Attenuation of SARS-CoV-2 replication and associated inflammation by concomitant targeting of viral and host cap 2'-O-ribose methyltransferases.

The SARS-CoV-2 infection cycle is a multistage process that relies on functional interactions between the host and the pathogen. Here, we repurposed antiviral drugs against both viral and host enzymes to pharmaceutically block methylation of the viral RNA 2'-O-ribose cap needed for viral immune escape. We find that the host cap 2'-O-ribose methyltransferase MTr1 can compensate for loss of viral NSP16 methyltransferase in facilitating virus replication. Concomitant inhibition of MTr1 and NSP16 efficiently suppresses SARS-CoV-2 replication. Using in silico target-based drug screening, we identify a bispecific MTr1/NSP16 inhibitor with anti-SARS-CoV-2 activity in vitro and in vivo but with unfavorable side effects. We further show antiviral activity of inhibitors that target independent stages of the host SAM cycle providing the methyltransferase co-substrate. In particular, the adenosylhomocysteinase (AHCY) inhibitor DZNep is antiviral in in vitro, in ex vivo, and in a mouse infection model and synergizes with existing COVID-19 treatments. Moreover, DZNep exhibits a strong immunomodulatory effect curbing infection-induced hyperinflammation and reduces lung fibrosis markers ex vivo. Thus, multispecific and metabolic MTase inhibitors constitute yet unexplored treatment options against COVID-19.

NTRC and thioredoxins m1/m2 underpin the light acclimation of plants on proteome and metabolome levels.

During photosynthesis, plants must manage strong fluctuations in light availability on different time scales, leading to long-term acclimation and short-term responses. However, little is known about the regulation and coordination of these processes and the modulators involved. In this study, we used proteomics, metabolomics, and reverse genetics to investigate how different light environmental factors, such as intensity or variability, affect long-term and short-term acclimation responses of Arabidopsis (Arabidopsis thaliana) and the importance of the chloroplast redox network in their regulation. In the wild type, high light, but not fluctuating light, led to large quantitative changes in the proteome and metabolome, accompanied by increased photosynthetic dynamics and plant growth. This finding supports light intensity as a stronger driver for acclimation than variability. Deficiencies in NADPH-thioredoxin reductase C (NTRC) or thioredoxins m1/m2, but not thioredoxin f1, almost completely suppressed the re-engineering of the proteome and metabolome, with both the induction of proteins involved in stress and redox responses and the repression of those involved in cytosolic and plastid protein synthesis and translation being strongly attenuated. Moreover, the correlations of protein or metabolite levels with light intensity were severely disturbed, suggesting a general defect in the light-dependent acclimation response, resulting in impaired photosynthetic dynamics. These results indicate a previously unknown role of NTRC and thioredoxins m1/m2 in modulating light acclimation at proteome and metabolome levels to control dynamic light responses. NTRC, but not thioredoxins m1/m2 or f1, also improves short-term photosynthetic responses by balancing the Calvin-Benson cycle in fluctuating light.

Over 89% of Patients Return to Work After Undergoing Arthroscopic or Open Latarjet Procedure for Anterior Shoulder Instability: A Systematic Review.

PURPOSE: To better define the rate of return to work in patients undergoing Latarjet surgery for anterior shoulder instability METHODS: A systematic review was performed in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines. PubMed, Cochrane Center Register of Controlled Trials (CENTRAL), and Scopus were queried in October 2023 with the following keywords: ((latarjet) OR (anterior shoulder instability)) AND ((work) or (compensation)). Articles were included if they reported return to work in patients (including military members) undergoing Latarjet surgery for traumatic anterior shoulder instability with bone loss and were written in English. This study was registered in Prospero (ID blinded). RESULTS: Six studies reporting on 419 shoulders were included in the review. Five studies reported on patients in the general population, and one reported on military members. Mean age ± standard deviation (SD) of patients ranged from 23.1 ± 5.8 - 32.0 ± 12.3 years old. Moreover, there were 286 primary Latarjet procedures, 131 revision Latarjet procedures, and 2 unspecified as primary or revision surgery. Mean glenoid bone loss ranged from 14.5% to 22.9%. Return to work rates ranged from 89.1% to 100%, with two studies reporting all patients were able to return to work. Among military members, 89.1% were able to return to duty. Mean time to return to work ranged from 8.69 - 34.8 weeks after surgery. Four studies also reported return to sport, which ranged from 60.9% to 100%. Mean time for returning to sport varied between 10.0 - 35.2 weeks after Latarjet procedure. CONCLUSION: Patients with anterior shoulder instability who undergo arthroscopic or open Latarjet procedure can expect high rates of return to work and sport. All studies reported return to work rates over 89% with 89.1% of military members were able to return to duty. LEVEL OF EVIDENCE: IV, systematic review of level III and Level IV studies.

