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Nicotine is a major alkaloid in tobacco plants and an addictive component of tobacco products. Some bacteria grow on tobacco plants and have evolved the ability to metabolize nicotine. As part of our microbiology teaching lab, we used minimal media with nicotine as the sole carbon source to isolate nicotine-degrading bacteria from tobacco leaves and commercial tobacco products. Students then identified these bacteria using 16S rRNA sequencing and biochemical assays and assessed their ability to catabolize nicotine using UV spectroscopy. Students were able to isolate and identify 14 distinct genera that can metabolize nicotine. This modification of the commonly used unknown project gave students firsthand experience using selective media, and students got the opportunity to work with largely uncharacterized microbes with a real-world connection to public health, which increased student engagement. Students had the opportunity to think critically about why nicotine-degrading microorganisms associate with tobacco plants, why there are different bacteria that use the same specialized metabolism, and how these organisms are isolated from other bacteria using selective media., Competing Interests: The authors declare no conflict of interest.

Background: Despite recent systematic reviews suggesting their benefit for postoperative nausea, vomiting, or both (PONV) prevention, benzodiazepines have not been incorporated into guidelines for PONV prophylaxis because of concerns about possible adverse effects. We conducted an updated meta-analysis to inform future practice guidelines., Methods: We included randomised controlled trials (RCTs) of all languages comparing benzodiazepines with non-benzodiazepine comparators in adults undergoing inpatient surgery. Our outcomes were postoperative nausea, vomiting, or both. We assessed risk of bias for RCTs using the Cochrane Risk of Bias tool. We pooled data using a random-effects model and assessed the quality of evidence for each outcome using the Grading of Recommendations, Assessment, Development and Evaluation (GRADE) approach., Results: We screened 31 413 abstracts and 950 full texts. We included 119 RCTs; 104 were included in quantitative synthesis. Based on moderate certainty evidence, we found that perioperative benzodiazepine administration reduced the incidence of PONV (52 studies, n=5086, relative risk [RR]: 0.77, 95% confidence interval [CI] 0.66-0.89; number needed to treat [NNT] 16; moderate certainty), postoperative nausea (55 studies, n=5916, RR: 0.72, 95% CI 0.62-0.83; NNT 21; moderate certainty), and postoperative vomiting (52 studies, n=5909, RR: 0.74, 95% CI 0.60-0.91; NNT 55; moderate certainty)., Conclusions: Moderate quality evidence shows that perioperative benzodiazepine administration decreases the incidence of PONV. The results of this systematic review and meta-analysis will inform future clinical practice guidelines., Systematic Review Protocol: The protocol for this systematic review was pre-registered with PROSPERO International Prospective Register of Systematic Reviews (CRD42022361088) and published in BMJ Open (PMID 31831540)., (Copyright © 2023 British Journal of Anaesthesia. Published by Elsevier Ltd. All rights reserved.)

The use of wastewater-based surveillance (WBS) for detecting pathogens within communities has been growing since the beginning of the COVID-19 pandemic with early efforts investigating severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) RNA in wastewater. Recent efforts have shed light on the utilization of WBS for alternative targets, such as fungal pathogens, like Candida auris, in efforts to expand the technology to assess non-viral targets. The objective of this study was to extend workflows developed for SARS-CoV-2 quantification to evaluate whether C. auris can be recovered from wastewater, inclusive of effluent from a wastewater treatment plant (WWTP) and from a hospital with known numbers of patients colonized with C. auris. Measurements of C. auris in wastewater focused on culture-based methods and quantitative PCR (qPCR). Results showed that C. auris can be cultured from wastewater and that levels detected by qPCR were higher in the hospital wastewater compared to the wastewater from the WWTP, suggesting either dilution or degradation of this pathogenic yeast at downstream collection points. The results from this study illustrate that WBS can extend beyond SARS-CoV-2 monitoring to evaluate additional non-viral pathogenic targets and demonstrates that C. auris isolated from wastewater is competent to replicate in vitro using fungal-specific culture media., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Occupational exposure to ototoxicants, substances that can cause hearing loss alone or exacerbate hearing loss when exposure occurs in combination with noise, is a workplace hazard that is poorly understood. A review of existing research indicates that some solvents and heavy metals may be ototoxic, but few studies have attempted to estimate the impact of ototoxicant exposure on the United States worker population. Researchers examined trends in workplace exposure to ototoxicants among workers in the United States by comparing exposure data collected by the Occupational Safety and Health Administration against worker hearing loss data provided by the Bureau of Labor Statistics (BLS) for 2012-2019. The study found that the noise exposure data was strongly correlated to the hearing loss data using Pearson's correlation ( p < .001), confirming that the exposure data collected by OSHA is predictive of the risk of occupational illness as reported by BLS. Chi-square analysis indicates that reported hearing loss was more common among industry subsectors with exposure to ototoxicants than those without exposure to ototoxicants. These findings suggest that workers with coexposure to ototoxicants and noise may be at a higher risk of experiencing hearing loss than those exposed to noise alone, and action should be taken to minimize this risk.

