Screening for latent tuberculosis in UK health care workers.

BACKGROUND: Active tuberculosis (TB) infection was diagnosed in two health care workers (HCWs) originally from high-incidence countries at a National Health Service (NHS) hospital trust in Northern England. In response, the trust screened current clinical employees from countries with a high TB prevalence for active and latent TB infection (LTBI). AIMS: To identify the number of HCWs, within the organization, who are at risk of TB infection. METHODS: Clinical employees from countries with a high TB prevalence (those described by the World Health Organization as having an incidence of >40 cases per 100000 populations) were reviewed. Employees were identified via human resource systems and occupational health records, from which nationality or country of birth was identified. A letter was sent to identified employees advising of the rationale for a reviewed screening process and inviting them to attend for an interferon-gamma release assay (IGRA) blood test. RESULTS: A total of 587 clinical staff were identified as fitting the criteria of clinical HCWs from high-incidence countries. Of 469 HCWs screened, 27% screened positive using IGRA. This represented 4% of the total clinical workforce for the organization. CONCLUSIONS: A considerable proportion of the workforce at this NHS hospital trust had previously undiagnosed LTBI, carrying a risk of conversion to active disease. Further action, such as treatment of LTBI or increased workforce awareness of symptoms of active disease, could help to reduce the risk of transmission of TB to patients and the need for associated 'look-back' exercises.

Rate of cyp51A mutation in Aspergillus fumigatus among lung transplant recipients with targeted prophylaxis.

OBJECTIVES: The most common mechanism of azole (itraconazole and voriconazole) resistance in Aspergillus fumigatus is a mutation at the cyp51A locus. The aim of our study was to determine the rate of cyp51A mutations in lung transplant recipients (LTR) undergoing targeted antifungal prophylaxis with 12 weeks of voriconazole. METHODS: We conducted a prospective study that included 22 LTR with A. fumigatus between October 2008 and November 2011. Of those, 10 LTR were colonized with A. fumigatus and 12 had invasive pulmonary aspergillosis. RESULTS: Four patients were found to have A. fumigatus isolates with a cyp51A mutation, two had colonization and two had invasive pulmonary aspergillosis. The remaining 18 LTR had WT cyp51A A. fumigatus isolates. All A. fumigatus isolates (except one due to mixed growth) were tested for antifungal susceptibility. A total of nine LTR were exposed to azoles prior to A. fumigatus isolation for a median duration of 249 (IQR 99-524) days. Azole exposure preceded the isolation of two mutant isolates and seven WT isolates. None of the cyp51A mutant isolates conferred phenotypic resistance to azoles. CONCLUSIONS: Targeted antifungal prophylaxis in LTR did not lead to cyp51A resistance mutations in this cohort. Data on larger cohorts who receive universal antifungal prophylaxis are needed.

An invertebrate model to evaluate virulence in Aspergillus fumigatus: the role of azole resistance.

The impact of different mutations in the Aspergillus fumigatus ergosterol biosynthesis pathway on pathogenesis has been evaluated using a simple invertebrate mini host, the caterpillar Galleria mellonella. A set of strains that includes clinical isolates and isogenic mutants with mutations at the cyp51A gene conferring azole resistance were studied. All strains demonstrated a similar in vitro growth pattern and are equally virulent against the insect larvae. These results suggest that in A. fumigatus acquisition of this particular azole-resistance mechanism would not imply any significant change in virulence. G. mellonella may provide a convenient and inexpensive model for the in vivo prescreening of mutants of A. fumigatus, contributing to the generation of a hypotheses that can be further tested in refined experiments in mammalian models.

Impact of Bolus dosing versus continuous infusion of Piperacillin and Tazobactam on the development of antimicrobial resistance in Pseudomonas aeruginosa.