Determining the Optimal Intramedullary Screw Canal Fill Ratio in Length Unstable Metacarpal Fractures: A Biomechanical Investigation.

PURPOSE: Intramedullary (IM) screw fixation is gaining popularity in the treatment of metacarpal fractures. Despite its rapid adoption, there is a paucity of evidence regarding parameters to optimize effectiveness. This study aimed to quantify the relationship between stability, IM screw size, and canal fill using a cadaveric model. METHODS: Thirty cadaveric metacarpals (14 index, 13 middle, and three ring fingers; mean age: 58.3 years, range: 48-70) were selected to allow for canal fill ratios of 0.7-1.1 for screws sized 3.0, 3.5, and 4.5 mm. Metacarpals underwent a 45° volar-dorsal osteotomy at the midpoint before fixation with an IM screw. Specimens were subjected to 100 cycles of loading at 10 N, 20 N, and 30 N before load-to-failure testing. Correlation coefficients for angular displacement on the final cycle at each load, peak load to failure, and average stiffness were assessed. RESULTS: Correlation coefficients for the angular displacement on the 100th cycle were as follows: 10 N, R = 0.62, 20 N, R = 0.57, and 30N, R = 0.58. Correlation values for peak load to failure as a function of canal fit were as follows: 3.0 mm, R = 0.5, 3.5 mm, R = 0.17, and 4.5 mm, R = 0.44. The canal fill ratio that intersected the line-of-best fit at an angular deformity of 10° was 0.74. Average peak forces for 3.0-, 3.5-, and 4.5-mm screws were 79.5, 136.5, and 179.6 N, respectively. Average stiffness for each caliber was 14.8, 33.4, and 52.3 N/mm. CONCLUSIONS: Increasing screw diameter and IM fill resulted in more stable fixation, but marginal gains were seen in ratios >0.9. A minimum fill ratio of 0.74 was sufficient to withstand forces of early active motion with angular deformity <10°. CLINICAL RELEVANCE: An understanding of the relationship of IM fill ratio of metacarpal screws to fracture stability may provide a framework for clinicians to optimally size these implants.

The impact of social deprivation on healthcare utilization patterns following rotator cuff repair.

BACKGROUND: Disparities in social determinants of health have been linked to worse patient reported outcomes, higher pain, and increased risk of revision surgery following rotator cuff repair. Identification of perioperative predictors of increased healthcare utilization is of particular interest to surgeons to improve outcomes and mitigate the total cost of care. The effect of social deprivation on healthcare utilization has not been fully characterized. METHODS: This is a retrospective review of a single institution's experience with primary rotator cuff repair between 2012 and 2020. Demographic variables (age, race, gender, American Society of Anesthesiologists (ASA) score) and healthcare utilization (hospital readmission, emergency department visits, follow-up visits, telephone calls) were recorded within 90 days of surgery. The Area Deprivation Index (ADI) was recorded, and patients were separated into terciles according to their relative level of social deprivation. Outcomes were then stratified based on ADI tercile and compared. RESULTS: A total of 1695 patients were included. The upper, middle, and lower terciles of ADI consisted of 410, 767, and 518 patients, respectively. The most deprived tercile had greater emergency department visitation and office visitation within 90 days of surgery relative to the least and intermediate deprived terciles. Higher levels of social deprivation were independent risk factors for increased emergency department (ED) visitation and follow-up visitation. There was no difference in 90-day readmission rates or telephone calls made between the least, intermediate, and most deprived patients. CONCLUSIONS: Patients with higher levels of deprivation demonstrated greater postoperative hospital utilization. We hope to use these results to identify risk factors for increased hospital use, guide clinical decision making, increase transparency, and manage patient outcomes following rotator cuff repair surgery.

The impact of social deprivation on rotator cuff repair outcomes.

