https://orcid.org/0000-0003-0949-592X
Correction
14 May 2021: The PLOS ONE Staff (2021) Correction: Synergistic antibacterial effects of colistin in combination with aminoglycoside, carbapenems, cephalosporins, fluoroquinolones, tetracyclines, fosfomycin, and piperacillin on multidrug resistant Klebsiella pneumoniae isolates. PLOS ONE 16(5): e0251994. https://doi.org/10.1371/journal.pone.0251994 View correction
Figures
Abstract
Multidrug resistant Enterobacterales have become a serious global health problem, with extended hospital stay and increased mortality. Antibiotic monotherapy has been reported ineffective against most drug resistant bacteria including Klebsiella pneumoniae, thus encouraging the use of multidrug therapies as an alternative antibacterial strategy. The present works assessed the antibacterial activity of colistin against K. pneumoniae isolates. Resistant isolates were tested against 16 conventional antibiotics alone and in combination with colistin. The results revealed that all colistin resistant isolates demonstrated multidrug resistance against the tested antibiotics except amikacin. At sub-inhibitory concentrations, combinations of colistin with amikacin, or fosfomycin showed synergism against 72.72% (8 of 11 isolates). Colistin with either of gentamicin, meropenem, cefoperazone, cefotaxime, ceftazidime, moxifloxacin, minocycline, or piperacillin exhibited synergism against 81.82% (9 of 11 isolates). Combinations of colistin with either of tobramycin or ciprofloxacin showed synergism against 45.45% (5 in 11 isolates), while combinations of colistin with imipenem or ceftolozane and tazobactam displayed 36.36% (4 of 11 isolates) and 63.64% (7 of 11 isolates) synergism. In addition, combinations of colistin with levofloxacin was synergistic against 90.91% (10 of 11 isolates). The results revealed that combinations of colistin with other antibiotics could effectively inhibit colistin resistant isolates of K. pneumoniae, and thus could be further explore for the treatment of multidrug resistant pathogens.
Citation: Ontong JC, Ozioma NF, Voravuthikunchai SP, Chusri S (2021) Synergistic antibacterial effects of colistin in combination with aminoglycoside, carbapenems, cephalosporins, fluoroquinolones, tetracyclines, fosfomycin, and piperacillin on multidrug resistant Klebsiella pneumoniae isolates. PLoS ONE 16(1): e0244673. https://doi.org/10.1371/journal.pone.0244673
Editor: Iddya Karunasagar, Nitte University, INDIA
Received: August 29, 2020; Accepted: December 14, 2020; Published: January 6, 2021
Copyright: © 2021 Ontong et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Data Availability: All relevant data are within the paper and its Supporting Information files.
Funding: This work was supported by Senior Research Scholar, the Thailand Research Fund, in the form of a grant awarded to SC (RTA6180006).
Competing interests: The authors declare that no competing interests exist.
Introduction
The spread of multidrug resistant bacteria has become a public health emergency that threatens the continued usage of antibiotics chemotherapy. Infections due to carbapenemase-producing Enterobacterales are fast spreading across the globe, rendering the healthcare systems ineffective. It is estimated that infections associated with drug-resistant pathogens currently cause about 700,000 deaths annual, and if the status quo prevails could increase to 10 million annual deaths by 2050 [1]. Klebsiella pneumoniae is a Gram-negative Enterobacterales member commonly associated with hospital acquired infections. In the past, carbapenems were used as choice drugs for the treatment of K. pneumoniae infection. However, with the emergence and spread of carbapenemase-producing Enterobacterales, colistin is considered a last-resort treatment option for infections caused by carbapenem resistant bacterial isolates.
Colistin also known as polymyxin E is a cationic peptide that targets the negatively charged lipopolysaccharide (LPS) of Gram-negative bacteria. Colistin competitively displaces divalent cations (Ca2+ and Mg2+) from the phosphate groups of membrane lipids, resulting in the rupture of cell membrane and leakage of intracellular components [2]. Emergence of colistin resistance in Enterobacterales including K. pneumoniae has been reported in many parts of the globe [3, 4] and is fast spreading [5]. Colistin resistance was generally thought to be mediated by alterations and modification of the LPS target through addition of positively charged 4-amino-4-deoxy-L-arabinose (L-Ara4N) and phosphoethanolamine (pEtN) cationic molecules, responsible for decrease in bacterial outer membrane negative charge and reduced interaction with colistin [6]. However, recent scientific enquiries have reported the role of efflux pumps [7–10] and plasmid encoded mcr 1–9 genes [4, 11, 12] in colistin resistance.
