Hotspots of Malaria Transmission in the Peruvian Amazon: Rapid Assessment through a Parasitological and Serological Survey

Background

With low and markedly seasonal malaria transmission, increasingly sensitive tools for better stratifying the risk of infection and targeting control interventions are needed. A cross-sectional survey to characterize the current malaria transmission patterns, identify hotspots, and detect recent changes using parasitological and serological measures was conducted in three sites of the Peruvian Amazon.

Material and Methods

After full census of the study population, 651 participants were interviewed, clinically examined and had a blood sample taken for the detection of malaria parasites (microscopy and PCR) and antibodies against P. vivax (PvMSP1₁₉, PvAMA1) and P. falciparum (PfGLURP, PfAMA1) antigens by ELISA. Risk factors for malaria infection (positive PCR) and malaria exposure (seropositivity) were assessed by multivariate survey logistic regression models. Age-specific seroprevalence was analyzed using a reversible catalytic conversion model based on maximum likelihood for generating seroconversion rates (SCR, λ). SaTScan was used to detect spatial clusters of serology-positive individuals within each site.

Results

The overall parasite prevalence by PCR was low, i.e. 3.9% for P. vivax and 6.7% for P. falciparum, while the seroprevalence was substantially higher, 33.6% for P. vivax and 22.0% for P. falciparum, with major differences between study sites. Age and location (site) were significantly associated with P. vivax exposure; while location, age and outdoor occupation were associated with P. falciparum exposure. P. falciparum seroprevalence curves showed a stable transmission throughout time, while for P. vivax transmission was better described by a model with two SCRs. The spatial analysis identified well-defined clusters of P. falciparum seropositive individuals in two sites, while it detected only a very small cluster of P. vivax exposure.

Conclusion

The use of a single parasitological and serological malaria survey has proven to be an efficient and accurate method to characterize the species specific heterogeneity in malaria transmission at micro-geographical level as well as to identify recent changes in transmission.     

A Library of Plasmodium vivax Recombinant Merozoite Proteins Reveals New Vaccine Candidates and Protein-Protein Interactions

Background

A vaccine targeting Plasmodium vivax will be an essential component of any comprehensive malaria elimination program, but major gaps in our understanding of P. vivax biology, including the protein-protein interactions that mediate merozoite invasion of reticulocytes, hinder the search for candidate antigens. Only one ligand-receptor interaction has been identified, that between P. vivax Duffy Binding Protein (PvDBP) and the erythrocyte Duffy Antigen Receptor for Chemokines (DARC), and strain-specific immune responses to PvDBP make it a complex vaccine target. To broaden the repertoire of potential P. vivax merozoite-stage vaccine targets, we exploited a recent breakthrough in expressing full-length ectodomains of Plasmodium proteins in a functionally-active form in mammalian cells and initiated a large-scale study of P. vivax merozoite proteins that are potentially involved in reticulocyte binding and invasion.

Methodology/Principal Findings

We selected 39 P. vivax proteins that are predicted to localize to the merozoite surface or invasive secretory organelles, some of which show homology to P. falciparum vaccine candidates. Of these, we were able to express 37 full-length protein ectodomains in a mammalian expression system, which has been previously used to express P. falciparum invasion ligands such as PfRH5. To establish whether the expressed proteins were correctly folded, we assessed whether they were recognized by antibodies from Cambodian patients with acute vivax malaria. IgG from these samples showed at least a two-fold change in reactivity over naïve controls in 27 of 34 antigens tested, and the majority showed heat-labile IgG immunoreactivity, suggesting the presence of conformation-sensitive epitopes and native tertiary protein structures. Using a method specifically designed to detect low-affinity, extracellular protein-protein interactions, we confirmed a predicted interaction between P. vivax 6-cysteine proteins P12 and P41, further suggesting that the proteins are natively folded and functional. This screen also identified two novel protein-protein interactions, between P12 and PVX_110945, and between MSP3.10 and MSP7.1, the latter of which was confirmed by surface plasmon resonance.

