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Despite having the highest risk of progressing to severe disease due to lack of acquired immunity, the youngest children living in areas of highly intense malaria transmission have long been observed to be infected at lower rates than older children. Whether this observation is due to reduced exposure to infectious mosquito bites from behavioral and biological factors, maternally transferred immunity, genetic factors, or enhanced innate immunity in the young child has intrigued malaria researchers for over half a century. Recent evidence suggests that maternally transferred immunity may be limited to early infancy and that the young child's own immune system may contribute to control of malarial symptoms early in life and prior to the development of more effective adaptive immunity. Prospective studies of active and passive detection of Plasmodium falciparum blood-stage infections have identified young children (<5 years old) who remain uninfected through a defined surveillance period despite living in settings of highly intense malaria transmission. Yet, little is known about the potential immunological basis for this 'aparasitemic' phenotype. In this review, we summarize the observational evidence for this phenotype in field studies and examine potential reasons why these children escape detection of parasitemia, covering factors that are either extrinsic or intrinsic to their developing immune system. We discuss the challenges of distinguishing malaria protection from lack of malaria exposure in field studies. We also identify gaps in our knowledge regarding cellular immunity in the youngest age group and propose directions that researchers may take to address these gaps.
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Malária Falciparum , Parasitemia , Plasmodium falciparum , Humanos , Pré-Escolar , Malária Falciparum/transmissão , Plasmodium falciparum/imunologia , Lactente , Malária/transmissão , Imunidade Inata , AnimaisRESUMO
A systems analysis was conducted to determine the potential molecular mechanisms underlying differential immunogenicity and protective efficacy results of a clinical trial of the radiation-attenuated whole-sporozoite PfSPZ vaccine in African infants. Innate immune activation and myeloid signatures at prevaccination baseline correlated with protection from P. falciparum parasitemia in placebo controls. These same signatures were associated with susceptibility to parasitemia among infants who received the highest and most protective PfSPZ vaccine dose. Machine learning identified spliceosome, proteosome, and resting DC signatures as prevaccination features predictive of protection after highest-dose PfSPZ vaccination, whereas baseline circumsporozoite protein-specific (CSP-specific) IgG predicted nonprotection. Prevaccination innate inflammatory and myeloid signatures were associated with higher sporozoite-specific IgG Ab response but undetectable PfSPZ-specific CD8+ T cell responses after vaccination. Consistent with these human data, innate stimulation in vivo conferred protection against infection by sporozoite injection in malaria-naive mice while diminishing the CD8+ T cell response to radiation-attenuated sporozoites. These data suggest a dichotomous role of innate stimulation for malaria protection and induction of protective immunity by whole-sporozoite malaria vaccines. The uncoupling of vaccine-induced protective immunity achieved by Abs from more protective CD8+ T cell responses suggests that PfSPZ vaccine efficacy in malaria-endemic settings may be constrained by opposing antigen presentation pathways.
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Imunidade Inata , Vacinas Antimaláricas , Malária Falciparum , Plasmodium falciparum , Esporozoítos , Vacinas Atenuadas , Vacinas Antimaláricas/imunologia , Vacinas Antimaláricas/administração & dosagem , Imunidade Inata/imunologia , Humanos , Animais , Malária Falciparum/prevenção & controle , Malária Falciparum/imunologia , Plasmodium falciparum/imunologia , Camundongos , Vacinas Atenuadas/imunologia , Vacinas Atenuadas/administração & dosagem , Esporozoítos/imunologia , Esporozoítos/efeitos da radiação , Linfócitos T CD8-Positivos/imunologia , Lactente , Proteínas de Protozoários/imunologia , Anticorpos Antiprotozoários/imunologia , Feminino , Parasitemia/imunologia , Parasitemia/prevenção & controle , Imunoglobulina G/imunologia , Imunoglobulina G/sangue , Eficácia de VacinasRESUMO
Malaria and schistosomiasis are two major parasitic diseases that remain leading causes of morbidity and mortality worldwide. Co-infections of these two parasites are common in the tropics, where both diseases are endemic. The clinical consequences of schistosomiasis and malaria are determined by a variety of host, parasitic, and environmental variables. Chronic schistosomiasis causes malnutrition and cognitive impairments in children, while malaria can cause fatal acute infections. There are effective drugs available to treat malaria and schistosomiasis. However, the occurrence of allelic polymorphisms and the rapid selection of parasites with genetic mutations can confer reduced susceptibility and lead to the emergence of drug resistance. Moreover, the successful elimination and complete management of these parasites are difficult due to the lack of effective vaccines against Plasmodium and Schistosoma infections. Therefore, it is important to highlight all current vaccine candidates undergoing clinical trials, such as pre-erythrocytic and erythrocytic stage malaria, as well as a next-generation RTS,S-like vaccine, the R21/Matrix-M vaccine, that conferred 77% protection against clinical malaria in a Phase 2b trial. Moreover, this review also discusses the progress and development of schistosomiasis vaccines. Furthermore, significant information is provided through this review on the effectiveness and progress of schistosomiasis vaccines currently under clinical trials, such as Sh28GST, Sm-14, and Sm-p80. Overall, this review provides insights into recent progress in malarial and schistosomiasis vaccines and their developmental approaches.