Background: Benzodiazepine use is associated with delirium, and guidelines recommend avoiding them in older and critically ill patients. Their perioperative use remains common because of perceived benefits., Methods: We searched CENTRAL, MEDLINE, CINAHL, PsycInfo, and Web of Science from inception to June 2021. Pairs of reviewers identified randomised controlled trials and prospective observational studies comparing perioperative use of benzodiazepines with other agents or placebo in patients undergoing surgery. Two reviewers independently abstracted data, which we combined using a random-effects model. Our primary outcomes were delirium, intraoperative awareness, and mortality., Results: We included 34 randomised controlled trials (n=4354) and nine observational studies (n=3309). Observational studies were considered separately. Perioperative benzodiazepines did not increase the risk of delirium (n=1352; risk ratio [RR] 1.43; 95% confidence interval [CI]: 0.9-2.27; I 2 =72%; P=0.13; very low-quality evidence). Use of benzodiazepines instead of dexmedetomidine did, however, increase the risk of delirium (five studies; n=429; RR 1.83; 95% CI: 1.24-2.72; I 2 =13%; P=0.002). Perioperative benzodiazepine use decreased the risk of intraoperative awareness (n=2245; RR 0.26; 95% CI: 0.12-0.58; I 2 =35%; P=0.001; very low-quality evidence). When considering non-events, perioperative benzodiazepine use increased the probability of not having intraoperative awareness (RR 1.07; 95% CI: 1.01-1.13; I 2 =98%; P=0.03; very low-quality evidence). Mortality was reported by one randomised controlled trial (n=800; RR 0.90; 95% CI: 0.20-3.1; P=0.80; very low quality)., Conclusions: In this systematic review and meta-analysis, perioperative benzodiazepine use did not increase postoperative delirium and decreased intraoperative awareness. Previously observed relationships of benzodiazepine use with delirium could be explained by comparisons with dexmedetomidine., Systematic Review Protocol: PROSPERO CRD42019128144., (Copyright © 2022 British Journal of Anaesthesia. Published by Elsevier Ltd. All rights reserved.)

The 16 th Workshop on Recent Issues in Bioanalysis (16 th WRIB) took place in Atlanta, GA, USA on September 26-30, 2022. Over 1000 professionals representing pharma/biotech companies, CROs, and multiple regulatory agencies convened to actively discuss the most current topics of interest in bioanalysis. The 16 th WRIB included 3 Main Workshops and 7 Specialized Workshops that together spanned 1 week in order to allow exhaustive and thorough coverage of all major issues in bioanalysis, biomarkers, immunogenicity, gene therapy, cell therapy and vaccines. Moreover, in-depth workshops on ICH M10 BMV final guideline (focused on this guideline training, interpretation, adoption and transition); mass spectrometry innovation (focused on novel technologies, novel modalities, and novel challenges); and flow cytometry bioanalysis (rising of the 3rd most common/important technology in bioanalytical labs) were the special features of the 16 th edition. As in previous years, WRIB continued to gather a wide diversity of international, industry opinion leaders and regulatory authority experts working on both small and large molecules as well as gene, cell therapies and vaccines to facilitate sharing and discussions focused on improving quality, increasing regulatory compliance, and achieving scientific excellence on bioanalytical issues. This 2022 White Paper encompasses recommendations emerging from the extensive discussions held during the workshop and is aimed to provide the bioanalytical community with key information and practical solutions on topics and issues addressed, in an effort to enable advances in scientific excellence, improved quality and better regulatory compliance. Due to its length, the 2022 edition of this comprehensive White Paper has been divided into three parts for editorial reasons. This publication (Part 2) covers the recommendations on LBA, Biomarkers/CDx and Cytometry. Part 1 (Mass Spectrometry and ICH M10) and Part 3 (Gene Therapy, Cell therapy, Vaccines and Biotherapeutics Immunogenicity) are published in volume 15 of Bioanalysis, issues 16 and 14 (2023), respectively.

Delirium is common after cardiac surgery and is associated with adverse outcomes. Administration of benzodiazepines before and after cardiac surgery is associated with delirium; guidelines recommend minimizing their use. Benzodiazepine administration during cardiac surgery remains common because of its recognized benefits. The B enzodiazepine -Free Cardiac Anesthesia for Reduction of Postoperative Delirium (B-Free) trial is a randomized cluster crossover trial evaluating whether an institutional policy of restricting intraoperative benzodiazepine administration (ie, ≥ 90% of patients do not receive benzodiazepines during cardiac surgery), as compared with a policy of liberal intraoperative benzodiazepine administration (ie, ≥ 90% of patients receive ≥ 0.03 mg/kg midazolam equivalent), reduces delirium. Hospitals performing ≥ 250 cardiac surgeries a year are included if their cardiac anesthesia group agrees to apply both benzodiazepine policies per their randomization, and patients are assessed for postoperative delirium every 12 hours in routine clinical care. Hospitals apply the restricted or liberal benzodiazepine policy during 12 to 18 crossover periods of 4 weeks each. Randomization for all periods takes place in advance of site startup; sites are notified of their allocated policy during the last week of each crossover period. Policies are applied to all patients undergoing cardiac surgery during the trial period. The primary outcome is the incidence of delirium at up to 72 hours after surgery. The B-Free trial will enroll ≥ 18,000 patients undergoing cardiac surgery at 20 hospitals across North America. Delirium is common after cardiac surgery, and benzodiazepines are associated with the occurrence of delirium. The B-Free trial will determine whether an institutional policy restricting the administration of benzodiazepines during cardiac surgery reduces the incidence of delirium after cardiac surgery. Clinicaltrials.gov registration number : NCT03928236 (First registered April 26, 2019)., (© 2023 The Authors.)

