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Objectives To determine the prevalence of penicillin susceptibility among MSSA causing bloodstream infections (BSIs) in 16 Spanish hospitals and to characterize the penicillin-susceptible MSSA (MSSA-PENS) isolates. Methods A total of 1011 Staphylococcus aureus isolates were collected from blood cultures in 16 Spanish hospitals during 2018–19 (6–12 months) and their susceptibility to 18 antimicrobials was determined. The MSSA-PENS isolates were selected and examined by PCR to determine the presence of the blaZ gene, other resistance genes and the genes lukF/lukS-PV, eta, etb and tst. The immune evasion cluster (IEC) type was also analysed. All the MSSA-PENS isolates were submitted to S. aureus protein A (spa) typing and the clonal complexes (CCs) were assigned according to their spa type. Results The prevalence of MSSA was 74.6% (754/1011) and 14.9% (151/1011) were MSSA-PENS-blaZnegative. MSSA-PENS-blaZnegative isolates (n = 151) were ascribed to 88 spa types and 11 CCs. The most frequent CCs were CC5 (35/151) and CC398 (25/151), with t002-CC5 and t571-CC398 being the most common lineages. Pan-susceptibility was identified in 117 of the 151 MSSA-PENS-blaZnegative isolates (77.5%). In the remaining isolates, erythromycin and clindamycin resistance was the most frequent resistance found, although tobramycin, ciprofloxacin, fusidic acid, mupirocin and/or tetracycline resistance was also detected. Thirty-eight MSSA-PENS-blaZnegative isolates were IEC negative and four isolates were Panton–Valentine leucocidin (‘PVL’) positive. Conclusions A high penicillin susceptibility rate was detected among MSSA, opening therapeutic opportunities for BSIs. The emergence of new successful MSSA-PENS clones could be responsible for these data. The detection among MSSA-PENS-blaZnegative isolates of the clonal lineage CC398 or the absence of an IEC raises questions about their possible animal origin, requiring further analysis.

Fusidane-type antibiotics represented by fusidic acid, helvolic acid, and cephalosporin P1 have very similar core structures, but they are produced by fungi belonging to different taxonomic groups. The origin and evolution of fusidane-type antibiotics biosynthetic gene clusters (BGCs) in different antibiotics producing strains remained an enigma. In this study, we investigated the distribution and evolution of the fusidane BGCs in 1,284 fungal genomes. We identified 12 helvolic acid BGCs, 4 fusidic acid BGCs, and 1 cephalosporin P1 BGC in Pezizomycotina fungi. Phylogenetic analyses indicated six horizontal gene transfer (HGT) events in the evolutionary trajectory of the BGCs, including 1) three transfers across Eurotiomycetes and Sordariomycetes classes; 2) one transfer between genera under Sordariomycetes class; and 3) two transfers within Aspergillus genus under Eurotiomycetes classes. Finally, we proposed that the ancestor of fusidane BGCs would be originated from the Zoopagomycota by ancient HGT events according to the phylogenetic trees of key enzymes in fusidane BGCs (OSC and P450 genes). Our results extensively clarify the evolutionary trajectory of fusidane BGCs by HGT among distantly related fungi and provide new insights into the evolutionary mechanisms of metabolic pathways in fungi.

Aims\ud \ud The aims of this study were to explore the development of bacterial resistance and cross‐resistance in four common human pathogens following realistic exposure to antibiotics found in over‐the‐counter (OTC) sore throat medicines: gramicidin, neomycin, bacitracin and tyrothricin.\ud Methods and Results\ud \ud Bacterial exposure to in‐use (concentration in the product before use) and diluted concentration (i.e. during use ) of antibiotic where conducted in broth for 24 h or until growth was visible. The changes in bacterial susceptibility profile before and after exposure was determined using standardized ISO microdilution broth. Antibiotic testing was performed according to EUCAST guidelines. We demonstrated that test bacteria were able to survive exposure to the in‐use concentrations of some antibiotics used in OTC medicines. Exposure to during use concentrations of bacitracin resulted in stable increase in minimal inhibitory concentration (MIC) (>8‐fold) in Staphylococcus aureus and Acinetobacter baumannii . Exposure to tyrothricin resulted in a stable increase in MIC (2·4‐fold) in Klebsiella pneumoniae , and exposure to neomycin resulted in a stable increase MIC (5000‐fold higher than the baseline) in Streptococcus pyogenes . Clinical cross‐resistance to other antibiotics (ciprofloxacin, fusidic acid, gentamicin, cefpodoxime, amoxicillin/clavulanic acid and cefotaxime) was also demonstrated following exposure to bacitracin or tyrothricin. Bacitracin exposure lead to a stable bacterial resistance after 10 passages.\ud Conclusions\ud \ud Our results indicate that OTC antibiotic medicines have the potential to drive resistance and cross‐resistance in vitro .\ud Significance and Impact of the Study\ud \ud Tackling antibiotic resistance is a high worldwide priority. It is widely accepted that the overuse and misuse of antibiotics increase the risk of the development and spread of antibiotic resistance within communities. A number of OTC sore throat products, widely available across the world for topical use in respiratory indications, contain locally delivered antibiotics. Our findings showed that these antibiotics in OTC medicines present a risk for emerging cross‐resistance in a number of bacterial respiratory pathogens.

Background Bacterial biofilm formation is a complicating factor in the antimicrobial treatment of bacterial infections. Objectives In this study, we assessed the impact of a novel hydrogel with the active antimicrobial compound JBC 1847 on eradication of preformed biofilms of Staphylococcus epidermidis, Cutibacterium acnes and MRSA in vitro, and evaluated the in vivo efficacy of MRSA wound treatment. Methods Biofilms were exposed to JBC 1847 for 24 h and subsequently the treatments were neutralized and surviving biofilm-associated bacteria recovered and enumerated. The efficacy of the hydrogel on post-treatment load of MRSA was determined in a murine model of MRSA wound infection, and skin samples of the infected mice were examined histologically to evaluate the degree of healing. Results A concentration-dependent eradication of biofilm-embedded bacteria by JBC 1847 was observed for all three pathogens, and the hydrogel caused a greater than four log reduction of cfu in all cases. In the mouse model, treatment with the hydrogel significantly reduced the cfu/mL of MRSA compared with treatment of MRSA-infected wounds with pure hydrogel. Histopathological analysis of the wounds showed that the JBC 1847 treatment group had a lower grade of inflammation, a higher mean score of re-epithelization and higher mean scores of parameters assessing the maturity of the newly formed epidermis, compared with both the fusidic acid 2% and vehicle treatment groups. Conclusions The novel hydrogel shows promising results as a candidate for future wound treatment, likely to be highly effective even in the case of biofilm-complicating infected wounds.

Background There is a lack of recently published data on impetigo presentation incidence and treatment practices in the routine Western European primary care setting. Objectives To investigate impetigo incidence, treatments and recurrence in primary care in the Netherlands. Methods A retrospective, observational study. Electronic records of patients treated for impetigo in 2015 at 29 general practices in Utrecht and surrounds were reviewed. An episode of impetigo was defined as one or more patient–doctor contacts within 8 weeks of the index consultation. Within an episode, patient demographics and prescribing patterns were analysed including number of treatments, and the category and sequence of individual medicines. Results A total of 1761 impetigo episodes were managed, with an incidence rate of 13.6 per 1000 person years. Impetigo peaked in summer. Most patients, the majority children, experienced a single episode (93%), and 25% had eczema as comorbidity. Topical antibiotics (primarily fusidic acid) were the most prescribed initial treatments (85%), followed by oral antibiotics (14%). Topical antibiotics were progressively used less over subsequent treatments, while there was an inverse increase in oral antibiotic use. Topical fusidic acid as the most common first line treatment seemed satisfactory as only 12% of initial treatments with this drug received further therapy. Repeat treatments generally occurred within 7 days. Conclusion This study of impetigo prescribing patterns in primary care highlighted that Dutch general practitioners were generally adherent to national treatment guidelines. Topical treatment, and if needed systemic small-spectrum antibiotic treatment, appeared satisfactory; these findings aid in antimicrobial stewardship.

