International Journal of Antimicrobial Agents 40 (2012) 177–181 Contents lists available at SciVerse ScienceDirect International Journal of Antimicrobial Agents jou rn al h om epa ge: h ttp://www.elsevier.com/locate/ijantimicag Short communication Nutrient dispersion enhances conventional antibiotic activity against Pseudomonas aeruginosa biofilms Stacy Sommerfeld Ross a , Jennifer Fiegel a,b,∗ a University of Iowa, College of Pharmacy, Department of Pharmaceutical Sciences and Experimental Therapeutics, Iowa City, IA 52242, USA b University of Iowa, College of Engineering, Department of Chemical and Biochemical Engineering, Iowa City, IA 52242, USA a r t i c l e i n f o Article history: Received 17 January 2012 Accepted 20 April 2012 Keywords: Biofilm Synergy Antibiotic Dispersion Pseudomonas aeruginosa a b s t r a c t Bacterial biofilms cause significant infections in the medical field. Antibiotics commonly used to treat these infections often do not achieve complete bacterial eradication. New approaches to eliminate biofilms have focused on dispersion compounds to entice the bacteria to actively escape or disperse from the biofilm, where the bacteria may become more susceptible to antibiotics. The aim of this study was to demonstrate that combining antibiotics with nutrient dispersion compounds can synergistically decrease the viability of Pseudomonas aeruginosa biofilms. The effects of various co-treatments were stud- ied on mature biofilms through qualitative and quantitative confocal microscopy. Combined treatment of P. aeruginosa biofilms with antibiotic and dispersion compounds resulted in a significant reduction in the live bacterial population compared with the untreated control in all cases, with four combi- nations displaying synergistic action (citrate with amikacin disulphate, colistin methanesulphonate or erythromycin, and succinic acid with colistin methanesulphonate). © 2012 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved. 1. Introduction Pseudomonas aeruginosa bacteria play a significant role in tra- cheal, urinary and wound infections. Pseudomonas aeruginosa can develop into biofilms where the bacteria secrete an external matrix of polysaccharides, extracellular DNA and proteins for added pro- tection. Conversion of bacteria from a non-mucoid phenotype to a mucoid phenotype provides a defence against antibiotics and the human immune system [1]. Therefore, mucoid P. aeruginosa biofilms are not easily eradicated with antibiotics, including mul- tiple antibiotics [2]. Because of the difficulty in treating P. aeruginosa with antibiotics alone, researchers have begun investigating methods to increase the effectiveness of antibiotics with dispersion-cueing agents. Dis- persion results when bacteria actively escape the biofilm, where they become more susceptible to antibiotics [3–5]. Nutrient dis- persion compounds, chelators, salt solutions, quorum-sensing inhibitors and altered pH have been shown to induce changes to the physicochemical environment surrounding the bacteria, lead- ing to bacterial dispersal [4,6,7]. Nutrient dispersion compounds, such as the conjugate bases of simple acids, provide a nutrient-rich environment around the biofilm [7]. Moulton and Montie showed ∗ Corresponding author. Present address: The University of Iowa, 115 S. Grand Ave., S215 PHAR, Iowa City, IA 52242, USA. Tel.: +1 319 335 8830; fax: +1 319 335 9349. E-mail address: jennifer-fiegel@uiowa.edu (J. Fiegel). nutrient compounds, such as organic acids, attract motile bacte- ria [8]. Sauer et al. observed that the bacteria in biofilms treated with dispersion compounds have increased expression of a flag- ella gene and decreased expression of a pili attachment gene [7]. Since released motile bacteria can easily be 100 times more suscep- tible to antibiotics, co-treatment with dispersion compounds and antimicrobials may enhance biofilm eradication. To date, nutrient dispersion compounds have not been investi- gated for synergistic biofilm reduction with antibiotics. However, previous studies have reported improved biofilm killing when com- bining antibiotics with other compounds that cue dispersion [3,4]. In a study by Rogers et al., synergistic eradication of P. aeruginosa biofilms was found with a 2-aminoimidazole-derived dispersion compound and tobramycin [3]. In another study, Brackman et al. showed increased bacterial killing with tobramycin and quorum- sensing inhibitors [4]. The objective of this study was to identify combinations of antibiotics and nutrient dispersion compounds that synergistically decrease P. aeruginosa biofilm viability using qualitative and quantitative confocal microscopy studies. 2. Materials and methods 2.1. Materials Difco TM nutrient agar and nutrient broth were purchased from BD Diagnostic Systems (Sparks, MD); glycerol, amikacin disulphate, tobramycin sulphate, erythromycin, colistin methanesulphonate, 0924-8579/$ – see front matter © 2012 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved. http://dx.doi.org/10.1016/j.ijantimicag.2012.04.015