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Extrachromosomal DNA (ecDNA) are circular DNA structures associated with cancer and drug resistance. One specific type, double minute (DM) chromosomes, has been studied since the 1960s using imaging techniques like cytogenetics and fluorescence microscopy. Specialized techniques such as atomic force microscopy (AFM) and scanning electron microscopy (SEM) offer micro to nano-scale visualization, but current sample preparation methods may not optimally preserve ecDNA structure. Our study introduces a systematic protocol using SEM for high-resolution ecDNA visualization. We have optimized the end-to-end procedure, providing a standardized approach to explore the circular architecture of ecDNA and address the urgent need for better understanding in cancer research.

Purpose Team-based care has been linked to key outcomes associated with the Quadruple Aim and a key driver of high-value patient-centered care. Use of the electronic health record (EHR) and machine learning have significant potential to overcome previous barriers to studying the impact of teams, including delays in accessing data to improve teamwork and optimize patient outcomes. Methods This study utilized a large EHR dataset (n = 316,542) from an urban health system to explore the relationship between team composition and patient activation, a key driver of patient engagement. Teams were operationalized using consensus definitions of teamwork from the literature. Patient activation was measured using the Patient Activation Measure (PAM). Results from multilevel regression analyses were compared to machine learning analyses using multinomial logistic regression to calculate propensity scores for the effect of team composition on PAM scores. Under the machine learning approach, a causal inference model with generalized overlap weighting was used to calculate the average treatment effect of teamwork. Results Seventeen different team types were observed in the data from the analyzed sample (n = 12,448). Team sizes ranged from 2 to 5 members. After controlling for confounding variables in both analyses, more diverse, multidisciplinary teams (team size of 4 or more) were observed to have improved patient activation scores. Conclusions This is the first study to explore the relationship between team composition and patient activation using the EHR and big data analytics. Implications for further research using EHR data and machine learning to study teams and other patient-centered care are promising and could be used to advance team science. (J Patient Cent Res Rev. 2024;11:18-28.)

Abstract Since their inception, small area studies intended to measure health system performance have been challenged by concerns that regional variation in health care may be primarily explained by differences in patient health risk. Controlling for regional population differences depends on appropriate risk adjustment, but the adequacy of the methods used in early analyses was contested. A novel response to these concerns was the development of end-of-life cohorts by Dartmouth Atlas investigators. These were used initially to control for differences in population health status in studies investigating relative efficiency across regions. Later, they became useful for studying hospital-level variation in chronic illness care, and for measuring utilization and patient experiences at the very end of life. Altogether, end-of-life cohorts have been invaluable for clarifying the contribution of health system and provider factors to health care variation and outcomes.

IntroductionClinical mastitis (CM) treatment decision-making is a multifaceted process that remains relatively understudied, despite CM being one of the most prevalent diseases on dairy farms worldwide, contributing greatly to the use of antimicrobials in the dairy industry. This study aimed to gain insights into decision-making mechanisms employed by dairy farmers in British Columbia and Alberta, Canada, when dealing with CM.MethodsInterviews were held with 15 dairy farmers in the Canadian provinces of British Columbia and Alberta and analyzed using reflexive thematic analysis to develop both the decision-pathway and overarching themes influencing the CM decisions by farmers in this region.Results and discussionThe analysis generated a decision-making process that begins with identification and classification of CM, guided by visual characteristics of milk and the udder, available milk production and quality data, presence of systemic signs, and additional diagnostics. Subsequently, CM cases are assessed based on the likelihood of cure, value of the cow, and herd goals to decide whether antimicrobial treatment is desired. Next, a treatment choice is made by evaluating severity and urgency of the case, availability of drugs and timing of the case. Finally, definition of treatment success and progression over time following the treatment decision guides the termination of treatment. Three overarching themes were generated that shape the decision-making process: ‘Personal attributes’, including personal approach and experiential knowledge; ‘Inter-actor dynamics’, such as shared decision-making and dynamics among producers, veterinarians, and milkers; and ‘Moving beyond protocols’, which highlights the dynamic nature of mastitis decision-making. These insights have the potential to inform the development of effective interventions to improve CM antimicrobial use that align with the reality of farming operations within Western Canada, and potentially beyond.

BackgroundTrachoma causes blindness due to repeated conjunctival infection by Chlamydia trachomatis (Ct). Transmission intensity is estimated, for programmatic decision-making, by prevalence of the clinical sign trachomatous inflammation-follicular (TF) in children aged 1-9 years. Research into complementary indicators to field-graded TF includes work on conjunctival photography, tests for ocular Ct infection, and serology. The perceived acceptability and feasibility of these indicators among a variety of stakeholders is unknown.MethodologyFocus group discussions (FGDs) with community members and in-depth interviews (IDIs) with public health practitioners in Tanzania were conducted. FGDs explored themes including participants' experience with, and thoughts about, different diagnostic approaches. The framework method for content analysis was used. IDIs yielded lists of perceived strengths of, and barriers to, implementation for programmatic use of each indicator. These were used to form an online quantitative survey on complementary indicators distributed to global stakeholders via meetings, mailing lists, and social media posts.ResultsSixteen FGDs and 11 IDIs were conducted in October-November 2022. In general, all proposed sample methods were deemed acceptable by community members. Common themes included not wanting undue discomfort and a preference for tests perceived as accurate. Health workers noted the importance of community education for some sample types. The online survey was conducted in April-May 2023 with 98 starting the questionnaire and 81 completing it. Regarding barriers to implementing diagnostics, the highest agreement items related to feasibility, rather than acceptability. No evidence of significant differences was found in responses pertaining to community acceptability based on participant characteristics.ConclusionsAll of the indicators included were generally deemed acceptable by all stakeholders in Tanzania, although community education around the benefits and risks of different sample types, as well as addressing issues around feasibility, will be key to successful, sustainable integration of these indicators into trachoma programs.

IntroductionExtracellular ATP (eATP) released from damaged cells activates the P2X7 receptor (P2X7R) ion channel on the surface of surrounding cells, resulting in calcium influx, potassium efflux and inflammasome activation. Inherited changes in the P2X7R gene (P2RX7) influence eATP induced responses. Single nucleotide polymorphisms (SNPs) of P2RX7 influence both function and signaling of the receptor, that in addition to ion flux includes pathogen control and immunity.MethodsSubjects (n = 105) were admitted to the ICU at the University Hospital Ulm, Germany between June 2018 and August 2019. Of these, subjects with a diagnosis of sepsis (n = 75), were also diagnosed with septic shock (n = 24), and/or pneumonia (n = 42). Subjects with pneumonia (n = 43) included those without sepsis (n = 1), sepsis without shock (n = 29) and pneumonia with septic shock (n = 13). Out of the 75 sepsis/septic shock patients, 33 patients were not diagnosed with pneumonia. Controls (n = 30) were recruited to the study from trauma patients and surgical patients without sepsis, septic shock, or pneumonia. SNP frequencies were determined for 16 P2RX7 SNPs known to affect P2X7R function, and association studies were performed between frequencies of these SNPs in sepsis, septic shock, and pneumonia compared to controls.ResultsThe loss-of-function (LOF) SNP rs17525809 (T253C) was found more frequently in patients with septic shock, and non-septic trauma patients when compared to sepsis. The LOF SNP rs2230911 (C1096G) was found to be more frequent in patients with sepsis and septic shock than in non-septic trauma patients. The frequencies of these SNPs were even higher in sepsis and septic patients with pneumonia. The current study also confirmed a previous study by our group that showed a five SNP combination that included the GOF SNPs rs208294 (C489T) and rs2230912 (Q460R) that was designated #21211 was associated with increased odds of survival in severe sepsis.DiscussionThe results found an association between expression of LOF P2RX7 SNPs and presentation to the ICU with sepsis, and septic shock compared to control ICU patients. Furthermore, frequencies of LOF SNPs were found to be higher in sepsis patients with pneumonia compared to those without pneumonia. In addition, a five SNP GOF combination was associated with increased odds of survival in severe sepsis. These results suggest that P2RX7 is required to control infection in pneumonia and that inheritance of LOF variants increases the risk of sepsis when associated with pneumonia. This study confirms that P2RX7 genotyping in pneumonia may identify patients at risk of developing sepsis. The study also identifies P2X7R as a target in sepsis associated with an excessive immune response in subjects with GOF SNP combinations.