Bacterial ribosome recycling requires breakdown of the post-termination complex (PoTC), comprising a messenger RNA (mRNA) and an uncharged transfer RNA (tRNA) cognate to the terminal mRNA codon bound to the 70S ribosome. The translation factors, elongation factor G and ribosome recycling factor, are known to be required for recycling, but there is controversy concerning whether these factors act primarily to effect the release of mRNA and tRNA from the ribosome, with the splitting of the ribosome into subunits being somewhat dispensable, or whether their main function is to catalyze the splitting reaction, which necessarily precedes mRNA and tRNA release. Here, we utilize three assays directly measuring the rates of mRNA and tRNA release and of ribosome splitting in several model PoTCs. Our results largely reconcile these previously held views. We demonstrate that, in the absence of an upstream Shine–Dalgarno (SD) sequence, PoTC breakdown proceeds in the order: mRNA release followed by tRNA release and then by 70S splitting. By contrast, in the presence of an SD sequence all three processes proceed with identical apparent rates, with the splitting step likely being rate-determining. Our results are consistent with ribosome profiling results demonstrating the influence of upstream SD-like sequences on ribosome occupancy at or just before the mRNA stop codon.

A topical pharmaceutical cream containing the active pharmaceutical ingredients (APIs) betamethasone-17-valerate and fusidic acid has been developed for the treatment of inflammatory skin conditions and associated secondary infections. In this work, a novel stability-indicating RP-HPLC method has been developed for the simultaneous quantitation of impurities of both APIs present in this cream. The HPLC column was a 150 mm × 4.6 mm I.D. YMC-Pack Pro C18 column with 3 µm particles. The column-oven temperature was maintained at 40°C and UV detection at 235 nm was used. A gradient programme was employed at a flow rate of 0.7 mL/min. Mobile phase A comprised of a 16:21:21:42 (v/v/v/v) mixture of methanol, 10 g/L phosphoric acid, HPLC grade water and acetonitrile. Mobile phase B comprised of a 24:5:5:66 (v/v/v/v) mixture of methanol, 10 g/L phosphoric acid, HPLC grade water and acetonitrile. The method has been validated according to current International Conference on Harmonisation (ICH) guidelines and applied during formulation development and stability studies. The procedure has been shown to be stability-indicating for the topical cream.

Objectives Increasing multidrug resistance amongst canine pathogenic staphylococci has renewed interest in topical antibacterial therapy for skin infections in the context of responsible veterinary prescribing. We therefore determined the activity in vitro of three clinically relevant topical agents and synergism between two of them against Staphylococcus pseudintermedius and Staphylococcus aureus. Methods The MICs of fusidic acid (n = 199), chlorhexidine (n = 198), miconazole (n = 198) and a 1:1 combination of miconazole/chlorhexidine (n = 198) were determined for canine isolates [50 MRSA and 49 methicillin-resistant S. pseudintermedius (MRSP), 50 MSSA and 50 methicillin-susceptible S. pseudintermedius (MSSP)] collected from the UK and Germany using an agar dilution method (CLSI VET01-A4). Fractional inhibitory concentration (FIC) indices were calculated to assess the interaction of miconazole with chlorhexidine. Results MICs of each drug/combination were significantly (P Conclusions Since the majority of these staphylococci, including methicillin-resistant isolates, had MICs that should be readily exceeded by topical skin application of these agents, their therapeutic efficacy for canine superficial pyoderma should be assessed. The synergistic interaction shown in vitro supports further clinical evaluation of miconazole/chlorhexidine combination therapy for staphylococcal infection.

Our aim was to assess national prescribing trends and determine longitudinal resistance patterns for topical antimicrobials in New Zealand. We observed a dramatic increase in fusidic acid (FA) resistance, and clonal expansion of FA-resistant Staphylococcus aureus. This increase was concurrent with a significant national increase in topical FA dispensing.

Translation elongation factor G (EF-G) in bacteria catalyses the translocation of transfer RNA on ribosomes in the elongation step as well as dissociation of post-termination state ribosomes into two subunits in the recycling step. In contrast, the dual functions of EF-G are exclusively divided into two different paralogues in human mitochondria, named EF-G1mt for translocation and EF-G2mt for ribosomal dissociation. Many of the two eukaryotic EF-G paralogues are phylogenetically associated with EF-G1mt and EF-G2mt groups. However, plant paralogues are associated with EF-G1mt and plastid EF-G, not with EF-G2mt. In this study, we phylogenetically and biochemically characterized Arabidopsis thaliana EF-G1mt (AtEF-G1mt) to clarify the factor responsible for the dissociation of ribosomes in plant mitochondria. We showed that eukaryotic EF-G1mts form one monophyletic group separated from bacterial EF-G and are classified into five sister groups. AtEF-G1mt is classified into a different group from its human counterpart. We also demonstrated that AtEF-G1mt catalyses both translocation and ribosomal dissociation, unlike in humans. Meanwhile, AtEF-G1mt is resistant to fusidic acid, an inhibitor of bacterial EF-G. Here, we propose that the functional division is not necessarily conserved among mitochondriate eukaryotes and also that EF-G1mt in organisms lacking EF-G2mt functions in two steps, similar to conventional bacterial EF-G.

Background Fusidic acid (FA) is an anti-staphylococcal agent used to treat chronic bone and joint infections (BJI) due to the availability of an oral formulation and its MRSA activity. Though used widely throughout the world for decades, FA is not approved in the USA. Methods To evaluate the safety and effectiveness of FA as chronic suppressive therapy in patients with staphylococcal BJI, we enrolled 30 patients in a prospective, single-arm, multi-center study in the USA. Eligible patients had refractory infections that could not be managed surgically and/or had not responded to previous antibiotic treatment. In Part A of the study, all patients received 6 months of oral FA treatment. In the first 1–2 weeks, patients could receive a companion antibiotic. Clinical success was based on lack of need for surgery or additional antibiotics. After all patients completed Part A of the study, an interim analysis was performed. In Part B of the study (ongoing), patients who completed Part A and require continued suppressive therapy may continue to receive FA for a total of 24 months. Results Most patients (83%) had orthopedic hardware infections. Therapy was considered successful at the 6-month visit in 18 patients (60%). Microbiological persistence was observed in eight patients, with three cases of decreasing FA susceptibility (including one case of resistance). Among 29 patients who experienced a treatment-emergent adverse event (TEAE), the most frequently reported events were: urinary tract infection (n = 9), peripheral edema/swelling (n = 8), nausea/dyspepsia (n = 7). Seven patients experienced TEAEs related to study drug; mild gastrointestinal disorders were most common. Two treatment-related events (unrelated to therapeutic failure) led to discontinuation of study drug Conclusion Patients with refractory BJI have few treatment options. In our study, 60% of infections were effectively suppressed for 6 months with FA treatment. The frequency of TEAEs was high, though not unexpected in this population with many chronic diseases. FA was well-tolerated with few patients experiencing treatment-related AEs leading to study drug discontinuation. FA administered chronically as monotherapy may lead to decreasing susceptibility and treatment failure in some patients; thus, combination therapy is warranted for this indication. Disclosures A. Sheets, Melinta Therapeutics: Employee, Salary. D. Graham, Cempra: Grant Investigator, Research grant. R. Darouiche, Cempra: Grant Investigator, Grant recipient. A. Strayer, Melinta Therapeutics, Inc.: Employee and Shareholder, Salary.

