Novel bio-recognition elements (BREs) for biosensing and differentiation of Gram-positive bacteria in hydrocarbon environments

Excessive microbial growth in fuel and fuel systems can lead to biofouling and biocorrosion, which makes early detection crucial. A wide range of phylogenetically and metabolically diverse microorganisms have been isolated from environments exposed to hydrocarbons with Gram-positive and Gram-negative bacteria being ubiquitous. The aim of this work was to identify specific biomarker-based bio-recognition elements (BREs) for sensing hydrocarbon-degrading Gram-positive bacteria. We used two bacterial surface targets that are conserved and specific for Gram-positive bacteria, Lipoteichoic acid (LTA) and a short surface-epitope region from the cell wall-associated secreted protein Fasciclin 1 (FASC1), from Gordonia sihwensis. Several peptide-based ligands to detect numerous Gram-positive bacteria in fuel were discovered by using phage display technology and the binding of these peptides to Gram-positive bacteria in the presence of hydrocarbons was validated using Microscale Thermophoresis (MST) and Western blot analysis. The binding peptides were conjugated to quantum dots and their specificity to Gram-positive bacteria was characterized by fluorescence imaging and fluorometric analysis in the presence of target and non-target microorganisms. It was shown that jet fuel did not interfere with binding of the peptides to the target biomarker in the bacteria, indicating that these BREs are suitable for functionalization of biosensors for detection of Gram-positive bacteria in hydrocarbon rich environments.