Purpose: To examine the life satisfaction outcomes after spinal cord injury (SCI) and to identify the factors associated with life satisfaction at 18 months and 10 years post-SCI in New Zealand (NZ)., Methods: Adults (16-64 years) were recruited between 2007 and 2009 from NZ's two spinal units following first admission for SCI. Interviews at 6 months, 18 months, and 10 years post-SCI examined demographic, physical, psychosocial, economic, and environmental characteristics. Multivariable regression models were used to identify predictors of life satisfaction at each timepoint., Results: Overall, 118 people participated at 6 months, 103 at 18 months, and 63 at 10 years post-SCI. Pre-SCI, 90% of participants were satisfied with life, 67% were satisfied at 18 months, and 78% at 10 years. At 18 months post-SCI, participants who reported: never or sometimes using a wheelchair, no problems with self-care, no problems with anxiety or depression, no/lesser disability, or fewer secondary health conditions (SHCs) at 6 months post-SCI were more likely to be satisfied (p < 0.05), compared to those without these characteristics. Participants who experienced considerable disability at 6 months post-SCI were 22% less likely to be satisfied 10 years post-SCI compared to those experiencing no/lesser disability (p = 0.028)., Conclusions: A higher proportion of participants were satisfied at both 18 months and 10 years post-SCI than not satisfied. To improve the likelihood of satisfaction with life, increased focus on reducing disability and providing supports for those using wheelchairs, experiencing anxiety/depression or problems with self-care, and effects of SHCs are promising for future potential interventions., (© 2023. The Author(s).)

Organoid cell culture methodologies are enabling the generation of cell models from healthy and diseased tissue. Patient-derived cancer organoids that recapitulate the genetic and histopathological diversity of patient tumours are being systematically generated, providing an opportunity to investigate new cancer biology and therapeutic approaches. The use of organoid cultures for many applications, including genetic and chemical perturbation screens, is limited due to the technical demands and cost associated with their handling and propagation. Here we report and benchmark a suspension culture technique for cancer organoids which allows for the expansion of models to tens of millions of cells with increased efficiency in comparison to standard organoid culturing protocols. Using whole-genome DNA and RNA sequencing analyses, as well as medium-throughput drug sensitivity testing and genome-wide CRISPR-Cas9 screening, we demonstrate that cancer organoids grown as a suspension culture are genetically and phenotypically similar to their counterparts grown in standard conditions. This culture technique simplifies organoid cell culture and extends the range of organoid applications, including for routine use in large-scale perturbation screens., (© 2022. The Author(s).)

In sweet cherry ( Prunus avium ), infection by ' Candidatus Phytoplasma pruni' results in small fruit with poor color and taste, rendering the fruit unmarketable. Yet the disease pathology is poorly understood, particularly at the cultivar level. Therefore, in this study we examined the physiological effects of Ca. P. pruni infection across a range of cultivars and locations in eastern Washington. We found that infection could be separated into early and established stages based on pathogen titer, which correlated with disease severity, including fruit size, color, and sugar and metabolite content. Furthermore, we observed that the effects of early-stage infections were largely indistinguishable from healthy, uninfected plants. Cultivar- and location-specific disease outcomes were observed with regard to size, color, sugar content, and citric acid content. This study presents the first in-depth assessment of X-disease symptoms and biochemical content of fruit from commercially grown sweet cherry cultivars known to be infected with Ca. P. pruni.

Purpose: Ponseti casting has universally been accepted as the gold standard for treatment of idiopathic clubfoot. Conversely, primary treatment for non-idiopathic clubfoot has not been established. The purpose of this study is to compare treatment outcomes following primary soft tissue release (STR) and Ponseti casting of non-idiopathic clubfoot. Methods: An IRB-approved retrospective study of patients treated for non-idiopathic clubfoot between 2005 and 2020 was conducted. Patients were included if they began treatment before the age of 2 and had at least 1 year of follow up. Patients were placed into either the STR group or Ponseti group and variables of interest were documented including reoccurrence of deformity, number of surgeries performed, type of surgeries performed, anesthesia time, and surgery time. Data was analyzed using Mann-Whitney U test for continuous variables. Results: A total of 33 children with 57 neuromuscular/syndromic clubfoot were identified of which 9 (15 feet) were treated with STR and 24 (42 feet) were treated with Ponseti casting. Average anesthesia and surgery time were found to be 291 and 179 min, respectively, for the STR group, and 113 and 67 min for the Ponseti group. The difference in operating time was determined to be significant ( p = 0.02, p = 0.01). Patients treated with STR were found to have significantly more surgeries performed over the course of treatment than those treated with Ponseti casting ( p = 0.001) with an average of 4.2 surgeries in the STR group and 1.5 surgeries in the Ponseti group. Extracapsular procedures were performed in 100% of the STR group and 97.6% of the Ponseti group ( p = 0.55). Intracapsular procedures were performed in 100% of the STR group and 50% of the Ponseti group ( p = 0.001). Conclusion: The Ponseti method should serve as the primary approach in the initial treatment of non-idiopathic clubfoot as it can reduce the risk of future invasive intracapsular surgery and shorten anesthesia and surgery times when surgical treatment is necessary. Level of Evidence: Level III retrospective case control study., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Abraham, Wall, Diab and Beaver.)

The 13th Global CRO Council (GCC) closed forum for bioanalysis was held in New Orleans, LA, USA on 5 April 2019. This GCC meeting was organized to discuss the contents of the 2019 ICH M10 Bioanalytical Method Validation Draft Guideline published in February 2019 and consolidate the feedback of the GCC members. While ICH M10 will cover requirements for reference standards, one of the biggest challenges facing the CRO community is the lack of consistency and completeness of Certificates of Analysis for reference standards used in regulated bioanalysis. Similar challenges exist with critical reagents (e.g., capture and detection antibodies) used for assays supporting biologics. The recommendations provided in this publication are the minimum requirements for the content that GCC members believe should be included in Certificates of Analysis for reference standards obtained from commercial vendors, sponsors and compendial suppliers, for use in regulated bioanalytical studies. In addition, recommendations for internal standards, metabolites and critical reagents are discussed.