OBJECTIVES: Vancomycin-intermediate Staphylococcus aureus (VISA) and heterogeneous VISA (hVISA) phenotypes are increasingly reported in methicillin-resistant S. aureus (MRSA) strains of distinct genetic backgrounds. This study tracked genetic evolution during the development of vancomycin non-susceptibility in a prevalent Asian community-associated MRSA clone of sequence type (ST) 59. METHODS: ST59 strains were consecutively isolated from a patient who failed chemotherapy for a septic knee over 15 months. The genetic mutations associated with the VISA phenotype were identified by whole-genome sequencing of two strains, which had the vancomycin-susceptible S. aureus (VSSA) and VISA phenotypes. The mutations were subsequently screened in other strains. By correlating the accumulated mutations with vancomycin susceptibility, genetic evolution was tracked at the whole-genome scale. RESULTS: Nine non-synonymous mutations and two steps of genetic evolution were identified during the development of the VISA phenotype. The first step involved a nonsense mutation in agrC and point mutations at five other loci, which were associated with the VSSA-to-hVISA conversion. Mutations of rpoB and fusA following the use of rifampicin and fusidic acid were identified in the second step of evolution, which corresponded to the development of dual resistance to rifampicin and fusidic acid and the conversion of hVISA to VISA. CONCLUSIONS: In vivo genetic evolution of S. aureus occurred in stepwise order during the development of incremental vancomycin non-susceptibility and was related to the use of antimicrobial agents.

Ribosomes, after one round of translation, must be recycled so that the next round of translation can occur. Complete disassembly of post-termination ribosomal complex (PoTC) in yeast for the recycling consists of three reactions: release of tRNA, release of mRNA and splitting of ribosomes, catalyzed by eukaryotic elongation factor 3 (eEF3) and ATP. Here, we show that translocation inhibitors cycloheximide and lactimidomycin inhibited all three reactions. Cycloheximide is a non-competitive inhibitor of both eEF3 and ATP. The inhibition was observed regardless of the way PoTC was prepared with either release factors or puromycin. Paromomycin not only inhibited all three reactions but also re-associated yeast ribosomal subunits. On the other hand, sordarin or fusidic acid, when applied together with eEF2/GTP, specifically inhibited ribosome splitting without blocking of tRNA/mRNA release. From these inhibitor studies, we propose that, in accordance with eEF3’s known function in elongation, the release of tRNA via exit site occurs first, then mRNA is released, followed by the splitting of ribosomes during the disassembly of post-termination complexes catalyzed by eEF3 and ATP.

Objectives Research in residential homes has been limited to date and the extent of systemic and topical antimicrobial prescribing is largely unknown. The aim of this study was to investigate antimicrobial prescribing in residential homes in Northern Ireland (NI). Methods Point prevalence studies (PPSs) were completed in November 2010 (PPS1) and April 2011 (PPS2) in 30 residential homes. Data were obtained from care plans, medication administration records and staff in relation to antimicrobial prescribing and facility and resident characteristics, and analysed descriptively. Results The point prevalence of systemic antimicrobial prescribing was 9.4% in PPS1 and 9.2% in PPS2 (range 0.0%-33.3% during both PPSs). Trimethoprim was the most commonly prescribed systemic antimicrobial and the main indication was the prevention of urinary tract infections. Almost 25% of systemic antimicrobials were prescribed at inappropriate doses. The point prevalence of topical antimicrobial prescribing was 6.4% (range 0.0%-22.2%) in PPS1 and 5.9% (range 0.0%-21.1%) in PPS2. The most commonly prescribed topical antimicrobials were chloramphenicol eye preparations in PPS1 and fusidic acid skin preparations in PPS2; treatment with these topical antimicrobials was generally prolonged. More than 25% of all systemic and 55% of all topical antimicrobials were initiated following telephone consultations as opposed to face-to-face consultations. Conclusions The prevalence of systemic antimicrobial prescribing in residential homes in NI is relatively high compared with care homes (particularly nursing homes) in other countries. Systemic and topical antimicrobial prescribing is not always appropriate in terms of the doses prescribed and the duration of use. It is apparent that current strategies employed in NI are insufficient to ensure prudent antimicrobial prescribing within this environment.

BACKGROUND: Emergence of multidrug-resistant Staphylococcus aureus has triggered a reassessment of fusidic acid (CEM-102, sodium fusidate). METHODS: Fusidic acid was examined for (1) activity against recent methicillin-resistant S. aureus (MRSA) isolates; (2) modulation of activity by acidic pH; and (3) accumulation by phagocytic cells and intracellular activity against methicillin-susceptible S. aureus (MSSA) and MRSA. RESULTS: About 96% of strains (N = 94) were susceptible (European Committee on Antimicrobial Susceptibility Testing breakpoint [≤ 1 mg/L]). Activity was enhanced at pH 5.5 (6 dilutions decrease for minimum inhibitory concentration) in parallel with an increase of drug bacterial accumulation (opposite effects for clindamycin; linezolid remained unaffected). Fusidic acid accumulated in THP-1 cells (about 5.5 fold), with further accumulation at pH 5.5 vs pH 7.4. The intracellular activity of Fusidic acid was similar to that of clindamycin and linezolid (maximal relative activity, 0.4-0.6 log(10) colony-forming unit decrease). No cross-resistance to vancomycin or daptomycin was observed. CONCLUSIONS: Fusidic acid is active against S. aureus in broth as well as intracellularly, with no cross-resistance to other antibiotics.

Fusidic acid has a long history of consistent activity against staphylococcal pathogens including methicillin-resistant Staphylococcus aureus (MRSA). Fusidic acid (CEM-102) was susceptibility tested against a surveillance study collection of 12,707 Gram-positive pathogens (2008-2009) from the United States. Reference broth microdilution method results demonstrated the following MIC(50/90) results: S. aureus (.12/.25 μg/mL), coagulase-negative staphylococci (.12/.25 μg/mL), enterococci (4/4 μg/mL), Streptococcus pyogenes (4/8 μg/mL), and viridans group Streptococcus spp. (>8/>8 μg/mL). At a proposed susceptible breakpoint (≤1 μg/mL), fusidic acid inhibited 99.7% of MRSA strains and 99.3% to 99.9% of multidrug-resistant phenotypes of S. aureus. Furthermore, S. aureus strains nonsusceptible to fusidic acid (.35%) generally had detectable resistance mechanisms (fusA, B, C, and E). Reviews of in vitro susceptibility test development confirm the accuracy and intermethod reproducibility of various fusidic acid methods. Fusidic acid is a promising oral therapy for staphylococcal skin and skin structure infections in the United States, where the contemporary S. aureus population remains without significant resistance.

