Long-Haulers: Spike Protein Present on Covid And in Vaccines Dangerously Modifies Genes Even After Exposure – Texas Tech Uni Study

study by the Texas Tech University published in Biochemistry and Molecular Biology section of the FASEB journal, has found that spike proteins which Covid-19 uses to penetrate cells and is also supplied in abundance by vaccines to stimulate the body’s immune response, modify the body’s genes.

The study coordinators, Nicholas Evans et al. found that even after exposure, these proteins stimulate continued gene expression and inflammatory events that may be behind the long haul Covid and post vaccine syndromes.

Quoting from a review of the study as we published earlier:

“Results from a new cell study at Texas Tech University Health Sciences Center, US, suggest that the SARS-CoV-2 Spike (S) protein can bring about long-term gene expression changes. The findings could help explain why some COVID-19 patients experience symptoms such as shortness of breath and dizziness long after clearing the infection, a condition known as long COVID.

“We found that exposure to the SARS-CoV-2 S protein alone was enough to change baseline gene expression in airway cells,” said Nicholas Evans, one of the researchers. “This suggests that symptoms seen in patients may initially result from the S protein interacting with the cells directly.””

https://www.drugtargetreview.com/news/90224/gene-changes-caused-by-spike-protein-could-explain-long-covid/

The study is titled “Lung Time No See”: SARS-Cov-2 Spike Protein Changes Genetic Expression in Human Primary Bronchial Epithelial Cells After Recovery” and is published in the Biochemistry and Molecular Biology section of the FASEB journal.

https://faseb.onlinelibrary.wiley.com/doi/abs/10.1096/fasebj.2021.35.S1.03097

Long Haul Covid and Long Haul Post-Vaccine Syndromes

Long haul covid or Long-COVID or COVID long-haulers according to a new review can present with as many as 55 long term symptoms. The most common of which are “fatigue (58%), headache (44%), attention disorder (27%), hair loss (25%), and dyspnea (24%)…Diseases such as stroke and diabetes mellitus were also present.” Psychiatric problems like dementia and insomnia are also included. Smell and taste deficiency may persist as also cough and lung abnormalities. Autoimmune problems where the body fights itself is also part of this plethora of presentations. Weight loss, palpitations, renal failure, mood disorders, throat pain and sputum, myocarditis, arrhythmia, OCD, intermittent fever, digestive problems are some more.

These same symptoms as well as symptoms of acute covid infection are now reported months after being vaccinated according to Dr. Bruce Patterson, a pioneer in figuring out Covid and Long haul syndromes.

The precise cause of Long-COVID and Long-Post-Vaccine is being investigated but it may be due to organ damage or persistent autoimmune or inflammatory damage after the infection. Another recent study found Epstein Bar virus reactivated in 73% of long haulers and blamed this for the chronic fatigue, raynaud’s phenomenon and other related symptoms in long haulers.

“We found over 73 percent of COVID-19 patients who were experiencing long COVID symptoms were also positive for EBV reactivation.”

The long-lasting gene changes that result from spike proteins as discovered by Texas Tech University may well be the culprits in these prolonged syndromes.

Watch Dr. Patterson Discuss Long Haul Post-Vaccine Syndrome on Dr. Bean

Video culled from https://www.youtube.com/watch?v=JwjJs5ZHKJI&t=1210s @ 20mins

Dr. Thomas E. Levy, MD, JD writes in OrthoMolecular that “depending on the cell types to which such spike proteins bind, a wide variety of diseases with autoimmune qualities can result.” He recommends treating both syndromes the same way with Vitamin C, Ivermectin, Quercetin and other agents.

“long-haul COVID syndrome likely represents a low-grade unresolved smoldering COVID infection with the same kind of spike protein persistence and clinical impact as is seen in many individuals after their COVID vaccinations (Mendelson et al., 2020; Aucott and Rebman, 2021; Raveendran, 2021).”

http://orthomolecular.org/resources/omns/v17n15.shtml

He further postulates an effect of the spike protein on the ACE2 receptor which has roles in essential pathways that protect blood vessels and other vital physiological processes.

“By itself, the disruption of ACE2 receptor function in so many areas of the body has resulted in an array of different side effects (Ashraf et al., 2021).” Dr Levy writes.

Salk researchers earlier in April found that the spike protein was dangerous to blood vessels by itself, their investigations “proving that the spike protein alone was enough to cause disease.”

Texas Tech University described the dangerous gene-modifying effects of the spike protein as being long-lived and leading to genetic inflammatory changes even after exposure.

“The researchers found that cultured human airway cells exposed to both low and high concentrations of purified S protein showed differences in gene expression that remained even after the cells recovered from the exposure. The top genes included ones related to inflammatory response.”

The study researcher, Nicholas Evans, et al. Concluded:

“Our preliminary results suggest that the SARS-CoV-2 spike protein is enough to change the baseline protein expression in primary HBECs. After recovery, genes related to immune response retained changes in gene expression, and these may indicate relevant long-term effects in asymptomatic patients. Additionally, the interplay between immune response and other pathways after SARS-CoV-2 spike protein exposure should be investigated in the future.”

NewsRescue experts opine: “It is plausible that candidates with lower immunities face more adverse reactions and are on greater risk of Long haul post-vaccine syndrome because it takes longer for their immune system to kick off and wash out the spike proteins produced in the body post vaccine. Studies should investigate any relationship between Long haul post-vaccine and immune compromise.

“Of the various vaccines, AstraZeneca distributed in Africa and the rest of the poorer third world is the worst for many reasons including the fact that it delivers the wild-type unmodified spike protein which has the capacity to transform to the ‘post-fusion’ state and enter the cell and as such has the potential to cause more gene modification or other side effects.”

Animal studies have found toxic effects of spike proteins.

“…those animal studies where Spike protein was produced by a pseudovirus, or the S1 subunit was administered directly. Both of these caused pathology in the animals all by itself, without coronavirus itself being present.”

https://blogs.sciencemag.org/pipeline/archives/2021/06/15/the-novavax-vaccine-data-and-spike-proteins-in-general

Vaccine Technology and the Spike Protein

All current vaccines introduce the spike protein in the recipient. Oxford/AstraZeneca and Janssen (Johnson and Johnson) use ‘vectors’, J&J uses adenovirus type 26 (Ad26) as its vector, while AstraZeneca uses a chimpanzee adenovirus to introduce the spike protein. Moderna, Pfizer/BioNTech vaccines introduce mRNA which is a sort of pre-protein. This mRNA is then read by your cells and translated into S1 and modified spike proteins.

While Oxford/AZ introduces the same exact spike protein the virus uses, Moderna, Pfizer/BioNTech and J&J make the body produce a modified spike protein which has the addition of two amino acids (these are the building blocks of proteins), which modify the spike protein so it stays stuck in the ‘pre-fusion’ state and does not switch to the ‘post-fusion’ state which can penetrate the cell.

– Read more: https://newsrescue.com/long-haul-post-vaxx-syndrome/

Certainly more studies shall continue to help humanity deal with these complex new long haul syndromes and to completely reverse the effects of the possible gene modification as reported.

Resolving “Long-Haul COVID” and Vaccine Toxicity: Neutralizing the Spike Protein

Authors: Dr. Thomas E. Levy, MD, JD   July 1, 2021

Although the mainstream media outlets might have you believe otherwise, the vaccines that continue to be administered for the COVID pandemic are emerging as substantial sources of morbidity and mortality themselves. While the degree to which these negative outcomes of the COVID vaccines can be debated, there is no question that enough disease and death have already occurred to warrant cessation of the administration of these vaccines until additional scientifically-based research can examine the balance between its now clear-cut side effects versus its potential (and still not yet clearly proven) ability to prevent new COVID infections.

Nevertheless, enough vaccinations have already been administered to warrant concern that a new “pandemic” of illness and death may well be emerging from the side effects that continue to be documented in steadily increasing numbers. The vaccine-induced “culprit” that is now receiving most of the attention and is the focus of much new research is the COVID virus fragment known as the spike protein. Its physiological impact appears to be doing far more harm than good (COVID antibody induction), and its manner of introduction appears to be fueling its ongoing replication with a continuing presence inside the body for an indefinite length of time.

The physical appearance of the COVID virus can been depicted as a central sphere of viral protein surrounded completely by spear-like appendages. Known as spike proteins, they are very analogous to the quills surrounding a porcupine. And just as the porcupine stabs its victim, these spike proteins penetrate into cell membranes throughout the body. After this penetration, protein-dissolving enzymes are activated, the cell membrane breaks down, the viral sphere enters the cytoplasm through this membrane breach, and the metabolism of the cell is subsequently “hijacked” to manufacture more viral particles. These spike proteins are the focus of a great deal of ongoing research examining vaccine side effects (Belouzard et al., 2012; Shang et al., 2020).

