RESUMEN

India experienced the second wave of SARS-CoV-2 infection from April 3 to June 10, 2021. During the second wave, Delta variant B.1617.2 emerged as the predominant strain, spiking cases from 12.5 million to 29.3 million (cumulative) by the end of the surge in India. Vaccines against COVID-19 are a potent tool to control and end the pandemic in addition to other control measures. India rolled out its vaccination programme on January 16, 2021, initially with two vaccines that were given emergency authorization-Covaxin (BBV152) and Covishield (ChAdOx1 nCoV- 19). Vaccination was initially started for the elderly (60+) and front-line workers and then gradually opened to different age groups. The second wave hit when vaccination was picking up pace in India. There were instances of vaccinated people (fully and partially) getting infected, and reinfections were also reported. We undertook a survey of staff (front line health care workers and supporting) of 15 medical colleges and research institutes across India to assess the vaccination coverage, incidence of breakthrough infections, and reinfections among them from June 2 to July 10, 2021. A total of 1876 staff participated, and 1484 forms were selected for analysis after removing duplicates and erroneous entries (n = 392). We found that among the respondents at the time of response, 17.6% were unvaccinated, 19.8% were partially vaccinated (received the first dose), and 62.5% were fully vaccinated (received both doses). Incidence of breakthrough infections was 8.7% among the 801 individuals (70/801) tested at least 14 days after the 2nd dose of vaccine. Eight participants reported reinfection in the overall infected group and reinfection incidence rate was 5.1%. Out of (N = 349) infected individuals 243 (69.6%) were unvaccinated and 106 (30.3%) were vaccinated. Our findings reveal the protective effect of vaccination and its role as an essential tool in the struggle against this pandemic.

RESUMEN

Iron-sulfur (Fe-S) cluster containing proteins have been assigned roles in various essential cellular processes, such as regulation of gene expression, electron transfer, sensing of oxygen and balancing free radical chemistry. However, their role as the drug target remains sparse. Recently the screening of protein alkylation targets for artemisinin in Plasmodium falciparum led to identification of Dre2, a protein involved in redox mechanism for the cytoplasmic Fe-S cluster assembly in different organisms. In the present study, to further explore the interaction between artemisinin and Dre2, we have expressed the Dre2 protein of both P. falciparum and P. vivax in E. coli. The opaque brown colour of the IPTG induced recombinant Plasmodium Dre2 bacterial pellet, suggested iron accumulation as confirmed by the ICP-OES analysis. In addition, overexpression of rPvDre2 in E. coli reduced its viability, growth and increased the ROS levels of bacterial cells, which in turn led to an increase in expression of stress response genes of E. coli such as recA, soxS, mazF. Moreover, the overexpression of rDre2 induced cell death could be rescued by treatment with Artemisinin derivatives suggesting their interaction. The interaction between DHA and PfDre2 was later demonstrated by CETSA and microscale thermophoresis. Overall, this study suggests that Dre2 is the probable target of Artemisinin and the antimalarial activity of DHA/Artemether could also be due to yet unidentified molecular mechanism altering the Dre2 activity in addition to inducing DNA and protein damage.

Asunto(s)

Artemisininas , Proteínas de Escherichia coli , Proteínas Hierro-Azufre , Plasmodium , Artemisininas/farmacología , Proteínas de Unión al ADN/metabolismo , Endorribonucleasas , Escherichia coli/genética , Escherichia coli/metabolismo , Proteínas de Escherichia coli/metabolismo , Hierro/metabolismo , Proteínas Hierro-Azufre/genética , Proteínas Hierro-Azufre/química , Proteínas Hierro-Azufre/metabolismo , Plasmodium/metabolismo , Azufre/metabolismo

RESUMEN

Objective: The objective of this subanalysis of data from centres across urban areas in India of the Global Asthma Network (GAN) was to study 1) the prevalence of symptoms of asthma in children and adults, 2) the change in prevalence of asthma and its trigger factors since the International Study of Asthma and Allergies in Childhood (ISAAC), and 3) current asthma treatment practice. Methods: In this cross-sectional, multicentre, school-based and self-administered questionnaire, responses from children aged 6-7 years and 13-14 years, and their respective parents, were analysed. Results: The GAN Phase I study included 20 084 children in the 6-7-year age group, 25 887 children in the 13-14-year age group and 81 296 parents. The prevalence of wheeze in the previous 12 months was 3.16%, 3.63% and 3.30% in the three groups, respectively. In comparison to the ISAAC studies, there was a significant reduction in the prevalence of current wheeze (p<0.001). Bivariate analysis revealed a significant reduction in the prevalence of trigger factors. Almost 82% of current wheezers and 70% of subjects with symptoms of severe asthma were not clinically diagnosed as having asthma. The daily use of inhaled corticosteroids (ICS) was less than 2.5% in subjects with current wheeze and those with symptoms of severe asthma but less than 1% used daily ICS when asthma remained undiagnosed. Conclusion: The prevalence of current wheeze and its causal factors showed a significant reduction compared to previous ISAAC studies. Among subjects with current wheeze and symptoms of severe asthma, the problem of under-diagnosis and under-treatment was widespread.

