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Aims and objectives. To compare the nursing time and cost required for preparation and administration of liposomal amphotericin B, amphotericin B deoxycholate and voriconazole. Design. Cost comparison study. Methods. Nurse activities associated with the preparation and administration of the three study drugs were divided into 11 tasks and timed by observers at five hospitals. Target tasks were defined as those likely to be affected by the differences between drugs and excluded those tasks likely to differ owing to site-specific factors. Mean times for administration of a single day of therapy for each study drug were compared. Costs of preparation and administration of a 14-day regimen were estimated. Results. Sixty-nine patients were observed receiving a total of 256 doses of study medications. Labour times were 20, 16, 14 and 3 minutes per day for liposomal amphotericin B, amphotericin B deoxycholate, intravenous voriconazole and oral voriconazole, respectively. Administration time was significantly lower for intravenous voriconazole compared with liposomal amphotericin B (p < 0·05), and for oral voriconazole compared with all intravenous regimens (p < 0·05). Preparation of medications took the longest time for intravenous formulations and was longer for liposomal amphotericin B than for the other drugs by 3-5 minutes. Average non-drug costs associated with preparation and administration of a 14-day regimen were greatest in the amphotericin B deoxycholate arm at US$ 335, followed by liposomal amphotericin B (US$ 310) and voriconazole (US$ 180). Conclusion. Intravenous voriconazole required less time to prepare and administer on a daily basis than liposomal amphotericin B, and was similar to amphotericin B deoxycholate. Measurements of intravenous vs. oral voriconazole administration suggest the opportunity to save 10-17 minutes per day with the oral formulation. Relevance to clinical practice. Oral voriconazole may provide significant savings in terms of nursing time compared with intravenous antifungal drugs. [ABSTRACT FROM AUTHOR]

Among long-stay critically ill patients in the adult intensive care unit (ICU), there are often marked changes in the complexity of the gut microbiota. However, it remains unclear whether such patients might benefit from enhanced surveillance or from interventions targeting the gut microbiota or the pathogens therein. We therefore undertook a prospective observational study of 24 ICU patients, in which serial faecal samples were subjected to shotgun metagenomic sequencing, phylogenetic profiling and microbial genome analyses. Two-thirds of the patients experienced a marked drop in gut microbial diversity (to an inverse Simpson's index of <4) at some stage during their stay in the ICU, often accompanied by the absence or loss of potentially beneficial bacteria. Intravenous administration of the broad-spectrum antimicrobial agent meropenem was significantly associated with loss of gut microbial diversity, but the administration of other antibiotics, including piperacillin/tazobactam, failed to trigger statistically detectable changes in microbial diversity. In three-quarters of ICU patients, we documented episodes of gut domination by pathogenic strains, with evidence of cryptic nosocomial transmission of Enterococcus faecium . In some patients, we also saw an increase in the relative abundance of apparent commensal organisms in the gut microbiome, including the archaeal species Methanobrevibacter smithii . In conclusion, we have documented a dramatic absence of microbial diversity and pathogen domination of the gut microbiota in a high proportion of critically ill patients using shotgun metagenomics.

Background: Studies have reported large scale overprescribing of antibiotics for urinary tract infection (UTI) in hospitalised older adults. Older adults often have asymptomatic bacteriuria, and clinicians have been found to diagnose UTIs inappropriately based on vague symptoms and positive urinalysis and microbiology. However, the joined perspectives of different staff groups and older adult patients on UTI diagnosis have not been investigated., Methods: Thematic analysis of qualitative interviews with healthcare staff ( n  = 27) and older adult patients ( n  = 14) in two UK hospitals., Results: Interviews featured a recurrent theme of discrepant understandings and gaps in communication or translation between different social groups in three key forms: First, between clinicians and older adult patients about symptom recognition. Second, between nurses and doctors about the use and reliability of point-of-care urinary dipsticks. Third, between nurses, patients, microbiologists and doctors about collection of urine specimens, contamination of the specimens and interpretation of mixed growth laboratory results. The three gaps in communication could all foster inappropriate diagnosis and antibiotic prescribing., Conclusion: Interventions to improve diagnosis and prescribing for UTIs in older adults typically focus on educating clinicians. Drawing on the sociological concept of translation and interviews with staff and patients our findings suggest that inappropriate diagnosis and antibiotic prescribing in hospitals can be fuelled by gaps in communication or translation between different staff groups and older adult patients, using different languages and technologies or interpreting them differently. We suggest that interventions in this area may be improved by also addressing discrepant understandings and communication about symptoms, urinary dipsticks and the process of urinalysis., Competing Interests: Competing interestsThe authors declare that they have no competing interests.