The Spike Protein’s Toxic Effects in the Body

The spike protein first attaches to ACE2 (angiotensin converting enzyme 2) receptors in the cell membranes (Pillay, 2020). This initial binding step is vital to triggering the subsequent sequence of events that brings the virus inside the cell. When this binding is blocked by competition or prompt enough displacement with an appropriate therapeutic agent, the virus cannot enter the cell, the infectious process is effectively stopped, and the immune defenses of the body are freed to mop up, metabolize, and eliminate the viral pathogens, or just the spike protein alone if free and no longer attached to a viral particle.

Although ACE2 is found in many different cells throughout the body, it is especially noteworthy to realize that it is the initial target bound by coronavirus on the epithelial cells lining the airways after pathogen inhalation (Hoffmann et al., 2020). ACE2 expression (concentration) is also especially high on lung alveolar epithelial cells (Alifano et al., 2020). This cell membrane-bound virus can then begin the process that eventually results in the severe acute respiratory syndrome (SARS) seen in clinically-advanced COVID infections (Perrotta et al., 2020; Saponaro et al., 2020). The SARS presentation manifests most clearly when the degree of oxidative stress in the lungs is very elevated. This stage of COVID infection-related extreme oxidative stress is often referred to in the literature as a cytokine storm, and left unchecked this invariably leads to death (Hu et al., 2021).

Increasing concern has focused on the continued presence of the spike protein in the blood by itself, unattached to a virion, following COVID vaccination. Supposedly intended to initiate an immune response to the entire virus particle, the spike protein injections are disseminating throughout the body rather than staying put in the upper arm at the vaccine site while the immune response to it evolves. Furthermore, it also appears that these circulating spike proteins can enter cells on their own and replicate themselves without attached virus particles. This not only wreaks havoc inside those cells, it helps to assure the indefinite presence of the spike protein throughout the body.

It has also been suggested that large amounts of spike protein are just binding ACE2 receptors and not proceeding any further into the cell, effectively blocking or disabling normal ACE2 function in a given tissue. Additionally, when the spike protein binds a cell wall and “stops” there, the spike protein serves as a hapten (antigen) which can then initiate an autoimmune (antibody or antibody-like) response to the cell itself, rather than to the virus particle to which it is usually attached. Depending on the cell types to which such spike proteins bind, a wide variety of diseases with autoimmune qualities can result.

Finally, another worrisome property of the spike protein which alone would be of great concern is that the spike protein itself appears to be highly toxic. This intrinsic toxicity, along with the apparent ability of the spike protein to replicate itself indefinitely within the cells it enters, probably represents the way in which the vaccine can inflict its worst long-term damage, as the production of this toxin can continue indefinitely without other external factors at play.

In fact, the long-haul COVID syndrome likely represents a low-grade unresolved smoldering COVID infection with the same kind of spike protein persistence and clinical impact as is seen in many individuals after their COVID vaccinations (Mendelson et al., 2020; Aucott and Rebman, 2021; Raveendran, 2021).

Post-Vaccine Complications

While the totality of the mechanisms involved are far from being completely understood and worked out, the increasing occurrence of post-vaccine clinical complications is nevertheless very clear-cut and must be addressed as rapidly and effectively as possible. By itself, the disruption of ACE2 receptor function in so many areas of the body has resulted in an array of different side effects (Ashraf et al., 2021). Such clinical complications being seen in different organ systems and areas of the body, can all occur in the following three clinical situations. All three are “spike protein syndromes,” although the acute infection always includes the entirety of the virus particles along with the spike protein during the initial phases of the infection.

  1. in an active COVID-19 infection,
  2. during the long-haul COVID syndrome, or
  3. in response to a spike protein-laden vaccine, include the following:
    • Heart failure, heart injury, heart attack, myocarditis (Chen et al., 2020; Sawalha et al., 2021)
    • Pulmonary hypertension, pulmonary thromboembolism and thrombosis, lung tissue damage, possible pulmonary fibrosis (McDonald, 2020; Mishra et al., 2020; Pasqualetto et al., 2020; Potus et al., 2020; Dhawan et al., 2021)
    • Increased venous and arterial thromboembolic events (Ali and Spinler, 2021)
    • Diabetes (Yang et al., 2010; Lima-Martinez et al., 2021)
    • Neurological complications, including encephalopathy, seizures, headaches, and neuromuscular diseases. Also, hypercoagulability and stroke (AboTaleb, 2020; Bobker and Robbins, 2020; Hassett et al., 2020; Hess et al., 2020)
    • Gut dysbiosis, inflammatory bowel disease, and leaky gut (Perisetti et al., 2020; Zeppa et al., 2020; Hunt et al., 2021)
    • Kidney damage (Han and Ye, 2021)
    • Impaired male reproductive capacity (Seymen, 2021)
    • Skin lesions and other cutaneous manifestations (Galli et al., 2020)
    • General autoimmune diseases, autoimmune hemolytic anemia (Jacobs and Eichbaum, 2021; Liu et al., 2021)
    • Liver injury (Roth et al., 2021)

In structuring a clinical protocol to stop the ravages of persistent spike protein presence throughout the body, it is first important to realize that the protocol should be able to effectively treat any aspect of COVID infection, including those periods during active infection, after “active” infection (long-haul COVID), and during ongoing spike protein presence secondary to either “chronic” COVID infection or resulting from COVID vaccine administration.

Treatment Protocols

As is the case with any treatment for any condition, factors of expense, availability, and patient compliance always play a role in determining what treatment a given patient will actually undergo for a given period of time. As such, no one specific protocol will be appropriate for all patients, even if the same pathology is present. Ideally, of course, the best protocol is to use all of the options discussed below.

When the entirety of the protocol is not possible or feasible, which is most often the case, the combination of HP nebulization, high-dose vitamin C, and appropriately-dosed ivermectin is an excellent way to effectively address long-haul COVID and persistent spike protein syndromes.

Much of the rationale of the protocols is based on what is known about the spike protein and how it appears to inflict its harm. The following aspects of spike protein pathophysiology need to all be considered in crafting an optimal treatment protocol:

  • The ongoing production of spike protein by the vaccine-supplied mRNA into the cells for the purpose of stimulating the production of neutralizing antibodies (Khehra et al., 2021)
  • The binding of the spike protein, with or without an attached virion, to an ACE2 binding site on the cell wall, as an initial step to dissolving that portion of the cell wall, permitting the spike protein (and attached virus particle if present) into the cell
  • The binding of the spike protein to an ACE2 binding site, but just remaining bound to that site and not initiating enzymatic degradation of the cell wall, with or without an attached virion
  • The degree to which circulating spike protein is present in the blood and actively disseminating throughout the body
  • The fact that the spike protein by itself is toxic (pro-oxidant in nature) and capable of generating disease-generating oxidative stress throughout the body. This is addressed most directly by persistent and highly-dosed vitamin C.

Therapeutic Agents and Their Mechanisms

A substantial number of agents have already been found to be highly effective in resolving COVID infections, and even more are continuing to be discovered as worldwide research efforts have so intensely focused on curing this infection (Levy, 2020). Some of the most effective agents and their mechanisms of actions include the following:

