Quorum Sensing

4 Notes

1. With respect to the Iglewski PAO1, we adjust the beads to a
   concentration of 1.5 108 CFU/ml.


2. A power analysis should be made to estimate the group sizes.


3. This is the concentration we use, but it has to be adjusted
   depending on the mouse strain. Please consult other references
   regarding this subject.


4. Usually the mice appear sick within the first 24 h after infection.
   We believe that this is due to the fact that they have been given
   pneumonia and anesthetics.


5. In case of the Iglewski PAO1, homogenates from most of the
   mice will display CFUs when diluted to 10^3. If no CFU can be
   counted the mice were inoculated with too little beads.


6. Bacteria from lung tissues containing mixed inoculates ofP.
   aeruginosaandE. colican be discriminated based on the colony
   color on the blue agar plates combined with the ability ofE. coli
   colonies to express GFP in the presence of exogenous AHLs.
   Therefore the plates can be supplemented with 10 nM 3-oxo-
   C 6 -HSL.


7. The experiments should not run for longer than 3 days since it
   is not possible to detect theE. colimonitor stain.


8. Make sure that the hexane is cold enough.


9. Ensure that what are observed under the microscope are in fact
   bacteria. This can be verified by the morphology and by look-
   ing at the bacteria under normal light.

Acknowledgement

We acknowledge the contributions made by Drs. Morten Hentzer,

Hong Wu, Thomas Bovbjerg Rasmussen, Jens Bo Andersen, and

Allan Bech Christensen with regard to the initial developments of

the model systems.

References

1. Costerton JW, Stewart PS, Greenberg EP
   (1999) Bacterial biofilms: a common cause of
   persistent infections. Science 284:1318–1322


2. Høiby N (1974) Pseudomonas aeruginosa
   infection in cystic fibrosis. Relationship
   between mucoid strains ofPseudomonas aeru-
   ginosaand the humoral immune response. Acta
   Pathol Microbiol Scand B Microbiol Immunol
   82:551–558
   3. Singh PK, Parsek MR, Greenberg EP, Welsh
   MJ (2002) A component of innate immunity
   prevents bacterial biofilm development. Nature
   417:552–555
   4. Bjarnsholt T, Jensen PO, Burmølle M, Hentzer
   M, Haagensen JA, Hougen HP et al (2005)
   Pseudomonas aeruginosatolerance to tobramy-
   cin, hydrogen peroxide and polymorphonuclear

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