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BACKGROUND: TP53 has been shown to play a role in inflammatory processes, including malaria. We previously found that p53 attenuates parasite-induced inflammation and predicts clinical protection to Plasmodium falciparum infection in Malian children. Here, we investigated whether p53 codon 47 and 72 polymorphisms are associated with differential risk of P. falciparum infection and uncomplicated malaria in a prospective cohort study of malaria immunity. METHODS: p53 codon 47 and 72 polymorphisms were determined by sequencing TP53 exon 4 in 631 Malian children and adults enrolled in the Kalifabougou cohort study. The effects of these polymorphisms on the prospective risk of febrile malaria, incident parasitemia, and time to fever after incident parasitemia over 6 months of intense malaria transmission were assessed using Cox proportional hazards models. RESULTS: Confounders of malaria risk, including age and hemoglobin S or C, were similar between individuals with or without p53 S47 and R72 polymorphisms. Relative to their respective common variants, neither S47 nor R72 was associated with differences in prospective risk of febrile malaria, incident parasitemia, or febrile malaria after parasitemia. CONCLUSIONS: These findings indicate that p53 codon 47 and 72 polymorphisms are not associated with protection against incident P. falciparum parasitemia or uncomplicated febrile malaria.
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Malária Falciparum , Malária , Criança , Adulto , Humanos , Estudos de Coortes , Estudos Prospectivos , Parasitemia/genética , Proteína Supressora de Tumor p53/genética , Plasmodium falciparum/genética , Malária/complicações , Malária Falciparum/epidemiologia , Malária Falciparum/genética , Malária Falciparum/complicações , Febre/etiologiaRESUMO
Conventional airborne virus measurement usually requires appreciable sampling and detection times. Viral aerosols should also be collected or prepared in a liquid medium whose volume typically ranges from milliliters to tens of milliliters; hence, many sampling and detection steps need to be taken with the unit horizontal or immobile. Moreover, viral aerosols need to be sufficiently enriched, which makes real-time monitoring difficult. Herein, we present a near real-time enrichment and quantification system of airborne viruses that consists of a wet-paper-based electrochemical immunosensor with a gel electrolyte and a modified electrostatic particle concentrator. A small amount of phosphate-buffered saline flowed on the electrode, which resulted in sensor electrodes that are barely wet (covered in a thin buffer film measuring several micrometers) to ensure antigen-antibody interaction and the removal of non-target particles on the electrode surface. This system ensures that airborne viruses are highly enriched on the working electrode of the immunosensor, and it is possible to measure the MS2 virus particle concentrations every 10 min for 60 min stably and selectively against non-target airborne viruses and bacteria at horizontal and tilted measurement configurations. This system thus has the potential to be used in the real-time mobile monitoring of airborne microorganisms.
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Técnicas Biossensoriais , Vírus , Eletricidade Estática , Imunoensaio , Aerossóis , FosfatosRESUMO
Growth tube collectors (GTCs) are used to sample virus aerosols because of their superior viable virus recovery among air samplers. However, a major limitation of such samplers is that they operate at low flow rates compared to many inertia-based air samplers. Herein, we demonstrated efficient measurements of airborne MS2 and T3 viruses using a GTC that can implement high flow velocities for higher flow rates per tube, which we refer to as the growth-based virus aerosol concentrator (GVC), via qPCR and the plaque assay technique. The GVC exhibited a flow rate of up to 6 L/min, where the average sampling flow velocity was 5.09 m/s, 22 times higher than those used in the GTCs, for a single tube with a diameter of 5 mm. The count median diameter of the size-increased particles at the exit of the initiator was measured to be 1.44 µm at 6 L/min, considerably smaller than those observed in conventional GTCs. Nevertheless, the measurement of airborne MS2 and T3 viruses using the GVC showed a high concentration (high enrichment ratio of 109,458 at 10-min sampling) of viruses in a sampling medium, with a high viable virus percentage (> 90%) and physical collection efficiency (> 90%) at 6 L/min, which shows the potential for rapid on-site detection of airborne viruses.