  1. Hydrogen peroxide (HP) nebulization. Correctly applied, this treatment eliminates acute COVID pathogen presence and any other chronic pathogen colonizations persisting in the aerodigestive tract. Also, a positive healing effect on the lower digestive tract is typically seen, as less pathogens and their associated pro-oxidant toxins are chronically swallowed. Stunning anecdotal evidence has already been seen documenting the ability of HP nebulization to cure even advanced COVID infections (20 of 20 cases) as a monotherapy. (Levy, 2021). All of the supporting research, scientific analysis, and practical suggestions on this therapy is available as a free eBook download [Rapid Virus Recovery] (Levy, 2021).
  2. Vitamin C. Vitamin C works synergistically with HP in eradicating pathogens. It gives strong general immune support, while working to support the optimal healing of damaged cells and tissues. Clinically, it is the most potent antitoxin ever described in the literature, and no reports of it failing to neutralize any acute intoxication when administered appropriately have been published. Continuing persistent and highly-dosed vitamin C in all its forms will prove to be the most useful intervention when there is a large amount of circulating toxic spike protein present. Intravenous, regular oral forms, and liposome-encapsulated oral forms are all very useful in resolving any infection and neutralizing any toxin (Levy, 2002). There is also a polyphenol-based supplement that appears to allow some humans to synthesize their own vitamin C, which could prove to be of enormous protective and healing capacity with COVID patients and vaccine recipients. (https://formula216.com/).
  3. Ivermectin. This agent has powerful antiparasitic and antiviral properties. Evidence indicates that ivermectin binds the ACE2 receptor site that the spike protein needs to bind to proceed with entry into the cell and the replication of viral protein (Lehrer and Rheinstein, 2020; Eweas et al., 2021). Also, under some circumstances, the binding of the spike protein to the ACE2 receptor does not activate the enzymes needed to enter the cell. Possibly, ivermectin might also competitively displace such bound spike protein from the cell walls as well when a sufficient dose is taken. It also appears that circulating spike protein can be bound up directly by ivermectin, rendering it inactive and making it accessible for metabolic processing and excretion (Saha and Raihan, 2021). Where there has been mass administration of ivermectin for parasitic diseases in Africa there has also been noted a significantly lower incidence of COVID-19 infection (Hellwig and Maia, 2021). Ivermectin is also very safe when administered appropriately (Munoz et al., 2018).
  4. Hydroxychloroquine (HCQ) and Chloroquine (CQ). Both HCQ and CQ have been shown to be very effective agents in resolving acute COVID-19 infections. They have also both been shown to be zinc ionophores that can increase intracellular zinc levels which can then inhibit the enzyme activity needed for viral replication. However, both HCQ and CQ have also been found to block the binding of COVID virus spike proteins to the ACE2 receptors needed to initiate viral entry into the cells, giving scientific support for their utility as more directly interfering with spike protein activity before the virus ever breaches the cell (Fantini et al., 2020; Sehailia and Chemat, 2020; Wang et al., 2020).
  5. Quercetin. Similar to HCQ and CQ, quercetin also serves as a zinc ionophore. And like HCQ and CQ, quercetin appears to also work to block the binding of COVID virus spike proteins to the ACE2 receptors, impairing spike protein-virus entry into the cell, or impairing spike protein alonef from entering the cells (Pan et al., 2020; Derosa et al., 2021). Many other phytochemicals and bioflavonoids are demonstrating this ACE2 binding capacity as well (Pandey et al., 2020; Maiti and Banerjee, 2021).
  6. Other Bio-Oxidative Therapies. These include ozone, ultraviolet blood irradiation, and hyperbaric oxygen therapy (in addition to hydrogen peroxide and vitamin C). These three therapies are highly effective in patients with acute COVID infections. It is less clear how effective they would be for long-haul COVID syndrome and patients suffering from ongoing vaccine-generated spike protein syndromes. That is not to say, however, that all three would not prove to be just as excellent for dealing with the spike protein as with the intact virus. It just remains to be determined.
  7. Baseline Vital Immune Support Supplementation. There are definitely hundreds, and perhaps thousands, of quality vitamin, mineral, and nutrient supplements that are all capable of making some contribution to reaching and maintaining optimal health, while minimizing the chances of contracting any kind of infectious disease. A baseline regimen of supplementation that factors in expense, overall health impact, and convenience should include vitamin C, vitamin D3, magnesium chloride (other forms good, but chloride form optimal for antiviral impact), vitamin K2, zinc, and an iodine supplement, such as Lugol’s solution or iodoral. More specific guidance in dosing can be found in Appendix A of Hidden Epidemic, also available as a free eBook download (Levy, 2017). Specifics on mixing up a solution of magnesium chloride for regular supplementation are also available (Levy, 2020).

[More detail on the therapeutic agents above is available in Chapter 10 of Rapid Virus Recovery]

The suggested optimal way to deal with acute COVID that has evolved into long-haul COVID, or with symptoms consistent with the toxic effects of circulating spike protein post-vaccination, is to always eliminate any active or chronic areas of pathogen proliferation with HP nebulization. Vitamin C supplementation should be optimized at the same time. 50-gram infusions of sodium ascorbate should be administered at least several times weekly as long as there is symptomatology attributable to long-haul COVID and circulating spike protein.

Initially, a 25-gram infusion of sodium ascorbate given three times a day should prove to be even more effective as circulating vitamin C is rapidly excreted. Oral vitamin C supplementation should be taken as well, either as several grams of liposome-encapsulated vitamin C daily, or as a teaspoon of sodium ascorbate powder several times daily. One capsule daily of Formula 216 can be added to this as well.

With the “foundation” of HP nebulization and vitamin C supplementation in place, the best prescription medicines to counter long-haul COVID and circulating spike protein would be with ivermectin first, and then HCQ or HQ if the clinical response is not acceptable. Dosages would need to be determined by the prescribing physician.

Along with the baseline immune support supplements noted above, quercetin, 500 to 1,000 mg daily, should be added as well.

Any and all of the above recommendations should be undertaken with the guidance of a trusted physician or other appropriately-trained health care professional.

Recap

Even as the COVID pandemic appears to be slowly subsiding, many individuals are now chronically ill with long-haul COVID and/or with the side effects of a COVID vaccination. It would appear that both clinical situations are primarily characterized by persistent presence of the spike protein and its negative impact on different tissues and organs.

Treatment is aimed at neutralizing the direct toxic impact of spike protein, while working to block its ability to bind the receptors needed to hijack the metabolism of the cell into making new viruses and/or more spike protein. At the same time, treatment measures are taken to assure that there is as complete an elimination of active or smoldering COVID infection remaining in the patient.

The views expressed in this article are the author’s and not necessarily those of the Orthomolecular Medicine News Service or all members of its Editorial Board. OMNS invites alternative viewpoints. Submissions may be sent directly to Andrew W. Saul, Editor, at the email contact address further below.

[Editor’s note: The information in this article is not meant to replace the advice of your doctor. Please consult with your personal physician before making any adjustments to your health care routine.]

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Durability of mRNA-1273 vaccine–induced antibodies against SARS-CoV-2 variants

  1. Authors: ViewAmarendra Pegu1,†, Sarah O’Connell1,† View ORCID ProfileStephen D. Schmidt1,, Sijy O’Dell1,View ORCID ProfileChloe A. Talana1, Lilin Lai2, Jim Albert Science  12 Aug 2021: eabj4176m OI: 10.1126/science.abj4176

Abstract

SARS-CoV-2 mutations may diminish vaccine-induced protective immune responses, particularly as antibody titers wane over time. Here, we assess the impact of SARS-CoV-2 variants B.1.1.7 (Alpha), B.1.351 (Beta), P.1 (Gamma), B.1.429 (Epsilon), B.1.526 (Iota), and B.1.617.2 (Delta) on binding, neutralizing, and ACE2-competing antibodies elicited by the vaccine mRNA-1273 over seven months. Cross-reactive neutralizing responses were rare after a single dose. At the peak of response to the second vaccine dose, all individuals had responses to all variants. Binding and functional antibodies against variants persisted in most subjects, albeit at low levels, for 6-months after the primary series of the mRNA-1273 vaccine. Across all assays, B.1.351 had the lowest antibody recognition. These data complement ongoing studies to inform the potential need for additional boost vaccinations.

SARS-CoV-2, the virus that causes COVID-19, has infected millions of people worldwide fueling the ongoing global pandemic (1). The combination of RNA virus mutation rates, replication and recombination, in a very large number of individuals is conducive to the emergence of viral variants with improved replication capacity and transmissibility, as well as immunological escape. Of particular interest are the Variants of Concern B.1.1.7 (20I/501Y.V1 or Alpha), B.1.351 (20H/501Y.V2 or Beta), P.1 (Gamma; first identified in Brazil), B.1.429 (Cal20 or Epsilon; first identified in California), and B.1.617.2 (Delta; first identified in India); and Variant of Interest B.1.526 (Iota; first identified in New York). In multiple studies, B.1.351 is the most resistant to neutralization by convalescent or vaccinee sera, with 6-15 fold less neutralization activity for sera from individuals immunized with vaccines based on the virus strain first described in January 2020 (Wuhan-Hu-1, spike also called WA1) (29). Most of these prior studies evaluated sera from vaccinated individuals at timepoints soon after the first or second dose, and had limited data on the durability of such responses. Likewise, clinical studies have reported somewhat reduced efficacy and effectiveness against the B.1.1.7, B.1.351, and B.1.617.2 variants (1012). Although such data provide critical insights into the performance of the vaccines against viral variants, they have not fully addressed the durability of cross-reactive binding and functional antibodies.

Here we investigate the impact of SARS-CoV-2 variants on recognition by sera from individuals who received two 100 mcg doses of the SARS-CoV-2 vaccine mRNA-1273. mRNA-1273 encodes the full-length stabilized spike protein of the WA1 and was administered as a two-dose series 28-days apart. We previously described the binding and neutralization activity against the WA1 SARS-CoV-2 spike longitudinally over 7 months from the first vaccination in volunteers from the Phase 1 trial of the mRNA-1273 vaccine (1316). In the current study, we demonstrate the utility of employing multiple methodologies to assess SARS-CoV-2 vaccine-elicited humoral immunity to variant viruses over time. We tested sera from a random sample of 8 volunteers in each of three age groups: 18-55, 55-70, and 71+ years of age, all of whom had samples available from four timepoints: 4 weeks after the first dose, and two weeks, 3 months, and 6 months after the second dose (Days 29, 43, 119, and 209 after the first dose, respectively).

