Acid exposure induces adaptive resistance in <i>Klebsiella pneumoniae</i> to combination treatment with antibiotics and biofilm-disrupting surgical lavage

ABSTRACT Surgical irrigation solutions remove bacteria and disrupt biofilms to prevent nosocomial infections. This study evaluated the impact of combined treatment of a biofilm-disrupting lavage solution and antibiotic therapy. Bacteria treated with antibiotics and/or lavage were evaluated via broth microdilution and checkerboard assays. Staphylococcus aureus was strongly inhibited by all doses of surgical lavage, while combination treatments of Escherichia coli increased antibiotic susceptibility by more than 10-fold. However, combination treatments of Klebsiella pneumoniae resulted in specific dose regimens with diluted lavage solution that eliminated the bacteriostatic effect, such that significant bacterial growth occurred at 4× minimum inhibitory concentration treatments. This effect was observed across multiple strains of K. pneumoniae and multiple classes of antibiotics. Analysis of the individual components of the lavage solution revealed this to be a pH-related effect. Klebsiella grown in pH 4.5 media replicated the observed increase in minimum inhibitory concentration (MIC) without the addition of the other lavage components. This adaptive resistance response was correlated with activated stress responses, alterations in membrane polarization, and altered transcription of outer membrane porins. Transcription of ompA and lpp , which stabilize the outer membrane, was upregulated, while transcription of the alternative maltoporin lamB was halted. Deletion of the iroA siderophore also enhanced growth in bacteriostatic concentrations of antibiotics at low pH. Together, these data strongly suggest that Klebsiella has a distinct ability to mount a rapid adaptive response to an acidic environment via changes in membrane permeability, porin, and siderophore gene expression to enhance survival in the presence of both disinfecting agents and antibiotics. IMPORTANCE Disinfecting washes that disrupt biofilms are commonly used in hospital settings to prevent the establishment of bacterial infection. This study investigated how combined exposure to a biofilm-disrupting surgical wash and antibiotics affected bacterial susceptibility to these bacteriostatic agents. While the response to Staphylococcus aureus and Escherichia coli was additive and fully bacteriostatic, we found specific combinations of antibiotic and diluted disinfecting agent that enhanced bacterial growth of Klebsiella pneumoniae , evidence of an adaptive resistance response. This response was found to be specific to Klebsiella , was initiated by a specific acidic pH range, and required altered transcription of major porins. These data indicate that Klebsiella can adapt to an acidic pH in ways that enhance resistance to both antibiotics and disinfectant agents. This adaptive response may be an important precursor to the evolution of fully resistant strains of Klebsiella via porin loss.