[Show abstract][Hide abstract] ABSTRACT: Malaria and undernutrition frequently coexist, especially in pregnant women and young children. Nutrient supplementation of these vulnerable groups might reduce their susceptibility to malaria by improving immunity.
Antibody immunity to antigens expressed by a placental-binding parasite isolate, a non-placental binding parasite isolate, merozoites and schizonts at enrolment (before 20 gestation weeks) and at 36 gestation weeks were measured in 1,009 Malawian pregnant women receiving a daily lipid-based nutrient supplement, multiple micronutrients or iron and folic acid, who were participants in a randomized clinical trial assessing the effects of nutrient supplementation on pregnancy outcomes and child development(registration ID: NCT01239693 ).
Antibodies to placental-binding isolates significantly increased while antibodies to most merozoite antigens declined over pregnancy. Overall, after adjustment for covariates, the type of supplementation did not influence antibody levels at 36 gestation weeks or the rate of change in antibody levels from enrolment to 36 weeks. A negative association between maternal body mass index and opsonizing antibodies to placental-binding antigens (coefficient (95% CI) -1.04 (-1.84, -0.24), was observed. Similarly, women with higher socioeconomic status had significantly lower IgG and opsonizing antibodies to placental-binding antigens. Neither of these associations was significantly influenced by the supplementation type.
In the current cohort nutrient supplementation did not affect anti-malarial antibody responses, but poor and undernourished mothers should be a priority group in future trials.
[Show abstract][Hide abstract] ABSTRACT: Understanding how malaria parasites move between populations is important, particularly given the potential for malaria to be reintroduced into areas where it was previously eliminated. We examine the distribution of malaria genetics across seven sites within the Democratic Republic of Congo (DRC) and two nearby countries, Ghana and Kenya, in order to understand how the relatedness of malaria parasites varies across space, and whether there are barriers to the flow of malaria parasites within the DRC or across borders. Parasite DNA was retrieved from dried blood spots from 7 Demographic and Health Survey sample clusters in the DRC. Malaria genetic characteristics of parasites from Ghana and Kenya were also obtained. For each of 9 geographic sites (7 DRC, 1 Ghana and 1 Kenya), a pair-wise RST statistic was calculated, indicating the genetic distance between malaria parasites found in those locations. Mapping genetics across the spatial extent of the study area indicates a complex genetic landscape, where relatedness between two proximal sites may be relatively high (RST > 0.64) or low (RST < 0.05), and where distal sites also exhibit both high and low genetic similarity. Mantel's tests suggest that malaria genetics differ as geographic distances increase. Principal Coordinate Analysis suggests that genetically related samples are not co-located. Barrier analysis reveals no significant barriers to gene flow between locations. Malaria genetics in the DRC have a complex and fragmented landscape. Limited exchange of genes across space is reflected in greater genetic distance between malaria parasites isolated at greater geographic distances. There is, however, evidence for close genetic ties between distally located sample locations, indicating that movement of malaria parasites and flow of genes is being driven by factors other than distance decay. This research demonstrates the contributions that spatial disease ecology and landscape genetics can make to understanding the evolutionary dynamics of infectious diseases.
[Show abstract][Hide abstract] ABSTRACT: Plasmodium falciparum parasites that are resistant to artemisinins have been detected in Southeast Asia. Resistance is associated with several polymorphisms in the parasite's K13-propeller gene. The molecular epidemiology of these artemisinin-resistant genotypes in African parasite populations is unknown. We developed an assay to quantify rare polymorphisms in parasite populations using a pooled deep-sequencing approach to score allele frequencies, validated it using mixtures of laboratory parasite strains, and then employed it to screen over 1,100 African P. falciparum infections collected since 2002 from 14 sites across sub-Saharan Africa. We found no mutations in African parasite populations that are associated with artemisinin resistance in Southeast Asian parasites. However, we observed 15 coding mutations, including 12 novel mutations, and limited allele sharing between parasite populations, consistent with a large reservoir of naturally-occurring K13-propeller variation. Though polymorphisms associated with artemisinin resistance in P. falciparum in Southeast Asia are not prevalent in sub-Saharan Africa, numerous K13-propeller coding polymorphisms circulate in Africa. Although their distributions do not support a widespread selective sweep for an artemisinin resistant phenotype, the impact of these mutations on artemisinin susceptibility is unknown and will require further characterization. Rapid, scalable molecular surveillance offers a useful adjunct in tracking and containing artemisinin resistance.
