The genome-wide <i>de novo</i> mutations and <i>icaA</i> gene expression levels in <i>Staphylococcus aureus</i> under long-term subinhibitory and semisubinhibitory nisin exposure

The emergence and rapid spread of multidrug-resistant pathogens have caused a need for alternative antimicrobials, and bacteriocins are considered promising alternatives due to their lower risk of resistance development. Regarding this, we aimed to investigate the long-term subinhibitory and semisubinhibitory concentrations of a commonly used bacteriocin (nisin) in Staphylococcus aureus using an experimental evolution approach followed by genome sequencing. We then performed RT-qPCR to examine changes in the expression level of the biofilm-related icaA gene in evolved lines. We found that while nisin treatment did not significantly elevate the base-substitution rates, there was a significant decrease in the insertion/deletion rate in the lines exposed to the subinhibitory concentration of nisin. We also revealed an increase in nonsynonymous mutations in specific genes (e.g. sarS and cap8) associated with resistance and virulence mechanisms. Importantly, we observed a transition bias in the nisin-treated lines for the first time, and it may be related to the resistance development to nisin. RT-qPCR analysis of the icaA gene showed a reduced expression levels in nisin-treated groups, although the results were not statistically significant. These findings show the potential outcomes of nisin exposure in S. aureus and emphasize the need for careful consideration of bacteriocins in clinical practice.