Risk factors and characterization of Plasmodium vivax-associated admissions to pediatric intensive care units in the Brazilian Amazon

Background

Plasmodium vivax is responsible for a significant proportion of malaria cases worldwide and is increasingly reported as a cause of severe disease. The objective of this study was to characterize severe vivax disease among children hospitalized in intensive care units (ICUs) in the Western Brazilian Amazon, and to identify risk factors associated with disease severity.

Methods and Findings

In this retrospective study, clinical records of 34 children, 0–14 years of age hospitalized in the 11 public pediatric and neonatal ICUs of the Manaus area, were reviewed. P. falciparum monoinfection or P. falciparum/P. vivax mixed infection was diagnosed by microscopy in 10 cases, while P. vivax monoinfection was confirmed in the remaining 24 cases. Two of the 24 patients with P. vivax monoinfection died. Respiratory distress, shock and severe anemia were the most frequent complications associated with P. vivax infection. Ninety-one children hospitalized with P. vivax monoinfections but not requiring ICU were consecutively recruited in a tertiary care hospital for infectious diseases to serve as a reference population (comparators). Male sex (p = 0.039), age less than five years (p = 0.028), parasitemia greater than 500/mm³ (p = 0.018), and the presence of any acute (p = 0.023) or chronic (p = 0.017) co-morbidity were independently associated with ICU admission. At least one of the WHO severity criteria for malaria (formerly validated for P. falciparum) was present in 23/24 (95.8%) of the patients admitted to the ICU and in 17/91 (18.7%) of controls, making these criteria a good predictor of ICU admission (p = 0.001). The only investigated criterion not associated with ICU admission was hyperbilirubinemia (p = 0.513)].

Conclusions

Our study points to the importance of P. vivax-associated severe disease in children, causing 72.5% of the malaria admissions to pediatric ICUs. WHO severity criteria demonstrated good sensitivity in predicting severe P. vivax infection in this small case series.     

Plasma Superoxide Dismutase-1 as a Surrogate Marker of Vivax Malaria Severity

Background

Severe outcomes have been described for both Plasmodium falciparum and P. vivax infections. The identification of sensitive and reliable markers of disease severity is fundamental to improving patient care. An intense pro-inflammatory response with oxidative stress and production of reactive oxygen species is present in malaria. Inflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha) and antioxidant agents such as superoxide dismutase-1 (SOD-1) are likely candidate biomarkers for disease severity. Here we tested whether plasma levels of SOD-1 could serve as a biomarker of severe vivax malaria.

Methodology/Principal Findings

Plasma samples were obtained from residents of the Brazilian Amazon with a high risk for P. vivax transmission. Malaria diagnosis was made by both microscopy and nested PCR. A total of 219 individuals were enrolled: non-infected volunteers (n = 90) and individuals with vivax malaria: asymptomatic (n = 60), mild (n = 50) and severe infection (n = 19). SOD-1 was directly associated with parasitaemia, plasma creatinine and alanine amino-transaminase levels, while TNF-alpha correlated only with the later enzyme. The predictive power of SOD-1 and TNF-alpha levels was compared. SOD-1 protein levels were more effective at predicting vivax malaria severity than TNF-alpha. For discrimination of mild infection, elevated SOD-1 levels showed greater sensitivity than TNF-alpha (76% vs. 30% respectively; p<0.0001), with higher specificity (100% vs. 97%; p<0.0001). In predicting severe vivax malaria, SOD-1 levels exhibited higher sensitivity than TNF-alpha (80% vs. 56%, respectively; p<0.0001; likelihood ratio: 7.45 vs. 3.14; p<0.0001). Neither SOD-1 nor TNF-alpha could discriminate P. vivax infections from those caused by P. falciparum.

Conclusion

SOD-1 is a powerful predictor of disease severity in individuals with different clinical presentations of vivax malaria.     

Cytokine Balance in Human Malaria: Does Plasmodium vivax Elicit More Inflammatory Responses than Plasmodium falciparum?

Background

The mechanisms by which humans regulate pro- and anti-inflammatory responses on exposure to different malaria parasites remains unclear. Although Plasmodium vivax usually causes a relatively benign disease, this parasite has been suggested to elicit more host inflammation per parasitized red blood cell than P. falciparum.