Case: A 13-year, 6-month-old boy sustained a Delbet type III femoral neck fracture with postoperative femoral head avascular necrosis (AVN) and subsequent capital femoral physeal separation (CFPS). Preoperative angiography revealed a patent artery of the ligamentum teres to the femoral head epiphysis, allowing our patient to undergo a modified Dunn procedure to maintain this artery and preserve his native hip., Conclusion: Preoperative angiography allows for real-time identification of femoral head epiphyseal blood supply in patients with femoral head AVN complicated by CFPS and guides surgical treatment for hip preservation.

The primary objective of this work was to optimize red wine phenolic prediction with models built from wine ultraviolet-visible absorbance spectra. Three major obstacles were addressed to achieve this, namely algorithm selection, spectral multicollinearity, and phenolic evolution over time. For algorithm selection, support vector regression, kernel ridge regression, and kernel partial least squares regression were compared. For multicollinearity, the spectrum of malvidin chloride was used as an external standard for spectral adjustment. For phenolic evolution, spectral data were collected during fermentation as well as once a week for four weeks after fermentation had ended. Support vector regression gave the most accurate predictions among the three algorithms tested. Additionally, malvidin chloride proved a useful standard for phenolic spectral transformation and isolation. As for phenolic evolution, models needed to be calibrated and validated throughout the aging process to ensure predictive accuracy. In short, red wine phenolic prediction by the models built in this work can be realistically achieved, although periodic model re-calibration and expansion from data obtained using known phenolic assays is recommended to maintain model accuracy.

Introduction: Perioperative benzodiazepines are used because of their anxiolytic, sedative and amnestic effects. Evidence has demonstrated an association of benzodiazepines with adverse neuropsychiatric effects. Nonetheless, because of their potential benefits, perioperative benzodiazepines continue to be used routinely. We seek to evaluate the body of evidence of the risks and benefits of benzodiazepine use during the perioperative period., Methods and Analysis: We will search Cochrane CENTRAL, MEDLINE, EMBASE, PsychINFO, CINAHL and Web of Science from inception to March 2019 for randomised controlled trials (RCTs) and observational studies evaluating the administration of benzodiazepine medications as compared with all other medications (or nothing) in patients undergoing cardiac and non-cardiac surgery. We will exclude studies assessing the use of benzodiazepines for procedural sedation or day surgery. We will examine the impact of giving these medications before, during and after surgery. Outcomes of interest include the incidence of delirium, duration of delirium, postprocedure cognitive change, the incidence of intraoperative awareness, patient satisfaction/quality of life/quality of recovery, length-of-stay (LOS) in the intensive care unit (ICU), hospital LOS and in-hospital mortality.Reviewers will screen references and assess eligibility using predefined criteria independently and in duplicate. Two reviewers will independently collect data using prepiloted forms. We will present results separately for RCTs and observational studies. We will pool data using a random effect model and present results as relative risk with 95% CIs for dichotomous outcomes and mean difference with 95% CI for continuous outcomes. We will pool adjusted ORs for observational studies. We will assess risk of bias for individual studies using the Cochrane Collaboration tool for RCTs. For observational studies, we will use tools designed by the Clinical Advances through Research and Information Translation group. Quality of evidence for each outcome will be assessed using the Grading of Recommendations Assessment, Development and Evaluation approach., Ethics and Dissemination: This systematic review involves no patient contact and no interaction with healthcare providers or systems. As such, we did not seek ethics board approval. We will disseminate the findings of our systematic review through the presentation at peer-reviewed conferences and by seeking publication in a peer-reviewed journal., Prospero Registration Number: CRD42019128144., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2019. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

The 2019 13 th Workshop on Recent Issues in Bioanalysis (WRIB) took place in New Orleans, LA on 1-5 April 2019 with an attendance of over 1000 representatives from pharmaceutical/biopharmaceutical companies, biotechnology companies, contract research organizations and regulatory agencies worldwide. WRIB was once again a 5-day, week-long event - a full immersion week of bioanalysis, biomarkers, immunogenicity and gene therapy. As usual, it was specifically designed to facilitate sharing, reviewing, discussing and agreeing on approaches to address the most current issues of interest including both small- and large-molecule bioanalysis involving LCMS, hybrid LBA/LCMS, LBA cell-based/flow cytometry assays and qPCR approaches. This 2019 White Paper encompasses recommendations emerging from the extensive discussions held during the workshop, and is aimed to provide the bioanalytical community with key information and practical solutions on topics and issues addressed, in an effort to enable advances in scientific excellence, improved quality and better regulatory compliance. Due to its length, the 2019 edition of this comprehensive White Paper has been divided into three parts for editorial reasons. This publication (Part 2) covers the recommendations on the 2018 FDA BMV guidance, 2019 ICH M10 BMV draft guideline and regulatory agencies' input on bioanalysis, biomarkers, immunogenicity and gene therapy. Part 1 (Innovation in small molecules and oligonucleotides and mass spectrometry method development strategies for large molecules bioanalysis) and Part 3 (New insights in biomarker assay validation, current and effective strategies for critical reagent management, flow cytometry validation in drug discovery and development and CLSI H62, interpretation of the 2019 FDA immunogenicity guidance and gene therapy bioanalytical challenges) are published in volume 10 of Bioanalysis , issues 22 and 24 (2019), respectively.