Conclusions/Significance

We produced a new library of recombinant full-length P. vivax ectodomains, established that the majority of them contain tertiary structure, and used them to identify predicted and novel protein-protein interactions. As well as identifying new interactions for further biological studies, this library will be useful in identifying P. vivax proteins with vaccine potential, and studying P. vivax malaria pathogenesis and immunity.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00663546","term_id":"NCT00663546"}}NCT00663546     

Contrasting Transmission Dynamics of Co-endemic Plasmodium vivax and P. falciparum: Implications for Malaria Control and Elimination

Background

Outside of Africa, P. falciparum and P. vivax usually coexist. In such co-endemic regions, successful malaria control programs have a greater impact on reducing falciparum malaria, resulting in P. vivax becoming the predominant species of infection. Adding to the challenges of elimination, the dormant liver stage complicates efforts to monitor the impact of ongoing interventions against P. vivax. We investigated molecular approaches to inform the respective transmission dynamics of P. falciparum and P. vivax and how these could help to prioritize public health interventions.

Methodology/ Principal Findings

Genotype data generated at 8 and 9 microsatellite loci were analysed in 168 P. falciparum and 166 P. vivax isolates, respectively, from four co-endemic sites in Indonesia (Bangka, Kalimantan, Sumba and West Timor). Measures of diversity, linkage disequilibrium (LD) and population structure were used to gauge the transmission dynamics of each species in each setting. Marked differences were observed in the diversity and population structure of P. vivax versus P. falciparum. In Bangka, Kalimantan and Timor, P. falciparum diversity was low, and LD patterns were consistent with unstable, epidemic transmission, amenable to targeted intervention. In contrast, P. vivax diversity was higher and transmission appeared more stable. Population differentiation was lower in P. vivax versus P. falciparum, suggesting that the hypnozoite reservoir might play an important role in sustaining local transmission and facilitating the spread of P. vivax infections in different endemic settings. P. vivax polyclonality varied with local endemicity, demonstrating potential utility in informing on transmission intensity in this species.

Conclusions/ Significance

Molecular approaches can provide important information on malaria transmission that is not readily available from traditional epidemiological measures. Elucidation of the transmission dynamics circulating in a given setting will have a major role in prioritising malaria control strategies, particularly against the relatively neglected non-falciparum species.     

Plasmodium vivax Malaria in Pregnant Women in the Brazilian Amazon and the Risk Factors Associated with Prematurity and Low Birth Weight: A Descriptive Study

Introduction

Plasmodium vivax is the most prevalent malaria species in the American region. Brazil accounts for the higher number of the malaria cases reported in pregnant women in the Americas. This study aims to describe the characteristics of pregnant women with malaria in an endemic area of the Brazilian Amazon and the risk factors associated with prematurity and low birth weight (LBW).

Methods/Principal Findings

Between December 2005 and March 2008, 503 pregnant women with malaria that attended a tertiary health centre were enrolled and followed up until delivery and reported a total of 1016 malaria episodes. More than half of study women (54%) were between 20–29 years old, and almost a third were adolescents. The prevalence of anaemia at enrolment was 59%. Most women (286/503) reported more than one malaria episode and most malaria episodes (84.5%, 846/1001) were due to P. vivax infection. Among women with only P. vivax malaria, the risk of preterm birth and low birth weight decreased in multigravidae (OR, 0.36 [95% CI, 0.16–0.82]; p = 0.015 and OR 0.24 [95% CI, 0.10–0.58]; p = 0.001, respectively). The risk of preterm birth decreased with higher maternal age (OR 0.43 [95% CI, 0.19–0.95]; p = 0.037) and among those women who reported higher antenatal care (ANC) attendance (OR, 0.32 [95% CI, 0.15–0.70]; p = 0.005).

Conclusion

This study shows that P. vivax is the prevailing species among pregnant women with malaria in the region and shows that vivax clinical malaria may represent harmful consequences for the health of the mother and their offsprings particularly on specific groups such as adolescents, primigravidae and those women with lower ANC attendance.     

Prevalence and Predictors of Asymptomatic Malaria Parasitemia among Pregnant Women in the Rural Surroundings of Arbaminch Town,South Ethiopia

Background

In Sub-Saharan African countries, including Ethiopia, malaria in pregnancy is a major public health threat which results in significant morbidities and mortalities among pregnant women and their fetuses. In malaria endemic areas, Plasmodium infections tend to remain asymptomatic yet causing significant problems like maternal anemia, low birth weight, premature births, and still birth. This study was conducted to determine the prevalence and predictors of asymptomatic Plasmodium infection among pregnant women in the rural surroundings of Arba Minch Town, Southern Ethiopia.

