Introduction
After its emergence in China, the COVID‐19 pandemic has spread rapidly around the world, affecting all of our lives. Over the past few months, different vaccines have been issued to the ...public following FDA approval. Since their release, the number of COVID‐19 cases in the United States have gone down significantly. However, there is a long way to go until the eradication of the virus. There has also been a strong emergence of delta variants across the world, which (for the most part) can still be partially blocked by vaccines. This is why it is so important everyone gets a vaccine for COVID‐19.
Purpose
The purpose of this project was to determine the public’s opinion of COVID‐19 and the COVID‐19 vaccines, while also determining correlation between different demographics and these opinions.
Hypothesis
There are many different factors within various demographics that cause a correlation to a certain belief regarding COVID‐19 and COVID‐19 vaccines.
Materials and Methods
This study was conducted in collaboration with Loyola University Chicago. Survey questions about COVID‐19 and COVID‐19 vaccinations have been sent electronically to the respondents. Initial results based on a pilot study where the answers from 250 respondents were compiled. Basic descriptive statistics were used to characterize the sample and study variables. Univariate analysis of associations between categorical variables and vaccination intent and acceptance were assessed by chi‐square statistics. Multivariate analysis performed by regression analysis to determine significant independent predictors. A p‐value of less than 0.05 was considered to indicate statistical significance; all tests were 2‐tailed.
Results
In our study population vaccine willingness have been found as 94,4%, the rate of receiving at least one dose of vaccination have been found as 90,8%. In univariate analysis, gender, believing that vaccinations protect others and having enough information about safety and efficacy of vaccines have been found significant factors for vaccine willingness. For receiving at least one dose of vaccination, gender, previous COVID‐19 infection, believing that vaccinations protect others, having enough information about safety and efficacy of vaccines, the degree of COVID‐19 related effects on person’s life, knowledge that COVID‐19 can cause blood clots and the degree of concerns about delta variant have been found significant factors for vaccine willingness. In multivariate analysis only believing that vaccinations protect others have been found as an independent factor for vaccine willingness. For receiving at least one dose of vaccination, believing that vaccinations protect others, the degree of COVID‐19 related effects on person’s life and having enough information about safety and efficacy of vaccines have been found independent factors in multivariate analysis.
Conclusion
There are several vaccines currently available in the United States. All of these vaccines provide immunity against COVID‐19 and its variants which have been identified in 2020 and 2021. The level of immunity and the reported adverse events vary with each of the vaccines. Despite minor issues the vaccination outweighs the benefits and provides a clear approach to control this pandemic.
Scientists continue to study SARS‐CoV‐2, the virus currently responsible for a worldwide pandemic, to determine the best methods to inactivate the virus on surfaces and reduce the risk of contact ...transmission. Its classification as a Biosafety Level (BSL) ‐3 pathogen complicates research since few facilities have the capacity for this level of containment. Human betacoronavirus OC43 (OC43) is a good surrogate for SARS‐CoV‐2 based on its genetic similarity and OC43’s BSL‐2 classification. Two industry specific surfaces were inoculated with OC43 and sampled at timepoints that reflect typical industry timelines for sanitation. The objective was to determine the stability of OC43 on typical meat facility surfaces.
OC43 was propagated on human ileocecal colorectal adenocarcinoma cells (HRT‐18, ATCC CCL‐244). Infectious virus was quantified by indirect immunofluorescence TCID50 assays on HRT‐18 cells. Stainless steel grade 316 and polyethylene (poly top) coupons were inoculated with 100 ul of 2.51 x 106 TCID50/ml virus by pipette in a grid pattern at room temperature (20ºC) in an active biosafety cabinet in triplicate for each timepoint. Coupons were rinsed with 5 ml of sterile filtered beef extract buffer (BEB7), composed of Tris base, glycine, magnesium chloride, and beef extract pH adjusted to 7.0, at 0, 1, 4, 8, and 16 hours post inoculation (hpi). The collected rinsate was aliquoted and stored at ‐80ºC until the TCID50 assays were conducted in batch. The 16 hpi stainless steel samples were created at the time of the poly top experiment. The TCID50 assays were run in duplicate for each sample and TCID50/ml was calculated and averaged for each replicate. Percent recovery of infectious virus was determined for each timepoint.
