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Abstract Background The prevention of type 2 diabetes (T2DM) is recognised as a health care priority in the UK. In people living with T2DM, lifestyle changes (e.g. increasing physical activity) have been shown to slow disease progression and protect from the development of associated comorbidities. The use of digital health technologies provides a strategy to increase physical activity in patients with chronic disease. Furthermore, behaviour economics suggests that financial incentives may be a useful strategy for increasing the maintenance and effectiveness of behaviour change intervention, including physical activity intervention using digital health technologies. The Milton Keynes Activity Rewards Programme (MKARP) is a 24-month intervention which combines the use of a mobile health app, smartwatch (Fitbit or Apple watch) and financial incentives to encourage people living with T2DM to increase physical activity to improve health. Therefore, this randomised controlled trial aims to examine the long-term acceptability, health effects and cost-effectiveness of the MKARP on HbA1c in patients living with T2DM versus a waitlist usual care comparator. Methods A two-arm, single-centre, randomised controlled trial aiming to recruit 1018 participants with follow-up at 12 and 24 months. The primary outcome is the change in HbA1c at 12 months. Secondary outcomes included changes in markers of metabolic, cardiovascular, anthropometric, and psychological health along with cost-effectiveness. Recruitment will be via annual diabetes review in general practices, retinal screening services and social media. Participants aged 18 or over, with a diagnosis of type 2 diabetes and a valid HbA1c measurement in the last 2 months are invited to take part in the trial. Participants will be individually randomised (1:1 ratio) to receive either the Milton Keynes Activity Rewards Programme or usual care. The intervention will last for 24 months with assessment for outcomes at baseline, 12 and 24 months. Discussion This study will provide new evidence of the long-term effectiveness of an activity rewards scheme focused on increasing physical activity conducted within routine care in patients living with type 2 diabetes in Milton Keynes, UK. It will also investigate the cost-effectiveness of the intervention. Trial registration ISRCTN 14925701. Registered on 30 October 2023.

The coadministration of xylazine, a veterinary tranquilizer, with illicit fentanyl has led to severe soft tissue injuries, ranging from superficial irritation to deep tissue necrosis and even bone involvement, because of multifactorial tissue toxicity. Despite its non-opioid nature, xylazine enhances and prolongs the euphoric effects of fentanyl, exacerbating the potential for abuse. The pathogenesis of the tissue damage from xylazine is multifactorial but most akin to a burn from local tissue injury. With illicit opioids increasingly adulterated with xylazine, particularly in urban areas like Philadelphia, the prevalence of associated wounds, especially in the upper extremities, is anticipated to rise. Managing these wounds demands a multidisciplinary approach, with hand surgeons and reconstructive surgeons playing a central role. This review summarizes the historical context, pharmacodynamics, initial evaluation, wound categorization, algorithmic treatment, and expected outcomes of xylazine-associated wounds.

Purpose: Opioid stewardship ensures effective pain management while avoiding overprescribing of opioids after surgery. This prospective randomized study investigates the efficacy of a multimodal postoperative pain regimen compared to a traditional opioid-only pain regimen following elective outpatient orthopedic hand surgery. We hypothesized that patients receiving multimodal pain management would consume fewer opioids and report greater satisfaction than patients receiving only opioids. Methods: Consecutive patients undergoing outpatient hand and upper extremity surgery performed by two board-certified fellowship-trained orthopedic hand surgeons at one institution were recruited and randomized into either a study or control group. The study group received a standing multimodal postoperative regimen consisting of scheduled oral acetaminophen and naproxen as well as oxycodone to be taken as needed. The control group received only oxycodone to be taken as needed. Postoperatively, daily pain levels, medication usage, refills, satisfaction, and adverse events were recorded. Descriptive statistics were performed. Results: Of the 112 patients enrolled, 54 were randomized to the control group, and 58 were randomized to the study group. Study and control group patients did not differ significantly based on daily average pain scores or daily worst pain scores. However, study group patients reported fewer average daily oxycodone intake and total oxycodone pill count (7.0 vs 2.4 total pills, P

Purpose: Metacarpal fractures are common injuries of the hand that often require operative repair. However, there is a paucity of data regarding the rate of reoperation and circumstances following metacarpal repair. Methods: A retrospective review of all metacarpal fracture cases performed at a single academic institution between 2017 and 2021 was performed. All patients with isolated, acute metacarpal fractures were included for review. Data on patient demographics, fracture morphology, surgical technique, rate of early reoperation, and reason for reoperation were collected. Results: A total of 499 patients were identified to have undergone operative treatment for an isolated metacarpal fracture with an average follow-up of 4.2 months. The rate of unplanned early reoperation was 8.0% (n = 40), with seven patients requiring revision fracture surgery and 33 patients undergoing removal of symptomatic hardware. Mean and median time to reoperation was 2.1 and 1.5 months, respectively. The rate of reoperation for fractures of the metacarpal shaft was significantly lower than that of other fracture locations. Among the 40 revision cases, one case was following percutaneous fixation while 39 cases were following open reduction and internal fixation. Other demographic factures and fracture characteristics failed to show significant correlations to the rate of reoperation. Conclusions: An unplanned early reoperation rate of 8.0% after operative fixation of acute metacarpal fractures was observed with the majority involving cases of removal of symptomatic hardware and an average time to reoperation of approximately 2.1 months. This information can be used to counsel patients and set expectations about the potential for metacarpal fracture surgeries. Type of Study/Level of Evidence: Prognosis 2b.