Methodology/Principal Findings

We measured plasma concentrations of seven cytokines and two soluble tumor necrosis factor (TNF)-α receptors, and evaluated clinical and laboratory outcomes, in Brazilians with acute uncomplicated infections with P. vivax (n = 85), P. falciparum (n = 30), or both species (n = 12), and in 45 asymptomatic carriers of low-density P. vivax infection. Symptomatic vivax malaria patients, compared to those infected with P. falciparum or both species, had more intense paroxysms, but they had no clear association with a pro-inflammatory imbalance. To the contrary, these patients had higher levels of the regulatory cytokine interleukin (IL)-10, which correlated positively with parasite density, and elevated IL-10/TNF-α, IL-10/interferon (IFN)-γ, IL-10/IL-6 and sTNFRII/TNF-α ratios, compared to falciparum or mixed-species malaria patient groups. Vivax malaria patients had the highest levels of circulating soluble TNF-α receptor sTNFRII. Levels of regulatory cytokines returned to normal values 28 days after P. vivax clearance following chemotherapy. Finally, asymptomatic carriers of low P. vivax parasitemias had substantially lower levels of both inflammatory and regulatory cytokines than did patients with clinical malaria due to either species.

Conclusions

Controlling fast-multiplying P. falciparum blood stages requires a strong inflammatory response to prevent fulminant infections, while reducing inflammation-related tissue damage with early regulatory cytokine responses may be a more cost-effective strategy in infections with the less virulent P. vivax parasite. The early induction of regulatory cytokines may be a critical mechanism protecting vivax malaria patients from severe clinical complications.     

Malaria in the Post-Partum Period; a Prospective Cohort Study

Background

Several studies have shown a prolonged or increased susceptibility to malaria in the post-partum period. A matched cohort study was conducted to evaluate prospectively the susceptibility to malaria of post-partum women in an area where P.falciparum and P.vivax are prevalent.

Methods

In an area of low seasonal malaria transmission on the Thai-Myanmar border pregnant women attending antenatal clinics were matched to a non-pregnant, non-post-partum control and followed up prospectively until 12 weeks after delivery.

Results

Post-partum women (n = 744) experienced significantly less P.falciparum episodes than controls (hazard ratio (HR) 0.39 (95%CI 0.21–0.72) p = 0.003) but significantly more P.vivax (HR 1.34 (1.05–1.72) p = 0.018). The reduced risk of falciparum malaria was accounted for by reduced exposure, whereas a history of P.vivax infection during pregnancy was a strong risk factor for P.vivax in post-partum women (HR 13.98 (9.13–21.41), p<0.001). After controlling for effect modification by history of P.vivax, post-partum women were not more susceptible to P.vivax than controls (HR: 0.33 (0.21–0.51), p<0.001). Genotyping of pre-and post-partum infections (n⊕ = ⊕10) showed that each post-partum P.falciparum was a newly acquired infection.

Conclusions

In this area of low seasonal malaria transmission post-partum women were less likely to develop falciparum malaria but more likely to develop vivax malaria than controls. This was explained by reduced risk of exposure and increased risk of relapse, respectively. There was no evidence for altered susceptibility to malaria in the post-partum period. The treatment of vivax malaria during and immediately after pregnancy needs to be improved.     

Features and prognosis of severe malaria caused by Plasmodium falciparum, Plasmodium vivax and mixed Plasmodium species in Papua New Guinean children

Background

Mortality from severe pediatric falciparum malaria appears low in Oceania but Plasmodium vivax is increasingly recognized as a cause of complications and death. The features and prognosis of mixed Plasmodium species infections are poorly characterized. Detailed prospective studies that include accurate malaria diagnosis and detection of co-morbidities are lacking.

Methods and Findings

We followed 340 Papua New Guinean (PNG) children with PCR-confirmed severe malaria (77.1% P. falciparum, 7.9% P. vivax, 14.7% P. falciparum/vivax) hospitalized over a 3-year period. Bacterial cultures were performed to identify co-incident sepsis. Clinical management was under national guidelines. Of 262 children with severe falciparum malaria, 30.9%, 24.8% and 23.2% had impaired consciousness, severe anemia, and metabolic acidosis/hyperlactatemia, respectively. Two (0.8%) presented with hypoglycemia, seven (2.7%) were discharged with neurologic impairment, and one child died (0.4%). The 27 severe vivax malaria cases presented with similar phenotypic features to the falciparum malaria cases but respiratory distress was five times more common (P = 0.001); one child died (3.7%). The 50 children with P. falciparum/vivax infections shared phenotypic features of mono-species infections, but were more likely to present in deep coma and had the highest mortality (8.0%; P = 0.003 vs falciparum malaria). Overall, bacterial cultures were positive in only two non-fatal cases. 83.6% of the children had alpha-thalassemia trait and seven with coma/impaired consciousness had South Asian ovalocytosis (SAO).

Conclusions

The low mortality from severe falciparum malaria in PNG children may reflect protective genetic factors other than alpha-thalassemia trait/SAO, good nutrition, and/or infrequent co-incident sepsis. Severe vivax malaria had similar features but severe P. falciparum/vivax infections were associated with the most severe phenotype and worst prognosis.     

