Methods to Evaluate Bacterial Motility and Its Role in Bacterial-Host Interactions

Abstract

Bacterial motility is a widespread characteristic that can provide several advantages for the cell, allowing it to move towards more favorable conditions and enabling host-associated processes such as colonization. There are different bacterial motility types, and their expression is highly regulated by the environmental conditions. Because of this, methods for studying motility under realistic experimental conditions are required. A wide variety of approaches have been developed to study bacterial motility. Here, we present the most common techniques and recent advances and discuss their strengths as well as their limitations. We classify them as macroscopic or microscopic and highlight the advantages of three-dimensional imaging in microscopic approaches. Lastly, we discuss methods suited for studying motility in bacterial-host interactions, including the use of the zebrafish model.

Keywords: bacteria; bacterial motility; bacterial–host interaction; flagella; microscopy; motility methods.

Figure 1

Some examples of methods to study bacterial motility and motility tracking. (a) Soft agar assay is the most common macroscopic method used to study motility. After inoculating 1–6 µL or a stab of bacterial culture in soft agar, motile bacteria will spread and blur the media. (b) Assessing motility using some common microscopic methods is based on tracking individual bacteria to obtain their 2D trajectories. If a cell leaves the focal plane (orange cell) the track ends. (c) Three-dimensional trajectories can be obtained by (i) stacks of 2D slices along the z-axis (z-stacking) or by (ii) projecting the 2D image in the z-axis according to certain parameters such as depth-dependent shape in the case of defocused imaging methods. (d) Intravital microscopy (IVM) aims to visualize phenomena occurring inside live animals. For example, exposing the tissue of an anesthetized mouse by doing small incisions while carefully preserving its physiological conditions, a glass coverslip can be placed in the knee joint [32] or an intestinal loop [33] to visualize the movement of fluorescently labeled bacteria. Bacteria (green) are shown inside knee joint microvasculature. (e) The optical transparency of zebrafish larvae allows non-invasive visualization of the in vivo motility of fluorescent bacteria with light-sheet fluorescence microscopy (LSFM) in which a focal plane is illuminated, exciting all points in the plane simultaneously.