Management of nosocomial pneumonia is frequently complicated by bacterial resistance. Extended infusions of beta-lactams are increasingly being used to improve clinical outcomes. However, the impact of this strategy on the emergence of antimicrobial resistance is not known. A hollow-fiber infection model with Pseudomonas aeruginosa (PAO1) was used. Pharmacokinetic (PK) profiles of piperacillin-tazobactam similar to those in humans were simulated over 5 days. Three dosages of piperacillin-tazobactam were administered over 0.5 h or 4 h, with redosing every 8 h. Two initial bacterial densities were investigated (∼10(4) CFU/ml and ∼10(7) CFU/ml). The time courses of the total bacterial population and the resistant subpopulation were determined. All data were described using a mathematical model, which was then used to define the relationship between drug concentrations, bacterial killing, and emergence of piperacillin resistance. There was logarithmic growth in controls in the initial 24 h, reaching a plateau of ∼9 log10 CFU/ml. Bacterial killing following administration of piperacillin via bolus dosing and that after extended infusions were similar. For the lower initial bacterial density, trough total plasma piperacillin concentration/MIC ratios of 3.4 and 10.4 for bolus and extended-infusion regimens, respectively, were able to suppress the emergence of piperacillin resistance. For the higher initial bacterial density, all regimens were associated with progressive growth of a resistant subpopulation. A stratified approach, according to bacterial density, is required to treat patients with nosocomial pneumonia. Antimicrobial monotherapy may be sufficient for some patients. However, for patients with a high bacterial burden, alternative therapeutic strategies are required to maximize bacterial killing and prevent antimicrobial resistance.

Pharmacokinetics and pharmacodynamics of anidulafungin for experimental Candida endophthalmitis: insights into the utility of echinocandins for treatment of a potentially sight-threatening infection.

Candida chorioretinitis and endophthalmitis are relatively common manifestations of disseminated candidiasis. Anidulafungin is increasingly used for the treatment of disseminated candidiasis, but its efficacy for Candida endophthalmitis is not known. A nonneutropenic model of hematogenous Candida endophthalmitis was used. Anidulafungin at 5, 10, and 20 mg/kg was initiated at 48 h postinoculation. The fungal densities in the kidney and vitreous humor were determined. Anidulafungin concentrations in the plasma and vitreous humor were measured using high-performance liquid chromatography (HPLC). A pharmacokinetic-pharmacodynamic model was used to link anidulafungin concentrations with the observed antifungal effect. The area under the concentration-time curve (AUC) associated with stasis was determined in the both the kidney and the vitreous humor. The results were bridged to humans to identify likely dosages that are associated with significant antifungal activity within the eye. Inoculation of Candida albicans resulted in logarithmic growth in both the vitreous humor and the kidney. The pharmacokinetics of anidulafungin were linear. There was dose-dependent penetration of the anidulafungin into the vitreous humor. The exposure-response relationships in the kidney and vitreous were completely discordant. AUCs of 270 and 100 were required for stasis in the eye and kidney, respectively. The currently licensed regimen results in an AUC for an average patient that is associated with stasis in the kidney but minimal antifungal activity in the eye. We conclude that anidulafungin penetrates the eye in a dose-dependent manner and that dosages higher than those currently licensed are required to achieve significant antifungal activity in the eye.

Major variations in Aspergillus fumigatus arising within aspergillomas in chronic pulmonary aspergillosis.

Aspergillomas develop from progressive layers of mycelial growth on the walls of pulmonary cavities over months. Aspergillomas are characteristic of chronic pulmonary aspergillosis and are a risk factor for azole resistance. We investigated genotypic and phenotypic alterations in Aspergillus fumigatus recovered from aspergillomas. Aspergillomas were removed from three patients (two at surgery, one at autopsy) and dissected. Overall 92 colonies of A. fumigatus were isolated. Microsatellite typing was conducted to determine genetic type. Itraconazole, voriconazole and posaconazole susceptibilities were performed. The cyp51A gene was sequenced in 22 isolates. Isolates from Patient 1 (n = 25) were azole susceptible and resistant, although all cyp51A sequences were wild type, the isolates split into two distinct clades. In Patient 2, isolates were less variable (n = 10), all were azole susceptible. In Patient 3 only azole-resistant strains (n = 57) were isolated, with M220K or M220T Cyp51A alterations, and microevolution was indicated. Marked diversity was observed in isolates from these patients; revealing differences in azole susceptibility, mechanism of resistance and genetic type. Importantly, routine sampling from respiratory specimens proved suboptimal in all cases; azole resistance was missed (Patient 1), cultures were negative (Patient 2) and high-level posaconazole resistance was not detected (Patient 3).

