Development of Non-Invasive Impedance Sensors for Real Time Monitoring of Bacteria Growth

Abstract

Mucosal surface in the gastrointestinal track and oral cavity are colonized by more than 700 types of microorganism. The bacteria interaction is depended on the location of its formation and dynamics. Streptococcus mutans, is known pathogenic bacteria present in oral microbiome, which metabolizes sucrose and other carbohydrates to produce lactic acid and lowers the pH 5.5 or less, which causes caries and periodontal diseases. Therefore, study of growth and dynamics of Sm bacteria is critical to understand oral cavity formation. Fluorescent in-situ hybridization probe (FISH) with confocal laser scanning microscopy (CLSM) is widely used for monitoring of biofilm with spatial resolution. However, these sophisticated instruments are not economically viable for multiple replication or high throughput applications. Here, we have used single frequency impedance spectroscopy as fast and non-invasive technique to monitor real time growth of Sm biofilm and kinetics of its formation.Real time monitoring of Sm biofilm was performed by the continuous recording of impedance while Sm bacteria was grown on glass substrate in BMM with 30 mM sucrose feeding. The cell-sensors impedance has been reported in the arbitrary unit as normalized impedance called cell-index (CI). Sensors were first tested in phosphate buffer solution (PBS) and then changed to BMM as background signal. After stabilization of sensors, streptococcus mutane was injected. There is increase of CI after Sm injection from zero to 0.3 at 10kHz frequency then decreased ~ 0.2. The decrease of CI at 10 kHz is lactic acid release by the Sm bacteria which causes decrease in solution resistance. The CI value at 1 Hz is increased from zero to ~ 1 after bacteria inoculation. The increase in CI at lower frequency is due to growth of Sm bacteria on the electrodes surface which causes increase of capacitance. After 56 hrs of growth of Sm, surfactant sodium dodecyl sulphate (SDS, CMC) was injected for investigation of sensitivity of impedancemetry sensors on destruction of bacteria biofilm. There is decrease in CI value at 1 Hz suggests that Sm biofilm matrix was partially collapsed. So, this study suggests that, high frequency impedance-based sensor can be used as solution resistance sensors and low frequency impedance based sensor can be used as capacitive based sensor for monitoring of biofilm growth.