Pseudomonas aeruginosa can be detected in a polymicrobial competition model using impedance spectroscopy with a novel biosensor

Electrochemical Impedance Spectroscopy (EIS) is a powerful technique that can be used to elicit information about an electrode interface. In this article, we highlight six principal processes by which the presence of microorganisms can affect impedance and show how one of these - the production of electroactive metabolites - changes the impedance signature of culture media containing Pseudomonas aeruginosa. EIS, was used in conjunction with a low cost screen printed carbon sensor to detect the presence of P. aeruginosa when grown in isolation or as part of a polymicrobial infection with Staphylococcus aureus. By comparing the electrode to a starting measurement, we were able to identify an impedance signature characteristic of P. aeruginosa. Furthermore, we are able to show that one of the changes in the impedance signature is due to pyocyanin and associated phenazine compounds. The findings of this study indicate that it might be possible to develop a low cost sensor for the detection of P. aeruginosa in important point of care diagnostic applications. In particular, we suggest that a development of the device described here could be used in a polymicrobial clinical sample such as sputum from a CF patient to detect P. aeruginosa

Impedimetric antimicrobial peptide-based sensor for the early detection of periodontopathogenic bacteria

Peri-implantitis, an inflammation caused by biofilm formation, constitutes a major cause of implant failure in dentistry. Thus, the detection of bacteria at the early steps of biofilm growth represents a powerful strategy to prevent implant-related infections. In this regard, antimicrobial peptides (AMPs) can be used as effective biological recognition elements to selectively detect the presence of bacteria. Thus, the aim of the present study was to combine the use of miniaturized and integrated impedimetric transducers and AMPs to obtain biosensors with high sensitivity to monitor bacterial colonization. Streptococcus sanguinis, which is one of the most prevalent strains in the onset of periodontal diseases, was used as a model of oral bacteria. To this end, a potent AMP derived from human lactoferrin was synthesized and covalently immobilized on interdigitated electrode arrays (IDEA). X-ray photoelectron spectroscopy (XPS) and electrochemical impedance spectroscopy (EIS) were employed to optimize and characterize the method of immobilization. Noteworthy, the interaction of Streptococcus sanguinis with AMP-coated sensors provoked significant changes in the impedance spectra, which were univocally associated with the presence of bacteria, proving the feasibility of our method. In this regard, the developed biosensor permits to detect the presence of bacteria at concentrations starting from 101 colony forming units (CFU) mL-1 in KCl and from 102 CFU mL-1 in artificial saliva. Moreover, the system was devoid of cytotoxicity for human fibroblasts. These results indicate that the proposed approach can be effective in the detection of initial stages of biofilm formation, and may be useful in the early prevention and treatment of peri-implantitisPeer ReviewedPostprint (author's final draft

Hierarchy and assortativity as new tools for affinity investigation: the case of the TBA aptamer-ligand complex

Aptamers are single stranded DNA, RNA or peptide sequences having the ability to bind a variety of specific targets (proteins, molecules as well as ions). Therefore, aptamer production and selection for therapeutic and diagnostic applications is very challenging. Usually they are in vitro generated, but, recently, computational approaches have been developed for the in silico selection, with a higher affinity for the specific target. Anyway, the mechanism of aptamer-ligand formation is not completely clear, and not obvious to predict. This paper aims to develop a computational model able to describe aptamer-ligand affinity performance by using the topological structure of the corresponding graphs, assessed by means of numerical tools such as the conventional degree distribution, but also the rank-degree distribution (hierarchy) and the node assortativity. Calculations are applied to the thrombin binding aptamer (TBA), and the TBA-thrombin complex, produced in the presence of Na+ or K+. The topological analysis reveals different affinity performances between the macromolecules in the presence of the two cations, as expected by previous investigations in literature. These results nominate the graph topological analysis as a novel theoretical tool for testing affinity. Otherwise, starting from the graphs, an electrical network can be obtained by using the specific electrical properties of amino acids and nucleobases. Therefore, a further analysis concerns with the electrical response, which reveals that the resistance sensitively depends on the presence of sodium or potassium thus posing resistance as a crucial physical parameter for testing affinity.Comment: 12 pages, 5 figure

