Investigations into the Life Cycle of the Bacterial Predator Bdellovibrio bacteriovorus 109J at an Interface by Atomic Force Microscopy

Abstract

Atomic force microscopy was used to image Bdellovibrio bacteriovorus 109J, a gram-negative bacterial predator that consumes a variety of other gram-negative bacteria. In predator-prey communities grown on filters at hydrated air-solid interfaces, repeated cycles of hunting, invasion, growth, and lysis occurred readily even though the cells were limited to near two-dimensional movement. This system allowed us to image the bacteria directly without extensive preparation or modification, and many of the cells remained alive during imaging. Presented are images of the life cycle in two species of prey organisms, both Escherichia coli (a small prey bacterium that grows two-dimensionally on a surface) and Aquaspirillum serpens (a large prey bacterium that grows three-dimensionally on a surface), including high-resolution images of invaded prey cells called bdelloplasts. We obtained evidence for multiple invasions per prey cell, as well as significant heterogeneity in morphology of bdellovibrios. Mutant host-independent bdellovibrios were observed to have flagella and to excrete a coating that causes the predators to clump together on a surface. Most interestingly, changes in the texture of the cell surface membranes were measured during the course of the invasion cycle. Thus, coupled with our preparation method, atomic force microscopy allowed new observations to be made about Bdellovibrio at an interface. These studies raise important questions about the ways in which bacterial predation at interfaces (air-solid or liquid-solid) may be similar to or different from predation in solution.

Spatially Organized Films from Bdellovibrio bacteriovorus Prey Lysates

“…Finding many bdelloplasts in the outer region is consistent with the results of our 2003 study (29) that showed an advancing tide of bdelloplasts when HD bdellovibrios were deposited on a extant, flat monolayer of E. coli prey. Flannagan et al (6) downregulated an motA motility gene, resulting in variously shaped bdelloplasts, reduction in the motility of progeny bdellovibrios, and a much delayed lysis release.…”

“…(E. coli cells can be long as shown in Fig. S1 in the supplemental material, and previous work with Aquaspirillum has yielded long bdelloplasts [29]). Two large bdelloplasts observed in overlay (with E. coli in green from the GAM42a probe) and bright-field images, respectively, are displayed in panels d and e. In Fig.…”

“…Standard silicon nitride tips were used for these measurements. Many details of the procedures and instrumental parameters for imaging bacterial cells by AFM are provided in our previous publications (1,27,29).…”

“…In previous studies, our collaboration has explored interactions between wild-type Bdellovibrio predators and their prey at interfaces in model biofilm communities. When trapped atop a nutrient-poor flat surface with Escherichia coli prey cells, bdellovibrios multiply robustly and quickly devour their prey (29). In model E. coli biofilms submerged in liquid, B. bacteriovorus hunts vigorously, decimating or eradicating its prey in a single-component biofilm (1).…”

Three-Dimensional Imaging of the Highly Bent Architecture of Bdellovibrio bacteriovorus by Using Cryo-Electron Tomography

“…Moreover, the contrast in the images originates primarily from the stain and not from the intrinsic density of target structures, limiting the resolution attainable by this approach. Atomic force microscopic studies have also been used to probe the bacterial surface, but detailed internal structures are not visible and the samples are also subjected to freeze-drying and staining (24,25). Cryo-EM and cryo-electron tomography are powerful alternatives for visualization of the global architecture of bacterial cells that have been preserved in their native state (4,19,29,39).…”