Ultrasound evidence of early fetal growth restriction after maternal malaria infection

Background

Intermittent preventive treatment (IPT), the main strategy to prevent malaria and reduce anaemia and low birthweight, focuses on the second half of pregnancy. However, intrauterine growth restriction may occur earlier in pregnancy. The aim of this study was to measure the effects of malaria in the first half of pregnancy by comparing the fetal biparietal diameter (BPD) of infected and uninfected women whose pregnancies had been accurately dated by crown rump length (CRL) before 14 weeks of gestation.

Methodology/Principal Findings

In 3,779 women living on the Thai-Myanmar border who delivered a normal singleton live born baby between 2001–10 and who had gestational age estimated by CRL measurement <14 weeks, the observed and expected BPD z-scores (<24 weeks) in pregnancies that were (n = 336) and were not (n = 3,443) complicated by malaria between the two scans were compared. The mean (standard deviation) fetal BPD z-scores in women with Plasmodium (P) falciparum and/or P.vivax malaria infections were significantly lower than in non-infected pregnancies; −0.57 (1.13) versus −0.10 (1.17), p<0.001. Even a single or an asymptomatic malaria episode resulted in a significantly lower z-score. Fetal female sex (p<0.001) and low body mass index (p = 0.01) were also independently associated with a smaller BPD in multivariate analysis.

Conclusions/Significance

Despite early treatment in all positive women, one or more (a)symptomatic P.falciparum or P.vivax malaria infections in the first half of pregnancy result in a smaller than expected mid-trimester fetal head diameter. Strategies to prevent malaria in pregnancy should include early pregnancy.     

Concurrent Helminthic Infection Protects Schoolchildren with Plasmodium vivax from Anemia

Background

Plasmodium vivax is responsible for a significant portion of malaria cases worldwide, especially in Asia and Latin America, where geo-helminthiasis have a high prevalence. Impact of the interaction between vivax malaria and intestinal helminthes has been poorly explored. The objective of this study was to evaluate the influence of intestinal helminthiasis on the concentration of hemoglobin in children with Plasmodium vivax malaria in rural areas in the municipality of Careiro, in the Western Brazilian Amazon.

Methodology/Principal Findings

A cohort study was conducted from April to November 2008, enrolling children from 5 to 14 years old in two rural areas endemic for malaria. A cross-sectional evaluation was performed in April to actively detect cases of malaria and document baseline hemoglobin and nutritional status. Children were followed-up for six months through passive case detection of malaria based on light microscopy. Throughout the follow-up interval, hemoglobin value and stool examination (three samples on alternate days) were performed on children who developed P. vivax malaria. For 54 schoolchildren with a single infection by P. vivax, hemoglobin during the malaria episode was similar to the baseline hemoglobin for children co-infected with Ascaris lumbricoides (n = 18), hookworm (n = 11) and Trichuris trichiura (n = 9). In children without intestinal helminthes, a significant decrease in the hemoglobin during the malarial attack was seen as compared to the baseline concentration. In the survival analysis, no difference was seen in the time (in days) from the baseline cross-sectional to the first malarial infection, between parasitized and non-parasitized children.

Conclusion/Significance

For the first time, a cohort study showed that intestinal helminthes protect against hemoglobin decrease during an acute malarial attack by P. vivax.     

The Plasmodium vivax Merozoite Surface Protein 3β Sequence Reveals Contrasting Parasite Populations in Southern and Northwestern Thailand

Background

Malaria control efforts have a significant impact on the epidemiology and parasite population dynamics. In countries aiming for malaria elimination, malaria transmission may be restricted to limited transmission hot spots, where parasite populations may be isolated from each other and experience different selection forces. Here we aim to examine the Plasmodium vivax population divergence in geographically isolated transmission zones in Thailand.

Methodology

We employed the P. vivax merozoite surface protein 3β (PvMSP3β) as a molecular marker for characterizing P. vivax populations based on the extensive diversity of this gene in Southeast Asian parasite populations. To examine two parasite populations with different transmission levels in Thailand, we obtained 45 P. vivax isolates from Tak Province, northwestern Thailand, where the annual parasite incidence (API) was more than 2%, and 28 isolates from Yala and Narathiwat Provinces, southern Thailand, where the API was less than 0.02%. We sequenced the PvMSP3β gene and examined its genetic diversity and molecular evolution between the parasite populations.

