Dear Yumeng,

Your research aims are very complex and it will be interesting to see how far you get!

We have in the past tried to separate different effects or phenotypes when assessing biofilm-formation by pseudomonads - our model bacterium, P. fluorescens SBW25 is motile, capable of swimming, twitching and swarming, it can form weak air-liquid interface biofilms when non-specifically induced by iron, and mutants capable of forming more robust biofilms usually appear within 3 days of incubation. The wild-type strain can express cellulose, PNAG, and alginate, and we know that cellulose and eDNA are present in biofilms - it also expresses a strong surfactant.

I think that the only way to try and work out whether a particular phenotype (i.e. cell behaviour or the expression of a particular compound) or outcome (i.e. growth in a microcosm or biofilm biomass) is important can only be determined by comparing a number of single and multiple mutants and under a variety of conditions, and to assess the relative importance of each using an appropriate statistical model.

Surface properties can be measured directly, and then by changing buffer conditions or culture age you will be able to see how sensitive these properties are to changes. Although it is very difficult to remove surfactants from a culture, it is possible to add back surfactants (semi-purified or by using culture supernatants) to a mutant that cannot produce these compounds. Depending on your bacterium, it might be possible to inhibit swimming using an ionophore (e.g. CCCP), but this will also impact on growth rates. Perhaps for this you will need a flagella mutant to experiment with.

We have also looked at the rheological properties of our biofilms, and found that even cultures of bacteria had measurable viscosity-elastic features.

Good luck! Andrew