RESUMEN

Background: The Global Asthma Network phase I study in India aimed to study the prevalence, time trends, and associated risk factors of allergic rhinitis and eczema among 6-7-year-old, and 13-14-year-old school children and their parents. Objectives: The objective of the study was to understand the current prevalence and associated risk factors of Allergic Rhinitis and Eczema in India among 6-7-year-olds, 13-14-year-olds and in their parents/guardians for newer directions to health care providers, policy makers and academicians. Methods: Cross-sectional, multicenter study using self- and parent-administered questionnaire among randomly selected school children aged 6 to 7 years from 8 centers and 13 to 14 years from 9 centers and their respective parents/guardians across India. Results: Prevalence of allergic rhinitis (AR) (%, 95% CI) among 20,084 6-7-year-olds (children), 25,887 13-14-year-olds (adolescents), and 81,296 adults/parents was 7.7% (7.4%-8.1%), 23.5% (23.0%-24.1%), and 9.8% (9.55%-9.96%) and that of eczema was 2.5% (2.3%-2.7%), 3.5% (3.27%-3.71%), and 9.9% (9.7%-10.1%), respectively. Among 6-7-year-olds, AR and eczema showed a significantly (P < .001) declining time trend since the International Study of Asthma and Allergies in school children phase III in 2002-2003. Among 13-14-year-olds, AR (P < .01) but not eczema showed a significant temporal decline. Coexisting atopic condition, parental history of atopy, and some environmental factors consistent with previous studies were significant risk factors among children and adolescents. AR or eczema in either parent strongly predicted the same atopic condition among their adolescent offspring. Among adults, coexisting atopic condition was the strongest predictor of either AR or eczema. Conclusions: There is a slight declining time trend of AR and eczema in India over 2 decades among children and adolescents. Nearly 10% of Indian adults suffer from AR and eczema. Although genetic factors had the strongest association for AR and eczema among all age groups, certain early-life and environmental exposures need consideration to devise preventative strategies.

RESUMEN

OBJECTIVE: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of corona virus-induced disease 19 (COVID-19) that was declared as a global pandemic in March 2020 by the world health organization (WHO). Two vaccines were granted for emergency use by the Central Drugs Standard Control Organization (CDSCO) in India, Covishield® (AstraZeneca's vaccine manufactured by Serum Institute of India) and Covaxin® (manufactured by Bharat Biotech Limited). Sputnik - V has been granted EUA in the month of April 2021. The purpose of this study is to determine the association of COVID-19 infection, its severity and outcome in COVID-19 vaccinated people. METHODS: This was a hospital based prospective cohort study done between March to June 2021 at PBM Associated Group of Hospitals (AGH), Bikaner, Raj. Total 1028 COVID suspected cases consulted in COVID OPD or hospitalized under department of medicine, out of which 146 satisfied the inclusion and exclusion criteria, out of these 146, first 100 cases who gave consent for part of study were selected. RESULTS: Among 100 COVID-19 infected cases, 49 received first dose while rest got both doses. After first dose of vaccination 42.86% had mild and 32.65% had severe clinical infection while after both doses 80.39% had mild and 11.76% had severe clinical infection. On evaluation of HRCT Chest, after first dose 8.16% had normal 40.82% were in severe category while those who got both doses it was 52.82% 3.92% respectively. Among 49 who got first dose, 10.20% recovered on just home based treatment without any need of hospitalization, while 89.8% got admitted in dedicated COVID hospital out of which 73.47% got recovered and 16.33% died. Among 51 who got both the doses, 66.67% recovered on just home based treatment, while 33.33% required hospitalization out of which 25.49% got recovered and 7.84% died. CONCLUSION: After 2nd dose of vaccine there is a significant risk reduction in need of hospitalization and getting severe infection and mortality when compared with first dose only.

Asunto(s)

COVID-19 , Atención a la Salud , Humanos , India/epidemiología , Estudios Prospectivos , SARS-CoV-2

RESUMEN

Management of severe malaria remains a critical global challenge. In this study, using a multiplexed quantitative proteomics pipeline we systematically investigated the plasma proteome alterations in non-severe and severe malaria patients. We identified a few parasite proteins in severe malaria patients, which could be promising from a diagnostic perspective. Further, from host proteome analysis we observed substantial modulations in many crucial physiological pathways, including lipid metabolism, cytokine signaling, complement, and coagulation cascades in severe malaria. We propose that severe manifestations of malaria are possibly underpinned by modulations of the host physiology and defense machinery, which is evidently reflected in the plasma proteome alterations. Importantly, we identified multiple blood markers that can effectively define different complications of severe falciparum malaria, including cerebral syndromes and severe anemia. The ability of our identified blood markers to distinguish different severe complications of malaria may aid in developing new clinical tests for monitoring malaria severity.