BACKGROUND: Rotator cuff tears are a common orthopedic injury, and the role of social determinants of health (SDoH) in surgical outcomes remains underexplored. The goal of this study was to investigate the correlation between social deprivation, measured by the Area Deprivation Index (ADI), and outcomes following arthroscopic rotator cuff repair. METHODS: We conducted a retrospective chart review on patients undergoing primary arthroscopic rotator cuff repair at a level 1 academic center between 2006 and 2019. Patient demographics (age, gender, race), comorbidities, ADIs, range of motion, visual analog pain scores, and patient-reported outcomes (Simple Shoulder Test [SST], American Shoulder and Elbow Surgeons Standardized Shoulder Assessment Form [ASES], and Quick Disabilities of the Arm, Shoulder, and Hand questionnaire [QuickDASH]) were collected. Patients were stratified into terciles based on their relative level of deprivation. Statistical analysis was performed using analysis of variance, t tests, χ2 tests, and univariate or multivariate logistic regression. RESULTS: A total of 322 patients were included in this study. The most deprived group had a higher prevalence of diabetes compared to the least and intermediately deprived group (P < .001). Massive tear occurrence was greater in the least deprived group (P = .003) compared to the most deprived group. There was no difference in objective outcomes between groups. Patient-reported outcomes (SST, ASES, and QuickDASH scores) were worse in the most deprived group compared with the least and intermediate deprived groups. CONCLUSION: Social deprivation significantly affects patient-reported outcomes in rotator cuff repair surgery. Although clinician-reported outcomes were consistent, patients' perceptions varied based on social determinants. Integrating SDoH considerations in orthopedic care is a promising next step in securing equitable approaches. However, more research is needed to validate and expand these findings.

SARS-CoV-2 Variants of Concern Hijack IFITM2 for Efficient Replication in Human Lung Cells.

It has recently been shown that an early SARS-CoV-2 isolate (NL-02-2020) hijacks interferon-induced transmembrane proteins (IFITMs) for efficient replication in human lung cells, cardiomyocytes, and gut organoids. To date, several "variants of concern" (VOCs) showing increased infectivity and resistance to neutralization have emerged and globally replaced the early viral strains. Here, we determined whether the five current SARS-CoV-2 VOCs (Alpha, Beta, Gamma, Delta, and Omicron) maintained the dependency on IFITM proteins for efficient replication. We found that depletion of IFITM2 strongly reduces viral RNA production by all VOCs in the human epithelial lung cancer cell line Calu-3. Silencing of IFITM1 had modest effects, while knockdown of IFITM3 resulted in an intermediate phenotype. Strikingly, depletion of IFITM2 generally reduced infectious virus production by more than 4 orders of magnitude. In addition, an antibody directed against the N terminus of IFITM2 inhibited SARS-CoV-2 VOC replication in induced pluripotent stem cell (iPSC)-derived alveolar epithelial type II cells, thought to represent major viral target cells in the lung. In conclusion, endogenously expressed IFITM proteins (especially IFITM2) are critical cofactors for efficient replication of genuine SARS-CoV-2 VOCs, including the currently dominant Omicron variant. IMPORTANCE Recent data indicate that SARS-CoV-2 requires endogenously expressed IFITM proteins for efficient infection. However, the results were obtained with an early SARS-CoV-2 isolate. Thus, it remained to be determined whether IFITMs are also important cofactors for infection of emerging SARS-CoV-2 VOCs that outcompeted the original strains in the meantime. This includes the Omicron VOC, which currently dominates the pandemic. Here, we show that depletion of endogenous IFITM2 expression almost entirely prevents productive infection of Alpha, Beta, Gamma, Delta, and Omicron SARS-CoV-2 VOCs in human lung cells. In addition, an antibody targeting the N terminus of IFITM2 inhibited SARS-CoV-2 VOC replication in iPSC-derived alveolar epithelial type II cells. Our results show that SARS-CoV-2 VOCs, including the currently dominant Omicron variant, are strongly dependent on IFITM2 for efficient replication, suggesting a key proviral role of IFITMs in viral transmission and pathogenicity.

A dominant mutation in ß-AMYLASE1 disrupts nighttime control of starch degradation in Arabidopsis leaves.