Background: Estimates of the prevalence of asymptomatically carried Clostridium difficile in elderly patients in long-term care range from 0% to 51%. Asymptomatic carriage is possibly a risk factor for the development of infection, and there is ongoing debate surrounding the role of asymptomatic carriage in transmission., Aim: To investigate the prevalence of asymptomatic carriage amongst patients residing in intermediate care (bedded) facilities (ICBFs), and to investigate whether asymptomatically carried C. difficile strains contribute to nosocomial C. difficile infection (CDI)., Methods: Stools were collected from eligible asymptomatic patients in ICBFs, and a subset was also processed from symptomatic patients accessing primary or secondary care outside of ICBFs. All samples were cultured for C. difficile, and resulting colonies were processed through whole genome sequencing., Findings: In total, 151 asymptomatic patients were sampled, 22 of which were positive for C. difficile through stool culture, representing a carriage rate of 14.6%. Sequencing of these isolates, alongside 14 C. difficile polymerase chain reaction and culture-positive isolates from symptomatic individuals, revealed that all asymptomatic patients were carrying toxigenic C. difficile, and these strains were genetically similar to those from symptomatic patients., Conclusion: This small study of asymptomatic carriage revealed a rectal asymptomatic carriage rate of 14.6% in patients nursed in ICBFs, and a high level of genetic similarity of these strains to those recovered from symptomatic patients. As such, asymptomatic carriers may be important for the transmission of symptomatic CDI, although it is acknowledged that this study was small, and many other factors govern whether C. difficile is carried asymptomatically or causes symptoms., (Copyright © 2018 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.)

Background: Carbapenemase-producing Enterobacteriaceae (CPE) pose a considerable threat to modern medicine. New treatment options and methods to limit spread need to be investigated. Blue light (BL) is intrinsically antimicrobial, and we have previously demonstrated significant antimicrobial effects on biofilms of a panel of isolates, including two CPEs.This study was performed to assess the antibacterial activity of 405 nm BL against a panel of CPE isolates (four encoding bla NDM , three bla KPC , two bla OXA-48 , and three encoding both NDM and OXA-48 carbapenemases)., Methods: In vitro experiments were conducted on 72 h old biofilms of CPEs which were exposed to 60 mW/cm 2 of BL. Changes to biofilm seeding were assessed by measuring the optical density of treated and untreated biofilms., Results: Twelve bacterial clinical isolates (comprising eight Klebsiella pnemoniae , one K. oxytoca , and three Escherichia coli ) were tested. BL was delivered for 5, 15 and 30 min, achieving doses of 162, 54, and 108 J/cm 2 , respectively.All of the CPEs were susceptible to BL treatment, with increasing reductions in seeding with increasing durations of exposure. At 30 min, reductions in biofilm seeding of ≥80% were observed for 11 of the 12 isolates, compared to five of 12 after 15 min. CPE&#95;8180 was less susceptible than the rest, with a maximum reduction in seeding of 66% at 30 min., Conclusions: BL is effective at reducing the seeding of mature CPE biofilms in vitro, and offers great promise as a topical decontamination/treatment agent for both clinical and environmental applications., Competing Interests: Not applicable.Not applicable.The authors declare that they have no competing interests.Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Background: The early diagnosis of infection or sepsis in burns are important for patient care. Globally, a large number of burn centres advocate quantitative cultures of wound biopsies for patient management, since there is assumed to be a direct link between the bioburden of a burn wound and the risk of microbial invasion. Given the conflicting study findings in this area, a systematic review was warranted., Methods: Bibliographic databases were searched with no language restrictions to August 2015. Study selection, data extraction and risk of bias assessment were performed in duplicate using pre-defined criteria. Substantial heterogeneity precluded quantitative synthesis, and findings were described narratively, sub-grouped by clinical question., Results: Twenty six laboratory and/or clinical studies were included. Substantial heterogeneity hampered comparisons across studies and interpretation of findings. Limited evidence suggests that (i) more than one quantitative microbiology sample is required to obtain reliable estimates of bacterial load; (ii) biopsies are more sensitive than swabs in diagnosing or predicting sepsis; (iii) high bacterial loads may predict worse clinical outcomes, and (iv) both quantitative and semi-quantitative culture reports need to be interpreted with caution and in the context of other clinical risk factors., Conclusion: The evidence base for the utility and reliability of quantitative microbiology for diagnosing or predicting clinical outcomes in burns patients is limited and often poorly reported. Consequently future research is warranted., (Crown Copyright © 2017. Published by Elsevier Ltd. All rights reserved.)

Objective: We previously reported on the ability of SurgihoneyRO (SHRO), an engineered honey, to prevent biofilm formation in vitro, but data were lacking regarding the activity against preformed biofilms. This study aims to assess whether SHRO has any antibacterial activity against mature, preformed biofilms and whether there is any evidence to support the observed clinical effectiveness when SHRO has been used anecdotally on acute and chronic wounds where biofilm is most likely present., Method: We tested the in vitro antibacterial activity of SHRO against the mature biofilms of 16 clinically relevant wound pathogens, in terms of impacts on biofilm seeding and biofilm biomass. The honey was serially double diluted from 1:3 down to 1:6144, and the lowest dilution achieving a statistically significant reduction in biomass of ≥50%, compared with untreated controls, was recorded., Results: All 16 bacterial isolates were susceptible to SHRO, with reduced biofilm seeding observed for all, and percentage reductions ranging from 58% (ACI&#95;C59) to 94.3% (MDR&#95;B) for the strongest concentration of honey (1:3). Furthermore at this concentration, biofilm seeding of the test biofilm was reduced by 80-94.3% (when compared with the positive control) for 12/16 isolates. We additionally demonstrated that SHRO has antibiofilm impacts, with the 24 hour exposure resulting in disruption of the biofilm, reduced seeding and reduced biomass., Conclusion: SHRO is effective at reducing seeding of preformed biofilms of clinically important wound pathogens in vitro, and also has antibiofilm activity. This supports the anecdotal clinical data for antibiofilm efficacy, and supports the use of SHRO as a promising topical wound care agent.

