Progress in the Treatments of COVID-19

Since the beginning of the COVID-19 pandemic, extensive research has focused on therapeutics. A combinational approach of traditional drug development and drug repurposing is necessary to make progress in treating and preventing COVID-19 in a timely manner. While developing new drugs is critical, repurposing existing molecules has been a major strategy. In the early stages of the COVID-19 pandemic, Remdesivir has been identified as a promising repurposing drug to treat COVID-19 patients (1). Indeed, it became the first treatment for COVID-19 to receive the U.S. Food and Drug Administration (FDA)’s Emergency Use Authorization (EUA) (2). Recently, the FDA issued an EUA for three more treatments. In this perspective, we will review these available treatments based on the findings of clinical trials. In addition, we are updating other COVID-19 therapeutics according to recent publications.

Remdesivir is an intravenous nucleotide prodrug of an adenosine analog and is recognized as an antiviral drug against various RNA viruses (3). It binds to the viral RNA-dependent RNA polymerase and incorporates into nascent viral RNA chains. This results in pre-mature termination of RNA transcription and inhibiting viral replication. The FDA approved remdesivir (Veklury) for use in adult and pediatric patients 12 years of age and older and weighing at least 40 kilograms (about 88 pounds) for the treatment of COVID-19 requiring hospitalization (2). In a recently published report, a double-blind, randomized, placebo-controlled trial of intravenous remdesivir was conducted with hospitalized adults having lower respiratory tract infection (4). A total of 1062 patients underwent randomization (with 541 assigned to remdesivir and 521 to placebo). The reported clinical effect of intravenous remdesivir was very modest. The recovery time was significantly shorter among patients who received remdesivir than among those who received placebo (10 days vs. 15 days). A trend toward lower mortality was observed among patients who received remdesivir than among those who received placebo, both at day 15 (6.7% vs. 11.9%) and at day 29 (11.4% vs. 15.2%), but the differences were not statistically significant. Therefore, the primary outcome was the time to recovery. In fact, WHO has issued a conditional recommendation against the use of remdesivir in hospitalized patients, regardless of disease severity (5). Consistent with this, in a huge WHO sponsored clinical study, remdesivir treatment did not improve mortality for hospitalized COVID-19 patients (6). This trial was conducted in 405 hospitals in 30 countries. A total of 11,266 adults were randomized, with 2750 allocated remdesivir, 954 hydroxychloroquine, 1411 lopinavir, 651 interferon plus lopinavir, 1412 interferon only, and 4088 no study drug. Interestingly, this study concluded that remdesivir, hydroxychloroquine, lopinavir and interferon regimens appeared to have little or no effect on hospitalized COVID-19, as indicated by overall mortality, initiation of ventilation and duration of hospital stay.

The pathogenesis of SARS-CoV-2 involves not only viral replication, but also immunomodulation and inflammation (7). Therefore, a combination therapy of remdesivir with other antivirals or antiinflammatory agents could enhance its efficacy. On November 19, 2020, the FDA authorized a combination of remdesivir (Veklury) and the JAK (Janus kinase) inhibitor, baricitinib (Olumiant) for the treatment of suspected or laboratory confirmed COVID-19 in hospitalized adults and pediatric patients 2 years of age or older requiring supplemental oxygen, invasive mechanical ventilation. This approval is based on data from the Adaptive COVID-19 Treatment Trial (ACTT-2), a randomized double-blind, placebo-controlled study to evaluate the efficacy and safety of baricitinib in combination with remdesivir versus placebo with remdesivir in hospitalized patients with or without oxygen requirements (8). The recommended dose for this approval is baricitinib 4-mg once daily for 14 days or until hospital discharge. Baricitinib has been already approved to treat moderate to severe rheumatoid arthritis. Baricitinib, taken orally, inhibits cytokine signaling in the body that play roles in causing inflammatory responses. Rational of this clinical study is that adding an anti-inflammatory agent to the remdesivir treatment can provide additional benefit for patients, including improving mortality outcomes (8). The putative benefit of baricitinib for COVID-19 has been described in a case series of critically ill patients who recovered from COVID-19. The data from this clinical study have not been published yet. It has been suggested that patients treated with baricitinib in combination with remdesivir had a significant reduction in median time to recovery from 8 to 7 days (12.5% improvement) compared to remdesivir (9). The proportion of patients who progressed to ventilation (non-invasive or invasive) or died by Day 29 was lower in baricitinib in combination with remdesivir (23%) compared to remdesivir (28%). The proportion of patients who died by Day 29 was 4.7% for baricitinib in combination with remdesivir vs. 7.1% for remdesivir. However, the safety of this treatment needs to be validated.

