Resisting the resistance: The antimicrobial peptide DGL13K selects for small colony variants of Staphylococcus aureus that show increased resistance to its stereoisomer LGL13K, but not to DGL13K

About 30% of the population are nasal carriers of Staphylococcus aureus , a leading cause of bacteremia, endocarditis, osteomyelitis, skin and soft tissue infections. Antibiotic-resistant bacteria, in particular, are an increasing problem in both hospital and community settings. In this study, we sought to determine the cellular consequences of long-term exposure of S. aureus to the antimicrobial peptide stereo-isomers, DGL13K and LGL13K. Both peptides selected for mutations in the chorismate/menaquinone biosynthetic pathway, which resulted in increased resistance to LGL13K but not DGL13K. DGL13K-selected isolates showed a mutation in aroF , while menA and menH were mutated in LGL13K-selected isolates. The latter also contained a mutation of frsA . The peptide-selected isolates exhibited golden coloration, suggesting increased production of the carotenoid staphyloxanthin, which could enhance resistance to cationic AMPs. The peptide-selected isolates grew as small colony variants, which have also been associated with resistance to AMPs. Addition of menaquinone to the growth medium reduced the generation time of DGL13K-selected mutants, but not LGL13K-selected mutants. Instead, the latter showed greatly reduced ATP-levels, suggesting defective electron transport, which is also associated with menadione auxotrophism. The mechanisms behind the differential effect of DGL13K and LGL13K on S. aureus resistance remain to be elucidated. The finding that DGL13K induced resistance to the stereo-isomer LGL13K but not to DGL13K itself, suggests that peptide primary structure is responsible for inducing bacterial defense mechanisms but the peptide secondary structure determines if the defense mechanisms are effective against each peptide.