Assessing Spns2-dependent S1P Transport as a Prospective Therapeutic Target.

S1P (sphingosine 1-phosphate) receptor modulator (SRM) drugs interfere with lymphocyte trafficking by downregulating lymphocyte S1P receptors. While the immunosuppressive activity of SRM drugs has proved useful in treating autoimmune diseases such as multiple sclerosis, that drug class is beset by on-target liabilities such as initial dose bradycardia. The S1P that binds to cell surface lymphocyte S1P receptors is provided by S1P transporters. Mice born deficient in one of these, spinster homolog 2 (Spns2), are lymphocytopenic and have low lymph S1P concentrations. Such observations suggest that inhibition of Spns2-mediated S1P transport might provide another therapeutically beneficial method to modulate immune cell positioning. We report here results using a novel S1P transport blocker (STB), SLF80821178, to investigate the consequences of S1P transport inhibition in rodents. We found that SLF80821178 is efficacious in a multiple sclerosis model but - unlike the SRM fingolimod - neither decreases heart rate nor compromises lung endothelial barrier function. Notably, although Spns2 null mice have a sensorineural hearing defect, mice treated chronically with SLF80821178 have normal hearing acuity. STBs such as SLF80821178 evoke a dose-dependent decrease in peripheral blood lymphocyte counts, which affords a reliable pharmacodynamic marker of target engagement. However, the maximal reduction in circulating lymphocyte counts in response to SLF80821178 is substantially less than the response to SRMs such as fingolimod (50% vs. 90%) due to a lesser effect on T lymphocyte sub-populations by SLF80821178. Finally, in contrast to results obtained with Spns2 deficient mice, lymph S1P concentrations were not significantly changed in response to administration of STBs at doses that evoke maximal lymphopenia, which indicates that current understanding of the mechanism of action of S1P transport inhibitors is incomplete.

Innate Immune Activation and Mitochondrial ROS Invoke Persistent Cardiac Conduction System Dysfunction after COVID-19.

Background: Cardiac risk rises during acute SARS-CoV-2 infection and in long COVID syndrome in humans, but the mechanisms behind COVID-19-linked arrhythmias are unknown. This study explores the acute and long term effects of SARS-CoV-2 on the cardiac conduction system (CCS) in a hamster model of COVID-19. Methods: Radiotelemetry in conscious animals was used to non-invasively record electrocardiograms and subpleural pressures after intranasal SARS-CoV-2 infection. Cardiac cytokines, interferon-stimulated gene expression, and macrophage infiltration of the CCS, were assessed at 4 days and 4 weeks post-infection. A double-stranded RNA mimetic, polyinosinic:polycytidylic acid (PIC), was used in vivo and in vitro to activate viral pattern recognition receptors in the absence of SARS-CoV-2 infection. Results: COVID-19 induced pronounced tachypnea and severe cardiac conduction system (CCS) dysfunction, spanning from bradycardia to persistent atrioventricular block, although no viral protein expression was detected in the heart. Arrhythmias developed rapidly, partially reversed, and then redeveloped after the pulmonary infection was resolved, indicating persistent CCS injury. Increased cardiac cytokines, interferon-stimulated gene expression, and macrophage remodeling in the CCS accompanied the electrophysiological abnormalities. Interestingly, the arrhythmia phenotype was reproduced by cardiac injection of PIC in the absence of virus, indicating that innate immune activation was sufficient to drive the response. PIC also strongly induced cytokine secretion and robust interferon signaling in hearts, human iPSC-derived cardiomyocytes (hiPSC-CMs), and engineered heart tissues, accompanied by alterations in electrical and Ca 2+ handling properties. Importantly, the pulmonary and cardiac effects of COVID-19 were blunted by in vivo inhibition of JAK/STAT signaling or by a mitochondrially-targeted antioxidant. Conclusions: The findings indicate that long term dysfunction and immune cell remodeling of the CCS is induced by COVID-19, arising indirectly from oxidative stress and excessive activation of cardiac innate immune responses during infection, with implications for long COVID Syndrome.

Troponin I Tyrosine Phosphorylation Beneficially Accelerates Diastolic Function.

