Targeting NAD+ regeneration enhances antibiotic susceptibility of Streptococcus pneumoniae during invasive disease.

Anaerobic bacteria are responsible for half of all pulmonary infections. One such pathogen is Streptococcus pneumoniae (Spn), a leading cause of community-acquired pneumonia, bacteremia/sepsis, and meningitis. Using a panel of isogenic mutants deficient in lactate, acetyl-CoA, and ethanol fermentation, as well as pharmacological inhibition, we observed that NAD(H) redox balance during fermentation was vital for Spn energy generation, capsule production, and in vivo fitness. Redox balance disruption in fermentation pathway-specific fashion substantially enhanced susceptibility to killing in antimicrobial class-specific manner. Blocking of alcohol dehydrogenase activity with 4-methylpyrazole (fomepizole), an FDA-approved drug used as an antidote for toxic alcohol ingestion, enhanced susceptibility of multidrug-resistant Spn to erythromycin and reduced bacterial burden in the lungs of mice with pneumonia and prevented the development of invasive disease. Our results indicate fermentation enzymes are de novo targets for antibiotic development and a novel strategy to combat multidrug-resistant pathogens.

Efficacy of Intraoperative Antiseptic Techniques in the Prevention of Periprosthetic Joint Infection: Superiority of Betadine.

BACKGROUND: Povidone-iodine (PI), chlorhexidine gluconate (CHG), and vancomycin (VANC) powder are common intrawound prophylactic agents to prevent periprosthetic joint infection during primary total joint arthroplasty. The aims of this study are (1) to determine the minimal inhibitory concentration (MIC) and time to death for PI, CHG, and VANC against multiple bacteria and (2) to determine time to death against bacteria dried on titanium discs. METHODS: A standard quantitative suspension assay was performed to determine the MIC for PI, CHG, and VANC against methicillin-resistant Staphylococcus aureus (MRSA), Staphylococcus epidermidis, Haemophilus influenzae, Pseudomonas aeruginosa, Burkholderia cepacia, and Escherichia coli. Time to death assay was performed with time points of 0, 3, 30, and 60 minutes. Concentrations of antiseptic agents for time to death assay were 1% PI, 0.05% CHG, and 5 µg/mL VANC. Dry-phase bacteria on titanium discs were treated in a similar fashion. RESULTS: The MIC of PI was 0.63%, CHG was 0.0031%, and VANC was 1.56 µg/mL. All 7 bacterial isolates were completely killed by PI at all times tested. CHG failed to kill MRSA and B cepacia at 0- and 3-minute exposures. Vancomycin completely killed MRSA and S epidermidis isolates between 18-20 hours of exposure. All bacterial isolates dried on titanium discs were eliminated by PI exposure on contact. E coli and S epidermidis were incompletely eliminated by CHG at 0 minutes, with all isolates eliminated at 3, 10, and 30 minutes. CONCLUSION: Our study suggests that PI kills all bacteria tested immediately on contact and that the exposure time is not the key factor.

Structure-Function Relationships in the Oligomeric NADPH-Dependent Assimilatory Sulfite Reductase.

The central step in the assimilation of sulfur is a six-electron reduction of sulfite to sulfide, catalyzed by the oxidoreductase NADPH-dependent assimilatory sulfite reductase (SiR). SiR is composed of two subunits. One is a multidomain flavin binding reductase (SiRFP) and the other an iron-containing oxidase (SiRHP). Both enzymes are primarily globular, as expected from their functions as redox enzymes. Consequently, we know a fair amount about their structures but not how they assemble. Curiously, both structures have conspicuous regions that are structurally undefined, leaving questions about their functions and raising the possibility that they are critical in forming the larger complex. Here, we used ultraviolet-visible and circular dichroism spectroscopy, isothermal titration calorimetry, proteolytic sensitivity tests, electrospray ionization mass spectrometry, and activity assays to explore the effect of altering specific amino acids in SiRFP on their function in the holoenzyme complex. Additionally, we used computational analysis to predict the propensity for intrinsic disorder within both subunits and found that SiRHP's N-terminus is predicted to have properties associated with intrinsic disorder. Both proteins also contained internal regions with properties indicative of intrinsic disorder. We showed that SiRHP's N-terminal disordered region is critical for complex formation. Together with our analysis of SiRFP amino acid variants, we show how molecular interactions outside the core of each SiR globular enzyme drive complex assembly of this prototypical oxidoreductase.

