Background: The ability of staphylococci to grow in a wide range of salt concentrations is well documented. In this study, we aimed to clarify the role of cardiolipin (CL) in the adaptation of Staphylococcus aureus to high salinity. Results: Using an improved extraction method, the analysis of phospholipid composition suggested that CL levels increased slightly toward stationary phase, but that this was not induced by high salinity. Deletion of the two CL synthase genes, SA1155 (cls1) and SA1891 (cls2), abolished CL synthesis. The cls2 gene encoded the dominant CL synthase. In a cls2 deletion mutant, Cls1 functioned under stress conditions, including high salinity. Using these mutants, CL was shown to be unnecessary for growth in either basal or high-salt conditions, but it was critical for prolonged survival in high-salt conditions and for generation of the L-form. Conclusions: CL is not essential for S. aureus growth under conditions of high salinity, but is necessary for survival under prolonged high-salt stress and for the generation of L-form variants.

SCCmec is a large mobile genetic element that includes the mecA gene and confers resistance to β-lactam antibiotics in methicillin-resistant Staphylococcus aureus (MRSA). There is evidence that SCCmec disseminates among staphylococci, but the transfer mechanisms are unclear. Here, we show that two-component systems mediate the upregulation of natural competence genes in S. aureus under biofilm growth conditions, and this enhances the efficiency of natural transformation. We observe SCCmec transfer via natural transformation from MRSA, and from methicillin-resistant coagulase-negative staphylococci, to methicillin-sensitive S. aureus. The process requires the SCCmec recombinase genes ccrAB, and the stability of the transferred SCCmec varies depending on SCCmec types and recipients. Our results suggest that natural transformation plays a role in the transfer of SCCmec and possibly other mobile genetic elements in S. aureus biofilms.

The bacterial nucleoid contains several hundred kinds of nucleoid-associated proteins (NAPs), which play critical roles in genome functions such as transcription and replication. Several NAPs, such as Hu and H-NS in Escherichia coli, have so far been identified. Log- and stationary-phase cells of E. coli, Pseudomonas aeruginosa, Bacillus subtilis, and Staphylococcus aureus were lysed in spermidine solutions. Nucleoids were collected by sucrose gradient centrifugation, and their protein constituents analyzed by liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS). Over 200 proteins were identified in each species. Envelope and soluble protein fractions were also identified. By using these data sets, we obtained lists of contaminant-subtracted proteins enriched in the nucleoid fractions (csNAP lists). The lists do not cover all of the NAPs, but included Hu regardless of the growth phases and species. In addition, the csNAP lists of each species suggested that the bacterial nucleoid is equipped with the species-specific set of global regulators, oxidation-reduction enzymes, and fatty acid synthases. This implies bacteria individually developed nucleoid associated proteins toward obtaining similar characteristics. Ours is the first study to reveal hundreds of NAPs in the bacterial nucleoid, and the obtained data set enabled us to overview some important features of the nucleoid. Several implications obtained from the present proteomic study may make it a landmark for the future functional and evolutionary study of the bacterial nucleoid.

Bacterial genomic DNA is packed within the nucleoid of the cell along with various proteins and RNAs. We previously showed that the nucleoid in log phase cells consist of fibrous structures with diameters ranging from 30 to 80 nm, and that these structures, upon RNase A treatment, are converted into homogeneous thinner fibers with diameter of 10 nm. In this study, we investigated the role of major DNA-binding proteins in nucleoid organization by analyzing the nucleoid of mutant Escherichia coli strains lacking HU, IHF, H–NS, StpA, Fis, or Hfq using atomic force microscopy. Deletion of particular DNA-binding protein genes altered the nucleoid structure in different ways, but did not release the naked DNA even after the treatment with RNase A. This suggests that major DNA-binding proteins are involved in the formation of higher order structure once 10-nm fiber structure is built up from naked DNA.