Have you ever wondered what the life of bacteria looks like? They eat, grow, divide, and continue that cycle. And just like us, they live in a community with many other bacteria. Their social interactions with others, however, are not always pleasant. Just like how lizards (predator) catch and eat insects (prey) for sustenance, bacteria can ambush and kill other bacteria too – they are indeed the smallest hunters on Earth!
A predatory bacterium chasing its much larger prey. Video by Welsh and Stocker/Stocker Lab, MIT and ETH Zurich
One of the most notorious predatory bacteria is Bdellovibrio bacteriovorus. Despite being a tiny, microscopic bacterium (Figure 1, top, comma-shaped with a flagellum), it can prey on a much larger bacterium (Figure 1, bottom, rod-shaped).
Figure 1. A false-color micrograph of Bdellovibrio bacteriovorus infecting a larger bacterium. Credit: Alfred Pasieka/Science Source
As it approaches and attacks its prey, B. bacteriovorus drills a hole in the prey’s cell wall, just small enough for it to squeeze in the space between the cell wall and the inner membrane (Figure 2, Attack phase). Once inside, it reseals and continues its life cycle within the prey. Much of a fierce invader, it steals the nutrients from the prey to grow and divide, then its offspring will burst out to find their next meals (Figure 2, Proliferative phase)
Figure 2. Predatory life cycle of Bdellovibrio bacteriovorus. Source: Laloux 2020
While bacteria-eating-bacteria are quite widespread in nature and can be detected in soil, rivers, the ocean, water treatment plants, crab gills, and oyster shells, etc., they have not gathered much attention from the research community. Scientists seem a lot more interested in viruses, protists, and animals that kill and eat bacteria, and even the small community of researchers who study predatory bacteria have not fully understood the dynamics of bacterial predator-prey interactions, and in particular, how B. bacteriovorus remodels the cell wall of its prey. Unfortunately, the predator and the prey have such a similar cell wall architecture that makes it impossible to visualize what happens at their points of contact under a black-and-white electron microscope.
So why are scientists curious about bacterial cell walls? If you were a bacterium, then the cell wall would be your must-have protective layer so that you could maintain your shape without swelling, shrinking, or even bursting in response to the water pressure from the surroundings. What makes the cell wall such a wonderful armor is the peptidoglycan composition, which is a molecular mesh consisting of chains of sugars called NAG and NAM as well as short chains of (unusual) amino acids (Figure 3).
Figure 3. Peptidoglycan composition of the bacterial cell wall. Source: Adapted from Figure 3.17, p. 89 in Microbiology: An Evolving Science, 2nd ed.; Joan L. Slonczweski and John W. Foster (2011)
Figure 4. Bacterial cell wall acts as a protective armor against lytic enzymes and helps bacteria escape from a host’s innate immune system. Credit: Felipe Cava, Umeå University
More interestingly, B. bacteriovorus can utilize special enzymes to exchange a range of naturally occurring D-amino acids (DAAs) with the fifth- and fourth-position D-alanines in the peptide crosslinks (Figure 3), which may strengthen the cell wall and make it even a better armor (Figure 4)! From this unique feature, scientists developed an innovative fluorescent imaging technique to visualize the dynamics of the peptidoglycan cell wall. Think of a marathon where different teams wear different color shirts. The color is like a tag to keep track of which member belongs to which team and where they are along the route. A similar approach is applied here – the D-amino acids which will be incorporated into the peptidoglycan cell wall are labeled with different fluorescent dyes for different cells. So even if B. bacteriovorus and its prey have similar cell wall architecture, we can distinguish them at their points of contact based on distinct fluorescent signals for these bacterial cells.
