Epigenetics may characterize asymptomatic COVID-19 infection.

RT-PCR is the foremost clinical test for diagnosis of COVID-19. Unfortunately, PCR-based testing has limitations and may not result in a positive test early in the course of infection before symptoms develop. Enveloped RNA viruses, such as coronaviruses, alter peripheral blood methylation and DNA methylation signatures may characterize asymptomatic versus symptomatic infection. We used Illumina's Infinium MethylationEPIC BeadChip array to profile peripheral blood samples from 164 patients who tested positive for SARS-CoV-2 by RT-PCR, of whom 8 had no symptoms. Epigenome-wide association analysis identified 10 methylation sites associated with infection and a quantile-quantile plot showed little inflation. These preliminary results suggest that differences in methylation patterns may distinguish asymptomatic from symptomatic infection.

Pulse oximeter plethysmograph waveform and automated oscillometric sphygmomanometer for ankle-brachial index measurement.

OBJECTIVES: There are limited non-invasive methods to assess lower extremity arterial injuries in the emergency department (ED) and pre-hospital setting. The ankle-brachial index (ABI) requires careful auscultation by Doppler, an approach made difficult in noisy environments. We sought to determine the agreement of the ABI measured using the pulse oximeter plethysmograph waveform (Pleth) with auscultation by Doppler in a controlled setting. A secondary outcome sought to examine the agreement of ABI by automated oscillometric sphygmomanometer (AOS) with Doppler. METHODS: We measured blood pressure in the right upper and lower extremities of healthy volunteers using: (1) Doppler and manual sphygmomanometer; (2) Pleth and manual sphygmomanometer; and (3) AOS. The Bland-Altman approach to assessing agreement between methods was used comparing mean differences between ABI pairs to their means for Doppler versus Pleth and Doppler versus AOS. The intraclass correlation coefficient (ICC) from mixed effects models examined intra- and inter-rater reliability. RESULTS: Among 100 participants with normal ABI the mean ABI (95%CI) were Doppler 1.11 (0.90-1.33), Pleth 1.10 (0.91-1.30), and AOS 1.10 (0.90-1.30). The ABI difference (95% CI for limits of agreement) were 0.01 (-0.20,0.18) for Doppler-Pleth and 0.02 (-0.26, 0.22) for Doppler-AOS. The ICC for the Doppler-Pleth comparison (ICC = 0.56, 95% CI 0.47-0.63) was greater than for the Doppler-AOS (ICC = 0.32, 95% CI 0.19-0.43). CONCLUSIONS: The ABI measured using the Pleth has a high level of agreement with measurement by Doppler. The AOS and Doppler have good agreement with greater measurement variability. Pleth and AOS may be reasonable alternatives to Doppler for ABI.

Two faces of patient advocacy: the current controversy in newborn screening.

Newborn screening programmes began in the 1960s, have traditionally been conducted without parental permission and have grown dramatically in the last decade. Whether these programmes serve patients' best interests has recently become a point of controversy. Privacy advocates, concerned that newborn screening infringes upon individual liberties, are demanding fundamental changes to these programmes. These include parental permission and limiting the research on the blood samples obtained, an agenda at odds with the viewpoints of newborn screening advocates. This essay presents the history of newborn screening in the USA, with attention to factors that have contributed to concerns about these programmes. The essay suggests that the rapid increase in the number of disorders screened for and the addition of research without either public knowledge or informed consent were critical to the development of resistance to mandatory newborn screening and research. Future newborn screening initiatives should include public education and comment to ensure continued support.

Correlation between locally versus centrally processed serum procalcitonin during emergency department research evaluation of febrile infants aged 0-60 days.