Purpose: Intramedullary screw fixation has emerged as a popular approach for the treatment of displaced metacarpal fractures. The purpose of this study was to investigate the functional and radiographic outcomes of a newly designed, headless noncompressive fully threaded intramedullary nail (TIMN) for the treatment of metacarpal fractures. Methods: A retrospective chart review was performed on patients who were treated with the INnate TIMN (ExsoMed) at a single academic institution with a minimum of 1-year follow-up. Patient-reported functional outcomes included Quick Disabilities for the Arm, Shoulder, and Hand (QuickDASH) questionnaires, return to work and physical activity time, and overall satisfaction. Radiographs were retrospectively reviewed to determine radiographic union, change in angulation, and metacarpal shortening. Results: A total of 49 patients (58 fractures) with a mean age of 36 years (range: 17–75 years) were included. The mean follow-up time was 2.7 years (range: 1.4–4.3 years). Overall, the mean patient satisfaction rating was 4.9 of 5 (range: 3–5). The mean return to work time was 7.2 weeks (range: 0.14–28 weeks), and the mean return to sport or activity was 8.3 weeks (range: 1–28 weeks). Average QuickDASH scores across all patients were 4 (range: 0–56.9). The median radiographic healing time was 6.1 weeks (range: 4.7–15.4 weeks). Mean postoperative shortening in the fifth metacarpal fracture was 3 mm (range: −4.2 to 8 mm) at the initial postoperative visit and 3.6 mm (range: −3.3 to 7.9 mm) at the final radiographic follow-up. Subgroup analysis showed that postoperative shortening was similar, regardless of the fracture pattern. The following four complications were reported: one case of persistent pain and stiffness, one case of carpal tunnel syndrome, one nonunion, and one fractured intramedullary nail. Conclusions: Our findings suggest that the TIMN allows for a reliable return to work and physical activity, high patient satisfaction, low complication rate, and minimal shortening at the final radiographic follow-up. Type of study/level of evidence: Therapeutic IV.

The human leukocyte antigen (HLA) system is a major factor controlling cancer immunosurveillance and response to immunotherapy, yet its status in pediatric cancers remains fragmentary. We determined high-confidence HLA genotypes in 576 children, adolescents and young adults with recurrent/refractory solid tumors from the MOSCATO-01 and MAPPYACTS trials, using normal and tumor whole exome and RNA sequencing data and benchmarked algorithms. There was no evidence for narrowed HLA allelic diversity but discordant homozygosity and allele frequencies across tumor types and subtypes, such as in embryonal and alveolar rhabdomyosarcoma, neuroblastoma MYCN and 11q subtypes, and high-grade glioma, and several alleles may represent protective or susceptibility factors to specific pediatric solid cancers. There was a paucity of somatic mutations in HLA and antigen processing and presentation (APP) genes in most tumors, except in cases with mismatch repair deficiency or genetic instability. The prevalence of loss-of-heterozygosity (LOH) ranged from 5.9 to 7.7% in HLA class I and 8.0 to 16.7% in HLA class II genes, but was widely increased in osteosarcoma and glioblastoma (~15-25%), and for DRB1-DQA1-DQB1 in Ewing sarcoma (~23-28%) and low-grade glioma (~33-50%). HLA class I and HLA-DR antigen expression was assessed in 194 tumors and 44 patient-derived xenografts (PDXs) by immunochemistry, and class I and APP transcript levels quantified in PDXs by RT-qPCR. We confirmed that HLA class I antigen expression is heterogeneous in advanced pediatric solid tumors, with class I loss commonly associated with the transcriptional downregulation of HLA-B and transporter associated with antigen processing (TAP) genes, whereas class II antigen expression is scarce on tumor cells and occurs on immune infiltrating cells. Patients with tumors expressing sufficient HLA class I and TAP levels such as some glioma, osteosarcoma, Ewing sarcoma and non-rhabdomyosarcoma soft-tissue sarcoma cases may more likely benefit from T cell-based approaches, whereas strategies to upregulate HLA expression, to expand the immunopeptidome, and to target TAP-independent epitopes or possibly LOH might provide novel therapeutic opportunities in others. The consequences of HLA class II expression by immune cells remain to be established. Immunogenetic profiling should be implemented in routine to inform immunotherapy trials for precision medicine of pediatric cancers.

Abstract Background Prescription opioids remain an important contributor to the United States opioid crisis and to the development of opioid use disorder for opioid-naïve individuals. Recent legislative actions, such as the implementation of state prescription drug monitoring programs (PDMPs), aim to reduce opioid morbidity and mortality through enhanced tracking and reporting of prescription data. The primary objective of our study was to describe the opioid prescribing trends in the state of Pennsylvania (PA) as recorded by the PA PDMP following legislative changes in reporting guidelines, and discuss the PDMP’s role in a multifactorial approach to opioid harm reduction. Methods State-level opioid prescription data summaries recorded by the PA PDMP for each calendar quarter from August 2016 through March 2020 were collected from the PA Department of Health. Data for oxycodone, hydrocodone, and morphine were analyzed by quarter for total prescription numbers and refills. Prescription lengths, pill quantities, and average morphine milliequivalents (MMEs) were analyzed by quarter for all 14 opioid prescription variants recorded by the PA PDMP. Linear regression was conducted for each group of variables to identify significant differences in prescribing trends. Results For total prescriptions dispensed, the number of oxycodone, hydrocodone, and morphine prescriptions decreased by 34.4, 44.6, and 22.3% respectively (p

Computational methods coupled with experimental validation play a critical role in the identification of novel inhibitory peptides that interact with viral antigenic determinants. The interaction between the receptor binding domain (RBD) of SARS-CoV-2 spike protein and the helical peptide of human angiotensin-converting enzyme-2 (ACE2) is a necessity for the initiation of viral infection. Herein, natural orthologs of human ACE2 helical peptide were evaluated for competitive inhibitory binding to the viral RBD by use of a computational approach, which was experimentally validated. A total of 624 natural ACE2 orthologous 32-amino acid long peptides were identified through a similarity search. Molecular docking was used to virtually screen and rank the peptides based on binding affinity metrics, benchmarked against human ACE2 peptide docked to the RBD. Molecular dynamics (MD) simulations were done for the human reference and the Nipponia nippon peptide as it exhibited the highest binding affinity (Gibbs free energy; −14 kcal/mol) predicted from the docking results. The MD simulation confirmed the stability of the assessed peptide in the complex (−12.3 kcal/mol). The top three docked-peptides (from Chitinophaga sancti, Nipponia nippon, and Mus musculus) and the human reference were experimentally validated by use of surface plasmon resonance technology. The human reference exhibited the weakest binding affinity (Kd of 318–441 pM) among the peptides tested, in agreement with the docking prediction, while the peptide from Nipponia nippon was the best, with 267–538-fold higher affinity than the reference. The validated peptides merit further investigation. This work showcases that the approach herein can aid in the identification of inhibitory biosimilar peptides for other viruses.