IntroductionLeptin and its receptors are expressed by the human placenta throughout gestation, yet the role of leptin in early human placental development is not well characterized. Leptin is overexpressed in the placentas from preeclamptic (PE) pregnancies. PE can result from the impaired invasion of fetal placental cells, cytotrophoblasts (CTBs), into the maternal decidua. We hypothesized that elevated leptin levels would impair human CTB invasion.MethodsThe effects of leptin on the invasion of human CTBs were evaluated in three cell models, HTR-8/SVneo cells, primary CTBs, and placental villous explants using invasion assays. Further, leptin receptor expression was characterized in all three cell models using RT-PCR. Further phosphokinase assays were performed in HTR-8/SVneo cells to determine signaling pathways involved in CTB invasion in response to differential leptin doses.ResultsWe found that, prior to 8 weeks gestation, leptin promoted CTB invasion in the explant model. After 11 weeks gestation in explants, primary CTBs and in HTR-8/SVneo cells, leptin promoted invasion at moderate but not at high concentrations. Further, leptin receptor characterization revealed that leptin receptor expression did not vary over gestation, however, STAT, PI3K and MAPK pathways showed different signaling in response to varied leptin doses.DiscussionThese data suggest that the excess placental leptin observed in PE may cause impaired CTB invasion as a second-trimester defect. Leptin’s differential effect on trophoblast invasion may explain the role of hyperleptinemia in preeclampsia pathogenesis.

Objective The objective of our study is to explore Nepali women’s beliefs about access to mammography screening, and motivations to get screened or not. This work was intended to be hypothesis generating for subsequent quantitative analysis and to inform policy and decision-making to improve access. Methods We conducted structured qualitative interviews among nine Nepali women in the Northeast of the United States receiving care at a local community health center and among nine white women receiving mammography care at a large academic medical center in the Northeast. We analyzed the transcripts using a mixed deductive (content analysis) and inductive (grounded theory) approach. Deductive codes were generated from the Health Belief Model which states that a person’s belief in the real threat of a disease with their belief in the effectiveness of the recommended health service or behavior or action will predict the likelihood the person will adopt the behavior. We compared and contrasted qualitative results from both groups. Results We found that eligible Nepali women who had not received mammography screening had no knowledge of its availability and its importance. Primary care physicians emerged as a critical link in addressing this disparity: trust was found to be high among Nepali women with their established primary care provider. Conclusion The findings of this study suggest that the role of primary care practitioners in conversations around the importance and eligibility for mammography screening is of critical importance, especially for underserved groups with limited health knowledge of screening opportunities and potential health benefits. Follow-up research should focus on primary care practices.

Abstract Cardiac physiologic pacing (CPP), encompassing cardiac resynchronization therapy (CRT) and conduction system pacing (CSP), has emerged as a pacing therapy strategy that may mitigate or prevent the development of heart failure (HF) in patients with ventricular dyssynchrony or pacing‐induced cardiomyopathy. This clinical practice guideline is intended to provide guidance on indications for CRT for HF therapy and CPP in patients with pacemaker indications or HF, patient selection, pre‐procedure evaluation and preparation, implant procedure management, follow‐up evaluation and optimization of CPP response, and use in pediatric populations. Gaps in knowledge, pointing to new directions for future research, are also identified.

Objectives: As the world transitions to COVID-19 endemicity, studies focusing on aerosol shedding of highly transmissible SARS-CoV-2 variants of concern (VOCs) are vital for the calibration of infection control measures against VOCs that are likely to circulate seasonally. This follow-up Gesundheit-II aerosol sampling study aims to compare the aerosol shedding patterns of Omicron VOC samples with pre-Omicron variants analyzed in our previous study. Design: Coarse and fine aerosol samples from 47 patients infected with SARS-CoV-2 were collected during various respiratory activities (passive breathing, talking, and singing) and analyzed using reverse transcription-quantitative polymerase chain reaction and virus culture. Results: Compared with patients infected with pre-Omicron variants, comparable SARS-CoV-2 RNA copy numbers were detectable in aerosol samples of patients infected with Omicron despite being fully vaccinated. Patients infected with Omicron also showed a slight increase in viral aerosol shedding during breathing activities and were more likely to have persistent aerosol shedding beyond 7 days after disease onset. Conclusion: This follow-up study reaffirms the aerosol shedding properties of Omicron and should guide continued layering of public health interventions even in highly vaccinated populations.

Rates of fluoroquinolone resistance in Escherichia coli, a key opportunistic human pathogen, are problematic. Taking a One Health approach, we investigated the excretion of fluoroquinolone-resistant (FQ-R) E. coli by 600 dogs (303 from rural and 297 from urban environments) recruited from a 50 × 50 km region where we have also surveyed FQ-R E. coli from cattle and from human urine. FQ-R E. coli were detected in faeces from 7.3% (rural) and 11.8% (urban) of dogs. FQ-R E. coli from rural dogs tended to be of sequence types (STs) commonly excreted by cattle, whilst those from urban dogs tended to carry plasmid-mediated quinolone resistance genes, common in human E. coli in our study region. Phylogenetic evidence was obtained for sharing FQ-R E. coli - particularly for STs 10, 162 and 744 - between cattle, dogs and humans. Epidemiological analysis showed a strong association between feeding dogs uncooked meat and the excretion of FQ-R E. coli, particularly for STs 10, 162 and 744. This practice, therefore, could serve as a transmission link for FQ-R E. coli from farmed animals entering the home so we suggest that dogs fed uncooked meat should be handled and housed using enhanced hygiene practices.

OBJECTIVES/GOALS: We designed the Biocascade Exhaled Breath Sampler (BEBS) to characterize viral aerosol shedding among individuals with influenza and other respiratory virus infections. We first aimed to test the BEBS on volunteer COVID-19 cases and report the aerodynamic size distribution of exhaled breath aerosol particles carrying SARS-CoV-2 RNA. METHODS/STUDY POPULATION: From June 15 through December 15, 2022, we recruited 27 PCR-confirmed COVID-19 cases from a college campus and the surrounding community to provide 30-minute breath samples into a well-validated Gesundheit-II (G-II) exhaled breath aerosol sampler. Among these individuals, 17 provided an additional exhaled breath sample into the newly designed BEBS. We quantified samples for viral RNA using reverse transcription digital polymerase chain reaction (RT-dPCR) and determined the viral RNA copies collected within two aerosol size fractions (≤5 µm and >5 µm in diameter) from the G-II, and four aerosol size fractions (8.2 µm) from the BEBS. RESULTS/ANTICIPATED RESULTS: Individuals with a SARS-CoV-2 Omicron BA.4 or BA.5 infection shed virus in aerosols at an average rate of 7.5x103 RNA copies per 30-minute G-II sample, with 78% of the total RNA in aerosols ≤5 µm in diameter. Among the BEBS samples, 10% of the total viral RNA was detected in aerosols 8.2 µm size fraction. Based on viral RNA loads, our results indicate that exhaled aerosols ≤3.2 µm contribute the majority of SARS-CoV-2 inhalation exposure. DISCUSSION/SIGNIFICANCE: Our data provide additional evidence that respirable aerosols contribute to the spread of SARS-CoV-2. Thus, our data suggest that mitigation measures designed to reduce infectious aerosol inhalation, such as ventilation and the use of air cleaners and respirators, are needed to control the spread.

