Methods: We grew, in tissue-culture wells, biofilms of the following organisms: methicillin-sensitive and –resistant Staphylococcus aureus (MSSA, MRSA), methicillin-resistant S. epidermidis (MRSE), vancomycin-resistant Enterococcus faecium (VRE), Pseudomonas aeruginosa, Enterobacter cloacae and Klebsiella pneumoniae. After 24 hours of incubation, the broth was replaced with either tryptic soy broth (TSB) containing NAC 80 mg/ml or TSB alone as control and then incubated for 24 hours. The wells were then rinsed and stained with propidium iodide (stains non-viable cells red) and FITC-labelled concanavalin A type IV (stains extracellular polysaccharide green). Using an upright confocal laser scanning microscope, five non-contiguous areas were viewed in each well. Each experiment was conducted thrice. We measured biofilm thickness, counted the number of non-viable bacterial cells, and measured green fluorescent intensity as a marker of extracellular polysaccharide in the biofilm matrix. All continuous variables were analyzed using a t-test.
Results: We noted statistically significant decrease in biofilm thickness of NAC-treated wells for all organisms (p<0.05) except VRE. We also noted an increase in the number of non-viable cells in the NAC-treated wells of gram-positive organisms. This was statistically significant for MRSA, MRSE, and VRE, with a trend for higher number of non-viable cells for MSSA. We noted scant cellularity in the NAC-treated gram-negative biofilms which also lacked complex 3-dimensional structures that were characteristic of the controls. Green fluorescent intensity was similar in the control and experimental arms.
Conclusion: NAC exposure leads to disruption of clinically relevant and drug-resistant bacterial biofilms. This involves increased cell death of biofilm-embedded gram-positive bacteria as well as disruption of complex structures. Polysaccharide content of the residual biofilm appears unchanged. NAC has the potential for use as a novel anti-infective agent for control of biofilm-related infections.
Baylor Coll. of Medicine (BCM): BCM owns the rights to a patent describing the use of N-acetylcysteine to combat device-related infections based on my invention. There is no licensing agreement regarding this patent between BCM and any company and hence no royalties. and Employee,
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