EGFR core fucosylation, induced by enveloped viruses, promotes TRIM40-mediated-RIG-I ubiquitination and suppresses interferon-I antiviral defenses (preprint)

Aberrant N-glycosylation has been implicated in viral diseases. Alpha-(1,6)-fucosyltransferase (FUT8) is the sole enzyme responsible for core fucosylation of N-glycans during glycoprotein biosynthesis. Here we found that multiple viral envelope proteins (including HCV-E2, VSV-G, SARS-COV-2-Spike and HIV-gp120) enhanced FUT8 expression and core fucosylation. HCV-E2 manipulates host transcription factor SNAIL to induce FUT8 expression through EGFR-AKT-SNAIL activation and SNAIL nuclear translocation. These aberrant increased-FUT8 expression promotes TRIM40-mediated RIG-I K48-ubiquitination and suppresses the antiviral interferon (IFN)-I response through core fucosylated EGFR-JAK1-STAT3-RIG-I signaling. FUT8 inhibitor 2FF, N-glycosylation site-specific mutation (Q352AT) of EGFR, and tissue-targeted Fut8 silencing significantly increased antiviral IFN-I responses and suppressed RNA viral replication, suggesting that core fucosylation mediated by FUT8 is critical for antiviral innate immunity. These findings reveal an immune invasion mechanism in which virus-induced FUT8 suppresses endogenous RIG-I-mediated antiviral defenses by enhancing core fucosylated EGFR-mediated activation. FUT8 may be a promising target for RNA viral therapies.

Online Education Isn’t the Best Choice: Evidence-Based Medical Education in the Post-epidemic Era—A Cross-Sectional Study (preprint)

Background The COVID-19 pandemic led many educational institutions to shift to online courses, making blended education a significant trend in teaching. We examined the effectiveness of blended learning in an evidence-based medicine course.Methods We compared the examination scores of a blended learning group, an online only group, and a traditional offline group and conducted a questionnaire survey on students’ preferences for different learning modes and the reasons for their preferences. A total of 2100 undergraduate students in clinical medicine were included in this cross-sectional study. Examination results were collected, and questionnaires were administered to the study participants. We compared the mean theoretical scores and exam pass rates of the three teaching groups using ANOVA and c2test for multiple comparisons.Results The blended group’s theoretical scores and pass rate were significantly higher than those of the offline and online groups. Furthermore, 71.6% preferred the blended teaching mode. Most students believed that blended teaching was the most effective mode—offline education: 7.86%; online education: 26.14%; blended education: 66%. Subsequently, in a questionnaire administered to a blended group of students, their foremost reason for liking online instruction was ‘flexible in time and space’ (99%), followed by ‘can be viewed repeatedly, facilitating a better understanding of knowledge points’ (98%). Their foremost reason for liking offline teaching was ‘helps to create a good learning atmosphere’ (97%), followed by ‘teachers can control students’ learning status in real time’ (89%).Conclusions This study explored the effectiveness of learning in evidence-based medicine courses by comparing the learning outcomes and personal perceptions of three different teaching modes. This is the first cross-sectional study in which three different teaching models are compared and discussed in an evidence-based medicine course. We also elaborate on the specific instructional protocols for each model. This study shows that using a blended education approach in evidence-based medicine courses can improve students’ learning motivation, autonomy, and satisfaction. It also enhances instructional efficiency, thereby improving students’ understanding of the course content.

Protective effect of plasma neutralization from prior SARS-CoV-2 Omicron infection against BA.5 subvariant symptomatic reinfection (preprint)