Arabidopsis (Arabidopsis thaliana) leaves possess a mechanism that couples the rate of nighttime starch degradation to the anticipated time of dawn, thus preventing premature exhaustion of starch and nighttime starvation. To shed light on the mechanism, we screened a mutagenized population of a starvation reporter line and isolated a mutant that starved prior to dawn. The mutant had accelerated starch degradation, and the rate was not adjusted to time of dawn. The mutation responsible led to a single amino acid change (S132N) in the starch degradation enzyme BETA-AMYLASE1 (BAM1; mutant allele named bam1-2D), resulting in a dominant, gain-of-function phenotype. Complete loss of BAM1 (in bam1-1) did not affect rates of starch degradation, while expression of BAM1(S132N) in bam1-1 recapitulated the accelerated starch degradation phenotype of bam1-2D. In vitro analysis of recombinant BAM1 and BAM1(S132N) proteins revealed no differences in kinetic or stability properties, but in leaf extracts, BAM1(S132N) apparently had a higher affinity than BAM1 for an established binding partner required for normal rates of starch degradation, LIKE SEX FOUR1 (LSF1). Genetic approaches showed that BAM1(S132N) itself is likely responsible for accelerated starch degradation in bam1-2D and that this activity requires LSF1. Analysis of plants expressing BAM1 with alanine or aspartate rather than serine at position 132 indicated that the gain-of-function phenotype is not related to phosphorylation status at this position. Our results strengthen the view that control of starch degradation in wild-type plants involves dynamic physical interactions of degradative enzymes and related proteins with a central role for complexes containing LSF1.

Absence of a molecular circadian clock in the preimplantation embryo is a conserved characteristic in the mammal.

In brief: It is not known when a functional circadian clock is established in the developing embryo. Lack of expression of key genes involved in the clock mechanism is indicative that a functional circadian clock mechanism is absent in the mammalian preimplantation embryo through the blastocyst stage of development. Abstract: An embryonic circadian clock could conceivably organize cellular and developmental events temporally and in synchrony with other circadian rhythms in the mother. The hypothesis that a functional molecular clock exists in the preimplantation bovine, pig, human, and mouse embryo was tested by using publicly available RNAseq datasets to examine developmental changes in expression of the core genes responsible for the circadian clock - CLOCK, ARNTL, PER1, PER2, CRY1, and CRY2. In general, the transcript abundance of each gene decreased as development advanced to the blastocyst stage. The most notable exception was for CRY2, where transcript abundance was low and constant from the two-cell or four-cell to the blastocyst stage. Developmental patterns were generally the same for all species although there were some species-specific patterns such as an absence of PER1 expression in the pig, an increase in ARNTL expression at the four-cell stage in human, and an increase in expression of Clock and Per1 from the zygote to two-cell stage in the mouse. Analysis of intronic reads (indicative of embryonic transcription) for bovine embryos indicated an absence of embryonic transcription. Immunoreactive CRY1 was not detected in the bovine blastocyst. Results indicate that the preimplantation mammalian embryo lacks a functional intrinsic clock although specific components of the clock mechanism could conceivably play a role in other functions in the embryo.

Ten rapid antigen tests for SARS-CoV-2 widely differ in their ability to detect Omicron-BA.4 and -BA.5.

Since late 2021, the variant landscape of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been dominated by the variant of concern (VoC) Omicron and its sublineages. We and others have shown that the detection of Omicron-BA.1 and -BA.2-positive respiratory specimens by rapid antigen tests (RATs) is impaired compared to Delta VoC-containing samples. Here, in a single-center retrospective laboratory study, we evaluated the performance of ten most commonly used RATs for the detection of Omicron-BA.4 and -BA.5 infections. We used 171 respiratory swab specimens from SARS-CoV-2 RNA-positive patients, of which 71 were classified as BA.4 and 100 as BA.5. All swabs were collected between July and September 2022. 50 SARS-CoV-2 PCR-negative samples from healthy individuals, collected in October 2022, showed high specificity in 9 out of 10 RATs. When assessing analytical sensitivity using clinical specimens, the 50% limit of detection (LoD50) ranged from 7.6 × 104 to 3.3 × 106 RNA copies subjected to the RATs for BA.4 compared to 6.8 × 104 to 3.0 × 106 for BA.5. Overall, intra-assay differences for the detection of these two Omicron subvariants were not significant for both respiratory swabs and tissue culture-expanded virus isolates. In contrast, marked heterogeneity was observed among the ten RATs: to be positive in these point-of-care tests, up to 443-fold (BA.4) and up to 56-fold (BA.5) higher viral loads were required for the worst performing RAT compared to the best performing RAT. True-positive rates for Omicron-BA.4- or -BA.5-containing specimens in the highest viral load category (Ct values < 25) ranged from 94.3 to 34.3%, dropping to 25.6 to 0% for samples with intermediate Ct values (25-30). We conclude that the high heterogeneity in the performance of commonly used RATs remains a challenge for the general public to obtain reliable results in the evolving Omicron subvariant-driven pandemic.