BACKGROUND: A healthy heart is able to modify its function and increase relaxation through post-translational modifications of myofilament proteins. While there are known examples of serine/threonine kinases directly phosphorylating myofilament proteins to modify heart function, the roles of tyrosine (Y) phosphorylation to directly modify heart function have not been demonstrated. The myofilament protein TnI (troponin I) is the inhibitory subunit of the troponin complex and is a key regulator of cardiac contraction and relaxation. We previously demonstrated that TnI-Y26 phosphorylation decreases calcium-sensitive force development and accelerates calcium dissociation, suggesting a novel role for tyrosine kinase-mediated TnI-Y26 phosphorylation to regulate cardiac relaxation. Therefore, we hypothesize that increasing TnI-Y26 phosphorylation will increase cardiac relaxation in vivo and be beneficial during pathological diastolic dysfunction. METHODS: The signaling pathway involved in TnI-Y26 phosphorylation was predicted in silico and validated by tyrosine kinase activation and inhibition in primary adult murine cardiomyocytes. To investigate how TnI-Y26 phosphorylation affects cardiac muscle, structure, and function in vivo, we developed a novel TnI-Y26 phosphorylation-mimetic mouse that was subjected to echocardiography, pressure-volume loop hemodynamics, and myofibril mechanical studies. TnI-Y26 phosphorylation-mimetic mice were further subjected to the nephrectomy/DOCA (deoxycorticosterone acetate) model of diastolic dysfunction to investigate the effects of increased TnI-Y26 phosphorylation in disease. RESULTS: Src tyrosine kinase is sufficient to phosphorylate TnI-Y26 in cardiomyocytes. TnI-Y26 phosphorylation accelerates in vivo relaxation without detrimental structural or systolic impairment. In a mouse model of diastolic dysfunction, TnI-Y26 phosphorylation is beneficial and protects against the development of disease. CONCLUSIONS: We have demonstrated that tyrosine kinase phosphorylation of TnI is a novel mechanism to directly and beneficially accelerate myocardial relaxation in vivo.

Development and validation of a theory-based questionnaire examining barriers and facilitators to discontinuing long-term benzodiazepine receptor agonist use.

BACKGROUND: Long-term use of benzodiazepine receptor agonists (BZRAs) is a persistent healthcare challenge and poses patient safety risks. Interventions underpinned by behaviour change theory are needed to support discontinuation of long-term BZRA use. The aim of this study was to develop and validate a questionnaire based on the Theoretical Domains Framework (TDF) to examine mediators of behaviour change relating to the discontinuation of long-term BZRA use. METHODS: An initial 52 item questionnaire was developed using the 14 domains of TDF version 2 and iteratively refined over two rounds. The questionnaire was disseminated online via online support groups that focused on BZRAs to community-based adults with either current or previous experience of taking BZRAs on a long-term basis (≥3 months). Confirmatory factor analysis was undertaken to assess the questionnaire's reliability, discriminant validity and goodness of fit. The Standardized Root Mean Square Residual (SRMR), Root Mean Square Error of Approximation (RMSEA) and Comparative Fit Index (CFI) were calculated. RESULTS: Following an iterative process of adjustment, the results obtained from confirmatory factor analysis resulted in the final questionnaire consisting of 29 items across nine theoretical domains. The internal consistency reliability values across these domains ranged from 0.62 to 0.85. For the final model, the SRMR was 0.23, the RMSEA was 0.11 and the CFI was 0.6. CONCLUSIONS: The questionnaire offers a potential tool that could be used to identify domains that need to be targeted as part of a behaviour change intervention at an individual patient level. Further research is needed to assess the questionnaire's acceptability and usability, and to develop a scoring system so that domains can be prioritised and subsequently targeted as part of an intervention.

Long-term consequences of benzodiazepine-induced neurological dysfunction: A survey.

BACKGROUND: Acute benzodiazepine withdrawal has been described, but literature regarding the benzodiazepine-induced neurological injury that may result in enduring symptoms and life consequences is scant. OBJECTIVE: We conducted an internet survey of current and former benzodiazepine users and asked about their symptoms and adverse life events attributed to benzodiazepine use. METHODS: This is a secondary analysis of the largest survey ever conducted with 1,207 benzodiazepine users from benzodiazepine support groups and health/wellness sites who completed the survey. Respondents included those still taking benzodiazepines (n = 136), tapering (n = 294), or fully discontinued (n = 763). RESULTS: The survey asked about 23 specific symptoms and more than half of the respondents who experienced low energy, distractedness, memory loss, nervousness, anxiety, and other symptoms stated that these symptoms lasted a year or longer. These symptoms were often reported as de novo and distinct from the symptoms for which the benzodiazepines were originally prescribed. A subset of respondents stated that symptoms persisted even after benzodiazepines had been discontinued for a year or more. Adverse life consequences were reported by many respondents as well. LIMITATIONS: This was a self-selected internet survey with no control group. No independent psychiatric diagnoses could be made in participants. CONCLUSIONS: Many prolonged symptoms subsequent to benzodiazepine use and discontinuation (benzodiazepine-induced neurological dysfunction) have been shown in a large survey of benzodiazepine users. Benzodiazepine-induced neurological dysfunction (BIND) has been proposed as a term to describe symptoms and associated adverse life consequences that may emerge during benzodiazepine use, tapering, and continue after benzodiazepine discontinuation. Not all people who take benzodiazepines will develop BIND and risk factors for BIND remain to be elucidated. Further pathogenic and clinical study of BIND is needed.