The 12th GCC Closed Forum was held in Philadelphia, PA, USA, on 9 April 2018. Representatives from international bioanalytical Contract Research Organizations were in attendance in order to discuss scientific and regulatory issues specific to bioanalysis. The issues discussed at the meeting included: critical reagents; oligonucleotides; certificates of analysis; method transfer; high resolution mass spectrometry; flow cytometry; recent regulatory findings and case studies involving stability and nonclinical immunogenicity. Conclusions and consensus from discussions of these topics are included in this article.

Over the last decade, the use of biomarker data has become integral to drug development. Biomarkers are not only utilized for internal decision-making by sponsors; they are increasingly utilized to make critical decisions for drug safety and efficacy. As the regulatory agencies are routinely making decisions based on biomarker data, there has been significant scrutiny on the validation of biomarker methods. Contract research organizations regularly use commercially available immunoassay kits to validate biomarker methods. However, adaptation of such kits in a regulated environment presents significant challenges and was one of the key topics discussed during the 12th Global Contract Research Organization Council for Bioanalysis (GCC) meeting. This White Paper reports the GCC members' opinion on the challenges facing the industry and the GCC recommendations on the classification of commercial kits that can be a win-win for commercial kit vendors and end users.

Vincristine is a cytotoxic chemotherapy agent classified as an antitumor alkaloid and is part of the vinca alkaloid family. Vincristine's mechanism of action is to primarily inhibit mitosis of the cancer cell and is given by IV route only for treatment. Accidental intrathecal administration of vincristine has lethal consequences for patients. To minimize the risk of accidental intrathecal administration of vincristine, 14 infusion centers participated in a quality improvement project to change the practice of vincristine administration from IV push to IV piggyback via minibag and gravity. After three months, all infusion centers successfully implemented the practice.

The 2018 12 th Workshop on Recent Issues in Bioanalysis took place in Philadelphia, PA, USA on April 9-13, 2018 with an attendance of over 900 representatives from pharmaceutical/biopharmaceutical companies, biotechnology companies, contract research organizations and regulatory agencies worldwide. WRIB was once again a 5-day full immersion in bioanalysis, biomarkers and immunogenicity. As usual, it was specifically designed to facilitate sharing, reviewing, discussing and agreeing on approaches to address the most current issues of interest including both small- and large-molecule bioanalysis involving LCMS, hybrid LBA/LCMS and LBA/cell-based assays approaches. This 2018 White Paper encompasses recommendations emerging from the extensive discussions held during the workshop and is aimed to provide the bioanalytical community with key information and practical solutions on topics and issues addressed, in an effort to enable advances in scientific excellence, improved quality and better regulatory compliance. Due to its length, the 2018 edition of this comprehensive White Paper has been divided into three parts for editorial reasons. This publication (Part 3) covers the recommendations for large molecule bioanalysis, biomarkers and immunogenicity using LBA and cell-based assays. Part 1 (LCMS for small molecules, peptides, oligonucleotides and small molecule biomarkers) and Part 2 (hybrid LBA/LCMS for biotherapeutics and regulatory agencies' inputs) are published in volume 10 of Bioanalysis , issues 22 and 23 (2018), respectively.

Background: Genome editing by CRISPR-Cas9 technology allows large-scale screening of gene essentiality in cancer. A confounding factor when interpreting CRISPR-Cas9 screens is the high false-positive rate in detecting essential genes within copy number amplified regions of the genome. We have developed the computational tool CRISPRcleanR which is capable of identifying and correcting gene-independent responses to CRISPR-Cas9 targeting. CRISPRcleanR uses an unsupervised approach based on the segmentation of single-guide RNA fold change values across the genome, without making any assumption about the copy number status of the targeted genes., Results: Applying our method to existing and newly generated genome-wide essentiality profiles from 15 cancer cell lines, we demonstrate that CRISPRcleanR reduces false positives when calling essential genes, correcting biases within and outside of amplified regions, while maintaining true positive rates. Established cancer dependencies and essentiality signals of amplified cancer driver genes are detectable post-correction. CRISPRcleanR reports sgRNA fold changes and normalised read counts, is therefore compatible with downstream analysis tools, and works with multiple sgRNA libraries., Conclusions: CRISPRcleanR is a versatile open-source tool for the analysis of CRISPR-Cas9 knockout screens to identify essential genes.

The 11th Global CRO Council Closed Forum was held in Universal City, CA, USA on 3 April 2017. Representatives from international CRO members offering bioanalytical services were in attendance in order to discuss scientific and regulatory issues specific to bioanalysis. The second CRO-Pharma Scientific Interchange Meeting was held on 7 April 2017, which included Pharma representatives' sharing perspectives on the topics discussed earlier in the week with the CRO members. The issues discussed at the meetings included cumulative stability evaluations, matrix stability evaluations, the 2016 US FDA Immunogenicity Guidance and recent and unexpected FDA Form 483s on immunogenicity assays, the bioanalytical laboratory's role in writing PK sample collection instructions, biosimilars, CRO perspectives on the use of chiral versus achiral methods, hybrid LBA/LCMS assays, applications of fit-for-purpose validation and, at the Global CRO Council Closed Forum only, the status and trend of current regulated bioanalytical practice in China under CFDA's new BMV policy. Conclusions from discussions of these topics at both meetings are included in this report.

