Cyclic di-adenosine monophosphate (c-di-AMP) is a signalling molecule that is important for the survival of Firmicutes. Previous work has shown that the gene(s) coding for the di-adenylate cyclase enzyme(s) could not be mutated when bacteria are grown in standard rich medium. My previous work has revealed that also in Staphylococcus aureus it is not possible to delete the dacA gene coding for the sole c-di-AMP cyclase in rich medium. However, it was possible to generate a dacA mutant strain in chemically defined medium. Using the severe growth defect of this mutant in rich medium, suppressor strains with improved growth under these conditions were obtained. Inactivating mutations that bypass the essentiality of dacA were identified in the alsT and opuD genes using a whole genome sequencing approach. As part of this work, I could show that these genes code for the main glutamine (AlsT) and glycine betaine (OpuD) transporters in S. aureus. Thus, these results revealed that S. aureus can survive without c-di-AMP when glutamine or glycine betaine uptake is prevented. Using bacterial cell size as a proxy for their osmotic balance, it was further shown that inactivation of OpuD helps bacteria to re-establish their osmotic balance, while inactivation of AlsT does not and bacteria remain enlarged. Glutamine and glutamate levels serve as a key indicator of nitrogen availability in bacterial cells. Therefore, with this work, not only a further connection between the c-di-AMP signalling network and osmotic regulation in S. aureus was established but also to central nitrogen metabolism. Other suppressor mutations were in hepS, hemB, ctaA and qoxB, which code for respiration related proteins. Expanding on this, I could show that dacA is dispensable in anaerobic growth conditions. This finding provides a novel link between the c-di-AMP signalling network and its requirement for aerobic but not anaerobic growth in S. aureus.