Asunto(s)

Malaria Falciparum/diagnóstico , Malaria Falciparum/patología , Proteómica/métodos , Anemia/diagnóstico , Anemia/patología , Biomarcadores/sangre , Dengue/diagnóstico , Dengue/metabolismo , Dengue/patología , Humanos , Malaria Falciparum/metabolismo , Malaria Vivax/sangre , Malaria Vivax/metabolismo , Malaria Vivax/patología

RESUMEN

Plasmodium vivax malaria is a neglected disease, particularly during pregnancy. Severe vivax malaria is associated with inflammatory responses but in pregnancy immune alterations make it uncertain as to what cytokine signatures predominate, and how the type and quantity of blood immune mediators influence delivery outcomes. We measured the plasma concentrations of a set of thirty-one biomarkers, comprising cytokines, chemokines and growth factors, in 987 plasma samples from a cohort of 572 pregnant women from five malaria-endemic tropical countries and related these concentrations to delivery outcomes (birth weight and hemoglobin levels) and malaria infection. Samples were collected at recruitment (first antenatal visit) and at delivery (periphery, cord and placenta). At recruitment, we found that P. vivax-infected pregnant women had higher plasma concentrations of proinflammatory (IL-6, IL-1ß, CCL4, CCL2, CXCL10) and TH1-related cytokines (mainly IL-12) than uninfected women. This biomarker signature was essentially lost at delivery and was not associated with birth weight nor hemoglobin levels. Antiinflammatory cytokines (IL-10) were positively associated with infection and poor delivery outcomes. CCL11 was the only biomarker to show a negative association with P. vivax infection and its concentration at recruitment was positively associated with hemoglobin levels at delivery. Birth weight was negatively associated with peripheral IL-4 levels at delivery. Our multi-biomarker multicenter study is the first comprehensive one to characterize the immunological signature of P. vivax infection in pregnancy thus far. In conclusion, data show that while TH1 and pro-inflammatory responses are dominant during P. vivax infection in pregnancy, antiinflammatory cytokines may compensate excessive inflammation avoiding poor delivery outcomes, and skewness toward a TH2 response may trigger worse delivery outcomes. CCL11, a chemokine largely neglected in the field of malaria, emerges as an important marker of exposure or mediator in this condition.

Asunto(s)

Citocinas/sangre , Malaria Vivax/sangre , Plasmodium vivax/fisiología , Complicaciones Parasitarias del Embarazo/sangre , Adolescente , Adulto , Estudios de Cohortes , Femenino , Humanos , Recién Nacido , Interleucina-10/sangre , Interleucina-1beta/sangre , Malaria Vivax/inmunología , Malaria Vivax/parasitología , Malaria Vivax/fisiopatología , Masculino , Embarazo , Complicaciones Parasitarias del Embarazo/inmunología , Complicaciones Parasitarias del Embarazo/parasitología , Complicaciones Parasitarias del Embarazo/fisiopatología , Resultado del Embarazo , Células Th2/inmunología , Adulto Joven

RESUMEN

Sphingosine-1-phosphate (S1P), a bioactive lipid mediator is involved in an array of biological processes and linked to pathological manifestations. Erythrocyte is known as the major reservoir for S1P as they lack S1P-degrading enzymes (S1P lyase and S1P phosphohydrolase) and harbor sphingosine kinase-1 (SphK-1) essential for sphingosine conversion to S1P. Reduced S1P concentration in serum was correlated with disease severity in patients with Plasmodium falciparum and Plasmodium vivax infections. Herein, we aimed to identify the underlying mechanism and contribution of host erythrocytes toward depleted S1P levels in Plasmodium-infected patients vs. healthy individuals. The level and activity of SphK-1 were measured in vitro in both uninfected and cultured P. falciparum-infected erythrocytes. Infected erythrocytes demonstrated a significant decrease in SphK-1 level in a time-dependent manner. We found that 10-42 h post invasion (hpi), SphK1 level was predominantly reduced to ∼50% in rings, trophozoites, and schizonts compared to uninfected erythrocytes. We next analyzed the phosphorylation status of SphK-1, a modification responsible for its activity and S1P production, in both uninfected control and Plasmodium-infected erythrocytes. Almost ∼50% decrease in phosphorylation of SphK-1 was observed that could be corroborated with significant reduction in the production and release of S1P in infected erythrocytes. Serum S1P levels were studied in parallel in P. falciparum (N = 15), P. vivax (N = 36)-infected patients, and healthy controls (N = 6). The findings revealed that S1P concentration was significantly depleted in uncomplicated malaria cases and was found to be lowest in complicated malaria and thrombocytopenia in both P. falciparum and P. vivax-infected groups (∗∗ p < 0.01). The lower serum S1P level could be correlated with the reduced platelet count defining the role of S1P level in platelet formation. In conclusion, erythrocyte SphK-1 and S1P levels were studied in Plasmodium-infected individuals and erythrocytes that helped in characterizing the complications associated with malaria and thrombocytopenia, providing insights into the contribution of host erythrocyte biology in malaria pathogenesis. Finally, this study proposes the use of S1P and its analog as a novel adjunct therapy for malaria complications.