Disseminated Candidiasis caused by Candida albicans with amino acid substitutions in Fks1 at position Ser645 cannot be successfully treated with micafungin.

The clinical utility of the echinocandins is potentially compromised by the emergence of drug resistance. We investigated whether Candida albicans with amino acid substitutions at position Ser645 in Fks1 can be treated with either a conventional or an elevated dosage of micafungin. We studied Candida albicans (wild-type SC5314; MIC, 0.06 mg/liter) and four fks1 mutants (one FKS1/fks1 heterozygote mutant [MIC, 0.5 mg/liter] and three fks1/fks1 homozygous mutants [MICs for all, 2 mg/liter]) with a variety of amino acid substitutions at Ser645. The pharmacokinetic and pharmacodynamic relationships were characterized in a persistently neutropenic murine model of disseminated candidiasis. A mathematical model was fitted to all pharmacokinetic and pharmacodynamic data. This mathematical model was then used to "humanize" the murine pharmacokinetics, and the predicted antifungal effect was determined. The estimated maximal rate of growth and ultimate fungal densities in the kidney for each of the strains were similar. The administration of micafungin at 1 mg/kg of body weight to the wild type resulted in moderate antifungal activity, whereas the administration of 5 and 20 mg/kg resulted in rapid fungicidal activity. In contrast, the FKS1/fks heterozygote was killed only with 20 mg/kg, and the homozygous fks1 mutants failed to respond to any dosage. The bridging study revealed that human dosages of 100 and 400 mg/day were active only against the wild type, with no activity against either the heterozygote or the homozygote mutants. Ser645 Fks1 Candida albicans mutants cannot be treated with either conventional or elevated dosages of micafungin and should be deemed resistant.

A prospective survey of Aspergillus spp. in respiratory tract samples: prevalence, clinical impact and antifungal susceptibility.

A three-month laboratory-based prospective survey was conducted at four major university hospitals covering one-third of the Danish population in order to determine the prevalence, significance, and susceptibility pattern of aspergilli in airway samples. Samples received in January-March 2007 for routine microbiologic investigation were examined for Aspergillus following routine procedures and with extended incubation (5 days). Identification was done by morphologic criteria and susceptibility testing using EUCAST method for azoles and amphotericin B E-test. Invasive aspergillosis (IA) was evaluated using modified EORTC/MSG criteria. A total of 11,368 airway samples were received. Growth of Aspergillus spp. was found in 129 and 151 patients using routine and extended incubation, respectively. Three patients had proven IA (2%), 11 probable (7%), four had allergic bronchopulmonary aspergillosis (ABPA) (3%), but the majority was colonised (88%). Underlying conditions were cystic fibrosis in 82 patients (55%), chronic obstructive pulmonary disease in 19 (13%) and haematological disorder in 11 (7%). Twenty-six patients (18%) were at intensive care unit and 69 (47%) received steroid treatment. Azole MICs were elevated for five isolates as follows (itraconazole, posaconazole, voriconazole MICs [mg/L]): two A. fumigatus isolates (>4; >4; 2 and >4; 0.125; 1), one A. lentulus isolate (2; 2; 0.5) and two A. terreus isolates (2; 2; 2 and 2; 0.125; 1). For four isolates the amphotericin B MIC was >1 µg/ml (3/112 A. fumigatus, 1/2 A. terreus). In conclusion, Aspergillus appears to be an important pathogen in Denmark. Elevated itraconazole MICs were detected in 4% of the isolates including a multi-azole resistant isolate.

Understanding the influence of 24-hour movement behaviours on the health and development of preschool children from low-income South African settings: the SUNRISE pilot study.