Three functional assays and two binding assays were used to assess the humoral immune response to the SARS-CoV-2 spike protein. SARS-CoV-2 neutralization was measured using both a lentivirus-based pseudovirus assay, and a live-virus focus reduction neutralization test (FRNT) (17). The third functional assay was a MSD-ECLIA (Meso Scale Discovery-Electrochemiluminescence immunoassay)-based ACE2 competition assay. This method measured the ability of mRNA-1273 vaccine-elicited antibodies to compete with labeled soluble ACE2 for binding to the specific RBD (WA1 or variant) spotted onto the MSD plate. Antibody binding to cell-surface expressed full-length spike was analyzed by flow cytometry. Binding to soluble protein was measured by interferometry in the MSD-ECLIA platform. All samples were assessed against WA1 and the B.1.1.7 and B.1.351 variants in each of these orthogonal serology assays. In addition, all samples were tested against WA1 containing the D614G mutation in both neutralization assays, as well as binding in the cell-surface assay. Further variants were tested in binding assays as follows: S-2P and RBD binding, P.1 against all samples; cell-surface spike binding, P.1, B.1.429, B.1.526, and B.1.617.2 against all samples. A subset of samples – Day 43 to capture the peak response, and Day 209 to look at durability – were evaluated by pseudovirus neutralization against P.1, B.1.429, B.1.526, and B.1.617.2. The specific sequences used in each assay are defined in table S1.

We first assessed the patterns of antibody activity over time. Consistently across assays, low-level recognition of all variants was observed after a single dose (Day 29) (Fig. 1). Activity against all variants peaked two weeks after the second dose (Day 43) with moderate declines over time through Day 209 (Fig. 1). Notably, the values obtained for each assay on a per-sample basis correlated with each other (fig. S1). We next evaluated the relative impact of each variant, considering all timepoints together. Employing the pseudovirus assay, the neutralizing activity was highest against D614G and lowest against B.1.351, with values for all other variants tested falling in between those two variants (Fig. 1A and Fig. 2A). Similar to previous reports from our group (15) and others (18), pseudovirus neutralization ID50s to D614G were 3-fold higher than to WA1 (Fig. 2G). In contrast, using the live-virus FRNT neutralization assay (Fig. 1B and Fig. 2B), titers to WA1 were higher than to D614G, consistent with previous reports for that assay (19). For all other variants, the impact in the live-virus and pseudovirus neutralization assays were concordant: titers against B.1.1.7 were similar to D614G and lower against B.1.351. ACE2 competition was highest for WA1 RBD, intermediate for B.1.1.7, and lowest for B.1.351 (Fig. 1C and Fig. 2C). Spike-binding antibodies were measured using two different methodologies. In the cell-surface spike binding assay, serum antibodies were bound to full-length, membrane-embedded spike on the surface of transfected cells and measured by flow cytometry (20). In this assay (Fig. 1D and Fig. 2D), WA1 and D614G were nearly indistinguishable, with ~1.5-fold reduced binding to B.1.1.7, B.1.526, B.1.617.2, and 2.4 to 3.0-fold reduced binding to P.1, and B.1.429, and B.1.351. We also used the MSD-ECLIA multiplex binding assay to simultaneously measure IgG binding against both the stabilized soluble spike protein S-2P (21) and RBD proteins derived from WA1 and the B.1.1.7, B.1.351, and P.1 variants. The ECLIA assay showed slightly reduced binding to the variant S-2P (Fig. 1E and Fig. 2E) and RBD (Fig. 1F and Fig. 2F) proteins, with the rank order of highest to lowest binding as follows: WA1, B.1.1.7, P.1, and B.1.351. The overall effect of each variant in each assay is tabulated in Fig. 2G, which shows the geometric mean of the ratios between values for WA1 and variant or D614G and variant. In all assays, B.1.351 was the variant that caused the greatest reduction in titers compared to WA1 or D614G.

For More Information: https://science.sciencemag.org/content/early/2021/08/11/science.abj4176.full

Risk of severe COVID-19 disease with ACE inhibitors and angiotensin receptor blockers: cohort study including 8.3 million people

  1. Julia Hippisley-Cox1, Duncan Young2,3, Carol Coupland4, Keith M Channon5, Pui San Tan6, David A Harrison7, Kathryn Rowan8,  Paul Aveyard6, Ian D Pavord9, Peter J Watkinson5,10
  2. Correspondence to Prof Julia Hippisley-Cox, Primary Care Health Sciences, University of Oxford, Oxford OX1 

Abstract

Background 

There is uncertainty about the associations of angiotensive enzyme (ACE) inhibitor and angiotensin receptor blocker (ARB) drugs with COVID-19 disease. We studied whether patients prescribed these drugs had altered risks of contracting severe COVID-19 disease and receiving associated intensive care unit (ICU) admission.

Methods 

This was a prospective cohort study using routinely collected data from 1205 general practices in England with 8.28 million participants aged 20–99 years. We used Cox proportional hazards models to derive adjusted HRs for exposure to ACE inhibitor and ARB drugs adjusted for sociodemographic factors, concurrent medications and geographical region. The primary outcomes were: (a) COVID-19 RT-PCR diagnosed disease and (b) COVID-19 disease resulting in ICU care.

Findings 

Of 19 486 patients who had COVID-19 disease, 1286 received ICU care. ACE inhibitors were associated with a significantly reduced risk of COVID-19 disease (adjusted HR 0.71, 95% CI 0.67 to 0.74) but no increased risk of ICU care (adjusted HR 0.89, 95% CI 0.75 to 1.06) after adjusting for a wide range of confounders. Adjusted HRs for ARBs were 0.63 (95% CI 0.59 to 0.67) for COVID-19 disease and 1.02 (95% CI 0.83 to 1.25) for ICU care.

There were significant interactions between ethnicity and ACE inhibitors and ARBs for COVID-19 disease. The risk of COVID-19 disease associated with ACE inhibitors was higher in Caribbean (adjusted HR 1.05, 95% CI 0.87 to 1.28) and Black African (adjusted HR 1.31, 95% CI 1.08 to 1.59) groups than the white group (adjusted HR 0.66, 95% CI 0.63 to 0.70). A higher risk of COVID-19 with ARBs was seen for Black African (adjusted HR 1.24, 95% CI 0.99 to 1.58) than the white (adjusted HR 0.56, 95% CI 0.52 to 0.62) group.

Interpretation 

ACE inhibitors and ARBs are associated with reduced risks of COVID-19 disease after adjusting for a wide range of variables. Neither ACE inhibitors nor ARBs are associated with significantly increased risks of receiving ICU care. Variations between different ethnic groups raise the possibility of ethnic-specific effects of ACE inhibitors/ARBs on COVID-19 disease susceptibility and severity which deserves further study.

Link between fever, diarrhea, severe COVID-19, and persistent anti-SARS-CoV-2 antibodies

Authors: By Dr. Liji Thomas, MD Jan 7 2021

Ever since the coronavirus disease 2019 (COVID-19) pandemic began, there have been many attempts to understand the nature and duration of immunity against the causative agent, the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2).

A new preprint research paper appearing on the medRxiv* server describes a link between the persistence of neutralizing antibodies against the virus, disease severity, and specific COVID-19 symptoms.

Permanent immunity is essential if the pandemic is to end. In the earlier SARS epidemic, antibodies were found to last for three or more years after infection in most patients. With the current virus, it may last for six or more months at least, as appears from some reports. Other researchers have concluded that immunity wanes rapidly over the same period, with some patients who were tested positive for antibodies becoming seronegative later on. This discrepancy may be traceable to variation in testing methods, sample sizes and testing time points, as well as disease severity.

Study details

The current study looked at a population of over a hundred convalescent COVID-19 patients, testing most of them for antibodies at five weeks and three months from symptom resolution.

The researchers used a multiplex assay that measured the Immunoglobulin G (IgG) levels against four SARS-CoV-2 antigens, one from SARS-CoV, and four from circulating seasonal coronaviruses. In addition, they carried out an inhibition assay against SARS-CoV-2 spike receptor-binding domain (RBD)-angiotensin-converting enzyme 2 (ACE2) binding and a neutralization assay against the virus. The antibody titers were then plotted against various clinical features and demographic factors.

Antibody titers higher in COVID-19 convalescents

The researchers found that severe disease is correlated with advanced age and the male sex. Patients with underlying vascular disease were more likely to be hospitalized with COVID-19, but those with asthma were relatively spared.

Convalescent COVID-19 patients had higher IgG levels against all four SARS-CoV-2 antigens, relative to controls, and in 98% of cases, at least one of the tests was likely to show higher binding compared to controls. IgGs targeting the viral spike and RBD were likely to be much more discriminatory between SARS-CoV-2 patients and controls. Interestingly, anti-SARS-CoV IgG, as well as anti-seasonal betacoronavirus antibodies, were likely to be higher in these patients.

