Enhanced Electrochemical Impedance Spectroscopy Analysis of Microbial Biofilms on an Electrochemically <i>In Situ</i> Generated Graphene Interface

Abstract

Biofilms can cause many bacterial diseases, such as dental disease. An <i>in vitro</i> detection of biofilms may help to screen antibiofilm drugs. An impedance measurement based on an Au electrode has been successfully used for <i>in vitro</i> real-time monitoring of animal and human cell growth. However, microbial growth on the Au electrode produced a poor signal because of the small size of microbial cells. We have recently demonstrated that graphene derivatives can be produced on a carbon electrode through facile electrochemical activation, thus forming a reduced graphene oxide-carbon electrode (rGO-CE). Based on this fact, we hypothesized that an <i>in vitro</i> formed rugose graphene layer of rGO-CE may provide a large surface area for the growth of microbial biofilms and can therefore produce a strong impedance signal in response to a change in the biomass. In this study, three oral bacteria, <i>Streptococcus mutans</i> (<i>S. mutans</i>), <i>Actinomyces viscosus</i> (<i>A. viscosus</i>), and <i>Lactobacillus fermentum</i> (<i>L. fermentum</i>), were cultured on the surfaces of rGO-CE. As a result, the impedance response signal of the rGO-CE for the growth of <i>S. mutans</i> and <i>A. viscosus</i> was found to be 3.3 times and 6.0 times stronger than that of the Au electrode at 1.17 and 54.7 kHz, respectively. In particular, the poorly adhering strain of <i>L. fermentum</i> also produced a detectable signal on the graphene electrode but not on the Au electrode at 1.17 kHz. Furthermore, destructions of the biofilms grown on the rGO-CE by cetylpyridinium chloride were successfully monitored by impedance changes. Overall, it is promising to develop a graphene-based impedance biosensor platform for biofilm study and antibiofilm drug screening.