Methods

A community based cross-sectional study comprising multistage sampling was conducted between April and June, 2013. Socio-demographic data were collected by using a semi-structured questionnaire. Plasmodium infection was diagnosed by using Giemsa-stained blood smear microscopy and a rapid diagnostic test (SD BIOLINE Malaria Ag Pf/Pv POCT, standard diagnostics, inc., Korea).

Results

Of the total 341 pregnant women participated in this study, 9.1% (31/341) and 9.7% (33/341) were confirmed to be infected with Plasmodium species by microscopy and rapid diagnostic tests (RDTs), respectively. The geometric mean of parasite density was 2392 parasites per microliter (μl); 2275/ μl for P. falciparum and 2032/ μl for P. vivax. Parasitemia was more likely to occur in primigravidae (Adjusted odds ratio (AOR): 9.4, 95% confidence interval (CI): 4.3–60.5), secundigravidae (AOR: 6.3, 95% CI: 2.9–27.3), using insecticide treated bed net (ITN) sometimes (AOR: 3.2, 95% CI: 1.8- 57.9), not using ITN at all (AOR: 4.6, 95% CI: 1.4–14.4) compared to multigravidae and using ITN always, respectively.

Conclusion

Asymptomatic malaria in this study is low compared to other studies’ findings. Nevertheless, given the high risk of malaria during pregnancy, pregnant women essentially be screened for asymptomatic Plasmodium infection and be treated promptly via the antenatal care (ANC) services.     

Multiplicity of Infection and Disease Severity in Plasmodium vivax

Background

Multiplicity of infection (MOI) refers to the average number of distinct parasite genotypes concurrently infecting a patient. Although several studies have reported on MOI and the frequency of multiclonal infections in Plasmodium falciparum, there is limited data on Plasmodium vivax. Here, MOI and the frequency of multiclonal infections were studied in areas from South America where P. vivax and P. falciparum can be compared.

Methodology/Principal Findings

As part of a passive surveillance study, 1,328 positive malaria patients were recruited between 2011 and 2013 in low transmission areas from Colombia. Of those, there were only 38 P. vivax and 24 P. falciparum clinically complicated cases scattered throughout the time of the study. Samples from uncomplicated cases were matched in time and location with the complicated cases in order to compare the circulating genotypes for these two categories. A total of 92 P. vivax and 57 P. falciparum uncomplicated cases were randomly subsampled. All samples were genotyped by using neutral microsatellites. Plasmodium vivax showed more multiclonal infections (47.7%) than P. falciparum (14.8%). Population genetics and haplotype network analyses did not detect differences in the circulating genotypes between complicated and uncomplicated cases in each parasite. However, a Fisher exact test yielded a significant association between having multiclonal P. vivax infections and complicated malaria. No association was found for P. falciparum infections.

Conclusion

The association between multiclonal infections and disease severity in P. vivax is consistent with previous observations made in rodent malaria. The contrasting pattern between P. vivax and P. falciparum could be explained, at least in part, by the fact that P. vivax infections have lineages that were more distantly related among them than in the case of the P. falciparum multiclonal infections. Future research should address the possible role that acquired immunity and exposure may have on multiclonal infections and their association with disease severity.     

Further Evidence of Increasing Diversity of Plasmodium vivax in the Republic of Korea in Recent Years

Background

Vivax malaria was successfully eliminated from the Republic of Korea (ROK) in the late 1970s but re-emerged in 1993. Two decades later as the ROK enters the final stages of malaria elimination, dedicated surveillance of the local P. vivax population is critical. We apply a population genetic approach to gauge P. vivax transmission dynamics in the ROK between 2010 and 2012.