In November 9, 2020, the FDA approved investigational monoclonal antibody therapy bamlanivimab for the treatment of mild-to-moderate COVID-19 in adult and pediatric patients (10). Bamlanivimab is authorized for the treatment of nonhospitalized patients with mild to moderate COVID-19 who are at high risk for progressing to severe disease and/or hospitalization This includes those who are 65 years of age or older, or who have certain chronic medical conditions. The approval is based on data from BLAZE-1, a randomized, double-blind, placebo-controlled Phase 2 study in patients with recently diagnosed mild to moderate COVID-19 in the outpatient setting (11). It is not authorized for patients who are hospitalized due to COVID-19 or require oxygen therapy due to COVID-19. Bamlanivimab (Lily)also known as LY-CoV555 and LY3819253) is a neutralizing monoclonal antibody that targets the receptor-binding domain of the spike protein of SARS-CoV-2. Because this drug may block SARS-CoV-2 entry into host cells, it is being evaluated for the treatment of COVID-19. This trial randomly assigned 452 patients to receive a single intravenous infusion of neutralizing antibody LY-CoV555 in one of three doses (700 mg, 2800 mg, or 7000 mg) or placebo (12). After undergoing randomization, patients received an infusion of LY-CoV555 or placebo within a median of 4 days after the onset of symptoms. The viral load at day 11 (the primary outcome) was lower than that in the placebo group only among those who received the 2800-mg dose. However, the evaluation of the effect of LY-CoV555 therapy on patients’ symptoms at earlier time points during treatment (e.g., on day 3) showed a possible treatment effect, with no substantial differences observed among the three doses. While the safety and effectiveness of this investigational therapy continues to be evaluated, bamlanivimab was shown in this trial to reduce COVID-19-related hospitalization or emergency room visits in patients at high risk for disease progression within 28 days after treatment when compared to placebo. For patients at high risk for disease progression, hospitalizations and emergency room visits occurred in 3% of bamlanivimab-treated patients on average compared to 10% in placebo-treated patients. However, a benefit of bamlanivimab treatment has not been shown in patients hospitalized due to COVID-19. Monoclonal antibodies, such as bamlanivimab, may be associated with worse clinical outcomes when administered to hospitalized patients with COVID-19 requiring high flow oxygen or mechanical ventilation (10).

In November 21, 2020, the FDA approved monoclonal antibodies, casirivimab and imdevimab (Regeneron) to be administered together by intravenous infusion for the treatment of mild to moderate COVID-19 in adults and pediatric patients (12 years of age or older weighing at least 40 kilograms) with positive results of direct SARS-CoV-2 viral testing and who are at high risk for progressing to severe COVID-19 (13). This includes those who are 65 years of age or older or who have certain chronic medical conditions. However, this treatment is not authorized for patients who are hospitalized due to COVID-19 or require oxygen therapy due to COVID-19. These monoclonal antibodies are specifically directed against the spike protein of SARS-CoV-2, designed to block the virus’ attachment and entry into human cells. This approval is based on a randomized, double-blind, placebo-controlled clinical trial in 799 non-hospitalized adults with mild to moderate COVID-19 symptoms. Of these patients, 266 received a cocktail of casirivimab and imdevimab (1,200 mg each), 267 received a cocktail of casirivimab and imdevimab (4,000 mg each), and 266 received a placebo intravenously, within three days of obtaining a positive SARS-CoV-2 test. Viral load reduction in patients treated with casirivimab and imdevimab was larger than in patients treated with placebo at day seven. Further, for patients at high risk for disease progression, hospitalizations and emergency room visits occurred in 3% of casirivimab and imdevimab-treated patients on average compared to 9% in placebo-treated patients. The findings of this study have not been published in a peer-reviewed journal yet. The safety and effectiveness of this investigational therapy for use in the treatment of COVID-19 continues to be evaluated.

