Potential pandemic risk of circulating swine H1N2 influenza viruses.

Influenza A viruses in swine have considerable genetic diversity and continue to pose a pandemic threat to humans due to a potential lack of population level immunity. Here we describe a pipeline to characterize and triage influenza viruses for their pandemic risk and examine the pandemic potential of two widespread swine origin viruses. Our analysis reveals that a panel of human sera collected from healthy adults in 2020 has no cross-reactive neutralizing antibodies against a α-H1 clade strain (α-swH1N2) but do against a γ-H1 clade strain. The α-swH1N2 virus replicates efficiently in human airway cultures and exhibits phenotypic signatures similar to the human H1N1 pandemic strain from 2009 (H1N1pdm09). Furthermore, α-swH1N2 is capable of efficient airborne transmission to both naïve ferrets and ferrets with prior seasonal influenza immunity. Ferrets with H1N1pdm09 pre-existing immunity show reduced α-swH1N2 viral shedding and less severe disease signs. Despite this, H1N1pdm09-immune ferrets that became infected via the air can still onward transmit α-swH1N2 with an efficiency of 50%. These results indicate that this α-swH1N2 strain has a higher pandemic potential, but a moderate level of impact since there is reduced replication fitness and pathology in animals with prior immunity.

Debulking of the Femoral Stem in a Primary Total Hip Joint Replacement: A Novel Method to Reduce Stress Shielding.

In current-generation designs of total primary hip joint replacement, the prostheses are fabricated from alloys. The modulus of elasticity of the alloy is substantially higher than that of the surrounding bone. This discrepancy plays a role in a phenomenon known as stress shielding, in which the bone bears a reduced proportion of the applied load. Stress shielding has been implicated in aseptic loosening of the implant which, in turn, results in reduction in the in vivo life of the implant. Rigid implants shield surrounding bone from mechanical loading, and the reduction in skeletal stress necessary to maintain bone mass and density results in accelerated bone loss, the forerunner to implant loosening. Femoral stems of various geometries and surface modifications, materials and material distributions, and porous structures have been investigated to achieve mechanical properties of stems closer to those of bone to mitigate stress shielding. For improved load transfer from implant to femur, the proposed study investigated a strategic debulking effort to impart controlled flexibility while retaining sufficient strength and endurance properties. Using an iterative design process, debulked configurations based on an internal skeletal truss framework were evaluated using finite element analysis. The implant models analyzed were solid; hollow, with a proximal hollowed stem; FB-2A, with thin, curved trusses extending from the central spine; and FB-3B and FB-3C, with thick, flat trusses extending from the central spine in a balanced-truss and a hemi-truss configuration, respectively. As outlined in the International Organization for Standardization (ISO) 7206 standards, implants were offset in natural femur for evaluation of load distribution or potted in testing cylinders for fatigue testing. The commonality across all debulked designs was the minimization of proximal stress shielding compared to conventional solid implants. Stem topography can influence performance, and the truss implants with or without the calcar collar were evaluated. Load sharing was equally effective irrespective of the collar; however, the collar was critical to reducing the stresses in the implant. Whether bonded directly to bone or cemented in the femur, the truss stem was effective at limiting stress shielding. However, a localized increase in maximum principal stress at the proximal lateral junction could adversely affect cement integrity. The controlled accommodation of deformation of the implant wall contributes to the load sharing capability of the truss implant, and for a superior biomechanical performance, the collared stem should be implanted in interference fit. Considering the results of all implant designs, the truss implant model FB-3C was the best model.

Therapist contribution, client reflective functioning, and alliance rupture-repair: A microprocess case study of psychodynamic therapy for pregnancy after loss.