Methodology/Principal Findings

P. vivax positive blood samples from 98 autochthonous cases were collected from patients attending health centers in the ROK in 2010 (n = 27), 2011 (n = 48) and 2012 (n = 23). Parasite genotyping was undertaken at 9 tandem repeat markers. Although not reaching significance, a trend of increasing population diversity was observed from 2010 (H_E = 0.50 ± 0.11) to 2011 (H_E = 0.56 ± 0.08) and 2012 (H_E = 0.60 ± 0.06). Conversely, linkage disequilibrium declined during the same period: I_AS = 0.15 in 2010 (P = 0.010), 0.09 in 2011 (P = 0.010) and 0.05 in 2012 (P = 0.010). In combination with data from other ROK studies undertaken between 1994 and 2007, our results are consistent with increasing parasite divergence since re-emergence. Polyclonal infections were rare (3% infections) suggesting that local out-crossing alone was unlikely to explain the increased divergence. Cases introduced from an external reservoir may therefore have contributed to the increased diversity. Aside from one isolate, all infections carried a short MS20 allele (142 or 149 bp), not observed in other studies in tropical endemic countries despite high diversity, inferring that these regions are unlikely reservoirs.

Conclusions

Whilst a number of factors may explain the observed population genetic trends, the available evidence suggests that an external geographic reservoir with moderate diversity sustains the majority of P. vivax infection in the ROK, with important implications for malaria elimination.     

Differential Diagnosis of Malaria on Truelab Uno?, a Portable,Real-Time,MicroPCR Device for Point-Of-Care Applications

Background

Sensitive and specific detection of malarial parasites is crucial in controlling the significant malaria burden in the developing world. Also important is being able to identify life threatening Plasmodium falciparum malaria quickly and accurately to reduce malaria related mortality. Existing methods such as microscopy and rapid diagnostic tests (RDTs) have major shortcomings. Here, we describe a new real-time PCR-based diagnostic test device at point-of-care service for resource-limited settings.

Methods

Truenat^® Malaria, a chip-based microPCR test, was developed by bigtec Labs, Bangalore, India, for differential identification of Plasmodium falciparum and Plasmodium vivax parasites. The Truenat Malaria tests runs on bigtec’s Truelab Uno^® microPCR device, a handheld, battery operated, and easy-to-use real-time microPCR device. The performance of Truenat^® Malaria was evaluated versus the WHO nested PCR protocol. The Truenat^® Malaria was further evaluated in a triple-blinded study design using a sample panel of 281 specimens created from the clinical samples characterized by expert microscopy and a rapid diagnostic test kit by the National Institute of Malaria Research (NIMR). A comparative evaluation was done on the Truelab Uno^® and a commercial real-time PCR system.

Results

The limit of detection of the Truenat Malaria assay was found to be <5 parasites/μl for both P. falciparum and P. vivax. The Truenat^® Malaria test was found to have sensitivity and specificity of 100% each, compared to the WHO nested PCR protocol based on the evaluation of 100 samples. The sensitivity using expert microscopy as the reference standard was determined to be around 99.3% (95% CI: 95.5–99.9) at the species level. Mixed infections were identified more accurately by Truenat Malaria (32 samples identified as mixed) versus expert microscopy and RDTs which detected 4 and 5 mixed samples, respectively.

Conclusion

The Truenat^® Malaria microPCR test is a valuable diagnostic tool and implementation should be considered not only for malaria diagnosis but also for active surveillance and epidemiological intervention.     

Parasite Specific Antibody Increase Induced by an Episode of Acute P. falciparum Uncomplicated Malaria

Introduction

There is no approved vaccine for malaria, and precisely how human antibody responses to malaria parasite components and potential vaccine molecules are developed and maintained remains poorly defined. In this study, antibody anamnestic or memory response elicited by a single episode of P. falciparum infection was investigated.

Methods

This study involved 362 malaria patients aged between 6 months to 60 years, of whom 19% were early-diagnosed people living with HIV/AIDS (PLWHA). On the day malaria was diagnosed and 42 days later, blood specimens were collected. Parasite density, CD4⁺ cells, and antibodies specific to synthetic peptides representing antigenic regions of the P. falciparum proteins GLURP, MSP3 and HRPII were measured.