Unlabelled: The blue wavelengths within the visible light spectrum are intrinisically antimicrobial and can photodynamically inactivate the cells of a wide spectrum of bacteria (Gram positive and negative) and fungi. Furthermore, blue light is equally effective against both drug-sensitive and -resistant members of target species and is less detrimental to mammalian cells than is UV radiation. Blue light is currently used for treating acnes vulgaris and Helicobacter pylori infections; the utility for decontamination and treatment of wound infections is in its infancy. Furthermore, limited studies have been performed on bacterial biofilms, the key growth mode of bacteria involved in clinical infections. Here we report the findings of a multicenter in vitro study performed to assess the antimicrobial activity of 400-nm blue light against bacteria in both planktonic and biofilm growth modes. Blue light was tested against a panel of 34 bacterial isolates (clinical and type strains) comprising Acinetobacter baumannii, Enterobacter cloacae, Stenotrophomonas maltophilia, Pseudomonas aeruginosa, Escherichia coli, Staphylococcus aureus, Enterococcus faecium, Klebsiella pneumoniae, and Elizabethkingia meningoseptica All planktonic-phase bacteria were susceptible to blue light treatment, with the majority (71%) demonstrating a ≥5-log10 decrease in viability after 15 to 30 min of exposure (54 J/cm(2) to 108 J/cm(2)). Bacterial biofilms were also highly susceptible to blue light, with significant reduction in seeding observed for all isolates at all levels of exposure. These results warrant further investigation of blue light as a novel decontamination strategy for the nosocomial environment, as well as additional wider decontamination applications., Importance: Blue light shows great promise as a novel decontamination strategy for the nosocomial environment, as well as additional wider decontamination applications (e.g., wound closure during surgery). This warrants further investigation., (© Crown copyright 2016.)

Enterococcal infections are common in liver transplantation and hepatopancreaticobiliary (HPB) surgery. Linezolid is frequently used to treat not only vancomycin-resistant Enterococcus (VRE), but also vancomycin-sensitive Enterococcus (VSE) infections, and resistance can develop. This study evaluated all the Liver Unit patients who developed infections with linezolid-resistant Enterococcus (LRE) in order to elicit the association with prior linezolid usage, to explore possible risk factors for infection, and to better understand the epidemiology of these isolates in this patient group. Between 2010 and 2015, infections with LRE developed in 10 patients (8 following liver transplantation and 2 following HPB surgery) after 22-108 days of treatment. Selected pulsed-field gel electrophoresis demonstrated that 2 out of 10 patients were cocolonized with different strains and indicated that cross-transmission may have occurred. In conclusion, in this group of patients with complex hepatobiliary infections, the optimal antibiotic strategies for the treatment of Enterococcus faecium infections are not clearly defined, and there is a significant risk of emergence of resistance to linezolid in E. faecium after exposure to this agent in patients, especially in the presence of a deep source of infection on a background of hepatic artery insufficiency. Caution is needed when using prolonged courses of linezolid in this setting, and further studies are necessary to determine the optimum treatment., (© 2015 American Association for the Study of Liver Diseases.)

Objective: Honey is recognised to be a good topical wound care agent owing to a broad-spectrum of antimicrobial activity combined with healing properties. Surgihoney RO (SH1) is a product based on honey that is engineered to produce enhanced reactive oxygen species (ROS) and has been reported to be highly antimicrobial. The objective was to investigate the ability of the engineered honey and its comparators to prevent biofilm formation in vitro., Method: We tested the ability of three medical-grade honeys SH1, Activon manuka honey (MH) and Medihoney manuka honey (Med), alongside five antimicrobial dressings (AMDs) to prevent the formation of biofilms by 16 isolates. Honeys were serially double diluted from 1:3 down to 1:6144 and the lowest dilution achieving a statistically significant reduction in biomass of at least 50%, compared with untreated controls, was recorded., Results: Although all the honeys were antibacterial and were able to prevent the formation of biofilms, SH1 was the most potent, with efficacy at lower dilutions than the medical honeys for five isolates, and equivalent dilutions for a further six. Additionally, SH1 was superior in antibacterial potency to three commercially available AMDs that contain honey., Conclusion: SH1 is effective at preventing bioflms from forming and is superior to medical honeys and AMDs in in vitro tests., Declaration of Interest: Surgihoney RO was provided free of charge for testing by Matoke Holdings, UK and the hospital pharmacy provided the other honeys and dressings. This paper presents independent research funded by the National Institute for Health Research (NIHR). The views expressed are those of the authors and not necessarily those of the NHS, the NIHR or the Department of Health.

