Wednesday, September 17, 2014

Should I Stay Or Should I Go

Biology concepts – bacteria, motility, flagella, quorum sensing, bacterial swarming, biofilms, pathogenesis

Nomads are wanderers. They come in different flavors.
Hunter-gatherers follow the animals as they graze in
different places. Pastoral nomads have animal herds and
move them around to where the grazing is best. But the
interesting ones are the peripatetic nomads. These are
people that move around within cities and other
populated areas, often to sell services or trades. Romanis,
or gypsies as they are sometimes called, are a
group of peripatetic nomads.
We humans have complex interactive behaviors with one another - these can make things better or, oh so much worse. We form herds as nomadic tribes, or we settle to form cities. Each has its own set of niches and behaviors that must be fulfilled by members of the group. But, it's important that we realize that we aren’t doing anything new, apparently bacteria have been roaming and settling for billions of years.

Our current series has been talking about flagella and how they help bacteria become motile (amongst other things). A relatively new discovery has opened our eyes to an exceptional movement by flagellated bacteria, swarming.

Swimming is when a bacterium on a liquid/surface interface or in liquid moves around by itself using its flagella as a propeller. But groups of bacteria can use their flagella to create a swarm; a mass of bacteria moving as one unit, often faster than the individuals can move on their own.

Bacteria moving as a unit is like tribes of humans moving from one place to another. But there are also those bacteria that choose to hunker down in one location and build a “city.” This prokaryotic Gotham is called a biofilm. We should do a whole series on biofilms, but for now let’s just talk about them in general.

When a number of bacteria of the same type, or sometimes even of different types, are in the same place at the same time, they may begin to form a biofilm. Certain bacteria will secrete proteins as filaments, polysaccharides in the form of slime, and some other structures. All of these together form a network of tunnels, tubules, cavities, and surfaces onto which the bacteria adhere. The biofilm also adheres to whatever surface is nearby. It’s a bacterial city.

The plaque on your teeth is a biofilm. The saliva and
crevicular fluid (between root and gum) provides some
proteins and sugars to build the film. Above is a
photomicrograph of plaque showing that yeast and
bacteria are both involved in mature plaque.
The biofilm matures over time, and different bacteria will have different jobs. The bacteria are stronger together than they are on their own, since the biofilm can prevent antibacterial agents from working. Biofilms are turning out to be important virulence factors (structures that enhance an organism’s ability to cause disease) and are crucial for pathogenesis (patho = disease, and genesis = beginning or course).

Some bacterial colonies settle to form cities and some move on en masse to another location – it really does sound like humans tribes. But biofilms and swarming are not mutually exclusive, in some cases you will see the bacteria at the edge of a biofilm start to swarm and expand, just like urban sprawl creates bigger cities.

There's organization to the swarm as well. Swarming isn’t an, “Everybody run!” kind of movement. Swarming requires controls, regulation, and numerous gene products spread out over the colony. Even though they work as a group, the bacteria might not all go the same direction.

Since bacteria divide by binary fission (one form of asexual reproduction), they tend to form masses in one location, often circular. When they give the signal(s) to swarm, some may take off in this direction, and some in another, based on where they are in the circle. Look at the picture below and right. Pretty, but it shows that colony swarm has multiple leading edges that will travel out into the unknown, and part of the colony will follow behind each.

Every once in a while, a new leading edge might branch off and swarm in a different direction, taking some followers with it. Other types of bacteria seem to swarm equally in all directions, forming concentric circles of new colonies.

This is a false color image showing the branching of a
bacteria colony in a swarm. Dr. Eshel Ben-Jacob from
Tel Aviv University produces these images as science
and art. See many of his images at this site.
The disease-causing bacteria Pseudomonas aeruginosa branches when it swarms, but even this is coordinated. A 2014 paper used a computer to model the branches seen in P. aeruginosa. They occur over a very narrow range of parameters. This means that the bacteria are limiting their activities and conducting themselves within a finely adapted range of behaviors and signals. Bottom line - their movement isn’t random.

Many behaviors occur in swarming bacteria that don’t occur in swimming bacteria. The leading edge cells may secrete surfactant, a combination of chemicals that reduce the surface tension on the plane so that the bacteria can move with less resistance.

The leading edge bacteria grow extra flagella, become elongated, and secrete slime for easy movement - but only the leading edge cells. They band together, becoming like rafts; in fact that’s what they’re called, rafts. The movement of the leading edge plows a furrow in the material they're moving across. This is partly due to the leading edge cells, but it has more to do with the cells behind them. The following cells form roiling masses, and together they push the leading edge along, like pushing a plow to form a ditch for planting seeds.

The furrows are then followed and expanded by the cells behind the leading edge, growing larger and easier to follow. That way, they can push the leading edge better. All these changes and functions lead to faster movement, which is why the swarm can move faster than individuals.

One amazing thing discovered in a 2013 series of experiments was that the leading edge cells secrete DNA. This nucleic acid doesn’t function as genetic material, but is apparently important for keeping the leading edge cells together and moving in the same direction, as well as stimulating movement at all. In experiments where this DNA was chewed by enzymes, the swarming movement stopped completely. Amazing - if they were a marching band in a parade, the DNA would be the banner carried by the drum majors that's emblazoned with their school and nickname. Everybody follows the banner and the drum major.

Integral to the concepts of swarming and biofilm development is the idea of multicellularity in bacteria. They're all clones of one another (except for mutation and any lateral gene transfer), but they work together and may take on different jobs, structures, and morphologies. They are working together to accomplish more than they could on their own. That sounds a lot like a multicellular organism where the different cell specialize into different types in order to perform different functions.

