Urinary tract infections affect >150 million individuals annually. Hospital-acquired UTIs are mainly caused by Escherichia coli and Enterococcus faecalis and result in intracellular and catheter associated biofilm formation. A strong correlation exists between bladder epithelia invasion and recurrent-UTIs, as intracellular bacteria often resurface and recolonise post-antibiotic treatment.
We investigated if the antioxidant N-acetyl cysteine (NAC) can prevent uropathogenic bladder invasion, given its proven potential as an antibiofilm agent. Bladder epithelial cells (BECs) were infected with 107 bacteria and exposed to NAC for two hours prior to enumeration of intracellular bacterial load. NAC completely inhibited BEC invasion by multiple E. coli and E. faecalis clinical strains in a dose-dependent manner. This was also evident when bacterial invasion was visualised using GFP-tagged E.coli. Therefore NAC prevented BEC invasion and subsequent intracellular bacterial colony (IBC) formation (ie., intracellular biofilms), preventing bacterial reservoirs from accumulating and recolonising post-treatment.
We also found that NAC displayed no cytotoxicity against bladder cells even at 50mM, despite its intrinsic acidity, with >90% cellular viability after 48hrs exposure. BECs may have greater acidic tolerance, as urine pH is ca. 4.5.
Moreover, NAC prevented biofilm formation by E. coli and E. faecalis, and significantly reduced bacterial loads in 48hr biofilms when combined with ciprofloxacin. NAC visibly damaged E. coli and E. faecalis bacterial membranes, with a three-fold increase in propidium iodide (dead-cell marker) stained cells following NAC exposure.
We also showed NAC has a direct significant effect on biofilm matrix components (extracellular-DNA and polysaccharide), suggesting a mechanism whereby NAC breaks down pre-formed biofilm matrices. Therefore, combining NAC with an antibiotic could enhance antibiotic activity by disrupting the biofilm matrix and increasing bacterial antibiotic exposure.
NAC is a non-toxic antibiofilm agent, and this ability to prevent cell invasion and IBC formation by uropathogens provides a novel and efficacious approach to UTI treatments. Combining it with an antibiotic provides a two-pronged approach to disrupt bacterial biofilms and eliminate residual bacteria.