From December 2022 to January 2023, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections caused by BA.5 and BF.7 subvariants of B.1.1.529 (Omicron) swept across mainland China. It is crucial to estimate the protective effect of the neutralizing antibodies generated by such mass infections against the next potential SARS-CoV-2 reinfection wave, especially if driven by CH.1.1 or XBB.1.5. Previously, we recruited and continuously followed a cohort of individuals that experienced Omicron BA.1, BA.2, and BA.5 breakthrough infections, as well as a control cohort with no history of SARS-CoV-2 infection. In the previously uninfected cohort, the total symptomatic infection rate surveyed during the outbreak was 91.6%, while the symptomatic reinfection rate was 32.9%, 10.5%, and 2.8% among individuals with prior Omicron BA.1, BA.2 and BA.5 infection, respectively, with median intervals between infections of 335, 225 and 94 days. Pseudovirus neutralization assays were performed in plasma samples collected from previously Omicron BA.1-infected individuals approximately 3 months before the outbreak. Results indicate a robust correlation between the plasma neutralizing antibody titers and the protective effect against symptomatic reinfection. The geometric mean of the 50% neutralizing titers (NT50) against D614G, BA.5, and BF.7 were 2.0, 2.5, and 2.3-fold higher in individuals without symptomatic reinfection than in those with symptomatic reinfection (p < 0.01). Low plasma neutralizing antibody titer (below the geometric mean of NT50) was associated with an enhanced cumulative risk of symptomatic reinfection, with a hazard ratio (HR) of 23.55 (95% CI: 9.23-60.06) against BF.7 subvariant. Importantly, neutralizing antibodies titers post one month after BF.7/BA.5 breakthrough infections against CH.1.1 and XBB.1.5 are similar to that against BF.7 from individuals with prior BA.1 infection while not experiencing a symptomatic BF.7/BA.5 reinfection (plasma collected 3 months before the outbreak), suggesting that the humoral immunity generated by the current BF.7/BA.5 breakthrough infection may provide protection against CH.1.1 and XBB.1.5 symptomatic reinfection wave for 4 months. Of note, the higher hACE2 binding of XBB.1.5 may reduce the protection period since the potential increase of infectivity.

Nirmatrelvir and the Risk of Post-Acute Sequelae of COVID-19 (preprint)

Long Covid -- the disease encompassing the post-acute sequelae of SARS-CoV-2 (PASC) -- affects millions of people around the world. Prevention of PASC is an urgent public health priority. In this work, we aimed to examine whether treatment with nirmatrelvir in the acute phase of COVID-19 is associated with reduced risk of post-acute sequelae. We used the healthcare databases of the US Department of Veterans Affairs to identify users of the health system who had a SARS-CoV-2 positive test between March 01, 2022 and June 30, 2022, were not hospitalized on the day of the positive test, had at least 1 risk factor for progression to severe COVID-19 illness and survived the first 30 days after SARS-CoV-2 diagnosis. We identify those who were treated with oral nirmatrelvir within 5 days after the positive test (n=9217) and those who received no COVID-19 antiviral or antibody treatment during the acute phase of SARS-CoV-2 infection (control group, n= 47,123). Inverse probability weighted survival models were used to estimate the effect of nirmatrelvir (versus control) on a prespecified panel of 12 post-acute COVID-19 outcomes and reported as hazard ratio (HR) and absolute risk reduction (ARR) in percentage at 90 days. Compared to the control group, treatment with nirmatrelvir was associated with reduced risk of PASC (HR 0.74 95% CI (0.69, 0.81), ARR 2.32 (1.73, 2.91)) including reduced risk of 10 of 12 post-acute sequelae in the cardiovascular system (dysrhythmia and ischemic heart disease), coagulation and hematologic disorders (deep vein thrombosis, and pulmonary embolism), fatigue, liver disease, acute kidney disease, muscle pain, neurocognitive impairment, and shortness of breath. Nirmatrelvir was also associated with reduced risk of post-acute death (HR 0.52 (0.35, 0.77), ARR 0.28 (0.14, 0.41)), and post-acute hospitalization (HR 0.70 (0.61, 0.80), ARR 1.09 (0.72, 1.46)). Nirmatrelvir was associated with reduced risk of PASC in people who were unvaccinated, vaccinated, and boosted, and in people with primary SARS-CoV-2 infection and reinfection. In sum, our results show that in people with SARS-CoV-2 infection who had at least 1 risk factor for progression to severe COVID-19 illness, treatment with nirmatrelvir within 5 days of a positive SARS-CoV-2 test was associated with reduced risk of PASC regardless of vaccination status and history of prior infection. The totality of findings suggests that treatment with nirmatrelvir during the acute phase of COVID-19 reduces the risk of post-acute adverse health outcomes.