As the fight against multidrug resistance continues, concerted efforts by agencies, health care systems and biomedical scientist are restlessly exploring possible alternatives that might suffice pending the discovery and development of novel antibacterial agents that could effectively curb the spread of antibiotics resistance. The use of antibiotics combinations [13–15], efflux pump inhibitors [16–18], and resistance modifying agents [19, 20] are suggested as temporary control measures to reverse microbial resistance or enhance the inactivation of resistant bacterial isolates. Antibiotics combination therapy is proposed as a reliable option with demonstrated results against multidrug resistant bacterial isolates.
The present study tested the antibiotic susceptibility of 85 K. pneumoniae isolates obtained from hospitals in the southern region of Thailand against colistin. The resistant isolates were further tested against several groups of antibiotics, and the synergistic antibacterial effects of combinations of colistin with other antibiotics were evaluated. The study presents In vitro experimental antibacterial data from broth micro-dilution technique and the checkerboard assay but did not monitor the time-kill kinetics.
Materials and methods
Chemicals and media
All culture media were purchased from Becton Dickinson & Co. DifcoTM (Franklin Lakes, NJ, USA). Colistin sulfate, piperacillin, minocycline hydrochloride, tobramycin, and moxifloxacin hydrochloride were obtained from Sigma-Aldrich, (Saint Louis, MO, USA). Ciprofloxacin, cefotaxime, and levofloxacin were purchased from Siam Bheasach Co., Ltd. (Bangkok, Thailand). Tigecycline and ceftaroline fosamil were Pfizer Inc. (Philadelphia, PA, USA). Ceftazidime was obtained from Reyoung Pharmaceutical Co., Ltd. (Shandong, China). Imipenem was obtained from Merck Sharp & Dohme Corp. (Elkton, VA, USA). Meropenem was obtained from M&H manufacturing Co., Ltd. (Samutprakarn, Thailand). Cefoperazone and sulbactam was obtained from L.B.S. Laboratory Ltd. (Bangkok, Thailand). Ceftolozane and tazobactam was obtained from Steri-Pharma, LLC (Syracuse, NY, USA). Fosfomycin was obtained from Meiji seikakaisna, Ltd. (Tokyo, Japan).
Bacterial strains
Isolates collected from hospitalized patients unresponsive to antibiotics chemotherapy, with prolonged hospital stay in 8 hospitals located in Southern Thailand were used in this study. All isolates were identified to species level using standard biochemical tests and MALDI-TOF-MS. A total of 85 K. pneumoniae isolates that exhibited colony on MacConkey agar supplemented with 6 μg/mL imipenem were taken as resistant to carbapenems. The surveillance study was conducted post antibiotics treatments. Escherichia coli ATCC 25922 was used as a quality control. All the bacterial cultures were stored in tryptic soy broth (TSB supplemented with 40% glycerol and kept at -80°C.
Antimicrobial susceptibility testing
Minimum inhibitory concentrations (MICs) were determined by the broth microdilution method in according to Clinical and Laboratory Standards Institute (CLSI) guidelines [21]. Briefly, serial two-fold dilutions of antibiotics were prepared in cation-adjusted Mueller-Hinton II broth (CA-MHBII). To investigate the effect of each antibiotic, an aliquot of 100 μL of diluted bacterial suspension (1x106 CFU/ml) was mixed with 100 μL antibiotic into each well and incubated at 37°C for 18 h. MIC was expressed as the lowest concentration of the antibiotic that inhibits visible growth after incubation. Following European Committee on Antimicrobial Susceptibility Testing (EUCAST) breakpoints, isolates with a colistin MIC ≤ 2 μg/mL were categorized as susceptible and those with a colistin MIC ≥4 μg/mL were categorized as resistant [22]. To determine the MBC, MIC and supra-MIC dilutions were spotted on an agar plate and incubated overnight at 37°C. Bacterial growth was observed, and MBC was defined as the lowest concentration that showed no visible bacterial regrowth.
Checkerboard technique
The synergistic effects of colistin and any of sixteen other antibiotics (amikacin, gentamicin, tobramycin, imipenem, meropenem, cefoperazone, cefotaxime, ceftazidime, ceftolozane and tazobactam, ciprofloxacin, levofloxacin, moxifloxacin, minocycline, tigecycline, fosfomycin, and piperacillin) on K. pneumoniae isolates were tested using the checkerboard technique. Each antibiotic was diluted to concentrations ranging from 1/64 MIC to 8 MIC of the previously determined MIC. Briefly, 50 μL of colistin and each antibiotic were assessed by adding 100 μL diluted bacterial suspension (1x106 CFU/mL) into a well containing 50 μL colistin and 50 μL one of sixteen other antibiotics. The test was then read after 18 h of incubation at 37 C. Each value was mean of triplicates from three independent experiments. The effects of the antimicrobial combinations were defined according to the fractional inhibitory concentration index (FICI) as following equation:
The FICI results for each combination were interpreted as follows: FICI ≤ 0.5, synergism; 0.5 < FICI < 1, additive; 1 ≤ FICI < 2, indifference; and FICI ≥ 2, antagonism. Escherichia coli ATCC 25922 was used as standard control strains for the assays [23].