The Journal of Infectious Diseases 09/2014; 211(5). DOI:10.1093/infdis/jiu467 · 5.78 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Iron deficiency and malaria have similar global distributions, and frequently co-exist in pregnant women and young children. Where both conditions are prevalent, iron supplementation is complicated by observations that iron deficiency anaemia protects against falciparum malaria, and that iron supplements increase susceptibility to clinically significant malaria, but the mechanisms remain obscure. Here, using an in vitro parasite culture system with erythrocytes from iron-deficient and replete human donors, we demonstrate that Plasmodium falciparum infects iron-deficient erythrocytes less efficiently. In addition, owing to merozoite preference for young erythrocytes, iron supplementation of iron-deficient individuals reverses the protective effects of iron deficiency. Our results provide experimental validation of field observations reporting protective effects of iron deficiency and harmful effects of iron administration on human malaria susceptibility. Because recovery from anaemia requires transient reticulocytosis, our findings imply that in malarious regions iron supplementation should be accompanied by effective measures to prevent falciparum malaria.
[Show abstract][Hide abstract] ABSTRACT: 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.
[Show abstract][Hide abstract] ABSTRACT: Background
Intermittent preventive treatment of malaria in pregnancy (IPTp) with sulphadoxine-pyrimethamine (SP) decreases adverse effects of malaria during pregnancy. Zambia implemented its IPTp-SP programme in 2003. Emergence of SP-resistant Plasmodium falciparum threatens this strategy. The quintuple mutant haplotype (substitutions in N51I, C59R, S108N in dhfr and A437G and K540E in dhps genes), is associated with SP treatment failure in non-pregnant patients with malaria. This study examined efficacy of IPTp-SP and presence of the quintuple mutant among pregnant women in Mansa, Zambia.
In Mansa, an area with high malaria transmission, HIV-negative pregnant women presenting to two antenatal clinics for the 1st dose of IPTp-SP with asymptomatic parasitaemia were enrolled and microscopy for parasitaemia was done weekly for five weeks. Outcomes were parasitological failure and adequate parasitological response (no parasitaemia during follow-up). Polymerase chain reaction assays were employed to distinguish recrudescence from reinfection, and identify molecular markers of SP resistance. Survival analysis included those who had reinfection and incomplete follow-up (missed at least one follow-up).
Of the 109 women included in the study, 58 (53%) completed all follow-up, 34 (31%) had incomplete follow-up, and 17 (16%) were lost to follow-up after day 0. Of those who had complete follow-up, 15 (26%, 95% confidence interval [CI] [16–38]) had parasitological failure. For the 92 women included in the survival analysis, median age was 20 years (interquartile range [IQR] 18–22), median gestational age was 22 weeks (IQR range 20–24), and 57% were primigravid. There was no difference in time to failure in primigravid versus multigravid women. Of the 84 women with complete haplotype data for the aforementioned loci of the dhfr and dhps genes, 53 (63%, 95% CI [50–70]) had quintuple mutants (two with an additional mutation in A581G of dhps). Among women with complete follow-up and quintuple mutants, 22% had parasitological failure versus 0% without (p = 0.44).
While underpowered, this study found 26% failure rates of SP given the moderate prevalence of the quintuple mutant haplotype. Despite the presence of resistance, SP retained some efficacy in clearing parasites in pregnant women, and may remain a viable option for IPTp in Zambia.
[Show abstract][Hide abstract] ABSTRACT: Imported malaria threatens control and elimination efforts in countries that have low rates of transmission. In 2010, an outbreak of Plasmodium falciparum malaria was reported among United Nations peacekeeping soldiers from Guatemala who had recently returned from the Democratic Republic of the Congo (DRC). Epidemiologic evidence suggested that the soldiers were infected in the DRC, but local transmission could not be ruled out in all cases. We used population genetic analyses of neutral microsatellites to determine the outbreak source. Genetic relatedness was compared among parasites found in samples from the soldiers and parasite populations collected in the DRC and Guatemala; parasites identified in the soldiers were more closely related to those from the DRC. A phylogenetic clustering analysis confirms this identification with >99.9% confidence. Thus, results support the hypothesis that the soldiers likely imported malaria from the DRC. This study demonstrates the utility of molecular genotyping in outbreak investigations.