Anti-spike and anti-nucleocapsid IgG levels, as well as neutralizing antibody titers, were higher in convalescent hospitalized COVID-19 patients than in convalescent non-hospitalized patients, and the titers were positively associated with disease severity.Antibodies against SARS-CoV-2 persist three months after COVID-19 symptom resolution. Sera from COVID-19 convalescent subjects (n=79) collected 5 weeks (w) and 3 months (m) after symptom resolution were subjected to multiplex assay to detect IgG that binds to SARS-CoV-2 S, NTD, RBD and N antigens (A), to RBD-ACE2 binding inhibition assay (B), and to SARS-CoV-2 neutralization assay (C). Dots, lines, and asterisks in red represent non-hospitalized (n=67) and in blue represent hospitalized (n=12) subjects with lines connecting the two time points for individual subjects (*p<0.05 and **p<0.01 by paired t test).Antibodies against SARS-CoV-2 persist three months after COVID-19 symptom resolution. Sera from COVID-19 convalescent subjects (n=79) collected 5 weeks (w) and 3 months (m) after symptom resolution were subjected to multiplex assay to detect IgG that binds to SARS-CoV-2 S, NTD, RBD and N antigens (A), to RBD-ACE2 binding inhibition assay (B), and to SARS-CoV-2 neutralization assay (C). Dots, lines, and asterisks in red represent non-hospitalized (n=67) and in blue represent hospitalized (n=12) subjects with lines connecting the two time points for individual subjects (*p<0.05 and **p<0.01 by paired t test).

Clinical correlates of higher antibody titer

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When antibody titers in non-hospitalized subjects were compared with clinical and demographic variables, they found that older males with a higher body mass index (BMI) and a Charlson Comorbidity Index score >2 were likely to have higher antibody titers. COVID-19 symptoms that correlated with higher antibody levels in these patients comprise fever, diarrhea, abdominal pain and loss of appetite. Chest tightening, headache and sore throat were associated with less severe symptoms.

The link between the specific symptoms listed above with higher antibody titers could indicate that they mark a robust systemic inflammatory response, which in turn is necessary for a strong antibody response. Diarrhea may mark severe disease, but it is strange that in this case, it was not more frequent in the hospitalized cohort. Alternatively, diarrhea may have strengthened the immune antibody response via the exposure of the virus to more immune cells via the damaged enteric mucosa. More study is required to clarify this finding.

Potential substitute for neutralizing assay

The binding assay showed that the convalescent serum at five weeks inhibited RBD-ACE2 binding much more powerfully than control serum. Neutralizing activity was also higher in these sera, but in 15% of cases, convalescent patients showed comparable neutralizing antibody titers to those in control sera. On the whole, however, there was a positive association between neutralizing antibody titer, anti-SARS-CoV-2 IgG titers, and inhibition of ACE2 binding.

Persistent immunity at three months

This study also shows that SARS-CoV-2 antibodies persist in these patients at even three months after symptoms subside, with persistent IgG titers against the SARS-CoV-2 spike, RBD, nucleocapsid and N-terminal domain antigens. Binding and neutralization assays remained highly inhibitory throughout this period. The same was true of antibodies against the other coronaviruses tested as well, an effect that has been seen with other viruses and could be the result of cross-reactive anti-SARS-CoV-2 antibodies. Alternatively, it could be due to the activation of memory B cells formed in response to infection by the seasonal beta-coronaviruses.

Conclusion

IgG titers, particularly against S and RBD, and RBD-ACE2 binding inhibition better differentiate between COVID-19 convalescent and naive individuals than the neutralizing assay,” the researchers concluded.

These could be combined into a single diagnostic test, they suggest, with extreme sensitivity and specificity. The correlation with neutralizing antibody titers could indicate that the neutralizing assay, which is more expensive, sophisticated and expensive, as well as more dangerous for the investigators, could be replaced by the other antibody tests without loss of value.

In short, the study shows that specific antibodies persist for three months at least following recovery; antibody titers correlate with COVID-19-related fever, loss of appetite, abdominal pain and diarrhea; and are also higher in older males with more severe disease, a higher BMI and CCI above 2. Further research would help understand the lowest protective titer that prevents reinfection, and the duration of immunity.

*Important Notice

medRxiv publishes preliminary scientific reports that are not peer-reviewed and, therefore, should not be regarded as conclusive, guide clinical practice/health-related behavior, or treated as established information.Journal reference:

The Epidemiology, Transmission, and Diagnosis of COVID-19

Authors: By: Neesha C. Siriwardane & Rodney Shackelford, DO, Ph.D. April 15, 2020

Introduction to COVID-19

Coronaviruses are enveloped single-stranded RNA viruses of the Coronaviridae family and order Nidovirales (1). The viruses are named for their “crown” of club-shaped S glycoprotein spikes, which surround the viruses and mediate viral attachment to host cell membranes (1-3). Coronaviruses are found in domestic and wild animals, and four coronaviruses commonly infect the human population, causing upper respiratory tract infections with mild common cold symptoms (1,4). Generally, animal coronaviruses do not spread within human populations, however rarely zoonotic coronaviruses evolve into strains that infect humans, often causing severe or fatal illnesses (4). Recently, three coronaviruses with zoonotic origins have entered the human population; severe acute respiratory syndrome coronavirus-2 (SARS) in 2003, Middle Eastern respiratory syndrome (MERS) in 2012, and most recently, coronavirus disease 2019 (COVID-19), also termed SARS-CoV-2, which the World Health Organization declared a Public Health Emergency of International Concern on January 31st, 2020 (4,5). 

COVID19 Biology, Spread, and Origin

COVID-19 replicates within epithelial cells, where the COVID-19 S glycoprotein attaches to the ACE2 receptor on type 2 pneumocytes and ciliated bronchial epithelial cells of the lungs. Following this, the virus enters the cells and rapidly uses host cell biochemical pathways to replicate viral proteins and RNA, which assemble into viruses that in turn infect other cells (3,5,6). Following these cycles of replication and re-infection, the infected cells show cytopathic changes, followed by various degrees of pulmonary inflammation, changes in cytokine expression, and disease symptoms (5-7). The ACE2 receptor also occurs throughout most of the gastrointestinal tract and a recent analysis of stool samples from COVID-19 patients revealed that up to 50% of those infected with the virus have a COVID-19 enteric infection (8).

COVID-19 was first identified on December 31st, 2020 in Wuhan China, when twenty-seven patients presented with pneumonia of unknown cause. Some of the patients worked in the Hunan seafood market, which sold both live and recently slaughtered wild animals (4,9).  Clusters of cases found in individuals in contact with the patients (family members and healthcare workers) indicated a human-to-human transmission pattern (9,10). Initial efforts to limit the spread of the virus were insufficient and the virus soon spread throughout China. Presently COVID-19 occurs in 175 countries, with 1,309,439 cases worldwide, with 72,638 deaths as of April 6th, 2020 (4). Presently, the most affected countries are the United States, Italy, Spain, and China, with the United States showing a rapid increase in cases, and as of April 6th, 2020 there are 351,890 COVID-19 infected, 10,377 dead, and 18,940 recovered (4).  In the US the first case presented on January 19th, 2020, when an otherwise healthy 35-year-old man presented to an urgent care clinic in Washington State with a four-day history of a persistent dry cough and a two-day history of nausea and vomiting.  The patient had a recent travel history to Wuhan, China. On January 20th, 2020 the patient tested positive for COVID-19.  The patient developed pneumonia and pulmonary infiltrates, and was treated with supplemental oxygen, vancomycin, and remdesivir. By day eight of hospitalization, the patient showed significant improvement (11). 

Sequence analyses of the COVID-19 genome revealed that it has a 96.2% similarity to a bat coronavirus collected in Yunnan province, China. These analyses furthermore showed no evidence that the virus is a laboratory construct (12-14). A recent sequence analysis also found that COVID-19 shows significant variations in its functional sites, and has evolved into two major types (termed L and S). The L type is more prevalent, is likely derived from the S type, and may be more aggressive and spread more easily (14,15). 

Transmission

While sequence analyses strongly suggest an initial animal-to-human transmission, COVID-19 is now a human-to-human contact spread worldwide pandemic (4,9-11). Three main transmission routes are identified; 1) transmission by respiratory droplets, 2) contract transmission, and 3) aerosol transmission (16). Transmission by droplets occurs when respiratory droplets are expelled by an infected individual by coughing and are inhaled or ingested by individuals in relatively close proximity.  Contact transmission occurs when respiratory droplets or secretions are deposited on a surface and another individual picks up the virus by touching the surface and transfers it to their face (nose, mouth, or eyes), propagating the infection. The exact time that COVID-19 remains infective on contaminated surfaces is unknown, although it may be up to several days (4,16). Aerosol transmission occurs when respiratory droplets from an infected individual mix with air and initiate an infection when inhaled (16). Transmission by respiratory droplets appears to be the most common mechanism for new infections and even normal breathing and speech can transmit the virus (4,16,17). The observation that COVID-19 can cause enteric infections also suggests that it may be spread by oral-fecal transmission; however, this has not been verified (8). A recent study has also demonstrated that about 30% of COIVID-19 patients present with diarrhea, with 20% having diarrhea as their first symptom. These patients are more likely to have COVID-19 positive stool upon testing and a longer, but less severe disease course (18).  Recently possible COVID-19 transmission from mother to newborns (vertical transmission) has been documented. The significance of this in terms of newborn health and possible birth defects is currently unknown (19). 

