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Proteomics by mass spectrometry (MS) allows for the identification of amino acid/peptide sequences in complex mixtures. Peptide analysis and quantitation enables screening of protein biomarkers and targeted protein biomarker analysis for clinical applications. Whereas miniature mass spectrometers have primarily demonstrated point-of-care analyses with simple procedures aiming at drugs and lipids, it would be interesting to explore their potential in analyzing proteins and peptides. In this work, we adapted a miniature MS instrument for peptide analysis. A mass range as wide as 100-2000 m/z was achieved for obtaining peptide spectra using this instrument with dual linear ion traps. MS2 and MS3 can be performed to analyze a wide range of peptides. The parameters of pressure, electric potentials, and solution conditions were optimized to analyze peptides with molecular weights between 900 and 1800 Da. The amino acid sequences were identified using both beam-type and in-trap collision-induced dissociation, and the results were comparable to those obtained by a commercial quadrupole time-of-flight mass spectrometer. With product ion monitoring scan mode, peptide quantitation was performed with a limit of detection of 20 nM achieved for the Met peptide. The method developed has also been applied to the analysis of the trypsin-digested cell lysate of SKBR3 cells with a low expression level of the Met gene.

Antibodies against posttranslational modifications (PTMs) such as lysine acetylation, ubiquitin remnants, or phosphotyrosine have resulted in significant advances in our understanding of the fundamental roles of these PTMs in biology. However, the roles of a number of PTMs remain largely unexplored due to the lack of robust enrichment reagents. The addition of N-acetylglucosamine to serine and threonine residues (O-GlcNAc) by the O-GlcNAc transferase (OGT) is a PTM implicated in numerous biological processes and disease states but with limited techniques for its study. Here, we evaluate a new mixture of anti-O-GlcNAc monoclonal antibodies for the immunoprecipitation of native O-GlcNAcylated peptides from cells and tissues. The anti-O-GlcNAc antibodies display good sensitivity and high specificity toward O-GlcNAc-modified peptides and do not recognize O-GalNAc or GlcNAc in extended glycans. Applying this antibody-based enrichment strategy to synaptosomes from mouse brain tissue samples, we identified over 1300 unique O-GlcNAc-modified peptides and over 1000 sites using just a fraction of sample preparation and instrument time required in other landmark investigations of O-GlcNAcylation. Our rapid and robust method greatly simplifies the analysis of O-GlcNAc signaling and will help to elucidate the role of this challenging PTM in health and disease., Graphical Abstract, Highlights • Anti-O-GlcNAc antibodies are fast and simple enrichment reagents. • Anti-O-GlcNAc antibodies are sensitive and achieve significant depth of coverage. • Anti-O-GlcNAc antibodies are specific for singular O-GlcNAc modifications. • Anti-O-GlcNAc antibody enrichment techniques can be applied to cells and tissues. • HCD product-triggered EThcD data acquisition improves depth of coverage., In Brief O-GlcNAc, the single N-acetylglucosamine coupled to serine and threonines of nucleocytoplasmic proteins, is an enigmatic posttranslational modification implicated in most cellular functions. Methods to study the O-GlcNAc modified proteome have been challenging, in part, due to limitations in its enrichment from native tissue. Here, we characterize novel antibodies for the enrichment of native O-GlcNAc modified peptides. Our enrichment strategy shows strong sensitivity and excellent specificity for O-GlcNAc. We expect these reagents to be a significant advancement for the field.

Purpose The main objective of this study was to investigate abuse of residents with either dementia or Alzheimer's disease in long-term care settings, to identify facilitators and barriers surrounding implementation of systems to prevent such occurrences, and to draw conclusions on combating the issue of abuse. Patients and methods A systematic review was conducted using the Medline, CINAHL, and Academic Search Ultimate databases. With the use of key terms via Boolean search, 30 articles were obtained which were determined to be germane to research objectives. The review was conducted and structured based on Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. Results Residents with dementia or Alzheimer's disease are at greater risk of abuse. The growing population could increase this problem exponentially. The most common facilitators were the introduction of policies/programs in the facility, education, and working conditions. The most cited barriers were poor training, lack of research, and working conditions in the long-term care setting. Conclusion The examples given would be useful in minimizing the potential for abuse in the long-term care setting. Leadership can take an active role in the prevention of abuse of the elderly through their actions, education of employees, and changes in the work environment.

Background Gram-negative organisms (GNOs) have increasing resistance rates to levofloxacin at Virginia Commonwealth University Health System (VCUHS), where levofloxacin is the most common agent added to provide double coverage of gram-negative infections. The goal of this study was to determine the adequacy of empiric gram-negative coverage for septic patients at our institution. Methods A retrospective review of patients admitted to VCUHS, from January 1, 2014, to December 31, 2014, with a diagnosis of sepsis, severe sepsis, or septic shock and documented infection, was performed to determine the adequacy of various empiric antibiotic combinations. Results Of 219 patients who met the inclusion criteria, 56% of patients received monotherapy and 21% of patients received combination therapy (2 antibiotics) covering GNOs. GNOs (84%) were susceptible to piperacillin-tazobactam. When used in combination with cefepime and meropenem, levofloxacin did not increase coverage. However, levofloxacin provided an 8% increase in coverage and gentamicin provided an additional 13% increase in coverage, respectively, when used in combination with piperacillin-tazobactam. Conclusions Among septic patients at VCUHS, gentamicin provided increased gram-negative coverage when compared with levofloxacin. Although susceptibility to piperacillin-tazobactam alone was relatively low, the combination of piperacillin-tazobactam and gentamicin provided nearly equivalent coverage to meropenem and gentamicin.