Applied electrokinetics for tissue engineering applications

Tissue engineering could provide an alternative source of transplant tissue regardless of donor supply and with reduced risk of immune rejection. Engineered tissue requires three critical components for successful development: the cells, growth factors, and the scaffold on which they will initially grow. The scaffold acts as a temporary extracellular matrix (ECM) allowing cell attachment and acclimation to the environment prior to synthesis and construction of the cell\u27s native ECM. Prior to cell seeding, the scaffold must be characterized to determine whether or not the pore geometry is conducive to cell implantation. Electrochemical impedance spectroscopy (EIS) provides a unique and effective tool for scaffold evaluation upon fabrication as well as following modification and cell seeding. Chitosan scaffolds were tested with EIS showing good comparative values to more common and proven characterization methods. In addition to characterization, modification may be necessary to improve the cell scaffold interaction. Previous attempts to modify chitosan scaffolds with microparticles during the fabrication have shown that these approaches compromise scaffold integrity; however, particles cannot be effectively forced into the scaffold without damaging it. In light of this difficult modification, electrophoretic particle treatment was implemented. Transport of iron oxide microparticles at low voltage was proven an effective modification method and might be capable of modifying a seeded scaffold. Finally, it is necessary to not only evaluate the scaffold prior to cell seeding but also subsequent to cell seeding. Successful seeding should lead to cellular proliferation, gene expression, and ECM production. EIS has become a common tool for tissue characterization and can, in turn, be used as an evaluative approach for the assessment of engineered tissues. While cell response and volume exclusion requires a large quantity of cells, varying protein concentrations were notable in scaffolds saturated with complete cell media. In summary, electrokinetics has shown its potential for tissue engineering applications from scaffold fabrication to online monitoring

Physico-electrochemical Characterization of Pluripotent Stem Cells during Self-Renewal or Differentiation by a Multi-modal Monitoring System.

Monitoring pluripotent stem cell behaviors (self-renewal and differentiation to specific lineages/phenotypes) is critical for a fundamental understanding of stem cell biology and their translational applications. In this study, a multi-modal stem cell monitoring system was developed to quantitatively characterize physico-electrochemical changes of the cells in real time, in relation to cellular activities during self-renewal or lineage-specific differentiation, in a non-destructive, label-free manner. The system was validated by measuring physical (mass) and electrochemical (impedance) changes in human induced pluripotent stem cells undergoing self-renewal, or subjected to mesendodermal or ectodermal differentiation, and correlating them to morphological (size, shape) and biochemical changes (gene/protein expression). An equivalent circuit model was used to further dissect the electrochemical (resistive and capacitive) contributions of distinctive cellular features. Overall, the combination of the physico-electrochemical measurements and electrical circuit modeling collectively offers a means to longitudinally quantify the states of stem cell self-renewal and differentiation

Establishment of surface functionalization methods for spore-based biosensors and implementation into sensor technologies for aseptic food processing