Ethical statement
This retrospective study was approved by the Institutional Review Board (IRB) of the Faculty of Medicine, Prince of Songkla University, Thailand (EC: 54-080-14-1-2.). The researchers were granted permission to extract the data from the database with waiver of consent because of the observational nature of the study. All data were fully anonymized before the researcher accessed and analyzed.
Results and discussion
Distribution of K. pneumoniae isolates
In this study K. pneumoniae isolates obtained from patients receiving treatment in tertiary hospitals were tested against colistin. The isolates were collected and tested for antibiotics susceptibility due to patient’s unresponsiveness to antibiotics treatments and suspicion of opportunistic role of drug-resistant bacterial colonizers in aggravating health conditions of immunocompromised patients. Table 1. shows the distribution of the isolates, collected from eight public hospitals located in the southern region of Thailand based on sample type and location of the hospital. The results revealed that all nine isolates from Narathiwat hospital were resistant to colistin, while one out of the three isolates from Songkhla and the only isolate from Trang were also resistant to colistin. Demographic data, clinical characteristics, and outcomes of the patients with colonization due to colistin-resistant and carbapenem-resistant K. pneumoniae are presented in S1 Table. The samples were collected from adult patients between the ages of 25–94 years, who were admitted in the ICU units of the hospitals. Most of the patients had underlining health conditions including diabetes mellitus, hypertension, dyslipidemia, coronary artery disease, cerebrovascular disease, chronic kidney disease, and chronic obstructive pulmonary disease.
Antibacterial activities of colistin against K. pneumoniae isolates
A total of 85 isolates were tested for susceptibility to colistin using broth microdilution assay (Table 2 and S2 Table). The results showed that 74 isolates (87.05%) were susceptible to colistin with MIC values ranging from 0.5 to 2.0 μg/mL and MBC values ranging from 0.5 to 4.0 μg/mL. However, 11 isolates (12.94%) demonstrated resistance to colistin with MIC and MBC values ranging from 256 to >1024 μg/mL and 512 to >1024 μg/mL, respectively. Colistin resistant K. pneumoniae has been reported in various countries and regions including Netherlands, America, Nigeria as well as Thailand [24–26]. Recently 213 of 280 (79.1%) K. pneumoniae isolates obtained from humans in Thailand showed colistin resistance [27]. The emergence of colistin resistance K. pneumoniae presents a major threat to public health, since colistin represents the last line drug of choice against carbapenem resistant K. pneumoniae.
Antibiogram of colistin resistant K. pneumoniae isolates
The eleven resistant isolates were evaluated for antibiogram against sixteen conventional antibiotics and colistin (Table 3). Although the agar dilution method is recommended as reference method for the determination of Fosfomycin MICs, using the reference agar dilution method in a checkerboard analysis is practically difficult. Thus, the broth microdilution with glucose-6-phosphate (G-6-P) was used [28]. Results of the assay were interpreted based on CLSI and EUCAST breakpoint standards [22, 29]. All colistin resistant isolates exhibited multi-drug resistance to aminoglycosides, carbapenems, cephalosporins, fluoroquinolones, fosfomycin, tetracyclines, and piperacillin, but were susceptible to amikacin with MIC ranging from 4 to 16 μg/mL. Previous researchers have reported 69.57 and 64.1% susceptibility of K. pneumoniae to amikacin [30, 31]. Resistance to carbapenems (imipenem and meropenem) was observed for all colistin resistant isolates. Two isolates (1SK5R and 1TR5R) were susceptible to cefoperazone, ceftolozane and tazobactam, levofloxacin, fosfomycin, and piperacillin. In addition, isolate 1TR5R was susceptible to tobramycin, ceftazidime, and minocycline. Multidrug resistant K. pneumoniae has been reported by previous researchers [32, 33], and is globally spreading unabated. This might lead to increased hospital stay and K. pneumoniae associated mortality.
Synergistic effects of colistin-antibiotics combinations
While the search for alternative antimicrobial agents that can effectively control the spread of multidrug resistance continues, various temporary measures are been employed for the treatment of infections caused by drug resistant bacterial isolates. Antibiotic combination therapy is a possible effective option which currently is attracting numerous research attention [34–36]. Combinations of antibiotics effectively inhibit microbial proliferation through a multi-target approach resulting in the microbial death. Hence, the synergistic effects of colistin with conventional antibiotics (amikacin, gentamicin, tobramycin, imipenem, meropenem, cefoperazone, cefotaxime, ceftazidime, ceftolozane and tazobactam, ciprofloxacin, levofloxacin, moxifloxacin, minocycline, tigecycline, fosfomycin, and piperacillin) against K. pneumoniae isolates were evaluated and classified based on FICI parameter (S3 Table).