While proteins can often be stabilized by maintaining them in the frozen state, water-to-ice transformation can also lead to degradation of protein molecules. A new method to study protein/ice interaction is presented herein, which is based on measuring the characteristic features of X-ray diffraction (XRD) patterns of hexagonal ice (Ih). Aqueous solutions of four different proteins and several small molecular weight solutes are studied using high-resolution synchrotron X-ray diffraction at the ID22 beamline at the European Synchrotron Radiation Facility. The beamline is optimized to eliminate the instrumental broadening of diffraction lines and reduce the preferred orientation effects, thereby enabling quantitative analysis of the XRD data. The analysis demonstrates that two pharmaceutical proteins, recombinant human albumin (rHA) and monoclonal antibody (mAb), have a pronounced effect on the properties of ice crystals. In particular, the size of the crystalline domains is significantly smaller, and the microstrain is larger, in the solutions of the pharmaceutical proteins, when compared with a model protein (lysozyme), an antifreeze protein, and sucrose and histidine. Neither of the proteins studied exhibit preferred interaction with specific crystalline faces of Ih. The results are consistent with indirect interaction of the pharmaceutical proteins with ice, in which protein molecules are accumulated in the quasi-liquid layer next to growing ice crystallization front. Direct interaction would indicate a sorption of protein molecules on ice crystals, whereas “indirect interaction” terminology is used to describe any interference of proteins with ice crystals without sorption involved. Lysozyme molecules, on the other hand, do not exhibit any evidence of interaction (either direct or indirect) with ice crystals. This is the first report, to the best of our knowledge, of major difference in protein/ice interaction between different types of non-antifreeze proteins. In addition, we report an unexpected finding of a second population of ice crystals, with a much smaller (a few nm) size of crystalline domains. The second (minor) population is tentatively identified as a high-pressure form of ice, possibly IceIII or IceIX. This observation highlights a potential role of mechanical stresses and local pressure in freeze-induced destabilization of proteins.

Antibodies against post-translational modifications (PTMs) such as lysine acetylation, ubiquitin remnants, or phosphotyrosine have resulted in significant advances in our understanding of the fundamental roles of PTMs in biology. However, the roles of a number of PTMs remain largely unexplored due to the lack of robust enrichment reagents. The addition of N-acetylglucosamine to serine and threonine residues (O-GlcNAc) by the O-GlcNAc transferase (OGT) is a PTM implicated in numerous biological processes and disease states but with limited techniques for its study. Here, we evaluate a new mixture of anti-O-GlcNAc monoclonal antibodies for the immunoprecipitation of native O-GlcNAcylated peptides from cells and tissues. The anti-O-GlcNAc antibodies display good sensitivity and high specificity toward O-GlcNAc-modified peptides, and do not recognize O-GalNAc or GlcNAc in extended glycans. Applying this antibody-based enrichment strategy to synaptosomes from mouse brain tissue samples, we identified over 1,300 unique O-GlcNAc-modified peptides and over 1,000 sites using just a fraction of sample preparation and instrument time required in other landmark investigations of O-GlcNAcylation. Our rapid and robust method greatly simplifies the analysis of O-GlcNAc signaling and will help to elucidate the role of this challenging PTM in health and disease.

BackgroundLysine acylations are important post-translational modifications that are present in both eukaryotes and prokaryotes, regulating diverse cellular functions. Our knowledge of the microbiome lysine acylation remains limited due to the lack of efficient analytical and bioinformatics methods for complex microbial communities.ResultsWe show that serial enrichment using motif antibodies successfully captures peptides containing lysine acetylation, propionylation and succinylation from human gut microbiome samples. A new bioinformatic workflow consisting of unrestricted database search confidently identified >60,000 acetylated, and ~20,000 propionylated and succinylated gut microbial peptides. Characterization of these identified modification-specific metaproteomes, i.e. meta-PTMomes, demonstrates that lysine acylations are differentially distributed in microbial species with different metabolic capabilities.ConclusionThis study provides an analytical framework, consisting of a serial immunoaffinity enrichment and an open database search strategy, for the study of lysine acylations in microbiome, which enables functional microbiome studies at the post-translational level.

Background: Data regarding outpatient antibiotic prescribing for urinary tract infections (UTIs) are limited, and they have never been formally summarized in Virginia. Objective: We describe outpatient antibiotic prescribing trends for UTIs based on gender, age, geographic region, insurance payer and International Classification of Disease, Tenth Revision (ICD-10) codes in Virginia. Methods: We used the Virginia All-Payer Claims Database (APCD), administered by Virginia Health Information (VHI), which holds data for Medicare, Medicaid, and private insurance. The study cohort included Virginia residents who had a primary diagnosis of UTI, had an antibiotic claim 0–3 days after the date of the diagnosis and who were seen in an outpatient facility in Virginia between January 1, 2016, and December 31, 2016. A diagnosis of UTI was categorized as cystitis, urethritis or pyelonephritis and was defined using the following ICD-10 codes: N30.0, N30.00, N30.01, N30.9, N30.90, N30.91, N39.0, N34.1, N34.2, and N10. The following antibiotics were prescribed: aminoglycosides, sulfamethoxazole/trimethoprim (TMP-SMX), cephalosporins, fluoroquinolones, macrolides, penicillins, tetracyclines, or nitrofurantoin. Patients were categorized based on gender, age, location, insurance payer and UTI type. We used χ2 and Cochran-Mantel-Haenszel testing. Analyses were performed in SAS version 9.4 software (SAS Institute, Cary, NC). Results: In total, 15,580 patients were included in this study. Prescriptions for antibiotics by drug class differed significantly by gender (P < .0001), age (P < .0001), geographic region (P < .0001), insurance payer (P < .0001), and UTI type (P < .0001). Cephalosporins were prescribed more often to women (32.48%, 4,173 of 12,846) than to men (26.26%, 718 of 2,734), and fluoroquinolones were prescribed more often to men (53.88%, 1,473 of 2,734) than to women (47.91%, 6,155 of 12,846). Although cephalosporins were prescribed most frequently (42.58%, 557 of 1,308) in northern Virginia, fluoroquinolones were prescribed the most in eastern Virginia (50.76%, 1677 of 3,304). Patients with commercial health insurance, Medicaid, and Medicare were prescribed fluoroquinolones (39.31%, 1,149 of 2,923), cephalosporins (56.33%, 1,326 of 2,354), and fluoroquinolones (57.36%, 5,910 of 10,303) most frequently, respectively. Conclusions: Antibiotic prescribing trends for UTIs varied by gender, age, geographic region, payer status and UTI type in the state of Virginia. These data will inform future statewide antimicrobial stewardship efforts. Funding: None Disclosures: Michelle Doll reports a research grant from Molnlycke Healthcare.

