental parasitemia prevalence declined from 23.5% to 5.0% and from 25.2% to 6.8% respectively. Smaller declines in prevalence of low birth weight and anemia were observed. Coverage of intermittent preventive treatment and bednets increased. Number of sulfadoxine-pyrimethamine doses received correlated inversely with placental parasitemia (Odds Ratio (95% CI): 0.79 (0.68, 0.91)), maternal anemia (0.81, (0.73, 0.90)) and low birth weight from 1997-2001 (0.63 (0.53, 0.75)), but not from 2002-2006. Bednet use protected from peripheral parasitemia (0.47, (0.37, 0.60)) and placental parasitemia (0.41, (0.31, 0.54)) and low birth weight (0.75 (0.59, 0.95)) but not anemia throughout the study. Compared to women without nets who did not receive 2-dose sulfadoxine-pyrimethamine, women using nets and receiving 2-dose sulfadoxine-pyrimethamine were less likely to have parasitemia or low birth weight babies. Women receiving 2-dose sulfadoxine-pyrimethamine alone had little evidence of protection whereas bednets alone gave intermediate protection. CONCLUSIONS/SIGNIFICANCE: Increased bednet coverage explains changes in parasitemia and birth weight among pregnant women better than sulfadoxine-pyrimethamine use. High bed net coverage, and sulfadoxine-pyrimethamine resistance, may be contributing to its apparent loss of effectiveness.

Mutations associated with sulfadoxine-pyrimethamine and chlorproguanil resistance in Plasmodium falciparum isolates from Blantyre, Malawi.

We conducted a prevalence study of mutations in Plasmodium falciparum that are associated with antifolate resistance in Blantyre, Malawi. The dihydrofolate reductase 164-Leu mutation, which confers resistance to both pyrimethamine and chlorproguanil, was found in 4.7% of the samples. Previously unreported mutations in dihydropteroate synthase were also found.