Automated antigen assays display a high heterogeneity for the detection of SARS-CoV-2 variants of concern, including several Omicron sublineages.

Diagnostic tests for direct pathogen detection have been instrumental to contain the severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) pandemic. Automated, quantitative, laboratory-based nucleocapsid antigen (Ag) tests for SARS-CoV-2 have been launched alongside nucleic acid-based test systems and point-of-care (POC) lateral-flow Ag tests. Here, we evaluated four commercial Ag tests on automated platforms for the detection of different sublineages of the SARS-CoV-2 Omicron variant of concern (VoC) (B.1.1.529) in comparison with "non-Omicron" VoCs. A total of 203 Omicron PCR-positive respiratory swabs (53 BA.1, 48 BA.2, 23 BQ.1, 39 XBB.1.5 and 40 other subvariants) from the period February to March 2022 and from March 2023 were examined. In addition, tissue culture-expanded clinical isolates of Delta (B.1.617.2), Omicron-BA.1, -BF.7, -BN.1 and -BQ.1 were studied. These results were compared to previously reported data from 107 clinical "non-Omicron" samples from the end of the second pandemic wave (February to March 2021) as well as cell culture-derived samples of wildtype (wt) EU-1 (B.1.177), Alpha VoC (B.1.1.7) and Beta VoC (B.1.351)). All four commercial Ag tests were able to detect at least 90.9% of Omicron-containing samples with high viral loads (Ct < 25). The rates of true-positive test results for BA.1/BA.2-positive samples with intermediate viral loads (Ct 25-30) ranged between 6.7% and 100.0%, while they dropped to 0 to 15.4% for samples with low Ct values (> 30). This heterogeneity was reflected also by the tests' 50%-limit of detection (LoD50) values ranging from 44,444 to 1,866,900 Geq/ml. Respiratory samples containing Omicron-BQ.1/XBB.1.5 or other Omicron subvariants that emerged in 2023 were detected with enormous heterogeneity (0 to 100%) for the intermediate and low viral load ranges with LoD50 values between 23,019 and 1,152,048 Geq/ml. In contrast, detection of "non-Omicron" samples was more sensitive, scoring positive in 35 to 100% for the intermediate and 1.3 to 32.9% of cases for the low viral loads, respectively, corresponding to LoD50 values ranging from 6181 to 749,792 Geq/ml. All four assays detected cell culture-expanded VoCs Alpha, Beta, Delta and Omicron subvariants carrying up to six amino acid mutations in the nucleocapsid protein with sensitivities comparable to the non-VoC EU-1. Overall, automated quantitative SARS-CoV-2 Ag assays are not more sensitive than standard rapid antigen tests used in POC settings and show a high heterogeneity in performance for VoC recognition. The best of these automated Ag tests may have the potential to complement nucleic acid-based assays for SARS-CoV-2 diagnostics in settings not primarily focused on the protection of vulnerable groups. In light of the constant emergence of new Omicron subvariants and recombinants, most recently the XBB lineage, these tests' performance must be regularly re-evaluated, especially when new VoCs carry mutations in the nucleocapsid protein or immunological and clinical parameters change.

Variable detection of Omicron-BA.1 and -BA.2 by SARS-CoV-2 rapid antigen tests.

During 2022, the COVID-19 pandemic has been dominated by the variant of concern (VoC) Omicron (B.1.1.529) and its rapidly emerging subvariants, including Omicron-BA.1 and -BA.2. Rapid antigen tests (RATs) are part of national testing strategies to identify SARS-CoV-2 infections on site in a community setting or to support layman's diagnostics at home. We and others have recently demonstrated an impaired RAT detection of infections caused by Omicron-BA.1 compared to Delta. Here, we evaluated the performance of five SARS-CoV-2 RATs in a single-centre laboratory study examining a total of 140 SARS-CoV-2 PCR-positive respiratory swab samples, 70 Omicron-BA.1 and 70 Omicron-BA.2, as well as 52 SARS-CoV-2 PCR-negative swabs collected from March 8th until April 10th, 2022. One test did not meet minimal criteria for specificity. In an assessment of the analytical sensitivity in clinical specimen, the 50% limit of detection (LoD50) ranged from 4.2 × 104 to 9.2 × 105 RNA copies subjected to the RAT for Omicron-BA.1 compared to 1.3 × 105 to 1.5 × 106 for Omicron-BA.2. Overall, intra-assay differences for the detection of Omicron-BA.1-containing and Omicron-BA.2-containing samples were non-significant, while a marked overall heterogeneity among the five RATs was observed. To score positive in these point-of-care tests, up to 22-fold (LoD50) or 68-fold (LoD95) higher viral loads were required for the worst performing compared to the best performing RAT. The rates of true-positive test results for these Omicron subvariant-containing samples in the highest viral load category (Ct values < 25) ranged between 44.7 and 91.1%, while they dropped to 8.7 to 22.7% for samples with intermediate Ct values (25-30). In light of recent reports on the emergence of two novel Omicron-BA.2 subvariants, Omicron-BA.2.75 and BJ.1, awareness must be increased for the overall reduced detection rate and marked differences in RAT performance for these Omicron subvariants.