The basic reproductive number or R0, measures the expected number of cases generated by one infection case within a population where all the individuals can become infected. Any number over 1.0 means that the infection can propagate throughout a susceptible population (4). For COVID-19, this value appears to be between 2.2 and 4.6 (4,20,21). Unpublished studies have stated that the COVID10 R0 value may be as high as 6.6, however, these studies are still in peer review. 

COVID-19 Prevention

There is no vaccine available to prevent COVID-19 infection, and thus prevention presently centers on limiting COVID-19 exposures as much as possible within the general population (22). Recommendations to reduce transmission within community include; 1) hand hygiene with simultaneous avoidance of touching the face, 2) respiratory hygiene, 3) utilizing personal protective equipment (PPE) such as facemasks, 4) disinfecting surfaces and objects that are frequently touched, and 5) limiting social contacts, especially with infected individuals  (4,9,17,22). Hand hygiene includes frequent hand-washing with soap and water for twenty seconds, especially after contact with respiratory secretions produced by activities such as coughing or sneezing. When soap and water are unavailable, hand sanitizer that contains at least 60% alcohol is recommended (4,17,22). PPE such as N95 respirators are routinely used by healthcare workers during droplet precaution protocols when caring for patients with respiratory illnesses. One retrospective study done in Hunan, China demonstrated N95 masks were extremely efficient at preventing COVID-19 transfer from infected patients to healthcare workers (4,22-24). It is also likely that wearing some form of mask protection is useful to prevent COVID19 spread and is now recommended by the CDC (25). 

Although transmission of COVID-19 is primarily through respiratory droplets, well-studied human coronaviruses such as HCoV, SARS, and MERS coronaviruses have been determined to remain infectious on inanimate surfaces at room temperature for up to nine days. They are less likely to persist for this amount of time at a temperature of 30°C or more (26). Therefore, contaminated surfaces can remain a potential source of transmission. The Environmental Protection Agency has produced a database of appropriate agents for COVID-19 disinfection (27). Limiting social contact usually has three levels; 1) isolating infected individuals from the non-infected, 2) isolating individuals who are likely to have been exposed to the disease from those not exposed, and 3) social distancing. The later includes community containment, were all individuals limit their social interactions by avoiding group gatherings, school closures, social distancing, workplace distancing, and staying at home (28,29). In an adapted influenza epidemic simulation model, comparing scenarios with no intervention to social distancing and estimated a reduction of the number of infections by 99.3% (28). In a similar study, social distancing was estimated to be able to reduce COVID-19 infections by 92% (29). Presently, these measured are being applied in many countries throughout the world and have been shown to be at least partially effective if given sufficient time (4,17,30). Such measures proved effective during the 2003 SARS outbreak in Singapore (30). 

Symptoms, Clinical Findings, and Mortality 

On average COVID-19 symptoms appear 5.2 days following exposure and death fourteen days later, with these time periods being shorter in individuals 70-years-old or older (31,32). People of any age can be infected with COVID-19, although infections are uncommon in children and most common between the ages of 30-65 years, with men more affected than women (32,33). The symptoms vary from asymptomatic/paucisymptomatic to respiratory failure requiring mechanical ventilation, septic shock, multiple organ dysfunction, and death (4,9,32,33). The most common symptoms include a dry cough which can become productive as the illness progresses (76%), fever (98%), myalgia/fatigue (44%), dyspnea (55%), and pneumoniae (81%), with less common symptoms being headache, diarrhea (26%), and lymphopenia (44%) (4,32,33). Rare events such as COVID-19 acute hemorrhagic necrotizing encephalopathy have been documented and one paper describes conjunctivitis, including conjunctival hyperemia, chemosis, epiphora, or increased secretions in 30% of COVID-19 patients (34,35). Interestingly, about 30-60% of those infected with COVID-19 also experience a loss of their ability to taste and smell (36). 

The clinical features of COVID-19 include bilateral lung involvement showing patchy shadows or ground-glass opacities identified by chest X-ray or CT scanning (34). Patients can develop atypical pneumoniae with acute lung injury and acute respiratory distress syndrome (33). Additionally, elevations of aspartate aminotransferase and/or alanine aminotransferase (41%), C-reactive protein (86%), serum ferritin (63%), and increased pro-inflammatory cytokines, whose levels correlate positively with the severity of the symptoms (4,31-33,37-39).

About 81% of COVID-19 infections are mild and the patients make complete recoveries (38). Older patients and those with comorbidities such as diabetes, cardiovascular disease, hypertension, and chronic obstructive pulmonary disease have a more difficult clinical course (31-33,37-39). In one study, 72% of patients requiring ICU treatment had some of these concurrent comorbidities (40). According to the WHO 14% of COVID-19 cases are severe and require hospitalization, 5% are very severe and will require ICU care and likely ventilation, and 4% will die (41). Severity will be increased by older age and comorbidities (4,40,41). If effective treatments and vaccines are not found, the pandemic may cause slightly less than one-half billion deaths, or 6% of the world’s population (41). Since many individuals infected with COVID-19 appear to show no symptoms, the actual mortality rate of COIVD-19 is likely much less than 4% (42). An accurate understanding of the typical clinical course and mortality rate of COVID-19 will require time and large scale testing.         

COVID-19 Diagnosis

COVID-19 symptoms are nonspecific and a definitive diagnosis requires laboratory testing, combined with a thorough patient history.  Two common molecular diagnostic methods for COVID-19 are real-time reverse polymerase chain reaction (RT-PCR) and high-throughput whole-genome sequencing.  RT-PCR is used more often as it is cost more effective, less complex, and has a short turnaround time. Blood and respiratory secretions are analyzed, with bronchoalveolar lavage fluid giving the best test results (43). Although the technique has worked on stool samples, as yet stool is less often tested (8,43). RT-PCR involves the isolation and purification of the COVID-19 RNA, followed by using an enzyme called “reverse transcriptase” to copy the viral RNA into DNA. The DNA is amplified through multiple rounds of PCR using viral nucleic acid-specific DNA primer sequences. Allowing in a short time the COVID-19 genome ti be amplified millions of times and then easily analyzed (43). RT-PCR COVID-19 testing is FDA approved and the testing volume in the US is rapidly increasing (44,45). The FDA has also recently approved a COVID-19 diagnostic test that detects anti-COVID-19 IgM and IgG antibodies in patient serum, plasma, or venipuncture whole blood (43). As anti-COVID-19 antibody formation takes time, so a negative result does not completely preclude a COVID-19 infection, especially early infections. Last, as COVID-19 often causes bilateral pulmonary infiltrates, correlating diagnostic testing results with lung chest CT or X-ray results can be helpful (4,31-33,37-39).  

Testing for COVID-19 is based on a high clinical suspicion and current recommendations suggest testing patients with a fever and/or acute respiratory illness. These recommendations are categorized into priority levels, with high priority individuals being hospitalized patients and symptomatic healthcare facility workers. Low priority individuals include those with mild disease, asymptomatic healthcare workers, and symptomatic essential infrastructure workers. The latter group will receive testing as resources become available (41,46,47). 

COVID-19 Possible Treatments

Presently research on possible COVIS-19 infection treatments and vaccines are underway (48). At the writing of this article many different drugs are being examined, however any data supporting the use of any specific drug treating COVID-19 is thin as best. A few drugs that might have promise are:  

Hydroxychloroquine

Hydroxychloroquine has been used to treat malarial infections for seventy years and in cell cultures it has anti-viral effects against COVID-19 (49). In one small non-randomized clinical trial in France, twenty individuals infected with COVID-19 who received hydroxychloroquine showed a reduced COVID-19 viral load, as measured on nasopharyngeal viral carriage, compared to untreated controls (50). Six individuals who also received azithromycin with hydroxychloroquine had their viral load lessened further (50). In one small study in China, a similar drug (chloroquine) was superior in reducing COVID-19 viral levels in treated individuals compared to untreated control individuals (51).  These results are preliminary, but promising. 

Remdesivir

Remdesivir is a drug that showed value in treating patients infected with SARS (52). COVID-19 and SARS show about 80% sequence similarity and since Remdesivir has been used to treat SARS, it might have value in treating COVID-19 (52). These trials are underway (48). Remdesivir was also used to treat the first case of COIVD-19 identified within the US (11). There are many other drugs being examined to treat COVID-19 infections, however, the data on all of them is presently slight to none, and research has only begun. There is an enormous research effort underway, and progress should be rapid (48). 