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Microbiologia do Ar , Vírus , Aerossóis/análise , Monitoramento Ambiental/métodos , Tamanho da PartículaRESUMO
In the present study, we have investigated the role of antimalarial drug halofantrine (HF) in inducing the sterile protection against challenges with sporozoites of the live infectious Plasmodium yoelii (Killick-Kendrick, 1967) in Swiss mice malaria model. We observed that during the first to third sequential sporozoite inoculation cycles, blood-stage patency remains the same in the control and chemoprophylaxis under HF drug cover (CPS-HF) groups. However, a delayed blood-stage infection was observed during the fourth and fifth sporozoite challenges and complete sterile protection was produced following the sixth sporozoite challenge in CPS-HF mice. We also noticed a steady decline in liver stage parasite load after 3th to 6th sporozoite challenge cycle in CPS-HF mice. CPS-HF immunisation results in a significant up-regulation of pro-inflammatory cytokines (IFN-γ, TNF-α, IL-12 and iNOS) and down-regulation of anti-inflammatory cytokines (IL-10 and TGF-ß) mRNA expression in hepatic mononuclear cells (HMNC) and spleen cells in the immunised CPS-HF mice (after 6th sporozoite challenge) compared to control. Overall, our study suggests that the repetitive sporozoite inoculation under HF drug treatment develops a strong immune response that confers protection against subsequent challenges with sporozoites of P. yoelii.
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Malária , Preparações Farmacêuticas , Plasmodium yoelii , Animais , Quimioprevenção , Imunização , Malária/prevenção & controle , Camundongos , Camundongos Endogâmicos BALB C , FenantrenosRESUMO
Many studies have been conducted on measuring avian influenza viruses and their hemagglutinin (HA) antigens via electrochemical principles; most of these studies have used gold electrodes on ceramic, glass, or silicon substrates, and/or labeling for signal enhancement. Herein, we present a paper-based immunosensor for label-free measurement of multiple avian influenza virus (H5N1, H7N9, and H9N2) antigens using flexible screen-printed carbon nanotube-polydimethylsiloxane electrodes. These flexible electrodes on a paper substrate can complement the physical weakness of the paper-based sensors when wetted, without affecting flexibility. The relative standard deviation of the peak currents was 1.88% when the electrodes were repeatedly bent and unfolded twenty times with deionized water provided each cycle, showing the stability of the electrodes. For the detection of HA antigens, approximately 10-µl samples (concentration: 100 pg/ml-100 ng/ml) were needed to form the antigen-antibody complexes during 20-30 min incubation, and the immune responses were measured via differential pulse voltammetry. The limits of detections were 55.7 pg/ml (0.95 pM) for H5N1 HA, 99.6 pg/ml (1.69 pM) for H7N9 HA, and 54.0 pg/ml (0.72 pM) for H9N2 HA antigens in phosphate buffered saline, and the sensors showed good selectivity and reproducibility. Such paper-based sensors are economical, flexible, robust, and easy-to-manufacture, with the ability to detect several avian influenza viruses.
