Bacteria can sense and adapt to environmental change using second messenger signaling networks. Among them, cyclic di-adenosine monophosphate (c-di-AMP) was discovered in 2008 and presents ubiquitously in Firmicutes and several Gram-negative bacteria as well as archaea. It plays numerous important roles in bacteria including osmoregulation, central metabolism, DNA integrity, cell wall homeostasis, virulence, biofilm formation, and antibiotic susceptibility in many bacteria through direct binding to a number of proteins and riboswitches. It also activates an immune response in hosts by interaction with the stimulator of interferon genes (STING) which could be potential for vaccine adjuvant.
In some pathogens, c-di-AMP was reported to regulate β-lactam antibiotic resistance, however, the mechanism is unclear. Here, we reveal an intimate connection between the osmolyte pool and cefuroxime resistance in L. lactis. Our results showed that c-di-AMP synthesis defective strains of the model bacterium Lactococcus lactis could be rescued from cefuroxime (CEF) sensitivity following suppressor mutation of the glutamine transporter (GlnPQ). Metabolite analyses revealed significantly lower free glutamate (Glu) and aspartate (Asp) levels in CEF resistant suppressors. The accumulation of these osmolytes causes greater cell lysis upon CEF-induced damage or in hypotonic conditions. This provides evidence that the over-accumulation of the major free amino acids in L. lactis elevates cell turgor pressure which is the major reason for CEF sensitivity in the cell with depleted c-di-AMP levels.