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Background: Antibody tests are critical in elucidating the epidemiology of the rapidly evolving Coronavirus Disease 2019 (COVID-19) global pandemic and in developing effective infection control measures. To avoid the significant risks posed by the use of tests with poor performance characteristics and associated supply chain disruptions, we developed an in-house SARS-CoV-2 total antibody test. The performance of our robustly validated test was compared with that of three commercial antibody tests, and our test was used to determine the seroprevalence of COVID-19 among healthcare workers and outpatients in Minnesota. Methods: To evaluate the clinical performance of our SARS-CoV-2 total antibody assay, 63 plasma and serum samples were analyzed from 34 SARS-CoV-2 PCR positive patients 4 - 35 days after symptom onset. Positive samples were analyzed for IgG titers in a follow up assay. Samples were analyzed from PCR negative (n=30) and presumed true negative pre-pandemic (n=207) patients. Among samples collected from patients >14 days after symptom onset, the assay had 100% diagnostic sensitivity (95% CI: 85·7 - 100·0%), 100% diagnostic specificity (95%CI: 98·5 - 100·0%), 100% positive predictive value (95% CI: 86·2 - 100·0%), and 100% negative predictive value (95%CI: 98·6 - 100·0%). The analytical specificity was 99·8% (95% CI: 98·92 - 99·97%), indicating minimal cross-reactivity. A screening study was conducted to ascertain the seroprevalence of COVID-19 among healthcare workers and outpatients in Minnesota. Findings: Serum collected from 1,282 healthcare workers (1,000 females, 280 males, 2 gender unreported; median age 41 years [range 18 - 73]) and 2,379 outpatients (1,460 females, 916 males, 3 gender unreported; median age 49 years [range 2 months - 93 years]) between April 13 and May 21, 2020 was analyzed using our SARS-CoV-2 total antibody assay. The COVID-19 seroprevalence was 2·96% (95% CI: 2·04 - 3·89%) among healthcare workers and 4·46% (95% CI: 3·63 - 5·28%) among outpatients from 35 clinics who had a previous COVID-19 exposure or symptoms. Interpretation: Rigorously validated serology tests with robust clinical and analytical performance, such as our in-house SARS-CoV-2 total antibody test, are of critical importance when conducting epidemiological studies to inform public health decisions during the COVID-19 pandemic. Funding: University of Minnesota Department of Laboratory Medicine and Pathology Declaration of Interests: The authors do not declare any competing interests. Ethics Approval Statement: The University of Minnesota Institutional Review Board (IRB) determined that this study was not considered human research.

Background We report a unique case of transient hyperphosphatasemia in a pediatric patient with a history of hepatic and skeletal abnormalities. Patient and Methods A 2-month old male was diagnosed with progressive familial intrahepatic cholestasis type-2 and osteoporosis after marked increases in liver function tests were noted at 1 month of age. He underwent a second liver transplantation at 1 y. The increased liver function test trend resolved a few weeks post-transplantation. Four months after successful liver transplantation, unexplained significant increases in alkaline phosphatase (ALP) were observed, and they persisted for almost 9 months. Among the etiologies under consideration for the isolated increased ALP activity were viral infections and macro-ALP. Results A persistent trend in abnormally increased ALP for 9 months was investigated leading to a confirmed diagnosis of transient hyperphosphatasemia (TH). Conclusion Pediatric post-liver transplant patients with skeletal and hepatic abnormalities including isolated markedly increased ALP activities represent a previously undescribed TH patient population. The 4.3% prevalence of TH in pediatric liver transplant recipients within our healthcare system is considerably higher than the previously reported prevalence of 2.1% for patients within the United States.

S-nitrosothiols (SNOs) are endogenous signaling molecules that have numerous beneficial effects on the airway via cyclic guanosine monophosphate-dependent and -independent processes. Healthy human airways contain SNOs, but SNO levels are lower in the airways of patients with cystic fibrosis (CF). In this study, we examined the interaction between SNOs and the molecular cochaperone C-terminus Hsc70 interacting protein (CHIP), which is an E3 ubiquitin ligase that targets improperly folded CF transmembrane conductance regulator (CFTR) for subsequent degradation. Both CFBE41o- cells expressing either wild-type or F508del-CFTR and primary human bronchial epithelial cells express CHIP. Confocal microscopy and IP studies showed the cellular colocalization of CFTR and CHIP, and showed that S-nitrosoglutathione inhibits the CHIP-CFTR interaction. SNOs significantly reduced both the expression and activity of CHIP, leading to higher levels of both the mature and immature forms of F508del-CFTR. In fact, SNO inhibition of the function and expression of CHIP not only improved the maturation of CFTR but also increased CFTR's stability at the cell membrane. S-nitrosoglutathione-treated cells also had more S-nitrosylated CHIP and less ubiquitinated CFTR than cells that were not treated, suggesting that the S-nitrosylation of CHIP prevents the ubiquitination of CFTR by inhibiting CHIP's E3 ubiquitin ligase function. Furthermore, the exogenous SNOs S-nitrosoglutathione diethyl ester and S-nitro-N-acetylcysteine increased the expression of CFTR at the cell surface. After CHIP knockdown with siRNA duplexes specific for CHIP, F508del-CFTR expression increased at the cell surface. We conclude that SNOs effectively reduce CHIP-mediated degradation of CFTR, resulting in increased F508del-CFTR expression on airway epithelial cell surfaces. Together, these findings indicate that S-nitrosylation of CHIP is a novel mechanism of CFTR correction, and we anticipate that these insights will allow different SNOs to be optimized as agents for CF therapy.