Principal Findings

Of 58 isolates containing single PvMSP3β alleles, 31 sequence types were identified. The overall haplotype diversity was 0.77±0.06 and nucleotide diversity 0.0877±0.0054. The northwestern vivax malaria population exhibited extensive haplotype diversity (HD) of PvMSP3β (HD = 1.0). In contrast, the southern parasite population displayed a single PvMSP3β allele (HD = 0), suggesting a clonal population expansion. This result revealed that the extent of allelic diversity in P. vivax populations in Thailand varies among endemic areas.

Conclusion

Malaria parasite populations in a given region may vary significantly in genetic diversity, which may be the result of control and influenced by the magnitude of malaria transmission intensity. This is an issue that should be taken into account for the implementation of P. vivax control measures such as drug policy and vaccine development.     

Antibody-mediated growth inhibition of Plasmodium falciparum: relationship to age and protection from parasitemia in Kenyan children and adults

Background

Antibodies that impair Plasmodium falciparum merozoite invasion and intraerythrocytic development are one of several mechanisms that mediate naturally acquired immunity to malaria. Attempts to correlate anti-malaria antibodies with risk of infection and morbidity have yielded inconsistent results. Growth inhibition assays (GIA) offer a convenient method to quantify functional antibody activity against blood stage malaria.

Methods

A treatment-time-to-infection study was conducted over 12-weeks in a malaria holoendemic area of Kenya. Plasma collected from healthy individuals (98 children and 99 adults) before artemether-lumefantrine treatment was tested by GIA in three separate laboratories.

Results

Median GIA levels varied with P. falciparum line (D10, 8.8%; 3D7, 34.9%; FVO, 51.4% inhibition). The magnitude of growth inhibition decreased with age in all P. falciparum lines tested with the highest median levels among children <4 years compared to adults (e.g. 3D7, 45.4% vs. 30.0% respectively, p = 0.0003). Time-to-infection measured by weekly blood smears was significantly associated with level of GIA controlling for age. Upper quartile inhibition activity was associated with less risk of infection compared to individuals with lower levels (e.g. 3D7, hazard ratio = 1.535, 95% CI = 1.012–2.329; p = 0.0438). Various GIA methodologies had little effect on measured parasite growth inhibition.

Conclusion

Plasma antibody-mediated growth inhibition of blood stage P. falciparum decreases with age in residents of a malaria holoendemic area. Growth inhibition assay may be a useful surrogate of protection against infection when outcome is controlled for age.     

Pyronaridine-artesunate versus chloroquine in patients with acute Plasmodium vivax malaria: a randomized, double-blind, non-inferiority trial

Background

New antimalarials are needed for P. vivax and P. falciparum malaria. This study compared the efficacy and safety of pyronaridine-artesunate with that of chloroquine for the treatment of uncomplicated P. vivax malaria.

Methods and Findings

This phase III randomized, double-blind, non-inferiority trial included five centers across Cambodia, Thailand, India, and Indonesia. In a double-dummy design, patients (aged >3–≤60 years) with microscopically confirmed P. vivax mono-infection were randomized (1∶1) to receive pyronaridine-artesunate (target dose 7.2∶2.4 mg/kg to 13.8∶4.6 mg/kg) or chloroquine (standard dose) once daily for three days. Each treatment group included 228 randomized patients. Outcomes for the primary endpoint, Day-14 cure rate in the per-protocol population, were 99.5%, (217/218; 95%CI 97.5, 100) with pyronaridine-artesunate and 100% (209/209; 95%CI 98.3, 100) with chloroquine. Pyronaridine was non-inferior to chloroquine: treatment difference −0.5% (95%CI −2.6, 1.4), i.e., the lower limit of the 2-sided 95%CI for the treatment difference was greater than −10%. Pyronaridine-artesunate cure rates were non-inferior to chloroquine for Days 21, 28, 35 and 42. Parasite clearance time was shorter with pyronaridine-artesunate (median 23.0 h) versus chloroquine (32.0 h; p<0.0001), as was fever clearance time (median 15.9 h and 23.8 h, respectively; p = 0.0017). Kaplan-Meier estimates of post-baseline P. falciparum infection incidence until Day 42 were 2.5% with pyronaridine-artesunate, 6.1% with chloroquine (p = 0.048, log-rank test). Post-baseline P. vivax or P. falciparum infection incidence until Day 42 was 6.8% and 12.4%, respectively (p = 0.022, log rank test). There were no deaths. Adverse events occurred in 92/228 (40.4%) patients with pyronaridine-artesunate and 72/228 (31.6%) with chloroquine. Mild and transient increases in hepatic enzymes were observed for pyronaridine-artesunate.

Conclusion

Pyronaridine-artesunate efficacy in acute uncomplicated P. vivax malaria was at least that of chloroquine. As pyronaridine-artesunate is also efficacious against P. falciparum malaria, this combination has potential utility as a global antimalarial drug.

