Miracle-dz/UniFAR-FTUs huggingface dataset:
Antimicrobial activity of the novel polymyxin derivative NAB739 tested against Gram-negative pathogens.

OBJECTIVES In spite of reported nephrotoxicity, polymyxins have been reinstated as the last-line therapy to treat infections caused by Gram-negative bacterial strains that are resistant to other agents. NAB739 has a cyclic portion identical to that of polymyxin B, but its linear peptide portion consists of threonyl-d-serinyl instead of diaminobutyryl-threonyl-diaminobutyryl. Therefore, NAB739 lacks both of the positive charges present in the linear part of polymyxin B. Here, we compare the antibacterial activity of NAB739 with that of polymyxin B against a representative collection of contemporary Gram-negative bacteria. METHODS NAB739 and polymyxin B MIC values were determined for 310 clinical isolates by the reference broth microdilution method according to CLSI document M07-A9 (2012). RESULTS MIC(90)s of NAB739 for the subset consisting of polymyxin-susceptible (MIC, ≤ 2 mg/L) clinical isolates of Escherichia coli (n=51), Klebsiella pneumoniae (n=50), Acinetobacter spp. (n=49) and Pseudomonas aeruginosa (n=49) were 2, 2, 8 and 16 mg/L, respectively. For polymyxin-non-susceptible strains of E. coli (n=12), K. pneumoniae (n=11), Acinetobacter spp. (n=11) and P. aeruginosa (n=14) the NAB739 MIC(90) was ≥ 64 mg/L. CONCLUSIONS The MIC(90) of NAB739 for polymyxin-susceptible strains of E. coli and K. pneumoniae was identical to and 2-fold higher than that of polymyxin B, respectively. For polymyxin-susceptible strains of Acinetobacter spp. and P. aeruginosa, the MIC(90) of NAB739 was 4-fold and 8-fold higher than that of polymyxin B, respectively. For polymyxin-non-susceptible strains of all these species, the MIC(90) values of NAB739 were high and 2- to 4-fold higher than those of polymyxin B.
Invasive candidiasis in intensive care units in China: a multicentre prospective observational study.

OBJECTIVES To describe the epidemiology, microbiology and management of invasive Candida infection (ICI) in intensive care units (ICUs) in China. METHODS A multicentre, prospective, observational study in 67 hospital ICUs across China. Patients were ≥18 years old with clinical signs of infection and at least one of the following diagnostic criteria: (i) histopathological, cytopathological or microscopic confirmation of yeast cells from a normally sterile site; (ii) at least one peripheral blood culture positive for Candida; and (iii) positive Candida culture from a normally sterile site. The China-SCAN study is registered with ClinicalTrials.gov (NCT T01253954). RESULTS ICI incidence was 0.32% (306 patients/96,060 ICU admissions) and median time between ICU admission and diagnosis was 10.0 days. Candida albicans was the most prevalent single isolate (41.8% of patients), although non-albicans species accounted for the majority of infections. Diagnostic confirmation was based solely on at least one positive blood culture in 290 (94.8%) cases. Treatment was initiated after diagnostic confirmation in 166/268 (61.9%) patients. Triazoles (62.7%) and echinocandins (34.2%) were the most commonly used antifungal agents; first-line therapy was typically fluconazole (37.7%). The median duration of antifungal therapy was 14 days. The mortality rate was 36.6% (112/306); the median time between diagnosis and death was 14.5 days. Mortality was higher in older individuals, those with solid tumours, those with recent invasive mechanical ventilation and those with a higher sequential organ failure assessment score at diagnostic confirmation. Susceptibility to first-line antifungals was associated with lower mortality than dose-dependent susceptibility or complete resistance (P=0.008). CONCLUSIONS More infections were caused by non-albicans than Candida albicans strains. The majority of patients were treated only after diagnostic confirmation, rather than empirically. First-line antifungal susceptibility was associated with lower mortality.
Membrane Vesicle ABC Transporter Assays for Drug Safety Assessment