The 2017 11th Workshop on Recent Issues in Bioanalysis took place in Los Angeles/Universal City, California, on 3-7 April 2017 with participation of close to 750 professionals from pharmaceutical/biopharmaceutical companies, biotechnology companies, contract research organizations and regulatory agencies worldwide. WRIB was once again a 5-day, week-long event - a full immersion week of bioanalysis, biomarkers and immunogenicity. As usual, it was specifically designed to facilitate sharing, reviewing, discussing and agreeing on approaches to address the most current issues of interest including both small- and large-molecule analysis involving LC-MS, hybrid ligand-binding assay (LBA)/LC-MS and LBA approaches. This 2017 White Paper encompasses recommendations emerging from the extensive discussions held during the workshop, and is aimed to provide the bioanalytical community with key information and practical solutions on topics and issues addressed, in an effort to enable advances in scientific excellence, improved quality and better regulatory compliance. Due to its length, the 2017 edition of this comprehensive White Paper has been divided into three parts for editorial reasons. This publication (Part 3) covers the recommendations for large-molecule bioanalysis, biomarkers and immunogenicity using LBA. Part 1 (LC-MS for small molecules, peptides and small molecule biomarkers) and Part 2 (hybrid LBA/LC-MS for biotherapeutics and regulatory agencies' inputs) are published in volume 9 of Bioanalysis, issues 22 and 23 (2017), respectively.

Loss of skeletal muscle mass and function occurs with increasing age. Calorie restriction (CR) increases the lifespan of C57Bl/6 mice, but not in the shorter-lived DBA/2 strain. There is some evidence that calorie restriction reduces or delays many of the age-related defects that occur in rodent skeletal muscle. We therefore investigated the effect of short (2.5 month) and longer term (8.5 and 18.5 months) CR on skeletal muscle in male and female C57Bl/6 and DBA/2 mice. We found that short-term CR increased the satellite cell number and collagen VI content of muscle, but resulted in a delayed regenerative response to injury.Consistent with this, the in vitro proliferation of satellite cells derived from these muscles was reduced by CR. The percentage of stromal cells, macrophages, hematopoietic stem cells and fibroadipogenic cells in the mononucleated cell population derived from skeletal muscle was reduced by CR at various stages. But overall, these changes are neither consistent over time, nor between strain and sex. The fact that changes induced by CR do not persist with time and the dissimilarities between the two mouse strains, combined with sex differences, urge caution in applying CR to improve skeletal muscle function across the lifespan in humans.

The 10th Global CRO Council (GCC) Closed Forum was held in Orlando, FL, USA on 18 April 2016. In attendance were decision makers from international CRO member companies offering bioanalytical services. The objective of this meeting was for GCC members to meet and discuss scientific and regulatory issues specific to bioanalysis. The issues discussed at this closed forum included reporting data from failed method validation runs, GCP for clinical sample bioanalysis, extracted sample stability, biomarker assay validation, processed batch acceptance criteria, electronic laboratory notebooks and data integrity, Health Canada's Notice regarding replicates in matrix stability evaluations, critical reagents and regulatory approaches to counteract fraud. In order to obtain the pharma perspectives on some of these topics, the first joint CRO-Pharma Scientific Interchange Meeting was held on 12 November 2016, in Denver, Colorado, USA. The five topics discussed at this Interchange meeting were reporting data from failed method validation runs, GCP for clinical sample bioanalysis, extracted sample stability, processed batch acceptance criteria and electronic laboratory notebooks and data integrity. The conclusions from the discussions of these topics at both meetings are included in this report.

A nursing quality consortium standardized nursing practice across 17 independently functioning ambulatory oncology sites. Programs were developed to validate both competences and competencies. One program assessed nine competences needed to develop systems of care to detect and treat treatment-related side effects. A second program was developed to assess competencies needed to prevent harm to oncology patients. This manuscript describes a successful approach to standardizing nursing practice across geographically distant academic and community sites.

Workplace fatigue is common among occupations that have prolonged work hours, rotating shifts, night-time work hours, inadequate time for rest during work, and insufficient time for recovery between shifts. Available evidence suggests that workplace fatigue poses a substantial threat to patient safety and contributes to worker injury and decreased vigilance. However, little is known about workplace fatigue among nursing personnel working in institutions dedicated solely to the care of patients with cancer. This study describes the scope and severity of workplace fatigue among nursing personnel working in the inpatient and ambulatory care divisions of a comprehensive cancer center.

N-acylphosphatidylethanolamines (NAPEs) are naturally occurring derivatives of phosphatidylethanolmine (PE) in which the PE amino group is attached to an acyl chain. Given their occurrence in natural systems, there is interest in knowing the effect of NAPEs on membrane dynamic structure and function. This study examines the ability of NAPEs to affect the association of the cytochrome c and Zn-heme cytochrome c with the surface of bilayer membranes. Fluorescence titration experiments show that cationic cytochrome c has the same high affinity for the surfaces of anionic vesicles that are rich in NAPEs or diplalmitoyphosphatidylglycerol (DPPG) but the protein/membrane interaction in each case is quite different. Cytochrome c adsorption to DPPG membranes is relatively irreversible due to the DPPG molecules adopting an extended conformation that promotes strong hydrophobic contact with the adsorbed protein. In contrast, cytochrome c association with N-acyl DPPE membranes is due primarily to reversible electrostatic interactions with the anionic headgroup, and not hydrophobic contact with the N-acyl chain. The presence of a small mole fraction of an N-propionyl derivative of DPPE (N-C3:0-DPPE) diminishes cytochrome c affinity for vesicles containing a large amount of DPPG apparently by relieving the membrane packing strain that drives the extended DPPG conformation.