Trial registration

Clinicaltrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00440999","term_id":"NCT00440999"}}NCT00440999     

Plasmodium vivax sub-patent infections after radical treatment are common in Peruvian patients: results of a 1-year prospective cohort study

Background

There is an increasing body of literature reporting treatment failure of the currently recommended radical treatment of Plasmodium vivax infections. As P. vivax is the main malaria species outside the African continent, emerging tolerance to its radical treatment regime could have major consequences in countries like Peru, where 80% of malaria cases are due to P. vivax. Here we describe the results of a 1-year longitudinal follow up of 51 confirmed P. vivax patients living around Iquitos, Peruvian Amazon, and treated according to the Peruvian national guidelines.

Methodology

Each month a blood sample for microscopy and later genotyping was systematically collected. Recent exposure to infection was estimated by detecting antibodies against the P. vivax circumsporozoite protein (CSP) and all PCR confirmed P. vivax infections were genotyped with 16 polymorphic microsatellites.

Results

During a 1-year period, 84 recurrent infections, 22 positive also by microscopy, were identified, with a median survival time to first recurrent infection of 203 days. Most of them (71%) were asymptomatic; in 13 patients the infection persisted undetected by microscopy for several consecutive months. The genotype of mostly recurrent infections differed from that at day 0 while fewer differences were seen between the recurrent infections. The average expected heterozygosity was 0.56. There was strong linkage disequilibrium (I_A^s = 0.29, p<1.10⁻⁴) that remained also when analyzing only the unique haplotypes, suggesting common inbreeding.

Conclusion

In Peru, the P. vivax recurrent infections were common and displayed a high turnover of parasite genotypes compared to day 0. Plasmodium vivax patients, even when treated according to the national guidelines, may still represent an important parasite reservoir that can maintain transmission. Any elimination effort should consider such a hidden reservoir.     

Artemisinin-Naphthoquine versus Artemether-Lumefantrine for Uncomplicated Malaria in Papua New Guinean Children: An Open-Label Randomized Trial

Background

Artemisinin combination therapies (ACTs) with broad efficacy are needed where multiple Plasmodium species are transmitted, especially in children, who bear the brunt of infection in endemic areas. In Papua New Guinea (PNG), artemether-lumefantrine is the first-line treatment for uncomplicated malaria, but it has limited efficacy against P. vivax. Artemisinin-naphthoquine should have greater activity in vivax malaria because the elimination of naphthoquine is slower than that of lumefantrine. In this study, the efficacy, tolerability, and safety of these ACTs were assessed in PNG children aged 0.5–5 y.

Methods and Findings

An open-label, randomized, parallel-group trial of artemether-lumefantrine (six doses over 3 d) and artemisinin-naphthoquine (three daily doses) was conducted between 28 March 2011 and 22 April 2013. Parasitologic outcomes were assessed without knowledge of treatment allocation. Primary endpoints were the 42-d P. falciparum PCR-corrected adequate clinical and parasitologic response (ACPR) and the P. vivax PCR-uncorrected 42-d ACPR. Non-inferiority and superiority designs were used for falciparum and vivax malaria, respectively. Because the artemisinin-naphthoquine regimen involved three doses rather than the manufacturer-specified single dose, the first 188 children underwent detailed safety monitoring. Of 2,542 febrile children screened, 267 were randomized, and 186 with falciparum and 47 with vivax malaria completed the 42-d follow-up. Both ACTs were safe and well tolerated. P. falciparum ACPRs were 97.8% and 100.0% in artemether-lumefantrine and artemisinin-naphthoquine-treated patients, respectively (difference 2.2% [95% CI −3.0% to 8.4%] versus −5.0% non-inferiority margin, p = 0.24), and P. vivax ACPRs were 30.0% and 100.0%, respectively (difference 70.0% [95% CI 40.9%–87.2%], p<0.001). Limitations included the exclusion of 11% of randomized patients with sub-threshold parasitemias on confirmatory microscopy and direct observation of only morning artemether-lumefantrine dosing.

Conclusions

Artemisinin-naphthoquine is non-inferior to artemether-lumefantrine in PNG children with falciparum malaria but has greater efficacy against vivax malaria, findings with implications in similar geo-epidemiologic settings within and beyond Oceania.

Trial registration

Australian New Zealand Clinical Trials Registry ACTRN12610000913077 Please see later in the article for the Editors'' Summary     

A Large Plasmodium vivax Reservoir and Little Population Structure in the South Pacific

Introduction

The importance of Plasmodium vivax in malaria elimination is increasingly being recognized, yet little is known about its population size and population genetic structure in the South Pacific, an area that is the focus of intensified malaria control.