Results

On the day of malaria diagnosis, Immunoglobulin (IgG) antibodies against GLURP, MSP3 and HRP II peptides were present in the blood of 75%, 41% and 60% of patients, respectively. 42 days later, the majority of patients had boosted their serum IgG antibody more than 1.2 fold. The increase in level of IgG antibody against the peptides was not affected by parasite density at diagnosis. The median CD4⁺ cell counts of PLWHAs and HIV negative individuals were not statistically different, and median post-infection increases in anti-peptide IgG were similar in both groups of patients.

Conclusion

In the majority (70%) of individuals, an infection of P. falciparum elicits at least 20% increase in level of anti-parasite IgG. This boost in anti-P. falciparum IgG is not affected by parasite density on the day of malaria diagnosis, or by HIV status.     

Recognition of Human Erythrocyte Receptors by the Tryptophan-Rich Antigens of Monkey Malaria Parasite Plasmodium knowlesi

Background

The monkey malaria parasite Plasmodium knowlesi also infect humans. There is a lack of information on the molecular mechanisms that take place between this simian parasite and its heterologous human host erythrocytes leading to this zoonotic disease. Therefore, we investigated here the binding ability of P. knowlesi tryptophan-rich antigens (PkTRAgs) to the human erythrocytes and sharing of the erythrocyte receptors between them as well as with other commonly occurring human malaria parasites.

Methods

Six PkTRAgs were cloned and expressed in E.coli as well as in mammalian CHO-K1 cell to determine their human erythrocyte binding activity by cell-ELISA, and in-vitro rosetting assay, respectively.

Results

Three of six PkTRAgs (PkTRAg38.3, PkTRAg40.1, and PkTRAg67.1) showed binding to human erythrocytes. Two of them (PkTRAg40.1 and PkTRAg38.3) showed cross-competition with each other as well as with the previously described P.vivax tryptophan-rich antigens (PvTRAgs) for human erythrocyte receptors. However, the third protein (PkTRAg67.1) utilized the additional but different human erythrocyte receptor(s) as it did not cross-compete for erythrocyte binding with either of these two PkTRAgs as well as with any of the PvTRAgs. These three PkTRAgs also inhibited the P.falciparum parasite growth in in-vitro culture, further indicating the sharing of human erythrocyte receptors by these parasite species and the biological significance of this receptor-ligand interaction between heterologous host and simian parasite.

Conclusions

Recognition and sharing of human erythrocyte receptor(s) by PkTRAgs with human parasite ligands could be part of the strategy adopted by the monkey malaria parasite to establish inside the heterologous human host.     

Micronutrient Deficiencies and Plasmodium vivax Malaria among Children in the Brazilian Amazon

Background

There is a growing body of evidence linking micronutrient deficiencies and malaria incidence arising mostly from P. falciparum endemic areas. We assessed the impact of micronutrient deficiencies on malaria incidence and vice versa in the Brazilian state of Amazonas.

Methodology/Principal Findings

We evaluated children <10 years old living in rural communities in the state of Amazonas, Brazil, from May 2010 to May 2011. All children were assessed for sociodemographic, anthropometric and laboratory parameters, including vitamin A, beta-carotene, zinc and iron serum levels at the beginning of the study (May 2010) and one year later (May 2011). Children were followed in between using passive surveillance for detection of symptomatic malaria. Those living in the study area at the completion of the observation period were reassessed for micronutrient levels. Univariate Cox-proportional Hazards models were used to assess whether micronutrient deficiencies had an impact on time to first P. vivax malaria episode. We included 95 children median age 4.8 years (interquartile range [IQR]: 2.3–6.6), mostly males (60.0%) and with high maternal illiteracy (72.6%). Vitamin A deficiencies were found in 36% of children, beta-carotene deficiency in 63%, zinc deficiency in 61% and iron deficiency in 51%. Most children (80%) had at least one intestinal parasite. During follow-up, 16 cases of vivax malaria were diagnosed amongst 13 individuals. Micronutrient deficiencies were not associated with increased malaria incidence: vitamin A deficiency [Hazard ratio (HR): 1.51; P-value: 0.45]; beta-carotene [HR: 0.47; P-value: 0.19]; zinc [HR: 1.41; P-value: 0.57] and iron [HR: 2.31; P-value: 0.16]). Upon reevaluation, children with al least one episode of malaria did not present significant changes in micronutrient levels.