As an experimental monoclonal antibody treatment, two ultrapotent SARS-CoV-2 human neutralizing antibodies (S2E12 and S2M11) were isolated and characterized for their potential use as a prophylaxis or therapy (14). Cryo–electron microscopy structures show that S2E12 and S2M11 competitively block angiotensin-converting enzyme 2 (ACE2) attachment. Furthermore, S2M11 is a distinct class of potent neutralizers of SARS-CoV-2 by additionally locking the spike in a closed conformation by recognition of a quaternary epitope spanning two adjacent receptor-binding domains. Combinations of monoclonal antibodies leveraging multiple distinct mechanisms of action with additive or synergistic effects could provide additional benefits for clinical application. Indeed, using cocktail of these antibodies enhanced antibody responses to SARS-CoV-2 S and conferred significant protection in hamster models. The mAb cocktails are also expected to take advantage of both ultrapotent neutralization, different mechanisms of action, and Fc-mediated effector functions to protect from a broad spectrum of circulating SARSCoV-2 isolates. The findings of this study also has important implication in implementing antibody cocktails for circumventing or limiting the emergence of viral escape mutants.

Ivermectin is an FDA-approved broad-spectrum antiparasitic agent with demonstrated antiviral activity against a number of DNA and RNA viruses, including SARS-CoV-2 (15). The inhibition of importin α/β1-mediated nuclear import of viral proteins is suggested as the probable mechanism underlying its antiviral activity. In addition, ivermectin could ultimately induce an ionic imbalance that disrupts the potential of the viral membrane, thereby threatening its integrity and functionality.  A recent study showed that a single dose of ivermectin was able to reduce the replication of an Australian isolate of SARS-CoV-2 in Vero/hSLAM cells by 5000-fold (16). In addition to the indication for antiviral therapy, anti-inflammatory intervention may also be necessary to prevent acute lung injury in SARS-CoV-2 infection. With regard to its anti-inflammatory properties, ivermectin have been shown to mitigate skin inflammation. A study evaluating the ability of ivermectin to inhibit lipopolysaccharide (LPS)-induced inflammation revealed significantly decreased production of TNF-alpha, IL-1ss and IL-6 in vivo and in vitro. Further studies may establish the role of ivermectin in inflammatory response caused by SARS-CoV-2. A number of clinical studies are being conducted in various countries. The data from these studies are not available yet. However, it has been suggested that the necessary inhibitory concentration may only be achieved via high dosage regimes in humans. Further, safety of using high-dose antiviral therapy needs to be evaluated. Currently, no commercially available injectable forms of ivermectin are available for human use. The development of ivermectin formulations presents challenges, primarily due to its property of poor water solubility. Novel delivery strategies are needed to optimize ivermectin bioavailability. Furthermore, efficacy of ivermectin for COVID-19 treatment needs to be defined with extensive in vivo study and clinical trials.

Lastly, antiparasitic drug nitazoxanide is widely available and exerts broad-spectrum antiviral activity in vitro. For its use for treatment of mild COVID-19 patients, randomized, double-blind, placebo-controlled was conducted in a multicenter (17). In patients with mild Covid-19, symptom resolution did not differ between the nitazoxanide and placebo groups after 5 days of therapy. However, at the 1-week follow-up, 78% in the nitazoxanide group and 57% in the placebo group reported complete resolution of symptoms. Further, viral load was also reduced after nitazoxanide treatment compared to placebo. This study showed only partially analyzed data from the clinical trials. Complete analysis of this study will be required to clearly assess the efficacy of nitazoxanide for the treatment.

Reinforcing the efficacy of drug repurposing can provide much shorter and less costly development process than developing a new drug. Yet, we have to recognize that beside the very modest activity of remdesivir nothing has so far emerged from these efforts. In fact, patients with respiratory symptoms due to COVID-19 are still best treated by combining the vintage steroids combined with oxygen and anticoagulants.  On the other hand, the monoclonal antibody-based treatments have now become available for people in high-risk groups and seem to be effective early in the course of the disease; yet there is a limitation in the production capacity of such therapies and their cost must not be a barrier to get them widely available in low- and middle-income countries. In general, a combination of two to three different therapeutic agents for the treatment has become a common approach. Finally, treatment which might potentially prevent infection (such as Nitazoxanide, Romark LLC) are being evaluated. Overall, despite many difficulties, there is now hope that innovative and cost-effective therapeutic agents can be effectively developed.