Combination of colistin with aminoglycosides
Table 4 presents the chequerboard results of colistin in combination with aminoglycoside (amikacin, gentamicin, and tobramycin) against the eleven colistin resistant K. pneumoniae isolates. The FICI ranged from 0.125 to 0.500 for K. pneumoniae isolates, except for isolates 1NT6R and 1NT6Th (CCU)/1 with amikacin FICI values of 0.078 and 0.062, respectively. However, combination of colistin with amikacin or tobramycin showed no synergy against isolate 1NT7R. Antibacterial synergistic effects have been demonstrated for gentamicin and amikacin combinations with polymyxin B and ceftazidime-avibactam [37, 38].
Combination of colistin with carbapenems
The antibacterial activities of colistin in combination with carbapenems (imipenem and meropenem) is presented in Table 5. The results demonstrated synergism with FICI values ranging from 0.250 to 0.500 for most isolates, except isolates 1SK5R and 1TR5R. Combinations of colistin and imipenem however showed no effects on isolates 1NT6Tu/1, 1NT8Th, 1NT6Ng (CCU/1), 1NT6Th (CCU)/1, and 1NT6R (CCU). The antibacterial activities of fosfomycin and meropenem combination and colistin with meropenem combinations effective inhibited NDM and carbapenemase producing K. pneumoniae [34, 39]. In addition, relebactam-imipenem combinations showed enhanced antibacterial activity against colistin resistant K. pneumoniae with potentials of restoring bacteria susceptibility to imipenem [40, 41].
Combination of colistin with cephalosporins
Colistin–cephalosporins (Cefoperazone, Cefotaxime, Ceftazidime and Ceftolozane and Tazobactam) combinations were also assessed for synergism against colistin resistant K. pneumoniae isolates Table 6. Combinatory effect of colistin with cephalosporins revealed FICIs ranging from 0.187 to 0.375 for all isolates, except 1SK5R and 1TR5R, while combination with ceftolozane and tazobactam were ineffective against 1NT8Th and 1NT6R (CCU). The results suggested collaborative disruption of the cell wall since both colistin and cephalosporins targets strategic components of the cell wall. Ceftazidime-avibactam in combination with colistin was previous reported to be effective against colistin non-susceptible strains of multidrug resistant (MDR) Pseudomonas aeruginosa [42, 43]. In addition, combinations of ceftazidime/avibactam and colistin, tobramycin, or tigecycline were effective against OXA-48-producing Enterobacterales strains [44].
Combination of colistin with fluoroquinolones
The effects of colistin combination with fluoroquinolones (ciprofloxacin, levofloxacin, moxifloxacin) are presented in Table 7. The FICIs (0.093 to 0.500) indicated synergistic effects against most isolates. However, combinations of colistin with all three antibiotics were ineffective on isolate 1TR5R, while ciprofloxacin or moxifloxacin were also not effective on isolate 1SK5R.
Combination of colistin with fosfomycin, tetracyclines (minocycline, and tigecycline) and penicillin (piperacillin)
In addition, effects of colistin in combination with fosfomycin, tetracyclines (minocycline, and tigecycline) and penicillin (piperacillin) resistant K. pneumoniae isolates is presented in Table 8. Combination of colistin with fosfomycin showed FICI ranging from 0.062 to 0.500 against most isolates except isolates 1NT6Tu/1, 1NT6R, and 1SK5R while combination of colistin with piperacillin range of 0.046 to 0.375 for most isolates except 1NT7R and 1SK5R. Against all isolates, combination of colistin with tigecycline was not synergistic, whereas combinations with minocycline demonstrated synergistic activity against most isolates with FICI value of 0.093, except for isolate 1NT6R (CCU) with FICI value of 0.062, and were not effective against 1SK5R and 1TR5R.
Discussion
The rapid spread of multidrug resistant pathogenic bacteria places an enormous responsibility on global health care systems. Moreover, the search and development of new antibiotics is far too slow with no novel discovery over the last 30 years. This shortage of effective therapeutic agents has encouraged trial of combinations of existing agents for synergistic activities against drug resistant isolates. The present study combined colistin a last line drug for multidrug resistant Enterobacterales with 16 other antibiotics that are not effective against K. pneumoniae. A total of 11 isolates (12.94%) presented resistance to colistin. Nine out of the 11 resistant isolates were obtained from Narathiwat hospital, with a 100% resistance prevalence. The results suggested high prevalence of resistance amongs K. pneumoniae in Narathiwat hospital which might be due to local tranmission within the hospital. Furthermore, 1 among 3 isolates obtained from Songkhla hospital was resistant, whereas the single isolate obtained from Trang hospital was also resistant. However, due to the few number of isolates used in the study, epidemiological statements on the prevalence of colistin resistant isolates in this regions might be baised. Colistin resistant K. pneumoniae has previously been reported in various countries and regions including Netherlands, America, Nigeria as well as Thailand [24–26]. Recently 213 of 280 (79.1%) K. pneumoniae isolates obtained from humans in Thailand showed colistin resistance [27].
