This study focuses on the gliding motility of Phormidium lacuna filaments. The methodology includes homogenizing the culture and observing the movement of filaments under a microscope.
Begin with a culture of Phormidium lacuna filaments.
Briefly homogenize the culture to break up filament clusters and generate a uniform suspension.
Transfer the homogenized sample to a Petri plate and allow it to rest.
Cover the plate with cellophane to reduce evaporation.
Place a slide on the microscope stage to serve as a support platform.
Position the plate containing the sample directly on top of the slide.
Use a low-power objective lens and focus on the bacterial filaments suspended in the medium to observe their movement.
The filaments display a slow, gliding motion across the surface, driven by type IV pili-the microscopic fibers assembled at the inner membrane and emerging through the outer membrane.
These fibers extend and retract to pull the cells forward, allowing them to glide.
This assay reveals the characteristic gliding motility of Phormidium lacuna in a liquid environment.
Cultivate P. lacuna in F2 medium under 50 RPM horizontal agitation in white light for around five days until the estimated optical density at 750 nanometers becomes 0.35. Store the sample at four degrees Celsius. Homogenize the filaments with ultrasound for one minute at maximum power and cycle of one.
Measure the optical density at 750 nanometers and transfer eight milliliters of the medium containing P. lacuna into a six-centimeter Petri dish. Wait for a few minutes until the sample reaches room temperature and then cover the Petri dish with cellophane foil. Place a microscope slide on the X-Y table of a standard microscope with a camera.
Switch on the microscope light and move a 4X or 10X objective into the path of the light. Then, place the Petri dish on top of the slide. Adjust single filaments or filament bundles by the table's X, Y, and Z movements.
Observe the movements of single filaments or bundles and record the movements with a standard microscope camera. Ensure that the objective lens does not touch the liquid.
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