Interleukin-6 Drives Mitochondrial Dysregulation and Accelerates Physical Decline: Insights From an Inducible Humanized IL-6 Knock-In Mouse Model.

Chronic activation of inflammatory pathways (CI) and mitochondrial dysfunction are independently linked to age-related functional decline and early mortality. Interleukin 6 (IL-6) is among the most consistently elevated chronic activation of inflammatory pathways markers, but whether IL-6 plays a causative role in this mitochondrial dysfunction and physical deterioration remains unclear. To characterize the role of IL-6 in age-related mitochondrial dysregulation and physical decline, we have developed an inducible human IL-6 (hIL-6) knock-in mouse (TetO-hIL-6mitoQC) that also contains a mitochondrial-quality control reporter. Six weeks of hIL-6 induction resulted in upregulation of proinflammatory markers, cell proliferation and metabolic pathways, and dysregulated energy utilization. Decreased grip strength, increased falls off the treadmill, and increased frailty index were also observed. Further characterization of skeletal muscles postinduction revealed an increase in mitophagy, downregulation of mitochondrial biogenesis genes, and an overall decrease in total mitochondrial numbers. This study highlights the contribution of IL-6 to mitochondrial dysregulation and supports a causal role of hIL-6 in physical decline and frailty.

Acupuncture, Nutritional Augmentation, and Physical Rehabilitation: Improving Recovery Following Spinal Fusion.

Spinal fusion is one of the most common procedures performed in spinal surgery. The increasing complexity and frequency with which this operation is performed has led to a rise in postoperative complications and delayed recovery. Perioperative care of patients with spinal conditions aims to adequately manage pain and accelerate the return to function following surgery. Traditional pharmaceutical interventions, especially opioid utilization, are associated with delayed recovery and significant adverse effects. As such, many studies are now evaluating the benefits and efficacy of non-pharmacological therapies for enhancing recovery after spinal fusion surgery. In this review, the mechanism of acupuncture relative to postoperative pain control and symptom reduction will be discussed. Additionally, this review examines the implications of malnutrition, current preclinical and clinical approaches for improving nutritional status, and various forms of physical rehabilitation that aim to combat postoperative complications and support recovery after spinal fusion surgery.

Development of BNBSL: A ß-ν spectra library for spectrometry applications.

While modern nuclear decay data can provide many details of a given nuclides ß-decay modes (branching ratios, decay heating etc.), knowledge of the emitted ß-energy spectrum is often not included. This limitation hampers the use of decay data in some analysis, such as ß-spectrometry of irradiated material, prediction of ß-decay Bremsstrahlung or antineutrino, ν̄, detection. To address this deficiency, and for increased ease of ß-spectrometry studies of complex samples, a library of ß, ν and Bremsstrahlung-spectra, called BNBSL (Beta-Neutrino-Bremsstrahlung spectra library), has been produced. It has been found that the content compares favourably with experimental data and methodologies for its application to complex nuclear inventories have been developed. BNBSL contains spectra for over 1500 nuclides, which is hoped will benefit applied nuclear, radiation and materials science studies.

The role of histone methyltransferases in neurocognitive disorders associated with brain size abnormalities.

Brain size is controlled by several factors during neuronal development, including neural progenitor proliferation, neuronal arborization, gliogenesis, cell death, and synaptogenesis. Multiple neurodevelopmental disorders have co-morbid brain size abnormalities, such as microcephaly and macrocephaly. Mutations in histone methyltransferases that modify histone H3 on Lysine 36 and Lysine 4 (H3K36 and H3K4) have been identified in neurodevelopmental disorders involving both microcephaly and macrocephaly. H3K36 and H3K4 methylation are both associated with transcriptional activation and are proposed to sterically hinder the repressive activity of the Polycomb Repressor Complex 2 (PRC2). During neuronal development, tri-methylation of H3K27 (H3K27me3) by PRC2 leads to genome wide transcriptional repression of genes that regulate cell fate transitions and neuronal arborization. Here we provide a review of neurodevelopmental processes and disorders associated with H3K36 and H3K4 histone methyltransferases, with emphasis on processes that contribute to brain size abnormalities. Additionally, we discuss how the counteracting activities of H3K36 and H3K4 modifying enzymes vs. PRC2 could contribute to brain size abnormalities which is an underexplored mechanism in relation to brain size control.