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Antígenos Virais/análise , Técnicas Biossensoriais/métodos , Dimetilpolisiloxanos , Técnicas Eletroquímicas/métodos , Eletrodos , Imunoensaio/métodos , Virus da Influenza A Subtipo H5N1/imunologia , Subtipo H7N9 do Vírus da Influenza A/imunologia , Vírus da Influenza A Subtipo H9N2/imunologia , Nanotubos de Carbono , Papel , Virologia/métodos , Animais , Aves , Humanos , Influenza Aviária/diagnóstico , Influenza Aviária/virologia , Influenza Humana/diagnóstico , Influenza Humana/virologia , Limite de Detecção , Reprodutibilidade dos TestesRESUMO
Malaria represents one of the major life-threatening diseases that poses a huge socio-economic impact, worldwide. Chemoprophylaxis vaccination using a relatively low number of wild-type infectious sporozoites represents an attractive and effective vaccine strategy against malaria. However, the role of immune responses to pre-erythrocytic versus blood-stage parasites in protection against different antimalarial drugs remains unclear. Here, in the present study, we explored the immune responses against the repetitive inoculation of live Plasmodium yoelii (P. yoelii) sporozoites in an experimental Swiss mouse model under antimalarial drug lumefantrine chemoprophylaxis (CPS-LMF). We monitored the liver stage parasitic load, pro/anti-inflammatory cytokines expression, and erythrocytic stage patency, following repetitive cycles of sporozoites inoculations. It was found that repetitive sporozoites inoculation under CPS-LMF results in delayed blood-stage infection during the fourth sporozoites challenge, while sterile protection was produced in mice following the fifth cycle of sporozoites challenge. Intriguingly, we observed a significant up-regulation of pro-inflammatory cytokines (IFN-γ, TNF-α and IL-12) and iNOS response and down-regulation of anti-inflammatory cytokines (IL-4, IL-10 and TGF-ß) in the liver HMNC (hepatic mononuclear cells) and spleen cells after 4th and 5th cycle of sporozoites challenge in the CPS-LMF mice. Meanwhile, we also noticed that the liver stage parasites load under CPS-LMF immunization has gradually reduced after 2nd, 3rd, 4th and 5th sporozoites challenge. Overall, our study suggests that chemoprophylaxis vaccination under LMF drug cover develops strong immune responses and confer superior long-lasting protection against P. yoelii sporozoites. Furthermore, this vaccination strategy can be used to study the protective and stage-specific immunity against new protective antigens. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13205-021-03022-0.
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Air-transmissible pathogenic viruses, such as influenza viruses and coronaviruses, are some of the most fatal strains and spread rapidly by air, necessitating quick and stable measurements from sample air volumes to prevent further spread of diseases and to take appropriate steps rapidly. Measurements of airborne viruses generally require their collection into liquids or onto solid surfaces, with subsequent hydrosolization and then analysis using the growth method, nucleic-acid-based techniques, or immunoassays. Measurements can also be performed in real time without sampling, where species-specific determination is generally disabled. In this review, we introduce some recent advancements in the measurement of pathogenic airborne viruses. Air sampling and measurement technologies for viral aerosols are reviewed, with special focus on the effects of air sampling on damage to the sampled viruses and their measurements. Measurement of pathogenic airborne viruses is an interdisciplinary research area that requires understanding of both aerosol technology and biotechnology to effectively address the issues. Hence, this review is expected to provide some useful guidelines regarding appropriate air sampling and virus detection methods for particular applications.
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Microbiologia do Ar , Vírus , Aerossóis , Manejo de EspécimesRESUMO
Airborne influenza viruses are responsible for serious respiratory diseases, and most detection methods for airborne viruses are based on extraction of nucleic acids. Herein, vertical-flow-assay-based electrochemical paper immunosensors were fabricated to rapidly quantify the influenza H1N1 viruses in air after sampling with a portable electrostatic particle concentrator (EPC). The effects of antibodies, anti-influenza nucleoprotein antibodies (NP-Abs) and anti-influenza hemagglutinin antibodies (HA-Abs), on the paper sensors as well as nonpulsed high electrostatic fields with and without corona charging on the virus measurement were investigated. The antigenicity losses of the surface (HA) proteins were caused by H2O2 via lipid oxidation-derived radicals and 1O2 via direct protein peroxidation upon exposure of a high electrostatic field. However, minimal losses in antigenicity of NP of the influenza viruses were observed, and the concentration of the H1N1 viruses was more than 160 times higher in the EPC than the BioSampler upon using NP-Ab based paper sensors after 60 min collection. This NP-Ab-based paper sensors with the EPC provided measurements comparable to quantitative polymerase chain reaction (qPCR) but much quicker, specific to the influenza H1N1 viruses in the presence of other airborne microorganisms and beads, and more cost-effective than enzyme-linked immunosorbent assay and qPCR.