The present study reports the prevalence of Paramphistomum spp. in small and large ruminants and their association with the histopathology of the infected rumens. A total of 384 animals were screened for Paramphistomum spp. The animals found positive for Paramphistomum spp. were divided into three groups according to the worm load/5 cm2 (G1: 10 – 20 worms/5 cm2 = Low, G2: 20 – 40 worms/5 cm2 = Medium, and G3: >41 worms/5 cm2 = High). Tissue slides were prepared from samples of the rumen (1 cm2) taken from animals positive for ruminal fluke to determine the histological parameters, including epithelial length or thickness, length and width of the ruminal papilla, and thickness of tunica submucosa and mucularis externae. The overall prevalence of Paramphistomum spp. in the ruminant population of district Narowal was 56.25 % with a significant (P < 0.05) variation among different species of ruminants. The highest prevalence was in cattle, followed in order by buffalo, goat, and sheep. Epithelium thickness was significantly correlated with parasite load in large ruminants and the most significant (P < 0.05) decrease in epithelium thickness was in Group B (31.12 ± 1.82 μm) and Group C (31.07 ± 1.68 μm) and a same trend was recorded in small ruminants. Histopathological changes due to Paramphistomum spp. are reported for the first time, which explained the histomorphological and physiological changes in Paramphistomum-infected rumens which might be associated with lowered feed efficiency and productivity in ruminants.

The United States’ opioid epidemic has taken an immense toll over the past 2 decades when assessed by morbidities, mortalities, and economic costs. Prescription opioids are a substantial contribution to this public health emergency, and it is critical for health care providers to practice good analgesic stewardship. Interventions have effectively curtailed opioid overuse, including prescription drug monitoring programs, educational initiatives, and multimodal analgesia strategies. Surgeons, particularly hand surgeons or those who perform musculoskeletal procedures, have been implicated as high-volume opioid prescribers. Guidelines for appropriate opioid dosing and analgesic management strategies after common hand and upper-extremity surgeries are sparse and offer an area for meaningful improvement. We sought to generate comprehensive, evidence-based recommendations for postoperative analgesia regimens for common hand and upper-extremity procedures.

Mahnoor Asif, Tanzila Siddiqui Karachi Medical and Dental College, Karachi, PakistanCorrespondence: Tanzila Siddiqui, Tel +923471159632, Email tanzilas716@gmail.com

Klebsiella pneumoniae is a Gram-negative pathogen of clinical relevance, which can provoke serious urinary and blood infections and pneumonia. This bacterium is a major public health threat due to its resistance to several antibiotic classes. Using a reverse vaccinology approach, 7 potential antigens were identified, of which 4 were present in most of the sequences of Italian carbapenem-resistant K. pneumoniae clinical isolates. Bioinformatics tools demonstrated the antigenic potential of these bacterial proteins and allowed for the identification of T and B cell epitopes. This led to a rational design and in silico characterization of a multiepitope vaccine against carbapenem-resistant K. pneumoniae strains. As adjuvant, the mycobacterial heparin-binding hemagglutinin adhesin (HBHA), which is a Toll-like receptor 4 (TLR-4) agonist, was included, to increase the immunogenicity of the construct. The multiepitope vaccine candidate was analyzed by bioinformatics tools to assess its antigenicity, solubility, allergenicity, toxicity, physical and chemical parameters, and secondary and tertiary structures. Molecular docking binding energies to TLR-2 and TLR-4, two important innate immunity receptors involved in the immune response against K. pneumoniae infections, and molecular dynamics simulations of such complexes supported active interactions. A codon optimized multiepitope sequence cloning strategy is proposed, for production of recombinant vaccine in classical bacterial vectors. Finally, a 3 dose-immunization simulation with the multiepitope construct induced both cellular and humoral immune responses. These results suggest that this multiepitope construct has potential as a vaccination strategy against carbapenem-resistant K. pneumoniae and deserves further validation.

Abstract Background The United States opioid epidemic is a devastating public health crisis fueled in part by physician prescribing. While the next generation of prescribers is crucial to the trajectory of the epidemic, medical school curricula designated to prepare students for opioid prescribing (OP) and pain management is often underdeveloped. In response to this deficit, we aimed to investigate the impact of an online opioid and pain management (OPM) educational intervention on fourth-year medical student knowledge, attitudes, and perceived competence. Methods Graduating students completing their final year of medical education at Sidney Kimmel Medical College of Thomas Jefferson University were sent an e-mail invitation to complete a virtual OPM module. The module consisted of eight interactive patient cases that introduced topics through a case-based learning system, challenging students to make decisions and answer knowledge questions about the patient care process. An identical pre- and posttest were built into the module to measure general and case-specific learning objectives, with responses subsequently analyzed using the Wilcoxon matched-pairs signed-rank test. Results Forty-three students (19% response rate) completed the module. All median posttest responses ranked significantly higher than paired median pretest responses (p

Mahnoor Asif,&ast; Syeda Sakina Zehra,&ast; Tanzila Siddiqui Karachi Medical and Dental College, Karachi, Pakistan&ast;These authors contributed equally to this workCorrespondence: Syeda Sakina Zehra, Tel +92 3322609216, Email szehra362@gmail.com

