Assessing Bacterial Swarming Using a Gradient Inhibitor Plate

Take a gradient swarm plate with double-layered agar. The bottom layer, containing an inhibitor, was poured to create a gradient of the inhibitor.

The plate contains isometrically spaced wells to provide appropriate space for bacterial swarming or collective migration, preventing swarm overlaps.

Inoculate the wells with a bacterial culture and incubate the plate.

Under low inhibitor concentrations, bacteria coordinate their flagellar movement, initiating collective migration or swarming.

Under high inhibitor concentrations, the inhibitor suppresses the expression of flagella-related genes.

This results in limited flagellar movement, leading to restricted swarming motility.

Image the plate under white light at regular intervals to capture the swarming pattern.

During imaging, adjust the exposure time to enhance the brightness of the swarms.

Modify the threshold settings to minimize background light interference and increase contrast.

Save the image for subsequent analysis of the effect of inhibitor concentration on bacterial swarming.

Add 80 microliters of the overnight growth culture into every well. Take the swarm plates out of the incubator one at a time every 12 hours and place them in th