The use of plasma membrane vesicles that overexpress the bile salt export pump (BSEP) or multidrug resistance−associated protein 2, 3, or 4 (MRP2‐4) with an in vitro vacuum filtration system offers a rapid and reliable means for screening drug candidates for their effects on transporter function in hepatocytes and thus their potential for causing drug‐induced liver injury (DILI). Comparison of transporter activity in the presence and absence of ATP allows for determination of a specific assay window for each transporter. This window is used to determine the degree to which each test compound inhibits transporter activity. This assay battery is helpful for prioritizing and rank‐ordering compounds within a chemical series with respect to each other and in the context of known inhibitors of transporter activity and/or liver injury. This model can be used to influence the drug development process at an early stage and provide rapid feedback regarding the selection of compounds for advancement to in vivo safety evaluations. A detailed protocol for the high‐throughput assessment of ABC transporter function is provided, including specific recommendations for curve‐fitting to help ensure consistent results. Curr. Protoc. Toxicol. 54:23.5. 1‐23.5. 24. © 2012 by John Wiley & Sons, Inc.
Assessment of Drug Transporter Function Using Fluorescent Cell Imaging

ATP‐binding cassette (ABC) proteins, including the breast cancer resistance protein (BCRP) and multidrug resistance proteins (MDRs), actively transport structurally diverse chemicals from a number of tissues and are being increasingly cited as mediators of clinically relevant drug‐drug interactions. The potential outcomes of concomitantly administering two drugs that interact at the same transporter include altered disposition and toxicity and/or efficacy of one or both of the drugs. Research demonstrating the role of transporters in clinical pharmacokinetics has shed light on the need for in vitro screening methods that detect drug‐transporter interactions during preclinical development. This unit describes cell‐based procedures for detecting functional inhibitors of BCRP and MDR1 by measuring fluorescent substrate accumulation in suspended cells using an automated cell counter, which offers convenience, sensitivity, and speed in measuring intracellular fluorescence and identifying new inhibitors. An alternative method is provided for making similar measurements using a spectrophotometer with fluorescence detection capabilities. Curr. Protoc. Toxicol. 57:21.12. 1‐21.12. 15. © 2013 by John Wiley & Sons, Inc.
Steady-state pharmacokinetics and metabolism of voriconazole in patients.

OBJECTIVES Voriconazole exhibits non-linear pharmacokinetics in adults and is said to be mainly metabolized by CYP2C19 and CYP3A4 to voriconazole-N-oxide. The aim of this study was to obtain data on steady-state pharmacokinetics after dosing for at least 14 days in patients taking additional medication and in vivo data on metabolites other than voriconazole-N-oxide. PATIENTS AND METHODS Thirty-one patients receiving voriconazole as regular therapeutic drug treatment during hospitalization participated in this prospective study. Pharmacokinetic profiles were obtained for the 12 h (dosing interval) after the first orally administered dose (400 mg) or (if possible and) after an orally administered maintenance dose (200 mg) following intake for at least 14 days (n = 14 after first dose; n = 23 after maintenance dose). Blood and urine samples were collected and the concentrations of voriconazole and three of its metabolites (the N-oxide, hydroxy-voriconazole and dihydroxy-voriconazole) were determined, as well as the CYP2C19 genotype of the patients. All other drugs taken by the participating patients were evaluated. RESULTS A high variability of exposure (AUC) after the first dose was slightly reduced during steady-state dosing for voriconazole (82% to 71%) and the N-oxide (86% to 56%), remained high for hydroxy-voriconazole (79%) and even increased for dihydroxy-voriconazole (97% to 127%). In 16 of the 22 steady-state patients, trough plasma concentrations were <2 μg/mL. N-oxide plasma concentrations during steady state stayed almost constant. Hydroxylations of voriconazole seem to be quantitatively more important in its metabolism than N-oxidation. CONCLUSIONS High variability in voriconazole exposure, as well as low steady-state trough plasma concentrations, suggest that the suggested steady-state dosage of 200 mg twice a day has to be increased to prevent disease progression. Therapeutic drug monitoring is probably necessary to optimize the voriconazole dose for individual patients.
Stress, coping and psychological well-being amongst healthcare professionals employed within forensic inpatient settings