Conclusion

Our understanding of COVID-19 is changing extremely rapidly and new findings come out daily. Combating COVID-19 effectively will require multiple steps; including slowing the spread of the virus through socially isolating and measures such as hand washing. The development of effective drug treatments and vaccines is already a priority and rapid progress is being made (48). Additionally, many areas of the world, such as South American and sub-Saharan Africa, will be affected by the COVID-19 pandemic and are likely to have their economies and healthcare systems put under extreme stress. Dealing with the healthcare crisis in these countries will be very difficult. Lastly, several recent viral pandemics (SARS, MERS, and COVID-19) have come from areas where wildlife is regularly traded, butchered, and eaten in conditions that favor the spread of dangerous viruses between species, and eventually into human populations. The prevention of new viral pandemics will require improved handling of wild species, better separation of wild animals from domestic animals, and better regulated and lowered trade in wild animals, such as bats, which are known to be a risk for carrying potentially deadly viruses to human populations (53). 

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Study reveals mouth as primary source of COVID-19 infection

While most COVID-19 research has focused on the nose and lungs, this is the first study to identify the mouth as a primary site for coronavirus infection and underscores the importance of wearing a face covering and physical distancing.

By University Communications, Thursday, October 29th, 2020

A team of researchers led by the University of North Carolina at Chapel Hill and the National Institute of Dental and Craniofacial Research reveals coronavirus can take hold in the salivary glands where it replicates, and in some cases, leads to prolonged disease when infected saliva is swallowed into the gastrointestinal tract or aspirated to the lungs where it can lead to pneumonia.

While most COVID-19 research has focused on the nose and lungs, this is the first study to identify the mouth as a primary site for coronavirus infection and underscores the importance of wearing a face covering and physical distancing. The results have not been peer-reviewed.

“Our results show oral infection of COVID-19 may be underappreciated,” said senior study author Kevin M. Byrd, research instructor at the UNC Adams School of Dentistry and the Anthony R. Volpe Research Scholar at the American Dental Association Science and Research Institute. “Like nasal infection, oral infection could underlie the asymptomatic spread that makes this disease so hard to contain.”

Byrd along with Blake Warner, chief of the Salivary Disorders Unit at the National Institute of Dental and Craniofacial Research, coordinated the research conducted at the National Institutes of Health, Wellcome Sanger Institute, UNC Marsico Lung Institute and the J. Craig Venter Institute.

Researchers are just beginning to explore the oral symptoms patients experience during COVID-19, such as loss of taste or smell and persistent dry mouth.

In the study, researchers report preliminary results from a clinical trial of 40 subjects with COVID-19 which showed sloughed epithelial cells lining the mouth can be infected with SARS-CoV-2, the coronavirus that causes COVID-19. The amount of virus in patient saliva was positively correlated with taste and smell changes, according to the study.

Relying on oral cell identity maps, researchers also looked at where in the mouth the virus infects. They surveyed oral tissues with the highest levels of ACE2, the receptor that helps coronavirus grab and invade human cells.

Based on ACE2 expression and analysis of cadaver tissue, the most likely sites of infection in the mouth are the salivary glands, tongue and tonsil, the study showed.

The findings provide more evidence of the role of saliva in COVID-19. COVID-19 infection, specifically in the mouth, can allow the virus to spread internally and to others as the infected person breathes, speaks and coughs.

For More Information: https://www.unc.edu/posts/2020/10/29/study-reveals-mouth-as-primary-source-of-covid-19-infection/

How does coronavirus kill? Clinicians trace a ferocious rampage through the body, from brain to toes

Authors: By Meredith WadmanJennifer Couzin-FrankelJocelyn KaiserCatherine MatacicApr. 17, 2020 , 6:45 PM

On rounds in a 20-bed intensive care unit one recent day, physician Joshua Denson assessed two patients with seizures, many with respiratory failure and others whose kidneys were on a dangerous downhill slide. Days earlier, his rounds had been interrupted as his team tried, and failed, to resuscitate a young woman whose heart had stopped. All shared one thing, says Denson, a pulmonary and critical care physician at the Tulane University School of Medicine. “They are all COVID positive.”

As the number of confirmed cases of COVID-19 surges past 2.2 million globally and deaths surpass 150,000, clinicians and pathologists are struggling to understand the damage wrought by the coronavirus as it tears through the body. They are realizing that although the lungs are ground zero, its reach can extend to many organs including the heart and blood vessels, kidneys, gut, and brain.

“[The disease] can attack almost anything in the body with devastating consequences,” says cardiologist Harlan Krumholz of Yale University and Yale-New Haven Hospital, who is leading multiple efforts to gather clinical data on COVID-19. “Its ferocity is breathtaking and humbling.”

Understanding the rampage could help the doctors on the front lines treat the fraction of infected people who become desperately and sometimes mysteriously ill. Does a dangerous, newly observed tendency to blood clotting transform some mild cases into life-threatening emergencies? Is an overzealous immune response behind the worst cases, suggesting treatment with immune-suppressing drugs could help? What explains the startlingly low blood oxygen that some physicians are reporting in patients who nonetheless are not gasping for breath? “Taking a systems approach may be beneficial as we start thinking about therapies,” says Nilam Mangalmurti, a pulmonary intensivist at the Hospital of the University of Pennsylvania (HUP).

What follows is a snapshot of the fast-evolving understanding of how the virus attacks cells around the body, especially in the roughly 5% of patients who become critically ill. Despite the more than 1000 papers now spilling into journals and onto preprint servers every week, a clear picture is elusive, as the virus acts like no pathogen humanity has ever seen. Without larger, prospective controlled studies that are only now being launched, scientists must pull information from small studies and case reports, often published at warp speed and not yet peer reviewed. “We need to keep a very open mind as this phenomenon goes forward,” says Nancy Reau, a liver transplant physician who has been treating COVID-19 patients at Rush University Medical Center. “We are still learning.”

The infection begins

When an infected person expels virus-laden droplets and someone else inhales them, the novel coronavirus, called SARS-CoV-2, enters the nose and throat. It finds a welcome home in the lining of the nose, according to a preprint from scientists at the Wellcome Sanger Institute and elsewhere. They found that cells there are rich in a cell-surface receptor called angiotensin-converting enzyme 2 (ACE2). Throughout the body, the presence of ACE2, which normally helps regulate blood pressure, marks tissues vulnerable to infection, because the virus requires that receptor to enter a cell. Once inside, the virus hijacks the cell’s machinery, making myriad copies of itself and invading new cells.

As the virus multiplies, an infected person may shed copious amounts of it, especially during the first week or so. Symptoms may be absent at this point. Or the virus’ new victim may develop a fever, dry cough, sore throat, loss of smell and taste, or head and body aches.

If the immune system doesn’t beat back SARS-CoV-2 during this initial phase, the virus then marches down the windpipe to attack the lungs, where it can turn deadly. The thinner, distant branches of the lung’s respiratory tree end in tiny air sacs called alveoli, each lined by a single layer of cells that are also rich in ACE2 receptors.

Normally, oxygen crosses the alveoli into the capillaries, tiny blood vessels that lie beside the air sacs; the oxygen is then carried to the rest of the body. But as the immune system wars with the invader, the battle itself disrupts this healthy oxygen transfer. Front-line white blood cells release inflammatory molecules called chemokines, which in turn summon more immune cells that target and kill virus-infected cells, leaving a stew of fluid and dead cells—pus—behind. This is the underlying pathology of pneumonia, with its corresponding symptoms: coughing; fever; and rapid, shallow respiration (see graphic). Some COVID-19 patients recover, sometimes with no more support than oxygen breathed in through nasal prongs.

But others deteriorate, often quite suddenly, developing a condition called acute respiratory distress syndrome (ARDS). Oxygen levels in their blood plummet and they struggle ever harder to breathe. On x-rays and computed tomography scans, their lungs are riddled with white opacities where black space—air—should be. Commonly, these patients end up on ventilators. Many die. Autopsies show their alveoli became stuffed with fluid, white blood cells, mucus, and the detritus of destroyed lung cells.

For More Information: https://www.sciencemag.org/news/2020/04/how-does-coronavirus-kill-clinicians-trace-ferocious-rampage-through-body-brain-toes

Blood molecular markers associated with COVID-19 immunopathology and multi-organ damage

Authors: Yan-Mei ChenYuanting ZhengYing YuYunzhi WangQingxia HuangFeng QianLei SunZhi-Gang SongZiyin ChenJinwen FengYanpeng AnJingcheng YangZhenqiang SuShanyue SunFahui DaiQinsheng ChenQinwei LuPengcheng LiYun LingZhong YangHuiru TangLeming ShiLi JinEdward C HolmesChen DingTong-Yu ZhuYong-Zhen Zhang

Abstract

COVID-19 is characterized by dysregulated immune responses, metabolic dysfunction and adverse effects on the function of multiple organs. To understand host responses to COVID-19 pathophysiology, we combined transcriptomics, proteomics, and metabolomics to identify molecular markers in peripheral blood and plasma samples of 66 COVID-19-infected patients experiencing a range of disease severities and 17 healthy controls. A large number of expressed genes, proteins, metabolites, and extracellular RNAs (exRNAs) exhibit strong associations with various clinical parameters. Multiple sets of tissue-specific proteins and exRNAs varied significantly in both mild and severe patients suggesting a potential impact on tissue function. Chronic activation of neutrophils, IFN-I signaling, and a high level of inflammatory cytokines were observed in patients with severe disease progression. In contrast, COVID-19-infected patients experiencing milder disease symptoms showed robust T-cell responses. Finally, we identified genes, proteins, and exRNAs as potential biomarkers that might assist in predicting the prognosis of SARS-CoV-2 infection. These data refine our understanding of the pathophysiology and clinical progress of COVID-19.