Methods

We have genotyped 13 microsatellite markers in 295 P. vivax isolates from four geographically distinct sites in Papua New Guinea (PNG) and one site from Solomon Islands, representing different transmission intensities.

Results

Diversity was very high with expected heterozygosity values ranging from 0.62 to 0.98 for the different markers. Effective population size was high (12′872 to 19′533 per site). In PNG population structuring was limited with moderate levels of genetic differentiation. F _ST values (adjusted for high diversity of markers) were 0.14–0.15. Slightly higher levels were observed between PNG populations and Solomon Islands (F _ST = 0.16).

Conclusions

Low levels of population structure despite geographical barriers to transmission are in sharp contrast to results from regions of low P. vivax endemicity. Prior to intensification of malaria control programs in the study area, parasite diversity and effective population size remained high.     

Whole Genome Sequencing of Field Isolates Reveals a Common Duplication of the Duffy Binding Protein Gene in Malagasy Plasmodium vivax Strains

Background

Plasmodium vivax is the most prevalent human malaria parasite, causing serious public health problems in malaria-endemic countries. Until recently the Duffy-negative blood group phenotype was considered to confer resistance to vivax malaria for most African ethnicities. We and others have reported that P. vivax strains in African countries from Madagascar to Mauritania display capacity to cause clinical vivax malaria in Duffy-negative people. New insights must now explain Duffy-independent P. vivax invasion of human erythrocytes.

Methods/Principal Findings

Through recent whole genome sequencing we obtained ≥70× coverage of the P. vivax genome from five field-isolates, resulting in ≥93% of the Sal I reference sequenced at coverage greater than 20×. Combined with sequences from one additional Malagasy field isolate and from five monkey-adapted strains, we describe here identification of DNA sequence rearrangements in the P. vivax genome, including discovery of a duplication of the P. vivax Duffy binding protein (PvDBP) gene. A survey of Malagasy patients infected with P. vivax showed that the PvDBP duplication was present in numerous locations in Madagascar and found in over 50% of infected patients evaluated. Extended geographic surveys showed that the PvDBP duplication was detected frequently in vivax patients living in East Africa and in some residents of non-African P. vivax-endemic countries. Additionally, the PvDBP duplication was observed in travelers seeking treatment of vivax malaria upon returning home. PvDBP duplication prevalence was highest in west-central Madagascar sites where the highest frequencies of P. vivax-infected, Duffy-negative people were reported.

Conclusions/Significance

The highly conserved nature of the sequence involved in the PvDBP duplication suggests that it has occurred in a recent evolutionary time frame. These data suggest that PvDBP, a merozoite surface protein involved in red cell adhesion is rapidly evolving, possibly in response to constraints imposed by erythrocyte Duffy negativity in some human populations.     

A High Force of Plasmodium vivax Blood-Stage Infection Drives the Rapid Acquisition of Immunity in Papua New Guinean Children

Background

When both parasite species are co-endemic, Plasmodium vivax incidence peaks in younger children compared to P. falciparum. To identify differences in the number of blood stage infections of these species and its potential link to acquisition of immunity, we have estimated the molecular force of blood-stage infection of P. vivax (_molFOB, i.e. the number of genetically distinct blood-stage infections over time), and compared it to previously reported values for P. falciparum.

Methods

P. vivax _molFOB was estimated by high resolution genotyping parasites in samples collected over 16 months in a cohort of 264 Papua New Guinean children living in an area highly endemic for P. falciparum and P. vivax. In this cohort, P. vivax episodes decreased three-fold over the age range of 1–4.5 years.

Results

On average, children acquired 14.0 new P. vivax blood-stage clones/child/year-at-risk. While the incidence of clinical P. vivax illness was strongly associated with _molFOB (incidence rate ratio (IRR) = 1.99, 95% confidence interval (CI95) [1.80, 2.19]), _molFOB did not change with age. The incidence of P. vivax showed a faster decrease with age in children with high (IRR = 0.49, CI95 [0.38, 0.64] p<0.001) compared to those with low exposure (IRR = 0.63, CI95[0.43, 0.93] p = 0.02).

Conclusion

P. vivax _molFOB is considerably higher than P. falciparum _molFOB (5.5 clones/child/year-at-risk). The high number of P. vivax clones that infect children in early childhood contribute to the rapid acquisition of immunity against clinical P. vivax malaria.     

Naturally Acquired Immune Responses to P. vivax Merozoite Surface Protein 3α and Merozoite Surface Protein 9 Are Associated with Reduced Risk of P. vivax Malaria in Young Papua New Guinean Children

Background

Plasmodium vivax is the most geographically widespread human malaria parasite. Cohort studies in Papua New Guinea have identified a rapid onset of immunity against vivax-malaria in children living in highly endemic areas. Although numerous P. vivax merozoite antigens are targets of naturally acquired antibodies, the role of many of these antibodies in protective immunity is yet unknown.

