ERP111720 PRJEB29420 ena-STUDY-IMPERIAL COLLEGE LONDON-29-10-2018-11:59:16:993-462 Inactivation of the monofunctional peptidoglycan glycosyltransferase SgtB allows Staphylococcus aureus to survive in the absence of lipoteichoic acid The cell wall of Staphylococcus aureus is composed of peptidoglycan and the anionic polymers lipoteichoic acid (LTA) and wall teichoic acid. LTA is required for growth and normal cell morphology in S. aureus. Strains lacking LTA are usually only viable when grown under osmotically stabilizing conditions or after the acquisition of compensatory mutations. LTA negative suppressor strains with inactivating mutations in gdpP, resulting in an increase in intracellular c-di-AMP levels, have been described previously. Here, we sought to identify factors other than c-di-AMP that allow S. aureus to survive without LTA. LTA-negative strains able to grow in un-supplemented medium were obtained and found to contain mutations in sgtB, mazE, clpX or vraT. The growth improvement through mutations in mazE and sgtB was confirmed by complementation analysis. We also show that an S. aureus sgtB transposon mutant, inactivated for the monofunctional peptidoglycan glycosyltransferase SgtB, displays a 4-fold increase in the MIC towards Oxacillin, suggesting that alteration in the peptidoglycan structure could help bacteria compensate for the lack of LTA. Muropeptide analysis of peptidoglycan isolated from a WT and sgtB mutant strains did not reveal any sizable alternations in the peptidoglycan structure. In contrast, the peptidoglycan isolated from an LTA-negative ltaS mutant strain showed a significant reduction in the fraction of highly crosslinked peptidoglycan, which was partially rescued in the sgtB ltaS double mutant suppressor strain. Taken together, these data point towards an important function of LTA in cell wall integrity through its requirement for proper peptidoglycan assembly. Staphylococcus aureus ltaS suppressor mutants The cell wall of Staphylococcus aureus is composed of peptidoglycan and the anionic polymers lipoteichoic acid (LTA) and wall teichoic acid. LTA is required for growth and normal cell morphology in S. aureus. Strains lacking LTA are usually only viable when grown under osmotically stabilizing conditions or after the acquisition of compensatory mutations. LTA negative suppressor strains with inactivating mutations in gdpP, resulting in an increase in intracellular c-di-AMP levels, have been described previously. Here, we sought to identify factors other than c-di-AMP that allow S. aureus to survive without LTA. LTA-negative strains able to grow in un-supplemented medium were obtained and found to contain mutations in sgtB, mazE, clpX or vraT. The growth improvement through mutations in mazE and sgtB was confirmed by complementation analysis. We also show that an S. aureus sgtB transposon mutant, inactivated for the monofunctional peptidoglycan glycosyltransferase SgtB, displays a 4-fold increase in the MIC towards Oxacillin, suggesting that alteration in the peptidoglycan structure could help bacteria compensate for the lack of LTA. Muropeptide analysis of peptidoglycan isolated from a WT and sgtB mutant strains did not reveal any sizable alternations in the peptidoglycan structure. In contrast, the peptidoglycan isolated from an LTA-negative ltaS mutant strain showed a significant reduction in the fraction of highly crosslinked peptidoglycan, which was partially rescued in the sgtB ltaS double mutant suppressor strain. Taken together, these data point towards an important function of LTA in cell wall integrity through its requirement for proper peptidoglycan assembly. ENA-FIRST-PUBLIC 2018-10-29 ENA-LAST-UPDATE 2018-10-29