In my experiment I study biofilm formed by Pseudomonas aeruginosa. And the strains include Pseudomonas aeruginosa PAO1 and several clinical strains and their ability to form biofilm remains stable. But in recent study, the biofilm suddenly decreases. I resuscitated new strains from the refrigerator, changed the 96-well plates and new medium. But they all did not work. So I hope to get some suggestion for what I need to do next and why. Thank you!
There can be non-mucoid reversion. May occur at higher frequency in some strains. Check the C:N ratio of your medium and its respective prep to ensure consistency. As medium is a common factor across your strains. Check plate batch to ensure same and that a different coating has not been used.
Conference Paper Effects of Growth Conditions and Pretreatment on the Iodine ...
SIMPLE THINGS TO CHECK: I immagine the experiments were "independent"... so the inoculums were different... even if you check O.D. , this can make a big difference.
Also, are you sure physical factors parameters were stable during incubation? If people keep on opeing and closing an incubator... temperatures flactuate a lot.... and this influences biofilm formation a lot for something like Pseudomonas aeruginosa...
What assay are you performing: crystal violet? Viable counts (MBEC)? etc..
To give you a comprehensive explanation for what might be happening I would need more information such as:
(1) what medium are you using and what did you use before
(2) current growth conditions vs previous growth conditions
(3) what assays did you use to determine biofilm formation?
Strains that are kept in a refrigerator comes a time when both remove them and put them to the refrigerator those successive changes in temperature cause bacteria to weaken, this may be a difference between your previous experiments and new. fresh strains obtained from infectious processes are generally good forming biofilms long as they comply with all the ideal conditions for this. Your old vines can not miss them back by successive passes in blood Agar to strengthen them before making the protocol. Something important in biofilm formation is concentration of glucose in the medium and the incubation temperature.
I hope I have helped you
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