Unlabelled: Antimicrobial medicated dressings (AMD) are often used to reduce bacterial infection of burns and other wounds. However, there is limited literature regarding comparative efficacies to inform effective clinical decision making., Objectives: Following on from a previous study where we demonstrated good antibiofilm properties of acetic acid (AA), we assessed and compared the in vitro anti-biofilm activity of a range of AMDs and non-AMDs to AA., Methods: Laboratory experiments determined the ability of a range of eleven commercial AMD, two nAMD, and AA, to prevent the formation of biofilms of a panel of four isolates of Pseudomonas aeruginosa and Acinetobacter baumannii., Results: There is a large variation in ability of different dressings to inhibit biofilm formation, seen between dressings that contain the same, and those that contain other antimicrobial agents. The best performing AMD were Mepilex(®) Ag and Acticoat. AA consistently prevented biofilm formation., Conclusions: Large variation exists in the ability of AMD to prevent biofilm formation and colonisation of wounds. A standardised in vitro methodology should be developed for external parties to examine and compare the efficacies of commercially available AMDs, along with robust clinical randomised controlled trials. This is essential for informed clinical decision-making and optimal patient management., (Copyright © 2015 Elsevier Ltd and ISBI. All rights reserved.)

Background: Sepsis from burn injuries can result from colonisation of burn wounds, especially in large surface area burns. Reducing bacterial infection will reduce morbidity and mortality, and mortality for severe burns can be as high as 15 %. There are various quantitative and semi-quantitative techniques to monitor bacterial load on wounds. In the UK, burn wounds are typically monitored for the presence or absence of bacteria through the collection and culture of swabs, but no absolute count is obtained. Quantitative burn wound culture provides a measure of bacterial count and is gaining increased popularity in some countries. It is however more resource intensive, and evidence for its utility appears to be inconsistent. This systematic review therefore aims to assess the evidence on the utility and reliability of different quantitative microbiology techniques in terms of diagnosing or predicting clinical outcomes., Methods/design: Standard systematic review methods aimed at minimising bias will be employed for study identification, selection and data extraction. Bibliographic databases and ongoing trial registers will be searched and conference abstracts screened. Studies will be eligible if they are prospective studies or systematic reviews of burn patients (any age) for whom quantitative microbiology has been performed, whether it is compared to another method. Quality assessment will be based on quality assessment tools for diagnostic and prognostic studies and tailored to the review as necessary. Synthesis is likely to be primarily narrative, but meta-analysis may be considered where clinical and methodological homogeneity exists., Discussion: Given the increasing use of quantitative methods, this is a timely systematic review, which will attempt to clarify the evidence base. As far as the authors are aware, it will be the first to address this topic., Trial Registration: PROSPERO, CRD42015023903.

Introduction: Localised infections, and burn wound sepsis are key concerns in the treatment of burns patients, and prevention of colonisation largely relies on biocides. Acetic acid has been shown to have good antibacterial activity against various planktonic organisms, however data is limited on efficacy, and few studies have been performed on biofilms., Objectives: We sought to investigate the antibacterial activity of acetic acid against important burn wound colonising organisms growing planktonically and as biofilms., Methods: Laboratory experiments were performed to test the ability of acetic acid to inhibit growth of pathogens, inhibit the formation of biofilms, and eradicate pre-formed biofilms., Results: Twenty-nine isolates of common wound-infecting pathogens were tested. Acetic acid was antibacterial against planktonic growth, with an minimum inhibitory concentration of 0.16-0.31% for all isolates, and was also able to prevent formation of biofilms (at 0.31%). Eradication of mature biofilms was observed for all isolates after three hours of exposure., Conclusions: This study provides evidence that acetic acid can inhibit growth of key burn wound pathogens when used at very dilute concentrations. Owing to current concerns of the reducing efficacy of systemic antibiotics, this novel biocide application offers great promise as a cheap and effective measure to treat infections in burns patients.

As part of the tigecycline evaluation and surveillance trial (TEST), bacterial isolates were collected from 39 centres in France, Germany, Italy, Spain and the UK between January 2004 and August 2006. Antimicrobial susceptibilities were determined according to CLSI guidelines. Italy had the highest rate of methicillin-resistant Staphylococcus aureus (36.4%), and was the only country to report vancomycin-resistant Enterococcus faecalis (8.6%). Tigecycline was the only agent to which all Gram-positive isolates were susceptible. For many of the Gram-negative organisms collected, antimicrobial susceptibilities were lowest among isolates from Italy and highest among isolates from Spain. The notable exception was Acinetobacter baumannii, where the poorest susceptibility profile was among isolates from Spain. For A. baumannii, MIC(90)s of imipenem varied from 1 mg/L for isolates in France and Germany to > or =32 mg/L for isolates from Italy and Spain. Tigecycline was the only agent to maintain an MIC(90) of < or =1 mg/L against isolates from all five countries. The in-vitro activity of tigecycline against both Gram-positive and Gram-negative isolates may make it valuable in the treatment of hospital infections, including those caused by otherwise antimicrobial-resistant organisms.

Meticillin-resistant Staphylococcus aureus (MRSA) persists in the hospital environment and conventional cleaning procedures do not necessarily eliminate contamination. A prospective study was conducted on an intensive care unit to establish the level of environmental contamination with MRSA, assess the effectiveness of hydrogen peroxide vapour (HPV) decontamination and determine the rate of environmental recontamination. MRSA was isolated from 11.2% of environmental sites in the three months preceding the use of HPV and epidemiological typing revealed that the types circulating within the environment were similar to those colonising patients. After patient discharge and terminal cleaning using conventional methods, MRSA was isolated from five sites (17.2%). After HPV decontamination but before the readmission of patients, MRSA was not isolated from the environment. Twenty-four hours after readmitting patients, including two colonized with MRSA, the organism was isolated from five sites. The strains were indistinguishable from a strain with which a patient was colonized but were not all confined to the immediate vicinity of the colonized patient. In the eight weeks after the use of HPV, the environment was sampled on a weekly basis and MRSA was isolated from 16.3% sites. Hydrogen peroxide vapour is effective in eliminating bacteria from the environment but the rapid rate of recontamination suggests that it is not an effective means of maintaining low levels of environmental contamination in an open-plan intensive care unit.