Summary: Background: While vaccines have established utility against COVID-19, phase 3 efficacy studies have generally not comprehensively evaluated protection provided by previous infection or hybrid immunity (previous infection plus vaccination). Individual patient data from US government-supported harmonized vaccine trials provide an unprecedented sample population to address this issue. We characterized the protective efficacy of previous SARS-CoV-2 infection and hybrid immunity against COVID-19 early in the pandemic over three-to six-month follow-up and compared with vaccine-associated protection. Methods: In this post-hoc cross-protocol analysis of the Moderna, AstraZeneca, Janssen, and Novavax COVID-19 vaccine clinical trials, we allocated participants into four groups based on previous-infection status at enrolment and treatment: no previous infection/placebo; previous infection/placebo; no previous infection/vaccine; and previous infection/vaccine. The main outcome was RT-PCR-confirmed COVID-19 >7–15 days (per original protocols) after final study injection. We calculated crude and adjusted efficacy measures. Findings: Previous infection/placebo participants had a 92% decreased risk of future COVID-19 compared to no previous infection/placebo participants (overall hazard ratio [HR] ratio: 0.08; 95% CI: 0.05–0.13). Among single-dose Janssen participants, hybrid immunity conferred greater protection than vaccine alone (HR: 0.03; 95% CI: 0.01–0.10). Too few infections were observed to draw statistical inferences comparing hybrid immunity to vaccine alone for other trials. Vaccination, previous infection, and hybrid immunity all provided near-complete protection against severe disease. Interpretation: Previous infection, any hybrid immunity, and two-dose vaccination all provided substantial protection against symptomatic and severe COVID-19 through the early Delta period. Thus, as a surrogate for natural infection, vaccination remains the safest approach to protection. Funding: National Institutes of Health.

Abstract Background Over the course of the COVID-19 pandemic, colleges and universities have focused on creating policies, such as mask mandates, to minimize COVID-19 transmission both on their campuses and in the surrounding community. Adherence to and opinions about these policies remain largely unknown. Methods The Centers for Disease Control and Prevention (CDC) developed a cross-sectional study, the Mask Adherence and Surveillance at Colleges and Universities Project (MASCUP!), to objectively and inconspicuously measure rates of mask use at institutes of higher education via direct observation. From February 15 through April 11, 2021 the University of Colorado Boulder (CU, n = 2,808 observations) and Colorado State University Fort Collins (CSU, n = 3,225 observations) participated in MASCUP! along with 52 other institutes of higher education (n = 100,353 observations) spanning 21 states and the District of Columbia. Mask use was mandatory at both Colorado universities and student surveys were administered to assess student beliefs and attitudes. Results We found that 91.7%, 93.4%, and 90.8% of persons observed at indoor locations on campus wore a mask correctly at University of Colorado, Colorado State University, and across the 52 other schools, respectively. Student responses to questions about masking were in line with these observed rates of mask use where 92.9% of respondents at CU and 89.8% at CSU believe that wearing masks can protect the health of others. Both Colorado universities saw their largest surges in COVID-19 cases in the fall of 2020, with markedly lower case counts during the mask observation window in the spring of 2021. Conclusion High levels of mask use at Colorado’s two largest campuses aligned with rates observed at other institutes across the country. These high rates of use, coupled with positive student attitudes about mask use, demonstrate that masks were widely accepted and may have contributed to reduced COVID-19 case counts. This study supports an emerging body of literature substantiating masks as an effective, low-cost measure to reduce disease transmission and establishes masking (with proper education and promotion) as a viable tactic to reduce respiratory disease transmission on college campuses.

Abstract HSP90 inhibitors destabilize oncoproteins associated with cell cycle, angiogenesis, RAS-MAPK activity, histone modification, kinases and growth factors. We evaluated the HSP90-inhibitor ganetespib in combination with standard chemotherapy in patients with high-risk early-stage breast cancer. I-SPY2 is a multicenter, phase II adaptively randomized neoadjuvant (NAC) clinical trial enrolling patients with stage II-III breast cancer with tumors 2.5 cm or larger on the basis of hormone receptors (HR), HER2 and Mammaprint status. Multiple novel investigational agents plus standard chemotherapy are evaluated in parallel for the primary endpoint of pathologic complete response (pCR). Patients with HER2-negative breast cancer were eligible for randomization to ganetespib from October 2014 to October 2015. Of 233 women included in the final analysis, 140 were randomized to the standard NAC control; 93 were randomized to receive 150 mg/m2 ganetespib every 3 weeks with weekly paclitaxel over 12 weeks, followed by AC. Arms were balanced for hormone receptor status (51–52% HR-positive). Ganetespib did not graduate in any of the biomarker signatures studied before reaching maximum enrollment. Final estimated pCR rates were 26% vs. 18% HER2-negative, 38% vs. 22% HR-negative/HER2-negative, and 15% vs. 14% HR-positive/HER2-negative for ganetespib vs control, respectively. The predicted probability of success in phase 3 testing was 47% HER2-negative, 72% HR-negative/HER2-negative, and 19% HR-positive/HER2-negative. Ganetespib added to standard therapy is unlikely to yield substantially higher pCR rates in HER2-negative breast cancer compared to standard NAC, and neither HSP90 pathway nor replicative stress expression markers predicted response. HSP90 inhibitors remain of limited clinical interest in breast cancer, potentially in other clinical settings such as HER2-positive disease or in combination with anti-PD1 neoadjuvant chemotherapy in triple negative breast cancer. Trial registration: www.clinicaltrials.gov/ct2/show/NCT01042379

The QT interval is a heritable electrocardiographic measure associated with arrhythmia risk when prolonged. Here, the authors used a series of genetic analyses to identify genetic loci, pathways, therapeutic targets, and relationships with cardiovascular disease.

Gastrointestinal (GI) cancers, including esophageal, gastroesophageal junction, gastric, duodenal and distal small bowel, biliary tract, pancreatic, colon, rectal, and anal cancer, comprise a heterogeneous group of malignancies that impose a significant global burden. Immunotherapy has transformed the treatment landscape for several GI cancers, offering some patients durable responses and prolonged survival. Specifically, immune checkpoint inhibitors (ICIs) directed against programmed cell death protein 1 (PD-1), either as monotherapies or in combination regimens, have gained tissue site-specific regulatory approvals for the treatment of metastatic disease and in the resectable setting. Indications for ICIs in GI cancer, however, have differing biomarker and histology requirements depending on the anatomic site of origin. Furthermore, ICIs are associated with unique toxicity profiles compared with other systemic treatments that have long been the mainstay for GI cancer, such as chemotherapy. With the goal of improving patient care by providing guidance to the oncology community, the Society for Immunotherapy of Cancer (SITC) convened a panel of experts to develop this clinical practice guideline on immunotherapy for the treatment of GI cancer. Drawing from published data and clinical experience, the expert panel developed evidence- and consensus-based recommendations for healthcare professionals using ICIs to treat GI cancers, with topics including biomarker testing, therapy selection, and patient education and quality of life considerations, among others.

