Exacerbation of virulence of multi-drug resistant Escherichia coli O104:H4 by subinhibitory concentrations of ampicillin

Little is known about how subinhibitory concentrations of antibiotics to which bacteria are resistant affect bacterial virulence. In this study, the effect of subinhibitory concentrations of ampicillin on the virulence of E. coli O104:H4 was analyzed. Bacteria were pre-exposed to 0.1, 0.3, or 0.5 mg/mL of ampicillin in LB media and incubated for 4 h at 37 °C. Transformation capacity (using plasmids and PCR-amplified DNA sequences), swarming motility, biofilm production, curli formation, and virulence gene expression were determined. Ampicillin increased the transformation of E. coli O104:H4, with the highest number of transformants (>104 CFU/ng DNA; p ≤ 0.05) detected after exposure to DNA sequences of spectinomycin. In addition, bacteria pre-treated with 0.5 mg/mL of ampicillin exhibited higher swarming motility (7.6 cm, vs 6.0 cm for control; p ≤ 0.05) and biofilm production (up to 1.9-fold; p ≤ 0.05) when subsequently exposed to 0.1 and 0.3 mg/mL of antibiotic compared with the control. Also, significant overexpression of the virulence-related genes flhC (≤16.1-fold), fliA (≤22.1-fold), csgA (≤3.6-fold), csgD (≤9.1-fold), stx2a (≤32.2-fold), and the antibiotic resistance gene blaTEM-1 (≤5.5-fold) was observed. In conclusion, ampicillin-resistant E. coli O104:H4 increased the expression of its virulence factors when exposed to most subinhibitory concentrations of ampicillin analyzed in this study.