Proteomics, metabolomics and RNAseq data map immune responses in COVID-19 patients with different disease severity, revealing molecular makers associated with disease progression and alterations of tissue-specific proteins.

  • A multi-omics profiling of the host response to SARS-CoV2 infection in 66 clinically diagnosed and laboratory confirmed COVID-19 patients and 17 uninfected controls.
  • Significant correlations between multi-omics data and key clinical parameters.
  • Alteration of tissue-specific proteins and exRNAs.
  • Enhanced activation of immune responses is associated with COVID-19 pathogenesis.
  • Biomarkers to predict COVID-19 clinical outcomes pending clinical validation as prospective marker.

Introduction

Coronaviruses (family Coronaviridae) are a diverse group of positive-sense single-stranded RNA viruses with enveloped virions (Masters & Perlman, 2013; Cui et al2019). Coronaviruses are well known due to the emergence of Severe Acute Respiratory Syndrome (SARS) in 2002–2003 and Middle East Respiratory Syndrome (MERS) in 2012, both of which caused thousands of cases in multiple countries (Ksiazek et al2003; Bermingham et al2012; Cui et al2019). Coronaviruses naturally infect a broad range of vertebrate hosts including mammals and birds (Cui et al2019). As coronavirus primarily target epithelial cells, they are generally associated with gastrointestinal and respiratory infections (Masters & Perlman, 2013; Cui et al2019). In addition, they cause hepatic and neurological diseases of varying severity (Masters & Perlman, 2013).

The world is currently experiencing a disease pandemic (COVID-19) caused by a newly identified coronavirus called SARS-CoV-2 (Wu et al2020a). At the time of writing, there have been more than ~25 million cases of SARS-CoV-2 and ~830,000 deaths globally (WHO, 2020). The disease leads to both mild and severe respiratory manifestations, with the latter prominent in the elderly and those with underlying medical conditions such as cardiovascular and chronic respiratory disease, diabetes, and cancer (Guan et al., 2020). In addition to respiratory syndrome, mild gastrointestinal and/or cardiovascular symptoms and neurological manifestations have been documented in hospitalized COVID-19-infected patients (Gupta et al2020; Mao et al2020). These data point to the complexity of COVID-19 pathogenesis, especially in patients experiencing severe disease.

SARS-CoV-2 is able to use angiotensin-converting enzyme 2 (ACE 2) as a receptor for cell entry (Hoffmann et al2020; Zheng et al2020a; Zhou et al2020b). Aside from lungs, ACE2 is expressed in other organs including heart, liver, kidney, pancreas, and small intestines (Li et al2020; Liu et al2020; Zou et al2020; Chen et al2020a). More recently, ACE2 expression has also been found in Leydig cells in the testes (Li et al2020; Wang & Xu, 2020) and neurological tissue (Baig et al2020; Bullen et al2020; Xu & Lazartigues, 2020). As such, it is possible that these organs might also be infected by SARS-CoV-2, and recent autopsy studies have also revealed multi-organ damage including heart, liver, intestine, pancreas, brain, kidney, and spleen in fatal COVID-19-infected patients (Lax et al2020; Menter et al2020; Varga et al2020; Wichmann et al2020; Wang et al2020c). The host immune response to SARS-CoV-2 may also impact pathogenicity, resulting in severe tissue damage and, occasionally, death (Tay et al2020). Indeed, several studies have reported lymphopenia, exhausted lymphocytes, and cytokine storms in COVID-19-infected patients, especially those with severe symptoms (Blanco-Melo et al2020; Cao, 2020; Chua et al2020; Liao et al2020). Numerous clinical studies have also observed the elevation of lactate dehydrogenase (LDH), IL-6, troponin I, inflammatory markers, and D-dimer in COVID-19-infected patients (Zhou et al2020a; Wang et al2020b). However, despite the enormous burden of morbidity and mortality due to COVID-19, we know little about its pathophysiology, even though this establishes the basis for successful clinical practice, vaccine development, and drug discovery.

Using a multi-omics approach employing cutting-edge transcriptomic, proteomic, and metabolomic technologies, we identified significant molecular alterations in patients with COVID-19 compared with uninfected controls in this study. Our results refine the molecular view of COVID-19 pathophysiology associated with disease progression and clinical outcome.

For More Information: https://www.embopress.org/doi/full/10.15252/embj.2020105896

Severe COVID-19: what have we learned with the immunopathogenesis?

Abstract

The COVID-19 outbreak caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has become a global major concern. In this review, we addressed a theoretical model on immunopathogenesis associated with severe COVID-19, based on the current literature of SARS-CoV-2 and other epidemic pathogenic coronaviruses, such as SARS and MERS. Several studies have suggested that immune dysregulation and hyperinflammatory response induced by SARS-CoV-2 are more involved in disease severity than the virus itself.

Immune dysregulation due to COVID-19 is characterized by delayed and impaired interferon response, lymphocyte exhaustion and cytokine storm that ultimately lead to diffuse lung tissue damage and posterior thrombotic phenomena.

Considering there is a lack of clinical evidence provided by randomized clinical trials, the knowledge about SARS-CoV-2 disease pathogenesis and immune response is a cornerstone to develop rationale-based clinical therapeutic strategies. In this narrative review, the authors aimed to describe the immunopathogenesis of severe forms of COVID-19.

Background

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a positive-sense single-stranded RNA-enveloped virus, is the causative agent of coronavirus disease 2019 (COVID-19), being first identified in Wuhan, China, in December 2019. Previously, other epidemic coronavirus such as severe acute respiratory syndrome coronavirus (SARS-CoV) in 2002 and the middle-east respiratory syndrome coronavirus (MERS-CoV) in 2012, had serious impact on human health and warned the world about the possible reemergence of new pathogenic strains [1]. Despite being a new virus, several common morpho-functional characteristics have been reported between SARS-CoV and the SARS-CoV-2, including the interaction of the viral spike (S) glycoprotein with the human angiotensin converting enzyme 2 (ACE2). These similarities may help understanding some pathophysiological mechanisms and pointing out possible therapeutic targets.

The first step for SARS-CoV-2 entry into the host cell is the interaction between the S glycoprotein and ACE2 on cell surface. Since the latter acts as a viral receptor, the virus will only infect ACE2 expressing cells, notably type II pneumocytes. These cells represent 83% of the ACE2-expressing cells in humans, but cells from other tissues and organs, such as heart, kidney, intestine and endothelium, can also express this receptor [2]. A host type 2 transmembrane serine protease, TMPRSS2, facilitates virus entry by priming S glycoprotein. TMPRSS2 entails S protein in subunits S1/S2 and S2´, allowing viral and cellular membrane fusion driven by S2 subunit [3]. Once inside the cell viral positive sense single strand RNA is translated into polyproteins that will form the replicase-transcriptase complex. This complex function as a viral factory producing new viral RNA and viral proteins for viral function and assembly [4]. Considering these particularities, the infection first begins on upper respiratory tract mucosa and then reaches the lungs. The primary tissue damage is related to the direct viral cytopathic effects. At this stage, the virus has the potential to evade the immune system, where an inadequate innate immune response can occur, depending on the viral load and other unknown genetic factors. Subsequently, tissue damage is induced by additional mechanisms derived from a dysregulated adaptive immune response [5].

Although most of COVID-19 cases have a mild clinical course, up to 14% can evolve to a severe form, with respiratory rate ≥ 30/min, hypoxemia with pulse oxygen saturation ≤ 93%, partial pressure of arterial oxygen to fraction of inspired oxygen ratio < 300 and/or pulmonary infiltrates involving more than 50% of lung parenchyma within 24 to 48 h. Up to 5% of the cases can be critical, evolving with respiratory failure, septic shock and/or multiple organ dysfunction, presumably driven by a cytokine storm [6]. Host characteristics, including aging (immunosenescence) and comorbidities (hypertension, diabetes mellitus, lung and heart diseases) may influence the course of the disease [7]. The false paradox between inflammation and immunodeficiency is highlighted by the severe form of COVID-19. Thus, severe pneumonia caused by SARS-CoV-2 is marked by immune system dysfunction and hyperinflammation leading to acute respiratory distress syndrome (ARDS), macrophage activation, hypercytokinemia and coagulopathy [8].

Herein, we aim to review the factors related to the dysregulated immune response against the SARS-CoV-2, along with its relation with severe forms of COVID-19, namely ARDS and cytokine storm (CS).

For More Information: https://advancesinrheumatology.biomedcentral.com/articles/10.1186/s42358-020-00151-7