Secondary zoonotic dog-to-human transmission of SARS-CoV-2 suggested by timeline but refuted by viral genome sequencing.

PURPOSE: The risk of secondary zoonotic transmission of SARS-CoV-2 from pet animals remains unclear. Here, we report on a 44 year old Caucasian male presenting to our clinic with COVID-19 pneumonia, who reported that his dog displayed respiratory signs shortly prior to his infection. The dog tested real-time-PCR (RT-PCR) positive for SARS-CoV-2 RNA and the timeline of events suggested a transmission from the dog to the patient. METHODS: RT-PCR and serological assays were used to confirm SARS-CoV-2 infection in the nasopharyngeal tract in the dog and the patient. We performed SARS-CoV-2-targeted amplicon-based next generation sequencing of respiratory samples from the dog and patient for sequence comparisons. RESULTS: SARS-CoV-2 infection of the dog was confirmed by three independent PCR-positive pharyngeal swabs and subsequent seroconversion. Sequence analysis identified two separate SARS-CoV-2 lineages in the canine and the patient's respiratory samples. The timeline strongly suggested dog-to-human transmission, yet due to the genetic distance of the canine and the patient's samples paired-transmission was highly unlikely. CONCLUSION: The results of this case support current knowledge about the low risk of secondary zoonotic dog-to-human transmissions of SARS-CoV-2 and emphasizes the strength of genomic sequencing in deciphering viral transmission chains.

Impaired immune responses and prolonged viral replication in lung allograft recipients infected with SARS-CoV-2 in the early phase after transplantation.

PURPOSE: Lung transplant recipients are at increased risk of severe disease following infection with severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) due to high-dose immunosuppressive drugs and the lung is the main organ affected by Coronavirus disease 2019 (COVID-19). Several studies have confirmed increased SARS-CoV-2-related mortality and morbidity in patients living with lung allografts; however, detailed immunological studies of patients with SARS-CoV-2 infection in the early phase following transplantation remain scarce. METHODS: We investigated patients who were infected with SARS-CoV-2 in the early phase (18-103 days) after receiving double-lung allografts (n = 4, LuTx) in comparison to immunocompetent patients who had not received solid organ transplants (n = 88, noTx). We analyzed SARS-CoV-2-specific antibody responses against the SARS-CoV-2 spike and nucleocapsid proteins using enzyme-linked immunosorbent assays (ELISA), chemiluminescence immunoassays (CLIA), and immunoblot assays. T cell responses were investigated using Elispot assays. RESULTS: One LuTx patient suffered from persistent infection with fatal outcome 122 days post-infection despite multiple interventions including remdesivir, convalescent plasma, and the monoclonal antibody bamlanivimab. Two patients experienced clinically mild disease with prolonged viral shedding (47 and 79 days), and one patient remained asymptomatic. Antibody and T cell responses were significantly reduced or undetectable in all LuTx patients compared to noTx patients. CONCLUSION: Patients in the early phase following lung allograft transplantation are vulnerable to infection with SARS-CoV-2 due to impaired immune responses. This patient population should be vaccinated before LuTx, protected from infection post-LuTx, and in case of infection treated generously with currently available interventions.

Multicenter clinical experience with non-invasive cell-free DNA screening for monosomy X and related X-chromosome variants.