Although forensic services are often regarded as stressful, dangerous and emotionally demanding environments, there has been a surprising lack of research into the phenomena of stress amongst forensic healthcare professionals, especially in the UK. There is mounting evidence to suggest that burnout influences a significant proportion of forensic healthcare professionals and can result in a range of detrimental consequences for the individual, their organization and clients. The small body of research evidence that has examined burnout amongst forensic healthcare professionals is reviewed. Methodological limitations are explored and questions regarding the generalisability and rigor of the available research findings are raised. The research study investigated stress, coping and psychological well-being amongst 135 forensic healthcare professionals employed within four Medium Secure Units in the UK. Background information and measures of psychological well-being, burnout, occupational stress, work satisfaction and coping were collected using a postal survey. The results showed that a substantial proportion of forensic healthcare professionals experienced markedly elevated levels of occupational stress and psychological distress, whilst moderate levels of burnout and a range of problem-focused, emotionfocused and palliative coping strategies were demonstrated. Contributions to theory, research and clinical practice are discussed. The strengths and limitations of the research study are further examined. Future research directions and potential implications for clinical practice are explored. Personal motivations and practical difficulties encountered during the research process are finally presented.
Enzymatic Digestion of Proteins in Gels for Mass Spectrometric Identification and Structural Analysis

Enzymatic digestion of proteins is a key technique used in protein identification. By combining the digestion with mass spectrometric detection, proteins at very low femtomole levels, and in some cases subfemtomole levels, can be identified. Typically, one‐ or two‐dimensional SDS‐PAGE is used to isolate the proteins of interest, followed by staining with Coomassie blue, digestion‐compatible silver stain, or Sypro Ruby for detection. Two‐dimensional (2‐D) fluorescence difference gel electrophoresis (DIGE), which uses Cy3 and Cy5 dyes for detection, allows comparison of two different sample states in order to locate proteins that are up‐ or down‐regulated. In each case, an in‐gel digestion, usually tryptic, is used with mass spectrometry to identify these proteins of interest. For large numbers of gel spots, robotic digestion can save time and money.
Heart Failure: Pathophysiology, Molecular Biology, and Clinical Management

Preface Overview, Definition, Historical Aspects Organ Physiology:The Failing Heart as a Weakened Pump Cell Biochemistry:Abnormal Excitation-Contraction Coupling,Contraction, and Relaxation Arrhythmias Energy Starvation The Hemodynamic Defense Reaction Neurohumoral Response:The Inflammatory Response The Hypertrophic Response:Programmed Cell Death Signal Transduction Within Cells of the Failing Heart Maladaptive Hypertrophy and the Cardiomyopathy of Overload:Familial Cardiomyopathies Therapeutic Strategies for Managing Heart Failure Care of the Heart Failure Patient Inde
Geographical clustering of mecC-positive Staphylococcus aureus from bovine mastitis in France.

French Agency for Food, Environmental and Occupational Health Safety (ANSES), Unité Antibiorésistance et Virulence Bactériennes, Lyon, France; National Reference Centre for Staphylococci, Laboratoire de Bactériologie, Hospices Civils de Lyon, Lyon, France; International Center for Infectiology Research, INSERM U1111, CNRS UMR5308, Université Lyon 1, ENS de Lyon, Lyon, France; Laboratoire Vétérinaire et Alimentaire de Meurthe-et-Moselle, Malzeville, France
Expression, Purification, and Characterization of Caspases

This unit describes a protocol to obtain milligram amounts of enzymatically active pure recombinant caspases. Specific details for the expression, purification of caspase‐3, ‐6, ‐7, ‐8, ‐9 and ‐10 are discussed along with strategies to obtain particular forms (e.g., the zymogen) of some of them.
Purification and Characterization of Gingipains

