Three-dimensional structure of the Z-ring as a random network of FtsZ filaments.

The spatial organization of the Z-ring, the central element of the bacterial division machinery, is not yet fully understood. Using optical tweezers and subpixel image analysis, we have recently shown that the radial width of the Z-ring in unconstricted Escherichia coli is about 100 nm. The relatively large width is consistent with the observations of others.

Oriented imaging of 3D subcellular structures in bacterial cells using optical tweezers.

Using oscillating optical tweezers, we show that controlled alignment of rod-shaped bacterial cells allows imaging fluorescently labeled three-dimensional (3D) subcellular structures from different, optimized viewpoints. To illustrate our method, we analyze the Z ring of E. coli.

Timing of Z-ring localization in Escherichia coli.

Bacterial cell division takes place in three phases: Z-ring formation at midcell, followed by divisome assembly and building of the septum per se. Using time-lapse microscopy of live bacteria and a high-precision cell edge detection method, we have previously found the true time for the onset of septation, τ(c), and the time between consecutive divisions, τ(g). Here, we combine the above method with measuring the dynamics of the FtsZ-GFP distribution in individual Escherichia coli cells to determine the Z-ring positioning time, τ(z).

Rotation of single bacterial cells relative to the optical axis using optical tweezers.

Using a single-beam, oscillating optical tweezers, we demonstrate trapping and rotation of rod-shaped bacterial cells with respect to the optical axis. The angle of rotation, θ, is determined by the amplitude of the oscillation. It is shown that θ can be measured from the longitudinal cell intensity profiles in the corresponding phase-contrast images.

Shape of nonseptated Escherichia coli is asymmetric.

The shape of Escherichia coli is approximately that of a cylinder with hemispherical caps. Since its size is not much larger than optical resolution, it has been difficult to quantify deviations from this approximation. We show that one can bypass this limitation and obtain the cell shape with subpixel accuracy.