Aseptic processing has become a popular technology to increase the shelf-life of packaged products and to provide non-contaminated goods to the consumers. In 2017, the global aseptic market was evaluated to be about 39.5 billion USD. Many liquid food products, like juice or milk, are delivered to customers every day by employing aseptic filling machines. They can operate around 12,000 ready-packaged products per hour (e.g., Pure-Pak® Aseptic Filling Line E-PS120A). However, they need to be routinely validated to guarantee contamination-free goods. The state-of-the-art methods to validate such machines are by means of microbiological analyses, where bacterial spores are used as test organisms because of their high resistance against several sterilants (e.g., gaseous hydrogen peroxide). The main disadvantage of the aforementioned tests is time: it takes at least 36-48 hours to get the results, i.e., the products cannot be delivered to customers without the validation certificate. Just in this example, in 36 hours, 432,000 products would be on hold for dispatchment; if more machines are evaluated, this number would linearly grow and at the end, the costs (only for waiting for the results) would be considerably high. For this reason, it is very valuable to develop new sensor technologies to overcome this issue. Therefore, the main focus of this thesis is on the further development of a spore-based biosensor; this sensor can determine the viability of spores after being sterilized with hydrogen peroxide. However, the immobilization strategy as well as its implementation on sensing elements and a more detailed investigation regarding its operating principle are missing. In this thesis, an immobilization strategy is developed to withstand harsh conditions (high temperatures, oxidizing environment) for spore-based biosensors applied in aseptic processing. A systematic investigation of the surface functionalization’s effect (e.g., hydroxylation) on sensors (e.g., electrolyte-insulator semiconductor (EIS) chips) is presented. Later on, organosilanes are analyzed for the immobilization of bacterial spores on different sensor surfaces. The electrical properties of the immobilization layer are studied as well as its resistance to a sterilization process with gaseous hydrogen peroxide. In addition, a sensor array consisting of a calorimetric gas sensor and a spore-based biosensor to measure hydrogen peroxide concentrations and the spores’ viability at the same time is proposed to evaluate the efficacy of sterilization processes

University libraries in a changing environment: experiences and the way forward in the new e-learning environment in Zimbabwe

The paper looks at the reasons prompting academic librarians to embrace e-learning and the benefits that can be derived from using electronic information services. The authors came up with some suggestions that academic librarians can positively participate in e-learning by providing and advocating for the use of electronic books and journals, promoting the open access initiative, imparting information literacy skills, providing selective dissemination of information and document delivery services and establishing and promoting the use of institutional repositories. The paper ends by giving recommendations on how society can get ahead with e-learning by involving academic librarians in the development nexus

Selling the JUBILEE Project: issues arising from a methodology

This article, inspired by JUBILEE project fieldwork, is influenced by van Leeuwen and Leeuw's work on the growing negative attitude towards market research. While the JUBILEE team, as academic researchers, were not making a sales pitch, they were trying to sell the project to putative participants. Issues discussed include length - and degree of difficulty - of questionnaire; how to sell a questionnaire; and how to identify key prospects for interviews and focus groups. Through the experiences gained, the JUBILEE researchers were able to identify key points of good fieldwork practice in relation to the issues discussed

Sensing with FETs - once, now and future

In this paper a short overview is given of the several FET-based sensor devices and the operational principle of the ISFET is summarized. Some of the shortcomings of the FET sensors were circumvented by an alternative operational mode, resulting in a device capable of acid/base concentration determination by coulometric titrant generation as well as in an original pH-static enzyme sensor. A more recent example is presented in which the ISFET is used for the on-line monitoring of fermentation processes. Future research is directed towards direct covalent coupling of organic monolayers on the silicon itself. In addition, the field-effect can be applied to the so-called semiconducting nanowire devices, ultimately making single molecule detection of charged species possible

Community sewage sensors towards evaluation of drug use trends: detection of cocaine in wastewater with DNA-directed immobilization aptamer sensors

Illicit drug use has a global concern and effective monitoring and interventions are highly required to combat drug abuse. Wastewater-based epidemiology (WBE) is an innovative and cost-effective approach to evaluate community-wide drug use trends, compared to traditional population surveys. Here we report for the first time, a novel quantitative community sewage sensor (namely DNA-directed immobilization of aptamer sensors, DDIAS) for rapid and cost-effective estimation of cocaine use trends via WBE. Thiolated single-stranded DNA (ssDNA) probe was hybridized with aptamer ssDNA in solution, followed by co-immobilization with 6-mercapto-hexane onto the gold electrodes to control the surface density to effectively bind with cocaine. DDIAS was optimized to detect cocaine at as low as 10 nM with a dynamic range from 10 nM to 5 μM, which were further employed for the quantification of cocaine in wastewater samples collected from a wastewater treatment plant in seven consecutive days. The concentration pattern of the sampling week is comparable with that from mass spectrometry. Our results demonstrate that the developed DDIAS can be used as community sewage sensors for rapid and cost-effective evaluation of drug use trends, and potentially implemented as a powerful tool for on-site and real-time monitoring of wastewater by un-skilled personnel