OBJECTIVE: We aimed to investigate how the presence of fetal anomalies and different X chromosome variants influences Cell-free DNA (cfDNA) screening results for monosomy X. METHODS: From a multicenter retrospective survey on 673 pregnancies with prenatally suspected or confirmed Turner syndrome, we analyzed the subgroup for which prenatal cfDNA screening and karyotype results were available. A cfDNA screening result was defined as true positive (TP) when confirmatory testing showed 45,X or an X-chromosome variant. RESULTS: We had cfDNA results, karyotype, and phenotype data for 55 pregnancies. cfDNA results were high risk for monosomy X in 48/55, of which 23 were TP and 25 were false positive (FP). 32/48 high-risk cfDNA cases did not show fetal anomalies. Of these, 7 were TP. All were X-chromosome variants. All 16 fetuses with high-risk cfDNA result and ultrasound anomalies were TP. Of fetuses with abnormalities, those with 45,X more often had fetal hydrops/cystic hygroma, whereas those with "variant" karyotypes had different anomalies. CONCLUSION: Both, 45,X or X-chromosome variants can be detected after a high-risk cfDNA result for monosomy X. When there are fetal anomalies, the result is more likely a TP. In the absence of fetal anomalies, it is most often an FP or X-chromosome variant.

Dupuytren Disease: Surgical Tips, Optimizing Outcomes, and Cost Considerations.

Dupuytren disease is associated with benign fibroproliferative changes to the palmar fascia of the hand sometimes resulting in progressive contractures of the fingers. The earliest descriptions of these contractures date back to the 18th century. Much has been learned about the condition since the clawing condition was first described; however, optimal treatment still poses significant challenges to modern-day surgeons. It is important to examine the treatment options for Dupuytren disease and highlight the current evidence, techniques, and cost considerations of open fasciectomy, needle aponeurotomy, and recently described minimally invasive treatment.

Intramedullary Metacarpal Fracture Fixation: A Biomechanical Study of Screw Diameter and Comparison With Intramedullary Wire Stabilization.

PURPOSE: Interest in intramedullary metacarpal fracture fixation (IMFF) with screws is increasing. However, the optimal screw diameter for fracture fixation is not yet established. In theory, larger screws should be more stable, but there is concern about long-term sequelae of larger metacarpal head defects and extensor mechanism injury created during insertion as well as implant cost. Therefore, the purpose of this study was to compare different diameter screws for IMFF to a popular and more cost-effective alternative of intramedullary wiring. METHODS: Thirty-two cadaveric metacarpals were used in a transverse metacarpal shaft fracture model. Treatment groups consisted of IMFF with 3.0 × 60 mm, 3.5 x 60 mm, and 4.5 x 60 mm screws as well as 4 1.1-mm intramedullary wires. Cyclic cantilever bending was performed with the metacarpals mounted at 45° to simulate physiologic loading. Cyclical loading at 10, 20, and 30 N was performed to determine fracture displacement, stiffness, and ultimate force. RESULTS: At 10, 20, and 30 N of cyclical loading, all screw diameters tested provided similar stability as measured by fracture displacement and were superior to the wire group. However, ultimate force under load to failure testing was similar between the 3.5- and 4.5-mm screws and superior to 3.0-mm screws and wires. CONCLUSIONS: For IMFF, 3.0, 3.5, and 4.5-mm diameter screws provide adequate stability for early active motion and are superior to wires. When comparing the different screw diameters, 3.5- and 4.5-mm diameter screws offer similar construct stability and strength superior to the 3.0-mm diameter screw. Therefore, to minimize metacarpal head morbidity, smaller screw diameters may be preferable. CLINICAL RELEVANCE: This study suggests that IMFF with screws is biomechanically superior to wires in cantilever bending strength in the transverse fracture model. However, smaller screws may be sufficient to permit early active motion while minimizing metacarpal head morbidity.

Reverse shoulder arthroplasty in upper extremity ambulators: a matched-cohort analysis.