Outcomes of SARS-CoV-2 Reinfection (preprint)

First infection with SARS-CoV-2 is associated with increased risk of acute and post-acute death and sequelae in the pulmonary and extrapulmonary organ systems. However, whether reinfection adds to the risk incurred after the first infection is not clear. Here we use the national health care databases of the US Department of Veterans Affairs to build a cohort of people with first infection (n = 257,427), reinfection (2 or more infections, n = 38,926), and a non-infected control group (n = 5,396,855) to estimate risks and 6-month burdens of all-cause mortality, hospitalization, and a set of pre-specified incident outcomes. We show that compared to people with first infection, reinfection contributes additional risks of all-cause mortality, hospitalization, and adverse health outcomes in the pulmonary and several extrapulmonary organ systems (cardiovascular disorders, coagulation and hematologic disorders, diabetes, fatigue, gastrointestinal disorders, kidney disorders, mental health disorders, musculoskeletal disorders, and neurologic disorders); the risks were evident in those who were unvaccinated, had 1 shot, or 2 or more shots prior to the second infection; the risks were most pronounced in the acute phase, but persisted in the post-acute phase of reinfection, and most were still evident at 6 months after reinfection. Compared to non-infected controls, assessment of the cumulative risks of repeated infection showed that the risk and burden increased in a graded fashion according to the number of infections. The constellation of findings show that reinfection adds non-trivial risks of all-cause mortality, hospitalization, and adverse health outcomes in the acute and post-acute phase of the reinfection. Reducing overall burden of death and disease due to SARS-CoV-2 will require strategies for reinfection prevention.

Disease profile and plasma neutralizing activity of post-vaccination Omicron BA.1 infection in Tianjin, China: a retrospective study (preprint)