All colistin–resistant isolates were also resistant to other antibiotics including aminoglycosides, carbapenems, cephalosporins, fluoroquinolones, fosfomycin, tetracyclines, and piperacillin, but were susceptible to amikacin with MIC ranging from 4 to 16 μg/mL. Previous researchers have reported 69.57 and 64.1% susceptibility of K. pneumoniae to amikacin [30, 31]. The results suggested that amikacin might be a drug option in the management of drug resistant Enterobacterales, and should be further explored. Resistance to carbapenems (imipenem and meropenem) was observed for all colistin resistant isolates. Two isolates (1SK5R and 1TR5R) were susceptible to cefoperazone, ceftolozane and tazobactam, levofloxacin, fosfomycin, and piperacillin. In addition, isolate 1TR5R was susceptible to tobramycin, ceftazidime, and minocycline. Multidrug resistance in K. pneumoniae has been reported by previous researchers [32, 33], resulting in increased hospital stay and K. pneumoniae associated mortality.
While the search for alternative and effective antimicrobial agents continues, various temporary measures are been employed for the treatment of infections caused by drug resistant bacterial isolates. Antibiotic combination therapy is a possible effective option which currently is attracting numerous research attention [34–36]. Combinations of antibiotics effectively inhibit microbial proliferation through a multi-target approach resulting in effective inactivation of cells and microbial death. Hence, the synergistic effects of colistin with conventional antibiotics (amikacin, gentamicin, tobramycin, imipenem, meropenem, cefoperazone, cefotaxime, ceftazidime, ceftolozane and tazobactam, ciprofloxacin, levofloxacin, moxifloxacin, minocycline, tigecycline, fosfomycin, and piperacillin) against multi-drug resistant K. pneumoniae isolates were evaluated and classified based on FICI parameter (S3 Table). Antibiotic combination demonstrated 5- to 64-fold reduction in MIC of colistin and 4-to 512-fold reduction in MIC of tested antibiotics. A summary of the results indicates that colistin with amikacin, or fosfomycin combinations were synergistic against 72.72% (8 of 11 isolates). Colistin with gentamicin, or meropenem, or cefoperazone, or cefotaxime, or ceftazidime, or moxifloxacin, or minocycline, or piperacillin exhibited synergism against 81.82% (9 of 11 isolates). Combinations of colistin with either of tobramycin or ciprofloxacin showed 45.45% (5 in 11 isolates), while combinations of colistin with imipenem or ceftolozane and tazobactam displayed 36.36% (4 of 11 isolates) and 63.64% (7 of 11 isolates) synergism. In addition, combinations of colistin with levofloxacin was synergistic against 90.91% (10 of 11 isolates), while colistin and tigecycline combination were overall not synergistic. The results revealed that colistin in combination with fifteen other antibiotics could effectively inhibit colistin resistant isolates of K. pneumoniae. The results suggested that combination therapies could be further explore for the treatment of multidrug resistant pathogens.
Conclusions
The present study combined colistin a last line drug for multidrug resistant Enterobacterales with 16 other antibiotics that are not effective against K. pneumoniae. The results revealed that colistin in combination with fifteen other antibiotics effectively inhibit colistin resistant isolates of K. pneumoniae. The results suggested that combination therapies could be further explore for the treatment of multidrug resistant pathogens.
Supporting information
S1 Table. Demographic data, clinical characteristics, and outcomes of the patients with colonization due to colistin-resistant and carbapenem-resistant K. Pneumoniae.
https://doi.org/10.1371/journal.pone.0244673.s001
(DOCX)
S2 Table. Minimum Inhibitory Concentration (MIC) and Minimum Bactericidal Concentration (MBC) of colistin against Klebsiella pneumoniae clinical isolates.
https://doi.org/10.1371/journal.pone.0244673.s002
(DOCX)
S3 Table. Synergistic effects of antibiotics and colistin combination against Klebsiella pneumoniae isolates from hospitalized patients.
https://doi.org/10.1371/journal.pone.0244673.s003
(DOCX)
References
- 1.
WHO. No Time to Wait: Securing the future from drug-resistant infections. World Health Organization: Geneva, Switzerland. 2019.
