Signature gene expression profile of triclosan-resistant Escherichia coli

Objectives: To gain further insight into the defence mechanisms against triclosan in a mutant derived from an Escherichia coli strain carrying the triclosan-resistant target enzyme, FabI(G93V). Methods: An E. coli imp4231 FabI(G93V) strain was constructed by replacing intact fabI with a linear DNA cas-sette, fabI(G93V)-CmR, that contains a single mutation, GGT to GTT, at codon 93 of fabI(G93V) and a chloram-phenicol resistance gene (CmR) as a marker for the mutant allele by a Red-mediated recombination system. Using this E. coli imp4231 FabI(G93V) strain, nitrosoguanidine (NTG) mutagenesis was performed to generate E. coli IFNs [imp4231 FabI(G93V) treated with NTG] displaying higher MICs of triclosan than its parent strain. The genes overexpressed in E. coli IFN4 were identified by DNA microarray analysis. Results: An E. coli imp4231 FabI(G93V) strain displays 400-fold increased MICs of triclosan (MIC8 mg/L) compared with the parent strain (MIC0.02 mg/L). Furthermore, E. coli IFN4 has the highest MIC of triclosan (MIC80 mg/L). DNA microarray analysis of E. coli IFN4 shows that many genes involved in the biosynthesis of membrane proteins, including transporters, reductases/dehydrogenases and stress response regulators, were highly expressed in the mutant. Conclusions: These results strongly indicate that E. coli IFN cells might protect themselves from triclosan by activating various defence mechanisms, such as (i) changing efflux activities; (ii) capturing the triclosan; and (iii) increasing the expression of important regulators or metabolic enzymes