Meta-analysis has found a significant relation between rupture-repair and client outcome (Eubanks et al., 2018). Rupture-repair processes may be particularly important in psychotherapy for pregnancy loss wherein ruptures related to client feelings of shame and inadequacy, the societal invalidation of perinatal grief, and reenactments in the therapy relationship of early attachment experiences have been theorized to be common and important events (Markin, 2024). Thus, it is important to understand what occurs on a microlevel during the process of therapy to ultimately explain the rupture resolution (RR) and treatment outcome association. In particular, while both the therapist and client are believed to contribute to ruptures and to their repair (Safran & Muran, 2000), little is known about how therapist contributions impact rupture events, rupture resolution, and treatment progress. Further, client reflective functioning (RF) may represent a set of capacities that contribute to and are increased by rupture resolution yet vary depending on the role of the therapist in the rupture. The current investigation examined how observer-rated therapist contribution to ruptures and client RF were related to rupture events, rupture resolution, and client-reported symptom change and session quality over 22 sessions of psychodynamic therapy for pregnancy after loss. Therapist contribution to ruptures predicted rupture significance, high and steady within-session client RF scores, and symptom change. Client RF and rupture resolution predicted symptom change differently, often depending on type of symptom. Importantly, client RF and rupture resolution may predict successful outcomes through ameliorating commonly reported symptoms during pregnancies after loss. (PsycInfo Database Record (c) 2024 APA, all rights reserved).

A protective and broadly binding antibody class engages the influenza virus hemagglutinin head at its stem interface.

Influenza infection and vaccination impart strain-specific immunity that protects against neither seasonal antigenic variants nor the next pandemic. However, antibodies directed to conserved sites can confer broad protection. Here we identify and characterize a class of human antibodies that engage a previously undescribed, conserved epitope on the influenza hemagglutinin (HA) protein. Prototype antibody S8V1-157 binds at the normally occluded interface between the HA head and stem. Antibodies to this HA head-stem interface epitope are non-neutralizing in vitro but protect against lethal influenza infection in mice. Antibody isotypes that direct clearance of infected cells enhance this protection. Head-stem interface antibodies bind to most influenza A serotypes and seasonal human variants, and are present at low frequencies in the memory B cell populations of multiple human donors. Vaccines designed to elicit these antibodies might contribute to "universal" influenza immunity.

Cost analysis and clinical outcomes of anatomic pre-contoured locking versus conventional plates for distal fibula ankle fractures.

PURPOSE: To analyze differences in union, complication rates and cost from surgical fixation of distal fibula fractures with fibular plating implants. METHODS: In total, 380 adult patients from 2012 to 2015 treated with 12 fibular plates from 4 different manufacturers utilized by 9 surgeons were retrospectively reviewed. They were stratified into a conventional one-third tubular fibular plate group, pre-contoured anatomic locking plate group, or a heterogeneous group including 3.5-mm reconstruction, one-third tubular locking, composite, and limited compression plates. The outcomes included failure of fixation, deep infection requiring debridement, time to union, anatomic reduction, superficial infection, hardware removal, and post-traumatic arthritis. Plate and screw costs were calculated from hospital billing records. RESULTS: Pre-contoured locking plates were used in older, female patients with a greater number of comorbidities. Open injuries and OTA 44B fractures were more likely to be an indication for pre-contoured plates. There was no difference noted in time to union between the different plating groups. Risk factors for deep infection requiring debridement included a history of tobacco use, open fractures, and pre-contoured locking plates relative to the conventional plating group. The pre-contoured plating group was on average $586 more expensive compared to the conventional group. CONCLUSION: Pre-contoured locking plates achieved similar radiographic outcomes compared to conventional plates with an increased risk of complications and higher cost. Surgeons should consider their choice of implant based on the patient's fracture pattern, underlying comorbidities, and risk for infection.

Protective human antibodies against a conserved epitope in pre- and postfusion influenza hemagglutinin.

Phylogenetically and antigenically distinct influenza A and B viruses (IAV and IBV) circulate in human populations, causing widespread morbidity. Antibodies (Abs) that bind epitopes conserved in both IAV and IBV hemagglutinins (HAs) could protect against disease by diverse virus subtypes. Only one reported HA Ab, isolated from a combinatorial display library, protects against both IAV and IBV. Thus, there has been so far no information on the likelihood of finding naturally occurring human Abs that bind HAs of diverse IAV subtypes and IBV lineages. We have now recovered from several unrelated human donors five clonal Abs that bind a conserved epitope preferentially exposed in the postfusion conformation of IAV and IVB HA2. These Abs lack neutralizing activity in vitro but in mice provide strong, IgG subtype-dependent protection against lethal IAV and IBV infections. Strategies to elicit similar Abs routinely might contribute to more effective influenza vaccines.

A new class of antibodies that overcomes a steric barrier to cross-group neutralization of influenza viruses.

