A lux/gfp dual label system for studying attachment and biofilm formation of Enterobacter sakazakii

Abstract

A significant feature of Enterobacter sakazakii is the ability to form biofilms enabling its persistence in manufacturing and neonatal environments, and implicating it as the primary cause of outbreaks of neonatal meningitis. Conventional methods of studying E. sakazakii are time consuming, inaccurate and damaging to cells, preventing further analysis. A novel method for detecting biofilm formation has been utilized that takes advantage of a dual lux/gfp reporter system cloned into cells for simultaneous quantification of bacterial metabolic activity and cell numbers respectively. To evaluate the effectiveness and accuracy of the novel method compared to the conventional method, strains of bacteria were allowed to form biofilms and were measured using both methods. Biofilm formation of E. sakazakii strains over 2 days was measured and peak formation and changes in biofilm development was determined for each strain. The lux reporter was utilized to determine metabolic activity of cells in the biofilm, correlating to biofilm formation, over 3 days. Biofilm formation of strains was measured using both methods and compared to detect trends. This study found that the novel method was effective at detecting changes in metabolic activity of cells in biofilm. Cell numbers were to be simultaneously detected via the gfp reporter but filters with correct detection wavelength were unavailable thus cell numbers were not obtained here. These observations determine that this dual reporter system is a promising tool for monitoring bacteria in situ and for further understanding of biofilm formation.