Enduring neurological sequelae of benzodiazepine use: an Internet survey.

Introduction: Benzodiazepine tapering and cessation has been associated with diverse symptom constellations of varying duration. Although described in the literature decades ago, the mechanistic underpinnings of enduring symptoms that can last months or years have not yet been elucidated. Objective: This secondary analysis of the results from an Internet survey sought to better understand the acute and protracted withdrawal symptoms associated with benzodiazepine use and discontinuation. Methods: An online survey (n = 1207) was used to gather information about benzodiazepine use, including withdrawal syndrome and protracted symptoms. Results: The mean number of withdrawal symptoms reported by a respondent in this survey was 15 out of 23 symptoms. Six percent of respondents reported having all 23 listed symptoms. A cluster of least-frequently reported symptoms (whole-body trembling, hallucinations, seizures) were also the symptoms most frequently reported as lasting only days or weeks, that is, short-duration symptoms. Symptoms of nervousness/anxiety/fear, sleep disturbances, low energy, and difficulty focusing/distractedness were experienced by the majority of respondents (⩾85%) and, along with memory loss, were the symptoms of longest duration. Prolonged symptoms of anxiety and insomnia occurred in many who have discontinued benzodiazepines, including over 50% who were not originally prescribed benzodiazepines for that indication. It remains unclear if these symptoms might be caused by neuroadaptive and/or neurotoxic changes induced by benzodiazepine exposure. In this way, benzodiazepine withdrawal may have acute and long-term symptoms attributable to different underlying mechanisms, which is the case with alcohol withdrawal. Conclusions: These findings tentatively support the notion that symptoms which are acute but transient during benzodiazepine tapering and discontinuation may be distinct in their nature and duration from the enduring symptoms experienced by many benzodiazepine users.

Inhibiting O-GlcNAcylation impacts p38 and Erk1/2 signaling and perturbs cardiomyocyte hypertrophy.

The dynamic cycling of O-linked GlcNAc (O-GlcNAc) on and off Ser/Thr residues of intracellular proteins, termed O-GlcNAcylation, is mediated by the conserved enzymes O-GlcNAc transferase (OGT) and O-GlcNAcase. O-GlcNAc cycling is important in homeostatic and stress responses, and its perturbation sensitizes the heart to ischemic and other injuries. Despite considerable progress, many molecular pathways impacted by O-GlcNAcylation in the heart remain unclear. The mitogen-activated protein kinase (MAPK) pathway is a central signaling cascade that coordinates developmental, physiological, and pathological responses in the heart. The developmental or adaptive arm of MAPK signaling is primarily mediated by Erk kinases, while the pathophysiologic arm is mediated by p38 and Jnk kinases. Here, we examine whether O-GlcNAcylation affects MAPK signaling in cardiac myocytes, focusing on Erk1/2 and p38 in basal and hypertrophic conditions induced by phenylephrine. Using metabolic labeling of glycans coupled with alkyne-azide "click" chemistry, we found that Erk1/2 and p38 are O-GlcNAcylated. Supporting the regulation of p38 by O-GlcNAcylation, the OGT inhibitor, OSMI-1, triggers the phosphorylation of p38, an event that involves the NOX2-Ask1-MKK3/6 signaling axis and also the noncanonical activator Tab1. Additionally, OGT inhibition blocks the phenylephrine-induced phosphorylation of Erk1/2. Consistent with perturbed MAPK signaling, OSMI-1-treated cardiomyocytes have a blunted hypertrophic response to phenylephrine, decreased expression of cTnT (key component of the contractile apparatus), and increased expression of maladaptive natriuretic factors Anp and Bnp. Collectively, these studies highlight new roles for O-GlcNAcylation in maintaining a balanced activity of Erk1/2 and p38 MAPKs during hypertrophic growth responses in cardiomyocytes.

Alteration in tyrosine phosphorylation of cardiac proteome and EGFR pathway contribute to hypertrophic cardiomyopathy.