We describe traditional antimicrobial stewardship program (ASP) activities with a discussion of how these activities can be refocused in the setting of the COVID-19 pandemic. Additionally, we discuss possible adverse consequences of ASP attention diversion on COVID-19 response efforts and overall implications for future pandemic planning. We also discuss ASP in collaboration with other groups within health systems and how COVID-19 may affect these relationships long term. Despite the paucity of literature on Antimicrobial Stewardship and COVID-19, the potential contributions of ASPs during a pandemic are numerous. ASPs can develop strategies to identify patients with COVID-19-like-illness; this is particularly useful when these patients are missed at the time of health system entry. ASPs can also play a critical role in the management of potential drug shortages, developing local treatment guidelines, optimizing the use of antibiotics, and in the diagnostic stewardship of COVID-19 testing, among other roles. Importantly, it is often difficult to ascertain whether critically ill patients who are hospitalized with COVID-19 have concurrent or secondary bacterial infections-ASPs are ideally situated to help optimize antimicrobial use for these patients via a variety of mechanisms. ASPs are uniquely positioned to aid in pandemic response planning and relief efforts. ASPs are already integrated into health systems and play a key role in optimizing antimicrobial prescribing. As ASPs assist in COVID-19 response, understanding the role of ASPs in pandemic relief efforts may mitigate damage from future outbreaks.

Intelligent use of rural residential land and sustainable construction is inexorably linked to cost, however, options exist that are eco-friendly and have a positive return on investment. In 2011, a research residence was built to evaluate various land-use and sustainable components. This Texas house has subsequently been used for both residential and research purposes. The purpose of this case study was to evaluate break-even construction considerations, to assess environmental impacts, and to evaluate qualitatively efficacy of sustainable options incorporated in the research residence. Some of the specific components discussed are home site placement (directional positioning), materiel acquisition (transportation), wood product minimization, rainwater harvesting, wastewater management, grid-tied solar array power, electric car charging via a solar array, geothermal heating and cooling, insulation selection, windows, fixtures, and appliance selection, and on-demand electric water heaters for guest areas. This study seeks to identify the impact of proper land use and sustainable techniques on the environment and return-on-investment in rural areas. Break-even and 15-year Net Present Value (NPV) analysis at 3% and 5% cost of capital were used to evaluate traditional construction, partially sustainable construction, and fully sustainable construction options for the case study house, which was built sustainably. The additional cost of sustainable construction is estimated at $54,329. At 3%, the analysis suggests a 15-year NPV of $334,355 (traditional) versus $250,339 million (sustainable) for a difference of $84K. At 5% cost of capital, that difference falls to $63K. The total estimated annual difference in carbon emissions is 4.326 million g/CO2e for this research residence. The results indicate that good choices for quick return-on-investment in rural construction would be the use of engineered lumber, Icynene foam, and Energy Star windows and doors. Medium-term options include photovoltaic systems (PVS) capable of powering the home and an electric car. Sustainable construction options should positively affect the environment and the pocketbook. Regulations and code should require adoption of short-range, break-even sustainable solutions in residential construction.

major consideration for consumers and the residential construction industry is the cost&ndash, benefit and break-even of various sustainable construction options. This research provides apublicly available simulation that allows users to compare baseline construction options versussustainable options and evaluates both break-even costs as well as environmental effects. This R ShinyMonte Carlo simulation uses common pseudo-random number streams for replicability and includesoptions for solar, rainwater harvesting, wells, Icynene foam, engineered lumber, Energy Star windowsand doors, low flow fixtures, aerobic/non-aerobic/city waste treatment, electric versus gasolinevehicles, and many other options. This is the first simulation to quantify multiple sustainableconstruction options, associated break-even points, and environmental considerations for public use.Using user default parameters, coupled with a 100% solar solution for a baseline 3,000 square foot/279square meter house with 2 occupants results in a break-even of 9 years. Results show that many of thesustainable options are both green for the environment and green for the pocketbook.

The question of building sustainable in a geographical locality is inexorably linked to cost. In 2011, one of the authors built a sustainable house that was (at the time) the highest certified sustainable home based on the National Association of Home Builder’s standards for sustainable construction. This Texas house has been used for residential and research purposes for the past decade. In this case study, the authors evaluate components of the construction and their effectiveness as well as unseen secondary and tertiary effects. Some of the specific components discussed are home site placement; rainwater harvesting (100% of residential requirements); aerobic septic system; grid-tied solar array power; electric car charging; geothermal heating and cooling; reclaimed wood framing; spray foam installation; selection of windows, fixtures, and appliances; on-demand electric water heaters for guest areas; generator backups; and use of local items. Electric bills and water system improvements are discussed in detail, as improvements were made as part of residential and research requirements. This case study suggests that the financial outlay is worth the extra up-front costs if residents in this geographical area and climate will occupy the residence 7 years.

The recently described proportion of carbapenem consumption metric was used to assess the effectiveness of formulary restriction for carbapenems for 2 units housing predominantly immunocompromised patients at a large academic medical center. Interrupted time series analysis revealed a significant decrease in meropenem use for hematology-oncology and bone marrow transplant units after restriction.

The impact of formulary restriction and preauthorization (FRPA) on prescribing trends was examined over a 5-year period at an academic medical center. Ordinary least squares regression was used to identify hospital units demonstrating statistically significant trends in prescription of restricted agents. Significant decreases in restricted drug use were seen on 2 of 7 medicine units subject to FRPA, whereas a significant increase was seen in 1 of 4 surgical units subject to FRPA.

Highlights • Fungal superinfection is a concern in patients who are hospitalized with COVID-19. • There is concern that the COVID-19 pandemic will lead to the overuse of antifungals. • Antifungal use was not increased early in the COVID-19 pandemic at an academic medical center.

Mobile medical apps are commonly used by health care professionals and could be used by antimicrobial stewardship programs to enhance adherence to local recommendations. We conducted a survey of health care workers to inform the design of an antimicrobial stewardship smartphone app.