Gingipains are cysteine proteases produced in large quantities by Porphyromonas gingivalis which together constitute important virulence factors in the pathogenesis of periodontal disease by that organism. Described is this unit is an efficient procedure for the purification of gingipains from the growth medium of P. gingivalis strain HG66, along with detailed protocols for growth of the organism and basic characterization of the purified proteases. The purification procedure consists of acetone precipitation followed by gel filtration to separate high‐molecular‐mass gingipains (Kgp and HRgpA) from RgpB. Kgp and HRgpA are further separated on Arg‐Sepharose by the virtue of differential elution from the affinity matrix with lysine (Kgp) and arginine (HRgpA) eluant. Obtained from these procedures, the gingipains are stable and can be stored at −80°C for years without loss of activity. Curr. Protoc. Protein Sci. 49:21.20. 1‐21.20. 27. © 2007 by John Wiley & Sons, Inc.
Role of SMG‐1‐mediated Upf1 phosphorylation in mammalian nonsense‐mediated mRNA decay

SMG‐1, a member of the PIKK (phosphoinositide 3‐kinase‐related kinase) family, plays a critical role in the mRNA quality control system known as nonsense‐mediated mRNA decay (NMD). NMD protects cells from the accumulation of aberrant mRNAs with premature termination codons (PTCs) which encode nonfunctional or potentially harmful truncated proteins. SMG‐1 directly phosphorylates Upf1 helicase, another key component of NMD, upon recognition of PTC on postspliced mRNA during the initial round of translation. Phosphorylated‐Upf1 recruits the SMG‐5/SMG‐7 complex to induce ribosome dissociation and decapping‐mediated decay. Phospho‐Upf1 also recruits the SMG‐6 endonuclease which might be involved in endo‐cleavage. Upf1 ATPase/helicase activities are likely required for the activation of other mRNA decay enzymes and the mRNA‐protein complex dissociation to complete NMD. At present, a variety of tools are available that can specifically suppress NMD, and it has become possible to examine the contribution of NMD in a variety of physiological and pathological conditions.
Three-dimensional ultrasound in obstetrics and gynecology

Basis and principles of three-dimensional ultrasound. Principles of three-dimension volume imaging in sonography. Clinical applications of three-dimensional ultrasound in obstetrics. Three-dimensional ultrasound in the evaluation of fetalmalformations. Three-dimensional fetal cardiac imaging, Fetal biometry with three-dimensional ultrasound. The psychological impact of three-dimensional fetal imaging on the fetomaternal relationship. Three-dimensional ultrasound of the uterus.Three-dimensional transvaginal ultrasoonography: applications in infertility. Three-dimensional ultrasound in urogynecology. Index.
Longitudinal trends and cross-sectional analysis of English national hospital antibacterial use over 5 years (2008-13): working towards hospital prescribing quality measures.

OBJECTIVES There is global concern that antimicrobial resistance is a major threat to healthcare. Antimicrobial use is a primary driver of resistance but little information exists about the variation in antimicrobial use in individual hospitals in England over time or comparative use between hospitals. The objective of this study was to collate, analyse and report issue data from pharmacy records of 158 National Health Service (NHS) acute hospitals. METHODS This was a cohort study of inpatient antibacterial use in acute hospitals in England analysed over 5 years through a data warehouse from IMS Health, a leading provider of information, services and technology for the healthcare industry. Around 98% of NHS hospitals were included in a country with a population of 50 million residents. RESULTS There was a dramatic change in the usage of different groups of antibacterials between 2009 and 2013 with a marked reduction in the use of first-generation cephalosporins by 24.7% and second-generation cephalosporins by 41%, but little change in the use of third-generation cephalosporins (+5.7%) and fluoroquinolones (+1.6%). In contrast, use of co-amoxiclav, carbapenems and piperacillin/tazobactam increased by 60.1%, 61.4% and 94.8%, respectively. There was wide variation in the total and relative amounts of antibacterials used between individual hospitals. CONCLUSIONS Longitudinal analysis of antibacterial use demonstrated remarkable changes in NHS hospitals, probably reflecting governmental and professional guidance to mitigate the risk of Clostridium difficile infection. The wide variation in usage between individual hospitals suggests potential for quality improvement and benchmarking. Quality measures of optimal hospital antimicrobial prescribing need urgent development and validation to support antimicrobial stewardship initiatives.
β-Lactam/β-lactamase inhibitors versus carbapenems for the treatment of sepsis: systematic review and meta-analysis of randomized controlled trials.