Alterations of serine/threonine phosphorylation of the cardiac proteome are a hallmark of heart failure. However, the contribution of tyrosine phosphorylation (pTyr) to the pathogenesis of cardiac hypertrophy remains unclear. We use global mapping to discover and quantify site-specific pTyr in two cardiac hypertrophic mouse models, i.e., cardiac overexpression of ErbB2 (TgErbB2) and α myosin heavy chain R403Q (R403Q-αMyHC Tg), compared to control hearts. From this, there are significant phosphoproteomic alterations in TgErbB2 mice in right ventricular cardiomyopathy, hypertrophic cardiomyopathy (HCM), and dilated cardiomyopathy (DCM) pathways. On the other hand, R403Q-αMyHC Tg mice indicated that the EGFR1 pathway is central for cardiac hypertrophy, along with angiopoietin, ErbB, growth hormone, and chemokine signaling pathways activation. Surprisingly, most myofilament proteins have downregulation of pTyr rather than upregulation. Kinase-substrate enrichment analysis (KSEA) shows a marked downregulation of MAPK pathway activity downstream of k-Ras in TgErbB2 mice and activation of EGFR, focal adhesion, PDGFR, and actin cytoskeleton pathways. In vivo ErbB2 inhibition by AG-825 decreases cardiomyocyte disarray. Serine/threonine and tyrosine phosphoproteome confirm the above-described pathways and the effectiveness of AG-825 Treatment. Thus, altered pTyr may play a regulatory role in cardiac hypertrophic models.

Tbx18 Orchestrates Cytostructural Transdifferentiation of Cardiomyocytes to Pacemaker Cells by Recruiting the Epithelial-Mesenchymal Transition Program.

Previously, we reported that heterologous expression of an embryonic transcription factor, Tbx18, reprograms ventricular cardiomyocytes into induced pacemaker cells (Tbx18-iPMs), though the key pathways are unknown. Here, we have used a tandem mass tag proteomic approach to characterize the impact of Tbx18 on neonatal rat ventricular myocytes. Tbx18 expression triggered vast proteome remodeling. Tbx18-iPMs exhibited increased expression of known pacemaker ion channels, including Hcn4 and Cx45 as well as upregulation of the mechanosensitive ion channels Piezo1, Trpp2 (PKD2), and TrpM7. Metabolic pathways were broadly downregulated, as were ion channels associated with ventricular excitation-contraction coupling. Tbx18-iPMs also exhibited extensive intracellular cytoskeletal and extracellular matrix remodeling, including 96 differentially expressed proteins associated with the epithelial-to-mesenchymal transition (EMT). RNAseq extended coverage of low abundance transcription factors, revealing upregulation of EMT-inducing Snai1, Snai2, Twist1, Twist2, and Zeb2. Finally, network diffusion mapping of >200 transcriptional regulators indicates EMT and heart development factors occupy adjacent network neighborhoods downstream of Tbx18 but upstream of metabolic control factors. In conclusion, transdifferentiation of cardiac myocytes into pacemaker cells entails massive electrogenic, metabolic, and cytostructural remodeling. Structural changes exhibit hallmarks of the EMT. The results aid ongoing efforts to maximize the yield and phenotypic stability of engineered biological pacemakers.

Supporting safe and gradual reduction of long-term benzodiazepine receptor agonist use: Development of the SAFEGUARDING-BZRAs toolkit using a codesign approach.

INTRODUCTION: Long-term benzodiazepine receptor agonist (BZRA) use persists in healthcare settings worldwide and poses risks of patient harm. OBJECTIVE: This study aimed to develop an intervention to support discontinuation of long-term BZRA use among willing individuals. METHODS: The intervention development process aligned with the UK Medical Research Council's complex intervention framework. This involved a previous systematic review of brief interventions targeting long-term BZRA use in primary care and qualitative interviews based on the Theoretical Domains Framework that explored barriers and facilitators to discontinuing long-term BZRA use. A codesign approach was used involving an active partnership between experts by experience, researchers and clinicians. Intervention content was specified in terms of behaviour change techniques (BCTs). RESULTS: The SAFEGUARDING-BZRAs (Supporting sAFE and GradUAl ReDuctIon of loNG-term BenZodiazepine Receptor Agonist uSe) toolkit comprises 24 BCTs and includes recommendations targeted at primary care-based clinicians for operationalizing each BCT to support individuals with BZRA discontinuation. CONCLUSION: The SAFEGUARDING-BZRAs toolkit has been developed using a systematic and theory-based approach that addresses identified limitations of previous research. Further research is needed to assess its usability and acceptability by service users and clinicians, as well as its potential to effectively support safe and gradual reduction of long-term BZRA use. PATIENT OR PUBLIC CONTRIBUTION: The qualitative interview phase included patients as participants. The codesign process included 'experts by experience' with either current or previous experience of long-term BZRA use as collaborators.