Abstract Background Biology has entered the era of big data with the advent of high-throughput omics technologies. Biological databases provide public access to petabytes of data and information facilitating knowledge discovery. Over the years, sequence data of pathogens has seen a large increase in the number of records, given the relatively small genome size and their important role as infectious and symbiotic agents. Humans are host to numerous pathogenic diseases, such as that by viruses, many of which are responsible for high mortality and morbidity. The interaction between pathogens and humans over the evolutionary history has resulted in sharing of sequences, with important biological and evolutionary implications. Results This study describes a large-scale, systematic bioinformatics approach for identification and characterization of shared sequences between the host and pathogen. An application of the approach is demonstrated through identification and characterization of the Flaviviridae-human share-ome. A total of 2430 nonamers represented the Flaviviridae-human share-ome with 100% identity. Although the share-ome represented a small fraction of the repertoire of Flaviviridae (~ 0.12%) and human (~ 0.013%) non-redundant nonamers, the 2430 shared nonamers mapped to 16,946 Flaviviridae and 7506 human non-redundant protein sequences. The shared nonamer sequences mapped to 125 species of Flaviviridae, including several with unclassified genus. The majority (~ 68%) of the shared sequences mapped to Hepacivirus C species; West Nile, dengue and Zika viruses of the Flavivirus genus accounted for ~ 11%, ~ 7%, and ~ 3%, respectively, of the Flaviviridae protein sequences (16,946) mapped by the share-ome. Further characterization of the share-ome provided important structural-functional insights to Flaviviridae-human interactions. Conclusion Mapping of the host-pathogen share-ome has important implications for the design of vaccines and drugs, diagnostics, disease surveillance and the discovery of unknown, potential host-pathogen interactions. The generic workflow presented herein is potentially applicable to a variety of pathogens, such as of viral, bacterial or parasitic origin.

Mehreen Bashir,1 Junaid Ahmad,1 Muhammad Asif,2 Salah-Ud-Din Khan,3 Muhammad Irfan,1 Asim Y Ibrahim,4 Sajid Asghar,1 Ikram Ullah Khan,1 Muhammad Shahid Iqbal,5 Abdul Haseeb,6 Syed Haroon Khalid,1 Mohammed AS Abourehab7,8 1Department of Pharmaceutics, Faculty of Pharmaceutical Sciences, Government College University, Faisalabad, 38000, Pakistan; 2Department of Pharmacology, Faculty of Pharmacy, The Islamia University of Bahawalpur, Bahawalpur, Pakistan; 3Department of Biochemistry, College of Medicine, Imam Mohammad Ibn Saud Islamic University, Riyadh, Saudi Arabia; 4Faculty of Pharmacy, Omdurman Islamic University, Omdurman, Sudan; 5Department of Clinical Pharmacy, College of Pharmacy, Prince Sattam Bin Abdulaziz University, Alkharj, Saudi Arabia; 6Department of Clinical Pharmacy, College of Pharmacy, Umm Al-Qura University, Makkah, 21955, Saudi Arabia; 7Department of Pharmaceutics, Faculty of Pharmacy, Umm Al-Qura University, Makkah, Saudi Arabia; 8Department of Pharmaceutics and Industrial Pharmacy, Faculty of Pharmacy, Minia University, Minia, EgyptCorrespondence: Syed Haroon KhalidDepartment of Pharmaceutics, Faculty of Pharmaceutical Sciences, Government College University, Faisalabad, 38000, PakistanTel +92-3166752223Email syedharoonkhalid@gcuf.edu.pkPurpose: Rheumatoid arthritis is an autoimmune disorder that directly affects joints. However, other body organs including heart, eyes, skin, blood vessels and lungs may also be affected. The purpose of this study was to design and evaluate a nanoemulgel formulation of diflunisal (DIF) and solubility enhanced diflunisal (DIF-IC) for enhanced topical anti-inflammatory activity.Methodology: Nanoemulsion formulations of both DIF and DIF-IC were prepared and incorporated in three different gelling agents, namely carboxymethylcellulose sodium (CMC-Na), sodium alginate (Na-ALG) and xanthan gum (XG). All the formulations were evaluated in term of particle size, pH, conductivity, viscosity, zeta potential and in vitro drug release. The formulation 2 (NE2) of both DIF and DIF-IC which expressed optimum release and satisfactory physicochemical properties was incorporated with gelling agents to produce final nanoemulgel formulations. The optimized nanoemulgel formulation was subjected to three different in vivo anti-inflammatory models including carrageenan-induced paw edema model, histamine-induced paw edema model and formalin-induced paw edema model.Results: DIF-IC-loaded nanoemulgel formulations yielded significantly enhanced in vitro skin permeation than DIF-loaded nanoemulgel. The nanoemulgel formulation of DIF-IC formulated with XG produced improved in vivo anti-inflammatory activity.Conclusion: It was recommended that DIF-IC-based nanoemulgel formulation prepared with XG could be a better option for effective topical treatment of inflammatory conditions.Keywords: diflunisal, nanoemulsion, pseudoternary phase diagram, in vitro skin permeation, anti-inflammatory activity, improved efficacy

The demand for telemedicine has been increasing over the past several years with the growth of technology and digital connectivity in our daily lives. With the impact of the global coronavirus disease 2019 pandemic, telemedicine implementation has become a necessity for many specialties because social distancing measures have greatly affected access to routine medical care. This article presents a detailed and systematic approach to conducting a hand physical examination during a video telemedicine encounter. Although the telemedicine physical examination has limitations, most components of the normal physical examination can be completed remotely with a systematic approach. We enumerate modifications to maximize examination remotely and present considerations for improved delivery of telemedicine care. These methods may be beneficial to providers incorporating telemedicine into their practice.

Purpose: Thumb basal joint arthritis is a common degenerative condition of the hand that is often managed with thumb basal joint arthroplasty (BJA). This procedure generally results in a high level of patient satisfaction; however, the rate and cause of early unplanned reoperation after thumb BJA are not well-understood. Therefore, we performed a review to better understand the rate and cause of early reoperation. Methods: A retrospective review of all thumb BJA cases performed at a single private academic center between 2014 and 2016 yielded 637 patients and 686 primary thumb BJAs with a minimum 1-year follow-up (mean, 2.4 years). Data collection included patient demographics, surgical technique and type of thumb BJA performed, time to reoperation, reason for early reoperation (within 2 years), and type of reoperation. Results: Of 686 patients undergoing thumb BJAs, 10 had unplanned early reoperation (1.5%). Mean duration between the index procedure and reoperation was 5.2 months (range, 0.5–14.3 months). Of the 10 unplanned early reoperations, 4 thumbs in 4 patients required revision arthroplasties owing to persistent pain. Time to reoperation for revision arthroplasty was 9.6 months (range, 3.9–14.3 months). Three of 10 reoperations resulted from early infection, 2 from unplanned early removal of symptomatic K-wires, and one from radial sensory neuritis. Conclusions: In this series of nearly 700 consecutive cases, we identified an unexpected early reoperation rate of 1.5%, with only a 0.6% reoperation rate specifically for painful subsidence requiring a revision arthroplasty. Mean time to revision was 9.6 months. These rates are lower than those published previously and should be considered by patients and surgeons when planning thumb BJA. Type of study/level of evidence: Prognostic IV. Key words: Basal joint arthroplasty, Reoperation, Revision, Thumb, Trapeziometacarpal arthritis