Background: Data regarding outpatient antibiotic prescribing for urinary tract infections (UTIs) are limited, and they have never been formally summarized in Virginia. Objective: We describe outpatient antibiotic prescribing trends for UTIs based on gender, age, geographic region, insurance payer and International Classification of Disease, Tenth Revision (ICD-10) codes in Virginia. Methods: We used the Virginia All-Payer Claims Database (APCD), administered by Virginia Health Information (VHI), which holds data for Medicare, Medicaid, and private insurance. The study cohort included Virginia residents who had a primary diagnosis of UTI, had an antibiotic claim 0–3 days after the date of the diagnosis and who were seen in an outpatient facility in Virginia between January 1, 2016, and December 31, 2016. A diagnosis of UTI was categorized as cystitis, urethritis or pyelonephritis and was defined using the following ICD-10 codes: N30.0, N30.00, N30.01, N30.9, N30.90, N30.91, N39.0, N34.1, N34.2, and N10. The following antibiotics were prescribed: aminoglycosides, sulfamethoxazole/trimethoprim (TMP-SMX), cephalosporins, fluoroquinolones, macrolides, penicillins, tetracyclines, or nitrofurantoin. Patients were categorized based on gender, age, location, insurance payer and UTI type. We used χ2 and Cochran-Mantel-Haenszel testing. Analyses were performed in SAS version 9.4 software (SAS Institute, Cary, NC). Results: In total, 15,580 patients were included in this study. Prescriptions for antibiotics by drug class differed significantly by gender (P < .0001), age (P < .0001), geographic region (P < .0001), insurance payer (P < .0001), and UTI type (P < .0001). Cephalosporins were prescribed more often to women (32.48%, 4,173 of 12,846) than to men (26.26%, 718 of 2,734), and fluoroquinolones were prescribed more often to men (53.88%, 1,473 of 2,734) than to women (47.91%, 6,155 of 12,846). Although cephalosporins were prescribed most frequently (42.58%, 557 of 1,308) in northern Virginia, fluoroquinolones were prescribed the most in eastern Virginia (50.76%, 1677 of 3,304). Patients with commercial health insurance, Medicaid, and Medicare were prescribed fluoroquinolones (39.31%, 1,149 of 2,923), cephalosporins (56.33%, 1,326 of 2,354), and fluoroquinolones (57.36%, 5,910 of 10,303) most frequently, respectively. Conclusions: Antibiotic prescribing trends for UTIs varied by gender, age, geographic region, payer status and UTI type in the state of Virginia. These data will inform future statewide antimicrobial stewardship efforts.Funding: NoneDisclosures: Michelle Doll reports a research grant from Molnlycke Healthcare.

We used multivariable analyses to assess whether meeting core elements was associated with antibiotic utilization. Compliance with 7 elements versus not doing so was associated with higher use of broad-spectrum agents for community-acquired infections [days of therapy per 1,000 patient days: 155 (39) vs 133 (29), P = .02] and anti-methicillin-resistant S. aureus agents [days of therapy per 1,000 patient days: 145 (37) vs 124 (30), P = .03].

Purpose The main objective of this study was to investigate abuse of residents with either dementia or Alzheimer’s disease in long-term care settings, to identify facilitators and barriers surrounding implementation of systems to prevent such occurrences, and to draw conclusions on combating the issue of abuse. Patients and methods A systematic review was conducted using the Medline, CINAHL, and Academic Search Ultimate databases. With the use of key terms via Boolean search, 30 articles were obtained which were determined to be germane to research objectives. The review was conducted and structured based on Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. Results Residents with dementia or Alzheimer’s disease are at greater risk of abuse. The growing population could increase this problem exponentially. The most common facilitators were the introduction of policies/programs in the facility, education, and working conditions. The most cited barriers were poor training, lack of research, and working conditions in the long-term care setting. Conclusion The examples given would be useful in minimizing the potential for abuse in the long-term care setting. Leadership can take an active role in the prevention of abuse of the elderly through their actions, education of employees, and changes in the work environment.

Robust methods for deep-scale enrichment and site-specific identification of ubiquitylation sites are necessary for characterizing the myriad roles of protein ubiquitylation. To this end we previously developed UbiFast, a sensitive method for highly multiplexed ubiquitylation profiling where K-ɛ-GG peptides are enriched with anti-K-e-GG antibody and labeled on-antibody with isobaric labeling reagents for sample multiplexing. Here, we present robotic automation of the UbiFast method using a magnetic bead-conjugated K-e-GG antibody (mK-e-GG) and a magnetic particle processor. We report the identification of ∼20,000 ubiquitylation sites from a TMT10-plex with 500 μg input per sample processed in ∼2 hours. Automation of the UbiFast method greatly increased the number of identified and quantified ubiquitylation sites, improved reproducibility and significantly reduced processing time. The automated method also significantly reduced variability across process replicates compared to the manual method. The workflow enables processing of up to 96 samples in a single day making it suitable to study ubiquitylation in large sample sets. Here we demonstrate the applicability of the method to profile small amounts of tissue using breast cancer patient-derived xenograft (PDX) tissue samples.

Background: In this study, we assessed whether meropenem restriction led to delays in administration for patients in a medical intensive care unit (MICU) at a large tertiary-care urban teaching hospital. Methods: The antimicrobial stewardship program (ASP) at Virginia Commonwealth University Health System (VCUHS) requires approval for restricted antimicrobial orders placed between 8 a.m. and 9 p.m. Between 8 a.m. and 5 p.m. (daytime), authorized approvers include ASP and infectious diseases (ID) physicians. From 5 p.m. to 9 p.m. (evening) orders are approved by ID fellows. Orders were entered as Stat, Now, and Routine. Between 9 p.m. and 8 a.m. (night), patients receive doses without approval. Meropenem restriction began in mid-January 2018. Pre- and postmeropenem restriction periods were defined as February–December 2017 and February–December 2018. Meropenem use data were compared for adult patients in the MICU. A multivariable Cox regression model was implemented to compare (1) time from order entry to approval; (2) time from order approval to patient administration; (3) total time from order entry to patient administration, adjusting for order priority, approver (ASP, ID consult, ID fellow, pharmacy); and (4) time of day of order placement (day, eve, night). The analyses were performed using SAS version 9.4 software (SAS Institute, Cary, NC). Result: Time from order approval to patient administration was significantly decreased in the postrestriction period (HR, 1.840; P < .001) (Table 1). Stat orders were faster compared to routine orders for order entry to approval (HR, 1.735; P < .001), approval to administration (HR, 2.610; P < .001), and total time from order entry to administration (HR, 2.812; P < .001). No significant differences were found in time to approval by approving service. Time from order entry to approval was faster for nighttime orders than for daytime orders (HR, 1.399; P = .037). Conclusions: Our data indicate that the time from order entry to administration decreased following meropenem restriction in our MICU. More research is needed to identify the reason for this finding, but we postulate that this is due to an effect on drug administration prioritization within nursing workflow. These data will inform our local meropenem restriction efforts.Funding: NoneDisclosures: Michelle Doll reports a research grant from Molnlycke Healthcare.