RESUMEN

The immune status of women changes during and after pregnancy, differs between blood compartments at delivery and is affected by environmental factors particularly in tropical areas endemic for multiple infections. We quantified the plasma concentration of a set of thirty-one TH1, TH2, TH17 and regulatory cytokines, pro-inflammatory and anti-inflammatory cytokines and chemokines, and growth factors (altogether biomarkers), in a cohort of 540 pregnant women from five malaria-endemic tropical countries. Samples were collected at recruitment (first antenatal visit), delivery (periphery, cord and placenta) and postpartum, allowing a longitudinal analysis. We found the lowest concentration of biomarkers at recruitment and the highest at postpartum, with few exceptions. Among them, IL-6, HGF and TGF-ß had the highest levels at delivery, and even higher concentrations in the placenta compared to peripheral blood. Placental concentrations were generally higher than peripheral, except for eotaxin that was lower. We also compared plasma biomarker concentrations between the tropical cohort and a control group from Spain at delivery, presenting overall higher biomarker levels the tropical cohort, particularly pro-inflammatory cytokines and growth factors. Only IL-6 presented lower levels in the tropical group. Moreover, a principal component analysis of biomarker concentrations at delivery showed that women from Spain grouped more homogenously, and that IL-6 and IL-8 clustered together in the tropical cohort but not in the Spanish one. Plasma cytokine concentrations correlated with Plasmodium antibody levels at postpartum but not during pregnancy. This basal profiling of immune mediators over gestation and in different compartments at delivery is important to subsequently understand response to infections and clinical outcomes in mothers and infants in tropical areas.

Asunto(s)

Quimiocinas/sangre , Citocinas/sangre , Péptidos y Proteínas de Señalización Intercelular/sangre , Malaria/sangre , Malaria/inmunología , Plasmodium/inmunología , Complicaciones Parasitarias del Embarazo/sangre , Adulto , Brasil/epidemiología , Estudios de Cohortes , Colombia/epidemiología , Femenino , Guatemala/epidemiología , Factor de Crecimiento de Hepatocito/sangre , Humanos , Inmunoglobulina G/inmunología , India/epidemiología , Interleucina-6/sangre , Interleucina-8/sangre , Malaria/parasitología , Papúa Nueva Guinea/epidemiología , Placenta/metabolismo , Embarazo , Mujeres Embarazadas , España , Factor de Crecimiento Transformador beta/sangre

RESUMEN

Malaria and dengue have overlapping clinical symptoms and are prevalent in the same geographic region (tropical and subtropical), hence precise diagnosis is challenging. The high mortality rate associated with both malaria and dengue could be attributed to "false", "delayed", or "missed" diagnosis. The present study thus aims to stratify malaria and dengue using Raman spectroscopy (RS). In total, 130 human sera were analyzed for model development and double-blinded testing. Principal components linear discriminant analysis (PC-LDA) of acquired RS-spectra could classify malaria and dengue with a minor overlap of 16.7%. Receiver operating characteristic (ROC) analysis of test samples showed sensitivity/specificity of 0.9529 for malaria vs healthy controls (HC) and 0.9584 for dengue vs HC. The Raman findings were complemented by mass spectroscopy (MS)-based metabolite analysis of 8 individuals, each from malaria, dengue, and HC. Several of the metabolites, including amino acids, cell-free DNA, creatinine, and bilirubin, assigned for the predominant RS-bands were also identified by MS and showed similar trends. Our data clearly indicates that RS-based serum analysis using a microprobe has immense potential for early, accurate, and automated detection and discrimination of malaria and dengue, and in the future, it could be extrapolated in field-settings combined with hand-held RS. Further, this approach might be extended to diagnose other closely related infections with similar clinical manifestations.