Background: Glioblastoma poses an inevitable threat to patients despite aggressive therapy regimes. It displays a great level of molecular heterogeneity and numerous substitutions in several genes have been documented. Next-generation sequencing techniques have identified various molecular signatures that have led to a better understanding of the molecular pathogenesis of glioblastoma. In this limited study, we sought to identify genetic variants in a small number of rare patients with aggressive glioblastoma. Methods: Five tumor tissue samples were isolated from four patients with rapidly growing glioblastoma. Genomic DNA was isolated and whole exome sequencing was used to study protein-coding regions. Generated FASTQ files were analyzed and variants were called for each sample. Variants were prioritized with different approaches and functional annotation was applied for the detrimental variants. Results: A total of 49,780 somatic variants were identified in the five glioblastoma samples studied, with the majority as missense substitutions. The top ten genes with the highest number of substitutions were MUC3A, MUC4, MUC6, OR4C5, PDE4DIP, AHNAK2, OR4C3, ZNF806, TTN, and RP1L1. Notably, variant prioritization after annotation indicated that the MTCH2 (Chr11: 47647265 A>G) gene sequence change was putative deleterious in all of the aggressive tumor samples. Conclusion: The MTCH2 (Chr11: 47647265 A>G) gene substitution was identified as putative deleterious in highly aggressive glioblastomas, which merits further investigation. Moreover, a high tumor mutation burden was observed, with a signature of the highest substitutions in MUC3A, MUC4, MUC6, OR4C5, PDE4DIP, AHNAK2, OR4C3, ZNF806, TTN, and RP1L1 genes. The findings provide critical, initial data for the further rational design of genetic screening and diagnostic approaches against aggressive glioblastoma.

The knowledge about the spatial distribution of soil organic carbon stock (SOCS) helps in sustainable land-use management and ecosystem functioning. No such study has been attempted in the complex topography and land use of Himalayas, which is associated with great spatial heterogeneity and uncertainties. Therefore, in this study digital soil mapping (DSM) was used to predict and evaluate the spatial distribution of SOCS using advanced geostatistical methods and a machine learning algorithm in the Himalayan region of Jammu and Kashmir, India. Eighty-three soil samples were collected across different land uses. Auxiliary variables (spectral indices and topographic parameters) derived from satellite data were used as predictors. Geostatistical methods—ordinary kriging (OK) and regression kriging (RK)—and a machine learning method—random forest (RF)—were used for assessing the spatial distribution and variability of SOCS with inter-comparison of models for their prediction performance. The best fit model validation criteria used were coefficient of determination (R2) and root mean square error (RMSE) with resulting maps validated by cross-validation. The SOCS concentration varied from 1.12 Mg/ha to 70.60 Mg/ha. The semivariogram analysis of OK and RK indicated moderate spatial dependence. RF (RMSE = 8.21) performed better than OK (RMSE = 15.60) and RK (RMSE = 17.73) while OK performed better than RK. Therefore, it may be concluded that RF provides better estimation and spatial variability of SOCS; however, further selection and choice of auxiliary variables and higher soil sampling density could improve the accuracy of RK prediction.

Abstract Background The sequence diversity of dengue virus (DENV) is one of the challenges in developing an effective vaccine against the virus. Highly conserved, serotype-specific (HCSS), immune-relevant DENV sequences are attractive candidates for vaccine design, and represent an alternative to the approach of selecting pan-DENV conserved sequences. The former aims to limit the number of possible cross-reactive epitope variants in the population, while the latter aims to limit the cross-reactivity between the serotypes to favour a serotype-specific response. Herein, we performed a large-scale systematic study to map and characterise HCSS sequences in the DENV proteome. Methods All reported DENV protein sequence data for each serotype was retrieved from the NCBI Entrez Protein (nr) Database (txid: 12637). The downloaded sequences were then separated according to the individual serotype proteins by use of BLASTp search, and subsequently removed for duplicates and co-aligned across the serotypes. Shannon’s entropy and mutual information (MI) analyses, by use of AVANA, were performed to measure the diversity within and between the serotype proteins to identify HCSS nonamers. The sequences were evaluated for the presence of promiscuous T-cell epitopes by use of NetCTLpan 1.1 and NetMHCIIpan 3.2 server for human leukocyte antigen (HLA) class I and class II supertypes, respectively. The predicted epitopes were matched to reported epitopes in the Immune Epitope Database. Results A total of 2321 nonamers met the HCSS selection criteria of entropy 0.8. Concatenating these resulted in a total of 337 HCSS sequences. DENV4 had the most number of HCSS nonamers; NS5, NS3 and E proteins had among the highest, with none in the C and only one in prM. The HCSS sequences were immune-relevant; 87 HCSS sequences were both reported T-cell epitopes/ligands in human and predicted epitopes, supporting the accuracy of the predictions. A number of the HCSS clustered as immunological hotspots and exhibited putative promiscuity beyond a single HLA supertype. The HCSS sequences represented, on average, ~ 40% of the proteome length for each serotype; more than double of pan-DENV sequences (conserved across the four serotypes), and thus offer a larger choice of sequences for vaccine target selection. HCSS sequences of a given serotype showed significant amino acid difference to all the variants of the other serotypes, supporting the notion of serotype-specificity. Conclusion This work provides a catalogue of HCSS sequences in the DENV proteome, as candidates for vaccine target selection. The methodology described herein provides a framework for similar application to other pathogens.

