High-speed multifocal plane fluorescence microscopy for three-dimensional visualisation of beating flagella

Area of Science:

• Cell Biology
• Microscopy
• Parasitology

Background:

• Flagellum and cilium beating are crucial for cell motility.
• Analyzing these movements in 3D using fluorescence microscopy offers powerful insights.
• Uniflagellate parasites like *Trypanosoma brucei* and *Leishmania mexicana* rely heavily on motility.

Purpose of the Study:

• To reconstruct and analyze the 3D movement of *Trypanosoma brucei* and *Leishmania mexicana* flagella.
• To investigate the planarity of *L. mexicana* flagellar beating.
• To elucidate the mechanism of flagellar beating during *T. brucei* tumbling.

Main Methods:

• Utilized high-speed multifocal plane fluorescence microscopy to capture simultaneous focal planes.
• Achieved high frame rates (200 Hz) for visualizing flagellar and cell movement.
• Employed fluorescent stains and genetically-encoded fluorescent proteins for visualization.

Main Results:

• Reconstructed 3D movement of *T. brucei* and *L. mexicana* flagella.
• *L. mexicana* exhibits significant deviations from planar flagellar beating.
• *T. brucei* flagella bend the cell and beat distally during tumbling for reorientation.

Conclusions:

• High-speed multifocal plane fluorescence microscopy is a potent tool for analyzing flagellar dynamics.
• The study provides novel insights into the complex flagellar mechanics of these parasites.
• Findings contribute to understanding parasite motility and behavior.