Asunto(s)

Dengue/diagnóstico , Malaria/diagnóstico , Espectrometría Raman/métodos , Adolescente , Adulto , Aminoácidos/sangre , Dengue/sangre , Femenino , Humanos , Malaria/sangre , Masculino , Metabolómica/métodos , Análisis de Componente Principal , Curva ROC , Sensibilidad y Especificidad , Adulto Joven

RESUMEN

Iron-sulfur (Fe-S) clusters are critical metallo-cofactors required for cell function. Assembly of these cofactors is a carefully controlled process in cells to avoid toxicity from free iron and sulfide. In Plasmodium, two pathways for these Fe-S cluster biogenesis have been reported; ISC pathway in the mitochondria and SUF pathway functional in the apicoplast. Amongst these, SUF pathway is reported essential for the apicoplast maintenance and parasite survival. Many of its components have been studied from P. falciparum and P. berghei in recent years, still few queries remain to be addressed; one of them being the assembly and transfer of Fe-S clusters. In this study, using P. vivax clinical isolates, we have shown the in vitro interaction of SUF pathway proteins SufS and SufE responsible for sulfur mobilization in the apicoplast. The sulfur mobilized by the SufSE complex assembles on the scaffold protein PvSufA along with iron provided by the external source. Here, we demonstrate in vitro transfer of these labile Fe-S clusters from the scaffold protein on to an apo-protein, PvIspG (a protein involved in penultimate step of Isoprenoids biosynthesis pathway) in order to provide an insight into the interaction of different components for the biosynthesis and transfer of Fe-S clusters. Our analysis indicate that inspite of the presence of variations in pathway proteins, the overall pathway remains well conserved in the clinical isolates when compared to that reported in lab strains.

Asunto(s)

Hierro/metabolismo , Plasmodium vivax/metabolismo , Azufre/metabolismo , Secuencia de Aminoácidos , Liasas de Carbono-Azufre/química , Liasas de Carbono-Azufre/genética , Liasas de Carbono-Azufre/metabolismo , Cicloserina/farmacología , Humanos , Hierro/química , Malaria Vivax/parasitología , Estructura Molecular , Fijación del Nitrógeno , Espectroscopía de Fotoelectrones , Plasmodium vivax/genética , Fosfato de Piridoxal/metabolismo , ARN Protozoario/aislamiento & purificación , Alineación de Secuencia , Azufre/química

RESUMEN

Biosynthesis of isoprenoids (MEP Pathway) in apicoplast has an important role during the erythrocytic stages of Plasmodium, as it is the sole pathway to provide the major isoprene units required as metabolic precursor for various housekeeping activities. With the intensifying need to identify a novel therapeutic drug target against Plasmodium, the MEP pathway and its components are considered as potential therapeutic targets, due to the difference in the isoprenoid synthesis route (MVA) functional in the host cells. While few major components have already been studied from this pathway for their potential as a drug target, IspD (2-C-methyl-D-erythritol-4-phosphate cytidyltransferase) enzyme, the enzyme catalyzing the third step of the pathway has only been tested against a synthetic compound from Malaria box called MMV008138, which also has not shown adequate inhibitory activity against P. vivax IspD. In the present study, to validate the potential of PvIspD as a drug target, various antimicrobial agents were screened for their inhibition possibilities, using in-vitro High Throughput Screening (HTS) technique. Shortlisted antimicrobial drug molecules like Cefepime, Tunicamycin and Rifampicin were further validated by in-vitro biochemical enzyme inhibition assays where they showed activity at nanomolar concentrations suggesting them or their derivatives as prospective future antimalarials. This study also confirmed the in-vivo expression of PvIspD protein during asexual stages by sub-cellular localization in apicoplast and explores the importance of the IspD enzyme in the development of new therapeutics.

Asunto(s)

Antimaláricos/uso terapéutico , Inhibidores Enzimáticos/uso terapéutico , Malaria Vivax/tratamiento farmacológico , Terapia Molecular Dirigida , Nucleotidiltransferasas/antagonistas & inhibidores , Plasmodium vivax/efectos de los fármacos , Secuencia de Aminoácidos , Inhibidores Enzimáticos/farmacología , Eritritol/análogos & derivados , Eritritol/química , Eritritol/farmacología , Humanos , Modelos Moleculares , Simulación de Dinámica Molecular , Nucleotidiltransferasas/química , Nucleotidiltransferasas/genética , Nucleotidiltransferasas/metabolismo , Filogenia , Plasmodium vivax/enzimología , Alineación de Secuencia , Fosfatos de Azúcar/química , Fosfatos de Azúcar/farmacología

RESUMEN

SCOPE: Haptoglobin (Hp), an acute phase inflammatory protein is associated with malaria pathogenesis in several proteomics and genomics studies. The Hp gene has two co-dominant alleles: Hp1 and Hp2 that produce three genotypes: Hp1/Hp1, Hp1/Hp2 and Hp2/Hp2. EXPERIMENTAL DESIGN: In this study, validation of the proteomics data with Multiple Reaction Monitoring Mass Spectroscopy (MRM-MS) is performed and the association of the Hp gene variants with severe, non-severe malaria and community (healthy) controls using genotyping PCRs and DNA sequencing is analysed. RESULTS: Highly significant values of Hp is observed in the MRM assay that show a correlation with severity of malaria and is clearly distinguished from another febrile disease, dengue. Moreover, the Hp2/Hp2 genotype is seen in high percentages in non-severe malaria patients (74%) and community controls (72%) whereas patients diagnosed with severe malaria show only (31%) of this genotype. Sequencing of the Hp promoter region reveals three SNPs along with 10 unique haplotypes, out of which five are associated with non-severe and three with severe malaria populations (χ2  = 130; df = 18; p < 0.0001). CONCLUSION AND CLINICAL RELEVANCE: This proteo-genomic study focuses on the correlation of the Hp protein and gene with malaria, thus highlighting the pivotal role of this acute phase immune gene in malaria pathogenesis.