Understanding the mechanisms of how predatory bacteria like B. bacteriovorus invade their prey is a crucial step toward deploying them to help beat back harmful microbes in the environment. With a three-dimensional fluorescence microscope and innovative fluorescent D-amino acids (FDAA) labeling approach, Kuru, Lambert and their colleagues were able to visualize the interactions between bacteria with similar cell wall architecture and illuminate earlier hypotheses about how B. bacteriovorous invades and modifies the prey’s cell wall (E. coli) to suit its needs. Their findings were published in Nature Microbiology in 2017.
Figure 5. Three-dimensional illumination microscopy images that show how B. bacteriovorus (false-colored red with BADA) modifies the membrane of E. coli (false-colored cyan with HADA) through cell wall modification. BADA and HADA are fluorescent D-amino acids that can be incorporated into the peptidoglycan units in bacterial cell walls. Source: Adapted from Figure 2a,b in Kuru et al. 2017
In Figure 5a, we can see that the contact point of B. bacteriovorus and prey E. coli, which is pointed out by the white arrowheads, has a brightly colored ring structure. This structure represents the location of rigorous peptidoglycan modification. Since the HADA and BADA are incorporated in the peptidoglycan of the cell wall, and the HADA was always observed on the prey cell wall regardless of the predator's location, it is likely that the prey’s cell wall has been modified upon contact with the predator. Furthermore, the ring structure has a slightly smaller width compared to the prey cell, which supports the previously suggested idea that the predator squeezes itself into a smaller entry pore on the prey surface. Also, Figure 5b shows that the prey cell wall has deformed around the predator entry pore, which might indicate that B. bacteriovorus produces enzymes that soften the prey cell wall, possibly to assist the invasion into the prey.
Credit: Yuming Wang, Mount Holyoke College
Figure 5. Source: Adapted from Figure 2c in Kuru et al. 2017
What’s more intriguing is that the predator seems to be able to modify the prey cell wall again to seal the entry pore. Among the post-invasion prey cells that were investigated, 27% showed HADA-labeled ring structures similar to the entry pore at the end of the predator cell, where it comes in contact with the prey cell wall. In addition to that, 4% of the samples had HADA-labeled filled discs at the point of prey-predator contact, which might suggest that B. bacteriovorus is able to seal the entry pore by modifying the prey cell wall and thus maintain a favorable environment for itself to grow and multiply.
Credit: Yuming Wang, Mount Holyoke College
B. bacteriovorus is also very unique in the way that it divides - it undergoes asymmetric division! After the B. bacteriovorus have successfully attacked and entered the prey cell, they start feeding on its constituents with the help of certain digestive enzymes. This allows the B. bacteriovorus to grow and start dividing. The process of division also involves modifications to their peptidoglycan cell walls. This dynamic process is visualized in this figure with the help of FDAA labeling.
Figure 6. Peptidoglycan cell wall modifications as the cell undergoes asymmetric division. Source: Adapted from Fig. 6 in Kuru et al. 2017
The figure shows that growth starts in patches along the length of the now elongated B. bacteriovorus cell. Lines of division can be seen growing synchronously at regular intervals across its body (much like sausage links!). Once the cells have successfully completed division and have separated from each other, they undergo more cell wall modifications and grow a flagellum on one pole and pili on the other. The flagellum and the pili are hair-like structures that protrude from the outer membrane and help bacteria gain motility required to swim and seek out new prey. These new daughter B. bacteriovorus cells are now ready to break out of their host cell and move on to the “attack phase”, where they move on to find, attack and feed off other prey cells.
B. bacteriovorus are quite ruthless and really good at what they do! Since they can destroy lots of different bacteria, they have the potential to be used as antimicrobial agents with lots of specialized advantages. They are highly specific to Gram-negative bacteria which they exclusively prey on, which means that they are less likely to harm other cells such as those in the human body. They can also be used as alternatives to antibiotics and help overcome the growing issue of antibiotic and drug resistance. There are multiple studies in progress exploring the use of B. bacteriovorus and other bacterial predators to treat and prevent human infections.
Stay tuned to find out how we may soon be deploying these microbial assassins for our benefit!
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