The role of GpsB in Staphylococcus aureus cell morphogenesis.

For decades, cells of the Gram-positive bacterial pathogen Staphylococcus aureus were thought to lack a dedicated elongation machinery. However, S. aureus cells were recently shown to elongate before division, in a process that requires a shape elongation division and sporulation (SEDS)/penicillin-binding protein (PBP) pair for peptidoglycan synthesis, consisting of the glycosyltransferase RodA and the transpeptidase PBP3. In ovococci and rod-shaped bacteria, the elongation machinery, or elongasome, is composed of various proteins besides a dedicated SEDS/PBP pair. To identify proteins required for S. aureus elongation, we screened the Nebraska Transposon Mutant Library, which contains transposon mutants in virtually all non-essential staphylococcal genes, for mutants with modified cell shape. We confirmed the roles of RodA/PBP3 in S. aureus elongation and identified GpsB, SsaA, and RodZ as additional proteins involved in this process. The gpsB mutant showed the strongest phenotype, mediated by the partial delocalization from the division septum of PBP2 and PBP4, two penicillin-binding proteins that synthesize and cross-link peptidoglycan. Increased levels of these PBPs at the cell periphery versus the septum result in higher levels of peptidoglycan insertion/crosslinking throughout the entire cell, possibly overriding the RodA/PBP3-mediated peptidoglycan synthesis at the outer edge of the septum and/or increasing stiffness of the peripheral wall, impairing elongation. Consequently, in the absence of GpsB, S. aureus cells become more spherical. We propose that GpsB has a role in the spatio-temporal regulation of PBP2 and PBP4 at the septum versus cell periphery, contributing to the maintenance of the correct cell morphology in S. aureus. IMPORTANCE: Staphylococcus aureus is a Gram-positive clinical pathogen, which is currently the second cause of death by antibiotic-resistant infections worldwide. For decades, S. aureus cells were thought to be spherical and lack the ability to undergo elongation. However, super-resolution microscopy techniques allowed us to observe the minor morphological changes that occur during the cell cycle of this pathogen, including cell elongation. S. aureus elongation is not required for normal growth in laboratory conditions. However, it seems to be essential in the context of some infections, such as osteomyelitis, during which S. aureus cells apparently elongate to invade small channels in the bones. In this work, we uncovered new determinants required for S. aureus cell elongation. In particular, we show that GpsB has an important role in the spatio-temporal regulation of PBP2 and PBP4, two proteins involved in peptidoglycan synthesis, contributing to the maintenance of the correct cell morphology in S. aureus.

Differences in the information about procedures after cold chain disruption provided by pharmaceutical industry to hospital and community pharmacies.

PURPOSE: To compare the response of the pharmaceutical industry to information requests from a hospital pharmacy and a community pharmacy regarding the inadvertent exposure of refrigerated medicines (2-8°C) to out-of-range temperatures. METHODS: A complete list of all authorised medicines labelled for refrigeration was obtained from the Portuguese Medicines Agency. A standard information request regarding cold chain disruption for refrigerated medicines was sent to pharmaceutical companies from a hospital and a community pharmacy in Portugal. For companies who did not provide a response within the first 45 days, a second request was sent and an additional 45 days were allowed before completing data collection. To compare information received from the drug industry with that contained in the official European labelling, the Summaries of Product Characteristics (SmPCs) were retrieved from the regulatory agencies' databases. Response rate, response time and information appropriateness were assessed. RESULTS: A total of 792 medicines marketed by 70 different pharmaceutical companies were included. The hospital pharmacy received 560 (70.7%) responses, with a mean response time of 6.5 days (SD=5.3) for the first request. The community pharmacy obtained a response for 411 (51.9%), with a mean response time of 15.5 days (SD=4.8). More appropriate information was provided to hospital pharmacy requests. SmPCs did not contain complete information regarding the inadvertent exposure of medicines to unrefrigerated conditions. CONCLUSIONS: When enquired about a specific piece of information, the pharmaceutical industry provided quicker, higher quality and more frequent responses to a hospital pharmacy compared with a community pharmacy.