Previous literature has shown how associate engagement has positively impacted on productivity, job satisfaction, safety, retention, consumer sentiment, and financial performance in hospitals and healthcare systems. However, a lack of research showing the relationship between associate engagement and job satisfaction within the long-term care environment has existed. Our objective was to investigate characteristics within the long-term care environment that promote and detract from associate job satisfaction and extrapolate the best practices in maintaining job satisfaction and engagement. This systematic review queried CINAHL, PubMed (MEDLINE), and Academic Search Ultimate databases for peer-reviewed publications for facilitators and barriers commensurate with employee job satisfaction in long-term care facilities using the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) and the Kruse Protocols. The authors identified 11 facilitators for job satisfaction and 18 barriers to job satisfaction in the 60 selected articles. The top four facilitators were Supportive Leadership, Capable and Motivated Employees, Positive Organizational Values, and Social Support Mechanisms. The top four barriers were condescending management style, high job demands, lack of self-care, and lack of training with medically complex patients. The systematic review revealed the importance of maintaining satisfied employees in the long-term care workplace through am emphasis leadership and on the facilitators identified to best serve their associates and improve care for residents.

BACKGROUND More than 18 million Americans are currently suffering from alcohol use disorder (AUD): a compulsive behavior of alcohol use as a result of a chronic, relapsing brain disease. With alcohol-related injuries being one of the leading causes of preventable deaths, there is a dire need to find ways to assist those suffering from alcohol dependence. There still exists a gap in knowledge as to the potential of telemedicine in improving health outcomes for those patients suffering from AUD. OBJECTIVE The purpose of this systematic review was to evaluate the measures of effectiveness, efficiency, and quality that result from the utilization of telemedicine in the management of alcohol abuse, addiction, and rehabilitation. METHODS This review was conducted utilizing the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines. The articles used in this analysis were gathered using keywords inclusive of both telemedicine and alcohol abuse, which were then searched in the Cumulative Index to Nursing and Allied Health Literature, Cochrane, and MEDLINE (PubMed) databases. A total of 22 articles were chosen for analysis. RESULTS The results indicated that telemedicine reduced alcohol consumption. Other common outcomes included reduced depression (4/35, 11%), increased patient satisfaction (3/35, 9%), increase in accessibility (3/35, 9%), increased quality of life (2/35, 6%), and decreased cost (1/35, 3%). Interventions included mobile health (11/22, 50%), electronic health (6/22, 27%), telephone (3/33, 14%), and 2-way video (2/22, 9%). Studies were conducted in 3 regions: the United States (13/22, 59%), the European Union (8/22, 36%), and Australia (1/22, 5%). CONCLUSIONS Telemedicine was found to be an effective tool in reducing alcohol consumption and increasing patients’ accessibility to health care services or health providers. The group of articles for analysis suggested that telemedicine may be effective in reducing health care costs and improving the patient’s quality of life. Although telemedicine shows promise as an effective way to manage alcohol-related disorders, it should be further investigated before implementation.

Set7/9 (also known as Set7, Set9, Setd7, and Kmt7) is a lysine methyltransferase that catalyzes the methylation of multiple substrates, including histone H3 and non-histone proteins. Although not essential for normal development and physiology, Set7/9-mediated methylation events play important roles in regulating cellular pathways involved in various human diseases, making Set7/9 a promising therapeutic target. Multiple Set7/9 inhibitors have been developed, which exhibit varying degrees of potency and selectivity in vitro. However, validation of these compounds in vivo has been hampered by the lack of a reliable cellular biomarker for Set7/9 activity. Here, we report the identification of Rpl29, a ribosomal protein abundantly expressed in all cell types, as a major substrate of Set7/9. We show that Rpl29 lysine 5 (Rpl29K5) is methylated exclusively by Set7/9 and can be demethylated by Lsd1 (also known as Kdm1a). Rpl29 is not a core component of the ribosome translational machinery and plays a regulatory role in translation efficiency. Our results indicate that Rpl29 methylation has no effect on global protein synthesis but affects Rpl29 subcellular localization. Using an Rpl29 methylation-specific antibody, we demonstrate that Rpl29K5 methylation is present ubiquitously and validate that (R)-PFI-2, a Set7/9 inhibitor, efficiently reduces Rpl29K5 methylation in cell lines. Thus, Rpl29 methylation can serve as a specific cellular biomarker for measuring Set7/9 activity.

This report examines the effectiveness of antimicrobial restriction at 1 tertiary care health care system by analyzing the consumption of restricted versus nonrestricted gram-positive agents over time for medical versus surgical units. Significant reductions in restricted antibiotic use were detected in 57% of medical units versus none of the surgical units. There were no significant reductions in nonrestricted antibiotic use. We think looking at antibiotic consumption by service line provides opportunities for targeted antibiotic restriction program refinement.