BackgroundSARS-CoV-2 Omicron variant BA.1 first emerged on the Chinese mainland in January 2022 in Tianjin and caused a large wave of infections. During mass PCR testing, a total of 430 cases infected with Omicron were recorded between January 8 and February 7, 2022, with no new infections detected for the following 16 days. Most patients had been vaccinated with SARS-CoV-2 inactivated vaccines. The disease profile associated with BA.1 infection, especially after vaccination with inactivated vaccines, is unclear. Whether BA.1 breakthrough infection after receiving inactivated vaccine could create a strong enough humoral immunity barrier against Omicron is not yet investigated. MethodsWe collected the clinical information and vaccination history of the 430 COVID-19 patients infected with Omicron BA.1. Re-positive cases and inflammation markers were monitored during the patients convalescence phase. Ordered multiclass logistic regression model was used to identify risk factors for COVID-19 disease severity. Authentic virus neutralization assays against SARS-CoV-2 wildtype, Beta and Omicron BA.1 were conducted to examine the plasma neutralizing titers induced after post-vaccination Omicron BA.1 infection, and were compared to a group of uninfected healthy individuals who were selected to have a matched vaccination profile. FindingsAmong the 430 patients, 316 (73.5%) were adults with a median age of 47 years, and 114 (26.5%) were under-age with a median age of 10 years. Female and male patients account for 55.6% and 44.4%, respectively. Most of the patients presented with mild (47.7%) to moderate diseases (50.2%), with only 2 severe cases (0.5%) and 7 (1.6%) asymptomatic infections. No death was recorded. 341 (79.3%) of the 430 patients received inactivated vaccines (54.3% BBIBP-CorV vs. 45.5% CoronaVac), 49 (11.4%) received adenovirus-vectored vaccines (Ad5-nCoV), 2 (0.5%) received recombinant protein subunit vaccines (ZF2001), and 38 (8.8%) received no vaccination. No vaccination is associated with a substantially higher ICU admission rate among Omicron BA.1 infected patients (2.0% for vaccinated patients vs. 23.7% for unvaccinated patients, P<0.001). Compared with adults, child patients presented with less severe illness (82.5% mild cases for children vs. 35.1% for adults, P<0.001), no ICU admission, fewer comorbidities (3.5% vs. 53.2%, P<0.001), and less chance of turning re-positive on nucleic acid tests (12.3% vs. 22.5%, P=0.019). For adult patients, compared with no prior vaccination, receiving 3 doses of inactivated vaccine was associated with significantly lower risk of severe disease (OR 0.227 [0.065-0.787], P=0.020), less ICU admission (OR 0.023 [0.002-0.214], P=0.001), lower re-positive rate on PCR (OR 0.240 [0.098-0.587], P=0.002), and shorter duration of hospitalization and recovery (OR 0.233 [0.091-0.596], P=0.002). At the beginning of the convalescence phase, patients who had received 3 doses of inactivated vaccine had substantially lower systemic immune-inflammation index (SII) and C-reactive protein than unvaccinated patients, while CD4+/CD8+ ratio, activated Treg cells and Th1/Th2 ratio were higher compared to their 2-dose counterparts, suggesting that receipt of 3 doses of inactivated vaccine could step up inflammation resolution after infection. Plasma neutralization titers against Omicron, Beta, and wildtype significantly increased after breakthrough infection with Omicron. Moderate symptoms were associated with higher plasma neutralization titers than mild symptoms. However, vaccination profiles prior to infection, whether 2 doses versus 3 doses or types of vaccines, had no significant effect on post-infection neutralization titer. Among recipients of 3 doses of CoronaVac, infection with Omicron BA.1 largely increased neutralization titers against Omicron BA.1 (8.7x), Beta (4.5x), and wildtype (2.2x), compared with uninfected healthy individuals who have a matched vaccination profile. InterpretationReceipt of 3-dose inactivated vaccines can substantially reduce the disease severity of Omicron BA.1 infection, with most vaccinated patients presenting with mild to moderate illness. Child patients present with less severe disease than adult patients after infection. Omicron BA.1 convalescents who had received inactivated vaccines showed significantly increased plasma neutralizing antibody titers against Omicron BA.1, Beta, and wildtype SARS-CoV-2 compared with vaccinated healthy individuals. FundingThis research is supported by Changping Laboratory (CPL-1233) and the Emergency Key Program of Guangzhou Laboratory (EKPG21-30-3), sponsored by the Ministry of Science and Technology of the Peoples Republic of China. Research in contextO_ST_ABSEvidence before this studyC_ST_ABSPrevious studies (many of which have not been peer-reviewed) have reported inconsistent findings regarding the effect of inactivated vaccines against the Omicron variant. On Mar 6, 2022, we searched PubMed with the query "(SARS-CoV-2) AND ((Neutralisation) OR (Neutralisation)) AND ((Omicron) OR (BA.1)) AND (inactivated vaccine)", without date or language restrictions. This search identified 18 articles, of which 13 were directly relevant. Notably, the participants in many of these studies have received only one or two doses of inactivated vaccine with heterologous booster vaccination; other studies have a limited number of participants receiving inactivated vaccines. Added value of this studyTo date, this is the first study to report on the protective effect of inactivated vaccines against the severe disease caused by the Omicron variant. We examine and compare the disease profile of adults and children. Furthermore, we estimate the effect of post-vaccination omicron infection on plasma neutralization titers against Omicron and other SARS-COV-2 variants. Specifically, the disease profile of Omicron convalescents who had received two-dose primary series of inactivated vaccines with or without a booster dose prior to infection is compared with unvaccinated patients. We also analyzed the effect of infection on neutralizing activity by comparing vaccinated convalescents with vaccinated healthy individuals with matched vaccination profiles. Implications of all the available evidenceCompared with adults, child patients infected with Omicron tend to present with less severe disease and are less likely to turn re-positive on nucleic acid tests. Receipt of two-dose primary series or three doses of inactivated vaccine is a protective factor against severe disease, ICU admission, re-positive PCR and longer hospitalization. The protection afforded by a booster dose is stronger than two-dose primary series alone. Besides vaccination, infection with Omicron is also a key factor for elevated neutralizing antibody titers, enabling cross-neutralization against Omicron, wildtype (WT) and the Beta variant.