Antibody titers that inhibit the influenza virus hemagglutinin (HA) from engaging its receptor are the accepted correlate of protection from infection. Many potent antibodies with broad, intra-subtype specificity bind HA at the receptor binding site (RBS). One barrier to broad H1-H3 cross-subtype neutralization is an insertion (133a) between positions 133 and 134 on the rim of the H1 HA RBS. We describe here a class of antibodies that overcomes this barrier. These genetically unrestricted antibodies are abundant in the human B cell memory compartment. Analysis of the affinities of selected members of this class for historical H1 and H3 isolates suggest that they were elicited by H3 exposure and broadened or diverted by later exposure(s) to H1 HA. RBS mutations in egg-adapted vaccine strains cause the new H1 specificity of these antibodies to depend on the egg adaptation. The results suggest that suitable immunogens might elicit 133a-independent, H1-H3 cross neutralization by RBS-directed antibodies.

Metabolic syndrome accelerates epigenetic ageing in older adults: Findings from The Irish Longitudinal Study on Ageing (TILDA).

Metabolic syndrome (MetS) is a risk factor for the development of diabetes, cardiovascular disease, and all-cause mortality. It has an estimated prevalence of 40 % among older adults. Epigenetic clocks, which measure biological age based on DNA methylation (DNAm) patterns, are a candidate biomarker for ageing. GrimAge is one such clock which is based on levels of DNAm at 100 Cytosine-phosphate-Guanine (CpG) sites. This study hypothesised that those with MetS have 'accelerated ageing' (biological age greater than their chronological age) as indexed by GrimAge. This study examined MetS's association with GrimAge age acceleration (AA) using data from a subsample of 469 participants of the Irish Longitudinal Study on Ageing (TILDA). MetS was defined by National Cholesterol Education Program Third Adult Treatment Panel (ATP III) and International Diabetes Foundation (IDF) criteria, operationalised using the conventional binary cut-off, and as a count variable ranging from 0 to 5, based on the presence of individual components. This study also explored inflammation (as measured by C reactive protein) and metabolic dysfunction (as measured by adiponectin) as possible mediating factors between MetS and GrimAge AA. We found that MetS as defined by IDF criteria was associated with GrimAge AA of 0.63 years. When MetS was treated as a count, each unit increase in MetS score was associated with over 0.3 years GrimAge AA for both ATP III and IDF criteria. Inflammation mediated approximately one third of the association between MetS and GrimAge AA, suggesting that chronic subclinical inflammation observed in MetS has a relationship with DNAm changes consistent with a faster pace of ageing. Metabolic dysfunction mediated the association between MetS and GrimAge AA to a lesser extent (16 %). These data suggest that chronic subclinical inflammation observed in MetS has a relationship with DNAm changes consistent with a greater pace of ageing.

Transient inhibition of lysosomal functions potentiates nucleic acid vaccines.

Nucleic acid vaccines have shown promising results in the clinic against infectious diseases and cancers. To robustly improve the vaccine efficacy and safety, we developed an approach to increase the intracellular stability of nucleic acids by transiently inhibiting lysosomal function in targeted tissues using sucrose. To achieve efficient and localized delivery of sucrose in animals, we designed a biomimetic lipid nanoparticle (LNP) to target the delivery of sucrose into mouse muscle cells. Using this approach, viral antigen expression in mouse muscle after DNA vaccination was substantially increased and prolonged without inducing local or systemic inflammation or toxicity. The same change in antigen expression would be achieved if the vaccine dose could be increased by 3,000 folds, which is experimentally and clinically impractical due to material restrictions and severe toxicity that will be induced by such a high dose of nucleic acids. The increase in antigen expression augmented the infiltration and activation of antigen-presenting cells, significantly improved vaccine-elicited humoral and T cell responses, and fully protected mice against the viral challenge at a low dose of vaccine. Based on these observations, we conclude that transient inhibition of lysosome function in target tissue by sucrose LNPs is a safe and potent approach to substantially improve nucleic acid-based vaccines.

One size does not fit all: an application of stochastic modeling to estimating primary healthcare needs in Ethiopia at the sub-national level.