Asunto(s)

Biomarcadores/sangre , Haptoglobinas/metabolismo , Malaria/sangre , Polimorfismo de Nucleótido Simple , Proteogenómica/métodos , Índice de Severidad de la Enfermedad , Adolescente , Adulto , Anciano , Estudios de Casos y Controles , Femenino , Genotipo , Haptoglobinas/genética , Humanos , India/epidemiología , Malaria/epidemiología , Malaria/parasitología , Masculino , Persona de Mediana Edad , Plasmodium falciparum/aislamiento & purificación , Adulto Joven

RESUMEN

Dengue fever (DF) is a major global health burden with a pathophysiology that is still incompletely understood. Biomarkers that predict and explain susceptibility to DF and its progression to its more severe hemorrhagic form are much needed. DF is endemic in tropical and subtropical regions of the world, with a rapidly increasing incidence of disease severity. We conducted a clinical biomarker discovery study using both a case-control and longitudinal study design. Plasma proteome alterations in patients with DF (n = 12) and dengue hemorrhagic fever (DHF, n = 24) were analyzed in comparison to healthy controls (HCs, n = 16), using the isobaric tags for relative and absolute quantification (iTRAQ)-based quantitative proteomics methodology (false discovery rate of 1%, ≥2 peptides). Several proteins such as the alpha-2 macroglobulin, angiotensinogen, apolipoprotein B-100, serotransferrin, and ceruloplasmin were upregulated (fold change >1.2) in all DHF cases, and downregulated in DF (fold change <0.83), compared with HCs. Plasma cytokine profiling (8 DF, 8 DHF, and 8 HC) on two consecutive time points, at day 0 (day of admission) and days 5-7, found significant elevation in IL-1RA, IL-7, TNF-α, MCP1-MCAF, and MIP-1ß levels, but only in the DHF cases, which is the severe disease, and not in DF, compared with HCs (p < 0.05). These new observations on changes in the plasma proteome and cytokine profiles in patients with dengue infection identify several putative molecular leads for future biomarker development and precision medicine in relation to forecasting DF disease severity.

Asunto(s)

Biomarcadores/sangre , Citocinas/sangre , Dengue/diagnóstico , Proteómica/métodos , Índice de Severidad de la Enfermedad , Adolescente , Adulto , Estudios de Casos y Controles , Dengue/sangre , Enfermedades Endémicas , Femenino , Humanos , Incidencia , Estudios Longitudinales , Masculino , Persona de Mediana Edad , Medicina de Precisión , Factores de Tiempo

Asunto(s)

Malaria Cerebral/parasitología , Malaria Vivax/parasitología , Plasmodium vivax/aislamiento & purificación , Adolescente , Humanos , Malaria Cerebral/diagnóstico , Malaria Vivax/diagnóstico , Masculino , Plasmodium vivax/genética , Plasmodium vivax/fisiología

RESUMEN

BACKGROUND: Despite that over 90 million pregnancies are at risk of Plasmodium vivax infection annually, little is known about the epidemiology and impact of the infection in pregnancy. METHODOLOGY AND PRINCIPAL FINDINGS: We undertook a health facility-based prospective observational study in pregnant women from Guatemala (GT), Colombia (CO), Brazil (BR), India (IN) and Papua New Guinea PNG). Malaria and anemia were determined during pregnancy and fetal outcomes assessed at delivery. A total of 9388 women were enrolled at antennal care (ANC), of whom 53% (4957) were followed until delivery. Prevalence of P. vivax monoinfection in maternal blood at delivery was 0.4% (20/4461) by microscopy [GT 0.1%, CO 0.5%, BR 0.1%, IN 0.2%, PNG 1.2%] and 7% (104/1488) by PCR. P. falciparum monoinfection was found in 0.5% (22/4463) of women by microscopy [GT 0%, CO 0.5%, BR 0%, IN 0%, PNG 2%]. P. vivax infection was observed in 0.4% (14/3725) of placentas examined by microscopy and in 3.7% (19/508) by PCR. P. vivax in newborn blood was detected in 0.02% (1/4302) of samples examined by microscopy [in cord blood; 0.05% (2/4040) by microscopy, and 2.6% (13/497) by PCR]. Clinical P. vivax infection was associated with increased risk of maternal anemia (Odds Ratio-OR, 5.48, [95% CI 1.83-16.41]; p = 0.009), while submicroscopic vivax infection was not associated with increased risk of moderate-severe anemia (Hb<8g/dL) (OR, 1.16, [95% CI 0.52-2.59]; p = 0.717), or low birth weight (<2500g) (OR, 0.52, [95% CI, 0.23-1.16]; p = 0.110). CONCLUSIONS: In this multicenter study, the prevalence of P. vivax infection in pregnancy by microscopy was overall low across all endemic study sites; however, molecular methods revealed a significant number of submicroscopic infections. Clinical vivax infection in pregnancy was associated with maternal anemia, which may be deleterious for infant's health. These results may help to guide maternal health programs in settings where vivax malaria is endemic; they also highlight the need of addressing a vulnerable population such as pregnant women while embracing malaria elimination in endemic countries.