Long-term Neurologic Outcomes of COVID-19 (preprint)

The neurologic manifestations during the acute phase of COVID-19 are well characterized, but a comprehensive evaluation of the risks and burdens of post-acute neurologic disorders at 1-year after the initial infection has not been yet undertaken. Here we use national healthcare databases from the US Department of Veterans Affairs to build a cohort of 154,068 individuals with COVID-19 as well as two sets of control cohorts with 5,638,795 (contemporary controls) and 5,859,621 (historical controls) individuals to estimate risks and burdens of a set of pre-specified incident neurologic disorders at 12 months. Our results show that in the post-acute phase of COVID-19, there was increased risk of an array of neurologic disorders including stroke, cognition and memory disorders, peripheral nervous system disorders, episodic disorders (e.g., migraine and seizure), extrapyramidal and movement disorders, musculoskeletal disorders, sensory disorders, and other disorders including Guillain-Barré, and encephalitis or encephalopathy. The risks and burdens were significantly elevated even in people who did not require hospitalization during the acute phase of COVID-19. Taken together, our results provide evidence of increased risk of broad array of long-term neurologic disorders in people with COVID-19. This evidence will inform the management of the ongoing pandemic and guide post-acute COVID-19 clinical care and health care systems planning.

Long Covid after Breakthrough COVID-19: the post-acute sequelae of breakthrough COVID-19 (preprint)

The post-acute sequelae of COVID-19 have been described 1 , but whether breakthrough COVID-19 (that is the disease that ensues following vaccine breakthrough SARS-CoV-2 infection) results in post-acute sequelae is not yet clear. Here we use the national healthcare databases of the US Department of Veterans Affairs to characterize 6-month risks of incident post-acute sequelae in people with breakthrough COVID-19 who survived for at least 30 days after diagnosis. We show that compared to people with no evidence of COVID-19, beyond the first 30 days of illness, people with breakthrough COVID-19 exhibit a higher risk of death and broad array of incident post-acute sequelae in the pulmonary system, as well as extrapulmonary sequelae that include cardiovascular disorders, coagulation disorders, gastrointestinal disorders, general disorders (e.g., fatigue), kidney disorders, mental health disorders, metabolic disorders, musculoskeletal disorders, and neurologic disorders. Our analyses by care setting of the acute phase of the disease show that people who were not hospitalized during the first 30 days after diagnosis with breakthrough COVID-19 exhibit a small but not insignificant increase in risk of death and post-acute sequelae; the risks are further increased in people who were hospitalized during the acute phase of the disease. Our comparative approach shows that people with breakthrough COVID-19 exhibit lower risks of death and post-acute sequelae than people with COVID-19 who were not previously vaccinated for it; and in analyses among individuals who were hospitalized during the acute phase of the disease, people with breakthrough COVID-19 exhibit higher risks of death and post-acute sequelae than people with seasonal influenza. Altogether, our findings show increased risks of death and post-acute sequalae in people with breakthrough COVID-19; the risks are evident among those who were not hospitalized during the acute phase of the disease. Our comparative approach provides context for understanding the risks in relation to COVID-19 without prior vaccination and seasonal influenza. The findings will inform the ongoing effort to optimize strategies for prevention of breakthrough SARS-CoV-2 infections and will guide development and optimization of post-acute care pathways for people with breakthrough COVID-19.

One-year Risks and Burdens of Incident Cardiovascular Disease in COVID-19: Cardiovascular Manifestations of Long COVID (preprint)

The cardiovascular complications of acute COVID-19 are well described; however, a comprehensive characterization of the post-acute cardiovascular manifestations of COVID-19 at one year has not been undertaken. Here we use the US Department of Veterans Affairs national healthcare databases to build a cohort of 151,195 people with COVID-19, 3,670,087 contemporary and 3,656,337 historical controls to estimate risks and 1-year burdens of a set of pre-specified incident cardiovascular outcomes. We show that beyond the first 30 days of infection, people with COVID-19 are at increased risk of incident cardiovascular disease spanning several categories including cerebrovascular disorders, dysrhythmias, ischemic and non-ischemic heart disease, pericarditis, myocarditis, heart failure, and thromboembolic disease. The risks and burdens were evident among those who were non-hospitalized during the acute phase of the infection and increased in a graded fashion according to care setting of the acute infection (non-hospitalized, hospitalized, and admitted to intensive care). Taken together, our results provide evidence that risk and 1-year burden of cardiovascular disease in survivors of acute COVID-19 are substantial. Care pathways of people who survived the acute episode of COVID-19 should include attention to cardiovascular health and disease.