Background More than 18 million Americans are currently suffering from alcohol use disorder (AUD): a compulsive behavior of alcohol use as a result of a chronic, relapsing brain disease. With alcohol-related injuries being one of the leading causes of preventable deaths, there is a dire need to find ways to assist those suffering from alcohol dependence. There still exists a gap in knowledge as to the potential of telemedicine in improving health outcomes for those patients suffering from AUD. Objective The purpose of this systematic review was to evaluate the measures of effectiveness, efficiency, and quality that result from the utilization of telemedicine in the management of alcohol abuse, addiction, and rehabilitation. Methods This review was conducted utilizing the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines. The articles used in this analysis were gathered using keywords inclusive of both telemedicine and alcohol abuse, which were then searched in the Cumulative Index to Nursing and Allied Health Literature, Cochrane, and MEDLINE (PubMed) databases. A total of 22 articles were chosen for analysis. Results The results indicated that telemedicine reduced alcohol consumption. Other common outcomes included reduced depression (4/35, 11%), increased patient satisfaction (3/35, 9%), increase in accessibility (3/35, 9%), increased quality of life (2/35, 6%), and decreased cost (1/35, 3%). Interventions included mobile health (11/22, 50%), electronic health (6/22, 27%), telephone (3/33, 14%), and 2-way video (2/22, 9%). Studies were conducted in 3 regions: the United States (13/22, 59%), the European Union (8/22, 36%), and Australia (1/22, 5%). Conclusions Telemedicine was found to be an effective tool in reducing alcohol consumption and increasing patients’ accessibility to health care services or health providers. The group of articles for analysis suggested that telemedicine may be effective in reducing health care costs and improving the patient’s quality of life. Although telemedicine shows promise as an effective way to manage alcohol-related disorders, it should be further investigated before implementation.

Background Gram-negative bacterial infections are associated with high mortality in immunocompromised hosts, and the presence of drug resistance further increases mortality. Antibiotic consumption is a key outcome measure for Antimicrobial Stewardship Programs. Proper utilization of antibiotics can help limit the development of antimicrobial resistance. Resistance in Gram-negative organisms such as Pseudomonas, Enterobacter, and Acinetobacter is a major issue given the paucity of new drugs in the antibiotic pipeline for these organisms. A novel relative carbapenem consumption metric (the Proportion of Carbapenem Consumption, or PoCC) was recently described in US academic medical centers. The PoCC is calculated as follows: PoCC = [(meropenem Days of therapy(DOT)/1,000 patient-days (PDs))/(meropenem DOT/1,000 PDs + cefepime DOT/1,000 PDs + piperacillin–tazobactam DOT/1,000 PDs)]. The regional mean PoCC for the South Atlantic region has previously been approximated at 17%. Methods We examined the PoCC for the Bone Marrow Transplant (BMT) and dedicated Hematology/Oncology (H/O) inpatient wards at an academic medical center from August 2012 to June 2017. Results Table 1: Average Use of Antibiotics Expressed in DOT/1,000 PDs. Ward Piperacillin– Tazobactam Cefepime Meropenem Total PoCC Hematology/oncology 105.1 134.4 76.6 316.1 0.24 Bone marrow transplant 34.3 201.0 127.4 362.7 0.35 National meansa 76.2 60.2 30.7 b 0.18 aAs described by Markley et al. Infect Control Hosp Epidemiol 2018;39:229–232. bData unavailable. Conclusion This is the first description of the PoCC metric for dedicated Hematology/Oncology and Bone Marrow Transplant wards. When compared with national and regional mean PoCC scores for academic medical centers, the PoCC for these units was higher. More research is needed to determine the optimal PoCC scores for these types of units. The PoCC can contextualize relative carbapenem use and may be a useful antibiotic consumption metric. However, it does not provide data on absolute consumption. Further studies are needed to determine the best use of the PoCC metric by Antimicrobial Stewardship Programs for Hematology/Oncology and Bone Marrow Transplant wards. Disclosures All authors: No reported disclosures.

The purpose of this study was to determine if there are differences in interventions (int) required and compliance (comp) to recommendations (rec) by medical and surgical services as determined by an antimicrobial stewardship program (ASP). Concurrently collected ASP data from January 2008 through May 2011 were evaluated to determine number of int required and response to int. Results for medical (med) and surgical (surg) services were compared. There were 2322 int, of which 1108 were for med and 1214 were for surg. Recs were as follows: 242 appropriate coverage (AC) med versus 212 AC surg and 866 selective pressure (SP) med and 1002 SP surg. Comp with recs was higher for med: AC: 90.5 per cent (219 of 242) med versus 82.1 per cent (174 of 212) surg ( P = 0.0086) and SP: 81.1 per cent (702 of 866) med versus 69.5 per cent (696 of 1002) surg ( P < 0.0001). Overall surgical comp with ASP recs was better for AC than for SP. Moreover, less than half (seven of 15) of the surgical subspecialties demonstrated a comp of 75 per cent or greater with SP recs with only one-fifth having a comp over 80 per cent. Surgical services demonstrate poor compliance with ASP recommendations; this is especially true for interventions targeting selective pressure. By identifying services that are less compliant, programs can target their educational efforts to improve outcomes.

The Society of Healthcare Epidemiology of America, the Centers for Disease Control and Prevention, and the President's Council of Advisors on Science and Technology recognize the need to combat antimicrobial resistance through the promotion of antimicrobial stewardship programs. Health care facilities in Virginia were surveyed using a 23-item survey focused on facility characteristics and antimicrobial stewardship strategies. Antimicrobial stewardship activities were highly variable and many are missing key personnel and resources.

Traditional methods for analysis of peptides using liquid chromatography and tandem mass spectrometry (LC-MS/MS) lack the specificity to comprehensively monitor specific biological processes due to the inherent duty cycle limitations of the MS instrument and the stochastic nature of the analytical platform. PTMScan Direct is a novel, antibody-based method that allows quantitative LC-MS/MS profiling of specific peptides from proteins that reside in the same signaling pathway. New PTMScan Direct reagents have been produced that target peptides from proteins involved in DNA Damage/Cell Cycle and Apoptosis/Autophagy pathways. Together, the reagents provide access to 438 sites on 237 proteins in these signaling cascades. These reagents have been used to profile the response to UV damage of DNA in human cell lines. UV damage was shown to activate canonical DNA damage response pathways through ATM/ATR-dependent signaling, stress response pathways and induce the initiation of apoptosis, as assessed by an increase in the abundance of peptides corresponding to cleaved, activated caspases. These data demonstrate the utility of PTMScan Direct as a multiplexed assay for profiling specific cellular responses to various stimuli, such as UV damage of DNA.

