Looking Down the Barrel: Super-Resolution Imaging of Bacterial Nanoweapons

The bacterial Type VI Secretion System (T6SS) is a nanoweapon, comparable to a molecular speargun, which can be used to shape microbial communities, recognise kin, and kill adjacent competing microbes. T6SS-mediated interactions were only thought to occur between adjacent bacterial cells; however, T6SS-mediated interactions have recently been reported between the bacterium, Serratia marcescens, and several fungal species. The dynamics of the T6SS are poorly understood given the importance of such interactions in the environment and the host. This is due to a lack of adequate imaging methods to visualise interaction events, the transient nature of the system, and the need for new molecular tools to enable fluorescent labelling of T6SS components. I will outline the construction of a strain bank specifically designed for super-resolution microscopy, discuss the difficulties of bacterial immunostaining methods, and demonstrate the application of super-resolution microscopy techniques that enable super-resolved 3D imaging and quantification of the T6SS in S. marcescens. These methods result in the first 3D visualisations of the T6SS and lay the groundwork for understanding the dynamics of the system in a competitive polymicrobial environment.