Virus was observed to have partially dried at 1 hpi and was completely dry at all following timepoints on both surfaces. Few infectious virions were lost between inoculation and 0 hpi collection from both surfaces. Mean percent recovery (MPR) of infectious OC43 from stainless steel and poly top at 0 hpi was 153% and 90%, respectively. At 1 hpi, the MPR from stainless steel was 65% while poly top was 123%. The variability of the assay explains these findings. Stainless steel and poly top had average recoveries of 60% and 25% at 4 hours, respectively. The lowest MPR for stainless steel was 17% at 8 hpi while the lowest MPR for poly top was 7% at 16 hpi. A higher MPR of 43% from stainless steel at 16 hpi reflects this timepoint being conducted on a different day.
Infectious OC43 is still detectable on common meat facility surfaces several hours after inoculation, but our current knowledge of enveloped viruses asserts that they are inviable after drying. A maximum decrease of only 1 log TCID50 between inoculation and the 16 hpi samples means that a person could potentially be infected between cleanings if it is possible to transfer dried infectious virus from a surface to a mucous membrane. However, this is a conjecture that requires further study since the infectious dose of SARS‐CoV‐2 is unknown and no experiment to determine if this contact transfer is possible has been conducted. Currently, similar experiments with SARS are underway.
The adaptive immune response is under circadian control, yet, the benefit of this rhythmicity for the organism is unknown. Furthermore, it is not understood why adaptive immune reactions continue to ...exhibit circadian changes over long periods of time. Using a combination of experimental and mathematical modelling approaches, we show here that dendritic cells (DCs) migrate from the skin to the draining lymph node (LN) in a time‐of‐day‐dependent manner, which provides an enhanced likelihood for functional interactions with T cells. Greater numbers of infiltrating DCs induce rhythmic expression of TNF‐α in the draining LN, which enhances ICAM‐1 expression in high endothelial venules (HEVs), resulting in lymphocyte infiltration and LN expansion. Icam1 is controlled by binding of the core circadian clock transcription factor BMAL1 to its promoter region, and rhythmic Icam1 expression is lost in mice lacking endothelial cell BMAL1. LN cellularity continues to be different for weeks after the initial time‐of‐day‐dependent challenge, which determines the immune response to vaccinations directed against Hepatitis A virus as well as SARS‐CoV‐2, as evidenced by rhythmic T cell reactions, germinal center formation and antibody production. Our results provide the mechanistic understanding of the time‐of‐day dependent development and maintenance of an adaptive immune response, demonstrating its dependency upon the timing of the initial challenge and the interactions of rhythmicity in multiple parameters. This provides a strategy for using time‐of‐day to optimize vaccination regimes.
Antibodies play an important part in combating SARS‐CoV‐2 infection whether generated by the infection or vaccination. However, the many epitopes generated by infection have not been fully ...investigated with only a few epitopes known and these being mostly limited to the S protein’s receptor binding domain (RBD) and the N‐terminal domain which limits vaccine and drug design 1‐4. The difference between epitopes generated by infection and vaccination has also not been studied. To address this, we employed a SARS‐CoV‐2 proteome microarray to screen for linear epitopes recognized by antibodies present in COVID‐19 patients and individuals vaccinated with the Pfizer‐BioNTech mRNA COVID‐19 Vaccine.
The proteome microarray consisted of S, N, and E proteins, as well as spotting peptides that were 15 amino acids in length with overlaps of 5‐amino acids, covering the entire SARS‐CoV‐2 proteome (MN908947.3) (Figure 1). Blood samples were incubated onto the arrays followed by an incubation of fluorescent secondary anti‐human antibodies. Fluorescent intensity data generated and normalized using the Z‐score method and then further analyzed for significance by parametric one‐way ANOVA with Dunnett's post hoc test (COVID‐19 cohort) and repeated measure ANOVAs with Dunnett's post hoc tests (vaccinated cohort).
