I'm working with Actinobacillus pleuropneumoniae, one that really knows how to form biofilm. My log growth curve is looking good the first hours but is then stagnating or even getting reduced some. after about 2 hour stagnation it is growing again. Can biofilm formation play a role in this? I don't really know if I can "trust" the growth and expect it to handle antibiotic in an optimal way.
If the walls of the growth vessel/ flask appeared to be hazy then yes ! Probably the culture began forming a biofilm...and if it is true you might experience issues of proper mixing prior to plating on solid culture medium.
Biofilm can definitely cause a difference in your growth curve. This would be especially problematic if you are using a 96-well plate and reading the OD of it to judge your growth. The typical determination of growth by optical density is best for planktonic cells which should be evenly distributed in your culture. Cells that clump or aggregate would affect your reading. If you are doing your experiment in culture test tubes or flask, its possible that vortexing you sample could reduce the clumping. Alternatively, you could try plating dilutions to confirm your viable cells at each time point.
thanks for your answers. I have a large continous culture (200ml) and My growth curve is viable count (Cfu/ml). Ap is a fastidious bacteria but I use lysed hotse blood, NAD and yeast extract as supplements. I see the special growth curve in My in vitro system. I am thinking that the biofilm has easier to form here because more uneven surfaces and less movement of media. I have outlet tube and sample tube in the vessel. The vessel is stansning in A waterbath with shake ( onedirectional..)
I noticed that you are using lysed horse blood to grow your bacteria. Is the horse blood in the sample you are using to measure OD? Probably is biofilm but also, could be still intact RBCs or partial RBCs floating around and interfering with the reading? Just a thought.
Of course biofilm would affect your determination of a growth curve. Please consider that the culture is not homogenous suspension of cells but at least two distinct populations -. Those that are biofilm-associated and those as single cells. The aliquot you remove is representative only of the latter population
Thanks for your answers! So I could publish results from kinetic time kill experiments although the growth control looks a bit strange?
I'm not counting the biofilm bacteria but I guess it's an equilibrium between the biofilm and the bacteria in solution. And if there were biofilm in the bottle before adding the antibiotic, this will probably be resistent and might change My results. The bacterial Killing is looking good but the regrowth is fast in all concentrations. Although the final Cfu at 24 h are different between concentrations. I bougth teflon equipment to reduce biofilm but it didnt help. Do you know any biofilm reducing material that I can coat
Dear Larberg,AP have minimal effect on growth curve.You should be more concerned with antibiotic resistant of biofilm forming bacteria.Microorganisms found in biofilms have their antibiotic resistance increased to up to 1000 times when compared to free living bacteria of same species.Good luck with your research.Rgd.Murtaza Mustafa
Dear Mustafa, thank you for your answer. I'm not quite sure I' understand "AP have minimal effect on growth curve" Regards Astrid
Do continual exposures of biofilm pathogens to a given disinfectant emerge their resistance to antibiotics? "
Hi this is my thesis question to develop. please can someone recommend, any useful journal. thank you
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