Conclusion

Micronutrient serum levels were not associated with a higher malaria incidence nor the malaria episode influenced micronutrient levels. Future studies targeting larger populations to assess micronutrients levels in P. vivax endemic areas are warranted in order to validate these results.     

Phase I Clinical Trial of a Recombinant Blood Stage Vaccine Candidate for Plasmodium falciparum Malaria Based on MSP1 and EBA175

Background

A phase I randomised, controlled, single blind, dose escalation trial was conducted to evaluate safety and immunogenicity of JAIVAC-1, a recombinant blood stage vaccine candidate against Plasmodium falciparum malaria, composed of a physical mixture of two recombinant proteins, PfMSP-1₁₉, the 19 kD conserved, C-terminal region of PfMSP-1 and PfF2 the receptor-binding F2 domain of EBA175.

Method

Healthy malaria naïve Indian male subjects aged 18–45 years were recruited from the volunteer database of study site. Fifteen subjects in each cohort, randomised in a ratio of 2:1 and meeting the protocol specific eligibility criteria, were vaccinated either with three doses (10μg, 25μg and 50μg of each antigen) of JAIVAC-1 formulated with adjuvant Montanide ISA 720 or with standard dosage of Hepatitis B vaccine. Each subject received the assigned vaccine in the deltoid muscle of the upper arms on Day 0, Day 28 and Day 180.

Results

JAIVAC-1 was well tolerated and no serious adverse event was observed. All JAIVAC-1 subjects sero-converted for PfF2 but elicited poor immune response to PfMSP-1₁₉. Dose-response relationship was observed between vaccine dose of PfF2 and antibody response. The antibodies against PfF2 were predominantly of IgG1 and IgG3 isotype. Sera from JAIVAC-1 subjects reacted with late schizonts in a punctate pattern in immunofluorescence assays. Purified IgG from JAIVAC-1 sera displayed significant growth inhibitory activity against Plasmodium falciparum CAMP strain.

Conclusion

Antigen PfF2 should be retained as a component of a recombinant malaria vaccine but PfMSP-1₁₉ construct needs to be optimised to improve its immunogenicity.

Trial Registration

Clinical Trial Registry, India CTRI/2010/091/000301     

Epidemiology and Clinical Burden of Malaria in the War-Torn Area,Orakzai Agency in Pakistan

Background

Military conflict has been a major challenge in the detection and control of emerging infectious diseases such as malaria. It poses issues associated with enhancing emergence and transmission of infectious diseases by destroying infrastructure and collapsing healthcare systems. The Orakzai agency in Pakistan has witnessed a series of intense violence and destruction. Military conflicts and instability in Afghanistan have resulted in the migration of refugees into the area and possible introduction of many infectious disease epidemics. Due to the ongoing violence and Talibanization, it has been a challenge to conduct an epidemiological study.

Methodology/Principal Findings

All patients were sampled within the transmission season. After a detailed clinical investigation of patients, data were recorded. Baseline venous blood samples were taken for microscopy and nested polymerase chain reaction (nPCR) analysis. Plasmodium species were detected using nested PCR (nPCR) and amplification of the small subunit ribosomal ribonucleic acid (ssrRNA) genes using the primer pairs. We report a clinical assessment of the epidemic situation of malaria caused by Plasmodium vivax (86.5%) and Plasmodium falciparum (11.79%) infections with analysis of complications in patients such as decompensated shock (41%), anemia (8.98%), hypoglycaemia (7.3%), multiple convulsions (6.7%), hyperpyrexia (6.17%), jaundice (5%), and hyperparasitaemia (4.49%).

Conclusions/Significance

This overlooked distribution of P. vivax should be considered by malaria control strategy makers in the world and by the Government of Pakistan. In our study, children were the most susceptible population to malaria infection while they were the least expected to use satisfactory prevention strategies in such a war-torn deprived region. Local health authorities should initiate malaria awareness programs in schools and malaria-related education should be further promoted at the local level reaching out to both children and parents.     

Ocular Dominance Is Associated with the Ganglion Cell-Inner Plexiform Layer Thickness Profile in the Macula

Purpose

To investigate the characteristics of macular ganglion cell-inner plexiform layer (GCIPL) thickness profiles associated with ocular dominance.