The full‐length S protein showed a significant increase in COVID‐19 patients at around 20‐23 days after symptom onset and vaccinated individuals over all time points in both IgM and IgG antibodies (Figure 2A). Linear mapping of the IgM epitopes revealed a degree of overlap between infected and vaccinated individuals (22.2%; 6/27 total) with both having epitopes in the RBD and fusion peptide (FP) (Figure 2B). Structural mapping on 3D models of the S protein showed that all epitopes where on the surface of the protein and that COVID‐19 generated epitopes have a different pattern than those generated by vaccination (Figure 2C).
An Epitope identified in this study with future prospects is epitope S481‐495 from COIVD‐19 patients that partially overlapped the binding site of two neutralizing antibodies previously isolated from COVID‐19 patients, S2H135 and F2B‐2F61, and contacted amino acids that interact with ACE2 receptor6,7. One epitope of note from the vaccinated individuals is epitope S811‐825 which mapped adjacent to the fusion‐peptide proximal region. These epitopes may be helpful in future vaccine and antibody therapy development.
1 Ju, B. et al. Nature 584, 115‐119.
2 Robbiani, D. F. et al. Nature 584, 437‐442.
3 Seydoux, E. et al. bioRxiv.
4 Wu, Y. et al. Science 368, 1274‐1278.
5 Piccoli, L. et al. Cell 183, 1024‐1042 e1021.
6 Casalino, L. et al. ACS Cent Sci 6, 1722‐1734.
7 Wang, Q. et al. Cell 181, 894‐904 e899.
Endothelial cells (ECs) are in the inner layer of blood vessels, and it controls the transportation of nutrition and essential molecules from the extracellular matrix to the intracellular space ...making a homeostasis environment. Vascular endothelium can be disrupted with viruses, which are among the main reasons for epidemic and pandemic outbreaks, and it leads to health issues such as cardiovascular‐related diseases. Wide range of viruses can potentially target the endothelium including Dengue virus estimated to infect 390 million people per year. Zika virus leads to pregnancy complications such as preterm birth and miscarriage. Tick‐borne encephalic damaged the central nervous system and, Hantavirus has a mortality rate of 38% through hantavirus pulmonary syndrome. Reviewing these several diseases, including SARS‐CoV‐2, has given an in‐depth look at what the endothelial cells are going through when these viruses are infecting the host bodies. Following the virus infection, ECs are going through activation mode, which is accompanied with glycocalyx degradation, plasma leakage, phosphorylation of P120, and cadherin, increased proinflammatory responses and, disruption of barrier integrity.
The SARS‐CoV‐2 pandemic has taken a global toll; With a total of 362 million cases, there are 5.63 million deaths occurring. With the usage of immunofluorescence and vascular permeability assay, we monitor how Human Umbilical Vascular Endothelial Cells (HUVEC) and Human Lung Microvascular Endothelial Cells (HLMVEC) are impacted when treated with UV‐inactivated & heat‐inactivated SARS‐CoV‐2, and tumor necrosis factor (TNF)‐α. TNF‐α is an inflammatory cytokine prominent during inflammation and its known to increase the permeability of endothelium. Like the TNF‐α, both cells treated with UV‐inactivated and heat‐inactivated SARS‐CoV2 demonstrated boosted permeability. With the considerable effect of SARS‐CoV2 on the permeability of ECs, it is essential to understand the mechanisms and pathways behind the boosted leak, including the impact of the virus on the expression of adhesion molecules like platelet endothelial cell adhesion molecule (PECAM‐1). One can see in the images of HUVEC and HLMVEC that SARS‐CoV‐2 diminish the localization of PECAM‐1. On a non‐treated negative control, cells are spaced evenly and are close to another since their junctions are not targeted. Once the cells are treated with either heat‐inactivated or UV‐inactivated SARS‐CoV‐2, PECAM‐1 localization is disrupted, which can be seen from the space between cells since the PECAM‐1 has been altered and is no longer holding the cells together. From those exact images, one can detect the enlarged and irregular shapes of nucleuses and change in morphology of these cells since the SARS‐CoV‐2 is not only altering the PECAM‐1 complex but it can also modify various proteins and mRNAs. By combining information of various viruses and knowledge from our studies, we can understand the mechanisms of ECs and take measurements to protect ECs from viruses. In future experiments, we will detect if the PECAM‐1 expression is lowered when treated with SARS‐CoV‐2 and TNF‐α or if PECAM‐1 is internalized in the cell. Moreover, we can do mRNA and immunoblotting analysis to elaborate the changes in protein pathways as a consequence of COVID‐19.