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Vírus da Influenza A Subtipo H1N1 , Influenza Humana , Ensaio de Imunoadsorção Enzimática , Humanos , Peróxido de Hidrogênio , Eletricidade EstáticaRESUMO
Acute myocardial infarction (AMI), also recognized as a "heart attack," is one leading cause of death globally, and cardiac myoglobin (cMb), an important cardiac biomarker, is used for the early assessment of AMI. This paper presents an ultrasensitive, label-free electrochemical aptamer-based sensor (aptasensor) for cMb detection using polyethylenimine (PEI)-functionalized reduced graphene oxide (PEI-rGO) thin films. PEI, a cationic polymer, was used as a reducing agent for graphene oxide (GO), providing highly positive charges on the rGO surface and allowing direct immobilization of negatively charged single-strand DNA aptamers against cMb via electrostatic interaction without any linker or coupling chemistry. The presence of cMb was detected on Mb aptamer-modified electrodes using differential pulse voltammetry via measuring the current change due to the direct electron transfer between the electrodes and cMb proteins (Fe3+/Fe2+). The limits of detection were 0.97 pg mL-1 (phosphate-buffered saline) and 2.1 pg mL-1 (10-fold-diluted human serum), with a linear behavior with logarithmic cMb concentration. The specificity and reproducibility of the aptasensors were also examined. This electrochemical aptasensor using polymer-modified rGO shows potential for the early assessment of cMb in point-of-care testing applications.
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Though the immunity to malaria has been associated with cellular immune responses, the exact function of the phenotypic cell population is still unclear. This study investigated the host immune responses elicited during the pre-erythrocytic stage, post-Plasmodium yoelii sporozoite infection in Swiss mice model. For this purpose, we analyzed the dynamics of different subsets of immune cells population and cytokine levels in the hepatic mononuclear and splenic cells population during pre-erythrocytic liver-stage infection. We observed a significant reduction in the effectors immune cells population including CD8+ T cell, F4/80+ macrophage and in plasmacytoid dendritic cells (CD11c+ B220+). Interestingly, substantial down-regulation was also noted in pro-inflammatory cytokines (i.e. IFN-γ, TNF-α, IL-12, IL-2, IL-17 and iNOS), while, up-regulation of anti-inflammatory cytokines (i.e. IL-10, IL-4 and TGF-ß) during asymptomatic pre-erythrocytic liver-stage infection. Collectively, this study demonstrated that during pre-erythrocytic development, Plasmodium yoelii sporozoite impaired the host activators of innate and adaptive immune responses by regulating the immune effector cells, gene expression and cytokines levels for the establishment of infection and subsequent development in the liver and spleen. The results in this study provided a better understanding of the events leading to malarial infection and will be helpful in supportive treatment and vaccine development strategy.
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Immunity that controls parasitemia and inflammation during Plasmodium falciparum (Pf) malaria can be acquired with repeated infections. A limited understanding of this complex immune response impedes the development of vaccines and adjunctive therapies. We conducted a prospective systems biology study of children who differed in their ability to control parasitemia and fever following Pf infection. By integrating whole-blood transcriptomics, flow-cytometric analysis, and plasma cytokine and antibody profiles, we demonstrate that a pre-infection signature of B cell enrichment, upregulation of T helper type 1 (Th1) and Th2 cell-associated pathways, including interferon responses, and p53 activation associated with control of malarial fever and coordinated with Pf-specific immunoglobulin G (IgG) and Fc receptor activation to control parasitemia. Our hypothesis-generating approach identified host molecules that may contribute to differential clinical outcomes during Pf infection. As a proof of concept, we have shown that enhanced p53 expression in monocytes attenuated Plasmodium-induced inflammation and predicted protection from fever.
Assuntos
Linfócitos B/imunologia , Proteínas Sanguíneas/metabolismo , Inflamação/metabolismo , Malária Falciparum/metabolismo , Plasmodium falciparum/fisiologia , Células Th1/imunologia , Células Th2/imunologia , Proteína Supressora de Tumor p53/metabolismo , Adolescente , Adulto , Animais , Anticorpos Antiprotozoários/metabolismo , Criança , Pré-Escolar , Resistência à Doença , Feminino , Perfilação da Expressão Gênica , Humanos , Lactente , Interferons/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Estudos Prospectivos , Receptores Fc/metabolismo , Transdução de Sinais , Proteína Supressora de Tumor p53/genética , Adulto JovemRESUMO
Rapid subtyping of influenza viruses in clinical laboratories has been increasingly important because three subtypes (seasonal H1N1, H3N2, and 2009 H1N1) of influenza A virus currently disseminated in humans have variable susceptibilities to antiviral drug. Herein, we present DNA aptamers for selective detection of influenza A H1N1 (seasonal and 2009 pandemic H1N1) viruses by targeting recombinant influenza A mini-hemagglutinin (mini-HA) protein (the stable stem region of HA) and whole H1N1 viruses. The dissociation constants (KD) of aptamer candidates V46 and V57 were 19.2â¯nM and 29.6â¯nM, respectively, according to electrochemical characterization (differential pulse voltammetry), demonstrating strong binding to mini-HA. In comparison, the KD of the influenza virus antibodies is in the range of 1⯵M-10â¯nM. Aptamer V46 showed higher specificity and binding affinity to the mini-HA protein and H1N1 subtypes, and it was also incorporated into an indium tin oxide-based electrochemical sensor, showing sensitive and specific detection of H1N1 viruses, with a limit of detection (LOD) of 3.7 plaque-forming units per mL. The binding affinity, specificity, and LOD achieved with the electrochemical sensor suggest that it can be used for rapid subtyping of H1N1. We also propose that this aptamer can be used for the neutralization of H1N1 subtypes, suggesting potential therapeutic and diagnostic applications.
