MECHANISMS In Vitro Synergistic Activity of Colistin and Ceftazidime or Ciprofloxacin Against Multidrug-Resistant Clinical Strains of Pseudomonas aeruginosa Beena Benita D’Souza, 1 Sunil Rao Padmaraj, 1 Panchapaddy Devasya Rekha, 2 Rouchelle Charmaine Tellis, 1 Sudharshan Prabhu, 2 and Priyanka Pothen 2 Infections caused by multidrug resistant (MDR) Pseudomonas aeruginosa are difficult to treat. Antibiotic development is dwindling in recent years. In order to develop new alternate therapies antimicrobial activity of different antibiotic combinations are being studied in vitro and in vivo. Sub-inhibitory concentrations of colistin were tested in combination with ceftazidime or ciprofloxacin by the checkerboard method against 25 MDR strains of P. aeruginosa. Synergy was observed for ceftazidime or ciprofloxacin antibiotic combinations with colistin among 73.3% of MDR3 (R AMK, GEN, TOB R CAZ R CIP ) strains and 100% of MDR4 (R AMK, GEN, TOB R CAZ R CIP R TZP ) strains. 6.6% strains of MDR3 and 14.3% strains of MDR5 (R AMK, GEN, TOB R CAZ R CIP R TZP R IPM ) phenotypes were inhibited by colistin and ceftazidime alone and 6.6% strains of MDR3 phenotypes were inhibited by colistin and ciprofloxacin alone. For the remaining strains, though synergy was not observed, significant reduction in minimum inhibitory concentration was evident. The results of this study are significant as sub-inhibitory concentrations of colistin have an advantage of reducing in vivo toxicity. These findings need further evaluation for clinical use. Introduction O ver time, the treatment of infections caused by Pseudomonas aeruginosa has become very challeng- ing to the clinicians. This is because of its extraordinary capacity in developing resistance to nearly all available antibiotics. The ability to develop resistance results from the selection of mutations in chromosomal genes and from the increasing prevalence of transferable resistant determi- nants, particularly those encoding metallo-beta-lactamases (MBLs) or extended-spectrum beta-lactamases. These re- sistant determinants are frequently transferred along with genes encoding aminoglycoside-modifying enzymes. 6 In the last years, there have been no promising new antibiotics in the drug development pipeline to treat infections caused by multiresistant P. aeruginosa. Clinicians and microbiologists have been therefore forced to reappraise the clinical value of colistin. Colistin, a relatively old polymyxin antibiotic was in clinical use in the 1950s and was withdrawn from treatment because of its toxicity. Recently, on-therapy resistance with colistin monotherapy has been reported. 13 Colistin sulfate (polymyxin E) is one of the cyclic polypeptide antibiotics known to interact primarily with LPS and phospholipids present at the outer membrane of Gram-negative bacteria with also the ability to disturb permeability of the cytoplas- mic membrane. This may allow the entry of hydrophobic and/ or large molecules, which ultimately leads to leakage of cellular contents. 15 Therefore, colistin can be used in com- bination with other antibiotics to obtain a synergistic effect. Synergy is defined as significantly greater activity provided by two agents combined, compared with that provided by the sum of each agent alone. Combined antimicrobial therapies can be used in clinical infections caused by bacterial strains that are susceptible to one or more antibiotics, or are resistant to all available antimicrobials. 11 Often, researchers may encounter high minimum inhibi- tory concentration (MIC) for antibiotics that are uninter- pretable. The ability of the bacteria to survive at a high concentration of antibiotics clearly indicates that these strains have very strong mechanisms for overcoming the antibiotic effect and hence increasing the dose over time certainly will not be beneficial in treatment. Alternate treatment options with antibiotic combinations may be used for effective man- agement of such infections. Hence, experiments were carried out to explore the potential for synergistic activity of colistin with ceftazidime or ciprofloxacin against multidrug-resistant (MDR) clinical strains of P. aeruginosa. 1 Department of Microbiology, Yenepoya Medical College, Yenepoya University, Mangalore, India. 2 Yenepoya Research Centre, Yenepoya University, Mangalore, India. MICROBIAL DRUG RESISTANCE Volume 20, Number 6, 2014 ª Mary Ann Liebert, Inc. DOI: 10.1089/mdr.2014.0006 1