Presented is a case of chronic methicillin-resistant Staphylococcus aureus osteomyelitis, which was unsuccessfully treated with multiple courses of debridement and potent antibiotic therapies. Amputation of the patient’s lower limb was believed to be the only option remaining. A compassionate access program, with approval from the US Food and Drug Administration and the institutional review board, enabled the patient to undergo a course of treatment with oral fusidic acid (CEM-102). The patient tolerated the drug well, with no significant toxicities noted to date. His infection improved rapidly, his flap healed, he has returned to work part-time, and he continues to take daily suppressive doses of oral CEM-102. Reported is a case of chronic methicillin-resistant Staphylococcus aureus (MRSA) femoral osteomyelitis. Despite multiple debridements and failed curative antibiotic courses, the use of suppressive fusidic acid (CEM-102)resulted in the sparing of the lower limb and resolution of acute infective symptoms. Although presently not registered in the United States, fusidic acid offers an attractive alternative for difficult-to-treat cases of MRSA infection as a well-tolerated, oral agent. It has been used widely since the 1960s and is currently available in Europe, Australia, New Zealand, and Israel, where it is available in oral and topical form. Increasing drug resistance, noted especially in staphylococcal isolates, has lead to revitalized interest in its use in countries where it is currently unavailable, such as the United States [1], as well as cautious attempts at restriction in places where it is widely used [2].

Fusidic acid binds to elongation factor G (EF-G), preventing its release from the ribosome, thus stalling bacterial protein synthesis. In staphylococci, high-level fusidic acid resistance is usually caused by mutations in the gene encoding EF-G, fusA, and low-level resistance is generally caused by the horizontally transferable mechanisms fusB and fusC that have a putative protective role on EF-G. In addition, fusD is responsible for intrinsic resistance in Staphylococcus saprophyticus, and alterations in the L6 portion of rplF (fusE) have a role in fusidic acid resistance. Fusidic acid has been used in Europe and Australia for decades. More recently, it has also been used in other countries and regions, but not in the United States. Worldwide fusidic acid resistance has been slow to develop, and the level of resistance and genetic mechanisms responsible generally reflect the time since introduction, indications for treatment, route of administration, and prescribing practices.

A phase 1 trial of fusidic acid (CEM-102), an oral fusidane class antibiotic under development for treatment of gram-positive acute bacterial skin and skin structure infections, evaluating pharmacokinetics and safety is described. A randomized, double-blinded, placebo-controlled, dose escalation study was conducted in healthy adult subjects in the fasting state. Plasma exposure after multiple doses was higher than for single doses, indicating accumulation. Loading doses designed to optimize pharmacodynamic effects were well tolerated and achieved near-steady state concentrations of CEM-102 at 24 h. CEM-102 was safe and generally well tolerated at all single, multiple, and loading doses administered.

Fusidic acid (FA), though used widely throughout the world for decades, has never been approved in the United States. There is now a great need for an oral methicillin-resistant Staphylococcus aureus (MRSA) antibiotic with a long track record of safety. Cempra Pharmaceuticals successfully encouraged passage of a congressional amendment to allow for Hatch-Waxman market exclusivity when this antibiotic is approved in the United States. A new dosing regimen has been patented, allowing FA to be used as monotherapy, and decreased resistance selectivity has been shown. With almost no resistance to FA in the United States, the time is right for introduction into this market.

From around year 2000, impetigo caused by fusidic acid-resistant Staphylococcus aureus was observed in countries of Northern Europe. The bacteria were found to represent a clone, the epidemic European fusidic acid-resistant impetigo clone (EEFIC). This study reports longitudinal data on the incidence and bacteriology of impetigo in a Norwegian island community during the years 2001-09.All encounters with general practitioners regarding impetigo were registered. Bacterial swabs were taken in a high percentage of cases. Annual incidence was calculated. Phenotypic characteristics of the bacteria were determined for the whole period, and in 2008 and 2009 we also performed PFGE and spa typing.Outbreaks of impetigo were observed in 2002, 2003 and 2004, but since then the incidence decreased greatly. S. aureus was cultured from the impetigo site in the majority of cases. The proportion of S. aureus isolates resistant to fusidic acid decreased from 80% in 2002-04 to 45% in 2008-09. For 28 S. aureus isolates analysed by molecular methods in 2008-09, we found that nearly all cases of fusidic acid resistance were due to the presence of the EEFIC.S. aureus resistance to fusidic acid in relation to impetigo is now less frequent in this population than at the start of the century. At present, most S. aureus bacteria resistant to fusidic acid in impetigo belong to the EEFIC.

To describe the emergence of linezolid-resistant methicillin-resistant Staphylococcus aureus (MRSA) of sequence type (ST)36 lineage in two paediatric patients with cystic fibrosis, after long-term low-dose linezolid treatment.Two paediatric males with cystic fibrosis had sputum samples quantitatively cultured during hospitalization. After the isolation of MRSA from both patients, oral treatment with 300 mg linezolid twice daily was initiated for periods of 1-2 months separated by up to 6 months. Isolates cultured 9 months after the start of treatment were tested for resistance to linezolid by agar dilution (BSAC). Resistant isolates were examined for 23S rDNA mutations, and typed by phage and macrorestriction with SmaI. Isolates from follow-up sputum samples were obtained until 44-51 months after treatment with linezolid.Colonization with MRSA was at a density of approximately 10(6) cfu/mL sputum for both subjects. Initial isolates were susceptible to linezolid, but, 9 months later, isolates from both patients were resistant (MICs16 mg/L). Both isolates were epidemic MRSA-16 variant A1 (ST36-MRSA-II), which is widespread in UK hospitals. Both isolates were heterozygous for a G2576T mutation in their 23S rDNA genes, but one was resistant to fusidic acid and tetracycline. In follow-up sampling, the younger patient yielded linezolid-resistant EMRSA-16 for a further 42 months, whilst the other lost the linezolid-resistant MRSA and had alternately Pseudomonas aeruginosa or linezolid-susceptible EMRSA-16 variant A1 isolated over 35 further months.Linezolid resistance emerged in two isolates of ST36 MRSA colonizing the lungs of two paediatric cystic fibrosis patients. Subtherapeutic levels of linezolid may have facilitated the selection of resistance.

Summary Background. High rates of fusidic acid (FA)-resistant Staphylococcus aureus (FRSA) in patients with skin disease have been previously attributed to high usage of topical FA. Aims. To assess whether local community guidelines to restrict topical FA has affected its prescription and use and the level of FRSA in patients with skin disease. Methods. Stapylococcus aureus isolates from microbiology samples received over a 4-month period in 2004 were tested for antibiotic sensitivities. Comparison was then made with the results of a previous study carried out in 2001. Results. A significant fall was seen in the use of topical FA in dermatology patients. In 2001, 62% of patients had used FA-containing preparations within the previous 6 months, compared with just 15% of patients in 2004 (P

A simple, specific, and precise high-performance liquid chromatographic method is developed and validated for the simultaneous determination of chlorocresol (CC), mometasone furoate (MF), and fusidic acid (FA) in a cream formulation. The isocratic mobile phase consists of 1.5% w/v aqueous ammonium acetate buffer-acetonitrile, 55:45 (v/v) of pH 3.8. The column contains octylsilyl chemically bonded to porous silica particle (Symmetry C8, 150x3.9 mm, 5 microm). The detection is carried out using variable wavelength UV-vis detector set at 240 nm. The solutions are chromatographed at a steady flow rate of 1.0 mL/min. The current method separates CC, MF, and FA in less than 8 min with good resolution and peak shapes, minimal tailing, and with retention factors between approximately 1 and 5. Linearity range and percent recoveries for CC, MF, and FA are 10-30, 10-30, and 200-600 microg/mL; and 100.31%, 100.38%, and 100.34%, respectively. The method is validated according to International Conference on Harmonization guidelines and proven to be suitable for stability testing, content uniformity testing, and quality control of these compounds in pharmaceutical preparations.

