Artificial sweeteners (AS) are synthetic sugar substitutes that are commonly consumed in the diet. Recent studies have indicated considerable health risks which links the consumption of AS with metabolic derangements and gut microbiota perturbations. Despite these studies, there is still limited data on how AS impacts the commensal microbiota to cause pathogenicity. The present study sought to investigate the role of commonly consumed AS on gut bacterial pathogenicity and gut epithelium-microbiota interactions, using models of microbiota ( Escherichia coli NCTC10418 and Enterococcus faecalis ATCC19433) and the intestinal epithelium (Caco-2 cells). Model gut bacteria were exposed to different concentrations of the AS saccharin, sucralose, and aspartame, and their pathogenicity and changes in interactions with Caco-2 cells were measured using in vitro studies. Findings show that sweeteners differentially increase the ability of bacteria to form a biofilm. Co-culture with human intestinal epithelial cells shows an increase in the ability of model gut bacteria to adhere to, invade and kill the host epithelium. The pan-sweet taste inhibitor, zinc sulphate, effectively blocked these negative impacts. Since AS consumption in the diet continues to increase, understanding how this food additive affects gut microbiota and how these damaging effects can be ameliorated is vital.