Objective: The study aimed to examine the presence of methicillin-resistant Staphylococcus aureus (MRSA) in the environment and its relationship to patients' acquisition of MRSA., Design: A prospective study was conducted in a 9-bed intensive care unit for 14 months. At every environmental screening, samples were obtained from the same 4 sites in each bed space. Patients were screened at admission and then 3 times weekly. All environmental and patient strains were typed using pulsed-field gel electrophoresis., Results: MRSA was isolated from the environment at every environmental screening, when both small and large numbers of patients were colonized. Detailed epidemiological typing of 250 environmental and 139 patient isolates revealed 14 different pulsed-field gel electrophoresis profiles, with variants of EMRSA-15 being the predominant type. On only 20 (35.7%) of 56 occasions were the strains isolated from the patients and the strains isolated from their immediate environment indistinguishable. There was strong evidence to suggest that 3 of 26 patients who acquired MRSA while in the intensive care unit acquired MRSA from the environment., Conclusions: This study reveals widespread contamination of the hospital environment with MRSA, highlights the complexities of the problem of contamination, and confirms the need for more-effective cleaning of the hospital environment to eliminate MRSA.

Staphylococcal interspersed repeat unit typing has previously been shown to have the ability to discriminate between epidemic methicillin-resistant Staphylococcus aureus strains in the United Kingdom. The current study illustrates its ability to distinguish between strains within an endemic setting thereby providing a rapid transportable typing method for the identification of transmission events.

Variable-number tandem repeats (VNTRs) have been shown to be a powerful tool in the determination of evolutionary relationships and population genetics of bacteria. The sequencing of a number of Staphylococcus aureus genomes has allowed the identification of novel VNTR sequences in S. aureus, which are similar to those used in the study of the evolution of Mycobacterium tuberculosis clades. Seven VNTRs, termed staphylococcal interspersed repeat units (SIRUs), distributed around the genome are described, occurring in both unique and multiple sites, and varying in length from 48 to 159 bp. Variations in copy numbers were observed in all loci, within both the sequenced genomes and the UK epidemic methicillin-resistant S. aureus (EMRSA) isolates. Clonally related UK EMRSA isolates were clustered using SIRUs, which provided a greater degree of discrimination than multi-locus sequence typing, indicating that VNTRs may be a more appropriate evolutionary marker for studying transmission events and the geographical spread of S. aureus clades.

Methicillin resistant Staphylococcus aureus (MRSA) is endemic within many hospitals worldwide. Critically ill patients on intensive care units have increased risk factors making them especially prone to nosocomially acquired infections. This review addresses the current situation regarding the evolution of MRSA and the techniques for identifying and epidemiologically typing it. It discusses specific risk factors, the morbidity and mortality associated with critically ill patients, and possibilities for future antibiotic treatments.

Aim: To develop and evaluate a TaqMan(TM) polymerase chain reaction (PCR) for the rapid identification and speciation of candida species., Methods: Species specific primer and probe sets were designed for the identification of Candida albicans, C. parapsilosis, C. tropicalis, C. krusei, C. kefyr, and C. glabrata from clinical isolates in a 5' exonuclease (TaqMan(TM)) assay. The probes were labelled with three fluorescent dyes to enable differentiation between species when three primer and probe sets were combined in two multiplexes. The specificity of these assays was evaluated against a range of National Collection of Pathogenic Fungi strains, clinical isolates of yeast, bacterial and viral pathogens., Results: The primer and probe sets have been shown to be 100% specific for their respective species; there was no crossreaction between any set and human DNA, or extracts from other candida species, fungal, bacterial, or viral pathogens tested. Extracts from two clinical isolates, originally identified as C albicans on the basis of germ tube formation, were not amplified by any of the primer and probe sets. These isolates have been putatively re-identified as C dubliniensis after sequencing of the variable internal transcribed spacer region ITS2 and lack of growth at 45 degrees C., Conclusion: This TaqMan assay provides a rapid alternative to conventional culture based techniques for the identification and speciation of the most frequently isolated candida species. The simple extraction method followed by TaqMan PCR can identify the six species mentioned in four hours.

Faeces received in a diagnostic laboratory were screened for glycopeptide-resistant enterococci (GRE) on modified Lewisham medium, with and without enrichment in Enterococcosel broth. Colonization by GRE was detected in 102/838 patients (12.2%). In 74 (73%) of colonized patients GRE were detected by both methods and in 28 (27%) they were detected only after enrichment. The carriage rate in hospitalized patients was 32% (93/289) compared with 2.3% (11/425) in the community (GP patients and food-handlers). Carriage of GRE increased with age. Clostridium difficile isolation was associated with GRE colonization, odds ratio 6.76 (P<0.001). Fifty-nine percent (60/102) of the GRE had the VanA phenotype and 41% (42/102) had the VanB phenotype. In the community VanA predominated (91%), whereas 64% (57/89) of the isolates from hospitalised patients were of the VanB phenotype., (Copyright 1999 The Hospital Infection Society.)