As COVID-19 vaccine coverage increases, screening programs that focus on the unvaccinated population test fewer individuals. This modeling study identifies vaccine coverage and effectiveness scenarios where such programs are (and are not) impactful.

Abstract Background Color groups of fruits and vegetables (FV) are part of a healthy diet, but evidence for an association with cardiometabolic outcomes is inconsistent. Objective To examine the association between intake of FV of different colors with incident diabetes and cardiometabolic risk biomarkers among U.S. Hispanics/Latinos. Subjects/methods We used data from 9206 adults ages 18–74 years who were free of diabetes at baseline (2008–2011) and had follow-up data at visit 2 (2014–2017) in the Hispanic Community Health Study/Study of Latinos (HCHS/SOL), a multicenter, prospective cohort study of self-identified Hispanics/Latinos. Dietary intake was assessed using two 24 h recalls at baseline. FV were categorized into five color groups: green, white, yellow/orange, red/purple, and uncategorized. Diabetes was defined based on laboratory measures and self-reported antihyperglycemic medication. We used survey logistic regression models to evaluate the association between FV color groups and incident diabetes and survey linear regression models to evaluate the association of FV color groups with cardiometabolic risk biomarkers at visit 2. Results During ~6 years of follow-up, 970 incident cases of diabetes were documented. The red/purple FV color group was the least consumed (0.21 servings/day), whereas white FV were the most consumed (0.92 servings/day). For each serving of total FV intake, body mass index (BMI) was lower by 0.24% (p = 0.03) and insulin by 0.69% (p = 0.03). For each serving of red/purple FV intake, HDL was 1.59% higher (p = 0.04). For each serving of white FV intake (with potato), post-OGTT was 0.83% lower (p = 0.04) and triglycerides 1.43% lower (p = 0.04). There was no association between FV intake and incident diabetes. Conclusions Specific FV colors were associated with cardiometabolic benefits though the associations were of relatively small magnitudes. Dietary recommendations could consider varying colors of FV intake, especially white and red/purple color groups, for a healthy diet.

Antimicrobials are essential in veterinary medicine to treat and control bacterial disease in animals. Their prudent use in food-producing animals has been encouraged to reduce the development and spread of antimicrobial resistance. National and international guidelines for responsible antimicrobial use have been developed as tools to guide and rationalise antimicrobial prescribing decisions by veterinarians and usage decisions by farmers. Yet, there is little understanding of whether these existing guidelines are fit for purpose. Accordingly, this study rigorously assessed 128 veterinary guidelines for antimicrobial use in ruminants in the UK, following established qualitative methodologies. Findings revealed four pertinent themes: validation of the veterinarian as the prescriber, visibility of responsible use realities, vagueness in interpretation and variation in directing behaviour. These themes encompassed the roles and responsibilities of the veterinarian and the realities of prescribing scenarios, alongside concerns relating to the specificity within and variation between guidelines. Resultant recommendations to inform and support the future development of guidelines include establishing species-specific and disease-specific guidelines, expanding guidelines to include disease prevention measures, including definitions to resolve vagueness and promoting congruence in interpretation, encouraging the development of practice-level guidelines to endorse collaboration and ownership, and fostering active working between stakeholders to align priorities and messaging.

Abstract I-SPY2 is an adaptively randomized phase 2 clinical trial evaluating novel agents in combination with standard-of-care paclitaxel followed by doxorubicin and cyclophosphamide in the neoadjuvant treatment of breast cancer. Ganitumab is a monoclonal antibody designed to bind and inhibit function of the type I insulin-like growth factor receptor (IGF-1R). Ganitumab was tested in combination with metformin and paclitaxel (PGM) followed by AC compared to standard-of-care alone. While pathologic complete response (pCR) rates were numerically higher in the PGM treatment arm for hormone receptor-negative, HER2-negative breast cancer (32% versus 21%), this small increase did not meet I-SPY’s prespecified threshold for graduation. PGM was associated with increased hyperglycemia and elevated hemoglobin A1c (HbA1c), despite the use of metformin in combination with ganitumab. We evaluated several putative predictive biomarkers of ganitumab response (e.g., IGF-1 ligand score, IGF-1R signature, IGFBP5 expression, baseline HbA1c). None were specific predictors of response to PGM, although several signatures were associated with pCR in both arms. Any further development of anti-IGF-1R therapy will require better control of anti-IGF-1R drug-induced hyperglycemia and the development of more predictive biomarkers.

Abstract Background The incidence of colorectal cancer in adults younger than age 50 has increased with rates expected to continue to increase over the next decade. The objective of this study is to examine the survival benefit of surgical resection (primary and/or metastatic) versus palliative therapy in this patient population. Methods We identified 6708 young adults aged 18–45 years diagnosed with metastatic colorectal cancer (mCRC) from 2004 to 2015 from the SEER database. Overall survival (OS) was analyzed using Kaplan–Meier estimation, log rank test, and multivariate Cox proportional hazards model. Results Sixty‐three percent of patients in our study underwent primary tumor resection (PTR), with 40% undergoing PTR alone and 23% undergoing both resection of primary disease and metastasectomy. The median OS for patients who underwent both PTR and metastasectomy was 36 months, compared to 13 months for those who did not receive any surgical intervention. The multivariate analysis showed significant OS benefit of receiving both PTR and metastasectomy (HR 0.34, 95% CI: 0.31–0.37, p

Background There is evidence of the occurrence of trachoma in Peru, and studies have shown that soil-transmitted helminthiases (STH) are affecting rural communities in the Amazon basin in Loreto Department. This study was done to estimate trachoma prevalence, STH prevalence, and the associated factors for both diseases in children aged 1–9 years in rural communities of Peru. Methodology A population-based cross-sectional survey was carried out in rural communities of Loreto. A standardized survey questionnaire with individual and household risk factors related to both diseases was used. Ocular examination was done for all participants aged one year and above, and eye swab samples were collected from children with follicular trachoma (TF). Anthropometric measurements, stool samples for STH, and blood samples for hemoglobin measurement were taken from children. Principal findings TF prevalence was 7.74% (95% CI 5.08–11.63%), STH prevalence was 49.49% (95% CI 25.00–52.43%), and prevalence of co-occurrence of both diseases was 5.06% (95% CI 2.80–8.98%) in children aged 1–9 years. Being at age 3–8 years old (AOR = 6.76; 95% CI 1.346–33.947), have an unclean face (AOR = 24.64; 95% CI 6.787–89.444), and having been dewormed in the last six months (AOR = 2.47; 95% CI 1.106–5.514), were risk factors of TF. Being a female (AOR = 0.22; 95% CI 0.103–0.457) was associated with decreased odds of TF. Having been dewormed in the last six months (AOR = 0.30; 95% CI 0.139–0.628) was a preventative factor for STH. Risk factors for children with both diseases mirrored the findings for risk factors for individual diseases. Conclusions Neglected tropical diseases and associated risk factors overlap in communities living in vulnerable conditions in the Amazon basin of Peru. These findings support the need to implement integrated interventions, including mass drug administration, water, sanitation, and hygiene for both diseases in the study area. Author summary Trachoma is an infectious disease caused by the bacterium Chlamydia trachomatis, transmitted through person-to-person contact, and it is the world’s leading infectious cause of blindness. Soil-transmitted helminths (STH) is a group of intestinal parasites composed of Ascaris lumbricoides (roundworms), Trichuris trichiura (whipworms), and Ancylostoma duodenale and Necator Americanus (hookworms), that are transmitted by fecal contamination of the soil and adversely affect nutritional status and impair cognitive processes in children. Although there is evidence of the occurrence of trachoma in several areas of Peru, an epidemiological study has not been carried out to determine if it is a public health problem. STH infections have been reported to affect children in areas of the Loreto Department in Peru, but prevalence surveys in rural communities bordering Brazil and Colombia have not been carried out. In this study, we present the results of an integrated survey for trachoma and STH in Loreto and the factors associated with the occurrence of these diseases. The results of this study show that both diseases and risk factors overlap, supporting the need to implement integrated interventions to eliminate them as public health problems in communities living in vulnerable conditions in the Amazon basin of Peru.

