Transcriptomic and metabolic responses of Staphylococcus aureus exposed to supra-physiological temperatures.

BACKGROUND: Previous evaluation by different molecular and physiological assays of Staphylococcus aureus (S. aureus) responses to heat shock exposure yielded a still fragmentary view of the mechanisms determining bacterial survival or death at supra-physiological temperatures. This study analyzed diverse facets of S. aureus heat-shock adjustment by recording global transcriptomic and metabolic responses of bacterial cultures shifted for 10 min from 37 degrees C to a sub-lethal (43 degrees C) or eventually lethal (48 degrees C) temperature. A relevant metabolic model of the combined action of specific stress response mechanisms with more general, energy-regulating metabolic pathways in heat-shocked S. aureus is presented. RESULTS: While S. aureus cultures shifted to 43 degrees C or left at 37 degrees C showed marginal differences in growth and survival rates, bacterial cultures exposed to 48 degrees C showed a rapid growth arrest followed by a subsequent decline in viable counts. The most substantial heat shock-induced changes at both 43 degrees C and 48 degrees C occurred in transcript levels of HrcA- and CtsR-regulated genes, encoding classical chaperones DnaK and GroESL, and some Hsp100/Clp ATPases components, respectively. Other metabolic pathways up-regulated by S. aureus exposure at 48 degrees C included genes encoding several enzymes coping with oxidative stress, and DNA damage, or/and impaired osmotic balance. Some major components of the pentose phosphate cycle and gluconeogenesis were also up-regulated, which reflected depletion of free glucose by bacterial cultures grown in Mueller-Hinton broth prior to heat shock. In contrast, most purine- and pyrimidine-synthesis pathway components and amino acyl-tRNA synthetases were down-regulated at 48 degrees C, as well as arginine deiminase and major fermentative pathway components, such as alcohol, lactate and formate dehydrogenases. Despite the heat-induced, increased requirements for ATP-dependent macromolecular repair mechanisms combined with declining energy sources, intracellular ATP levels remained remarkably constant during heat shock. CONCLUSION: The sequential loss of replication and viability at 48 degrees C cannot be explained by significant reductions in intracellular ATP levels, but may reflect ATP rerouting for macromolecular repair mechanisms and cell survival. Our metabolic model also suggests that heat-stressed S. aureus should down-regulate the production of potential, DNA-damaging reactive oxygen species that might result from electron transport-generated ATP, involving excessive levels of free heavy metals, in particular iron.

Vancomycin-induced DRESS syndrome in a female patient.

BACKGROUND: DRESS syndrome (drug rash with eosinophilia and systemic symptoms) is a hypersensitivity reaction with skin rashes, eosinophilia, fever, lymph node enlargement and internal organ involvement. CASE REPORT: A 60-year-old diabetic woman was hospitalized at the University Hospitals of Geneva for mid-leg amputation due to peripheral arterial occlusive disease. No drug allergy was reported. Because of a wound infection by methicillin-resistant Staphylococcus aureus, treatment with vancomycin (2 g/day) in continuous perfusion was initiated. Approximately 2 weeks later, she developed a toxidermia with fever, a progressive maculopapular skin rash, eosinophilia and acute renal insufficiency. The skin biopsy revealed a necrosis with lymphocytic and eosinophilic infiltrations, supporting the suspicion of DRESS syndrome. A cure was achieved by the withdrawal of vancomycin and the administration of methylprednisolone (1 g/day), antihistaminics and topical mometasone, without the introduction of other antibiotics. CONCLUSION: Vancomycin can be a cause of DRESS syndrome. A high index of suspicion is warranted in order not to miss this potentially lethal disease.

A global view of Staphylococcus aureus whole genome expression upon internalization in human epithelial cells.

BACKGROUND: Staphylococcus aureus, a leading cause of chronic or acute infections, is traditionally considered an extracellular pathogen despite repeated reports of S. aureus internalization by a variety of non-myeloid cells in vitro. This property potentially contributes to bacterial persistence, protection from antibiotics and evasion of immune defenses. Mechanisms contributing to internalization have been partly elucidated, but bacterial processes triggered intracellularly are largely unknown. RESULTS: We have developed an in vitro model using human lung epithelial cells that shows intracellular bacterial persistence for up to 2 weeks. Using an original approach we successfully collected and amplified low amounts of bacterial RNA recovered from infected eukaryotic cells. Transcriptomic analysis using an oligoarray covering the whole S. aureus genome was performed at two post-internalization times and compared to gene expression of non-internalized bacteria. No signs of cellular death were observed after prolonged internalization of Staphylococcus aureus 6850 in epithelial cells. Following internalization, extensive alterations of bacterial gene expression were observed. Whereas major metabolic pathways including cell division, nutrient transport and regulatory processes were drastically down-regulated, numerous genes involved in iron scavenging and virulence were up-regulated. This initial adaptation was followed by a transcriptional increase in several metabolic functions. However, expression of several toxin genes known to affect host cell integrity appeared strictly limited. CONCLUSION: These molecular insights correlated with phenotypic observations and demonstrated that S. aureus modulates gene expression at early times post infection to promote survival. Staphylococcus aureus appears adapted to intracellular survival in non-phagocytic cells.