Severe Acute Respiratory Syndrome Coronavirus 2 (SARS‐CoV‐2), more commonly known as COVID‐19, is a virus that has spread at a rapid rate beginning in 2019 causing a pandemic. Currently, we remain in ...this pandemic, and as this virus progresses it is creating mutations of itself, called variants. With these variants, it has become increasingly difficult to control COVID‐19 and keep track of new variants that continue to infect people everywhere. In Orange County, California, COVID‐19 and its variants have enabled county wide shut downs and thousands of infections and deaths. To investigate its devastating effect, five positive COVID‐19 nasal samples were collected between February 2021 to July 2021, and the RNA was extracted. After extraction, a Reverse Transcription Polymerase Chain Reaction (RT‐PCR) was done to amplify 1,727 base pairs of the S‐ Gene within the COVID‐19 genome and was sent to sequencing. These sequences were then aligned using the Basic Local Alignment Search Tool (BLAST) and compared to the original COVID‐19 S‐Gene genome to analyze possible variations. Within the five samples, 30 different mutations were detected and a phylogenetic tree was generated. Awareness of the genetic variation of SARS‐CoV‐2 provides potential future development for more effective targeted research.
Severe viral pneumonia caused by the severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) is characterized by a hyperinflammatory state typified by elevated circulating pro‐inflammatory ...cytokines, frequently leading to potentially lethal vascular complications including thromboembolism, disseminated intracellular coagulopathy and vasculitis. Though endothelial infection and subsequent endothelial damage have been described in patients with fatal COVID‐19, the mechanism by which this occurs remains elusive, particularly given that, under naïve conditions, pulmonary endothelial cells demonstrate minimal cell surface expression of the SARS‐CoV‐2 binding receptor ACE2. Herein we describe SARS‐CoV‐2 infection of the pulmonary endothelium in postmortem lung samples from individuals who died of COVID‐19, demonstrating both heterogeneous ACE2 expression and endothelial damage (Figure). In primary endothelial cell cultures, we show that SARS‐CoV‐2 infection is dependent on the induction of ACE2 protein expression and that this process is facilitated by type 1 interferon‐alpha (IFNα) or ‐beta(β) ‐ two of the main anti‐viral cytokines induced in severe SARS‐CoV‐2 infection ‐ but not significantly by other cytokines (including interleukin 6 and interferon g /λ). Our findings suggest that the stereotypical anti‐viral interferon response may paradoxically facilitate the propagation of COVID‐19 from the respiratory epithelium to the vasculature, raising concerns regarding the use of exogenous IFNα/β in the treatment of patients with COVID‐19.
Introduction
Coronavirus disease 2019 (COVID‐19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS‐CoV2). The portal of entry for the virus is peptidase ACE2, a major key player of ...the renin‐angiotensin‐aldosterone system (RAAS), resulting in severe lung injury. Although COVID‐19 mainly manifests as an acute respiratory distress syndrome (ARDS), there is increasing evidence of neurological symptoms in patients infected by COVID‐19. Yet, there is limited understanding of how COVID‐19 impacts the central nervous system (CNS). We speculate that such neurological symptoms maybe a consequence of a dysfunction of the blood‐brain barrier (BBB) with our central hypothesis that the neurological effects of SARS CoV‐2 are driven by chronic hypoxic stress‐impaired ACE2 at the BBB.
Methods
An in‐vitro human induced pluripotent stem cells (hiPSCs) BBB model was used in the study. Such model was exposed to various levels of hypoxia (1,5 and 10%) for up to 24 hours. In addition, normoxic cells were treated with Angiotensin II (AngII) or Angiotensin 1‐7 (degradation by product of AngII by ACE2). Changes in the barrier function was assessed using TEER, permeability to fluorescein and tight junction staining. Changes in ACE2 and MasR expression was assessed by immunofluorescence, whereas ACE2 shedding and HIF‐1 alpha expression was assessed by ELISA.
