Magnetic Separation and Centri-Chronoamperometric Detection of Foodborne Bacteria Using Antibiotic-Coated Metallic Nanoparticles.

Quality and food safety represent a major stake and growing societal challenge in the world. Bacterial contamination of food and water resources is an element that pushes scientists to develop new means for the rapid and efficient detection and identification of these pathogens. Conventional detection tools are often bulky, laborious, expensive to buy, and, above all, require an analysis time of a few hours to several days. The interest in developing new, simple, rapid, and nonlaborious bacteriological diagnostic methods is therefore increasingly important for scientists, industry, and regulatory bodies. In this study, antibiotic-functionalized metallic nanoparticles were used to isolate and identify the foodborne bacterial strains Bacillus cereus and Shigella flexneri. With this aim, a new diagnostic tool for the rapid detection of foodborne pathogenic bacteria, gold nanoparticle-based centri-chronoamperometry, has been developed. Vancomycin was first stabilized at the surface of gold nanoparticles and then incubated with the bacteria B. cereus or S. flexneri to form the AuNP@vancomycin/bacteria complex. This complex was separated by centrifugation, then treated with hydrochloric acid and placed at the surface of a carbon microelectrode. The gold nanoparticles of the formed complex catalyzed the hydrogen reduction reaction, and the generated current was used as an analytical signal. Our results show the possibility of the simple and rapid detection of the S. flexneri and B. cereus strains at very low numbers of 3 cells/mL and 12 cells/mL, respectively. On the other hand, vancomycin-capped magnetic beads were easily synthesized and then used to separate the bacteria from the culture medium. The results show that vancomycin at the surface of these metallic nanoparticles is able to interact with the bacteria membrane and then used to separate the bacteria and to purify an inoculated medium.

Casein-Conjugated Gold Nanoparticles for Amperometric Detection of Leishmania infantum.

Sensitive and reliable approaches targeting the detection of Leishmania are critical for effective early diagnosis and treatment of leishmaniasis. In this frame, this paper describes a rapid quantification assay to detect Leishmania parasites based on the combination of the electrocatalytic ability of gold nanoparticles (AuNPs) to act as a catalyst for the hydrogen formation reaction along with the specificity of the interaction between casein and the major surface protease of the Leishmania parasite, GP63. First, pure and casein-modified AuNPs were prepared and characterized by scanning electron microscopy and ultraviolet-visible spectroscopy. Then, casein-conjugated AuNPs were incubated with Leishsmania parasites in solution; the formed complex was collected by centrifugation, treated by acidic solution, and the pelleted AuNPs were placed on screen-printed carbon electrodes (SPCEs) and chronoamperometric measurements were carried out. Our results suggest that it is possible to detect Leishmania parasites, with a limit less than 1 parasite/mL. A linear response over a wide concentration interval, ranging from 2 × 10-2 to 2 × 105 parasites/mL, was achieved. Additionally, a pretreatment of Leishmania parasites with Amphotericin B, diminished their interaction with casein. This findings and methodology are very useful for drug efficacy assessment.

Ligand-Capped Ultrapure Metal Nanoparticle Sensors for the Detection of Cutaneous Leishmaniasis Disease in Exhaled Breath.

Human cutaneous leishmaniasis, although designated as one of the most neglected tropical diseases, remains underestimated due to its misdiagnosis. The diagnosis is mainly based on the microscopic detection of amastigote forms, isolation of the parasite, or the detection of Leishmania DNA, in addition to its differential clinical characterization; these tools are not always available in routine daily practice, and they are expensive and time-consuming. Here, we present a simple-to-use, noninvasive approach for human cutaneous leishmaniasis diagnosis, which is based on the analysis of volatile organic compounds in exhaled breath with an array of specifically designed chemical gas sensors. The study was realized on a group of n = 28 volunteers diagnosed with human cutaneous leishmaniasis and a group of n = 32 healthy controls, recruited in various sites from Tunisia, an endemic country of the disease. The classification success rate of human cutaneous leishmaniasis patients achieved by our sensors test was 98.2% accuracy, 96.4% sensitivity, and 100% specificity. Remarkably, one of the sensors, based on CuNPs functionalized with 2-mercaptobenzoxazole, yielded 100% accuracy, 100% sensitivity, and 100% specificity for human cutaneous leishmaniasis discrimination. While AuNPs have been the most extensively used in metal nanoparticle-ligand sensing films for breath sensing, our results demonstrate that chemical sensors based on ligand-capped CuNPs also hold great potential for breath volatile organic compounds detection. Additionally, the chemical analysis of the breath samples with gas chromatography coupled to mass spectrometry identified nine putative breath biomarkers for human cutaneous leishmaniasis.

Electrochemical detection of influenza virus H9N2 based on both immunomagnetic extraction and gold catalysis using an immobilization-free screen printed carbon microelectrode.

Influenza is a viral infectious disease considered as a source of many health problems and enormous socioeconomic disruptions. Conventional methods are inadequate for in-field detection of the virus and generally suffer from being laborious and time-consuming. Thus, studies aiming to develop effective alternatives to conventional methods are urgently needed. In this work, we developed an approach for the isolation and detection of influenza A virus subtype H9N2. For this aim, two specific influenza receptors were used. The first, anti-matrix protein 2 (M2) antibody, was attached to iron magnetic nanoparticles (MNPs) and used for the isolation of the virus from allantoic fluid. The second biomolecule, Fetuin A, was attached to an electrochemical detectable label, gold nanoparticles (AuNPs), and used to detect the virus tacking advantage from fetuin-hemagglutinin interaction. The MNP-Influenza virus-AuNP formed complex was isolated and treated by an acid solution then the collected gold nanoparticles were deposited onto a screen printed carbon electrode. AuNPs catalyzes the hydrogen ions reduction in acidic medium while applying an appropriate potential, and the generated current signal was proportional to the virus titer. This approach allows the rapid detection of influenza virus A/H9N2 at a less than 16 HAU titer.

Detection of ESAT-6 by a label free miniature immuno-electrochemical biosensor as a diagnostic tool for tuberculosis.

Tuberculosis is a worldwide disease considered as a major health problem with high morbidity and mortality rates. Poor detection of Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis remains a major obstacle to the global control of this disease. Here we report the development of a new test based on the detection of the major virulent factor of Mtb, namely the early secreted antigenic target 6-kDa protein or ESAT-6. A label free electrochemical immunosensor using an anti-ESAT-6 monoclonal antibody as a bio-receptor is described herein. Anti-ESAT-6 antibodies were first covalently immobilized on the surface of a gold screen-printed electrode functionalized via a self-assembled thiol monolayer. Interaction between the bio-receptor and ESAT-6 antigen was evaluated by square wave voltammetry method using [Fe(CN)6]3-/4- as redox probe. The detection limit of ESAT-6 antigen was 7ng/ml. The immunosensor has also been able to detect native ESAT-6 antigen secreted in cell culture filtrates of three pathogenic strains of Mtb (CDC1551, H37RV and H8N8). Overall, this work describes an immune-electrochemical biosensor, based on ESAT-6 antigen detection, as a useful diagnostic tool for tuberculosis.