 

References

  1. Wang M, Cao R, Zhang L, et al. Remdesivir and chloroquine effectively inhibit the recently emerged novel coronavirus (2019-nCoV) in vitro. Cell Res. 2020;30(3):269-271. Available at: https://www.ncbi.nlm.nih.gov/pubmed/32020029.
  2. Food and Drug Administration. FDA Approves First Treatment for COVID-19. 2020. Available at: https://www.fda.gov/news-events/press-announcements/fda-approves-first-treatment-covid-19.
  3. Warren TK, et al. Therapeutic efficacy of the small molecule GS-5734 against Ebola virus in rhesus monkeys. Nature. 2016;531:381–385. doi: 10.1038/nature17180.
  4. Beigel JH, Tomashek KM, Dodd LE, et al. Remdesivir for the treatment of Covid-19 — final report. N Engl J Med 2020;383:1813-1826.
  5. World Health Organization. WHO recommends against the use of remdesivir in COVID-19 patients. Available at: https://www.who.int/news-room/feature-stories/detail/who-recommends-against-the-use-of-remdesivir-in-covid-19-patients.
  6. WHO Solidarity Trial Consortium. Repurposed antiviral drugs for covid-19—interim WHO Solidarity trial results. 15 Oct 2020. doi:10.1101/2020.10.15.20209817.
  7. Mehta P, McAuley DF, Brown M, Sanchez E, Tattersall RS, Manson JJ. COVID-19: consider cytokine storm syndromes and immunosuppression. Lancet 2020;395:1033-1034.
  8. National Institute of Health. NIH clinical trial testing antiviral remdesivir plus anti-inflammatory drug baricitinib for COVID-19 begins. Available at: https://www.nih.gov/news-events/news-releases/nih-clinical-trial-testing-antiviral-remdesivir-plus-anti-inflammatory-drug-baricitinib-covid-19-begins.
  9. Lilly. Baricitinib Receives Emergency Use Authorization from the FDA for the Treatment of Hospitalized Patients with COVID-19. Available at: https://investor.lilly.com/news-releases/news-release-details/baricitinib-receives-emergency-use-authorization-fda-treatment
  10. Food and Drug Administration. Fact sheet for healthcare providers: emergency use authorization (EUA) of bamlanivimab. 2020. Available at: https://www.fda.gov/media/143603/download.
  11. Lilly. Lilly’s neutralizing antibody bamlanivimab (LY-CoV555) receives FDA emergency use authorization for the treatment of recently diagnosed COVID-19. Available at: https://investor.lilly.com/news-releases/news-release-details/lillys-neutralizing-antibody-bamlanivimab-ly-cov555-receives-fda.
  12. Chen P, Nirula A, Heller B, et al. SARS-CoV-2 neutralizing antibody LY-CoV555 in outpatients with COVID-19. N Engl J Med. 2020; Available at: https://www.ncbi.nlm.nih.gov/pubmed/33113295.
  13. Food and Drug Administration Coronavirus (COVID-19) Update: FDA Authorizes Monoclonal Antibodies for Treatment of COVID-19. Available at: https://www.fda.gov/news-events/press-announcements/coronavirus-covid-19-update-fda-authorizes-monoclonal-antibodies-treatment-covid-19.
  14. M. ALEJANDRA TORTORICI et al., A potent antibody cocktail blocks attachment of SARS-CoV-2 to the host receptor and activates a protective immune response. SCIENCE20 NOV 2020 : 950-957.
  15. Fabio Rocha Formiga, Roger Leblanc, Juliana de Souza Rebouças, Leonardo Paiva Farias, Ronaldo Nascimento de Oliveira, Lindomar Pena, Ivermectin: an award-winning drug with expected antiviral activity against COVID-19, Journal of Controlled Release, 2020, https://doi.org/10.1016/j.jconrel.2020.10.009.
  16. L. Caly, J.D. Druce, M.G. Catton, D.A. Jans, K.M. Wagstaff. The FDA-approved drug Ivermectin inhibits the replication of SARS-CoV-2 in vitro. Antivir. Res., 178 (2020), p. 104787, 10.1016/j.antiviral.2020.104787.
  17. Patricia R. M. Rocco et al. Early use of nitazoxanide in mild Covid-19 disease: randomized, placebocontrolled trial. medRxiv preprint doi: https://doi.org/10.1101/2020.10.21.20217208.

