Penicillin‐binding protein PBP2a provides variable levels of protection toward different β‐lactams in Staphylococcus aureus RN4220

Methicillin‐resistant Staphylococcus aureus (MRSA) is resistant to most β‐lactams due to the expression of an extra penicillin‐binding protein, PBP2a, with low β‐lactam affinity. It has long been known that heterologous expression of the PBP2a‐encoding mecA gene in methicillin‐sensitive S. aureus (MSSA) provides protection towards β‐lactams, however, some reports suggest that the degree of protection can vary between different β‐lactams. To test this more systematically, we introduced an IPTG‐inducible mecA into the MSSA laboratory strain RN4220. We confirm, by growth assays as well as single‐cell microfluidics time‐lapse microscopy experiments, that PBP2a expression protects against β‐lactams in S. aureus RN4220. By testing a panel of ten different β‐lactams, we conclude that there is also a great variation in the level of protection conferred by PBP2a. Expression of PBP2a resulted in an only fourfold increase in minimum inhibitory concentration (MIC) for imipenem, while a 32‐fold increase in MIC was observed for cefaclor and cephalexin. Interestingly, in our experimental setup, PBP2a confers the highest protection against cefaclor and cephalexin—two β‐lactams that are known to have a high specific affinity toward the transpeptidase PBP3 of S. aureus. Notably, using a single‐cell microfluidics setup we demonstrate a considerable phenotypic variation between cells upon β‐lactam exposure and show that mecA‐expressing S. aureus can survive β‐lactam concentrations much higher than the minimal inhibitory concentrations. We discuss possible explanations and implications of these results including important aspects regarding treatment of infection.