BackgroundGreat progress has been made toward the elimination of trachoma as a public-health problem. Mathematical and statistical models have been used to forecast when the program will attain the goal of the elimination of active trachoma, defined as prevalence of trachomatous inflammation-follicular in 1-9 year olds (TF1-9) Methodology/principal findingsWe calculated the mean number of years (95% CI) observed for an implementation unit (IU) to move from a baseline TF1-9 prevalence ≥5% to the elimination threshold, based on the region (Ethiopia vs. non-Ethiopia) and baseline prevalence category. Ethiopia IUs had significantly different rates of reaching the TF1-9 elimination threshold after a trachoma impact survey (TIS) compared to non-Ethiopia IUs across all baseline categories. We used those estimates to predict when remaining active trachoma-endemic IUs (TF1-9 ≥5%) would have their last round of mass drug administration (MDA) based on the mean number of years required and number of MDA rounds already completed. Our model predicts that elimination of TF1-9 will be achieved in 2028 in Ethiopia (95% CI: 2026-2033) and 2029 outside of Ethiopia (95% CI: 2023-2034), with some IUs in East Africa predicted to be the last requiring MDA globally.Conclusions/significanceOur empirical estimate is similar to those resulting from previous susceptible-infectious-susceptible (SIS) and mathematical models, suggesting that the forecast achievement of TF1-9 elimination is realistic with the caveat that although disease elimination progress can be predicted for most IUs, there is an important minority of IUs that is not declining or has not yet started trachoma elimination activities. These IUs represent an important barrier to the timely global elimination of active trachoma.

We report a survey (August 2017 to March 2018) and risk factor analysis of faecal carriage of antibacterial-resistant (ABR) Escherichia coli in 223 16-week-old dogs in the United Kingdom. Raw feeding was associated with the presence of fluoroquinolone-resistant (FQ-R) E. coli and those resistant to tetracycline, amoxicillin, and streptomycin, but not to cefalexin. Whole genome sequencing of 36 FQ-R E. coli isolates showed a wide range of sequence types (STs), with almost exclusively mutational FQ-R dominated by ST744 and ST162. Comparisons between E. coli isolates from puppies known to be located within a 50 × 50 km region with those isolated from human urinary tract infections (isolated in parallel in the same region) identified an ST744 FQ-R lineage that was carried by one puppy and caused one urinary tract infection. Accordingly, we conclude that raw feeding is associated with carriage of ABR E. coli in dogs even at 16 weeks of age and that bacteria carried by puppies are shared with humans. We therefore suggest that those who feed their dogs raw meat seriously consider the potential ABR-transmission threat their pet may become as a result and deploy appropriate hygiene practices in mitigation.

Dengue virus causes a range of inflammatory pathology but understanding critical phases of the infection during human infection has been challenging. Here the author’s present immunotranscriptomic changes during the acute and clearance phases of a dengue virus serotype 2 human challenge model.

Antimicrobial resistance (AMR) is a global health and development threat, with calls for the optimisation of antimicrobial use (AMU) in the treatment of both humans and animals prevalent across national and international policy. Rapid, low-cost and readily available diagnostics that specifically identify pathogens and their antimicrobial susceptibility profiles have been identified as essential parts of this optimisation process, yet questions over the assumed utility of novel rapid technology as a cornerstone of tackling agricultural AMU still exist. To understand whether this technology may support the optimisation of AMU in the treatment of animal disease, this study qualitatively examines the discourse between veterinarians, laboratory representatives, veterinary researchers and (cattle) farmers within three participatory events concerning diagnostic testing on UK farms, to offer a critical examination of the interaction between veterinary diagnostic practice and agricultural AMU. Veterinarian-led discussion suggested that veterinary rationales for engaging with diagnostic testing are nuanced and complex, where veterinarians (i) were driven by both medical and non-medical motivators; (ii) had a complex professional identity influencing diagnostic-test engagement; and (iii) balanced a multitude of situated contextual factors that informed “gut feelings” on test choice and interpretation. In consequence, it is suggested that data-driven diagnostic technologies may be more palatable for veterinarians to promote to their farm clients in the pursuit of better and more sustainable AMU, whilst also being in synergy with the emerging preventative role of the farm veterinarian.

Multivalent vaccines that confer protection to multiple serotypes of Dengue virus have been established. Here the authors examine the presence of vaccine induced multivalent antibodies and how these link to protection in a human challenge model of Dengue virus.

Glucocorticoids (GC) are commonly used to suppress undesirable inflammatory responses. Here the authors show, using hi-dimensional flow cytometry data, that GC treatment following major surgeries alters adaptive immunity without significant modulation of innate immune responses or pain/functional impairment.

Purpose: People of color often face challenges in accessing equitable healthcare. Disparities in healthcare pose very real moral and ethical social justice dilemmas for society, and prevent efforts to improve the nation's health and manage escalating healthcare costs. A diverse healthcare workforce is necessary as a means to help care for an increasingly diverse patient population. Method: This paper focuses on programmatic and research information that is a collaborative effort between a number of researchers and educators in schools of medicine and allied healthcare. The paper looks at the current state of racial and ethnic diversity in the health professions and describes the social justice implications of a representative healthcare workforce. Using a “pipeline to practice” model, the authors will present information spanning the pipeline from encouraging high school students of color to enter the allied healthcare professions to introducing undergraduate and graduate students in health professions program to responsive policy making and cross-cultural communication. The authors reviewed the research literature across multiple institutions and professional health programs, and include illustrative case studies. Results: The authors found that overall, the healthcare workforce is becoming more diverse however, with the majority of people of color in healthcare jobs remaining in entry-level and often lower paying jobs. The need to increase the diversity of the healthcare workforce in all fields of allied health is a continuing need. The most promising practices tended to be comprehensive programs that include a combination of social support, academic support, and financial support. Discussion: This information has great significance for health professions education programs as they strive to diversify their student populations, retain students of color, and provide culturally responsive education and training. This interdisciplinary collaborative perspective illustrates what can be learned from varied health professional programs as well as making new connections across often disconnected practice settings.

Fusion-positive rhabdomyosarcoma (FP-RMS) is driven by a translocation that creates the chimeric transcription factor PAX3-FOXO1 (P3F), which assembles de novo super enhancers to drive high levels of transcription of other core regulatory transcription factors (CRTFs). P3F recruits co-regulatory factors to super enhancers such as BRD4, which recognizes acetylated lysines via BET bromodomains. In this study, we demonstrate that inhibition or degradation of BRD4 leads to global decreases in transcription, and selective downregulation of CRTFs. We also show that the BRD4 degrader ARV-771 halts transcription while preserving RNA Polymerase II (Pol2) loops between super enhancers and their target genes, and causes the removal of Pol2 only past the transcriptional end site of CRTF genes, suggesting a novel effect of BRD4 on Pol2 looping. We finally test the most potent molecule, inhibitor BMS-986158, in an orthotopic PDX mouse model of FP-RMS with additional high-risk mutations, and find that it is well tolerated in vivo and leads to an average decrease in tumor size. This effort represents a partnership with an FP-RMS patient and family advocates to make preclinical data rapidly accessible to the family, and to generate data to inform future patients who develop this disease.

