JAC Advance Access originally published online on March 10, 2006
Journal of Antimicrobial Chemotherapy 2006 57(5):1010-1012; doi:10.1093/jac/dkl072
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Correspondence |
Survival and resistance to imipenem of Pseudomonas aeruginosa on latex gloves
1 Department of Microbiology, School of Medicine, University of Seville, 41080 Seville, Spain; 2 University Hospital Virgen Macarena, Seville, Spain
* Corresponding author. Tel: +34-954552862; Fax: +34-954377413; E-mail: mudarra{at}us.es
Keywords: antimicrobial resistance , zinc , biofilms
Sir,
Pseudomonas aeruginosa is a significant cause of urinary tract infections in patients with urinary catheters. Latex is the biomaterial most commonly used to make urinary catheters. We have observed that eluates from siliconized latex urinary catheters decrease the activity of carbapenems against P. aeruginosa.1 This effect is related to changes in the outer membrane protein (OMP) profile.2 This reversible phenomenon is caused by zinc eluted from siliconized latex urinary catheters, which triggers expression of the heavy metal efflux pump CzcCBA.3,4 This mechanism does not affect other antimicrobial agents. The clinical relevance of this phenomenon is currently under investigation. The potential effect of latex gloves on the characteristics of nosocomial pathogens could have certain epidemiological value. The purpose of this study is to evaluate the effect of latex gloves on the activity of imipenem against P. aeruginosa.
To determine bacterial survival on latex gloves, a 1 cm2 fragment of sterile powder-free latex glove (sempercare®, Austria) was placed on a layer of humid sterile gauze in sterile Petri dishes (humid chamber). On to each latex fragment was dropped 0.025 mL of a 0.5 McFarland suspension of P. aeruginosa PAO1, and the humid chambers were left at room temperature for up to 4 weeks. At specific intervals (24 and 48 h; 1, 2, 3 and 4 weeks), latex fragments were removed to tubes containing 2 mL of ice-cold sterile water, vortexed and sonicated in an ultrasonic bath. Bacterial survival was determined by colony counts on Mueller–Hinton (MH) agar plates after 24 h at 35°C. P. aeruginosa PAO1 was able to survive for a long time on the surface of clean latex gloves in the absence of any external nutrient source. After 28 days, >6 x 106 cfu per square centimetre of latex glove was obtained. It can be assumed that P. aeruginosa is able to utilize nutrients leaching from the latex glove to support growth.5
The effect of latex eluates on the in vitro activity of imipenem (Merck, Madrid, Spain) was determined by microdilution, according to the Clinical and Laboratory Standards Institute guidelines.6 MICs of imipenem determined in MH broth and latex eluates increased 8-fold (from 1 to 8 mg/L) when P. aeruginosa PAO1 was grown in latex eluates. To determine whether this effect was due to high zinc concentration, the amount of zinc in latex glove eluates was studied. For this purpose, eluates from latex gloves were prepared by incubating segments of latex glove in sterile cation-adjusted MH broth at 37°C for 24 h (one 5 cm2 segment in 100 mL of medium). The zinc contents in MH and in the eluates were determined using inductively coupled plasma atomic emission spectrometry with a sequential multielement instrument (ARL 3410; FISONS Instruments, USA). The zinc contents in MH and latex glove eluates were 0.28 ± 0.2 and 21.7 ± 2.1 mg/L, respectively. The MIC of imipenem in MH broth containing 22 mg/L zinc was also 8 mg/L. This effect, also observed with siliconized latex urinary catheters, was mediated by the high zinc concentrations reached on such surfaces and was partially due to the loss of OprD2.
It was recently shown that sublethal zinc concentrations induced in P. aeruginosa a reversible resistance to zinc, other metals and imipenem, whereas lethal zinc concentrations selected stable mutants resistant to these compounds.4 To study the OMP profile of P. aeruginosa biofilm on latex, bacterial biofilms were obtained by incubating three sterile 5 cm2 latex segments in 100 mL of MH broth inoculated with 
5 x 105 cfu/mL. After incubation (24 h, 35°C) latex fragments were washed four times in ice-cold sterile water to remove non-adherent bacteria, deposited in tubes containing 5 mL of 0.1 M phosphate buffer (pH 7.0) and sonicated in an ultrasonic bath. Bacteria were disrupted by sonication and cell membranes were recovered by ultracentrifugation. OMPs were obtained after treatment of cell membranes with N-lauryl sarcosine and ultracentrifugation. The OMP profile was determined using SDS–PAGE with 10% (w/v) acrylamide and 0.1% (w/v) bis-acrylamide in the running gel. The gels were stained with Coomassie Blue. P. aeruginosa biofilm and bacteria grown in MH broth supplemented with 22 mg/L zinc lost OprD2 and expressed OprD3 (Figure 1). This phenomenon was reversible upon re-growth of the cells in zinc-free medium. The underlying mechanism for the OprD2 loss in the presence of zinc seems to be the simultaneous expression of the efflux pump, CzcCBA, which belongs to the resistance, nodulation, cell division (RND) transporter family, and the down-regulation of the expression of OprD2, in order to prevent intracellular accumulation of zinc.4 Co-regulation between heavy metal and antibiotic resistance might have important clinical implications—a possibility that is currently under investigation.
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In summary, zinc eluted from latex gloves induced resistance to imipenem in P. aeruginosa. Whether this reversible phenomenon has any relevance to the prevalence of P. aeruginosa resistant to imipenem in hospitals remains unknown, but it is possible that resistance to imipenem can be induced on different sorts of supports in the hospital environment.
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None to declare.
Acknowledgements
This work was supported by the Dirección General de Investigación del Ministerio de Ciencia y Tecnología, Spain (project SAF2003-01241), and it was developed under the auspices of REIPI (Spanish Network for Research in Infectious Diseases C013/14), Instituto de Salud Carlos III, Ministerio de Salud y Consumo, Spain.
References
1.
Martinez-Martinez L, Pascual A, Conejo MC et al. Resistance of Pseudomonas aeruginosa to imipenem induced by eluates from siliconized latex urinary catheters is related to outer membrane protein alterations. Antimicrob Agents Chemother 1999; 43: 397–99.
2. Conejo MC, Martinez-Martinez L, Garcia I et al. Effect of siliconized latex urinary catheters on the activity of carbapenems against Pseudomonas aeruginosa strains with defined mutations in ampC, oprD, and genes coding for efflux systems. Int J Antimicrob Agents. 2003; 22: 122–27.[Medline]
3.
Conejo MC, Garcia I, Martinez-Martinez L et al. Zinc eluted from siliconized latex urinary catheters decreases OprD expression, causing carbapenem resistance in Pseudomonas aeruginosa. Antimicrob Agents Chemother 2003; 47: 2313–15.
4.
Perron K, Caille O, Rossier C et al. CzcR-CzcS, a two-component system involved in heavy metal and carbapenem resistance in Pseudomonas aeruginosa. J Biol Chem 2004; 279: 8761–8.
5. Martinez-Martinez L, Pascual A, Perea EJ. Effect of three plastic catheters on survival and growth of Pseudomonas aeruginosa. J Hosp Infect 1990; 16: 311–18.[CrossRef][Medline]
6. Clinical and Laboratory Standards Institute. Performance Standards for Antimicrobial Susceptibility Testing: Fifteenth Informational Supplement M100-S15. CLSI, Wayne, PA, USA, 2005.
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