[Show abstract][Hide abstract] ABSTRACT: Malaria parasite infections that are only detectable by molecular methods are highly prevalent and represent a potential transmission reservoir. The methods used to detect these infections are not standardized and their operating characteristics are often unknown. We designed a proficiency panel of Plasmodium spp. in order to compare the accuracy of parasite detection of molecular protocols used by labs in a clinical trial consortium. Ten dried blood spots (DBS) were assembled containing P. falciparum, P. vivax, P. malariae, and P. ovale; DBS contained either a single species or a species mixed with P. falciparum. DBS panels were tested in 9 participating laboratories in masked fashion. Of 90 tests, 68 (75.6%) were correct; there were 20 false-negative results and 2 false-positives. The detection rate was 77.8% (49/63) for P. falciparum, 91.7% (11/12) for P. vivax, 83.3% (10/12) for P. malariae, and 70% (7/10) for P. ovale. Most false-negative P. falciparum results were from samples with estimated ≤ 5 parasites per μL blood. Between labs, accuracy ranged from 100% to 50%. In one lab, the inability to detect species in mixed-species infections prompted a re-design and improvement of the assay. Most PCR-based protocols were able to detect P. falciparum and P. vivax at higher densities but these assays may not reliably detect parasites in samples with low P. falciparum densities. Accordingly, formal quality assurance for PCR should be employed whenever this method is used for diagnosis or surveillance. Such efforts will be important if PCR is to be widely employed to assist malaria elimination efforts.
[Show abstract][Hide abstract] ABSTRACT: In the Democratic Republic of the Congo (DRC), artesunate-amodiaquine is first-line therapy for falciparum malaria; little is known about the prevalence of molecular markers of parasite drug resistance. Across the DRC, we genotyped 166 parasites in Plasmodium falciparum chloroquine resistance transporter (pfcrt) using polymerase chain reaction (PCR) and sequencing. Of these parasites, 73 (44%) parasites were pure wild-type CVMNK, 55 (31%) parasites were chloroquine-resistant CVIET, 35 (21.1%) parasites were mixed CVMNK and CVIET, and 3 parasites were other genotypes. Ninety-two infections (55.4%) harbored the pfcrt K76T substitution that is highly correlated with chloroquine failure. The amodiaquine-resistant SVMNT haplotype was absent. Geographically, pfcrt haplotypes were not clearly clustered. Chloroquine accounted for 19.4% of antimalarial use, and amodiaquine accounted for 15.3% of antimalarial use; there were no associations between drug use and mutant haplotype prevalence. In the DRC, our molecular survey indicates that resistance to chloroquine is substantial but that resistance to amodiaquine is absent. These contrasting findings highlight the need for molecular surveillance of drug resistance to inform malaria control policies.
The American journal of tropical medicine and hygiene 04/2014; 90(6). DOI:10.4269/ajtmh.13-0378 · 2.74 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Multiple red cell variants are known to confer protection from malaria. Here, we review advances in identifying new variants that modulate malaria risk and in defining molecular mechanisms that mediate malaria protection.
New red cell variants, including an innate variant in the red cell's major Ca pump and the acquired state of iron deficiency, have been associated with protection from clinical falciparum malaria. The polymorphisms hemoglobin C (HbC) and hemoglobin S (HbS) - known to protect carriers from severe falciparum malaria - enhance parasite passage to mosquitoes and may promote malaria transmission. At the molecular level, substantial advances have been made in understanding the impact of HbS and HbC upon the interactions between host microRNAs and Plasmodium falciparum protein translation; remodeling of red cell cytoskeletal components and transport of parasite proteins to the red cell surface; and chronic activation of the human innate immune system, which induces tolerance to blood-stage parasites. Several polymorphisms have now been associated with protection from clinical vivax malaria or reduced Plasmodium vivax density, including Southeast Asian ovalocytosis and two common forms of glucose-6-phosphate dehydrogenase deficiency.