Burdens of Post-acute Sequelae of COVID-19 by Age, Race, Sex, and Health Status (preprint)

The Post-Acute Sequelae of SARS-CoV-2 infection (PASC) have been characterized; however, the burden of PASC remains unknown. And whether the burden of individual sequela varies in different population groups is also not clear. Here we estimate that PASC — defined as the presence of at least one sequela in excess of non-infected controls — was 73.43 (72.10, 74.72) per 1000 persons at 6 months. The burden of PASC was 44.51 (43.09, 45.85), 217.08 (212.43, 222.23), and 360.16 (350.53, 369.38) among non-hospitalized, hospitalized, and those who required intensive care during the first 30-days of infection. Burdens of some sequelae were more pronounced in younger individuals, and some were more pronounced in older adults; the same picture was evident in analyses across race, and sex groups. The burden of individual sequela was consistently higher in people with poorer baseline health and increased in a graded fashion according to care setting of the acute infection. In sum, the burden of PASC is substantial; however, PASC is non-monolithic with sequelae that are differentially expressed in various population groups. Collectively, our results may be useful in informing health systems capacity planning and care strategies of people with PASC.

High Dimensional Characterization of Post-acute Sequalae of COVID-19: analysis of health outcomes and clinical manifestations at 6 months (preprint)

The coronavirus disease 2019 (COVID-19) is a viral illness caused by the severe acute respiratory syndrome coronavirus 2. The acute clinical manifestations of COVID-19 are well characterized. The post-acute sequalae of COVID-19 have not been comprehensively described. Here, we use the national healthcare databases of the US Department of Veterans Affairs to undertake a high dimensional approach to comprehensively identify 6-months outcomes of incident clinical manifestations including diagnoses, medication use, and laboratory abnormalities in people who survived the first 30 days of COVID-19. We first describe the clinical manifestations in people with COVID-19 compared to users of the Veterans Affairs healthcare system. We then provide a comparative evaluation of the post-acute sequalae in 30-day survivors who were hospitalized for COVID-19 vs. seasonal influenza. We show that beyond the first 30 days of illness, people with COVID-19 are at higher risk of death and health resource utilization. Our approach identifies incident clinical manifestations in the respiratory system and several other manifestations including the nervous system and neurocognitive disorders, mental health disorders, metabolic disorders, cardiovascular disorders, gastrointestinal disorders, and signs and symptoms related to poor generalized wellbeing including malaise, fatigue, musculoskeletal pain, and anemia. There was increased incident use of pain medications (opioids and non-opioids), antidepressants, anxiolytics, antihypertensives, antihyperlipidemic, insulin, and several other medication classes. The findings show that beyond the first 30 days of illness, substantial burden of health loss — spanning pulmonary and several extrapulmonary organ systems — is experienced by COVID-19 survivors. The results provide a roadmap to inform health system planning and development of multidisciplinary care strategies aimed at reducing chronic health loss and optimizing wellness among COVID-19 survivors.

The MERS-CoV Receptor DPP4 as a Candidate Binding Target of the SARS-CoV-2 Spike (preprint)

The ongoing outbreak of the novel coronavirus pneumonia COVID-19 has caused great number of cases and deaths, but our understanding about the pathogen SARS-CoV-2 remains largely unclear. The attachment of the virus with the cell-surface receptor and a co-factor is the firstly important step for the infection. Here, bioinformatics approaches combining human-virus protein interaction prediction and protein docking based on crystal structures have revealed the high affinity between human dipeptidyl peptidase 4 (DPP4) and the spike (S) receptor-binding domain of SARS-CoV-2. Intriguingly, the crucial binding residues of DPP4 at the interface are identical to those as bound to the MERS-CoV spike. Moreover, E484 insertion and adjacent substitutions should be most essential for this DPP4-binding ability acquirement of SARS-CoV-2-S compared with that of SARS-CoV-S, which does not interact with DPP4. This potential utilization of DPP4 as a coreceptor or binding target for SARS-CoV-2 may offer novel insight into the viral pathogenesis, and help the surveillance and therapeutics strategy for meeting the challenge of COVID-19.Funding: This work was supported by the National Key Research and Development Program and the National Natural Science Foundations of China (2017YFC1200204, 2017YFC1200205, 31670171 and 81974427).Declaration of Interest: The authors declare no conflict of interest.