Although the explanation for sudden infant death syndrome (SIDS) remains unknown, an increasing body of evidence now exists to suggest a possible role for bacterial toxins in the aetiology, and a number of investigators have considered that endotoxaemia could explain some of the associated features. Following the development of an animal model which confirmed that endotoxaemia could be detected after death, we studied endotoxin levels in blood and tissue samples taken at autopsy from SIDS infants, child controls and adult controls. There were significant correlations between endotoxin levels in blood and the various organs sampled particularly in SIDS cases and child controls, and blood endotoxin levels in SIDS cases were higher in those infants where there was histological evidence of mild to moderate inflammation. However, overall no significant differences were found between endotoxin levels in blood or tissue in the three study groups. Further studies into possible actions or interactions of endotoxin in SIDS are required.

The aim of the study was to test the following hypotheses: (i) that endotoxin injected 40 min prior to death can be detected in rat organs post mortem and (ii) that endotoxin levels do not change with increasing time post mortem. Rats were injected with or without endotoxin in buffered saline, 40 min prior to being killed. Endotoxin levels in rat organs were assessed using a Limulus amoebocyte assay. The effect of storage time post mortem was assessed by following various storage regimes at 25 degrees C and 8 degrees C. Significant differences (P = < 0.001) in endotoxin levels of all samples tested were found between rats injected with and without endotoxin. A significant increase in detectable endotoxin was observed between 0 h and 6 h post mortem in rats injected with or without endotoxin. No difference in detectable endotoxin levels in the kidney, liver and spleen was observed from 30 h to 102 h post mortem in rats injected with or without endotoxin. In rats injected with endotoxin, detectable endotoxin levels in the heart were raised between 0 h and 6 h, 6 h and 54 h, and 30 h and 78 h. Endotoxin injected into rats 40 min prior to death can be detected post mortem. For rats injected with saline or endotoxin prior to death levels in the kidney, liver and spleen were not affected by storage at 8 degrees C for 30-102 h, after initial storage at room temperature for 6 h. Levels of endotoxin detected in the hearts of rats injected with saline were not affected by storage up to 102 h. In rats injected with endotoxin prior to death, detectable levels in the heart were significantly affected by increasing time in storage.

Bacteriophage typing is currently the recognised methodology for the typing of methicillin-resistant Staphylococcus aureus (MRSA) in the UK. Bacteriophage typing is less discriminatory and does not type all isolates compared with some molecular methods for typing MRSA. Chromosomal genotyping by pulsed-field gel electrophoresis (PFGE) is increasingly recognised as an improved method for typing MRSA, providing increased discrimination and typability. In this study the results of a comparison of bacteriophage typing and PFGE typing and subtyping are presented for a large collection of isolates from the North-West of England. Isolates belonging to the most frequently isolated epidemic methicillin-resistant Staphylococcus aureus (EMRSA) bacteriophage types 15 and 16 were typed by PFGE with further discrimination of common PFGE types possible into a number of subtypes. These results for a large collection of isolates demonstrate the improved typing of MRSA with PFGE. The widespread acceptance of PFGE for typing MRSA isolates has been hampered by the lack of standardised methodologies. Recently, a standardised PFGE strain typing system, known as the GenePath system has become available. The results of an inter-laboratory comparison of PFGE typing for a collection of isolates demonstrated good reproducibility with this system.

Methicillin-resistant Staphylococcus aureus (MRSA) is increasingly common in hospital and community populations, making the recognition of true nosocomial outbreaks more difficult. We have used pulsed-field gel electrophoresis (PFGE) with Sma I digestion to analyse retrospectively two perceived outbreaks of epidemic methicillin-resistant Staphylococcus aureus 15 (EMRSA 15) colonization. The first cluster of cases in patients and staff on a general ward (ward D) revealed three different antibiograms based on differences in ciprofloxacin and rifampicin sensitivities. All isolates typed using PFGE, which was more discriminatory than phage-typing. One PFGE banding profile labelled type 5 was predominant, but 12 isolates proved to be subtypes of type 5 and two were PFGE type 11. Four staff members carried a strain not found in patients, three carried strains found in patients and transient carriage was highlighted as a problem when screening staff. PFGE enhanced the epidemiological data and proved that the cases on this ward did not comprise one large outbreak but numerous sporadic cases and smaller clusters. In contrast, isolates from a second cluster of cases which occurred on ward F were indistinguishable using antibiograms, phage-typing and PFGE, confirming this was more likely to be a true outbreak of colonization. We conclude that PFGE usefully augments epidemiological information and allows more logical infection control decisions to be made, with better utilization of scarce resources.