Red cell variants that modulate malaria risk can serve as models to identify clinically relevant mechanisms of pathogenesis, and thus define parasite and host targets for next-generation therapies.
Current opinion in hematology 03/2014; 21(3). DOI:10.1097/MOH.0000000000000039 · 4.05 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Pneumocystis jirovecii is a symbiotic respiratory fungus that causes pneumonia (PcP) in immunosuppressed patients. Because P. jirovecii cannot be reliably cultured in vitro, it has proven difficult to study and gaps in our understanding of the organism persist. The release of a draft genome for the organism opens the door for the development of new genotyping approaches for studying its molecular epidemiology and global population structure. We identified and validated 8 putatively neutral microsatellite markers and 1 microsatellite marker linked to the dihydropteroate synthase gene (dhps), the enzymatic target of sulfa drugs used for PcP prevention and treatment. Using these tools, we analyzed P. jirovecii isolates from HIV-infected patients from three geographically distant populations: Uganda, the United States, and Spain. Among the 8 neutral markers, we observed high levels of allelic heterozygosity (average He = 0.586 - 0.842). Consistent with past reports, we observed limited global population structuring, with only Ugandan isolates showing minor differentiation from the other two populations. In Ugandan isolates that harbored mutations in dhps, the microsatellite locus linked to dhps demonstrated a depressed He, consistent with positive directional selection for sulfa-resistance mutations. Using a subset of these microsatellites, analyses of individual and paired samples from infections in San Francisco showed reliable typeability within a single infection and high discriminatory power between infections. These features suggest this novel microsatellite typing approach will be an effective tool for molecular-epidemiological investigations into P. jirovecii population structure, transmission, and drug resistance.
[Show abstract][Hide abstract] ABSTRACT: Anemia is common in women of child-bearing age in the Democratic Republic of the Congo (DRC). As part of the 2007 DRC Demographic and Health Survey (DHS), 4638 women of childbearing age (including 526 pregnant women) were tested for HIV and had the hemoglobin content of their blood recorded. We used the leftover dried blood spots to assess malaria prevalence using PCR assays. The DHS provided extensive information on individuals, as well as the geographic coordinates of household clusters which enabled us to derive several variables that characterize the spatial context of these clusters. Multilevel analyses were conducted to determine individual and contextual risk factors for anemia. Prevalence varied geographically; the odds of anemia were associated with both one's ethnic group and the amount and type of nearby agriculture. The odds were not affected by HIV or malaria status.
Health & Place 08/2013; 24C:54-64. DOI:10.1016/j.healthplace.2013.07.009 · 2.44 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Molecular surveillance for drug-resistant malaria parasites requires reliable, timely, and scalable methods. These data may be efficiently produced by genotyping parasite populations using second-generation sequencing (SGS). We designed and validated a SGS protocol to quantify mutant allele frequencies in the Plasmodium falciparum genes dhfr and dhps in mixed isolates. We applied this new protocol to field isolates from children and compared it to standard genotyping using Sanger sequencing. The SGS protocol accurately quantified dhfr and dhps allele frequencies in a mixture of parasite strains. SGS of DNA that was extracted and then pooled from individual isolates estimated mutant allele frequencies that were closely correlated to those estimated by Sanger sequencing (correlations>0.98). SGS obviated most molecular steps in traditional methods, and is cost-saving for parasite populations larger than 50. This SGS genotyping method efficiently and reproducibly estimates parasite allele frequencies within populations of P. falciparum for molecular epidemiologic studies.
