Use of a stainless steel washer platform to study Acinetobacter baumannii adhesion and biofilm formation on abiotic surfaces

Abstract

Acinetobacter baumannii is a frequent cause of hospital-acquired pneumonia, and has recently increased in incidence as the causative agent of severe disease in troops wounded in Afghanistan and Iraq. Clinical approaches are limited since A. baumannii strains isolated from patients are extremely resistant to current antimicrobials. A. baumannii can survive desiccation and during outbreaks has been recovered from various sites in the patients' environment. To better understand its prevalence in hospital settings, we used a stainless steel washer (SSW) platform to investigate A. baumannii biofilm formation on abiotic surfaces. Scanning electron microscopy demonstrated that A. baumannii forms strong biofilms on stainless steel surfaces. This platform was combined with a colorimetric 2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenylamino)carbonyl]-2H-tetrazolium hydroxide (XTT) reduction assay to observe the metabolic activity of bacterial cells, and to facilitate the manipulation and comparison of multiple A. baumannii clinical strains. A strong correlation between XTT and c.f.u. assays was demonstrated. To complement the cell viability assays, A. baumannii biofilm mass was measured by crystal violet staining. Furthermore, the effect of commonly used disinfectants and environmental stressors on A. baumannii biofilms and planktonic cells was compared and characterized. Biofilms on SSWs were significantly more resistant than their planktonic counterparts, providing additional evidence that may allow us to understand the high prevalence of this microbe in hospital settings. Our results validate that SSWs are a simple, versatile and innovative method to study A. baumannii biofilms in vitro.

Microbicides Alter the Expression and Function of RND-Type Efflux Pump AdeABC in Biofilm-Associated Cells of Acinetobacter baumannii Clinical Isolates

“…Biofilm-forming bacteria are more resistant to antimicrobials and disinfectants. A. baumannii biofilms can survive desiccation (1)(2)(3)9), and microorganisms have been recovered from the patients' environments, including bed curtains, furniture, and hospital equipment, during outbreaks (10). Disinfection of patient rooms has been successful in reducing outbreaks of A. baumannii, which underscores the role of the hospital environment as a reservoir for A. baumannii (10).…”

“…The isolates were derived from blood or wound cultures at the Walter Reed Medical Center (Washington, DC) or the Montefiore Medical Center (Bronx, NY). The antimicrobial susceptibility profile for each clinical isolate tested in this study was published previously (2). The strains were stored at Ϫ80°C in brain heart infusion (BHI) broth (BD, Franklin Lakes, NJ), with 40% glycerol, until use.…”

“…SSWs were used as a substrate to grow A. baumannii biofilms, as described previously (2). Briefly, for each strain, an inoculum of 4 ϫ 10 6 to 5 ϫ 10 6 A. baumannii cells was suspended in a 50-ml Erlenmeyer flask with 25 ml of TSB containing 6-mm-diameter SSWs.…”

“…We used SSWs as hard abiotic surfaces to grow the biofilms because this platform mimics the substrates found in hospital settings. This model was recently validated for study of the effects of commonly used disinfectants and environmental stressors on A. baumannii cells within biofilms (2). Furthermore, we investigated whether the expression and function of the AdeABC efflux pump in A. baumannii isolates was modified after treatment with microbicides, to identify the molecular mechanisms involved in microbial susceptibility to these antimicrobial agents.…”

“…The principal concern regarding A. baumannii is its ability to persist in the hospital environment, on various abiotic materials (1)(2)(3). This allows susceptible patients to come into contact with the microbe, which often results in outbreaks of ventilator-associated pneumonia, meningitis, septicemia, urinary tract infections, and wound infections (4).…”