Muhammad Adeel,1 Muhammad Asif,1 Muhammad Naeem Faisal,2 Muhammad Hasanain Chaudary,3 Muhammad Sheraz Malik,4 Muhammad Khalid5 1Department of Computer Science, Faculty of Science, National Textile University, Faisalabad, Punjab, Pakistan; 2Faculty of Veterinary Science, Institute of Pharmacy, Physiology, and Pharmacology, University of Agriculture, Faisalabad, Punjab, Pakistan; 3Department of Computer Science, COMSATS University, Islamabad, Lahore Campus, Lahore, Punjab, Pakistan; 4Department of Information Technology, Government College University, Faisalabad, Punjab, Pakistan; 5Oncology Department, Faisalabad Medical University, Allied Hospital, Faisalabad, Punjab, Pakistan Purpose: This retrospective study presents a comparative analysis of the overall survival and toxicities, as side effects, of docetaxel plus cyclophosphamide (TC) and doxorubicin plus cyclophosphamide (AC). The study measured their efficacies during adjuvant chemotherapy, treating Pakistani breast cancer patients by validating the results obtained, with the published analysis of the same treatment given to US patients.Patients and methods: Between June 2015 and September 2017, for four chemotherapy cycles, 189 patients out of 358 received TC (75 mg/m2 of docetaxel, 600 mg/m2 of cyclophosphamide) and 169 were treated with AC (60 mg/m2 of doxorubicin, 600 mg/m2 of cyclophosphamide). On the basis of using pathological markers to assess patients, toxicities, as side effects, (due to docetaxel, doxorubicin, and cyclophosphamide) were listed in the database of this study. Common factors with respect to common terminology criteria for adverse events version 5.0 and side effects listed in MedlinePlus, NIH US database, and from the database of this study were then separated to be included in comparison for this study. Statistically, chi-squared test was used at α=0.05.Results: There was no statistically significant difference between the proportions of patients with vomiting, extreme tiredness, diarrhea, mild anemia, stability, and overall survival because P-value >0.05. However, AC remained less toxic (P-value

Readmissions for pulmonary hypertension are poorly understood and understudied. We sought to determine national estimates and risk factors for 30-day readmission after pulmonary hypertension-related hospitalizations. We utilized the Healthcare Cost and Utilization Project Nationwide Readmission Database, which has weighted estimates of roughly 35 million discharges in the US. Adult patients with primary International Classification of Disease, Ninth Revision, Clinical Modification diagnosis codes of 416.0 and 416.8 for primary and secondary pulmonary hypertension with an index admission between 2012 and 2014 and any readmission within 30 days of the index event were identified. Predictors of 30-day readmission were identified using multivariable logistic regression with adjustment for covariates. Results showed that the national estimate for Primary Pulmonary Hypertension vs Secondary Pulmonary Hypertension-related index events between 2012 and 2014 with 30-day readmission was 247 vs 2550 corresponding to a national readmission risk estimate of 17% vs 18.3%, respectively. The presence of fluid and electrolyte disorders, renal failure, and alcohol abuse were associated with increased risk of readmission in Primary Pulmonary Hypertension, while factors associated with Secondary Pulmonary Hypertension readmissions included anemia, congestive heart failure, lung disease, fluid and electrolyte disorders, renal failure, diabetes, and liver disease. The median cost of Primary Pulmonary Hypertension admissions and readmissions were $46,132 (IQR: $25,384–$85,647) and $41,604.50 (IQR: $22,481.50–$84,420.50), respectively. The median costs of Secondary Pulmonary Hypertension admissions and readmissions were $34,893 (IQR: $19,670–$66,143) and $36,279 (IQR: $19,059–$74,679), respectively. In conclusion, approximately 19% of Primary Pulmonary Hypertension and Secondary Pulmonary Hypertension hospitalizations result in 30-day readmission, with significant costs accrued during the index hospitalization and readmission. With evolving clinical terminology and diagnostic codes, future study will need to better clarify underlying factors associated with readmissions amongst pulmonary hypertension sub-types, and identify methods and procedures to minimize readmission risk.

Background Influenza A (H5N1) virus is a global concern with potential as a pandemic threat. High sequence variability of influenza A viruses is a major challenge for effective vaccine design. A continuing goal towards this is a greater understanding of influenza A (H5N1) proteome sequence diversity in the context of the immune system (antigenic diversity), the dynamics of mutation, and effective strategies to overcome the diversity for vaccine design. Methods Herein, we report a comprehensive study of the dynamics of H5N1 mutations by analysis of the aligned overlapping nonamer positions (1–9, 2–10, etc.) of more than 13,000 protein sequences of avian and human influenza A (H5N1) viruses, reported over at least 50 years. Entropy calculations were performed on 9,408 overlapping nonamer position of the proteome to study the diversity in the context of immune system. The nonamers represent the predominant length of the binding cores for peptides recognized by the cellular immune system. To further dissect the sequence diversity, each overlapping nonamer position was quantitatively analyzed for four patterns of sequence diversity motifs: index, major, minor and unique. Results Almost all of the aligned overlapping nonamer positions of each viral proteome exhibited variants (major, minor, and unique) to the predominant index sequence. Each variant motif displayed a characteristic pattern of incidence change in relation to increased total variants. The major variant exhibited a restrictive pyramidal incidence pattern, with peak incidence at 50% total variants. Post this peak incidence, the minor variants became the predominant motif for majority of the positions. Unique variants, each sequence observed only once, were present at nearly all of the nonamer positions. The diversity motifs (index and variants) demonstrated complex inter-relationships, with motif switching being a common phenomenon. Additionally, 25 highly conserved sequences were identified to be shared across viruses of both hosts, with half conserved to several other influenza A subtypes. Discussion The presence of distinct sequences (nonatypes) at nearly all nonamer positions represents a large repertoire of reported viral variants in the proteome, which influence the variability dynamics of the viral population. This work elucidated and provided important insights on the components that make up the viral diversity, delineating inherent patterns in the organization of sequence changes that function in the viral fitness-selection. Additionally, it provides a catalogue of all the mutational changes involved in the dynamics of H5N1 viral diversity for both avian and human host populations. This work provides data relevant for the design of prophylactics and therapeutics that overcome the diversity of the virus, and can aid in the surveillance of existing and future strains of influenza viruses.