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Técnicas Biossensoriais , Técnicas Eletroquímicas , Hemaglutininas Virais/análise , Vírus da Influenza A Subtipo H1N1/química , Vírus da Influenza A Subtipo H1N1/isolamento & purificação , Proteínas Recombinantes/análise , Aptâmeros de Nucleotídeos/química , Humanos , Compostos de Estanho/químicaRESUMO
This study investigates the safe carrying load limit in relation to gender of workers in the sugar industry. A total of 632 workers (male and female) were interviewed following a physiological and psycho-physiological methodology. All of these workers were in the age groups of 21-30, 31-40 and ≥41 years. In this study, for the physiological methodology, both field and laboratory data were collected, which included heart rate, heart rate variability, energy expenditure and maximum rate of oxygen consumption. Information regarding hemoglobin content, weight, age, lean body weight, etc., was recorded to know the physical health of the workers. In the psycho-physiological evaluation, load-carrying limits were determined by the psychophysical rating of the load by workers from the selected age groups. By considering the physiological stresses and psycho-physiological evaluation, the safe carrying load limit was determined in head and back mode for male and female workers as per their age groups.
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Fazendeiros , Remoção , Saúde Ocupacional/normas , Estresse Fisiológico/fisiologia , Suporte de Carga/fisiologia , Adulto , Ergonomia/métodos , Feminino , Frequência Cardíaca/fisiologia , Humanos , Índia , Masculino , Consumo de Oxigênio/fisiologia , Esforço Físico/fisiologia , Fatores Sexuais , Açúcares , Inquéritos e QuestionáriosRESUMO
We present a novel vertical flow-based paper immunosensor for the rapid and sensitive electrochemical and colorimetric detection of influenza H1N1 viruses using a different pore size (DP) sample pad. The DP sample pad consisted of two different pore size papers: larger pores (diameter: 11⯵m) facing the inlet, and smaller pores (diameter: 0.45⯵m) facing the conjugate pad. This sample pad offered moderate and uniform flows, and hence concentrated horseradish peroxidase tagged antibodies (HRP-Abs)-H1N1 complexes from 40⯵L of sample volumes on a conjugate pad for 2â¯min after sample injection, thereby providing fast detection (6â¯min for both detection methods) with 100⯵L of flushing afterwards, high sensitivity, and the simplicity of the sensor. The filtration characteristics of the DP sample pad were evaluated using fluorescent beads, indicating that only small-sized bio-particles such as viruses can pass through the sample pad. Sandwich immunoreactions of HRP-Ab-H1N1-Ab were performed on the gold paper electrode of the immunoStrip, which was determined by electrochemical impedance spectroscopy (EIS) measurements. Simultaneously, the color signal of free HRP-Ab captured on the colorimetric zone was obtained using a scanner, and the intensity was analyzed using ImageJ. This immunosensor detected H1N1 virus concentration as low as 3.3 plaque forming units (PFU)/mL (phosphate buffer saline; PBS) and 4.7â¯PFU/mL (saliva) by EIS, and 1.34â¯PFU/mL (PBS) and 2.27â¯PFU/mL (saliva) by the colorimetric method. Furthermore, integrating these two detection methods can reduce false results with double assurance, and this device can provide a simple and economical on-site detection platform.