Invasive fungal infections are an increasingly common problem in cancer patients and in other vulnerable groups. Invasive pulmonary aspergillosis (IPA) in the neutropenic host presents particular challenges in terms of diagnosis and therapy. Against the background of a recognized problem of invasive aspergillosis in haematology/oncology patients treated at the Christie Hospital, we undertook a prospective study in patients at risk for IPA. The aim of the study was to improve outcome by using the linked strategies of first, early diagnosis, and secondly, early aggressive therapy with a lipid-associated formulation of amphotericin B, amphotericin B colloidal dispersion ('Amphocil'). Early investigation comprised the use of high-resolution computerized tomography scanning of the thorax and fibreoptic bronchoscopy to obtain bronchoalveolar lavage specimens, processed using conventional detection and culture methods. Using this approach, the incidence of proven or probable IPA in patients with acute leukaemia was 9%. Prompt initiation of amphotericin B colloidal dispersion therapy led to a successful outcome in 11 of 13 patients, compared with a mortality of 100% in historical controls.

In a comparative study, isolates of methicillin-resistant Staphylococcus aureus (MRSA) with known pulsed-field gel electrophoresis (PFGE) and bacteriophage type were analysed by polymerase chain reaction (PCR) and restriction fragment length polymorphisms (RFLP) for additional discriminatory subtyping information. PFGE was previously performed using standardized, commercially available kits and pre-programmed software. Isolates were examined for coagulase (coa) and protein A (spa) gene polymorphisms following PCR amplification of the coa hypervariable and spa repeat regions. Coa gene RFLPs produced a total of 38 distinct combined patterns after digestion with HaeIII and AluI and identified the predominant epidemic (EMRSA) types 15 and 16. A unique HaeIII restriction site was identified by RFLP and sequence analysis in the coa gene for EMRSA 15 but not EMRSA 16. The spa gene PCR yielded a total of 14 different profiles ranging from 3-18 repeats with the 2 predominant EMRSA types falling into 2 distinct groups. PCR detection of coa and spa polymorphisms offer a rapid preliminary strain identification and discriminatory subtyping information for surveillance of MRSA.

Aim: To evaluate a newly developed aspergillus mitochondrial gene PCR-ELISA assay for the early diagnosis of invasive pulmonary aspergillosis (IPA) in neutropenic patients., Methods: The aspergillus mitochondrial gene was chosen for the amplification target for use with a solution hybridisation assay with colorimetric end stage detection in microtitre plate format (PCR-ELISA). The study group comprised neutropenic patients undergoing febrile episodes not responding to standard antibacterial antibiotics. Patients underwent computed tomography and bronchoscopy. Bronchoalveolar lavage (BAL) fluids were examined by culture and PCR., Results: The aspergillus mitochondrial gene PCR-ELISA was both sensitive (100%) and specific (100%) for IPA in neutropenic patients. All 12 patients with definite or probable IPA had PCR positive BAL fluids. None of the patients with undiagnosed or confirmed infections of other aetiologies were mitochondrial PCR positive. Speciation based upon amplicon size difference was possible., Conclusions: Aspergillus mitochondrial DNA PCR-ELISA on BAL fluid is useful in the early diagnosis of IPA in neutropenic patients alone or, potentially, as an indication for thoracic computed tomography.

Aim: To assess whether the information provided by automated continuous monitoring blood culture systems could aid in the diagnosis of catheter related sepsis., Methods: Serial dilutions of a strain of coagulase negative staphylococcus were inoculated into the BacT/Alert blood culture system. Blood culture results for seven patients with possible catheter related sepsis from coagulase negative staphylococci were reviewed., Results: Time to positivity and length of lag period were strongly related to the concentration of bacteria inoculated (average decrease of 1.5 hours to positivity for each 10-fold increase in concentration). Time to positivity and length of lag period were significantly shorter for central line blood cultures than for those taken from peripheral sites., Conclusions: Using data already measured by continuous monitoring blood culture systems may provide a simple alternative to quantitative blood cultures for the diagnosis of catheter related sepsis.

Over the past 20 years there has been a dramatic shift in the pattern of infection in neutropenic patients. During the 1970s Gram-negative organisms caused approximately 70% of all bacteraemias, but by the late 1980s the situation had reversed and approximately 70% of bacteraemias were due to Gram-positive organisms. The main contributors to this increase in Gram-positive infections have been the coagulase-negative staphylococci and the viridans streptococci. More recently, enterococci have emerged as significant pathogens in this patient group, and the development of glycopeptide resistance in the enterococci is of particular concern since this class of antibiotics is widely used in neutropenic patients. Among Gram-negative organisms, the emergence of resistance to fluoroquinolones, particularly in Escherichia coli, is a worrying feature which may lead to a reassessment of the use of quinolone prophylaxis in this setting.

Finer discrimination between strains of methicillin resistant Staphylococcus aureus (MRSA) than phage typing can provide is needed to identify and characterise spread of infection in outbreaks. This study compares three molecular methods with each other and with phage typing. Pulsed field gel electrophoresis provides the greatest discrimination, but finer discrimination is obtainable by combining methods.

Increasing numbers of elderly people are being treated in hospitals and are at particular risk of acquiring infections. The incidence, risk factors and types of hospital-acquired infection (HAI) in the elderly are reviewed. Special reference is made to urinary tract infections, respiratory tract infections, gastrointestinal infections including Clostridium difficile, bacteraemia, skin and soft tissue infections and infections with antibiotic-resistant organisms.

The AromaScan system was used to analyse vaginal swabs from 68 women attending a genitourinary clinic. Using clinical criteria, subjects were assessed for bacterial vaginosis. After training the AromaScan system to recognise patterns generated from four patients with and four patients without bacterial vaginosis, 16 of the 17 (94%) remaining subjects were correctly identified as having the condition. The positive predictive value of the test was 61.5%. These results indicate that the AromaScan technology may be of value as a screening test for bacterial vaginosis.

