Rapid method of luxS and pfs gene inactivation in enterotoxigenic Escherichia coli and the effect on biofilm formation.

Abstract

Rapid and efficient inactivation of a target gene in Escherichia coli chromosomes is required to investigate metabolic engineering. In the present study, a multiple gene inactivation approach was demonstrated in four strains of enterotoxigenic E. coli (ETEC), which are the predominant pathogenic bacteria causing piglet diarrhea, mediated by λ Red and Xer recombination. The chromosomal genes, luxS and pfs were inactivated using the multiple gene inactivation approach in the wild‑type strains of E. coli, K88, K99, 987P and F41. This indicated that dif sites may be reused to inactivate multiple chromosomal genes when no antibiotic‑resistant selectable markers remain. Following inactivation of luxS and pfs, the ability of ETEC to produce the quorum sensing signal, and induce auto‑inducer 2 activity and biofilm formation were significantly reduced. Furthermore, the multiple gene inactivation approach also exhibits a high recombination efficiency and follows a simple process.