Background Intrinsic thrombosis and stenosis are complications associated with the use of neck-remodeling devices in the treatment of intracranial aneurysms. Objective To examine the technical and anatomic factors that predict short- and long-term stent patency. Methods We undertook a retrospective review of 161 patients who underwent coil embolization of 168 ruptured and unruptured aneurysms assisted by the use of a neck-remodeling device. One hundred twenty-seven patients had catheter-based angiographic follow-up to evaluate 133 stent-coil constructs (mean, 15.4 months; median, 12.7 months). The technique of microcatheter jailing was used in a majority of patients; nonstandard stent configurations were also used. Results Clinical follow-up for all patients who had catheter-based angiograms demonstrated that among 133 stent constructs, a total of 9 (6.8%) had an in-stent event: 6 acute or subacute thrombosis (4.5%) and 3 delayed stenosis or occlusion (2.3%). Seven of these constructs were associated with a symptomatic event (5.3%). A significantly higher rate of in-stent events was seen with the use of constructs to treat anterior communicating artery aneurysms. When all patients are considered, including those who did not receive catheter-based follow-up imaging, 2 of 168 procedures (1.2%) resulted in the death of a patient, and procedural morbidity was 14.9%. Conclusion From these results and those in the published literature, in-stent complication rates are low in carefully selected patients. The use of dual antiplatelet therapy, sensitivity assays, and glycoprotein IIb/IIIa inhibitors may decrease the rate of acute and chronic in-stent complications.

Background Antimicrobial Stewardship Programs (ASPs) promote the optimal use of antimicrobial agents with the goal of preserving the effectiveness of existing drugs. One core ASP strategy is formulary restriction and preauthorization (PA). We evaluated restricted antimicrobial use at an academic medical center in Virginia; our program has a comprehensive restriction program that applies to adults only. Methods Data from August 2012 to June 2017 were evaluated at the hospital unit level by month in days of therapy (DOT) per 1,000 patient-days. Ordinary least squares regression was used to compare the time trend of restricted use with that of nonrestricted agents within the same unit. Results Across the study period significant decreases in restricted antibiotic use were detected for medical and pediatric units with no significant increases in use. However, significant increases were identified for surgical units. Table 1: Results of Time Trend Analysis by Unit for Restricted Drugs. Restricted Type Unit Time Trend P Medical Oncology 0.64 0.0924 Acute Care Medicine 0.39 0.4058 Cardiac ICU −0.37 0.1057 Medical ICU −2.06 0.0002 Bone Marrow Transplant 0.52 0.5002 Digestive Health −0.14 0.3004 Progressive Care −0.93 0.0002 Pediatric General Pediatrics −0.36 0.1363 Neonatal ICU −0.29 0.0007 Pediatric ICU −0.12 0.7849 Progressive Care −0.29 0.0993 Surgical Acute Care Surgery −0.30 0.1574 Burn ICU 0.84 0.0021 Cardiac Surgery ICU −0.50 0.2766 Surgical Trauma ICU −0.52 0.1019 Table 2: Significant Increases/ Decreases in Restricted Antimicrobials by Unit Type Restricted Unit Type Increase Decrease Medical 0/7 (0%) 2/7 (29%) Pediatric 0/4 (0%) 1/4 (25%) Surgical 1/4 (25%) 0/4 (0%) Conclusion These data suggest that the PA strategy for medical wards was effective across the time period whereas the PA strategy for surgical wards was suboptimal. However, it is unclear why pediatric wards (that were not subject to PA activities) also saw reductions in use; more research into this is needed. These data will help us to refine our PA strategy by targeting use on surgical wards. We believe that this type of analysis may be useful for other ASPs utilizing the PA strategy. Disclosures All authors: No reported disclosures.

Purpose. The implementation and evaluation of vancomycin nomogram guidelines in a computerized prescriber-order-entry (CPOE) system are described. Methods. Initial vancomycin orders for patients over age 18 years who received vancomycin between August 1 and September 30, 2006 (preimplementation), and between March 1 and April 30, 2007 (postimplementation), were compared with vancomycin nomogram recommendations to determine if the vancomycin regimen ordered coincided with the nomogram recommendation. The numbers of regimen changes and vancomycin serum concentrations measured during the first five days of therapy were also assessed. A multivariate logistic regression model assessed independent predictors of an initial vancomycin order that met the nomogram recommendation Results. A total of 522 vancomycin orders were included in the analysis (279 in the preimplementation group and 243 in the postimplementation group). A significant difference was observed in the percentage of initial vancomycin orders that met nomogram recommendations in the postimplementation group compared with the preimplementation group (36% versus 24%, p = 0.0028). No difference was noted between the two groups in the number of regimen changes or serum vancomycin concentrations measured during the first five days of therapy. In a multivariate analysis, age ( p = 0.02) and weight ( p < 0.0001) were negatively associated with a vancomycin order that met nomogram recommendations, while the postimplementation group was positively associated with an order that met nomogram recommendations ( p = 0.001). Conclusion. A vancomycin nomogram implemented into a CPOE system increased the likelihood of patients receiving an initial vancomycin regimen that coincided with the nomogram’s recommendations.

Anaplastic large cell lymphoma represents a subset of neoplasms caused by translocations that juxtapose the anaplastic lymphoma kinase (ALK) to dimerization partners. The constitutive activation of ALK fusion proteins leads to cellular transformation through a complex signaling network. To elucidate the ALK pathways sustaining lymphomagenesis and tumor maintenance, we analyzed the tyrosine-kinase protein profiles of ALK-positive cell lines using 2 complementary proteomic-based approaches, taking advantage of a specific ALK RNA interference (RNAi) or cell-permeable inhibitors. A well-defined set of ALK-associated tyrosine phosphopeptides, including metabolic enzymes, kinases, ribosomal and cytoskeletal proteins, was identified. Validation studies confirmed that vasodilator-stimulated phosphoprotein and 5-aminoimidazole-4-carboxamide ribonucleotide formyltransferase/inosine monophosphate cyclohydrolase (ATIC) associated with nucleophosmin (NPM)–ALK, and their phosphorylation required ALK activity. ATIC phosphorylation was documented in cell lines and primary tumors carrying ALK proteins and other tyrosine kinases, including TPR-Met and wild type c-Met. Functional analyses revealed that ALK-mediated ATIC phosphorylation enhanced its enzymatic activity, dampening the methotrexate-mediated transformylase activity inhibition. These findings demonstrate that proteomic approaches in well-controlled experimental settings allow the definition of informative proteomic profiles and the discovery of novel ALK downstream players that contribute to the maintenance of the neoplastic phenotype. Prediction of tumor responses to methotrexate may justify specific molecular-based chemotherapy.