Ozone (O3) is an air pollutant that primarily damages the lungs, but growing evidence supports the idea that O3 also harms the brain; acute exposure to O3 has been linked to central nervous system (CNS) symptoms such as depressed mood and sickness behaviors. However, the mechanisms by which O3 inhalation causes neurobehavioral changes are limited. One hypothesis is that factors in the circulation bridge communication between the lungs and brain following O3 exposure. In this study, our goals were to characterize neurobehavioral endpoints of O3 exposure as they relate to markers of systemic and pulmonary inflammation, with a particular focus on serum amyloid A (SAA) and kynurenine as candidate mediators of O3 behavioral effects. We evaluated O3-induced dose-, time- and sex-dependent changes in pulmonary inflammation, circulating SAA and kynurenine and its metabolic enzymes, and sickness and depressive-like behaviors in Balb/c and CD-1 mice. We found that 3 parts per million (ppm) O3, but not 2 or 1 ppm O3, increased circulating SAA and lung inflammation, which were resolved by 48 h and was worse in females. We also found that indoleamine 2,3-dioxygenase (Ido1) mRNA expression was increased in the brain and spleen 24 h after 3 ppm O3 and that kynurenine was increased in blood. Sickness and depressive-like behaviors were observed at all O3 doses (1–3 ppm), suggesting that behavioral responses to O3 can occur independently of increased SAA or neutrophils in the lungs. Using SAA knockout mice, we found that SAA did not contribute to O3-induced pulmonary damage or inflammation, systemic increases in kynurenine post-O3, or depressive-like behavior but did contribute to weight loss. Together, these findings indicate that acute O3 exposure induces transient symptoms of sickness and depressive-like behaviors that may occur in the presence or absence of overt pulmonary neutrophilia and systemic increases of SAA. SAA does not appear to contribute to pulmonary inflammation induced by O3, although it may contribute to other aspects of sickness behavior, as reflected by a modest effect on weight loss.

Abstract Background Myogenesis is driven by specific changes in the transcriptome that occur during the different stages of muscle differentiation. In addition to controlled transcriptional transitions, several other post-transcriptional mechanisms direct muscle differentiation. Both alternative splicing and miRNA activity regulate gene expression and production of specialized protein isoforms. Importantly, disruption of either process often results in severe phenotypes as reported for several muscle diseases. Thus, broadening our understanding of the post-transcriptional pathways that operate in muscles will lay the foundation for future therapeutic interventions. Methods We employed bioinformatics analysis in concert with the well-established C2C12 cell system for predicting and validating novel miR-1 and miR-206 targets engaged in muscle differentiation. We used reporter gene assays to test direct miRNA targeting and studied C2C12 cells stably expressing one of the cDNA candidates fused to a heterologous, miRNA-resistant 3′ UTR. We monitored effects on differentiation by measuring fusion index, myotube area, and myogenic gene expression during time course differentiation experiments. Results Gene ontology analysis revealed a strongly enriched set of putative miR-1 and miR-206 targets associated with RNA metabolism. Notably, the expression levels of several candidates decreased during C2C12 differentiation. We discovered that the splicing factor Srsf9 is a direct target of both miRNAs during myogenesis. Persistent Srsf9 expression during differentiation impaired myotube formation and blunted induction of the early pro-differentiation factor myogenin as well as the late differentiation marker sarcomeric myosin, Myh8. Conclusions Our data uncover novel miR-1 and miR-206 cellular targets and establish a functional link between the splicing factor Srsf9 and myoblast differentiation. The finding that miRNA-mediated clearance of Srsf9 is a key myogenic event illustrates the coordinated and sophisticated interplay between the diverse components of the gene regulatory network.

We examined a collection of 386 animal, 451 human, and 109 archived bioaerosol samples with a new pan-species coronavirus molecular assay. Thirty-eight (4.02%) of 946 specimens yielded evidence of human or animal coronaviruses. Our findings demonstrate the utility of employing the pan-CoV RT-PCR assay in detecting varied coronavirus among human, animal, and environmental specimens. This RT-PCR assay might be employed as a screening diagnostic for early detection of coronaviruses incursions or prepandemic coronavirus emergence in animal or human populations.

A continuing challenge in modern medicine is the identification of safer and more efficacious drugs. Precision therapeutics, which have one molecular target, have been long promised to be safer and more effective than traditional therapies. This approach has proven to be challenging for multiple reasons including lack of efficacy, rapidly acquired drug resistance, and narrow patient eligibility criteria. An alternative approach is the development of drugs that address the overall disease network by targeting multiple biological targets ('polypharmacology'). Rational development of these molecules will require improved methods for predicting single chemical structures that target multiple drug targets. To address this need, we developed the Multi-Targeting Drug DREAM Challenge, in which we challenged participants to predict single chemical entities that target pro-targets but avoid anti-targets for two unrelated diseases: RET-based tumors and a common form of inherited Tauopathy. Here, we report the results of this DREAM Challenge and the development of two neural network-based machine learning approaches that were applied to the challenge of rational polypharmacology. Together, these platforms provide a potentially useful first step towards developing lead therapeutic compounds that address disease complexity through rational polypharmacology.

Preeclampsia is a pregnancy-specific disorder involving placental abnormalities. Elevated placental Sialic acid immunoglobulin-like lectin (Siglec)-6 expression has been correlated with preeclampsia. Siglec-6 is a transmembrane receptor, expressed predominantly by the trophoblast cells in the human placenta. It interacts with sialyl glycans such as sialyl-TN glycans as well as binds leptin. Siglec-6 overexpression has been shown to influence proliferation, apoptosis, and invasion in the trophoblast (BeWo) cell model. However, there is no direct evidence that Siglec-6 plays a role in preeclampsia pathogenesis and its signaling potential is still largely unexplored. Siglec-6 contains an immunoreceptor tyrosine-based inhibitory motif (ITIM) and an ITIM-like motif in its cytoplasmic tail suggesting a signaling function. Site-directed mutagenesis and transfection were employed to create a series of Siglec-6 expressing HTR-8/SVneo trophoblastic cell lines with mutations in specific functional residues to explore the signaling potential of Siglec-6. Co-immunoprecipitation and inhibitory assays were utilized to investigate the association of Src-kinases and SH-2 domain-containing phosphatases with Siglec-6. In this study, we show that Siglec-6 is phosphorylated at ITIM and ITIM-like domains by Src family kinases. Phosphorylation of both ITIM and ITIM-like motifs is essential for the recruitment of phosphatases like Src homology region 2 containing protein tyrosine phosphatase 2 (SHP-2), which has downstream signaling capabilities. These findings suggest Siglec-6 as a signaling molecule in human trophoblasts. Further investigation is warranted to determine which signaling pathways are activated downstream to SHP-2 recruitment and how overexpression of Siglec-6 in preeclamptic placentas impacts pathogenesis.

Objectives: Antibacterial resistance (ABR) is a major global health security threat, with a disproportionate burden on lower-and middle-income countries (LMICs). It is not understood how ‘One Health’, where human health is co-dependent on animal health and the environment, might impact the burden of ABR in LMICs. Thailand's 2017 “National Strategic Plan on Antimicrobial Resistance” (NSP-AMR) aims to reduce AMR morbidity by 50% through 20% reductions in human and 30% in animal antibacterial use (ABU). There is a need to understand the implications of such a plan within a One Health perspective. Methods: A model of ABU, gut colonisation with extended-spectrum beta-lactamase (ESBL)-producing bacteria and transmission was calibrated using estimates of the prevalence of ESBL-producing bacteria in Thailand. This model was used to project the reduction in human ABR over 20 years (2020–2040) for each One Health driver, including individual transmission rates between humans, animals and the environment, and to estimate the long-term impact of the NSP-AMR intervention. Results: The model predicts that human ABU was the most important factor in reducing the colonisation of humans with resistant bacteria (maximum 65.7–99.7% reduction). The NSP-AMR is projected to reduce human colonisation by 6.0–18.8%, with more ambitious targets (30% reductions in human ABU) increasing this to 8.5–24.9%. Conclusions: Our model provides a simple framework to explain the mechanisms underpinning ABR, suggesting that future interventions targeting the simultaneous reduction of transmission and ABU would help to control ABR more effectively in Thailand.