The L1 Global Harmonization Team provides recommendations specifically for run acceptance of ligand binding methods used in bioanalysis of macromolecules in support of pharmacokinetics. The team focused on standard curve calibrators and quality controls for use in both pre-study validation and in-study sample analysis, including their preparation and acceptance criteria. The team also considered standard curve editing and the concept of total error.

The 2014 8th Workshop on Recent Issues in Bioanalysis (8th WRIB), a 5-day full immersion in the evolving field of bioanalysis, took place in Universal City, California, USA. Close to 500 professionals from pharmaceutical and biopharmaceutical companies, contract research organizations and regulatory agencies worldwide convened to share, review, discuss and agree on approaches to address current issues of interest in bioanalysis. The topics covered included both small and large molecules, and involved LCMS, hybrid LBA/LCMS, LBA approaches and immunogenicity. From the prolific discussions held during the workshop, specific recommendations are presented in this 2014 White Paper. As with the previous years' editions, this paper acts as a practical tool to help the bioanalytical community continue advances in scientific excellence, improved quality and better regulatory compliance. Due to its length, the 2014 edition of this comprehensive White Paper has been divided into three parts for editorial reasons. This publication (Part 2) covers the recommendations for Hybrid LBA/LCMS, Electronic Laboratory Notebook and Regulatory Agencies' Input. Part 1 (Small molecules bioanalysis using LCMS) was published in the Bioanalysis issue 6(22) and Part 3 (Large molecules bioanalysis using LBA and Immunogenicity) will be published in the Bioanalysis issue 6(24).

Study Design: Longitudinal cohort study., Objectives: To estimate socioeconomic and work outcomes over 2 and a half years following spinal cord injury (SCI), and to compare those in receipt of compensation (Accident Compensation Corporation, ACC) and those not., Setting: People admitted to the two spinal units in 2007-2009 in New Zealand, where there is a unique no-fault compensation scheme for injury., Methods: Interviews were conducted at ∼6, 18 and 30 months after SCI and data collected on pre-SCI and post-SCI health and socioeconomic characteristics. Poisson regression, quantile regression and a linear mixed model regression were used to compare differences in outcomes., Results: Of the 162 eligible people, 118 (73%) participated and 91(77%) were followed to 30 months; 79% received ACC. Median personal income, self-reported standard of living and household income adequacy all fell slightly to 18 months and then stabilized at 30 months. At that time, 49% had returned to paid work. Among those not eligible for ACC, income fell to less than half the ACC group (P<0.006 after adjustment), and return to work was lower (29% versus 54%)., Conclusion: The findings that most people retained their economic status and that return to work was relatively high appear to be due to the proportion entitled to the ACC no-fault compensation scheme for injury; with earnings-related compensation, a focus on rehabilitation to work and non-means-tested support services. This situation should mitigate against the downward spiral into poverty and further ill-health.

This paper estimates the incidence (all ages) of spinal cord neurological impairment (SCI; traumatic and non-traumatic) in New Zealand and describes pre-SCI characteristics and early post-SCI outcomes for participants (16-64 years) in this longitudinal study. Demographic and clinical data on all people admitted to New Zealand's two spinal units (mid-2007 to mid-2009) were included for the estimate of incidence. Participants in this longitudinal study were asked at first interview about pre-SCI socio-demographic, health and behavioural characteristics, and about post-SCI symptoms, general health status (EQ-5D) and disability (WHODAS 12-item). Age-adjusted incidence rates (95% CI) for European, Māori, Pacific and 'Other' ethnicities were 29 (24-34), 46 (30-64), 70 (40-100) and 16 (9-22) per million, respectively. Interviews with 118 (73%) participants (16-64 years), occurred 6.5 months post-SCI. Most reported bother with symptoms, and problems with health status and disability. Compared with Europeans, the incidence of SCI is high among Māori and particularly high among Pacific people. Six months after SCI, proximate to discharge from the spinal units, considerable symptomatic, general health and disability burden was borne by people with SCI.

This paper reviews what is known about the challenges of implementing quality improvement programs and draws on data from a systematic continuous quality improvement (CQI) project in remote communities in Australia and Fiji, known as Audit and Best practice for Chronic Disease, to synthesise lessons and discuss the potential for broader application in low and middle income countries, including Pacific Island countries and territories. Although a number of systematic reviews have indicated that quality improvement programs can be effective in changing professional practice and improving the quality of care and patient outcomes, little is known about the key ingredients for change or how services use and implement different strategies to achieve improvements. We identify key features of an innovative CQI model and factors related to implementation that support improvement in diabetes service delivery and intermediate outcomes. Requirements for supporting CQI are identified and the potential for wider application discussed. It is argued that the participatory action research approach supports innovation and broad-based change and the evidence it has produced extends the current knowledge base and facilitates the translation of knowledge into action, for both policy and practice., (© 2011 The Authors. Australian Journal of Rural Health © National Rural Health Alliance Inc.)

Unlabelled: Pain management is often described as a component of child life specialists' work. No research has described the specific pain management strategies used by child life specialists. The objectives of this study were to determine child life specialists' use of nonpharmacological strategies, to describe the perceived efficacy of these strategies, to determine how much training child life specialists had in these various strategies, and to determine what demographic characteristics predict the use of evidence-based techniques. Six hundred seven child life specialists from hospitals and health centers across North America responded to an online survey (response rate: 85.4%). Results indicate that child life specialists use a variety of techniques with varying degrees of perceived efficacy. The most commonly endorsed techniques were providing information/preparation, comforting/reassurance, and positive reinforcement. Respondents reported receiving substantial training in some techniques (eg, providing information/preparation, medical play) and high interest in receiving additional training in all techniques. Certification status, the proportion of patients for whom participants reported providing pain management services, and participants' perceived levels of knowledge and skill emerged as significant predictors of the use of evidence-based strategies. The results of this survey suggest that child life specialists are actively involved in pediatric pain management., Perspective: American and Canadian child life specialists were surveyed to assess their involvement in managing the pain of pediatric patients. Findings of the survey indicate that child life specialists are involved in the management of pediatric pain and are receptive to additional training in evidence-based techniques.