Methicillin-resistant Staphylococcus aureus (MRSA) and multiresistant Staphylococcus pseudintermedius (MRSP) have emerged as important pathogens in animal infections. Associated therapeutic problems and the zoonotic potential of staphylococci have renewed interest in topical antibiotics for treatment and carrier decolonization. Fusidic acid and mupirocin are used topically in humans and animals but resistant strains isolated from people are increasing. This study investigates the in vitro activity of fusidic acid and mupirocin against coagulase-positive staphylococci from pets.A collection of 287 staphylococci was examined, comprising 102 MRSA, 102 methicillin-susceptible S. aureus, 71 S. pseudintermedius and 12 MRSP from canine and feline infections and carrier sites isolated in the UK and Germany. MICs were determined by the agar dilution method according to CLSI (formerly NCCLS) standards.The majority (89.7%) of all MICs were/=0.25 mg/L. High MICs were observed for seven MRSA isolates (five with an MIC of fusidic acid of 512 mg/L, one with an MIC of fusidic acid of 1024 mg/L and one with with an MIC of mupirocin of 16 mg/L). MICs of both antibiotics were/=2 mg/L for all MRSP. Infection isolates had higher MICs than those isolated from carriage sites for both antibiotics (P/= 0.001).In all but seven MRSA isolates, MICs were below the concentrations achievable experimentally at application sites suggesting therapeutic efficacy of both antibiotics in infections involving multiresistant staphylococci and for decolonization of carriers. However, the seven MRSA with high MICs, all of the dominant UK human hospital lineages, highlight the importance of monitoring treatment success as resistant strains may occur in animals.

The fusidic acid steady-state concentrations in serum (S) and skin suction blister fluid (SBF) after oral doses of 250 and 500 mg administered twice daily for 6 days, as film-coated tablets of sodium fusidate, were studied in eight healthy subjects. The mean peak serum concentrations after the 250- and 500- mg regimens were 39 +/- 5 and 102 +/- 11 mg/l, respectively, obtained between 2 and 3 h after drug intake. The corresponding values for SBF, obtained later, between 2 and 12 h after drug intake, were 21 +/- 5 and 79 +/- 11 mg/l. As measured by the ratio of area under the concentration vs. time curve (SBF/S), fusidic acid penetration is 69-75%, whatever the dose. With either regime, the fusidic acid SBF and serum levels exceed the minimal inhibitory concentration of bacteria usually found in skin infections, especially Staphylococcus species. A dose of 250 mg twice a day appears sufficient to treat these infections and could be tested in clinical studies.

To determine the prevalence and mechanisms of resistance to fusidic acid in clinical isolates of Staphylococcus epidermidis.MICs of fusidic acid were determined for S. epidermidis isolates collected from the Leeds General Infirmary and from around Europe. Fusidic acid-resistant isolates were probed for the presence of the horizontally acquired resistance determinants fusB and fusC by a novel multiplex PCR assay. Mutations in the gene encoding the drug target (fusA) were detected by PCR and DNA sequencing. Resistant isolates were subjected to typing using the repeat region of the aap gene.Of 50 S. epidermidis isolates screened, 23 (46%) exhibited resistance to fusidic acid. The most common resistance determinant was fusB, found in 18 of the 23 isolates. Of the remaining isolates, two harboured fusC and three carried an identical mutation in fusA, leading to the substitution L(461)K in the target protein, elongation factor G. Molecular typing showed that this collection of isolates was genetically diverse.This study suggests a high prevalence of resistance to fusidic acid in clinical isolates of S. epidermidis. As in Staphylococcus aureus, resistance to fusidic acid in S. epidermidis is commonly associated with the fusB determinant.

Background: The morbidity and mortality associated with Panton–Valentine leucocidin (PVL)-positive Staphylococcus aureus suggest that this toxin is a key marker of disease severity. Nevertheless, the importance of PVL in the pathogenesis of primary bacteraemia caused by S. aureus is uncertain. We have determined the prevalence of PVL-encoding genes among isolates of S. aureus from bacteraemic patients. Methods: Consecutive bacteraemia isolates of S. aureus (n 5 244) from patients hospitalized in 25 centres in the UK and Ireland during 2005 were screened for PVL and mecA genes. PVL-positive isolates were characterized by toxin gene profiling, PFGE, spa-typing and MIC determinations for a range of antimicrobials. Results: Four out of 244 isolates (1.6%) were PVL-positive and susceptible to oxacillin [methicillinsusceptible S. aureus (MSSA)]. Eighty-eight out of 244 (36%) were oxacillin-resistant (methicillin-resistant S. aureus), but none was PVL-positive. The four patients (two males: 30 and 33 years; two females: 62 and 80 years) had infection foci of: skin and soft tissue, unknown, indwelling line, and surgical site, and were located at one centre in Wales, one in England and two in Ireland. One of four PVL-positive isolates was resistant to penicillin and fusidic acid, the remainder were susceptible to all antibiotics tested. Genotypic analyses showed that the four isolates represented three distinct strains; the two isolates from Ireland were related. Conclusions: We found that 1.6% of S. aureus (all MSSA) from bacteraemic patients were PVL-positive. This low incidence suggests that PVL-positive S. aureus are of no particular significance as causative agents of S. aureus bacteraemia.

Summary Background Little is known about incidence and natural variation of impetigo in general populations. Objectives To investigate the natural course of impetigo in a well-defined population, and to study the resistance pattern of the causal bacteria over time. Methods This is a population-based incidence study in Austevoll, an island community of 4457 inhabitants in Norway, in the years 2001–2005. Incidence rates are given as events per person-year. Epidemic periods were identified by statistical process-control analyses. Results The incidence rate of impetigo for the whole study period was 0·017 events per person-year, corresponding to a total of 334 cases. The incidence rates were 0·009, 0·026, 0·019, 0·016 and 0·009 in the years 2001, 2002, 2003, 2004 and 2005, respectively. Three epidemics were identified, starting in August of 2002, 2003 and 2004, lasting for 11, 11 and 5 weeks, respectively. Incidence rates in these epidemic periods were 0·099, 0·045 and 0·074, respectively. In epidemic periods, Staphylococcus aureus was the causal bacterium in 89% (117/132) of cases, while this proportion was 68% (84/123) in nonepidemic periods (P

Background: Linezolid is a bacteriostatic oxazolidinone antibiotic that has been proven to be effective for the treatment of patients with pneumonia, skin and soft tissue infections, and possibly bacteraemia, due to Gram-positive cocci. However, the drug is sometimes used for the treatment of patients with endocarditis due to Gram-positive cocci resistant to other antibiotics. Methods: We carried out a review of the available literature to evaluate whether linezolid is also effective for the treatment of patients with infective endocarditis. Results: We identified 23 case reports and 3 case series reporting the experience with 56 patients with endocarditis treated with linezolid. Evaluable data for 33 patients who received linezolid and for whom individual patient data were reported were further analysed. Prosthetic valve infective endocarditis accounted for 25% of the reviewed cases. Methicillin-resistant Staphylococcus aureus and vancomycin-intermediate S. aureus were the most commonly isolated cocci (24.2% and 30.3% of cases, respectively). Linezolid alone was administered to 66.7% of patients while the rest received the antibiotic in combination with rifampicin, gentamicin, fusidic acid or amikacin. A total of 63.6% (21/33) of patients with endocarditis were cured after linezolid administration. The overall and endocarditis-related mortality was 33.3% (11/33) and 12.1% (4/33), respectively. Thrombocytopenia developed in 30.8% (8/26) of patients for whom relevant data were available. Conclusions: The limited available evidence suggests that linezolid may be considered as a therapeutic option for the treatment of patients with endocarditis due to multidrug-resistant Gram-positive cocci. However, further published experience is needed to answer the question of whether a bacteriostatic antibiotic could be proven beneficial for patients with an infection for which bactericidal antibiotics have been traditionally used.