Setting

Private practice, Seoul, Republic of Korea.

Design

Comparative case-control study.

Methods

Both eyes of 199 participants with no ophthalmic abnormalities were included. Participants were imaged by spectral-domain optical coherence tomography, and underwent dominant eye testing using a hole-in-a-card test (sighting dominance) at the same visit. Macular GCIPL, as well as circumpapillary retinal nerve fiber layer (RNFL) thickness were compared for individual patients, according to ocular dominance.

Results

Ocular dominance occurred predominantly in the right eye (right vs. left: 72.36 vs. 27.60%; P < 0.001). In the comparison of macular GCIPL thickness, the average (81.27±5.01 μm vs. 80.66±6.31 μm in dominant vs. non-dominant eyes), inferonasal (81.39±5.47μm vs. 80.33±6.82μm, and inferior sectors (77.95±6.05μm vs. 76.97±8.15μm) were significantly different between dominant and non-dominant eyes (P = 0.040, 0.005, and 0.032, respectively). Significant predictors of average GCIPL thickness were spherical equivalent (β = 1.37, P<0.001), astigmatic power (β = 1.44, P = 0.009), disc area (β = 3.90, P < 0.001), average RNFL thickness (β = 0.22, P<0.001), average cup-to-disc ratio (β = 5.74, P = 0.002), difference between the inferior and superior quadrant RNFL thicknesses (β = 0.08, P = 0.024), and ocular dominance (β = 2.10, P = 0.020). On multivariate regression analysis, ocular dominance was correlated with average GCIPL thickness after adjusting for potential confounders (β = 1.63, P = 0.048).

Conclusions

Dominant eyes accompanied significantly thicker average macular GCIPL. This information suggests that macular GCIPL thickness may provide an indicator of the relative dominance of an eye.     

Heterologous Protection against Malaria after Immunization with Plasmodium falciparum Sporozoites

Background

Sterile protection in >90% of volunteers against homologous Plasmodium falciparum infection has been achieved only using the controlled human malaria infection (CHMI) model. This efficient model involves whole parasite immunizations under chloroquine prophylaxis (CPS-immunization), requiring only 30–45 mosquitoes bites infected with P. falciparum-sporozoites. Given the large diversity of P. falciparum parasites, it is essential to assess protection against heterologous parasite strains.

Methods

In an open-label follow-up study, 16 volunteers previously CPS-immunized and challenged with P. falciparum NF54 (West-Africa) in a dose de-escalation and challenge trial were re-challenged with clone NF135.C10 (Cambodia) at 14 months after the last immunization (NCT01660854).

Results

Two out of thirteen NF54 protected volunteers previously fully protected against NF54 were also fully protected against NF135.C10, while 11/13 showed a delayed patency (median prepatent period of 10.5 days (range 9.0–15.5) versus 8.5 days in 5 malaria-naïve controls (p = 0.0005). Analysis of patency by qPCR indicated a 91 to >99% estimated reduction of liver parasite load in 7/11 partially protected subjects. Three volunteers previously not protected against NF54, were also not protected against NF135.C10.

Conclusion

This study shows that CPS-immunization can induce heterologous protection for a period of more than one year, which is a further impetus for clinical development of whole parasite vaccines.

Trial Registration

Clinicaltrials.gov NCT01660854     

Presence of Plasmodium falciparum DNA in Plasma Does Not Predict Clinical Malaria in an HIV-1 Infected Population

Background

HIV-1 and Plasmodium falciparum malaria cause substantial morbidity in Sub-Saharan Africa, especially as co-infecting pathogens. We examined the relationship between presence of P. falciparum DNA in plasma samples and clinical malaria as well as the impact of atazanavir, an HIV-1 protease inhibitor (PI), on P. falciparum PCR positivity.

Methods

ACTG study A5175 compared two NNRTI-based regimens and one PI-based anti-retroviral (ARV) regimen in antiretroviral therapy naïve participants. We performed nested PCR on plasma samples for the P. falciparum 18s rRNA gene to detect the presence of malaria DNA in 215 of the 221 participants enrolled in Blantyre and Lilongwe, Malawi. We also studied the closest sample preceding the first malaria diagnosis from 102 persons with clinical malaria and randomly selected follow up samples from 88 persons without clinical malaria.