The Journal of Infectious Diseases 08/2013; DOI:10.1093/infdis/jit392 · 5.78 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Plasmodium falciparum malaria kills over 500,000 children every year and has been a scourge of humans for millennia. Owing to the co-evolution of humans and P. falciparum parasites, the human genome is imprinted with polymorphisms that not only confer innate resistance to falciparum malaria, but also cause hemoglobinopathies. These genetic traits-including hemoglobin S (HbS), hemoglobin C (HbC), and α-thalassemia-are the most common monogenic human disorders and can confer remarkable degrees of protection from severe, life-threatening falciparum malaria in African children: the risk is reduced 70% by homozygous HbC and 90% by heterozygous HbS (sickle-cell trait). Importantly, this protection is principally present for severe disease and largely absent for P. falciparum infection, suggesting that these hemoglobinopathies specifically neutralize the parasite's in vivo mechanisms of pathogenesis. These hemoglobin variants thus represent a "natural experiment" to identify the cellular and molecular mechanisms by which P. falciparum produces clinical morbidity, which remain partially obscured due to the complexity of interactions between this parasite and its human host. Multiple lines of evidence support a restriction of parasite growth by various hemoglobinopathies, and recent data suggest this phenomenon may result from host microRNA interference with parasite metabolism. Multiple hemoglobinopathies mitigate the pathogenic potential of parasites by interfering with the export of P. falciparum erythrocyte membrane protein 1 (PfEMP1) to the surface of the host red blood cell. Few studies have investigated their effects upon the activation of the innate and adaptive immune systems, although recent murine studies suggest a role for heme oxygenase-1 in protection. Ultimately, the identification of mechanisms of protection and pathogenesis can inform future therapeutics and preventive measures. Hemoglobinopathies slice the "Gordian knot" of host and parasite interactions to confer malaria protection, and offer a translational model to identify the most critical mechanisms of P. falciparum pathogenesis.
[Show abstract][Hide abstract] ABSTRACT: Pneumocystis jirovecii dihydropteroate synthase (DHPS) gene mutations are well-reported. Although sulfa prophylaxis generally is associated with DHPS mutant infection, whether mutant infection is associated with poorer clinical outcomes is less clear. The differing definitions of sulfa prophylaxis and the different mortality endpoints used in these studies may be one explanation for the conflicting study results. Applying different definitions of prophylaxis, mortality endpoints and DHPS mutant to 301 HIV-infected patients with Pneumocystis pneumonia, we demonstrate that prophylaxis, irrespective of definition, increased the risk of infection with pure mutant (any prophylaxis: AOR 4.00, 95% CI: 1.83-8.76, P < 0.001) but not mixed genotypes (any prophylaxis: AOR 0.78, 95% CI: 0.26-2.36, P = 0.65). However, infection with mutant DHPS, irrespective of definition, was not associated with increased mortality (all-cause or PCP death) at the three time-intervals examined (all P > 0.05). Future studies should standardize key variables associated with DHPS mutant infection as well as examine DHPS mutant subtypes (pure mutant vs. mixed infections) - perhaps even individual DHPS mutant genotypes - so that data can be pooled to better address this issue.
Medical mycology: official publication of the International Society for Human and Animal Mycology 03/2013; 51(6). DOI:10.3109/13693786.2013.770604 · 2.26 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Understanding the spatial clustering of Plasmodium falciparum populations can assist efforts to contain drug-resistant parasites and maintain the efficacy of future drugs. We sequenced single nucleotide polymorphisms (SNPs) in the dihydropteroate synthase gene (dhps) associated with sulfadoxine resistance and 5 microsatellite loci flanking dhps in order to investigate the genetic backgrounds, genetic relatedness, and geographic clustering of falciparum parasites in the Democratic Republic of the Congo (DRC). Resistant haplotypes were clustered into subpopulations: one in the northeast DRC, and the other in the balance of the DRC. Network and clonal lineage analyses of the flanking microsatellites indicate that geographically-distinct mutant dhps haplotypes derive from separate lineages. The DRC is therefore a watershed for haplotypes associated with sulfadoxine resistance. Given the importance of central Africa as a corridor for the spread of antimalarial resistance, the identification of the mechanisms of this transit can inform future policies to contain drug-resistant parasite strains.
[Show abstract][Hide abstract] ABSTRACT: In the context of an Intermittent preventive treatment (IPTp) trial for pregnant women in Malawi, Plasmodium falciparum samples from 85 women at enrolment and 35 women at delivery were genotyped for mutations associated with sulfadoxine–pyrimethamine resistance. The prevalence of the highly resistant haplotype with mutations at codons 51 and 108 of dihydrofolate reductase (dhfr) and codons 437 and 540 of dihydropteroate synthase (dhps) increased from 81% at enrolment to 100% at delivery (P = 0.01). Pregnant women who were smear-positive at enrolment were more likely to have P. falciparum parasitemia at delivery. These results lend support to concerns that IPTp use may lead to increased drug resistance in pregnant women during pregnancy and emphasise the importance of screening pregnant women for malaria parasites in areas with prevalent SP resistance even when they are already on IPTp.