Background: The International Study of Movement Behaviours in the Early Years, SUNRISE, was initiated to assess the extent to which young children meet movement behaviour guidelines (physical activity, sedentary behaviour, screen time, sleep). Objective: The South African SUNRISE pilot study assessed movement behaviours in preschool children from two low-income settings, and associations between these movement behaviours, adiposity, motor skills and executive function (EF). Methods: Preschool child/parent pairs (n = 89) were recruited from preschools in urban Soweto and rural Sweetwaters. Height and weight were measured to assess adiposity. Physical activity was assessed using accelerometers while sedentary behaviour, screen time and sleep were assessed via parent report. Fine and gross motor development were measured using the Ages and Stages Questionnaire-3, and EF was assessed using the Early Years Toolbox. Results: The proportion of children meeting the physical activity guideline was 84%, 66% met the sleep guideline, 48% met the screen time guideline, and 26% met all three guidelines. Rural children were more active, but spent more time on screens compared to urban children. Most children were on track for gross (96%) and fine motor (73%) development, and mean EF scores were in the expected range for all EF measures. EF was negatively associated with screen time, and gross motor skills were positively associated with physical activity. Conclusion: The South African SUNRISE study contributes to the growing literature on 24-hour movement behaviours in SA preschool children, and highlights that these behaviours require attention in this age group.

Calibration of magnetic probes in the vicinity of a conducting well.

Measuring magnetic fields near the edge of a plasma device can be complicated by the geometric effects of the ports through which such measurements are made. The primary effect is an attenuation of the magnetic field at the probe coil due to the field expanding into the finite sized conducting well of the port. In addition, it is possible to determine the correspondence between the location of a field line as it intersects the probe coil inside the well, with its location far from the perturbation of the well. Here we explore several methods of experimentally characterizing the magnetic fields in the vicinity of the magnetic probe ports of a vacuum vessel, with the aim of improving the interpretation of magnetic measurements needed for experiments in plasma physics.

EUCAST technical note on isavuconazole breakpoints for Aspergillus, itraconazole breakpoints for Candida and updates for the antifungal susceptibility testing method documents.

The Subcommittee on Antifungal Susceptibility Testing (AFST) of the ESCMID European Committee for Antimicrobial Susceptibility Testing (EUCAST) has determined breakpoints for isavuconazole and Aspergillus and for itraconazole and Candida spp., released a new document summarizing existing and new minimum inhibitory concentration ranges for quality control strains and revised the method documents for yeast and mould susceptibility testing. This technical note is based on the EUCAST isavuconazole and itraconazole rationale documents, version 1.0 of the routine and extended internal quality control for antifungal susceptibility testing as recommended by EUCAST, and the E.Def 7.3, E.Def 9.2 and E.Def 9.3 method documents (http://www.eucast.org).

Behavioral and fMRI evidence of the differing cognitive load of domain-specific assessments.

Standards-referenced educational reform has increased the prevalence of standardized testing; however, whether these tests accurately measure students' competencies has been questioned. This may be due to domain-specific assessments placing a differing domain-general cognitive load on test-takers. To investigate this possibility, functional magnetic resonance imaging (fMRI) was used to identify and quantify the neural correlates of performance on current, international standardized methods of spelling assessment. Out-of-scanner testing was used to further examine differences in assessment results. Results provide converging evidence that: (a) the spelling assessments differed in the cognitive load placed on test-takers; (b) performance decreased with increasing cognitive load of the assessment; and (c) brain regions associated with working memory were more highly activated during performance of assessments that were higher in cognitive load. These findings suggest that assessment design should optimize the cognitive load placed on test-takers, to ensure students' results are an accurate reflection of their true levels of competency.

Incidence and predictors of bulbar palsy after surgery for acoustic neuroma.