Gram-positive organisms, particularly staphylococci and streptococci, are responsible for the majority of bone and joint infections. Treatment of these infections can be difficult, usually involving a prolonged course of antibiotics, often with surgical intervention. The selection of antibiotics depends on sensitivity profile, patient tolerance and long-term goals, e.g. cure or suppression, but there are few randomized controlled trials in patients comparing efficacy of different antibiotics. Different degrees of bone penetration and clinical outcome for specific antibiotics, e.g. the beta-lactams, clindamycin and quinolones, have been described, although the methodology in these studies is not standardized and findings cannot always be applied directly to patients. The effect of attaining minimum serum bactericidal concentrations in patients has also been studied but this is no longer routinely recommended in clinical practice. Comparative clinical trials are few but have demonstrated efficacy of oral fluoroquinolones in combination with either rifampicin or fusidic acid for selected Gram-positive infections. In the past decade, increasingly resistant organisms, e.g. methicillin-resistant Staphylococcus aureus and vancomycin-resistant enterococci have been recognized as causes of orthopaedic infection. Individual case reports describe successful treatment using the newer antibiotics, e.g. linezolid and quinupristin/dalfopristin, but results of clinical trials are awaited.

Poor adherence with therapy is a major cause of treatment failure in atopic dermatitis. Reasons given are multifactorial, and include fear of real or imaginary side-effects, under-prescribing, failure to renew prescriptions on time, lack of time, and child refusal of therapy. Most important, however, is lack of knowledge about treatment, in particular the use of topical corticosteroid (TCS) therapy. We conducted a questionnaire-based study to determine the level of use and knowledge of commonly prescribed TCS preparations amongst parents or carers of 100 children attending paediatric outpatient clinics. Weakly potent TCSs were the most commonly used (86%), but poorly understood. Only 35 (41%) who had used hydrocortisone were aware that it was weakly potent, and 44% graded it as moderately potent. Of 65 who had used the moderately potent TCS clobetasone butyrate 0.05% (Eumovate); Glaxo Wellcome, Uxbridge, UK), 19 (29%) graded it as potent and eight (12%) as weak. Of 50 who had used betamethasone valerate 0.1% (Betnovate); Glaxo Wellcome, Uxbridge, UK), 42% did not grade it as potent. Understanding of TCS/antimicrobial combinations was generally worse. The hydrocortisone 1%/fusidic acid 2% combination (Fucidin H(R); Leo, Risborough, Bucks, UK) was graded as moderate or strong by 88% of the 74 who had used it. Over half (53%) of the 34 using the combination of clobetasone butyrate 0.05%/nystatin 100000 i.u./g tetracycline 3% (Trimovate); Glaxo Wellcome, Uxbridge, UK) assumed that it was a potent TCS. Forty-nine had used Fucibet (betamethasone valerate 0.1%, fusidic acid 2%; Leo, Risborough, Bucks, UK) but 34.5% did not grade it as potent. There was poor knowledge of the strengths of some of the most commonly used TCSs, and all steroid/antimicrobial combinations were perceived as being of greater potency than the constituent steroid alone. Fusidic acid was thought to be a steroid by almost half (46.9%) of the respondents. The packaging of the different products by some pharmaceutical companies is remarkably similar and labelling contains information on the compound and percentage rather than potency of the TCS. This may be a source of confusion. We recommend that manufacturers clearly label TCS products by potency as mild, moderate, potent or very potent and that packaging is sufficiently different for each strength of TCS or emollient to avoid confusion. In order to achieve optimal topical treatment for atopic dermatitis, patients and their carers must receive adequate information and training in how and when to use topical therapies in conjunction with written care plans.

Summary Background Staphylococcus aureus has a role in the pathophysiology of atopic eczema. Topical fusidic acid is widely used in its treatment. There is concern that topical use of fusidic acid may be driving the selection and dissemination of fusidic acid-resistant (FusR) S. aureus. Objectives To test the hypothesis that treatment of atopic eczema for 2 weeks with topical fusidic acid/steroid combination can increase carriage of FusRS. aureus. Methods Forty-six patients with atopic eczema were allocated randomly to one of two treatment groups. Group 1 (28 patients) were treated with topical 2% fusidic acid plus 0·1% betamethasone cream, and group 2 (18 patients) with topical 2% mupirocin and 0·1% betamethasone cream. The clinical response and nasal and skin colonization with S. aureus were recorded before treatment and after 1 and 2 weeks of therapy. Results Baseline samples from the site of worst eczema showed S. aureus (sensitive and resistant) in 76% of patients, and FusRS. aureus in 26%, with no significant difference between treatment groups. After 1 and 2 weeks, both groups showed similar significant clinical improvement. The overall median clinical improvement was paralleled by a reduction in prevalence and population density of S. aureus (sensitive and resistant) at the worst eczema site (P

SummaryBackground Glycocalyx collapses during dehydration to produce electron-dense accretions. Confocal laser scanning microscopy (CLSM) may be used to visualize fully hydrated microbial biofilms. Objectives Using CLSM, to analyse glycocalyx production by Staphylococcus aureus cells in skin lesions of bullous impetigo, atopic dermatitis and pemphigus foliaceus. A second objective was to compare numbers of S. aureus cells in tissue sections prepared by different methods for routine light microscopy. Methods S. aureus cells in skin lesions of impetigo, atopic dermatitis and pemphigus were stained with safranin, and positive staining with fluorescein isothiocyanate-conjugated concanavalin A was considered to indicate the presence of glycocalyx. Results All S. aureus cells tested in skin lesions of impetigo, atopic dermatitis and pemphigus were covered with glycocalyx and formed microcolonies. The numbers of S. aureus cells in a routine light microscopy section were significantly lower than those in a frozen section that had not been dehydrated with ethanol. Conclusions S. aureus cells generally produce glycocalyx in skin lesions of bullous impetigo, atopic dermatitis and pemphigus foliaceus, which accounts for the difficulty of removing S. aureus cells from these skin lesions. The glycocalyx may collapse during dehydration and most of the S. aureus cells may be carried away during preparation of routine light microscope sections.