Results
Mild (10%) hypoxia was sufficient to induce the loss of barrier function. Secretion of ACE2 under hypoxia followed a biphasic pattern, with highest levels at 5% and 10%. Ang II and Ang1‐7 had little effect on the barrier function under normoxic condition. The hypoxic exposure induced shedding of the membrane bound ACE2 and molecular mechanism of hypoxic exposure in regulation of ACE2 occurs in a HIF1α‐ dependent manner.
Discussion
Our preliminary data suggest that our human model of the BBB responds to hypoxia and express critical components of RAAS. Both ACE2 and MasR negatively respond to mild hypoxia followed by a decreased barrier function with no changes in tight junction complex. Such loss was correlated with increased ACE2 shedding in HIF1α‐ dependent manner. We are currently investigating role of Ang 1‐7 in rescuing barrier function under hypoxic stress.
Parkinsonism, the clinical term for a disorder with prominent bradykinesia and variable associated extrapyramidal signs and symptoms, is accompanied by degeneration of the nigrostriatal dopaminergic ...system, with neuronal loss and reactive gliosis in the substantia nigra found at autopsy. Parkinsonism is pathologically heterogeneous, with the most common pathologic substrates related to abnormalities in the presynaptic protein α-synuclein or the microtubule binding protein tau. In idiopathic Parkinson's disease (PD), α-synuclein accumulates in neuronal perikarya (Lewy bodies) and neuronal processes (Lewy neurites). The disease process is multifocal and involves select central nervous system neurons and peripheral autonomic nervous system neurons. The particular set of neurons affected determines nonmotor clinical presentations. Multiple system atrophy (MSA) is the other major α-synucleinopathy. It is also associated with autonomic dysfunction and in some cases with cerebellar signs. The hallmark histopathologic feature of MSA is accumulation of α-synuclein within glial cytoplasmic inclusions (GCI). The most common of the Parkinsonian tauopathies is progressive supranuclear palsy (PSP), which is clinically associated with severe postural instability leading to early falls. The tau pathology of PSP also affects both neurons and glia. Given the population frequency of PD, α-synuclein pathology similar to that in PD, but not accompanied by neuronal loss, is relatively common (10% of people over 65 years of age) in neurologically normal individuals, leading to proposed staging schemes for PD progression. Although MSA-like and PSP-like pathology can be detected in neurologically normal individuals, such cases are too infrequent to permit assessment of patterns of disease progression.
Autopsies of deceased with a confirmed severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection can provide important insights into the novel disease and its course. Furthermore, ...autopsies are essential for the correct statistical recording of the coronavirus disease 2019 (COVID-19) deaths. In the northern German Federal State of Hamburg, all deaths of Hamburg citizens with ante- or postmortem PCR-confirmed SARS-CoV-2 infection have been autopsied since the outbreak of the pandemic in Germany. Our evaluation provides a systematic overview of the first 80 consecutive full autopsies. A proposal for the categorisation of deaths with SARS-CoV-2 infection is presented (category 1: definite COVID-19 death; category 2: probable COVID-19 death; category 3: possible COVID-19 death with an equal alternative cause of death; category 4: SARS-CoV-2 detection with cause of death not associated to COVID-19). In six cases, SARS-CoV-2 infection was diagnosed postmortem by a positive PCR test in a nasopharyngeal or lung tissue swab. In the other 74 cases, SARS-CoV-2 infection had already been known antemortem. The deceased were aged between 52 and 96 years (average 79.2 years, median 82.4 years). In the study cohort, 34 deceased were female (38%) and 46 male (62%). Overall, 38% of the deceased were overweight or obese. All deceased, except for two women, in whom no significant pre-existing conditions were found autoptically, had relevant comorbidities (in descending order of frequency): (1) diseases of the cardiovascular system, (2) lung diseases, (3) central nervous system diseases, (4) kidney diseases, and (5) diabetes mellitus. A total of 76 cases (95%) were classified as COVID-19 deaths, corresponding to categories 1–3. Four deaths (5%) were defined as non-COVID-19 deaths with virus-independent causes of death. In eight cases, pneumonia was combined with a fulminant pulmonary artery embolism. Peripheral pulmonary artery embolisms were found in nine other cases. Overall, deep vein thrombosis has been found in 40% of the cases. This study provides the largest overview of autopsies of SARS-CoV-2-infected patients presented so far.
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