Introduction: Procalcitonin (PCT) is a useful biomarker in the initial evaluation of febrile infants for serious bacterial infections (SBIs). However, PCT is not always available locally and must at times be frozen and shipped to a reference laboratory for research studies. We sought to compare PCT measured locally versus centrally at a reference laboratory during a research study. Materials and methods: This was a secondary analysis of a multicenter study of febrile infants ≤60 days evaluated for SBIs from June 2016 to April 2019. A PCT cutoff value of 0.5 ng/mL was used to stratify infants at low-versus high-risk of SBIs. Statistical analyses consisted of Spearman's correlation, Bland-Altman difference plotting, Passing-Bablok regression, Deming regression, and Fisher's exact testing at the 0.5 ng/mL threshold. Results: 241 febrile infants had PCT levels measured both locally and at the reference laboratory. PCT levels measured locally on 5 different platforms and from the frozen research samples demonstrated strong Spearman's correlation (ρ = 0.83) and had similar mean PCT values with an average relative difference of 0.02%. Eleven infants with SBIs had PCT values < 0.5 ng/mL in both the clinical and research samples. Six other infants had differences in SBI prediction based on PCT values at the 0.5 ng/mL threshold between the clinical and research platforms. Conclusions: We found no significant differences in detection of febrile infants at high risk for SBI based on locally (on multiple platforms) versus centrally processed PCT. Testing at a central reference laboratory after freezing and shipping is an accurate and reliable alternative for research studies or when rapid turnaround is not required.

Accessing and utilizing clinical and genomic data from an electronic health record data warehouse.

Electronic health records (EHRs) and linked biobanks have tremendous potential to advance biomedical research and ultimately improve the health of future generations. Repurposing EHR data for research is not without challenges, however. In this paper, we describe the processes and considerations necessary to successfully access and utilize a data warehouse for research. Although imperfect, data warehouses are a powerful tool for harnessing a large amount of data to phenotype disease. They will have increasing relevance and applications in clinical research with growing sophistication in processes for EHR data abstraction, biobank integration, and cross-institutional linkage.

Immune mechanisms associated with sex-based differences in severe COVID-19 clinical outcomes.

BACKGROUND: Although biological males and females are equally likely to become infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), evidence has mounted that males experience higher severity and fatality compared to females. MAIN: The objective of this review is to examine the existing literature on biological mechanisms underlying sex-based differences that could contribute to SARS-CoV-2 infection clinical outcomes. Sex-based differences in immunologic response and hormonal expression help explain the differences in coronavirus disease 2019 (COVID-19) outcomes observed in biological males and females. X inactivation facilitates a robust immune response to COVID-19 in females, who demonstrate a more profound antibody response and faster recovery when compared to males. Low testosterone levels also help explain the dysregulated inflammatory response and poor outcomes observed in some males with COVID-19. Gender differences in health expression and behaviors further compound these observed differences. CONCLUSION: Understanding the biology of sex-based differences in COVID-19 severity and mortality could help inform preventative measures, treatment decisions, and development of personalized, sex-specific therapies.

Metabolomic Evaluation of N-Acetyl-p-Benzoquinone Imine Protein Adduct Formation with Therapeutic Acetaminophen Administration: Sex-based Physiologic Differences.

BACKGROUND: Acetaminophen (APAP)-associated transaminase elevation, induced by N-acetyl-p-benzoquinone imine (NAPQI) protein adduction, remains an area of research interest. Distinct from known genetic, physiologic, and dosage associations dictating severity of hepatic injury, no known factors predict an absence of protein adduct formation at therapeutic APAP dosing. HYPOTHESIS: Sex-based physiology is predictive of APAP-induced protein adduct formation and differential metabolite expression at therapeutic doses. METHODS: This retrospective study interrogated serum samples collected for a prior study investigating fluctuations of alanine aminotransferase (ALT) over time with 4G daily APAP dosing for ≥ 16 days in subjects from Denver, Colorado. Subjects were grouped by adduct formation (n = 184) vs no adducts (n = 20). Samples were run on ultra-high-performance liquid chromatography mass spectrometry from study days 0, 7, 16, and 31. Significant metabolite expressions were identified using t-tests with false discovery rate correction (FDR), partial least squares discriminant, and ANOVA simultaneous comparison analyses. Demographic and clinical data were explored using t-tests with FDR (age, weight, BMI, ALT) and Chi-square (sex, ethnicity, race) analyses. RESULTS: In pre-treatment samples, relative quantitation caprylic acid was expressed ninefold higher and 6-carboxyhexanoate was expressed threefold lower in subjects who did not develop adducts. Lactate had greater expression in the no adducts group (p = 0.001). Using absolute quantitation, glutathione was expressed 2.6-fold greater among no adduct subjects. Odds of males developing NAPQI protein adducts at therapeutic APAP dosing were 5.91 times lower than females (95% CI = 2.3-14.9; p = 0.0001). CONCLUSION: Multiple metabolites were differentially expressed based on adduct group and sex. Metabolites were identified unique to adduct development independent of sex. At therapeutic APAP dosing, males were less likely to develop APAP protein adducts. Further research into lipid biosynthesis and metabolism may provide further insight into physiology associated with adduct production.