Counteracting epigenetic mechanisms regulate the structural development of neuronal circuitry in human neurons.

Autism spectrum disorders (ASD) are associated with defects in neuronal connectivity and are highly heritable. Genetic findings suggest that there is an overrepresentation of chromatin regulatory genes among the genes associated with ASD. ASH1 like histone lysine methyltransferase (ASH1L) was identified as a major risk factor for ASD. ASH1L methylates Histone H3 on Lysine 36, which is proposed to result primarily in transcriptional activation. However, how mutations in ASH1L lead to deficits in neuronal connectivity associated with ASD pathogenesis is not known. We report that ASH1L regulates neuronal morphogenesis by counteracting the catalytic activity of Polycomb Repressive complex 2 group (PRC2) in stem cell-derived human neurons. Depletion of ASH1L decreases neurite outgrowth and decreases expression of the gene encoding the neurotrophin receptor TrkB whose signaling pathway is linked to neuronal morphogenesis. The neuronal morphogenesis defect is overcome by inhibition of PRC2 activity, indicating that a balance between the Trithorax group protein ASH1L and PRC2 activity determines neuronal morphology. Thus, our work suggests that ASH1L may epigenetically regulate neuronal morphogenesis by modulating pathways like the BDNF-TrkB signaling pathway. Defects in neuronal morphogenesis could potentially impair the establishment of neuronal connections which could contribute to the neurodevelopmental pathogenesis associated with ASD in patients with ASH1L mutations.

Evidence of sheep and abattoir environment as important reservoirs of multidrug resistant Salmonella and extended-spectrum beta-lactamase Escherichia coli.

The increase in antimicrobial-resistant (AMR) foodborne pathogens, including E. coli and Salmonella in animals, humans, and the environment, is a growing public health concern. Among animals, cattle, pigs, and chicken are reservoirs of these pathogens worldwide. There is a knowledge gap on the prevalence and AMR of foodborne pathogens in small ruminants (i.e., sheep and goats). This study investigates the prevalence and antimicrobial resistance of extended-spectrum beta-lactamase (ESBL) E. coli and Salmonella from sheep and their abattoir environment in North Carolina. We conducted a year-round serial cross-sectional study and collected a total of 1128 samples from sheep (n = 780) and their abattoir environment (n = 348). Sheep samples consisted of feces, cecal contents, carcass swabs, and abattoir resting area feces. Environmental samples consisted of soil samples, lairage swab, animal feed, and drinking water for animals. We used CHROMAgar EEC with 4 µg/ml of Cefotaxime for isolating ESBL E. coli, and ESBL production was confirmed by double-disk diffusion test. Salmonella was isolated and confirmed using standard methods. All of the confirmed isolates were tested against a panel of 14 antimicrobials to elucidate susceptibility profiles. The prevalence of ESBL E. coli and Salmonella was significantly higher in environmental samples (47.7% and 65.5%) compared to the sheep samples (19.5% and 17.9%), respectively (P < 0.0001). We recovered 318 ESBL E. coli and 368 Salmonella isolates from sheep and environmental samples. More than 97% (310/318) of ESBL E. coli were multidrug-resistant (MDR; resistant to ≥3 classes of antimicrobials). Most Salmonella isolates (77.2%, 284/368) were pansusceptible, and 10.1% (37/368) were MDR. We identified a total of 24 different Salmonella serotypes by whole genome sequencing (WGS). The most common serotypes were Agona (19.8%), Typhimurium (16.2%), Cannstatt (13.2%), Reading (13.2%), and Anatum (9.6%). Prevalence and percent resistance of ESBL E. coli and Salmonella isolates varied significantly by season and sample type (P < 0.0001). The co-existence of ESBL E. coli in the same sample was associated with increased percent resistance of Salmonella to Ampicillin, Chloramphenicol, Sulfisoxazole, Streptomycin, and Tetracycline. We presumed that the abattoir environment might have played a great role in the persistence and dissemination of resistant bacteria to sheep as they arrive at the abattoir. In conclusion, our study reaffirms that sheep and their abattoir environment act as important reservoirs of AMR ESBL E. coli and MDR Salmonella in the U.S. Further studies are required to determine associated public health risks.