Objective: This paper provides an overview of the services developed in response to the unique mental health needs of a remote Aboriginal community. We describe an evolving service on the Tiwi Islands in the Northern Territory and the challenges that need to be addressed if the community is to continue to take a leading role in dealing with mental health issues., Conclusions: The Tiwi Mental Health Service demonstrates that community members are able to identify needs and respond accordingly if they are provided with the relevant information and supported in their decision-making process. The establishment of social governance mechanisms and the long-term commitment by a change agent to facilitate the empowerment process are important keys to success. The main challenge in establishing services in rural Aboriginal communities is to identify and support community strengths, including leaders and cultural practices.

Objective: To compare hospital costs of Aboriginal and non-Aboriginal patients having haemodialysis treatment and forecast the future treatment cost., Methods: The costs of patients with HD in the "Top End" of Australia's Northern Territory were estimated for the financial years 1996/97 and 1997/98 using a hospital costing model. We used an Autoregression Integrated Moving Average model to predict future demand., Results: 165 patients (101 Aboriginal and 64 non-Aboriginal) were treated at a total cost of $12.4 million in this two-year period. These 165 patients represented 0.7% of inpatients, 8.8% of total inpatient costs and 31.6% of total inpatient episodes of care in the Top End region. $9.5 million (77%) was spent on routine haemodialysis treatment and $2.9m (23%) on other hospitalisations. The average cost per routine haemodialysis treatment over the two-year period was $527, or $78 600 per patient treatment year. Hospitalisations for comorbidities occurred in 86% of Aboriginal and 39% of non-Aboriginal patients. Average cost per patient, number of admissions and length of hospital stays were all significantly greater for Aboriginals. We predict an average increase in the number of treatments of 12% each year over the next five years and a five-year cost of $49.8m., Conclusions: A multipronged strategy designed to reduce the prevalence and costs of renal failure is required.

This article is concerned with the methodological issues of assessing the effects of casemix funding on hospital utilisation. Time-series analysis and intervention analysis are proposed to ascertain the effects. It was found there had been a decline in average length of stay and number of bed-days, an increase in weighted separations for teaching and non-teaching hospitals, and no apparent increase of costliness in terms of a comprehensive casemix index. No evidence of decline in quality of care can be established in terms of readmission rates. The long-term effects of casemix funding, and specific issues in terms of the funding model used, patients and cost shifting between hospital services and community health services, remain to be studied.

The Northern Territory intends to make use of Australian National Diagnosis Related Groups (DRGs) and their cost relativities as the basis for the allocation of budgets among public hospitals. The study reported here attempted to assess the extent to which there are variations in severity of illness and socio-economic status which are not adequately explained by DRG alone and, if so, to develop a DRG payment adjustment index by use of routinely available data items. The investigation was undertaken by use of a database containing all discharges between July 1992 and June 1995. Hospital length of stay was used as a proxy for cost. Multivariate analysis was undertaken and it was found that several variables were associated with cost variations within DRGs. Stepwise multiple linear regression was used to develop a model in which 14 variables were able to explain 45% of the variations. Index values were subsequently computed from the regression model for each of eight categories of admitted patient episodes which are the intersections of three binary variables: Aborigine or non-Aborigine, rural or urban usual place of residence of the patient and hospital type (teaching or other). It is intended that these index values will be used to compute differential funding rates for each hospital in the Territory.

Wide differentials continue to exist in mortality rates and other health outcomes between Aboriginal and non-Aboriginal Australians. In the Northern Territory (NT), where Aborigines make up 24% of the population, the all-causes age-adjusted Standardised Mortality Ratio for Aborigines compared to non-Aborigines has remained above 3 since the late 1970s, with significant regional variations. During 1995 an expenditure analysis was undertaken for primary health care (PHC) services in different regions of the NT and compared to mortality ratios. At the same time a method for needs-based funding was being developed which could replace the existing historical funding arrangements. In the first instance, the application of a simplified version of this Resource Allocation Formula (RAF) resulted in a significant shift of resources for new prevention program funding to regions of relatively high mortality and low per capita PHC expenditure. However, developing RAFs to redistribute at the margin within the NT is likely to generate further inequities between losing NT programs and counterparts in other states. If outcomes-based resource allocation is to be meaningful nationally, the reference point for the RAF should be national average PHC expenditure rather than existing state averages. There is a need for a combined approach to outcomes-based planning which takes into account both the equity arguments of resource allocation models and efficacy arguments to maximise health gains. Some of these arguments are explored in this paper.

In the 77-year-old woman described, atherectomy of a circumflex artery with the Rotablator device was complicated by "no reflow." Ten days later, a 75% right coronary artery stenosis was successfully managed by balloon angioplasty. On the following day, acute closure of this vessel resulted in death. Gross examination of the heart showed features of recent posterior infarction, and microscopic study revealed atheroemboli in myocardial arterioles. We conclude that high-speed pulverization of atherosclerotic plaque can cause clinically significant emboli.