Influence of overexpression of the phosphodiesterase YhjH on E. coli biofilm formation and composition

To better survive, most bacterial species gather in communities called biofilms which form on most every type of surface. Although biofilm is beneficial for the survival of these microbes, bacteria in biofilms are notably more tolerant to antibiotics, and their biomass can occlude pipes and contaminate storage containers, posing serious problems for public health and industry. We are interested in manipulating signaling pathways and studying their impact on biofilm formation and composition. Earlier work by our laboratory demonstrated that overexpression of the transcription factor RpoF significantly increased biofilm growth at warmer temperatures. The E. coli cyclic-di-GMP phosphodiesterase YhjH, a downstream target in the RpoF signaling pathway, has two distinct and opposite roles in biofilm formation. YhjH hydrolyzes the second messenger cyclic-di-GMP, leading to the inhibition of curli, extracellular structures that promote the adhesion and maturation of biofilm, thus ultimately inhibiting biofilm growth. Secondly, it can boost the growth of Type I pili, extracellular structures that promote biofilm growth. Here, we overexpress the phosphodiesterase YhjH in E. coli and measure how it affects biofilm growth and composition of the extracellular polymeric substance (EPS). We grew biofilm on glass wool and measured its growth over time using a crystal violet assay. We supplemented colorimetric biochemistry assays with confocal microscopy to measure the concentrations of different biomolecules in the EPS, such as carbohydrates, proteins, and extracellular DNA. Significantly less biofilm grew with YhjH overexpression at both warmer (37 C) and cooler (28 C) growth temperatures, and preliminary results show minimal differences in the composition of the EPS with these growth conditions. These results suggest that the RpoF overexpression\u27s influence on increased biofilm growth happens via a mechanism separate from YhjH activity