Results

PCR positivity was observed in 18 (8%) baseline samples and was not significantly associated with age, sex, screening CD4+ T-cell count, baseline HIV-1 RNA level or co-trimoxazole use within the first 8 weeks. Neither baseline PCR positivity (p = 0.45) nor PCR positivity after initiation of antiretroviral therapy (p = 1.0) were significantly associated with subsequent clinical malaria. Randomization to the PI versus NNRTI ARV regimens was not significantly associated with either PCR positivity (p = 0.5) or clinical malaria (p = 0.609). Clinical malaria was associated with a history of tuberculosis (p = 0.006) and a lower BMI (p = 0.004).

Conclusion

P. falciparum DNA was detected in 8% of participants at baseline, but was not significantly associated with subsequent development of clinical malaria. HIV PI therapy did not decrease the prevalence of PCR positivity or incidence of clinical disease.     

A Cholera Conjugate Vaccine Containing O-specific Polysaccharide (OSP) of V. cholerae O1 Inaba and Recombinant Fragment of Tetanus Toxin Heavy Chain (OSP:rTTHc) Induces Serum,Memory and Lamina Proprial Responses against OSP and Is Protective in Mice

Background

Vibrio cholerae is the cause of cholera, a severe watery diarrhea. Protection against cholera is serogroup specific. Serogroup specificity is defined by the O-specific polysaccharide (OSP) component of lipopolysaccharide (LPS).

Methodology

Here we describe a conjugate vaccine for cholera prepared via squaric acid chemistry from the OSP of V. cholerae O1 Inaba strain PIC018 and a recombinant heavy chain fragment of tetanus toxin (OSP:rTTHc). We assessed a range of vaccine doses based on the OSP content of the vaccine (10-50 μg), vaccine compositions varying by molar loading ratio of OSP to rTTHc (3:1, 5:1, 10:1), effect of an adjuvant, and route of immunization.

Principle Findings

Immunized mice developed prominent anti-OSP and anti-TT serum IgG responses, as well as vibriocidal antibody and memory B cell responses following intramuscular or intradermal vaccination. Mice did not develop anti-squarate responses. Intestinal lamina proprial IgA responses targeting OSP occurred following intradermal vaccination. In general, we found comparable immune responses in mice immunized with these variations, although memory B cell and vibriocidal responses were blunted in mice receiving the highest dose of vaccine (50 μg). We found no appreciable change in immune responses when the conjugate vaccine was administered in the presence or absence of immunoadjuvant alum. Administration of OSP:rTTHc resulted in 55% protective efficacy in a mouse survival cholera challenge model.

Conclusion

We report development of an Inaba OSP:rTTHc conjugate vaccine that induces memory responses and protection against cholera in mice. Development of an effective cholera conjugate vaccine that induces high level and long-term immune responses against OSP would be beneficial, especially in young children who respond poorly to polysaccharide antigens.     

Seasonal and Spatial Dynamics of the Primary Vector of Plasmodium knowlesi within a Major Transmission Focus in Sabah,Malaysia

Background

The simian malaria parasite Plasmodium knowlesi is emerging as a public health problem in Southeast Asia, particularly in Malaysian Borneo where it now accounts for the greatest burden of malaria cases and deaths. Control is hindered by limited understanding of the ecology of potential vector species.

Methodology/Principal Findings

We conducted a one year longitudinal study of P. knowlesi vectors in three sites within an endemic area of Sabah, Malaysia. All mosquitoes were captured using human landing catch. Anopheles mosquitoes were dissected to determine, oocyst, sporozoites and parous rate. Anopheles balabacensis is confirmed as the primary vector of. P. knowlesi (using nested PCR) in Sabah for the first time. Vector densities were significantly higher and more seasonally variable in the village than forest or small scale farming site. However An. balabacensis survival and P. knowlesi infection rates were highest in forest and small scale farm sites. Anopheles balabacensis mostly bites humans outdoors in the early evening between 1800 to 2000hrs.

Conclusions/Significance

This study indicates transmission is unlikely to be prevented by bednets. This combined with its high vectorial capacity poses a threat to malaria elimination programmes within the region.