Methodology/Principal Findings

In a cohort of children aged 1–3 years, antibodies to different regions of Merozoite Surface Protein 3α (PvMSP3α) and Merozoite Surface Protein 9 (PvMSP9) were measured and related to prospective risk of P. vivax malaria during 16 months of active follow-up. Overall, there was a low prevalence of antibodies to PvMSP3α and PvMSP9 proteins (9–65%). Antibodies to the PvMSP3α N-terminal, Block I and Block II regions increased significantly with age while antibodies to the PvMSP3α Block I and PvMSP9 N-terminal regions were positively associated with concurrent P. vivax infection. Independent of exposure (defined as the number of genetically distinct blood-stage infection acquired over time (_molFOB)) and age, antibodies specific to both PvMSP3α Block II (adjusted incidence ratio (aIRR) = 0.59, p = 0.011) and PvMSP9 N-terminus (aIRR = 0.68, p = 0.035) were associated with protection against clinical P. vivax malaria. This protection was most pronounced against high-density infections. For PvMSP3α Block II, the effect was stronger with higher levels of antibodies.

Conclusions

These results indicate that PvMSP3α Block II and PvMSP9 N-terminus should be further investigated for their potential as P. vivax vaccine antigens. Controlling for _molFOB assures that the observed associations are not confounded by individual differences in exposure.     

Is Plasmodium vivax Malaria a Severe Malaria?: A Systematic Review and Meta-Analysis

Background

Plasmodium vivax is one of the major species of malaria infecting humans. Although emphasis on P. falciparum is appropriate, the burden of vivax malaria should be given due attention. This study aimed to synthesize the evidence on severe malaria in P. vivax infection compared with that in P. falciparum infection.

Methods/Principal Findings

We searched relevant studies in electronic databases. The main outcomes required for inclusion in the review were mortality, severe malaria (SM) and severe anaemia (SA). The methodological quality of the included studies was assessed using the Newcastle-Ottawa Scale. Overall, 26 studies were included. The main meta-analysis was restricted to the high quality studies. Eight studies (n = 27490) compared the incidence of SM between P. vivax infection and P. falciparum mono-infection; a comparable incidence was found in infants (OR: 0.45, 95% CI:0.04–5.68, I ²:98%), under 5 year age group (OR: 2.06, 95% CI: 0.83–5.1, I ²:83%), the 5–15 year-age group (OR: 0.6, 95% CI: 0.31–1.16, I ²:81%) and adults (OR: 0.83, 95% CI: 0.67–1.03, I ²:25%). Six studies reported the incidences of SA in P. vivax infection and P. falciparum mono-infection; a comparable incidence of SA was found among infants (OR: 3.47, 95%:0.64–18.94, I ²: 92%), the 5–15 year-age group (OR:0.71, 95% CI: 0.06–8.57, I ²:82%). This was significantly lower in adults (OR:0.75, 95% CI: 0.62–0.92, I ²:0%). Five studies (n = 71079) compared the mortality rate between vivax malaria and falciparum malaria. A lower rate of mortality was found in infants with vivax malaria (OR:0.61, 95% CI:0.5–0.76, I ²:0%), while this was comparable in the 5–15 year- age group (OR: 0.43, 95% CI:0.06–2.91, I ²:84%) and the children of unspecified-age group (OR: 0.77, 95% CI:0.59–1.01, I ²:0%).

Conclusion

Overall, the present analysis identified that the incidence of SM in patients infected with P. vivax was considerable, indicating that P. vivax is a major cause of SM. Awareness of the clinical manifestations of vivax malaria should prompt early detection. Subsequent treatment and monitoring of complications can be life-saving.     

Microsatellite DNA Analysis Revealed a Drastic Genetic Change of Plasmodium vivax Population in the Republic of Korea During 2002 and 2003

Background

Vivax malaria was successfully eliminated in the Republic of Korea (South Korea) in the late 1970s, but it was found to have re-emerged from 1993. In order to control malaria and evaluate the effectiveness of malaria controls, it is important to develop a spatiotemporal understanding of the genetic structure of the parasite population. Here, we estimated the population structure and temporal dynamics of the transmission of Plasmodium vivax in South Korea by analyzing microsatellite DNA markers of the parasite.

