Ultrasensitive and Highly Selective Detection of Staphylococcus aureus at the Single-Cell Level Using Bacteria-Imprinted Polymer and Vancomycin-Conjugated MnO2 Nanozyme.

Pathogenic bacterial infections, even at extremely low concentrations, pose significant threats to human health. However, the challenge persists in achieving high-sensitivity bacterial detection, particularly in complex samples. Herein, we present a novel sandwich-type electrochemical sensor utilizing bacteria-imprinted polymer (BIP) coupled with vancomycin-conjugated MnO2 nanozyme (Van@BSA-MnO2) for the ultrasensitive detection of pathogenic bacteria, exemplified by Staphylococcus aureus (S. aureus). The BIP, in situ prepared on the electrode surface, acts as a highly specific capture probe by replicating the surface features of S. aureus. Vancomycin (Van), known for its affinity to bacterial cell walls, is conjugated with a Bovine serum albumin (BSA)-templated MnO2 nanozyme through EDC/NHS chemistry. The resulting Van@BSA-MnO2 complex, serving as a detection probe, provides an efficient catalytic platform for signal amplification. Upon binding with the captured S. aureus, the Van@BSA-MnO2 complex catalyzes a substrate reaction, generating a current signal proportional to the target bacterial concentration. The sensor displays remarkable sensitivity, capable of detecting a single bacterial cell in a phosphate buffer solution. Even in complex milk matrices, it maintains outstanding performance, identifying S. aureus at concentrations as low as 10 CFU mL-1 without requiring intricate sample pretreatment. Moreover, the sensor demonstrates excellent selectivity, particularly in distinguishing target S. aureus from interfering bacteria of the same genus at concentrations 100-fold higher. This innovative method, employing entirely synthetic materials, provides a versatile and low-cost detection platform for Gram-positive bacteria. In comparison to existing nanozyme-based bacterial sensors with biological recognition materials, our assay offers distinct advantages, including enhanced sensitivity, ease of preparation, and cost-effectiveness, thereby holding significant promise for applications in food safety and environmental monitoring.

Reusable and universal impedimetric sensing platform for the rapid and sensitive detection of pathogenic bacteria based on bacteria-imprinted polythiophene film.

The rapid and sensitive detection of pathogenic bacteria is highly demanded for early warning of infectious disease epidemics and protection of human health. Herein, a reusable and universal impedimetric sensing platform based on a bacteria-imprinted polythiophene film (BIF) is proposed for the rapid and sensitive detection of pathogenic bacteria using Staphylococcus aureus (S. aureus) as a model analyte. Monomer screening among four 3-substituted thiophenes was first performed based on the imprinting factor, and 3-thiopheneethanol (TE) was eventually selected. The BIF as a recognition layer was quickly deposited in an environmentally friendly process on a glassy carbon electrode via electro-copolymerization of the S. aureus template and TE monomer followed by in situ template removal. Upon rebinding of S. aureus on the BIF, the impedance increased. Under optimal conditions, the BIF-based sensor can quantitatively detect S. aureus in a wide linear range of 10 to 107 CFU mL-1 with a low detection limit of 4 CFU mL-1. Additionally, the sensor exhibits excellent selectivity, capable of identifying S. aureus from multi-bacterial strain mixtures. It also demonstrates applicability in the analysis of real lettuce and shrimp samples with good recoveries. Most significantly, the BIF sensing interface can be reused up to five times with good signal retention. Compared with most reported methods, this sensor is more rapid with a much shorter total assay time of 30 min, including the BIF preparation, bacterial rebinding, and impedance detection. This assay may hold great potential to help in the rapid, sensitive, and label-free detection of pathogenic bacteria in fields of food safety and public health.

Rapid, sensitive and label-free detection of pathogenic bacteria using a bacteria-imprinted conducting polymer film-based electrochemical sensor.

The rapid and sensitive detection of pathogenic bacteria is very important for timely prevention and treatment of foodborne disease. Here, a bacteria-imprinted conductive poly(3-thiopheneacetic acid) (BICP) film-based impedimetric sensor was developed for the rapid, sensitive and label-free detection of staphylococcus aureus (S. aureus). The BICP film preparation was very convenient and eco-friendly, which was in situ deposited on gold electrode surface without the use of toxic organic solvents and cross-linkers. The process of imprinting and recognition were characterized by electrochemical technique and scanning electron microscope. The BICP had a novel structure without cocci-shaped cavities formed in the poly(3-thiopheneacetic acid) (PTAA) matrices. To obtain the optimal sensing performance, a set of factors affecting the imprinting and recognition were investigated. Under the optimized conditions, an extremely rapid recognition within 10 min, a very low limit of detection (LOD) of 2 CFU/mL, and wide linear range from 10 to 108 CFU/mL were achieved by the BICP film-based impedimetric sensor. The sensor also demonstrated high selectivity, and good universality and repeatability. Furthermore, the feasibility of its application has also been demonstrated in the analysis of real milk samples. This sensor offered a simple and universal method for rapid, sensitive, and selective detection of pathogenic bacteria, which could hold great potentials in fields like food safety.