Objective: To investigate the possibility that the increased prevalence of fusidic acid-resistant Staphylococcus aureus in Norway is caused by clonal spread. Methods: Fusidic acid-resistant and -susceptible clinical isolates of S. aureusfrom patients with skin infections in the Norwegian county of Telemark and fusidic acid-resistant isolates from other parts of Scandinavia were compared. MICs of fusidic acid for bacterial isolates and pulsed-field gel electrophoresis (PFGE) patterns were investigated. Prevalence data for fusidic acid-resistant S. aureus for the period 1992-2001 were obtained. Results: The prevalence of fusidic acid resistance in S. aureus increased from 1992 to 2001. Eighty per cent of the resistant isolates investigated shared an identical PFGE pattern. The same pattern was found in fusidic acid-resistant isolates from other parts of Scandinavia. Fusidic acid-resistant S. aureus was typically found in impetigo bullosa-like skin disease in children mostly in the summer months. Conclusions: Fusidic acid resistance among S. aureus is increasing in Norway and is predominantly caused by one clone of S. aureus. The clone may spread further to other countries, and dissemination may be facilitated by extensive use of topical fusidic acid.

SummaryBackground Bacteria that adhere to damaged tissues encase themselves in a hydrated matrix of polysaccharides, forming a slimy layer known as a biofilm. This is the first report of detection of glycocalyx production by Staphylococcus aureus using confocal laser scanning microscopy (CLSM) on damaged skin tissues. Objectives To analyse glycocalyx production by S. aureus cells on damaged skin tissues and the influence of polymorphonuclear leucocytes (PMNs) and various antimicrobial agents on its production using CLSM in cyclophosphamide (Cy)-treated (neutropenic) or non-Cy-treated (normal) mice. Methods S. aureus cells were inoculated on damaged skin tissues in neutropenic or normal mice with or without topical application of antimicrobial agents. S. aureus cells were stained with safranine, and positive staining with fluorescein isothiocyanate-conjugated concanavalin A was considered to indicate the presence of glycocalyx. Results All S. aureus cells tested on damaged skin tissues formed microcolonies encircled by glycocalyx. The colony counts of S. aureus cells on croton oil dermatitis in normal mice treated with 2% fusidic acid ointment were about 100 times lower than those in neutropenic mice (control). Conclusions As S. aureus cells can generally produce a biofilm on damaged skin tissues, antimicrobial agents may not eradicate S. aureus cells without the help of PMNs. S. aureus glycocalyx may play a crucial role in colonization and adherence to damaged skin tissues.

We report the cloning of the fusidic acid and cadmium resistance determinants from Staphylococcus aureus plasmid pUB101. The pUB101 fusidic acid resistance determinant was located on a 2.9 kb HindIII fragment. Sequencing of this fragment revealed three putative open reading frames (ORFs) of 213 (far1), 152 (orf152) and 170 amino acids (orf170), which are flanked by the right-hand end of insertion sequence IS431/257 (IS431/257RH) and a partial ORF. Far1 and Orf152 demonstrated homology with a chromosomally encoded fibronectin-binding protein of Listeria monocytogenes and the putative protein YosT, found on the SP beta c2 prophage of Bacillus subtilis, respectively. Transformation of S. aureus with a construct containing a 949 bp far1-specific amplicon led to the isolation of a fusidic acid-resistant transformant, thereby identifying the pUB101 fusidic acid resistance structural gene. Between orf152 and far1 we identified a unique 113 bp symmetrical element and other repeat elements that may be involved with the control of orf152 and/or far1 expression. Hybridization of Southern blots revealed that far1 was not located on the chromosome or plasmid content of a limited number of Australian, UK and Hong Kong fusidic acid-resistant isolates. The pUB101 cadmium resistance determinant was located on a 3.6 kb HindIII fragment that carried a cadDX operon, remnants of two putative plasmid replication protein genes and IS431/257RH. Sequence analysis also demonstrated the presence of a single-stranded origin of replication, normally found on rolling circle replicating plasmids, within the putative promoter region of the cadDX operon.

SummaryBackground Neomycin, clioquinol and fusidic acid are all topical antibiotics widely used in dermatological practice in the U.K., either as a single agent or in combination with topical corticosteroids. However, an adverse effect of topical antibiotics is contact sensitization. Objectives To examine the frequency of positive patch test reactions to fusidic acid, clioquinol and neomycin. Methods To compare the frequency of allergic patch test reactions over 1 year, we patch tested all patients attending the St John's Institute of Dermatology contact dermatitis clinic for one calendar year with fusidic acid, neomycin and clioquinol. Results We patch tested 1119 patients. Postitive patch test reactions to neomycin were noted in 40 patients (3·6%), compared with eight patients (0·7%) to clioquinol and three patients (0·3%) to fusidic acid. The frequency of medicament allergy to neomycin was thus five times more common than to clioquinol and ten times more common than to fusidic acid. Although fusidic acid is not part of our extended standard series, it is in our medicaments series. Therefore, in the second part of our study, we reviewed all cases of positive patch test reactions to fusidic acid over the last 20 years. We found that the frequency of hypersensitivity has decreased since the early 1980s despite increasing usage; the current average frequency being 1·62 patch-tested patients per year (1·45%) of those patch tested to the medicaments series). The most common diagnosis in such patients was stasis dermatitis (54·2%). Conclusions The frequency of fusidic acid allergy in an eczema population is low and is comparable with published data from over 10 years ago.

A retrospective study was carried out to investigate possible reasons for a marked increase in fusidic acid-resistant Staphylococcus aureus (FusR S. aureus) identified by our routine hospital microbiology service. Information was obtained on a sample of 64 consecutive patients from whom resistant S. aureus had been cultured. The source of isolates was found to be diffuse within the hospital and community, The site of sample was most frequently chronic cutaneous infections (68%), All the S. aureus isolates were resistant to both fusidic acid and penicillin and many were resistant to multiple antibiotics. Topical fusidic acid had been used by 40% of patients in the preceding 6 months and none had received oral fusidic acid (sodium fusidate). Most (80%) had received an oral antibiotic in the preceding 2 years. Information from the Prescriptions Pricing Authority revealed that the total number of prescriptions for fusidic acid-containing preparations for the period September 1997 to August 1998 was markedly higher in Harrogate than in five other local areas where increases in (FusR) S. aureus have not been observed.

Screening facultative sheep-rumen bacteria which inhibit growth of Escherichia coli produced 11 strains of Pseudomonas aeruginosa. The isolates showed three different pulsed-field gel electrophoresis patterns and strains from different sheep produced pyocins that varied in strain specificity. Representative strains were resistant to ampicillin, methicillin, erythromycin, fusidic acid and augmentin, but not to tetracycline or nalidixic acid. Tested strains attached in large numbers to cultured rumen epithelial cells, potentially providing a means of survival in this ecosystem.

Salicylate, acetyl-salicylate, benzoate and ibuprofen increased fusidic acid MICs for fusidic acid-resistant and -susceptible strains of Staphylococcus aureus representing six genetic lineages. The effects of these substances on fusidic acid resistance levels occurred in a strain-dependent manner. The weak acid acetate, and acetaminophen did not alter fusidic acid resistance levels, while the addition of saligenin, the alcohol of salicylate, reduced gradient plate MICs for all strains studied. These findings indicate that a benzoic acid structure is required for the induction of increased intrinsic fusidic acid resistance levels. When 2 mM salicylate was added to media used in population analyses, the number of cells able to survive on high concentrations of fusidic acid increased. This increase in cell survival was observed in two unrelated fusidic acid-resistant strains, with chromosomal (WBG8287) or plasmid (WBG1576) mediated resistance determinants and two unrelated susceptible strains. The salicylate-induced increase in fusidic acid resistance was phenotypic at low fusidic acid concentrations (relative to resistance phenotype) for WBG8287 and a fusidic acid-susceptible strain. On media containing salicylate and high fusidic acid concentrations, the mutation frequency to higher fusidic acid resistance levels was greater for WBG8287, compared with unsupplemented fusidic acid-containing media. These experiments provide evidence for a novel salicylate inducible fusidic acid resistance mechanism in S. aureus.