Tolerance of staphylococci to bactericidal antibiotics.

Antibiotic therapy for deep-seated staphylococcal infections, especially when they are associated with artificial devices used for orthopedic surgery is often associated with failure. Standard anti-staphylococcal bactericidal antibiotics, such as semi-synthetic penicillins, cephalosporins, or glycopeptides, are effective when given prophylactically in clinical conditions or experimental trials of implant-related infections. However, the efficacy of all anti-staphylococcal agents is seriously diminished on already established implant-related deep-seated infections, which then frequently require surgical implant removal to obtain a cure. The failure of antibiotic therapy to cure established staphylococcal foreign-body infections may arise in part from a broad-spectrum phenotypic tolerance expressed in vivo to different classes of antimicrobial agents. The molecular and physiological mechanisms of this in vivo tolerance remain poorly understood.

The discriminatory power of MALDI-TOF mass spectrometry to differentiate between isogenic teicoplanin-susceptible and teicoplanin-resistant strains of methicillin-resistant Staphylococcus aureus.

To explore the discriminatory power of matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) for detecting subtle differences in isogenic isolates, we tested isogenic strains of Staphylococcus aureus differing in their expression of resistance to methicillin or teicoplanin. More important changes in MALDI-TOF MS spectra were found with strains differing in methicillin than in teicoplanin resistance. In comparison, very minor or no changes were recorded in pulsed-field gel electrophoresis profiles or peptidoglycan muropeptide digest patterns of these strains, respectively. MALDI-TOF MS might be useful to detect subtle strain-specific differences in ionizable components released from bacterial surfaces and not from their peptidoglycan network.

Evidence of an intracellular reservoir in the nasal mucosa of patients with recurrent Staphylococcus aureus rhinosinusitis.

Severe infections due to Staphylococcus aureus require prolonged therapy for cure, and relapse may occur even years after the first episode. Persistence of S. aureus may be explained, in part, by nasal carriage of S. aureus, which occurs in a large percentage of healthy humans and represents a major source of systemic infection. However, the persistence of internalized S. aureus within mucosal cells has not been evaluated in humans. Here, we provide the first in vivo evidence of intracellular reservoirs of S. aureus in humans, which were assessed in endonasal mucosa specimens from patients suffering from recurrent S. aureus rhinosinusitis due to unique, patient-specific bacterial clonotypes. Heavily infected foci of intracellular bacteria located in nasal epithelium, glandular, and myofibroblastic cells were revealed by inverted confocal laser scan fluorescence and electron microscopic examination of posttherapy intranasal biopsy specimens from symptom-free patients undergoing surgery on the sinuses. Intracellular residence may provide a sanctuary for pathogenic bacteria by protecting them from host defense mechanisms and antibiotic treatment during acute, recurrent S. aureus rhinosinusitis.

Trends in the treatment of orthopaedic prosthetic infections.

The most commonly used therapy for prosthetic joint infection is a two-stage prosthetic exchange separated by 6 weeks of intravenous antibiotic therapy. This often results in long periods of hospitalization, morbidity, severe functional impairment and sometimes increased mortality. Therefore novel and challenging therapeutic approaches have been attempted, particularly in hip prosthetic infection. This includes, whenever possible, according to the type of microorganism, antibacterial susceptibility and clinical presentation (including age and comorbidities): (i) less aggressive surgical techniques (debridement and prosthesis retention, or re-implantation with a single-stage exchange arthroplasty); and (ii) antibiotic combinations active against biofilm-associated bacteria, including rifampicin (particularly with quinolones) with excellent bio-availability which allow prolonged and efficient oral therapy.

An outbreak of Staphylococcus aureus in a pediatric cardiothoracic surgery unit.

OBJECTIVE: To investigate an outbreak of Staphylococcus aureus surgical-site infections. DESIGN: Case-control study. SETTING: Pediatric cardiothoracic surgery service of a tertiary-care university medical center. METHOD: Molecular typing was used to identify healthcare workers who carried the epidemic strain. RESULTS: Three children acquired surgical-site infections caused by a single strain of S. aureus. Fourteen (25%) of the staff members in the operating room and 17 (11%) on nursing units carried the epidemic strain (P = .01). A case-control study identified 4 healthcare workers who were associated statistically with the outbreak, 2 of whom (a cardiothoracic surgeon and a perfusionist) carried the epidemic strain in their nares. The surgeon also carried the epidemic strain on his hands. Each staff member who carried the epidemic strain was treated with mupirocin; those carrying the strain on their hands were required to wash their hands with chlorhexidine. The surgeon was not allowed to perform surgery until 2 of his hand cultures did not grow S. aureus. CONCLUSIONS: Only three children were infected with the epidemic strain, but it was disseminated widely among staff who cared for children who underwent cardiothoracic surgery. No additional cases were identified after staff members who carried the epidemic strain were decolonized. Both classic epidemiologic methods and molecular typing techniques were necessary to identify the source and extent of this outbreak.