- 2. Biswas S, Brunel J-M, Dubus J-C, Reynaud-Gaubert M, Rolain J-M. Colistin: an update on the antibiotic of the 21st century. Expert Review of Anti-Infective Therapy. 2012;10(8):917–34. pmid:23030331
- 3. Roy S, Das P, Das S, Roy S, Pal S, Joy VM, et al. Detection of the emergence of mcr-1–mediated colistin-resistant Escherichia coli and Klebsiella pneumoniae through a hospital-based surveillance in an oncology center in eastern India. Infection Control & Hospital Epidemiology. 2020;41(3):378–80.
- 4. Gelbicova T, Kolackova I, Krutova M, Karpiskova R. The emergence of mcr-1-mediated colistin-resistant Escherichia coli and Klebsiella pneumoniae in domestic and imported turkey meat in the Czech Republic 2017–2018. Folia Microbiologica. 2020;65(1):211–6. pmid:31001764
- 5. Lalaoui R, Bakour S, Livnat K, Assous MV, Diene SM, Rolain J-M. Spread of Carbapenem and Colistin-resistant Klebsiella pneumoniae ST512 clinical isolates in Israel: a cause for vigilance. Microbial Drug Resistance. 2019;25(1):63–71. pmid:30129868
- 6. Trent MS. Biosynthesis, transport, and modification of lipid A. Biochemistry and Cell Biology. 2004;82(1):71–86. pmid:15052329
- 7. Sun S, Gao H, Liu Y, Jin L, Wang R, Wang X, et al. Co-existence of a novel plasmid-mediated efflux pump with colistin resistance gene mcr in one plasmid confers transferable multidrug resistance in Klebsiella pneumoniae. Emerging Microbes & Infections. 2020;(just-accepted):1–41. pmid:32401163
- 8. Srinivasan VB, Rajamohan G. KpnEF, a new member of the Klebsiella pneumoniae cell envelope stress response regulon, is an SMR-type efflux pump involved in broad-spectrum antimicrobial resistance. Antimicrobial Agents and Chemotherapy. 2013;57(9):4449–62. pmid:23836167
- 9. Singh SK, Mishra M, Sahoo M, Patole S, Mohapatra H. Efflux mediated colistin resistance in diverse clones of Klebsiella pneumoniae from aquatic environment. Microbial Pathogenesis. 2017;102:109–12. pmid:27914962
- 10. Naha S, Sands K, Mukherjee S, Roy C, Rameez MJ, Saha B, et al. KPC-2-producing Klebsiella pneumoniae ST147 in a neonatal unit: Clonal isolates with differences in colistin susceptibility attributed to AcrAB-TolC pump. International Journal of Antimicrobial Agents. 2020;55(3):105903. pmid:31954832
- 11. Salloum T, Panossian B, Bitar I, Hrabak J, Araj GF, Tokajian S. First report of plasmid-mediated colistin resistance mcr-8.1 gene from a clinical Klebsiella pneumoniae isolate from Lebanon. Antimicrobial Resistance & Infection Control. 2020;9(1):1–7. pmid:32586402
- 12. Nabti LZ, Sahli F, Ngaiganam EP, Radji N, Mezaghcha W, Lupande-Mwenebitu D, et al. Development of real-time PCR assay allowed describing the first clinical Klebsiella pneumoniae isolate harboring plasmid-mediated colistin resistance mcr-8 gene in Algeria. Journal of Global Antimicrobial Resistance. 2020;20:266–71. pmid:31476479
- 13. Wistrand-Yuen P, Olsson A, Skarp K-P, Friberg LE, Nielsen EI, Lagerbäck P, et al. Evaluation of polymyxin B in combination with 13 other antibiotics against carbapenemase-producing Klebsiella pneumoniae in time-lapse microscopy and time-kill experiments. Clinical Microbiology and Infection. 2020. pmid:32224200
- 14. Bakthavatchalam YD, Shankar A, Muthuirulandi Sethuvel DP, Asokan K, Kanthan K, Veeraraghavan B. Synergistic activity of fosfomycin–meropenem and fosfomycin–colistin against carbapenem resistant Klebsiella pneumoniae: an in vitro evidence. Future Science OA. 2020;6(4):FSO461. pmid:32257374
- 15. Aye SM, Galani I, Yu H, Wang J, Chen K, Wickremasinghe H, et al. Polymyxin triple combinations against polymyxin-resistant, multidrug-resistant KPC-producing Klebsiella pneumoniae. Antimicrobial Agents and Chemotherapy. 2020. pmid:32393492
- 16. Sundaramoorthy NS, Mohan HM, Subramaniam S, Raman T, Ganesan SS, Sivasubamanian A, et al. Ursolic acid inhibits colistin efflux and curtails colistin resistant Enterobacteriaceae. AMB Express. 2019;9(1):27. pmid:30778773
- 17. Ni W, Li Y, Guan J, Zhao J, Cui J, Wang R, et al. Effects of efflux pump inhibitors on colistin resistance in multidrug-resistant Gram-negative bacteria. Antimicrobial Agents and Chemotherapy. 2016;60(5):3215–8. pmid:26953203
- 18. Baron SA, Rolain J-M. Efflux pump inhibitor CCCP to rescue colistin susceptibility in mcr-1 plasmid-mediated colistin-resistant strains and Gram-negative bacteria. Journal of Antimicrobial Chemotherapy. 2018;73(7):1862–71. pmid:29718423
- 19. Witherell KS, Price J, Bandaranayake AD, Olson J, Call DR. Circumventing colistin resistance by combining colistin and antimicrobial peptides to kill colistin-resistant and multidrug-resistant Gram-negative bacteria. Journal of Global Antimicrobial Resistance. 2020. pmid:32512236
- 20. Hashemi MM, Rovig J, Weber S, Hilton B, Forouzan MM, Savage PB. Susceptibility of colistin-resistant, Gram-negative bacteria to antimicrobial peptides and ceragenins. Antimicrobial Agents and Chemotherapy. 2017;61(8). pmid:28584137
- 21.