BACKGROUND: Upper extremity ambulators (UEAs) who require prolonged use of assistive devices for mobility have a high incidence of shoulder pathology secondary to increased stress across the shoulder joint with upper extremity weight-bearing. Reverse shoulder arthroplasty (RSA) for rotator cuff arthropathy has historically been associated with increased complications in UEA, but more recent studies have shown more promising outcomes. The objective of this study is to evaluate clinical outcomes and complication rates between these 2 groups to define the relative risk of RSA in the UEA population and identify opportunities to improve treatment outcomes. METHODS: An institutional review board-approved retrospective chart review was performed in patients who underwent RSA at our institution by the senior author from 2004 to 2019. UEAs were defined as patients who used regular upper extremity assistive devices for community ambulation before initial consultation for the surgical extremity. Pre- and postoperative range of motion, visual analog scale scores, American Shoulder and Elbow Surgeons scores, Constant-Murley scores, and Simple Shoulder Test scores were measured at defined intervals. Complications including infection, instability, and need for revision surgery were also compared. All patients were followed for a minimum of 2 years postoperatively. RESULTS: A total of 159 RSA procedures (70 UEAs, 89 controls) were performed during the study period. On average, UEA patients had more preoperative pain and less shoulder function than controls, with statistically significant differences in visual analog scores (6.897 vs. 5.532, P = .0010) and American Shoulder and Elbow Surgeons scores (33.50 vs. 40.20, P = .0290), respectively. Despite the lower baseline values, UEA patients experienced excellent postoperative improvement, leading to similar postoperative pain and shoulder function except for a lower average forward flexion in the UEA group (127° vs. 135°, P = .0354). Notching and complication rates were also similar between the 2 groups, with notching rates of 59% and 50% and complication rates of 14.3% and 13.5% in the UEA and control groups, respectively. CONCLUSIONS: RSA in the UEA population can achieve similar pain and functional outcomes as compared with age-matched controls without a significant increase in complication rates; however, further studies are required to assess long-term comparative outcomes in this challenging patient population.

Volar versus dorsal approach for supinator to posterior interosseous nerve transfer: An anatomical study in cadavers.

INTRODUCTION: Supinator to posterior interosseous nerve (SPIN) transfer allows reconstruction of finger/thumb extension and thumb abduction for low radial nerve palsy, incomplete C6 tetraplegia, and brachial plexus injury affecting C7-T1. No study has compared dorsal versus volar approach to perform SPIN transfer. This comparison is studied in the present work, assessing supinator motor branch length and ability to achieve nerve transfer from either approach. METHODS: Ten fresh frozen cadavers were randomly allocated to receive either a dorsal or volar approach to PIN and supinator radial and ulnar branches (RB = radial, UB = ulnar). Supinator head innervation patterns were documented. RB and UB lengths, forearm lengths measured from ulnar styloid to olecranon, visualization of extensor carpi radialis brevis (ECRB) motor nerve without additional dissection, and ability to perform tension-free nerve transfer were assessed. RESULTS: Nine of 10 specimens had supinator branches innervating both heads. The ECRB nerve was visualized in all volar but only one dorsal approach. No significant differences in forearm length were found. Volar with elbow extended: mean RB length was 35 ± 7.8 mm and UB was 37.8 ± 9.3 mm. Dorsal with elbow extended: mean RB length was 30 ± 4.1 mm and UB was 38.8 ± 7.3 mm. Dorsal with elbow flexed 90°: RB was 25.6 ± 3.8 mm and UB was 34.8 ± 4.8 mm. No significant differences were found in branch lengths between approaches (dorsal vs. volar UB, p = .339; dorsal vs. volar RB, p = .117). All limbs achieved tension-free coaptation. CONCLUSION: Neither approach demonstrated superiority in achieving tension-free nerve transfer. Volar permitted immediate identification of ECRB nerve whereas this was only visualized in one dorsal specimen without additional dissection. Overall, the volar approach allows direct coaptation in elbow extension, mimicking maximal physiologic tension for neurorrhaphy. It simultaneously permits additional procedures for pinch reconstruction via single exposure, circumventing limb/microscope maneuvering, dorsal dissection, and increased operative time. Ultimate choice of approach should depend on surgeon familiarity and potential need for additional simultaneous transfers.

Multielectrode Arrays at Wafer-Level for Miniaturized Sensors Applications: Electrochemical Growth of Ag/AgCl Reference Electrodes.

In this study, a range of miniaturized Ag/AgCl reference electrodes with various layouts were successfully fabricated on wafer-level silicon-based substrates with metallic intermediate layers by precisely controlling the electrochemical deposition of Ag, followed by electrochemical chlorination of the deposited Ag layer. The structure, as well as the chemical composition of the electrode, were characterized with SEM & EDS. The results showed that the chlorination is very sensitive to the applied electric field and background solution. Potentiostatic chlorination, in combination with an adjusted mushroom-shaped Ag sealing deposition, enabled the formation of electrochemical usable Ag/AgCl layers. The stability of the electrodes was tested using open circuit potential (OCP) measurement. The results showed that the reference electrodes stayed stable for 300 s under 3 M KCl solution. The first stage study showed that the stability of the Ag/AgCl reference electrode in a chip highly depends on chip size design, chlorination conditions, and a further protection layer.