A major question regarding the sensitivity of solid tumors to targeted kinase inhibitors is why some tumors respond and others do not. The observation that many tumors express EGF receptor (EGFR), yet only a small subset with EGFR-activating mutations respond clinically to EGFR inhibitors (EGFRIs), suggests that responsive tumors uniquely depend on EGFR signaling for their survival. The nature of this dependence is not understood. Here, we investigate dependence on EGFR signaling by comparing non-small-cell lung cancer cell lines driven by EGFR-activating mutations and genomic amplifications using a global proteomic analysis of phospho-tyrosine signaling. We identify an extensive receptor tyrosine kinase signaling network established in cells expressing mutated and activated EGFR or expressing amplified c-Met. We show that in drug sensitive cells the targeted tyrosine kinase drives other RTKs and an extensive network of downstream signaling that collapse with drug treatment. Comparison of the signaling networks in EGFR and c-Met-dependent cells identify a “core network” of ≈50 proteins that participate in pathways mediating drug response.

Activated tyrosine kinases have been frequently implicated in the pathogenesis of cancer, including acute myeloid leukemia (AML), and are validated targets for therapeutic intervention with small-molecule kinase inhibitors. To identify novel activated tyrosine kinases in AML, we used a discovery platform consisting of immunoaffinity profiling coupled to mass spectrometry that identifies large numbers of tyrosine-phosphorylated proteins, including active kinases. This method revealed the presence of an activated colony-stimulating factor 1 receptor (CSF1R) kinase in the acute megakaryoblastic leukemia (AMKL) cell line MKPL-1. Further studies using siRNA and a small-molecule inhibitor showed that CSF1R is essential for the growth and survival of MKPL-1 cells. DNA sequence analysis of cDNA generated by 5′RACE from CSF1R coding sequences identified a novel fusion of the RNA binding motif 6 (RBM6) gene to CSF1R gene generated presumably by a t(3;5)(p21;q33) translocation. Expression of the RBM6-CSF1R fusion protein conferred interleukin-3 (IL-3)–independent growth in BaF3 cells, and induces a myeloid proliferative disease (MPD) with features of megakaryoblastic leukemia in a murine transplant model. These findings identify a novel potential therapeutic target in leukemogenesis, and demonstrate the utility of phosphoproteomic strategies for discovery of tyrosine kinase alleles.

Gaining insight into normal cellular signaling and disease biology is a critical goal of proteomic analyses. The ability to perform these studies successfully to extract the maximum value and discovery of biologically relevant candidate biomarkers is therefore of primary importance. Many successful studies in the past have focused on total proteome analysis (changes at the protein level) combined with phosphorylation analysis by metal affinity enrichment (changes at the PTM level). Here, we use the gastric carcinoma cell line MKN-45 treated with the c-Met inhibitor SU11274 and PKC inhibitor staurosporine to investigate the most efficient and most comprehensive strategies for both total protein and PTM analysis. Under the conditions used, total protein analysis yielded few changes in response to either compound, while analysis of phosphorylation identified thousands of sites that changed differentially between the two treatments. Both metal affinity and antibody-based enrichments were used to assess phosphopeptide changes, and the data generated by the two methods was largely complementary (non-overlapping). Label-free quantitation of peptide peak abundances was used to accurately determine fold-changes between control and treated samples. Protein interaction network analysis allowed the data to be placed in a biologically relevant context, and follow-up validation of selected findings confirmed the accuracy of the proteomic data. Together, this study provides a framework for start-to-finish proteomic analysis of any experimental system under investigation to maximize the value of the proteomic study and yield the best chance for uncovering actionable target candidates.

A robust method was developed and optimized for enrichment and quantitative analysis of posttranslational modifications (PTMs) in serum/plasma samples by combining immunoaffinity purification and LC-MS/MS without depletion of abundant proteins. The method was used to survey serum samples of patients with acute myeloid leukemia (AML), breast cancer (BC), and nonsmall cell lung cancer (NSCLC). Peptides were identified from serum samples containing phosphorylation, acetylation, lysine methylation, and arginine methylation. Of the PTMs identified, lysine acetylation (AcK) and arginine mono-methylation (Rme) were more prevalent than other PTMs. Label-free quantitative analysis of AcK and Rme peptides was performed for sera from AML, BC, and NSCLC patients. Several AcK and Rme sites showed distinct abundance distribution patterns across the three cancer types. The identification and quantification of posttranslationally modified peptides in serum samples reported here can be used for patient profiling and biomarker discovery research.

Kinase domain (KD) mutations of Bcr-Abl interfering with imatinib binding are the major mechanism of acquired imatinib resistance in patients with Philadelphia chromosome-positive leukemia. Mutations of the ATP binding loop (p-loop) have been associated with a poor prognosis. We compared the transformation potency of five common KD mutants in various biological assays. Relative to unmutated (native) Bcr-Abl, the ATP binding loop mutants Y253F and E255K exhibited increased transformation potency, M351T and H396P were less potent, and the performance of T315I was assay dependent. The transformation potency of Y253F and M351T correlated with intrinsic Bcr-Abl kinase activity, whereas the kinase activity of E255K, H396P, and T315I did not correlate with transforming capabilities, suggesting that additional factors influence transformation potency. Analysis of the phosphotyrosine proteome by mass spectroscopy showed differential phosphorylation among the mutants, a finding consistent with altered substrate specificity and pathway activation. Mutations in the KD of Bcr-Abl influence kinase activity and signaling in a complex fashion, leading to gain- or loss-of-function variants. The drug resistance and transformation potency of mutants may determine the outcome of patients on therapy with Abl kinase inhibitors.