Asunto(s)

Malaria Vivax/complicaciones , Plasmodium vivax , Complicaciones Parasitarias del Embarazo/patología , Adolescente , Adulto , Brasil/epidemiología , Colombia/epidemiología , Femenino , Sangre Fetal , Guatemala/epidemiología , Humanos , India/epidemiología , Recién Nacido , Transmisión Vertical de Enfermedad Infecciosa , Malaria Vivax/epidemiología , Papúa Nueva Guinea/epidemiología , Embarazo , Complicaciones Parasitarias del Embarazo/epidemiología , Adulto Joven

RESUMEN

A vaccine to eliminate malaria would need a multi-stage and multi-species composition to achieve robust protection, but the lack of knowledge about antigen targets and mechanisms of protection precludes the development of fully efficacious malaria vaccines, especially for Plasmodium vivax (Pv). Pregnant women constitute a risk population who would greatly benefit from a vaccine preventing the adverse events of Plasmodium infection during gestation. We hypothesized that functional immune responses against putative targets of naturally acquired immunity to malaria and vaccine candidates will be associated with protection against malaria infection and/or poor outcomes during pregnancy. We measured (i) IgG responses to a large panel of Pv and Plasmodium falciparum (Pf) antigens, (ii) the capacity of anti-Pv ligand Duffy binding protein (PvDBP) antibodies to inhibit binding to Duffy antigen, and (iii) cellular immune responses to two Pv antigens, in a subset of 1,056 pregnant women from Brazil, Colombia, Guatemala, India, and Papua New Guinea (PNG). There were significant intraspecies and interspecies correlations for most antibody responses (e.g., PfMSP119 versus PfAMA1, Spearman's rho = 0.81). Women from PNG and Colombia had the highest levels of IgG overall. Submicroscopic infections seemed sufficient to boost antibody responses in Guatemala but not antigen-specific cellular responses in PNG. Brazil had the highest percentage of Duffy binding inhibition (p-values versus Colombia: 0.040; Guatemala: 0.047; India: 0.003, and PNG: 0.153) despite having low anti-PvDBP IgG levels. Almost all antibodies had a positive association with present infection, and coinfection with the other species increased this association. Anti-PvDBP, anti-PfMSP1, and anti-PfAMA1 IgG levels at recruitment were positively associated with infection at delivery (p-values: 0.010, 0.003, and 0.023, respectively), suggesting that they are markers of malaria exposure. Peripheral blood mononuclear cells from Pv-infected women presented fewer CD8+IFN-γ+ T cells and secreted more G-CSF and IL-4 independently of the stimulus used in vitro. Functional anti-PvDBP levels at recruitment had a positive association with birth weight (difference per doubling antibody levels: 45 g, p-value: 0.046). Thus, naturally acquired binding-inhibitory antibodies to PvDBP might confer protection against poor outcomes of Pv malaria in pregnancy.

RESUMEN

The prokaryotic type Methyl Erythritol phosphate (MEP) pathway functional in the apicoplast of Plasmodium is indispensable for the erythrocytic stages of the parasite. It is the sole process of isoprenoids biosynthesis in the parasite and is different from that in humans. Among the seven enzymes known to be functional in the MEP pathway in prokaryotes, most enzymes from Plasmodium are yet uncharacterized. The penultimate enzyme of this pathway 4-hydroxy-3-methylbut-2-en-1-yl diphosphate synthase (IspG), has been shown to act as a key target molecule in prokaryotes, where its deletion results in impairment of many housekeeping functions. The present study is the first detailed report of IspG enzyme from any Plasmodium sp. We report here that the protein is highly conserved across apicomplexans and prokaryotes and it localizes to the apicoplast as evident from the immune-localization studies performed on P. vivax infected blood smears made from clinical patients. The biochemical reconstitution and in silico docking of [4Fe-4S] clusters on the protein indicate their importance for the activity of enzyme. In-silico screening of different drug entities suggested the inhibitory role of alkyne diphosphate analogues and fosmidomycin against the IspG enzyme, suggesting the potential role of this enzyme as an antimalarial target.