CLSI. Performance standards for antibiotics susceptibility testing. M100S. CLSI Wayne, PA; 2016.
- 22.
EUCAST. "The European Committee on Antimicrobial Susceptibility Testing. Breakpoint tables for interpretation of MICs and zone diameters. European Society of Clinical Microbiology and Infectious Diseases. Version 10.0; 2020.
- 23. Kheshti R, Pourabbas B, Mosayebi M, Vazin A. In vitro activity of colistin in combination with various antimicrobials against Acinetobacter baumannii species, a report from South Iran. Infection and Drug Resistance. 2019;12:129. pmid:30643441
- 24. Weterings V, Zhou K, Rossen J, van Stenis D, Thewessen E, Kluytmans J, et al. An outbreak of colistin-resistant Klebsiella pneumoniae carbapenemase-producing Klebsiella pneumoniae in the Netherlands (July to December 2013), with inter-institutional spread. European Journal of Clinical Microbiology & Infectious Diseases. 2015;34(8):1647–55.
- 25. Olaitan AO, Diene SM, Kempf M, Berrazeg M, Bakour S, Gupta SK, et al. Worldwide emergence of colistin resistance in Klebsiella pneumoniae from healthy humans and patients in Lao PDR, Thailand, Israel, Nigeria and France owing to inactivation of the PhoP/PhoQ regulator mgrB: an epidemiological and molecular study. International Journal of Antimicrobial Agents. 2014;44(6):500–7. pmid:25264127
- 26. Marchaim D, Chopra T, Pogue JM, Perez F, Hujer AM, Rudin S, et al. Outbreak of colistin-resistant, carbapenem-resistant Klebsiella pneumoniae in metropolitan Detroit, Michigan. Antimicrobial Agents and Chemotherapy. 2011;55(2):593–9. pmid:21115786
- 27. Eiamphungporn W, Yainoy S, Jumderm C, Tan-arsuwongkul R, Tiengrim S, Thamlikitkul V. Prevalence of the colistin resistance gene mcr-1 in colistin-resistant Escherichia coli and Klebsiella pneumoniae isolated from humans in Thailand. Journal of Global Antimicrobial Resistance. 2018;15:32–5. pmid:29935331
- 28. Flamm RK, Rhomberg PR, Lindley JM, Sweeney K, Ellis-Grosse E, Shortridge D. Evaluation of the bactericidal activity of fosfomycin in combination with selected antimicrobial comparison agents tested against gram-negative bacterial strains by using time-kill curves. Antimicrobial Agents and Chemotherapy. 2019;63(5). pmid:30858207
- 29.
CLSI. Performance standards for antimicrobial susceptibility testing. Clinical and Laboratory Standards Institute Wayne, PA; 2017.
