Characterisation and optimisation of biofilm and pellicle formation by Escherichia coli K-12

Bacteria form biofilms on solid surfaces by using surface adhesins and by secreting extracellular matrix components. While many biofilms are harmful, for example in clinical and industrial settings, biofilms have been shown to improve yield in biocatalysis reactions generating pharmaceutical precursors. In this work, biofilm formation was studied in Escherichia coli K-12 PHL644, which is a known biofilm- forming strain that has been previously used for biocatalysis. In order to generate a physically robust biofilm, promoter-green fluorescent protein reporters were used to optimise the expression of curli, an adhesin important in biofilm formation, in response to a variety of stimuli and physical conditions. Curli expression was greatest in planktonic cells grown in minimal medium supplemented with 10 mM glucose at 30 °C and 70 rpm shaking. The effect of aromatic amino acids on biofilm formation was investigated, and the addition of phenylalanine to cultures was found to increase curli expression and optical density. It was hypothesised that phenylalanine forms amyloid- like fibres which seed curli formation in E. coli; however this needs to be investigated further. Potentially, phenylalanine may be used to make biofilm formation a tuneable system. During work to further improve biofilm formation, a biofilm floating at the air- liquid interface, called a pellicle, was observed. E. coli K-12 pellicle formation was optimised and characterised in terms of curli expression, motility, and secretion of matrix components and it was found that pellicle formation requires curli. The potential for a biofilm at the air-liquid interface may be advantageous in certain industrial settings