BACKGROUND: Primary healthcare systems require adequate staffing to meet the needs of their local population. Guidelines typically use population ratio targets for healthcare workers, such as Ethiopia's goal of two health extension workers for every five thousand people. However, fixed ratios do not reflect local demographics, fertility rates, disease burden (e.g., malaria endemicity), or trends in these values. Recognizing this, we set out to estimate the clinical workload to meet the primary healthcare needs in Ethiopia by region. METHODS: We utilize the open-source R package PACE-HRH for our analysis, which is a stochastic Monte Carlo simulation model that estimates workload for a specified service package and population. Assumptions and data inputs for region-specific fertility, mortality, disease burden were drawn from literature, DHS, and WorldPop. We project workload until 2035 for seven regions and two charted cities of Ethiopia. RESULTS: All regions and charted cities are expected to experience increased workload between 2021 and 2035 for a starting catchment of five thousand people. The expected (mean) annual clinical workload varied from 2,930 h (Addis) to 3,752 h (Gambela) and increased by 19-28% over fifteen years. This results from a decline in per capita workload (due to declines in fertility and infectious diseases), overpowered by total population growth. Pregnancy, non-communicable diseases, sick child care, and nutrition remain the largest service categories, but their priority shifts substantially in some regions by 2035. Sensitivity analysis shows that fertility assumptions have major implications for workload. We incorporate seasonality and estimate monthly variation of up to 8.9% (Somali), though most services with high variability are declining. CONCLUSIONS: Regional variation in demographics, fertility, seasonality, and disease trends all affect the workload estimates. This results in differences in expected clinical workload, the level of uncertainty in those estimates, and relative priorities between service categories. By showing these differences, we demonstrate the inadequacy of a fixed population ratio for staffing allocation. Policy-makers and regulators need to consider these factors in designing their healthcare systems, or they risk sub-optimally allocating workforce and creating inequitable access to care.

Facilitating Active Collaborative Learning in Medical Education; a Literature Review of Peer Instruction Method.

The call for educational reform by the Carnegie Foundation for the Advancement of Teaching marked a pivotal juncture in the trajectory of medical education in the United States. The call underscored the imperative for educational restructuring to equip forthcoming physicians with the requisite skills to engage in lifelong learning. Among the several active teaching methods is the Peer Instruction (PI), a brainchild of Eric Mazur, empowering students to steer their own education and wield knowledge adeptly into real-world scenarios. In this review paper, we delve into the core elements of PI which involves the combination of four dynamic pedagogical approaches which are: Just-in-Time Teaching, ConcepTest, Audience Response System, and Think-Pair-Share technique. PIs effectiveness notwithstanding, it is not exempt from limitations such as its flexible implementation, lengthy time, the level of expertise required for instructional design, among others. While Peer Instruction has become increasingly popular among educators across other disciplines, with proven educational benefits with positive outcomes, PIs footprint in gradate and postgraduate medical education remains inchoate, evidenced by a paucity of scholarly references. This underscores a crucial gap - despite its proven potency in fueling engagement and learning, PI still lacks formal recognition and acknowledgement as a distinct instructional method in medical education. Within these boundaries, the promise of heightened education and amplified engagement beckons further exploration of PI as a medical educational model, warranting more consideration and research.

Population-Level Risk Factors Related to Measles Case Fatality: A Conceptual Framework Based on Expert Consultation and Literature Review.

A better understanding of population-level factors related to measles case fatality is needed to estimate measles mortality burden and impact of interventions such as vaccination. This study aimed to develop a conceptual framework of mechanisms associated with measles case fatality ratios (CFRs) and assess the scope of evidence available for related indicators. Using expert consultation, we developed a conceptual framework of mechanisms associated with measles CFR and identified population-level indicators potentially associated with each mechanism. We conducted a literature review by searching PubMed on 31 October 2021 to determine the scope of evidence for the expert-identified indicators. Studies were included if they contained evidence of an association between an indicator and CFR and were excluded if they were from non-human studies or reported non-original data. Included studies were assessed for study quality. Expert consultation identified five mechanisms in a conceptual framework of factors related to measles CFR. We identified 3772 studies for review and found 49 studies showing at least one significant association with CFR for 15 indicators (average household size, educational attainment, first- and second-dose coverage of measles-containing vaccine, human immunodeficiency virus prevalence, level of health care available, stunting prevalence, surrounding conflict, travel time to major city or settlement, travel time to nearest health care facility, under-five mortality rate, underweight prevalence, vitamin A deficiency prevalence, vitamin A treatment, and general malnutrition) and only non-significant associations for five indicators (antibiotic use for measles-related pneumonia, malaria prevalence, percent living in urban settings, pneumococcal conjugate vaccination coverage, vitamin A supplementation). Our study used expert consultation and a literature review to provide additional insights and a summary of the available evidence of these underlying mechanisms and indicators that could inform future measles CFR estimations.