We present a retrospective review of the perioperative management and complications of 102 cases of acoustic neuroma operated on at an English teaching hospital over the last 9 years. Nine patients had a bulbar palsy postoperatively; five of those patients developed pulmonary complications. A bulbar palsy was more likely to occur in those with tumours > or = 3 cm. The occurrence of intraoperative bradycardias, present in nearly half of the bulbar palsy cases, did not help predict who would sustain this injury postoperatively. However, episodes of intraoperative hypotension served to better predict a bulbar palsy complication in conjunction with the presence of a large tumour. Post-operative airway management is an area of possible conflict of interest: the desire for early extubation to avoid the possibility of coughing and bucking on the endotracheal tube and the need to protect the airway of the patient with a lower cranial nerve deficit. Almost 10% of our patients did have such a deficit after surgery; and over half of them developed respiratory complications making this an important cause of postoperative morbidity. We recommend that the timing of extubation be judged on an individual basis for each patient. Those with tumours > or = 3 cm warrant particular concern. All patients should receive careful observation in a high dependency area for at least the first postoperative day.

Azole resistance in allergic bronchopulmonary aspergillosis and Aspergillus bronchitis.

Oral azole antifungal therapy is used extensively for all forms of aspergillosis, including allergic bronchopulmonary aspergillosis (ABPA). However, long-term therapy may increase the risk of resistance. Here we report itraconazole and voriconazole resistance with reduced susceptibility to posaconazole in Aspergillus fumigatus in two patients exposed to itraconazole. Patients were diagnosed with ABPA and Aspergillus bronchitis related to innate immune defects. An azole susceptible strain was initially isolated from patient 1, but later a genetically different azole-resistant strain was cultured, possibly related to sub-therapeutic itraconazole levels, which could be a trigger for selection of resistance. The mechanism of resistance identified in this case was an L98H change in Cyp51A, accompanied by a tandem repeat in the promoter region of cyp51A leading to increased expression. No cyp51A mutation was found in azole-resistant isolates recovered from patient 2. Both patients responded to posaconazole, with plasma levels of >1.0 mg/L. Subsequently, susceptible strains of different molecular types were cultured from both patients, suggesting eradication and replacement.

Flucytosine therapeutic monitoring: 15 years experience from the UK.

BACKGROUND: There is uniform consensus that flucytosine blood concentrations should be measured to avoid toxicity and ensure adequate efficacy. OBJECTIVES AND METHODS: The purpose of this study was to evaluate all flucytosine levels performed in a regional centre in the UK from October 1991 to May 2006. Concentrations were measured by bioassay. RESULTS: We reviewed 1071 flucytosine levels in 233 patients, including 33 neonates. Overall, only 20.5% of levels were in the expected therapeutic range. Low levels were observed in 40.5%, of which 5.1% were undetectable levels (<12.5 mg/L). High levels occurred in 38.9%, of which 9.9% were considered potentially toxic (>100 mg/L). High flucytosine levels occurred more frequently amongst neonates, which could be related to an immature renal system resulting in drug accumulation. CONCLUSIONS: Our findings reveal that the vast majority of patients were out of range for flucytosine levels. These data emphasize the importance of monitoring flucytosine levels.

Fungal contamination of bedding.

BACKGROUND: It is currently believed that most fungal exposure occurs external to the home. AIMS: To enumerate the fungal flora of used synthetic and feather pillows and the dust vacuumed from them, in the UK. METHODS: 10 pillows aged between 1.5 and >20 years in regular use were collected and quantitatively cultured for fungi. Swatches were taken from nine sections of the pillow and dust was also collected by vacuum from five pillows. Pillow vacuuming was carried out prior to pillow culture. All were cultured at room temperature, 30 and 37 degrees C for 7 days in broth before plating, and a subset were also cultured for 24 h in broth and then plated. Fungi were identified by standard morphological methods. RESULTS: The commonest three species isolated were Aspergillus fumigatus (n = 10), Aureobasidium pullulans (n = 6) and Rhodotorula mucilaginosa (n = 6). Another 47 species were isolated from pillows and vacuum dust. The number of species isolated per pillow varied from 4 to 16, with a higher number from synthetic pillows. Compared with the nonallergenic A. pullulans, more A. fumigatus was found in synthetic than feather pillows. CONCLUSIONS: We have examined pillows for fungal contamination, and show that the typical used pillow contains a substantial load of many species of fungi, particularly A. fumigatus. Given the time spent sleeping, and the proximity of the pillow to the airway, synthetic and feather pillows could be the primary source of fungi and fungal products. This has important implications for patients with respiratory disease, and especially asthma and sinusitis.