On the left is a cartoon that illustrates how the electron
donor hydrogen sulfide can’t donate electrons unless
something is available to accept them. The oxygen is the
acceptor, and the bacteria provide the cable to connect
them. The filaments of bacteria are shown on the right.
Photocredit to Nils Risgaard-Petersen.
One example comes from a 2012 study. Sea floor bacteria that bridge an area of high oxygen and low hydrogen sulfide to one of low oxygen and high hydrogen sulfide actually form filaments that act as power cables. Electron pass long a length of millions of cells to complete a circuit between the two sets of cells and this provides the energy to make ATP. Bacteria seem to work together in tough environments better than humans do on our best day.

We don’t know all the bacteria that are capable of swarming, but it's probably many more than we have found so far. And we aren't sure just why do they do it. Perhaps it's to leave an area of poor food value behind and strike out for better hunting grounds. Moving faster than they would as individuals might be important when trying to find, and then take advantage of a new food source. Eat up before someone else finds it.

Perhaps swarming is for protection. Like for biofilms, there is evidence that bacteria are less susceptible to antibiotics when swarming. Or it may have something to do with the best way to achieve full biochemical development. There are many studies that suggest that infectious organisms must swarm in order to create disease. Please remember, they aren’t trying to cause disease, but it shows that swarming must be important in their colonial development and a byproduct of this may be disease.

Three colonies of the same bacteria that were not clonal (not
from same exact ancestor - A, B, and C) were grown on the same
plate and they expanded in a swarm-like behavior. Where the
different colonies meet is the Dienes line. On the right is a false
color close up of a Dienes line, showing the battlefield. The
black line is 50 µm long.
They may also swarm to protect a new environment. Bacteria from one colony that grow and begin to swarm can tell their brethren apart. They can even discriminate between bacteria of the same type that have come from separate colony. When the two colonies swarm, they set a boundary between them, called a Dienes Line. A 2013 study showed that in Proteus mirabilis, a bacterium that causes urinary tract infections (UTIs), this boundary is really a battleground.

P. mirabilis has the ability to produce a type VI secretion system that acts as a needle. It punctures an adjacent bacterium and injects toxins. When a swarming colony invades another colony, they all start to produce their type VI secretion needles.

They attack any cell that makes contact with them, in a preemptive sort of fashion. There are many friendly fire incidents, but kin will survive the attack while cells from the other colony will be killed (they aren't immune to the specific toxin). The deeper invader is usually the dominant colony and will kill off the other colony, even though they may be of the same strain. Man - bacteria can be ruthless.

The key to both biofilm development and swarming is quorum sensing (quorum is from Latin qui meaning who, it means the number of members that must be present to transact business). The bacteria sense when their numbers reach a certain tipping point because the levels of certain chemicals reach critical concentrations.

We aren’t sure just why one behavior happens instead of the other, the situations that will induce either biofilm formation or swarming, but the number of bacteria and the state of their environment is key. Therefore, if you can stop the quorum sensing, you can stop swarming or biofilm formation, or both. This would be key to battling some pretty nasty infectious organisms since we said they are often important for pathogenesis.

Proteus mirabilis is a bacteria that swarms in concentric
circles. It causes urinary tract infections in both men and
women. In the lower image you can see the many flagella
of the organism – and this is before it starts to swarm and
leading edge organisms differentiate.
Several recent studies (here and here for example) have shown that certain natural or man made chemicals have the ability to interrupt quorum sensing or swarming/biofilms. Even cranberries seem to do the job.

We have discussed in prior posts about the amazing ability of cranberry to prevent UTIs. A 2013 paper shows that at least part of the cranberry's action on UTI-causing P. mirabilis is through the prevention of swarmer cell differentiation. Work with other bacteria shows that it is quorum sensing that is disrupted by the cranberry compounds, so the swarm in P. mirabilis might be stopped via the bacteria not knowing how many of their brothers are around. Bacteria won't pick a fight unless they know their gang is big enough - it's West Side Story in your bladder.

Next week - some prokaryotes don't move. Just like couch potatoes, they wait for someone to bring them their dinner.

Gloag ES, Turnbull L, Huang A, Vallotton P, Wang H, Nolan LM, Mililli L, Hunt C, Lu J, Osvath SR, Monahan LG, Cavaliere R, Charles IG, Wand MP, Gee ML, Prabhakar R, & Whitchurch CB (2013). Self-organization of bacterial biofilms is facilitated by extracellular DNA. Proceedings of the National Academy of Sciences of the United States of America, 110 (28), 11541-6 PMID: 23798445

Deng P, de Vargas Roditi L, van Ditmarsch D, & Xavier JB (2014). The ecological basis of morphogenesis: branching patterns in swarming colonies of bacteria. New journal of physics, 16, 15006-15006 PMID: 24587694

McCall J, Hidalgo G, Asadishad B, & Tufenkji N (2013). Cranberry impairs selected behaviors essential for virulence in Proteus mirabilis HI4320. Canadian journal of microbiology, 59 (6), 430-6 PMID: 23750959

Alteri CJ, Himpsl SD, Pickens SR, Lindner JR, Zora JS, Miller JE, Arno PD, Straight SW, & Mobley HL (2013). Multicellular bacteria deploy the type VI secretion system to preemptively strike neighboring cells. PLoS pathogens, 9 (9) PMID: 24039579

For more information or classroom activities, see:

Quorum sensing –

Biofilms –

Bacterial swarming -