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Técnicas Biossensoriais/instrumentação , Colorimetria/instrumentação , Técnicas Imunoenzimáticas/instrumentação , Vírus da Influenza A Subtipo H1N1/isolamento & purificação , Influenza Humana/virologia , Papel , Anticorpos Imobilizados/química , Técnicas Eletroquímicas/instrumentação , Desenho de Equipamento , Humanos , Influenza Humana/diagnóstico , Limite de Detecção , PorosidadeRESUMO
Dielectrophoresis (DEP) is usually effective close to the electrode surface. Several techniques have been developed to overcome its drawbacks and to enhance dielectrophoretic particle capture. Here we present a simple technique of superimposing alternating current DEP (high-frequency signals) and electroosmosis (EO; low-frequency signals) between two coplanar electrodes (gap: 25 µm) using a lab-made voltage adder for rapid and selective concentration of bacteria, viruses, and proteins, where we controlled the voltages and frequencies of DEP and EO separately. This signal superimposition technique enhanced bacterial capture (Escherichia coli K-12 against 1-µm-diameter polystyrene beads) more selectively (>99%) and rapidly (~30 s) at lower DEP (5 Vpp) and EO (1.2 Vpp) potentials than those used in the conventional DEP capture studies. Nanometer-sized MS2 viruses and troponin I antibody proteins were also concentrated using the superimposed signals, and significantly more MS2 and cTnI-Ab were captured using the superimposed signals than the DEP (10 Vpp) or EO (2 Vpp) signals alone (p < 0.035) between the two coplanar electrodes and at a short exposure time (1 min). This technique has several advantages, such as simplicity and low cost of electrode fabrication, rapid and large collection without electrolysis.
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Eletro-Osmose/instrumentação , Escherichia coli K12/isolamento & purificação , Levivirus/isolamento & purificação , Poliestirenos/química , Proteínas/isolamento & purificação , Eletricidade , Eletrodos , Desenho de EquipamentoRESUMO
Although many studies concerning the detection of influenza virus have been published, a paper-based, label-free electrochemical immunosensor has never been reported. Here, we present a cost-effective, handmade paper-based immunosensor for label-free electrochemical detection of influenza virus H1N1. This immunosensor was prepared by modifying paper with a spray of hydrophobic silica nanoparticles, and using stencil-printed electrodes. We used a glass vaporizer to spray the hydrophobic silica nanoparticles onto the paper, rendering it super-hydrophobic. The super-hydrophobicity, which is essential for this paper-based biosensor, was achieved via 30-40 spray coatings, corresponding to a 0.39-0.41 mg cm-2 coating of nanoparticles on the paper and yielding a water contact angle of 150° ± 1°. Stencil-printed carbon electrodes modified with single-walled carbon nanotubes and chitosan were employed to increase the sensitivity of the sensor, and the antibodies were immobilized via glutaraldehyde cross-linking. Differential pulse voltammetry was used to assess the sensitivity of the sensors at various virus concentrations, ranging from 10 to 104 PFU mL-1, and the selectivity was assessed against MS2 bacteriophages and the influenza B viruses. These immunosensors showed good linear behaviors, improved detection times (30 min), and selectivity for the H1N1 virus with a limit of detection of 113 PFU mL-1, which is sufficiently sensitive for rapid on-site diagnosis. The simple and inexpensive methodologies developed in this study have great potential to be used for the development of a low-cost and disposable immunosensor for detection of pathogenic microorganisms, especially in developing countries.
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Imunoensaio , Anticorpos Imobilizados , Técnicas Biossensoriais , Técnicas Eletroquímicas , Eletrodos , Ouro , Vírus da Influenza A Subtipo H1N1 , Limite de Detecção , Nanotubos de CarbonoRESUMO
Primaquine (PQ) is the only drug used to prevent relapse of malaria due to P. vivax and P. ovale, by eradicating the dormant liver form of the parasite (hypnozoites). The side-effects associated with PQ limits is uses in treatment of malaria. To overcome the premature oxidative deamination and to increase the life span of drug in the biological system, the novel glyco-conjugates of PQ were synthesized by coupling of primaquine with hexoses in phosphate buffer. The saccharide part of the hybrid molecules thought to direct the drug to the liver, where hypnozoites resides. All the synthesized compounds were fully characterized and evaluated for their radical curative activities. The three compounds viz glucoside (15a), galactoside (15b) and mannoside (15c) with high activity were tested for their activity in rhesus monkeys where the most active compound 15b showed twofold activity (100% radical curative activity at 1.92 mmol/kg) than the standard drug PQ diphosphate (3.861 mmol/kg). It is proposed that results from these studies may be advantageous to develop a new potent tissue schizonticide antimalarial compound.