Methodology/Principal Findings

We analyzed 14 microsatellite DNA loci of the P. vivax genome from 163 South Korean isolates collected from 1994 to 2008. Allelic data were used to analyze linkage disequilibrium (LD), genetic differentiation and population structure, in order to make a detailed estimate of temporal change in the parasite population. The LD analysis showed a gradual decrease in LD levels, while the levels of genetic differentiation between successive years and analysis of the population structure based on the Bayesian approach suggested that a drastic genetic change occurred in the South Korean population during 2002 and 2003.

Conclusions/Significance

Although relapse and asymptomatic parasite carriage might influence the population structure to some extent, our results suggested the continual introduction of P. vivax into South Korea through other parasite population sources. One possible source, particularly during 2002 and 2003, is North Korea. Molecular epidemiology using microsatellite DNA of the P. vivax population is effective for assessing the population structure and temporal dynamics of parasite transmission; information that can assist in the elimination of vivax malaria in endemic areas.     

Plasmodium vivax adherence to placental glycosaminoglycans

Background

Plasmodium vivax infections seldom kill directly but do cause indirect mortality by reducing birth weight and causing abortion. Cytoadherence and sequestration in the microvasculature are central to the pathogenesis of severe Plasmodium falciparum malaria, but the contribution of cytoadherence to pathology in other human malarias is less clear.

Methodology

The adherence properties of P. vivax infected red blood cells (PvIRBC) were evaluated under static and flow conditions.

Principal Findings

P. vivax isolates from 33 patients were studied. None adhered to immobilized CD36, ICAM-1, or thrombospondin, putative ligands for P. falciparum vascular cytoadherence, or umbilical vein endothelial cells, but all adhered to immobilized chondroitin sulphate A (CSA) and hyaluronic acid (HA), the receptors for adhesion of P. falciparum in the placenta. PvIRBC also adhered to fresh placental cells (N = 5). Pre-incubation with chondroitinase prevented PvIRBC adherence to CSA, and reduced binding to HA, whereas preincubation with hyaluronidase prevented adherence to HA, but did not reduce binding to CSA significantly. Pre-incubation of PvIRBC with soluble CSA and HA reduced binding to the immobilized receptors and prevented placental binding. PvIRBC adhesion was prevented by pre-incubation with trypsin, inhibited by heparin, and reduced by EGTA. Under laminar flow conditions the mean (SD) shear stress reducing maximum attachment by 50% was 0.06 (0.02) Pa but, having adhered, the PvIRBC could then resist detachment by stresses up to 5 Pa. At 37°C adherence began approximately 16 hours after red cell invasion with maximal adherence at 30 hours. At 39°C adherence began earlier and peaked at 24 hours.

Significance

Adherence of P. vivax-infected erythrocytes to glycosaminoglycans may contribute to the pathogenesis of vivax malaria and lead to intrauterine growth retardation.     

Angiopoietin-2 and Angiopoietin-2/Angiopoietin-1 Ratio as Indicators of Potential Severity of Plasmodium vivax Malaria in Patients with Thrombocytopenia

Introduction

Angiogenic factors such as angiopoietin 1 (Ang-1) and angiopoietin 2 (Ang-2) are biomarkers produced during activation and dysfunction of the vascular endothelium in several infectious diseases. The aim of this study was to determine the serum levels of Ang-1 and Ang-2 and to establish their relationship with the main indicators of worst-case prognosis in patients with P. vivax malaria.

Methods

This is a retrospective case-control study nested within a cohort of symptomatic malaria patients. A potentially severe case was defined as a patient that presented at least one of the main indicators of the worst-case prognosis for falciparum malaria, as established by the World Health Organization. Ang-2 and Ang-1 and the Ang-2/Ang-1 ratio were used to analyze the role of angiopoietins as biomarkers in signaling potentially severe vivax malaria. ROC curves were generated to identify a cut-off point discriminating between the angiopoietin concentrations that were most strongly associated with potential infection severity.

Results

The serum levels of Ang-2 and the Ang-2/Ang-1 ratio were higher in the case group. In contrast, the serum levels of Ang-1 were lower in the cases than in the control patients. The blood count for platelets showed a positive correlation with Ang-1 and a negative correlation with Ang-2 and with the Ang-2/Ang-1 ratio. The area under the ROC curve (AUC) for serum angiopoietins, as an indicator of worst-case prognosis in a potentially severe P. vivax malarial infection, was larger in the subgroup of patients with platelet counts <75,000/µL.

Conclusion

This study showed that patients with predictors of worst-case prognoses for P. vivax malaria have lower Ang-1 and higher Ang-2 serum levels (and higher values for the Ang-2/Ang-1 ratio) than controls. Elevated serum levels of Ang-2 and high values for the Ang-2/Ang-1 ratio may potentially be used as predictors of worst-case prognoses for P. vivax malaria, especially in patients with thrombocytopenia.