Purpose: Limited research examining the relationship between team-based models of care and patient satisfaction in the hospital setting is available. The purpose of this literature review was to explore this relationship as well as the relationships between team composition, team-based interventions, patient satisfaction, and other outcomes of care when measured as part of the study. Methods: A systematic appraisal of research studies published through February 2017 was conducted using PubMed, Cochrane Library, CINAHL, Embase, Ovid, gray literature and Google Scholar. Inclusion criteria were 1) experimental (randomized control trials), quasi-experimental, or non-experimental (cross-sectional) study design; 2) team-based care interventions; 3) hospital setting; 4) patient satisfaction measured as an outcome; and 5) published in English. Results: The literature search yielded 15,247 citations. In total, 142 articles were retrieved for full-text screening; 21 studies met inclusion criteria. Overall, 57% of the studies identified a statistically significant improvement in patient satisfaction associated with team-based care. Team-based care interventions ranged from single team activities such as multidisciplinary rounds to comprehensive team-based models of care. Patient satisfaction scores were greater with teams that had more than two professions and more comprehensive team-based models. About one-quarter of studies that measured patient satisfaction and at least one additional outcome demonstrated improvement in both. Conclusions: Team-based care may positively affect patient satisfaction. Team composition and type of team intervention appears to influence the strength of the relationship. Improvements in satisfaction are not consistently accompanied by improvements in other outcomes.

Background: Acute viral infections induce a rapid and transient increase in antibody-secreting plasmablasts. At convalescence, memory B cells (MBC) and long-lived plasma cells (LLPC) are responsible for long-term humoral immunity. Following an acute viral infection, the specific properties and relationships between antibodies produced by these B cell compartments are poorly understood. Methods: We utilized a controlled human challenge model of primary dengue virus serotype 2 (DENV2) infection to study acute and convalescent B-cell responses. Findings: The level of DENV2 replication was correlated with the magnitude of the plasmablast response. Functional analysis of plasmablast-derived monoclonal antibodies showed that the DENV2-specific response was dominated by cells producing DENV2 serotype-specific antibodies. DENV2-neutralizing antibodies targeted quaternary structure epitopes centered on domain III of the viral envelope protein (EDIII). Functional analysis of MBC and serum antibodies from the same subjects six months post-challenge revealed maintenance of the serotype-specific response in both compartments. The serum response mainly targeted DENV2 serotype-specific epitopes on EDIII. Interpretation: Our data suggest overall functional alignment of DENV2-specific responses from the plasmablast, through the MBC and LLPC compartments following primary DENV2 inflection. These results provide enhanced resolution of the temporal and specificity of the B cell compartment in viral infection and serve as framework for evaluation of B cell responses in challenge models. Funding: This study was supported by the Bill and Melinda Gates Foundation and the National Institutes of Health. Keywords: Dengue, Viral infection, Humoral immunity, Antibody

The P2X7 receptor (P2X7R) is an ATP-gated membrane ion channel that is expressed by multiple cell types. Following activation by extracellular ATP, the P2X7R mediates a broad range of cellular responses including cytokine and chemokine release, cell survival and differentiation, the activation of transcription factors, and apoptosis. The P2X7R is made up of three P2X7 subunits that contain specific domains essential for the receptor’s varied functions. Alternative splicing produces P2X7 isoforms that exclude one or more of these domains and assemble in combinations that alter P2X7R function. The modification of the structure and function of the P2X7R may adversely affect cellular responses to carcinogens and pathogens, and alternatively spliced (AS) P2X7 isoforms have been associated with several cancers. This review summarizes recent advances in understanding the structure and function of AS P2X7 isoforms and their associations with cancer and potential role in modulating the inflammatory response.

In this paper we consider the shifting role, practice and context of veterinary diagnosis in addressing concerns over what is, in the context of the growing threat of antimicrobial resistance, considered unnecessary or excessive antimicrobial medicine use in UK livestock farms. With increasing policy and regulatory interest in diagnostic practices and technologies, coupled with an expanding focus on the development and deployment of new rapid and point-of-care on-farm diagnostic testing, this paper investigates current diagnostic practices amongst veterinarians working on dairy, pig and poultry farms in Great Britain (England, Wales, and Scotland) and, more specifically, veterinarians' use and perceptions of new and emerging rapid and point-of-care diagnostic tests. Drawing on a series of 30 semi-structured interviews with farm animal veterinary professionals across the three sectors, this paper examines the manner in which such tests are both used and anticipated in clinical farm animal veterinary practice and the possible impact rapid test technologies might have on broader farm animal health management and disease control. Analysis of the transcribed interviews reveals a number of complexities around the use of rapid and point-of-care diagnostic tests. The relative rapidity and simplification of such tests, facilitating immediate treatment responses, is held in balance against both the accuracy and the more detailed and documented procedures of established laboratory testing routes. In situations of multifaceted on-farm etiologies, respondents maintained that rapid tests may offer restricted diagnostic capabilities, though in other situations they were found to offer ready confirmation of disease presence. A third complexity arising from the growth of rapid and point-of-care testing and revealed in this study relates to the shifting distribution of responsibilities in animal health care within contemporary food chains. The growing availability of rapid and point-of-care tests effectively diversifies the range of diagnostic actors with consequences for the flow of diagnostic and disease information. The veterinarians in this study identified areas where new rapid and point-of-care tests would be of particular value to them in their clinical practice particularly in addressing concerns over inappropriate antimicrobial use in animal treatment. However, despite the considerable policy advocacy on rapid and point-of-care tests as key tools in shifting diagnostic practice and reducing unnecessary antimicrobial use, veterinarians in this study, while recognizing the potential future role of such tools and technologies, nonetheless viewed diagnostic practice as a far more complex process for which rapid tests might constitute only a part.