- 30. Lin L, Xiao X, Wang X, Xia M, Liu S. In Vitro Antimicrobial Susceptibility Differences Between Carbapenem-Resistant KPC-2-Producing and NDM-1-Producing Klebsiella pneumoniae in a Teaching Hospital in Northeast China. Microbial Drug Resistance. 2020;26(2):94–9. pmid:31433255
- 31. Jacobs MR, Good CE, Hujer AM, Abdelhamed AM, Rhoads DD, Hujer KM, et al. ARGONAUT II study of the in vitro activity of plazomicin against carbapenemase-producing Klebsiella pneumoniae. Antimicrobial Agents and Chemotherapy. 2020;64(5). pmid:32152078
- 32. Xu H, Huo C, Sun Y, Zhou Y, Xiong Y, Zhao Z, et al. Emergence and molecular characterization of multidrug-resistant Klebsiella pneumoniae isolates harboring blaCTX-M-15 extended-spectrum β-lactamases causing ventilator-associated pneumonia in China. Infection and Drug Resistance. 2019;12:33. pmid:30588046
- 33. Palmeiro JK, De Souza RF, Schörner MA, Araujo HP, Grazziotin AL, Vidal NM, et al. Molecular epidemiology of multidrug-resistant Klebsiella pneumoniae isolates in a Brazilian tertiary hospital. Frontiers in Microbiology. 2019;10:1669. pmid:31396186
- 34. Yu L, Zhang J, Fu Y, Zhao Y, Wang Y, Zhao J, et al. Synergetic effects of combined treatment of colistin with meropenem or amikacin on carbapenem-resistant Klebsiella pneumoniae in vitro. Frontiers in Cellular and Infection Microbiology. 2019;9:422. pmid:31921701
- 35. Kilic U, Koroglu M, Olmez M, Altindis M. Investigation of the In Vitro Effectiveness of Aztreonam/Avibactam, Colistin/Apramycin, and Meropenem/Apramycin Combinations Against Carbapenemase-Producing, Extensively Drug-Resistant Klebsiella pneumoniae Strains. Microbial Drug Resistance. 2020.
- 36. Erdem F, Abulaila A, Aktas Z, Oncul O. In vitro evaluation of double carbapenem and colistin combinations against OXA-48, NDM carbapenemase-producing colistin-resistant Klebsiella pneumoniae strains. Antimicrobial Resistance & Infection Control. 2020;9(1):1–9. pmid:32430058
- 37. Mikhail S, Singh NB, Kebriaei R, Rice SA, Stamper KC, Castanheira M, et al. Evaluation of the synergy of ceftazidime-avibactam in combination with meropenem, amikacin, aztreonam, colistin, or fosfomycin against well-characterized multidrug-resistant Klebsiella pneumoniae and Pseudomonas aeruginosa. Antimicrobial Agents and Chemotherapy. 2019;63(8):e00779–19. pmid:31182535
- 38. Firmo EF, Júnior JBO, Scavuzzi AML, Alves LC, Brayner FA, Veras DL, et al. In vitro activity of polymyxin B in combination with meropenem, amikacin and gentamicin against Klebsiella pneumoniae clinical isolates coharboring AMEs, blaNDM-1 and blaKPC-2. Journal of Global Antimicrobial Resistance. 2020.
- 39. Erturk Sengel B, Altinkanat Gelmez G, Soyletir G, Korten V. In vitro synergistic activity of fosfomycin in combination with meropenem, amikacin and colistin against OXA-48 and/or NDM-producing Klebsiella pneumoniae. Journal of Chemotherapy. 2020:1–7. pmid:32228228
- 40. Galani I, Souli M, Nafplioti K, Adamou P, Karaiskos I, Giamarellou H, et al. In vitro activity of imipenem-relebactam against non-MBL carbapenemase-producing Klebsiella pneumoniae isolated in Greek hospitals in 2015–2016. European Journal of Clinical Microbiology & Infectious Diseases. 2019;38(6):1143–50.
- 41. Carpenter J, Neidig N, Campbell A, Thornsberry T, Truex T, Fortney T, et al. Activity of imipenem/relebactam against carbapenemase-producing Enterobacteriaceae with high colistin resistance. Journal of Antimicrobial Chemotherapy. 2019;74(11):3260–3. pmid:31430370
- 42. Montero M, Ochoa SD, López-Causapé C, VanScoy B, Luque S, Sorlí L, et al. Efficacy of Ceftolozane-Tazobactam in Combination with Colistin against Extensively Drug-Resistant Pseudomonas aeruginosa, Including High-Risk Clones, in an In Vitro Pharmacodynamic Model. Antimicrobial Agents and Chemotherapy. 2020;64(4).
- 43. Mataraci Kara E, Yilmaz M, İstanbullu Tosun A, Özbek Çelik B. Synergistic activities of ceftazidime-avibactam in combination with different antibiotics against colistin-nonsusceptible clinical strains of Pseudomonas aeruginosa. Infectious Diseases. 2020:1–9. pmid:32427010
- 44. Mataraci Kara E, Yilmaz M, Istanbullu Tosun A, Özbek Çelik B. Evaluation of the synergy of ceftazidime/avibactam in combination with colistin, doripenem, levofloxacin, tigecycline, and tobramycin against OXA-48 producing Enterobacterales. Journal of Chemotherapy. 2020:1–8.