(Invited) Electrochemical Bacterial Sensing
Authors: Shiigi Hiroshi 
Published: December 2024

A better understanding of the biological functions of microorganisms is required to reduce their threats and increase their usefulness. Therefore, the importance of real-time evaluation of bacterial activity increases for various purposes such as hygiene management, development of antibacterial agents, and effective utilization of bacterial resources.1 This necessitates a quantitative assessment of metabolic processes, including growth and respiration. Here we would like to introduce the development of electrochemical methods for assessing bacterial activity.
Electrochemical detection of viable bacterial cells was performed using cell membrane permeable electron mediator and redox-active pigment. On the other hand, a tetrazolium salt (MTT), was converted to an insoluble reduction form (formazan) through the respiration of microbial cells.2 The insolubility of this formazan was effectively exploited as a surface-confined redox event. The electrochemical detection of formazan was effectively coupled with the thermal lysis of microbes. The sensitivity of the present technique is up to 10,000-fold higher than that of MTT colorimetry and requires an incubation time of only 1 h, which is approximately 1/4 of that required for other metabolism-based techniques. On the other hand, Considering that oxidized MTT, which has excellent cell membrane permeability, changes into insoluble reduced formazan within cells, it is also possible to estimate the number of viable cells by focusing on the reduction current of MTT remaining in the suspension. was. Dissolved oxygen is an important substance for bacterial activity. When live bacteria in suspension were observed in real-time, it was found that uptake of MTT into bacteria was completed within 10 minutes, including the lag period. Furthermore, we also observed that the current response was dependent on viable cell density, regardless of the bacterial species present. This method allows us to quickly estimate the number of viable bacteria, making it possible to confirm the safety of food before it leaves the factory and prevent food poisoning.3
Furthermore, the measurement of the reduction current of dissolved oxygen provides an effective means for assessing the respiratory activity of bacteria in suspension.4

Organometallic nanohybrids (NHs), in which many small metal nanoparticles are encapsulated within a conductive polymer matrix, are useful as sensitive electrochemical labels because their components produce characteristic oxidation current responses. NHs made of metal nanoparticles such as gold or copper and polyaniline did not interfere with each other in terms of electrochemical signals obtained on the same electrode. Antibodies were introduced into these NHs that served as electrochemical labels to target specific bacteria. Electrochemical measurements using screen-printed electrodes dry-fixed with NH-labeled bacterial cells allowed the bacterial species and number to be estimated within minutes based on the distinct current response of the label. Our proposed method achieved simultaneous detection of enterohemorrhagic E. coli and Staphylococcus aureus in real samples. These NHs are expected to be a powerful tool as electrochemical labels and useful for rapid testing at food and pharmaceutical manufacturing sites.5

1) T. Kinoshita, K. Ishiki, D. Q. Nguyen, H. Shiigi, T. Nagaoka, Anal. Chem., 90, 4098 (2018).
2) K. Ishiki, D. Q. Nguyen, A. Morishita, H. Shiigi, T. Nagaoka, Anal. Chem., 90, 10903 (2018).
3) H. Ikeda, A. Tokonami, S. Nishii, X. Shan, Y. Yamamoto, Y. Sadanaga, Z. Chen, and H. Shiigi, Anal. Chem., 95, 12358-12364 (2023).
4) M. Saito, K. Ishiki, D. Q. Nguyen, H. Shiigi, Anal. Chem., 91, 12793-12798 (2019).
5) S. Itagaki, A. Nakao, S. Nakamura, M. Fujita, S. Nishii, Y. Yamamoto, Y. Sadanaga, H. Shiigi, Anal. Chem., 96, 3787-3793 (2024).