Embedded biofilm, a new biofilm model based on the embedded growth of bacteria

A variety of systems have been developed to study biofilm formation. However, most systems are based on the surface-attached growth of microbes under shear stress. In this study, we designed a microfluidic channel device, called a microfluidic agarose channel (MAC), and found that microbial cells in the MAC system formed an embedded cell aggregative structure (ECAS). ECASs were generated from the embedded growth of bacterial cells in an agarose matrix and better mimicked the clinical environment of biofilms formed within mucus or host tissue under shear-free conditions. ECASs were developed with the production of extracellular polymeric substances (EPS), the most important feature of biofilms, and eventually burst to release planktonic cells, which resembles the full developmental cycle of biofilms. Chemical and genetic effects have also confirmed that ECASs are a type of biofilm. Unlike the conventional biofilms formed in the flow cell model system, this embedded-type biofilm completes the developmental cycle in only 9 to 12 h and can easily be observed with ordinary microscopes. We suggest that ECASs are a type of biofilm and that the MAC is a system for observing biofilm formation.