Sub-cutaneous phaeohyphomycosis caused by Cladophialophora devriesii in a United Kingdom resident.

An 83-year-old diabetic man receiving corticosteroids developed a forearm lesion. Histology confirmed the presence of a dematiaceous fungus, with associated granulomatous inflammation. Culture of a biopsy yielded fungal colonies with branching chains of single-celled, melanised, dry, sympodial conidia, which were identified as Cladophialophora devriesii on the basis of morphology and rDNA gene sequencing. To date, C. devriesii has been a relatively rare cause of human disease. To our knowledge, this is only the second case to be described, and the first report of infection in a UK resident.

Fast algorithm for implementing the minimum-negativity constraint for Fourier spectrum extrapolation. Part 2.

The basic theory and essential ideas of a new algorithm based on the minimum-negativity constraint for restoring the Fourier spectrum and improving resolution were set forth in an earlier paper. The method was proved to work for artificial noise-free data of 1024 data points. In the present paper the efficacy of the method will be demonstrated for an experimental data set of 32K data points in length (K = 1024). Further details of the new algorithm will be discussed. A brief comparison will be made between this method and some of the other popular methods for Fourier spectrum extrapolation. Also, a modification of the algorithm that will allow the computations to be implemented on a parallel-processing computer will be presented.

Improved restoration using a modification of the minimum-negativity iterative algorithm.

A method is reported for bringing about more rapid and complete convergence of the equations that implement the minimum-negativity constraint. The method utilizes an improved spatial function constructed by leaving the negative values intact and replacing the positive values with the differences between their values for the present and previous iteration. (This difference is found to converge after several iterations.) The advantage of this new spatial function is that convergence may be obtained for recalcitrant data using fast approximate procedures requiring the FFT. Several improvements were made in restoring inverse-filtered data. One is to impose an upper bound constraint (in the form of a tapered window) on the Fourier spectrum most degraded by noise and other errors. This provides stability, and the window can be used on the starting estimate and all succeeding iterations. Errors in the low frequency spectrum (below the cutoff radius) are determined and corrected by a slight modification of the restoration procedure described in this paper. Results are shown for simulated CO(2) interferograms, experimental interferometer data, and experimental IRAS data.

Synthesis and application of a poly(ethylene glycol)-antibody affinity ligand for cell separations in aqueous polymer two-phase systems.

The possibility of producing biospecific affinity ligands for separating cells in two polymer aqueous phase systems on the basis of cell surface antigens was investigated. Rabbit anti-human erythrocyte IgG was reacted with cyanuric chloride-activated monomethyl poly(ethylene glycol) (PEG) fractions (molecular weights approximately 200, 1900, and 5000) at various molar ratios of PEG to protein lysine groups. The partition coefficient of the protein in a Dextran/PEG two-phase system increased with increasing degree of modification and increasing PEG molecular weight. There was a concomitant loss in ability to agglutinate human erythrocytes. The ability of the modified IgG to bind to a DEAE-cellulose column was almost eliminated by reaction with the PEG 5000, and was decreased to a lesser extent by PEG 1900. This PEG 1900-modified IgG substantially increased the partition of fresh or fixed human erythrocytes into the PEG-rich phase of a suitable phase system, while having no effect on rabbit cell partition. The partition increase could be inhibited by unmodified anti-human red cell IgG but not by nonspecific unmodified human IgG, demonstrating that the ligand effects were specific for the cell type against which the antibody was raised. A mixture of rabbit and human erythrocytes, which ordinarily have very similar partitions in the phase systems used, could be separated on a countercurrent distribution apparatus using the modified IgG. These results demonstrate the feasibility of producing immunologically specific affinity partition ligands for cell separation.