Abstract
The interplay between membrane permeability alterations and the enzymatic barrier contributes to Klebsiella pneumoniae multidrug resistance. We assessed the specific effect of the efflux levels of the main efflux pumps (AcrAB and OqxAB), alone and associated with the loss of the main porins (OmpK35 and OMPK36), on the activity of various antibiotics by constructing a set of K. pneumoniae isogenic strains, including strains with plasmid-mediated β-lactamases (DHA-1, CTX-M-15, and OXA-48). The two pumps contributed to intrinsic chloramphenicol resistance and AcrAB to that of nalidixic acid and cefoxitin, whereas they had no impact on the activity of the other 11 antibiotics tested. We confirmed the expulsion of these three antibiotics by the two overproduced pumps and that of tigecycline by overproduced AcrAB, and showed that overproduced AcrAB also expelled ertapenem, piperacillin, ceftolozane, and ceftazidime. The sole loss of porins did not significantly affect the activity of the tested antibiotics, except ertapenem. The effect of efflux increases and porin loss on β-lactam activity was the highest in plasmid-mediated β-lactamase-producing strains. Thus, DHA-1-producing strains became non-susceptible (NS) to (i) ertapenem when there was an increase in efflux or porin loss, (ii) imipenem and ceftazidime+avibactam when the two mechanisms were associated, and (iii) temocillin when AcrAB was overproduced. The CTX-M-15-producing strains became NS to (i) ertapenem when there was no porin, (ii) ceftolozane+tazobactam when there was either overproduced OqxAB or porin loss, and (iii) temocillin when AcrAB was overproduced. OXA-48-producing strains known to be NS to temocillin were also NS to ceftolozane and they became NS to imipenem when the two pumps were overproduced or there was porin loss. Overall, this study shows that the balance between influx and efflux differentially modulates the activity of the tested antibiotics, an important point for evaluating the activity of future antibiotics or new combinations.
Highlights
Klebsiella pneumoniae is a pathogen responsible for a wide range of nosocomial infections (Podschun and Ullmann, 1998; Wyres and Holt, 2016)
Imipenem, piperacillin, piperacillin+tazobactam, ceftolozane, ceftolozane+tazobactam, ceftazidime, ceftazidime+avibactam, and temocillin susceptibility of isogenic strains transformed with plasmids encoding the most commonly identified β-lactamases of AmpC type, i.e., DHA-1, ESBL type, i.e., CTX-M-15, and carbapenemase type, i.e., OXA-48-like, in European K. pneumoniae isolates (Baraniak et al, 2013; Potron et al, 2013; Freitas et al, 2014; Rodrigues et al, 2014; Ruiz-Garbajosa et al, 2016) in terms of clinical categorization
In addition to the diversity of β-lactamases involved in the acquired resistance to β-lactams in K. pneumoniae clinical isolates, membrane permeability alterations have been observed in isolates resistant to multiple antibiotics (Hasdemir et al, 2004; Dahmen et al, 2012; López-Camacho et al, 2014)
Summary
Klebsiella pneumoniae is a pathogen responsible for a wide range of nosocomial infections (Podschun and Ullmann, 1998; Wyres and Holt, 2016). It is the enterobacterial species in which plasmid-mediated resistance to extended-spectrum β-lactams related to extended-spectrum β-lactamases (ESBL) (Jarlier et al, 1988), cephalosporinases (Park et al, 2013; Freitas et al, 2014), and carbapenemases (Nordmann et al, 2011; Robert et al, 2014) first emerged and became widely disseminated It has been included in the ESKAPE group, which clusters the main bacterial species (i.e., Enterococcus faecium, Staphylococcus aureus, K. pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species) with a propensity for multidrug resistance (Pendleton et al, 2013). Several new variants were designed to assess the previously suggested role of the regulators of efflux pump gene expression on the production of general porins in K. pneumoniae (De Majumdar et al, 2013, 2015): RamA and RamR, which activate or repress acrAB gene expression, respectively, and RarA and OqxR, which activate or repress, oqxAB gene expression, respectively (Bialek-Davenet et al, 2013, 2015)
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