Sensor system for analysis of biofilm sensitivity to ampicillin.

The resistance of biofilms to antibiotics is a key factor that makes bacterial infections unsusceptible to antimicrobial therapy. The results of classical tests of cell sensitivity to antibiotics cannot be used to predict therapeutic success in infections associated with biofilm formation. We describe a simple and rapid method for the real-time evaluation of bacterial biofilm sensitivity to antibiotics, with Pseudomonas putida and ampicillin as examples. The method uses an electric biosensor to detect the difference between changes in the biofilm electric polarizability, thereby evaluating antibiotic sensitivity. The electric signals showed that P. putida biofilms were susceptible to ampicillin and that at high antibiotic concentrations, the biofilms differed markedly in their susceptibility (dose-dependent effect). The sensor also detected differences between biofilms before and after ampicillin treatment. The electric-signal changes enabled us to describe the physical picture of the processes occurring in bacterial biofilms in the presence of ampicillin. The approach used in this study is promising for evaluating the activity of various compounds against biofilms, because it permits a conclusion about the antibiotic sensitivity of biofilm bacteria to be made in real time and in a short period (analysis time, not longer than 20 min). An added strong point is that analysis can be done directly in liquid, without preliminary sample preparation. KEY POINTS: • Sensor system to analyze biofilm antimicrobial susceptibility is described. • The signal change depended on the ampicillin concentration (dose-dependent effect). • The sensor allows real-time determination of the antibiofilm effect of ampicillin.

Antimicrobial Resistance and Current Methods for its Detection.

Infection diagnosis and antibiotic sensitivity testing are important aspects of clinical microbiology that are in dire need of improvement owing to the inadequate current standards in the early detection of bacterial response to antibiotics. The increasing antimicrobial resistance is a serious global threat to human health. Current resistance-detecting methods, using the phenotypic antibiotic sensitivity test, which measures bacterial growth as affected by antibiotics, have long analysis times. Therefore, new and rapid methods are needed to detect antibiotic resistance. Here, we review the methods used to detect antibiotic resistance in bacteria, including that caused by biofilm development, and we look at the development of rapid methods for evaluating antimicrobial resistance (AMR).

Antibody Phage Display Technology for Sensor-Based Virus Detection: Current Status and Future Prospects.

Viruses are widespread in the environment, and many of them are major pathogens of serious plant, animal, and human diseases. The risk of pathogenicity, together with the capacity for constant mutation, emphasizes the need for measures to rapidly detect viruses. The need for highly sensitive bioanalytical methods to diagnose and monitor socially significant viral diseases has increased in the past few years. This is due, on the one hand, to the increased incidence of viral diseases in general (including the unprecedented spread of a new coronavirus infection, SARS-CoV-2), and, on the other hand, to the need to overcome the limitations of modern biomedical diagnostic methods. Phage display technology antibodies as nano-bio-engineered macromolecules can be used for sensor-based virus detection. This review analyzes the commonly used virus detection methods and approaches and shows the prospects for the use of antibodies prepared by phage display technology as sensing elements for sensor-based virus detection.

Fourier Transform Infrared (FTIR) Spectroscopic Study of Biofilms Formed by the Rhizobacterium Azospirillum baldaniorum Sp245: Aspects of Methodology and Matrix Composition.

Biofilms represent the main mode of existence of bacteria and play very significant roles in many industrial, medical and agricultural fields. Analysis of biofilms is a challenging task owing to their sophisticated composition, heterogeneity and variability. In this study, biofilms formed by the rhizobacterium Azospirillum baldaniorum (strain Sp245), isolated biofilm matrix and its macrocomponents have for the first time been studied in detail, using Fourier transform infrared (FTIR) spectroscopy, with a special emphasis on the methodology. The accompanying novel data of comparative chemical analyses of the biofilm matrix, its fractions and lipopolysaccharide isolated from the outer membrane of the cells of this strain, as well as their electrophoretic analyses (SDS-PAGE) have been found to be in good agreement with the FTIR spectroscopic results.

Recombinant antibodies by phage display for bioanalytical applications.

Antibody phage display, aimed at preparing antibodies to defined antigens, is a useful replacement for hybridoma technology. The phage system replaces all work stages that follow animal immunization with simple procedures for manipulating DNA and bacteria. It enables the time needed to generate stable antibody-producing clones to be shortened considerably, making the process noticeably cheaper. Antibodies prepared by phage display undergo several affinity selection steps and can be used as selective receptors in biosensors. This article briefly describes the techniques used in the making of phage antibodies to various antigens. The possibilities and prospects are discussed of using phage antibodies as selective agents in analytical systems, including biosensors.

Microfluidic bioanalytical system for biofilm formation indication.

The formation of biofilms is a key factor that researchers must consider when they work with bacterial cultures. We describe a new microfluidic bioanalytical sensory system for indicating biofilm formation. The method is demonstrated with Pseudomonas bacteria as an example and is based on the real-time recording of cell-polarizability changes caused by an alternating electric field. Control experiments using phase-contrast microscopy and traditional microbiological plating were done that proved biofilms had formed. The physical picture was described of the sensor-signal changes during cell transition from planktonic to biofilm growth. This transition was indicated by the appearance of a peak-shaped signal at 500 kHz and by an increase in the recorded relaxation time. Phenomena of increase in the signal relaxation time from 2.4 s for planktonic to 25.4 s for biofilm cells. The proposed microfluidic sensor system for indicating biofilm formation holds much promise, because it ensures an analysis time of about 20-30 min. An added bonus is that for this system there is no need to grow bacterial biofilms in a sensor and the flow cell is reusable.

Docking and antibacterial activity of novel nontoxic 5-arylidenepyrimidine-triones as inhibitors of NDM-1 and MetAP-1.

Background: Antibiotic resistance, which occurs through the action of metallo-ß-lactamases (NDM-1), is a serious problem in the treatment of infectious diseases. Therefore, the discovery of new NDM-1 inhibitors and promising antibacterial agents as inhibitors of alternative targets (MetAP-1) is important. Method & results: In this study, a virtual library of 5-arylidene barbituric acids was created and molecular docking was performed for identification of novel possible inhibitors of NDM-1 and MetAP-1. Antibacterial activity (agar well-diffusion assay) and cytotoxicity (alamarBlue assay) of perspective compounds were evaluated. Pharmacokinetic profiles and molecular properties were predicted. Conclusion: We have identified possible novel inhibitors of NDM-1 and MetAP-1 with bacteriostatic activity, most of which are not cytotoxic and have potential excellent drug-likeness properties.

Bacteria-based electro-optical platform for ampicillin detection in aquatic solutions.

We have shown for the first time that it is possible to use a bacteria-based sensory system consisting of the bacterium Pseudomonas putida TSh-18 and an electro-optical sensor to detect ampicillin in the concentration range 0.5-600 µg/mL. Changes in the anisotropy of cell polarizability were detected at 900 and 2100 kHz; these represented the state of the cytoplasm and of the cell membrane, respectively. The changes indicate the quickest cell response to changes in the characteristics of the bacterial culture exposed to ampicillin. We have also shown that it is possible to monitor the ampicillin in the presence of kanamycin. In control experiments, we examined the effects of ampicillin and kanamycin on bacterial cells by phase-contrast microscopy and by standard microbiological tests on solid media. P. putida TSh-18 is recommended as a sensor system for ampicillin detection. Electro-optical analysis ensures detection of ampicillin in aquatic solutions in real-time, takes 10 min, and offers a lower limit of ampicillin detection of 0.5 µg/mL, which is lower than the European Community's maximum residue limit standards for penicillin antibiotics.