Effects of l-arabinose efflux on λ Red recombination-mediated gene knockout in multiple-antimicrobial-resistant Salmonella enterica serovar Choleraesuis

• Published in: Archives of microbiology Vol. 200; no. 2; pp. 219 - 225
• Main Authors: Liao, Shi-Wei, Lee, Jen-Jie, Ptak, Christopher P., Wu, Ying-Chen, Hsuan, Shih-Ling, Kuo, Chih-Jung, Chen, Ter-Hsin
• Format: Journal Article
• Language: English
• Published: Berlin/Heidelberg Springer Berlin Heidelberg 01.03.2018; Springer Nature B.V
• Subjects: Antibiotic resistance; Antibiotics; Antiinfectives and antibacterials; Antimicrobial agents; Arabinose; bacteriophages; Biochemistry; Biomedical and Life Sciences; Biotechnology; Cell Biology; dose response; Ecology; Efflux; fluorescent dyes; Functional analysis; Gene expression; gene targeting; Genetic analysis; genetic engineering; Life Sciences; Microbial Ecology; Microbiology; Original Paper; Recombinase; Recombination; Salmonella; Salmonella choleraesuis Choleraesuis; Strains (organisms); Substrates; Swine; transporters; Virulence; Choleraesuis; Arabinose; λ Red recombination; Antimicrobial resistance; Gene knockout; Salmonella Choleraesuis; L-Arabinose
• ISSN: 0302-8933; 1432-072X; 1432-072X
• DOI: 10.1007/s00203-017-1436-4

In this study, six swine-derived multiple-antimicrobial-resistant (MAR) strains of Salmonella Choleraesuis ( S . Choleraesuis) were demonstrated to possess higher efflux pump activity than the wild-type (WT). l -Arabinose, a common inducer for gene expression, modulated S . Choleraesuis efflux pump activity in a dose-dependent manner. At low l -arabinose concentrations, increasing l -arabinose led to a corresponding increase in fluorophore efflux, while at higher l -arabinose concentrations, increasing l -arabinose decreased fluorophore efflux activity. The WT S. Choleraesuis that lacks TolC (Δ tolC ), an efflux protein associated with bacterial antibiotic resistance and virulence, was demonstrated to possess a significantly reduced ability to extrude l -arabinose. Further, due to the rapid export of l -arabinose, an efficient method for recombination-mediated gene knockout, the l -arabinose-inducible bacteriophage λ Red recombinase system, has a reduced recombination frequency (~ 12.5%) in clinically isolated MAR Salmonella strains. An increased recombination frequency (up to 60%) can be achieved using a higher concentration of l -arabinose (fivefold) for genetic manipulation and functional analysis for MAR Salmonella using the λ Red system. The study suggests that l -arabinose serves not only as an inducer of the TolC-dependent efflux system but also acts as a competitive substrate of the efflux system. In addition, understanding the TolC-dependent efflux of l -arabinose should facilitate the optimization of l -arabinose induction in strains with high efflux activity.