December turned out to be a rather hectic month for several reasons, so I decided to take a break from blogging. Now the holidays are over, I will back to regular blogging for 2013!

In a previous post I wrote about how two-component systems evolved in bacteria while dying out in animals, so for this week I thought I’d give a recent example of work on a two-component system in Pseudomonas aeruginosa. P. aeruginosa is a dangerous human pathogen that is particularly notorious for forming biofilms – a large bacterial city, consisting of lots of bacteria all held together within a sticky mess.

P. Aeruginosa from wikimedia commons.

P.  aeruginosa has a large number of two-component systems, many of which are involved in managing the transition from the free-floating planktonic state to the colony-based community lifestyles of the biofilm. In particular the researchers were looking at a system labelled “PprAB” where PprA is the sensor (which recieves the input) and PprB is the reciever (which creates the output by turning on certain genes).

Activating this PprAB system switches the bacteria from free-living single cells to a large multi-cellular biofilm. PprB activation is also associated with increased susceptibility to antibiotics and reduced virulence. As most biofilms are incredibly antibiotic resistant and usually increase virulence, the researchers decided to take a closer look at the PprAB system and what it was activating.

In order to do this, the researchers used a mutant strain of the bacteria labelled PprBK which constantly expressed PprB. They compared the growth of this strain to a normal wild type bacteria labelled PAO1. They found that the cells constantly expressing PprB (the PprBK strain) formed biofilms a lot faster than the wild type,with cells clumping together after just one day:

Biofilm formation after one day - wild-type cells on the left, PprB expressing cells on the right, which are already starting to clump! Image from the reference.

They also tested the virulence of the two strains in human HeLa cells using an LDH release assay. The more LDH is released, the more the human cells are damaged and dying. The graph below shows the effects of both strains on cell death over the course of three hours, with the wild-type PAO1 far more virulent and leading to more cell death:

Statistical differences evaluated using a t-test (*:<0.05, **:<0.0.01, ***:<0.001). Image from the reference below.

Another observation that the researchers made was that this biofilm is more susceptible to the antibiotic tobramycin. Several of the genes that the PprAB system activates are used to increase the permeability of the cell membrane (to allow signal molecules in so all the cells can talk to each other and form a biofilm at the same time) and to change the composition of the cell matrix. it looks like trying to form a biofilm quickly makes each cell more permeable, allowing antibiotic molecules easier access inside. The speed of biofilm formation may also be related to the loss of virulence - rather than forming a coordinated attack, the cells are just trying to build a city as quickly as possible.

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Reference: de Bentzmann S, Giraud C, Bernard CS, Calderon V, Ewald F, et al. (2012) Unique Biofilm Signature, Drug Susceptibility and Decreased Virulence in Drosophila through thePseudomonas aeruginosa Two-Component System PprAB. PLoS Pathog 8(11): e1003052. doi:10.1371/journal.ppat.1003052