Host methylation predicts SARS-CoV-2 infection and clinical outcome.

BACKGROUND: Since the onset of the SARS-CoV-2 pandemic, most clinical testing has focused on RT-PCR1. Host epigenome manipulation post coronavirus infection2-4 suggests that DNA methylation signatures may differentiate patients with SARS-CoV-2 infection from uninfected individuals, and help predict COVID-19 disease severity, even at initial presentation. METHODS: We customized Illumina's Infinium MethylationEPIC array to enhance immune response detection and profiled peripheral blood samples from 164 COVID-19 patients with longitudinal measurements of disease severity and 296 patient controls. RESULTS: Epigenome-wide association analysis revealed 13,033 genome-wide significant methylation sites for case-vs-control status. Genes and pathways involved in interferon signaling and viral response were significantly enriched among differentially methylated sites. We observe highly significant associations at genes previously reported in genetic association studies (e.g. IRF7, OAS1). Using machine learning techniques, models built using sparse regression yielded highly predictive findings: cross-validated best fit AUC was 93.6% for case-vs-control status, and 79.1%, 80.8%, and 84.4% for hospitalization, ICU admission, and progression to death, respectively. CONCLUSIONS: In summary, the strong COVID-19-specific epigenetic signature in peripheral blood driven by key immune-related pathways related to infection status, disease severity, and clinical deterioration provides insights useful for diagnosis and prognosis of patients with viral infections.

Host methylation predicts SARS-CoV-2 infection and clinical outcome.

BACKGROUND: Since the onset of the SARS-CoV-2 pandemic, most clinical testing has focused on RT-PCR1. Host epigenome manipulation post coronavirus infection2-4 suggests that DNA methylation signatures may differentiate patients with SARS-CoV-2 infection from uninfected individuals, and help predict COVID-19 disease severity, even at initial presentation. METHODS: We customized Illumina's Infinium MethylationEPIC array to enhance immune response detection and profiled peripheral blood samples from 164 COVID-19 patients with longitudinal measurements of disease severity and 296 patient controls. RESULTS: Epigenome-wide association analysis revealed 13,033 genome-wide significant methylation sites for case-vs-control status. Genes and pathways involved in interferon signaling and viral response were significantly enriched among differentially methylated sites. We observe highly significant associations at genes previously reported in genetic association studies (e.g. IRF7, OAS1). Using machine learning techniques, models built using sparse regression yielded highly predictive findings: cross-validated best fit AUC was 93.6% for case-vs-control status, and 79.1%, 80.8%, and 84.4% for hospitalization, ICU admission, and progression to death, respectively. CONCLUSIONS: In summary, the strong COVID-19-specific epigenetic signature in peripheral blood driven by key immune-related pathways related to infection status, disease severity, and clinical deterioration provides insights useful for diagnosis and prognosis of patients with viral infections.

Viral infections affect the body in many ways, including via changes to the epigenome, the sum of chemical modifications to an individual's collection of genes that affect gene activity. Here, we analyzed the epigenome in blood samples from people with and without COVID-19 to determine whether we could find changes consistent with SARS-CoV-2 infection. Using a combination of statistical and machine learning techniques, we identify markers of SARS-CoV-2 infection as well as of severity and progression of COVID-19 disease. These signals of disease progression were present from the initial blood draw when first walking into the hospital. Together, these approaches demonstrate the potential of measuring the epigenome for monitoring SARS-CoV-2 status and severity.