BACKGROUND Data on the relative efficacy of β-lactam/β-lactamase inhibitors (BL/BLIs) versus carbapenems are scant. METHODS This is a systematic review and meta-analysis of randomized controlled trials (RCTs) comparing any BL/BLI versus any carbapenem for the treatment of sepsis. The primary outcome was all-cause mortality. A broad search was conducted with no restrictions on language, publication status or date. Two reviewers independently applied the inclusion criteria and extracted the data. Assessment of risk of bias was performed using the domain-based approach. Subgroup analyses were used to investigate heterogeneity and focus on patient groups more likely to harbour ESBL-positive bacteria. Risk ratios (RRs) with 95% CIs were calculated and pooled. RESULTS Thirty-one RCTs were included. There was no difference between BL/BLIs and carbapenems in terms of mortality (RR 0.98, 95% CI 0.79-1.20), without heterogeneity. No differences were observed with regard to clinical or microbiological failure and bacterial superinfections. The results were not affected by risk of bias. No differences were detected in the subgroups of patients with nosocomial infections, Gram-negative infections and neutropenic fever. Adverse events requiring discontinuation were more common with BL/BLIs, on account of an increased incidence of diarrhoea. However, Clostridium difficile-associated diarrhoea (RR 0.29, 95% CI 0.10-0.87) was more frequent with carbapenems and seizures were more frequent with imipenem (RR 0.21, 95% CI 0.05-0.93). CONCLUSIONS No differences in efficacy between BL/BLIs and carbapenems exist in RCTs including patient populations with a certain, albeit unknown, rate of ESBL-positive bacteria causing infections.
Role of 100S ribosomes in bacterial decay period

Ribosomal proteins S10 and S2 were each fused with GFP to track the fates of these proteins in the stationary growth phase and the following decay period in Escherichia coli. The fused proteins localized mainly in the cytoplasm, and their amounts were proportional to the colony‐forming unit. S10‐GFP strains that lacked genes responsible for regulating 100S ribosomes and S2‐GFP strain that was unable to form 100S both showed shortened stationary phases. This result indicates that these strains exhibit earlier death in the absence of 100S formation (S2‐GFP, S10‐GFP∆rmf and S10‐GFP∆hpf) and breakdown (S10‐GFP∆yfiA). Therefore, in addition to the mere presence of 100S, the correct timing of 100S formation and breakdown is required to maintain viability. We propose a model in which 100S acts as a tentative repository of ribosomes that are protected from degradation and provide a source of amino acids in later growth period.
Evolutionarily distinct bacteriophage endolysins featuring conserved peptidoglycan cleavage sites protect mice from MRSA infection.

OBJECTIVES In the light of increasing drug resistance in Staphylococcus aureus, bacteriophage endolysins [peptidoglycan hydrolases (PGHs)] have been suggested as promising antimicrobial agents. The aim of this study was to determine the antimicrobial activity of nine enzymes representing unique homology groups within a diverse class of staphylococcal PGHs. METHODS PGHs were recombinantly expressed, purified and tested for staphylolytic activity in multiple in vitro assays (zymogram, turbidity reduction assay and plate lysis) and against a comprehensive set of strains (S. aureus and CoNS). PGH cut sites in the staphylococcal peptidoglycan were determined by biochemical assays (Park-Johnson and Ghuysen procedures) and MS analysis. The enzymes were tested for their ability to eradicate static S. aureus biofilms and compared for their efficacy against systemic MRSA infection in a mouse model. RESULTS Despite similar modular architectures and unexpectedly conserved cleavage sites in the peptidoglycan (conferred by evolutionarily divergent catalytic domains), the enzymes displayed varying degrees of in vitro lytic activity against numerous staphylococcal strains, including cell surface mutants and drug-resistant strains, and proved effective against static biofilms. In a mouse model of systemic MRSA infection, six PGHs provided 100% protection from death, with animals being free of clinical signs at the end of the experiment. CONCLUSIONS Our results corroborate the high potential of PGHs for treatment of S. aureus infections and reveal unique antimicrobial and biochemical properties of the different enzymes, suggesting a high diversity of potential applications despite highly conserved peptidoglycan target sites.