Asunto(s)

Transferasas Alquil y Aril/metabolismo , Antimaláricos/farmacología , Terapia Molecular Dirigida , Plasmodium vivax/efectos de los fármacos , Plasmodium vivax/enzimología , Transferasas Alquil y Aril/química , Antimaláricos/metabolismo , Secuencia Conservada , Humanos , Hierro/metabolismo , Simulación del Acoplamiento Molecular , Dominios Proteicos , Análisis de Secuencia , Azufre/metabolismo

RESUMEN

P. vivax infection during pregnancy has been associated with poor outcomes such as anemia, low birth weight and congenital malaria, thus representing an important global health problem. However, no vaccine is currently available for its prevention. Vir genes were the first putative virulent factors associated with P. vivax infections, yet very few studies have examined their potential role as targets of immunity. We investigated the immunogenic properties of five VIR proteins and two long synthetic peptides containing conserved VIR sequences (PvLP1 and PvLP2) in the context of the PregVax cohort study including women from five malaria endemic countries: Brazil, Colombia, Guatemala, India and Papua New Guinea (PNG) at different timepoints during and after pregnancy. Antibody responses against all antigens were detected in all populations, with PNG women presenting the highest levels overall. P. vivax infection at sample collection time was positively associated with antibody levels against PvLP1 (fold-increase: 1.60 at recruitment -first antenatal visit-) and PvLP2 (fold-increase: 1.63 at delivery), and P. falciparum co-infection was found to increase those responses (for PvLP1 at recruitment, fold-increase: 2.25). Levels of IgG against two VIR proteins at delivery were associated with higher birth weight (27 g increase per duplicating antibody levels, p<0.05). Peripheral blood mononuclear cells from PNG uninfected pregnant women had significantly higher antigen-specific IFN-γ TH1 responses (p=0.006) and secreted less pro-inflammatory cytokines TNF and IL-6 after PvLP2 stimulation than P. vivax-infected women (p<0.05). These data demonstrate that VIR antigens induce the natural acquisition of antibody and T cell memory responses that might be important in immunity to P. vivax during pregnancy in very diverse geographical settings.

Asunto(s)

Anticuerpos Antiprotozoarios/sangre , Antígenos de Protozoos/inmunología , Inmunoglobulina G/sangre , Malaria Vivax/inmunología , Plasmodium vivax/inmunología , Complicaciones Infecciosas del Embarazo/inmunología , Células TH1/inmunología , Adulto , Peso al Nacer , Brasil/epidemiología , Estudios de Cohortes , Coinfección/inmunología , Coinfección/parasitología , Colombia/epidemiología , Citocinas/metabolismo , Enfermedades Endémicas , Femenino , Guatemala/epidemiología , Humanos , Memoria Inmunológica , India/epidemiología , Interferón gamma/metabolismo , Leucocitos Mononucleares/inmunología , Malaria Falciparum/inmunología , Malaria Vivax/epidemiología , Papúa Nueva Guinea/epidemiología , Plasmodium falciparum/genética , Plasmodium falciparum/inmunología , Plasmodium vivax/genética , Plasmodium vivax/patogenicidad , Embarazo , Complicaciones Infecciosas del Embarazo/epidemiología , Proteínas Protozoarias/genética , Proteínas Protozoarias/inmunología , Proteínas Protozoarias/aislamiento & purificación

RESUMEN

High density oligonucleotide microarrays have been used on Plasmodium vivax field isolates to estimate whole genome expression. However, no microarray platform has been experimentally optimized for studying the transcriptome of field isolates. In the present study, we adopted both bioinformatics and experimental testing approaches to select best optimized probes suitable for detecting parasite transcripts from field samples and included them in designing a custom 15K P. vivax microarray. This microarray has long oligonucleotide probes (60mer) that were in-situ synthesized onto glass slides using Agilent SurePrint technology and has been developed into an 8X15K format (8 identical arrays on a single slide). Probes in this array were experimentally validated and represents 4180 P. vivax genes in sense orientation, of which 1219 genes have also probes in antisense orientation. Validation of the 15K array by using field samples (n=14) has shown 99% of parasite transcript detection from any of the samples. Correlation analysis between duplicate probes (n=85) present in the arrays showed perfect correlation (r2=0.98) indicating the reproducibility. Multiple probes representing the same gene exhibited similar kind of expression pattern across the samples (positive correlation, r≥0.6). Comparison of hybridization data with the previous studies and quantitative real-time PCR experiments were performed to highlight the microarray validation procedure. This array is unique in its design, and results indicate that the array is sensitive and reproducible. Hence, this microarray could be a valuable functional genomics tool to generate reliable expression data from P. vivax field isolates.

Asunto(s)

Perfilación de la Expresión Génica/instrumentación , Malaria Vivax/genética , Análisis de Secuencia por Matrices de Oligonucleótidos/instrumentación , Plasmodium vivax/genética , Humanos , Reacción en Cadena en Tiempo Real de la Polimerasa , Reproducibilidad de los Resultados