Introduction: High-resolution temporal bone computed tomography (CT) is considered the gold standard for diagnosing superior semicircular canal dehiscence (SCD). However, CT has been shown over-detect SCD and provide results that may not align with patient-reported symptoms. Ocular vestibular-evoked myogenic potentials (oVEMPs)—most commonly conducted at 500 Hz stimulation—are increasingly used to support the diagnosis and management of SCD. Previous research reported that stimulation at higher frequencies such as 4 kHz can have near-perfect sensitivity and specificity in detecting radiographic SCD. With a larger cohort, we seek to understand the sensitivity and specificity of 4 kHz oVEMPs for detecting clinically significant SCD, as well as subgroups of radiographic, symptomatic, and surgical SCD. We also investigate whether assessing the 4 kHz oVEMP n10-p15 amplitude rather than the binary n10 response alone would optimize the detection of SCD.Methods: We conducted a cross-sectional study of patients who have undergone oVEMP testing at 4 kHz. Using the diagnostic criteria proposed by Ward et al., patients were determined to have SCD if dehiscence was confirmed on temporal bone CT by two reviewers, patient-reported characteristic symptoms, and if they had at least one positive vestibular or audiometric test suggestive of SCD. Receiver operating characteristic (ROC) analysis was conducted to identify the optimal 4 kHz oVEMP amplitude cut-off. Comparison of 4 kHz oVEMP amplitude across radiographic, symptomatic, and surgical SCD subgroups was conducted using the Mann-Whitney U test.Results: Nine hundred two patients (n, ears = 1,804) underwent 4 kHz oVEMP testing. After evaluating 150 temporal bone CTs, we identified 49 patients (n, ears = 61) who had radiographic SCD. Of those, 33 patients (n, ears = 37) were determined to have clinically significant SCD. For this study cohort, 4 kHz oVEMP responses had a sensitivity of 86.5% and a specificity of 87.8%. ROC analysis demonstrated that accounting for the inter-amplitude of 4 kHz oVEMP was more accurate in detecting SCD than the presence of n10 response alone (AUC 91 vs. 87%). Additionally, using an amplitude cut-off of 15uV reduces false positive results and improves specificity to 96.8%. Assessing 4 kHz oVEMP response across SCD subgroups demonstrated that surgical and symptomatic SCD cases had significantly higher amplitudes, while radiographic SCD cases without characteristic symptoms had similar amplitudes compared to cases without evidence of SCD.Conclusion: Our results suggest that accounting for 4 kHz oVEMP amplitude can improve detection of SCD compared to the binary presence of n10 response. The 4 kHz oVEMP amplitude cut-off that maximizes sensitivity and specificity for our cohort is 15 uV. Our results also suggest that 4 kHz oVEMP amplitudes align better with symptomatic SCD cases compared to cases in which there is radiographic SCD but no characteristic symptoms.

BackgroundDespite the success of immune checkpoint blockade therapy in the treatment of certain cancer types, only a small percentage of patients with solid malignancies achieve a durable response. Consequently, there is a need to develop novel approaches that could overcome mechanisms of tumor resistance to checkpoint inhibition. Emerging evidence has implicated the phenomenon of cancer plasticity or acquisition of mesenchymal features by epithelial tumor cells, as an immune resistance mechanism.MethodsTwo soluble factors that mediate tumor cell plasticity in the context of epithelial-mesenchymal transition are interleukin 8 (IL-8) and transforming growth factor beta (TGF-β). In an attempt to overcome escape mechanisms mediated by these cytokines, here we investigated the use of a small molecule inhibitor of the IL-8 receptors CXCR1/2, and a bifunctional agent that simultaneously blocks programmed death ligand 1 (PD-L1) and traps soluble TGF-β.ResultsWe demonstrate that simultaneous inhibition of CXCR1/2, TGF-β, and PD-L1 signaling synergizes to reduce mesenchymal tumor features in murine models of breast and lung cancer, and to markedly increase expression of tumor epithelial E-cadherin while reducing infiltration with suppressive granulocytic myeloid-derived suppressor cells, significantly enhancing T-cell infiltration and activation in tumors, and leading to improved antitumor activity.ConclusionsThis study highlights the potential benefit of combined blockade of CXCR1/2 and TGF-β signaling for modulation of tumor plasticity and potential enhancement of tumor responses to PD-L1 blockade. The data provide rationale for the evaluation of this novel approach in the clinic.

Dengue virus (DENV) is a mosquito-borne flavivirus that causes serious human disease. The current lack of an effective vaccine to simultaneously protect against the four serotypes of DENV in seronegative individuals is a major unmet medical need. Further, the immunological basis for protective immunity in the setting of DENV infection or vaccination is not fully understood. Our team has developed a live attenuated tetravalent dengue virus vaccine that provides complete protection in a human model of dengue virus challenge. The goal of this study was to define, in the context of protective human vaccination, the quality of vaccine-induced DENV-specific CD8+ and CD4+ T cells and the temporal dynamics associated with their formation and maintenance. Multifunctional, DENV-specific CD8+ and CD4+ T cells developed 8–14 days after vaccination and were maintained for at least 6 months. Virus-specific CD8 T+ cells were a mixture of effector memory T cells (TEM) and effector memory T cells re-expressing CD45RA (TEMRA), with TEM cells predominating until day 21 post-vaccination and TEMRA cells thereafter. The majority of virus-specific CD4+ T cells were TEM with a small fraction being TEMRA. The frequency of virus-specific CD8+ and CD4+ T cells were further skewed to the TEMRA phenotype following either a second dose of the tetravalent vaccine or challenge with a single serotype of DENV. Collectively, our study has defined the phenotypic profile of antiviral CD8+ and CD4+ T cells associated with protective immunity to DENV infection and the kinetics of their formation and maintenance.

Severe acute respiratory illness (SARI) is a major cause of death and morbidity in low- and middle-income countries, however, the etiologic agents are often undetermined due to the lack of molecular diagnostics in hospitals and clinics. To examine evidence for select viral infections among patients with SARI in northern Vietnam, we studied 348 nasopharyngeal samples from military and civilian patients admitted to 4 hospitals in the greater Hanoi area from 2017-2019. Initial screening for human respiratory viral pathogens was performed in Hanoi, Vietnam at the National Institute of Hygiene and Epidemiology (NIHE) or the Military Institute of Preventative Medicine (MIPM), and an aliquot was shipped to Duke-NUS Medical School in Singapore for validation. Patient demographics were recorded and used to epidemiologically describe the infections. Among military and civilian cases of SARI, 184 (52.9%) tested positive for one or more respiratory viruses. Influenza A virus was the most prevalent virus detected (64.7%), followed by influenza B virus (29.3%), enterovirus (3.8%), adenovirus (1.1%), and coronavirus (1.1%). Risk factor analyses demonstrated an increased risk of influenza A virus detection among military hospital patients (adjusted OR, 2.0; 95% CI, 1.2-3.2), and an increased risk of influenza B virus detection among patients enrolled in year 2017 (adjusted OR, 7.9; 95% CI, 2.7-22.9). As influenza A and B viruses were commonly associated with SARI and are treatable, SARI patients entering these hospitals would benefit if the hospitals were able to adapt onsite molecular diagnostics.

Abstract Objectives There is an incomplete understanding of the host humoral immune response to severe acute respiratory syndrome (SARS)‐coronavirus (CoV)‐2, which underlies COVID‐19, during acute infection. Host factors such as age and sex as well as the kinetics and functionality of antibody responses are important factors to consider as vaccine development proceeds. The receptor‐binding domain of the CoV spike (RBD‐S) protein mediates host cell binding and infection and is a major target for vaccine design to elicit neutralising antibodies. Methods We assessed serum anti‐SARS‐CoV‐2 RBD‐S IgG, IgM and IgA antibodies by a two‐step ELISA and neutralising antibodies in a cross‐sectional study of hospitalised COVID‐19 patients of varying disease severities. Anti‐RBD‐S IgG levels were also determined in asymptomatic seropositives. Results We found equivalent levels of anti‐RBD‐S antibodies in male and female patients and no age‐related deficiencies even out to 93 years of age. The anti‐RBD‐S response was evident as little as 6 days after onset of symptoms and for at least 5 weeks after symptom onset. Anti‐RBD‐S IgG, IgM and IgA responses were simultaneously induced within 10 days after onset, with anti‐RBD‐S IgG sustained over a 5‐week period. Anti‐RBD‐S antibodies strongly correlated with neutralising activity. Lastly, anti‐RBD‐S IgG responses were higher in symptomatic COVID‐19 patients during acute infection compared with asymptomatic seropositive donors. Conclusion Our results suggest that anti‐RBD‐S IgG reflect functional immune responses to SARS‐CoV‐2, but do not completely explain age‐ and sex‐related disparities in COVID‐19 fatalities.