We tested the effects of various putative efflux pump inhibitors on colistin resistance in multidrug-resistant Gram-negative bacteria. Addition of 10 mg/liter cyanide 3-chlorophenylhydrazone (CCCP) to the test medium could significantly decrease the MICs of colistin-resistant strains. Time-kill assays showed CCCP could reverse colistin resistance and inhibit the regrowth of the resistant subpopulation, especially in Acinetobacter baumannii and Stenotrophomonas maltophilia. These results suggest colistin resistance in Gram-negative bacteria can be suppressed and reversed by CCCP. Colistin is regarded as the last resort antibiotic for infections caused by multidrug-resistant (MDR) and extensively drugresistant (XDR) Gram-negative bacteria. However due to colistin's widespread use, resistant strains are increasingly being isolated in the clinic (1). Resistance to colistin has been reported to be only chromosomally mediated. However, very recently, the finding of a transferable plasmid-mediated colistin resistance gene, mcr-1, suggests the possibility of quick acquisition of resistance (2, 3). If the mcr-1 gene is similar to the case of NDM-1, colistin-resistant bacteria may soon become endemic in the world.Therefore, it is of paramount importance to prevent the dissemination of colistin resistance in an era that lacks new antibiotics against resistant Gram-negative pathogens. Besides, the finding and development of agents that effectively reverse resistance may be a promising strategy. Efflux pump inhibitors (EPIs) are potential agents in this category, and there have been initial reports on EPIs. A recent study showed that the addition of cyanide 3-chlorophenylhydrazone (CCCP) could decrease the MICs of colistin in Acinetobacter baumannii (4), but whether this phenomenon is strain specific or CCCP can reverse colistin resistance in Gram-negative bacteria deserves further investigation, and whether other commonly used EPIs have the same effects remains unknown. In order to answer these questions, we evaluated the effect of various putative EPIs on resistance to colistin in multidrug-resistant Gram-negative bacteria.Nonduplicate colistin-susceptible and colistin-resistant clinical isolates of Klebsiella pneumoniae, A. baumannii, Pseudomonas aeruginosa, and Stenotrophomonas maltophilia that also show resistance to at least other three antibiotic classes were collected between May 2014 and October 2015 from separate patients in three tertiary hospitals of Beijing, China. Colistin, CCCP, phenylarginine-␤-naphthylamide (PA␤N), 1-(1-naphthylmethyl)-piperazine (NMP), omeprazole, and verapamil standards were purchased from Sigma-Aldrich (Shanghai, China). Reserpine standards were obtained from the National Institute for the Control of Pharmaceutical and Biological Products, China (Beijing, China).MICs of EPIs, colistin, and colistin combined with EPIs were determined by the agar dilution method according to CLSI performance and interpretive guidelines (5). CCCP, NMP, and omeprazole were dissolved in dimethyl sulfoxide (DMS...

Few effective therapeutic options are available for treating severe infections caused by extensively drug-resistant Acinetobacter baumannii (XDR-AB). Using a murine thigh-infection model, we examined the in vivo efficacy of colistin in combination with meropenem, tigecycline, fosfomycin, fusidic acid, rifampin, or sulbactam against 12 XDR-AB strains. Colistin, tigecycline, rifampin, and sulbactam monotherapy significantly decreased bacterial counts in murine thigh infections compared with those observed in control mice receiving no treatment. Colistin was the most effective agent tested, displaying bactericidal activity against 91.7% of strains at 48 h post-treatment. With strains showing a relatively low minimum inhibitory concentration (MIC) for meropenem (MIC ≤ 32 mg/L), combination therapy with colistin plus meropenem caused synergistic inhibition at both 24 h and 48 h post-treatment. However, when the meropenem MIC was ≥64 mg/L, meropenem did not significantly alter the efficacy of colistin. The addition of rifampin and fusidic acid significantly improved the efficacy of colistin, showing a synergistic effect in 100% and 58.3% of strains after 24 h of treatment, respectively, while the addition of tigecycline, fosfomycin, or sulbactam did not show obvious synergistic activity. No clear differences in activities were observed between colistin-rifampin and colistin-fusidic acid combination therapy with most strains. Overall, our in vivo study showed that administering colistin in combination with rifampin or fusidic acid is more efficacious in treating XDR-AB infections than other combinations. The colistin-meropenem combination may be another appropriate option if the MIC is ≤32 mg/L. Further clinical studies are urgently needed to confirm the relevance of these findings.