BACKGROUND: Implantable medical devices and hardware are prolific in medicine, but hardware associated infections remain a major issue. OBJECTIVE: To develop and evaluate a novel, biologic antimicrobial coating for medical implants. METHODS: Electrochemically compacted collagen sheets with and without crosslinked heparin were synthesized per a protocol developed by our group. Sheets were incubated in antibiotic solution (gentamicin or moxifloxacin) overnight, and in vitro activity was assessed with five-day diffusion assays against Pseudomonas aeruginosa. Antibiotic release over time from gentamicin-infused sheets was determined using in vitro elution and high performance liquid chromatography (HPLC). RESULTS: Collagen-heparin-antibiotic sheets demonstrated larger growth inhibition zones against P. aeruginosa compared to collagen-antibiotic alone sheets. This activity persisted for five days and was not impacted by rinsing sheets prior to evaluation. Rinsed collagen-antibiotic sheets did not produce any inhibition zones. Elution of gentamicin from collagen-heparin-gentamicin sheets was gradual and remained above the minimal inhibitory concentration for gentamicin-sensitive organisms for 29 days. Conversely, collagen-gentamicin sheets eluted their antibiotic load within 24 hours. Overall, heparin-associated sheets demonstrated larger inhibition zones against P. aeruginosa and prolonged elution profile via HPLC. CONCLUSION: We developed a novel, local antibiotic delivery system that could be used to coat medical implants/hardware in the future and reduce post-operative infections.

The global prevalence of type-two diabetes mellitus (T2DM) makes it a disease of public health concern. T2DM is strongly linked with insulin resistance caused by increased levels of visceral fat. Visceral fat secretes several adipocytokines that regulate body metabolism. Resistin is one of these adipocytokines which is encoded by the RETN gene. Herein, we tested the association of the RETN +299(G>A) and −420(C>G) single nucleotide polymorphisms (SNPs) with T2DM. T2DM patients (n=282) and healthy subjects (n=125) were included in the study. Subjects with metabolic syndromes other than diabetes were excluded. Genotyping of subjects was performed using PCR-RFLP. The +299(G>A) SNP was associated with T2DM (P=0.038). The AA genotype was higher in T2DM (17%) compared to controls (8%) with an odd ratio of 2.16 and 95% CI of 1.34 to 4.56. With respect to −420(C>G) SNP, no significant association was found with the risk of T2DM (P=0.128). The haplotype analysis of the examined SNPs indicated that the CA haplotype of the −420 and +299 SNPs in RETN was associated with T2DM risk (P=0.004; odd ration 4.0, 95% CI: 1.56–10.0). In conclusion, the present findings suggest a role of the RETN locus in modulating the risk of T2DM.

BackgroundThe LIAISON® QuantiFERON®-TB Gold Plus (QFT-Plus) assay, a fully automated chemiluminescence immunoassay (CLIA) system, has recently received FDA approval for the detection of interferon-γ (IFN-γ) on the LIAISON XL analyzer. Here, we evaluated the diagnostic performance of the LIAISON’s CLIA method in comparison to the widely used ELISA method using the Qiagen QuantiFERON-TB Gold Plus Blood Collection Tubes.MethodsHeparinized blood samples from 329 participants were categorized into 3 cohorts, including 15 with confirmed tuberculosis (TB) (active TB cohort), 129 non-TB (low-risk cohort), and 185 potential TB (mixed risk cohort). Samples were analyzed with both assays and results were interpreted according to the manufacturers’ criteria.ResultsThe LIAISON CLIA assay showed an overall agreement with the Qiagen ELISA assay in 13/14 (92.8%) samples among the active cohort, 93/95 (97.9%) among the low-risk cohort, with a Cohen’s kappa value of 0.76 and 0.74, respectively. Test results for 185 mixed risk cohort participants showed 97.8% agreement between the LIAISON and the Qiagen, with 17 positive, 163 negative, and 1 indeterminate. Four samples were discrepant; 3 of these were negative on the ELISA but positive on the CLIA assay.ConclusionOverall, the results were comparable (>92% agreement) in our study cohorts. In addition, our mixed risk cohort results showed an excellent agreement of 0.88 (Cohen’s kappa value) between the 2 assays. These findings suggest that the automated LIAISON QFT-Plus assay has a comparable diagnostic performance to the Qiagen assay and can be used for latent TB infection (LTBI) diagnosis.

The mechanisms by which transepithelial pressure changes observed during exercise and airway clearance can benefit lung health are challenging to study. Here, we have studied 117 mature, fully ciliated airway epithelial cell filters grown at air-liquid interface grown from 10 cystic fibrosis (CF) and 19 control subjects. These were exposed to cyclic increases in apical air pressure of 15 cmH2O for varying times. We measured the effect on proteins relevant to lung health, with a focus on the CF transmembrane regulator (CFTR). Immunoflourescence and immunoblot data were concordant in demonstrating that air pressure increased F508Del CFTR expression and maturation. This effect was in part dependent on the presence of cilia, on Ca2+ influx, and on formation of nitrogen oxides. These data provide a mechanosensory mechanism by which changes in luminal air pressure, like those observed during exercise and airway clearance, can affect epithelial protein expression and benefit patients with diseases of the airways.