Dans le cadre d'un essai de traitement préventif intermittent (TPI) pour les femmes enceintes au Malawi, des échantillons de P. falciparum prélevés chez 85 femmes à l'inscription et 35 femmes lors de l'accouchement ont été génotypés pour les mutations associées à la résistance au sulfadoxine-pyriméthamine. La prévalence de l'haplotype hautement résistant avec des mutations au niveau des codons 51 et 108 de la dihydrofolate réductase (dhfr) et des codons 437 et 540 de la dihydroptéroate synthase (dhfs) a augmenté de 81% lors de l'inscription à 100% à l'accouchement (P = 0,01). Les femmes enceintes avec des frottis positifs à l'inscription étaient plus susceptibles d'avoir une parasitémie à P. falciparum à l'accouchement. Ces résultats appuient les préoccupations que l'utilisation du TPI peut mener à une résistance accrue aux médicaments chez les femmes pendant la grossesse et mettent l'accent sur l'importance du dépistage des parasites du paludisme chez les femmes enceintes dans les zones avec une résistance répandue au sulfadoxine-pyriméthamine, même quand elles sont déjà sous TPI.
En el contexto de un ensayo de tratamiento preventivo intermitente (IPTp) en mujeres embarazadas en Malawi, se obtuvieron muestras de P. falciparum de 85 mujeres en el momento de ingresar en el ensayo y de 35 mujeres en el momento del parto y se genotiparon para ver mutaciones asociadas con resistencias a la sulfadoxina-pirimetamina. La prevalencia de los haplotipos altamente resistentes, con mutaciones en los codones 51 y 108 de la dihidrofolato reductasa (dhfr) y los codones 437 y 540 de la dihidropteroato sintetasa (dhps), aumentó del 81% en el momento de ingresar en el estudio al 100% en el momento del parto (P = 0.01). Las mujeres embarazadas que tenían una lámina positiva al comienzo del estudio tenían una mayor probabilidad de tener una parasitemia por P. falciparum en el momento del parto. Estos resultados apoyan la preocupación de que el uso del IPTp podría llevar a un aumento en la resistencia a medicamentos en mujeres embarazadas durante el embarazo, y enfatizar la importancia de monitorizar a las mujeres embarazadas para parásitos de malaria en áreas con prevalencia de la resistencia, aún cuando ya están recibiendo IPTp.
Tropical Medicine & International Health 11/2012; 18(2). DOI:10.1111/tmi.12028 · 2.30 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Pneumocystis jirovecii pneumonia (PCP) is an important opportunistic infection in patients infected with HIV, but its burden is incompletely characterized in those areas of sub-Saharan Africa where HIV is prevalent. We explored the prevalence of both PCP in HIV-infected adults admitted with pneumonia to a tertiary-care hospital in Uganda and of putative P. jirovecii drug resistance by mutations in fungal dihydropteroate synthase (dhps) and dihydrofolate reductase (dhfr). In 129 consecutive patients with sputum smears negative for mycobacteria, 5 (3.9%) were diagnosed with PCP by microscopic examination of Giemsa-stained bronchoalveolar lavage fluid. Concordance was 100% between Giemsa stain and PCR (dhps and dhfr). PCP was more prevalent in patients newly-diagnosed with HIV (11.4%) than in patients with known HIV (1.1%; p = 0.007). Mortality at 2 months after discharge was 29% overall: 28% among PCP-negative patients, and 60% (3 of 5) among PCP-positive patients. In these 5 fungal isolates and an additional 8 from consecutive cases of PCP, all strains harbored mutant dhps haplotypes; all 13 isolates harbored the P57S mutation in dhps, and 3 (23%) also harbored the T55A mutation. No non-synonymous dhfr mutations were detected. PCP is an important cause of pneumonia in patients newly-diagnosed with HIV in Uganda, is associated with high mortality, and putative molecular evidence of drug resistance is prevalent. Given the reliability of field diagnosis in our cohort, future studies in sub-Saharan Africa can investigate the clinical impact of these genotypes.
PLoS ONE 11/2012; 7(11):e49991. DOI:10.1371/journal.pone.0049991 · 3.23 Impact Factor