Investigation of the intraoperative cortical responses to spinal motor mapping in a patient with chronic pain.

Intraoperative neurophysiological monitoring (IONM) in spinal cord stimulation (SCS) surgery for chronic pain is shown to provide effective guidance during device placement. Electromyography (EMG) is used to determine the laterality of the paddle. In some SCS cases, laterality cannot be obtained via EMG due to patient physiology. Electroencephalography (EEG) is already used in IONM to monitor cortical responses. Here, we show proof-of-concept of assessing the responses of epidurally evoked EMGs simultaneously with EEGs to determine laterality during IONM using a high-resolution (HR) SCS paddle. An 8-column HR-SCS paddle was acutely placed at T9-T10 interspace in patients with failed back surgery syndrome. EMG signals from 18 muscle groups were recorded simultaneously with 60-channel EEG signals at various stimulation amplitudes (0-10 mA). Particular attention was paid to regions associated with pain including the somatosensory cortex (S1), prefrontal cortex (PFC), and motor cortex (M1). When left and right lateral contacts were stimulated at low amplitudes (1-2 mA), significant changes were seen in θ, α, and ß powers in the contralateral PFC but not in M1 or S1. There was a significant correlation between M1 and contralateral contacts in α power. At higher currents (7-8 mA), right-sided contacts resulted in α power change. We found significant differences in α, θ, and ß powers in PFC for contralateral stimulation of the lateral SCS contacts at low amplitudes and in α power at higher amplitudes. The changes in PFC suggest the potential of EEG for understanding a cortical mechanism of action of SCS and provide insight into the pathophysiology of chronic pain.NEW & NOTEWORTHY Here, we present proof of concept of assessing the responses of epidurally evoked electromyography simultaneously with scalp electroencephalography to determine whether both laterality and insights into pain mechanisms can be elucidated. With stimulation, significant changes were seen in θ, α, and ß band power in the contralateral prefrontal cortex and in α power in the motor cortex. We provide insight into the mechanism of action of SCS in preventing pain in this patient.

European Respiratory Society/American Thoracic Society technical statement: standardisation of the measurement of lung volumes, 2023 update.

This document updates the 2005 European Respiratory Society (ERS) and American Thoracic Society (ATS) technical standard for the measurement of lung volumes. The 2005 document integrated the recommendations of an ATS/ERS task force with those from an earlier National Heart, Lung, and Blood Institute workshop that led to the publication of background papers between 1995 and 1999 and a consensus workshop report with more in-depth descriptions and discussion. Advancements in hardware and software, new research and emerging approaches have necessitated an update to the 2005 technical standard to guide laboratory directors, physiologists, operators, pulmonologists and manufacturers. Key updates include standardisation of linked spirometry, new equipment quality control and validation recommendations, generalisation of the multiple breath washout concept beyond nitrogen, a new acceptability and grading system with addition of example tracings, and a brief review of imaging and other new techniques to measure lung volumes. Future directions and key research questions are also noted.

Efficacy and moderators of short-term psychodynamic psychotherapy for depression: A systematic review and meta-analysis of individual participant data.

BACKGROUND: Short-term psychodynamic psychotherapy (STPP) is frequently used to treat depression, but it is unclear which patients might benefit specifically. Individual participant data (IPD) meta-analyses can provide more precise effect estimates than conventional meta-analyses and identify patient-level moderators. This IPD meta-analysis examined the efficacy and moderators of STPP for depression compared to control conditions. METHODS: PubMed, PsycInfo, Embase, and Cochrane Library were searched September 1st, 2022, to identify randomized trials comparing STPP to control conditions for adults with depression. IPD were requested and analyzed using mixed-effects models. RESULTS: IPD were obtained from 11 of the 13 (84.6%) studies identified (n = 771/837, 92.1%; mean age = 40.8, SD = 13.3; 79.3% female). STPP resulted in significantly lower depressive symptom levels than control conditions at post-treatment (d = -0.62, 95%CI [-0.76, -0.47], p < .001). At post-treatment, STPP was more efficacious for participants with longer rather than shorter current depressive episode durations. CONCLUSIONS: These results support the evidence base of STPP for depression and indicate episode duration as an effect modifier. This moderator finding, however, is observational and requires prospective validation in future large-scale trials.