Abstract Background Newcastle disease (ND) outbreaks are global challenges to the poultry industry. Effective management requires rapid identification and virulence prediction of the circulating Newcastle disease viruses (NDV), the causative agent of ND. However, these diagnostics are hindered by the genetic diversity and rapid evolution of NDVs. Methods An amplicon sequencing (AmpSeq) workflow for virulence and genotype prediction of NDV samples using a third-generation, real-time DNA sequencing platform is described here. 1D MinION sequencing of barcoded NDV amplicons was performed using 33 egg-grown isolates, (15 NDV genotypes), and 15 clinical swab samples collected from field outbreaks. Assembly-based data analysis was performed in a customized, Galaxy-based AmpSeq workflow. MinION-based results were compared to previously published sequences and to sequences obtained using a previously published Illumina MiSeq workflow. Results For all egg-grown isolates, NDV was detected and virulence and genotype were accurately predicted. For clinical samples, NDV was detected in ten of eleven NDV samples. Six of the clinical samples contained two mixed genotypes as determined by MiSeq, of which the MinION method detected both genotypes in four samples. Additionally, testing a dilution series of one NDV isolate resulted in NDV detection in a dilution as low as 101 50% egg infectious dose per milliliter. This was accomplished in as little as 7 min of sequencing time, with a 98.37% sequence identity compared to the expected consensus obtained by MiSeq. Conclusion The depth of sequencing, fast sequencing capabilities, accuracy of the consensus sequences, and the low cost of multiplexing allowed for effective virulence prediction and genotype identification of NDVs currently circulating worldwide. The sensitivity of this protocol was preliminary tested using only one genotype. After more extensive evaluation of the sensitivity and specificity, this protocol will likely be applicable to the detection and characterization of NDV.

Muhammad Asif,1,2 Iqbal Ahmad Alvi,1,3 Shafiq Ur Rehman1 1Department of Microbiology and Molecular Genetics, University of the Punjab, Lahore, Pakistan; 2Department of Pathology, King Edward Medical University, Lahore, Pakistan; 3Department of Microbiology, Hazara University, Mansehra, Pakistan Abstract: Acinetobacter baumannii, once considered a low-category pathogen, has emerged as an obstinate infectious agent. The scientific community is paying more attention to this pathogen due to its stubbornness to last resort antimicrobials, including carbapenems, colistin, and tigecycline, its high prevalence of infections in the hospital setting, and significantly increased rate of community-acquired infections by this organism over the past decade. It has given the fear of pre-antibiotic era to the world. To further enhance our understanding about this pathogen, in this review, we discuss its taxonomy, pathogenesis, current treatment options, global resistance rates, mechanisms of its resistance against various groups of antimicrobials, and future therapeutics. Keywords: antibiotic resistance, colistin, tigecycline, phage therapy

Abstract Background Ebolavirus (EBOV) is responsible for one of the most fatal diseases encountered by mankind. Cellular T-cell responses have been implicated to be important in providing protection against the virus. Antigenic variation can result in viral escape from immune recognition. Mapping targets of immune responses among the sequence of viral proteins is, thus, an important first step towards understanding the immune responses to viral variants and can aid in the identification of vaccine targets. Herein, we performed a large-scale, proteome-wide mapping and diversity analyses of putative HLA supertype-restricted T-cell epitopes of Zaire ebolavirus (ZEBOV), the most pathogenic species among the EBOV family. Methods All publicly available ZEBOV sequences (14,098) for each of the nine viral proteins were retrieved, removed of irrelevant and duplicate sequences, and aligned. The overall proteome diversity of the non-redundant sequences was studied by use of Shannon’s entropy. The sequences were predicted, by use of the NetCTLpan server, for HLA-A2, -A3, and -B7 supertype-restricted epitopes, which are relevant to African and other ethnicities and provide for large (~86%) population coverage. The predicted epitopes were mapped to the alignment of each protein for analyses of antigenic sequence diversity and relevance to structure and function. The putative epitopes were validated by comparison with experimentally confirmed epitopes. Results & discussion ZEBOV proteome was generally conserved, with an average entropy of 0.16. The 185 HLA supertype-restricted T-cell epitopes predicted (82 (A2), 37 (A3) and 66 (B7)) mapped to 125 alignment positions and covered ~24% of the proteome length. Many of the epitopes showed a propensity to co-localize at select positions of the alignment. Thirty (30) of the mapped positions were completely conserved and may be attractive for vaccine design. The remaining (95) positions had one or more epitopes, with or without non-epitope variants. A significant number (24) of the putative epitopes matched reported experimentally validated HLA ligands/T-cell epitopes of A2, A3 and/or B7 supertype representative allele restrictions. The epitopes generally corresponded to functional motifs/domains and there was no correlation to localization on the protein 3D structure. These data and the epitope map provide important insights into the interaction between EBOV and the host immune system.

An amendment to this paper has been published and can be accessed via the original article.

The study of viral diversity is imperative in understanding sequence change and its implications for intervention strategies. The widely used alignment-dependent approaches to study viral diversity are limited in their utility as sequence dissimilarity increases, particularly when expanded to the genus or higher ranks of viral species lineage. Herein, we present an alignment-independent algorithm, implemented as a tool, UNIQmin, to determine the effective viral sequence diversity at any rank of the viral taxonomy lineage. This is done by performing an exhaustive search to generate the minimal set of sequences for a given viral non-redundant sequence dataset. The minimal set is comprised of the smallest possible number of unique sequences required to capture the diversity inherent in the complete set of overlapping k-mers encoded by all the unique sequences in the given dataset. Such dataset compression is possible through the removal of unique sequences, whose entire repertoire of overlapping k-mers can be represented by other sequences, thus rendering them redundant to the collective pool of sequence diversity. A significant reduction, namely ~44%, ~45%, and ~53%, was observed for all reported unique sequences of species Dengue virus, genus Flavivirus, and family Flaviviridae, respectively, while still capturing the entire repertoire of nonamer (9-mer) viral peptidome diversity present in the initial input dataset. The algorithm is scalable for big data as it was applied to ~2.2 million non-redundant sequences of all reported viruses. UNIQmin is open source and publicly available on GitHub. The concept of a minimal set is generic and, thus, potentially applicable to other pathogenic microorganisms of non-viral origin, such as bacteria.