Objectives Carbapenem-resistant Klebsiella pneumoniae (CR-KP) infections represent severe threats to public health worldwide. The aim of this study was to assess potential synergistic interaction between tigecycline and aminoglycosides via in vitro and in vivo studies. Methods Antibiotic resistance profiles and molecular characteristics of 168 CR-KP clinical isolates were investigated by susceptibility testing, PCR and MLST. Chequerboard tests and time–kill assays were performed for 20 CR-KP isolates to evaluate in vitro synergistic effects of tigecycline combined with aminoglycosides. A tissue-cage infection model of rats was established to evaluate in vivo synergistic effects. Different doses of tigecycline and aminoglycosides alone or in combination were administered for 7 days via tail vein injection. Antibiotic efficacy was evaluated in tissue-cage fluid and emergence of resistance was screened. Results The chequerboard tests showed that this combination displayed synergistic or partial synergistic activity against CR-KP. The time–kill assays further demonstrated that strong synergistic effects of such a combination existed against isolates that were susceptible to both drugs but for resistant isolates no synergy was observed if clinical pharmacokinetics were taken into consideration. The in vivo study showed that the therapeutic effectiveness of combination therapies was better than that of monotherapy for susceptible isolates, suggesting in vivo synergistic effects. Furthermore, combinations of tigecycline with an aminoglycoside showed significant activity in reducing the occurrence of tigecycline-resistant mutants. Conclusions Compared with single drugs, tigecycline combined with aminoglycosides could exert synergistic effects and reduce the emergence of tigecycline resistance. Such a combination might be an effective alternative when treating CR-KP infections in clinical practice.

Background: Cefiderocol is a promising antimicrobial agent against carbapenem-resistant, Gram-negative bacteria, but susceptibility data from the Chinese mainland are lacking. The aim of the present study was to test the susceptibility of cefiderocol against carbapenem-resistant, Gram-negative bacteria collected from Beijing, China. Methods: Carbapenem-resistant Klebsiella pneumoniae (CR-KP; n=105), carbapenem-resistant Acinetobacter baumannii (CR-AB; n=126), carbapenem-resistant Pseudomonas aeruginosa (CR-PA; n=74), and Stenotrophomonas maltophilia (SM; n=72) isolates were collected from inpatients at 4 tertiary hospitals in Beijing, China. Minimum inhibitory concentrations (MICs) for cefiderocol were determined using irondepleted cation-adjusted Mueller Hinton broth (CAMHB), and for comparators using CAMHB, according to the recommended Clinical and Laboratory Standards Institute (CLSI) methodology. Carbapenemase and other β-lactamase gene profiles were determined using polymerase chain reaction (PCR).Results: Cefiderocol inhibited 100% of CR-KP and CR-PA, and 98.6% of the SM isolates at the susceptibility breakpoint concentration of 4 mg/L. However, the susceptibility rate for cefiderocol against CR-AB was only 62.7%, with MIC 90 values as high as 128 mg/L. Nearly all the cefiderocol-susceptible CR-AB isolates were found to be positive for bla OXA-23 and bla TEM , whereas all the cefiderocol-resistant CR-AB isolates were found to be positive for the bla PER genes, in addition to bla OXA-23 and bla TEM .Conclusions: Cefiderocol showed potent in vitro activity against CR-KP, CR-PA, and SM isolates collected from Beijing, China. However, the resistance rate for cefiderocol against CR-AB was higher than that reported by other research centers, and the presence of bla PER might contribute to resistance in nonsusceptible CR-AB isolates.

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