Modeling juvenile sea turtle bycatch risk in commercial and recreational fisheries.

Understanding the drivers of fisheries bycatch is essential for limiting its impacts on vulnerable species. Here we present a model to estimate the relative magnitude of sea turtle bycatch in major coastal fisheries across the southeastern US based on spatiotemporal variation in fishing effort and the simulated distributions of juvenile Kemp's ridley (Lepidochelys kempii) and green (Chelonia mydas) sea turtles recruiting from oceanic to nearshore habitats. Over the period modeled (1996-2017), bycatch in recreational fisheries was estimated to be greater than the sum of bycatch that occurred in commercial fisheries that have historically been considered high risks to turtles (e.g., those using trawls, gillnets, and bottom longlines). Prioritizing engagement with recreational anglers to reduce bycatch could be especially beneficial to sea turtle populations. Applying lessons learned from efforts to protect turtles in commercial fisheries may help meet the challenges that arise from the large, diffuse recreational fishing sector.

Association between metabolic syndrome and risk of both prevalent and incident frailty in older adults: Findings from The Irish Longitudinal Study on Ageing (TILDA).

OBJECTIVES: Metabolic syndrome (MetS) is a risk factor for cardiovascular disease, diabetes, and all-cause mortality. Frailty is a condition of decreased multi-system physiological reserve where one has increased vulnerability to stressors. This study aimed to examine if MetS is associated with prevalent and incident frailty over a 4-year follow-up period in an aged population. METHODS: This study used data from waves 1 (2009-2011) and 3 (2014-2015) of The Irish Longitudinal Study on Ageing. Those aged <50 years or without baseline health assessment data were excluded. Baseline MetS status was determined using the National Cholesterol Education Program Third Adult Treatment Panel criteria. Frailty status was identified at both waves, operationalised using Fried's frailty phenotype (FP) and Rockwood's frailty index (FI). Ordinal logistic regression examined the cross-sectional association between MetS and prevalent frailty status. Those with prevalent pre-frailty or frailty were excluded and ordinal logistic regression models examined the association between MetS and incident frailty. Lastly, MetS' longitudinal associations with the five individual components of Fried's FP were examined. Models were adjusted for age, sex, education, smoking, chronic disease history and renal function. RESULTS: Ordinal logistic regression models (n > 5100), showed MetS was associated with prevalent frailty as assessed by both FP (odds ratio (OR) 1.29, p < 0.001) and FI (OR 1.65, p < 0.001). Of those who were non-frail at baseline, 2247 participants had longitudinal FP data, while 3546 participants had longitudinal FI data. Models demonstrated that MetS was associated with an increased likelihood of incident frailty for both FP (OR 1.57, p < 0.001) and FI (OR 1.29, p = 0.014). MetS was found to be associated with incident low physical activity (OR 1.57, p = 0.001) and incident unintentional weight loss (OR 1.59, p = 0.025). CONCLUSIONS: MetS in those ≥50 years was found to be associated with an increased likelihood of incident frailty over a 4-year period, by 57 % when measured by FP and 29 % by FI. MetS should be considered a risk factor for frailty and be taken into considered in any comprehensive geriatric assessment given frailty's dynamic nature and MetS being potentially modifiable.

Development and characterization of a new monoclonal antibody against SARS-CoV-2 NSP12 (RdRp).

SARS-CoV-2 NSP12, the viral RNA-dependent RNA polymerase (RdRp), is required for viral replication and is a therapeutic target to treat COVID-19. To facilitate research on SARS-CoV-2 NSP12 protein, we developed a rat monoclonal antibody (CM12.1) against the NSP12 N-terminus that can facilitate functional studies. Immunoblotting and immunofluorescence assay (IFA) confirmed the specific detection of NSP12 protein by this antibody for cells overexpressing the protein. Although NSP12 is generated from the ORF1ab polyprotein, IFA of human autopsy COVID-19 lung samples revealed NSP12 expression in only a small fraction of lung cells including goblet, club-like, vascular endothelial cells, and a range of immune cells, despite wide-spread tissue expression of spike protein antigen. Similar studies using in vitro infection also generated scant protein detection in cells with established virus replication. These results suggest that NSP12 may have diminished steady-state expression or extensive posttranslation modifications that limit antibody reactivity during SARS-CoV-2 replication.