Repurposing sunscreen as an antibiotic: zinc-activated avobenzone inhibits methicillin-resistant Staphylococcus aureus.

Methicillin-resistant Staphylococcus aureus (MRSA) is a major healthcare concern with associated healthcare costs reaching over ${\$}$1 billion in a single year in the USA. Antibiotic resistance in S. aureus is now observed against last line of defense antibiotics, such as vancomycin, linezolid, and daptomycin. Unfortunately, high throughput drug discovery approaches to identify new antibiotics effective against MRSA have not resulted in much tangible success over the last decades. Previously, we demonstrated the feasibility of an alternative drug discovery approach, the identification of metallo-antibiotics, compounds that gain antibacterial activity only after binding to a transition metal ion and as such are unlikely to be detected in standard drug screens. We now report that avobenzone, the primary active ingredient of most sunscreens, can be activated by zinc to become a potent antibacterial compound against MRSA. Zinc-activated avobenzone (AVB-Zn) potently inhibited a series of clinical MRSA isolates [minimal inhibitory concentration (MIC): 0.62-2.5 µM], without pre-existing resistance and activity without zinc (MIC: >10 µM). AVB-Zn was also active against clinical MRSA isolates that were resistant against the commonly used zinc-salt antibiotic bacitracin. We found AVB-Zn exerted no cytotoxicity on human cell lines and primary cells. Last, we demonstrate AVB-Zn can be deployed therapeutically as lotion preparations, which showed efficacy in a mouse wound model of MRSA infection. AVB-Zn thus demonstrates Zn-activated metallo-antibiotics are a promising avenue for future drug discovery.

Predictors of patient satisfaction with pain management and improvement 3 months after burn injury.

UNLABELLED: Very little is known about what influences patient satisfaction with burn pain management. The aim of this prospective study was to examine predictors of patient satisfaction with pain management following burn injury. METHODS: Participants were 97 adult burn patients admitted to Royal Perth Hospital, Western Australia between June 2007 and February 2009. Key patient satisfaction domains were: pain treatment in hospital, pain medication provided, and improvement since hospitalisation. Acute pain medication use, pain severity, psychological symptoms, and patient expectations were assessed as potential predictors of patient satisfaction. RESULTS: Patients reported moderate to high levels of satisfaction across all domains. Whether treatment matched patients' expectations was a significant predictor of satisfaction with pain treatment, pain medication and improvement (p < .001, p < .001, p < .05 respectively). Current pain at follow-up was a significant predictor of satisfaction with pain treatment and improvement (p < .01, p < .001 respectively). Acute pain medication use, depressive symptoms and reductions in average pain at three months were not significant independent predictors of patient satisfaction with pain management and improvement in this sample (p > .35). Yet, severity of posttrauma symptoms at three months was a significant predictor of satisfaction with pain medication and was moderately and positively associated with satisfaction with pain treatment (p < .05, p = .07, respectively). DISCUSSION: These findings suggest that acute medication use and reductions in perceived pain symptoms are less closely related to patient satisfaction compared with treatment expectations, current pain and posttrauma symptoms. Collectively, these findings indicate a need to proactively address treatment expectations about pain management, and manage current pain and psychological distress following burn injury in order to improve patient satisfaction with care received.

The impact of personality and coping on the development of depressive symptoms in adult burns survivors.

This prospective study examined the extent to which the personality traits neuroticism, extraversion and agreeableness and coping styles approach, avoidant and ambivalent contribute to the development of depressive symptoms in adult burns survivors at three months post-injury. Participants were 70 adult burns survivors admitted to Royal Perth Hospital in Western Australia between June 2007 and February 2008. Personality was assessed using the NEO Personality Inventory-Revised (NEO-PI-R), coping was evaluated with the Coping with Burns Questionnaire (CBQ) and depressive symptoms were measured using The Centre for Epidemiologic Studies Depression Scale (CES-D). Twenty one percent of retained participants at three months (n=29) reported clinically significant depressive symptoms. There were no significant relationships between depressive symptoms at three months and demographic or burn characteristics. Neuroticism significantly predicted depressive symptoms at three-month follow-up and this relationship was significantly mediated by avoidant coping. In addition, extraversion, avoidant coping and approach coping were all significant and independent predictors of depressive symptoms at three months. These findings suggest that burns patients at greatest risk of developing clinically significant depressive symptoms may be identifiable in the acute recovery phase.