A pilot study using electronic aroma detection was performed over a six-month period to assess the aroma of chronic non-healing venous leg ulcers and the effect of appropriate antibiotic therapy on modification of the aroma. Deep infection with pathogenic organisms was found on biopsy culture in 13 out of 15 patients. Odour analysis was performed at weekly intervals on the ulcer dressings using an AromaScan instrument. Data points on the aroma maps moved from their pre-treatment presentation. Alterations in aroma data correlated well with the progress of the ulcers. Aroma analysis is shown to be a potential tool in monitoring the progress towards healing of chronic venous ulcers.

Antibodies to the core region of endotoxin (endotoxin core antibodies, EndoCAb), which cross-react with endotoxin from a range of Gram-negative bacteria, are maintained in relative homeostasis in health, but undergo marked changes in a number of different diseases associated directly or indirectly with endotoxaemic or septicaemic states. The levels of EndoCAb IgG in the blood and cerebrospinal fluid (CSF) of 35 late-stage sleeping sickness patients and 9 control individuals were measured by ELISA. EndoCAb levels were significantly elevated in the patient blood (mean EndoCAb value 290 MU/ml cf. control 182 MU/ml, P < 0.001), and CSF (mean EndoCAb value 254 MU/ml cf. control 150 MU/ml, P < 0.001). EndoCAb IgG levels correlated with endotoxin levels in patient blood (r = 0.78, P < 0.001), but not in the CSF and were not reduced 6 weeks following chemotherapy, unlike the endotoxin levels. It is concluded that late-stage sleeping sickness is associated with chronic exposure to endotoxins from Gram-negative bacteria.

OBJECTIVE: To compare four vancomycin-containing agar media for the isolation of glycopeptide-resistant enterococci (GRE) from clinical fecal specimens: kanamycin---aesculin---azide (KAA) agar; bile---aesculin---polymixin (BAP) agar; aztreonam---amphotericin blood (CBAA) agar; and neomycin blood (CBN) agar. METHODS: Fecal specimens from 125 patients were inoculated onto each medium. Media were examined for enterococci after incubation for up to 48 h. Enterococci were identified to species level, and glycopeptide phenotypes were determined by measuring minimum inhibitory concentrations of vancomycin and teicoplanin. RESULTS: GRE were isolated from 44/125 samples. Enterococcus faecalis and Enterococcus faecium isolates, expressing glycopeptide resistance of the VanA or VanB phenotypes, were recovered from 27/33 (82%) specimens on BAP medium, 26/33 (79%) on KAA medium, and 21/33 (64%) on CBN and CBAA media. Enterococcus gallinarum and Enterococcus casseliflavus isolates expressing low-level glycopeptide resistance (VanC phenotype) were recovered from 14/15 (93%) specimens on CBAA medium, 7/15 (47%) on KAA and CBN media, and 6/15 (40%) on BAP medium. CONCLUSIONS: The media tested in this study, with the exception of CBN medium, detected at least 75% of patients colonized by GRE. Further development of BAP, CBAA and KAA media is warranted to improve growth and selectivity.

Penicillin has long been recognized as the antibiotic of choice for treatment of meningococcal infections, but clinicians have recently become concerned about the susceptibility of meningococci to penicillin and other antibiotics used in the management of meningococcal disease. Strains relatively resistant to penicillin (minimum inhibitory concentrations ranging from 0.1 mg/L to 1.28 mg/L) have been reported from a large number of countries, although the frequency with which such isolates are found varies widely. The mechanism of relative resistance to penicillin involves, at least in part, the production of altered forms of one of the penicillin-binding proteins. Although treatment with penicillin is still effective against these relatively resistant strains, there is evidence that low-dose treatment regimens can fail. beta-Lactamase production in meningococci is extremely rare but has been reported, and this finding is of great concern. Resistance to sulfonamides and rifampin is of particular concern in regard to the management of contacts of patients with meningococcal disease.

The clinical and molecular epidemiology of two clusters of colonization and infection of patients by glycopeptide-resistant enterococci (GRE) on a leukaemia and bone marrow transplantation unit was studied over a two-and-a half-year period. Thirty-five patients became colonized, of whom six developed clinical infections. Of the 53 isolates of GRE, 49 were Enterococcus faecium, multiply-resistant to vancomycin and ampicillin. DNA fingerprinting of 48 E. faecium isolates by pulsed-field gel electrophoresis identified six DNA types. One strain of VanB phenotype E. faecium predominated during the initial outbreak, and an unrelated strain of the VanA phenotype was present in a second cluster. Environmental and patient isolates of E. faecium were indistinguishable by DNA typing. The VanA phenotype enterococci probably arose by transfer from the renal ward at a nearby hospital, and a patient with persistent diarrhoea may have contributed to contamination and cross-infection. GRE may cause significant infections in immunocompromised patients, and are readily transmitted between them. GRE were controlled, but not eradicated on the unit; infection control measures included improved environmental cleaning and modification of antibiotic use. In order to control GRE, it is necessary to educate healthcare workers and implement the traditional, effective values of good personal hygiene and environmental cleanliness.