Abstract Background Viral vaccine target discovery requires understanding the diversity of both the virus and the human immune system. The readily available and rapidly growing pool of viral sequence data in the public domain enable the identification and characterization of immune targets relevant to adaptive immunity. A systematic bioinformatics approach is necessary to facilitate the analysis of such large datasets for selection of potential candidate vaccine targets. Results This work describes a computational methodology to achieve this analysis, with data of dengue, West Nile, hepatitis A, HIV-1, and influenza A viruses as examples. Our methodology has been implemented as an analytical pipeline that brings significant advancement to the field of reverse vaccinology, enabling systematic screening of known sequence data in nature for identification of vaccine targets. This includes key steps (i) comprehensive and extensive collection of sequence data of viral proteomes (the virome), (ii) data cleaning, (iii) large-scale sequence alignments, (iv) peptide entropy analysis, (v) intra- and inter-species variation analysis of conserved sequences, including human homology analysis, and (vi) functional and immunological relevance analysis. Conclusion These steps are combined into the pipeline ensuring that a more refined process, as compared to a simple evolutionary conservation analysis, will facilitate a better selection of vaccine targets and their prioritization for subsequent experimental validation.

Background and Purpose: To describe the final results of the TARGET Registry, a multicenter, real-world study of patients with intracranial aneurysms treated with new generation TARGET Coils.Methods: The TARGET Registry is a prospective, single-arm study with independent medical event monitoring and core-lab adjudication. Patients with de novo intracranial aneurysms were embolized with either TARGET-360° or helical coils in 12 US centers. The primary outcome was aneurysm packing density (PD), which was assessed immediately post-procedure. The secondary outcomes were immediate and long-term aneurysm occlusion rate using the Raymond Scale, and independent functional outcome using the modified Rankin Scale (mRS). A secondary analysis investigated the influence of the use of 100% 360-complex coils on clinical and angiographic outcomes.Results: 148 patients with 157 aneurysms met the inclusion and exclusion criteria. 58 (39.2%) patients with ruptured and 90 (61.8%) with unruptured aneurysms were treated using TARGET 360°, helical Coils, or both. Median age was 58.3 (IQR 48.1–67.4), 73% female, and 71.6% were Caucasian. Median follow-up time was 5.9 (IQR 4.0–6.9) months. The majority were treated with TARGET 360-coils (63.7%), followed by mixed and helical coils only. Peri-procedural morbidity and mortality was seen in 2.7% of patients. A good outcome at discharge (mRS 0–2) was seen in 89.9% of the full cohort, and in 84.5 and 93.3% in the ruptured and unruptured patients, respectively. The median packing density was 28.8% (IQR 20.3–41.1). Long-term complete and near complete occlusion rate was seen in 90.4% of aneurysms and complete obliteration was seen in 66.2% of the aneurysms. No significant difference in clinical and angiographic outcomes were noted between the pure 360-complex coiling vs. mixed 360-complex/Helical coiling strategies. In a multivariate analysis, predictors for long-term aneurysm occlusion were aneurysm location, immediate occlusion grade, and aneurysm size. The long-term independent functional outcome was achieved in 128/135 (94.8%) patients and all-cause mortality was seen in 3/148 (2%) patients.Conclusion: In the multicenter TARGET Registry, two-thirds of aneurysms achieved long-term complete occlusion and 91.0% achieved complete or near complete occlusion with excellent independent functional outcome.Clinical Trial Registration:www.ClinicalTrials.gov, identifier: NCT01748903

There is a growing attention toward personalized medicine. This is led by a fundamental shift from the ‘one size fits all’ paradigm for treatment of patients with conditions or predisposition to diseases, to one that embraces novel approaches, such as tailored target therapies, to achieve the best possible outcomes. Driven by these, several national and international genome projects have been initiated to reap the benefits of personalized medicine. Exome and targeted sequencing provide a balance between cost and benefit, in contrast to whole genome sequencing (WGS). Whole exome sequencing (WES) targets approximately 3% of the whole genome, which is the basis for protein-coding genes. Nonetheless, it has the characteristics of big data in large deployment. Herein, the application of WES and its relevance in advancing personalized medicine is reviewed. WES is mapped to Big Data “10 Vs” and the resulting challenges discussed. Application of existing biological databases and bioinformatics tools to address the bottleneck in data processing and analysis are presented, including the need for new generation big data analytics for the multi-omics challenges of personalized medicine. This includes the incorporation of artificial intelligence (AI) in the clinical utility landscape of genomic information, and future consideration to create a new frontier toward advancing the field of personalized medicine.

Avian influenza virus A (H7N9), after circulating in avian hosts for decades, was identified as a human pathogen in 2013. Herein, amino acid substitutions possibly essential for human adaptation were identified by comparing the 4706 aligned overlapping nonamer position sequences (1–9, 2–10, etc.) of the reported 2014 and 2017 avian and human H7N9 datasets. The initial set of virus sequences (as of year 2014) exhibited a total of 109 avian-to-human (A2H) signature amino acid substitutions. Each represented the most prevalent substitution at a given avian virus nonamer position that was selectively adapted as the corresponding index (most prevalent sequence) of the human viruses. The majority of these avian substitutions were long-standing in the evolution of H7N9, and only 17 were first detected in 2013 as possibly essential for the initial human adaptation. Strikingly, continued evolution of the avian H7N9 virus has resulted in avian and human protein sequences that are almost identical. This rapid and continued adaptation of the avian H7N9 virus to the human host, with near identity of the avian and human viruses, is associated with increased human infection and a predicted greater risk of human-to-human transmission.