Received 27 May 2008; returned 1 June 2008; revised 4 June 2008; accepted 5 June 2008Objectives: We determined the in vitro activity of retapamulin, a novel pleuromutilin antibiotic, against664 Staphylococcus aureus isolates from the UK, including many resistant to fusidic acid and/orhighly resistant to mupirocin.Methods: MICs were determined on Mueller–Hinton agar in accordance with the CLSI guidelines.Susceptibility was categorized using CLSI criteria, where available; otherwise the European Committee forAntimicrobial Susceptibility Testing (EUCAST)/BSAC criteria were used (for mupirocin and fusidic acid).Mutationsinthe rplCgene,whichencodesribosomalproteinL3,weresoughtbyPCRandDNAsequencing.Results:TheS. aureus included 488 (73%) methicillin-resistant isolates (oxacillin MICs >2 mg/L), 336 iso-lates (51%) resistant to fusidic acid (MICs >1 mg/L) and 254 (38%) with high-level mupirocin resistance(MICs >256 mg/L); 103 (16%) isolates were resistant both to fusidic acid and to high levels of mupirocin.Retapamulininhibited663(99.9%)isolatesat 0.25 mg/L.Asinglemethicillin-resistant S.aureus isolate,alsowith high-level mupirocin resistance, required a retapamulin MIC of 2 mg/L, but its reduced susceptibility toretapamulinwasnotassociatedwithanymutationinribosomalproteinL3.Conclusions : Retapamulin demonstrated excellent activity in vitro against S. aureus isolates, irrespective oftheir level of resistance to other antibacterials. These results support the EUCAST epidemiological cut-offvalueforretapamulinof 0.5 mg/Lagainst S.aureus.Keywords: S. aureus, MRSA, mechanisms

BackgroundNew therapies designed to downregulate the aberrant immune response associated with severe acute necrotizing pancreatitis (ANP) are being increasingly investigated in different experimental models of ANP. The aim of this study was to test the potential effects of sodium fusidate on the course of severe ANP in rabbits.MethodsANP was induced in 20 rabbits by retrograde injection of 5 per cent chenodeoxycholic acid into the pancreatic duct followed by duct ligation. The rabbits were allocated to pretreatment with intravenous physiological saline or sodium fusidate 80 mg/kg 30 min before the induction of ANP. Levels of serum amylase, lipase, tumour necrosis factor (TNF) α, interleukin (IL) 8, glucose and calcium, and leucocyte count were measured every 3 h for a total of 12 h. At the end of the experiment, ascitic fluid was collected and the pancreatic, lung and kidney tissues were obtained for histological examination.ResultsPretreatment with sodium fusidate reduced the mortality rate from six of ten to three of ten (P < 0·05) and reduced the output of ascitic fluid from 5·2 to 2·0 ml/h (P ConclusionPretreatment with sodium fusidate resulted in a considerable reduction in mortality rate and ascitic fluid output in rabbits with bile-induced ANP, probably by lowering the TNF-α and IL-8 blood levels. However, pretreatment with sodium fusidate did not alter the local or systemic manifestations of ANP. Thus, cytokines other than TNF-α and IL-8 are likely to mediate the local and systemic symptoms of ANP.

Iclaprim is a novel diaminopyrimidine for which a human plasma binding level of approximately 93% has been reported. The purpose of this study was to evaluate the effect of human plasma on the in vitro activity of iclaprim and to compare it with that of fusidic acid, teicoplanin and vancomycin, antibiotics with protein binding to human plasma of 97%,90% and 55%, respectively.MICs were determined using 40 methicillin-susceptible Staphylococcus aureus (MSSA) and 38 methicillin-resistant S. aureus (MRSA) isolates in Mueller-Hinton broth (MHB) alone or in the presence of 50% human plasma.MICs of iclaprim were not affected by the addition of human plasma. MIC ranges (MIC(90)) for iclaprim against MSSA and MRSA wereor =0.016-0.06 mg/L (MIC(90) 0.06 mg/L) andor =0.016-0.5 mg/L (MIC(90) 0.06 mg/L), respectively, in MHB andor =0.016-0.125 mg/L (MIC(90) 0.06 mg/L) andor =0.016-0.25 mg/L (MIC(90) 0.125 mg/L), respectively, in the presence of human plasma. As expected, the antimicrobial activity of fusidic acid was greatly affected by the presence of human plasma (MIC elevations of 4- to128-fold), whereas MICs of vancomycin remained unchanged. By contrast, despite the high protein binding, MICs of teicoplanin were only marginally affected by the presence of plasma with an MIC elevation of maximum 8-fold for two strains.This study demonstrates that human plasma does not affect the MIC of iclaprim in vitro.

The effect of dicloxacillin and fusidic acid used alone and in combination on the extracellular and intracellular killing of four isolates of Staphylococcus aureus in the presence of serum was studied. At the extracellular level, dicloxacillin (8 mg/L) had a bactericidal effect on all four isolates, whereas fusidic acid (64 mg/L) had a bacteriostatic effect on two isolates and no effect on the two other isolates. Fusidic acid significantly inhibited the extracellular bactericidal effect of dicloxacillin on two isolates. Intracellular killing was measured in human neutrophil granulocytes. Dicloxacillin (8 mg/L) significantly increased the intracellular killing of all four isolates, while fusidic acid (64 mg/L) significantly increased the intracellular killing of three isolates, but the killing was significantly lower than that of dicloxacillin. When the antibiotics were combined the intracellular killing of three of the isolates was significantly lower than that of dicloxacillin alone. The viability of the granulocytes and their ability to produce superoxide anion were not affected by the antibiotics. In conclusion, we found that the increased intracellular killing of S. aureus by dicloxacillin was inhibited by fusidic acid.

Methicillin-resistant Staphylococcus aureus (MRSA) is endemic in teaching hospitals in eastern Australian states, with prevalence rates averaging 25-30% of all S. aureus. Between 1990 and 1995, 1467 non-duplicate MRSA isolates from clinically infected sites were tested in Sydney, Melbourne, and Brisbane as part of a national survey of staphylococcal susceptibility. We reviewed the differing evolution of resistance to ciprofloxacin, rifampicin, and fusidic acid. Despite similarities in community and hospital antibiotic use and MRSA prevalence rates, trends in resistance to the oral antibiotics in these cities have progressed independently of each other. In the 1995 survey in individual hospitals in Melbourne, 16-24% of strains were ciprofloxacin-resistant, compared with 80-100% in Sydney and 30-44% in Brisbane. There was great diversity of phage type patterns for ciprofloxacin-resistant strains, suggesting heterogeneous development of resistance. Rifampicin resistance was more closely associated with distinct dominant epidemic phage types, common to institutions in the same city, but without spread to the other cites. Between 1990 and 1995, these comprised 30-60% of all MRSA in Brisbane, compared with 5-10% in Melbourne and < 25% in Sydney. Fusidic acid resistance was uncommon and sporadic (< 5%), and was distributed equally between methicillin-resistant and methicillin-susceptible strains. Resistance to the oral agents in MRSA is due to a complex mix of antibiotic selection pressures and cross-infection with local and epidemic strains in closely related institutions. Each of these mechanisms can predominate, dependent on local factors and the antibiotics used.