GVN STATEMENT ON RECENT COVID-19 VACCINE DATA SUCCESS

Baltimore, Maryland, USA, November 17, 2020: The Global Virus Network (GVN), , which comprises global preeminent human and animal virologists from 57 Centers of Excellence and 11 Affiliates in 33 countries, said today that the 94.5% efficacy rate of the Moderna vaccine, according to early data announced yesterday, is a significant step towards developing an effective vaccine to mitigate the COVID-19 pandemic. The GVN added that the announcement from Moderna combined with the recent vaccine data from Pfizer and BioNTech, which demonstrated 90% efficacy, represent major breakthroughs in the global effort to develop an effective vaccine against COVID-19.  The GVN, which has brought together the world’s foremost virologists to collaboratively evaluate the immune response against SARS-CoV-2, congratulates Moderna, Pfizer and BioNtech for their pioneering work and for achieving this milestone.

The two vaccines are mRNA based vaccine and require two-doses of vaccinations. Upon the FDA’s approval for emergency use authorization, Moderna and Pfizer project to produce about 20 million and 50 million doses, respectively, by the end of the year. Widespread vaccinations would be beneficial in mitigating the ongoing pandemic. As emphasized by the companies, the GVN suggests that an important next step is to evaluate the duration of protection, safety, and protective efficacy of vaccines in preventing viral transmission. GVN scientists will continue to contribute to these important milestones to curb, and ultimately control, the COVID-19 pandemic.

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About the Global Virus Network (GVN)

The Global Virus Network (GVN) is essential and critical in the preparedness, defense and first research response to emerging, exiting and unidentified viruses that pose a clear and present threat to public health, working in close coordination with established national and international institutions. It is a coalition comprised of eminent human and animal virologists from 57 Centers of Excellence and 11 Affiliates in 33 countries worldwide, working collaboratively to train the next generation, advance knowledge about how to identify and diagnose pandemic viruses, mitigate and control how such viruses spread and make us sick, as well as develop drugs, vaccines and treatments to combat them. No single institution in the world has expertise in all viral areas other than the GVN, which brings together the finest medical virologists to leverage their individual expertise and coalesce global teams of specialists on the scientific challenges, issues and problems posed by pandemic viruses. The GVN is a non-profit 501(c)(3) organization. For more information, please visit www.gvn.org. Follow us on Twitter @GlobalVirusNews

GVN MEDIA CONTACTS:
Nora Samaranayake
Phone: 410-706-1966; Email: [email protected]

 

Global Virus Network (GVN) and the University of South Florida Launch Online Course “Microbiomes and Their Impact on Viral Infections”

GVN Offers Four Scholarships to Self-Paced Online Training Course

Baltimore, Maryland, USA, November 17, 2020: The Global Virus Network (GVN), together with the University of South Florida (USF) Institute on Microbiomes, recently launched the self-paced online course “Microbiomes and Their Impact on Viral Infections.Taught by world-renowned instructors, this course will provide students, academics, and health professionals with the latest knowledge of the importance and role of microbiomes in preventing, mitigating, and treating diseases. The initiative also supports GVN’s mission to train the next generation of virologists and better prepare mankind for future viral threats.

“This course is timely as virologists around the world work to further their investigations into the causes, catalysts, and prevention mechanisms of viral infection,” said Dr. Christian Bréchot, president of GVN and professor at the USF Health Morsani College of Medicine. “We are pleased to collaborate with the USF Institute on Microbiomes, which houses the online, trans-disciplinary program. It is a terrific example of a much-needed training partnership critical to mitigating viral threats.”

Microbiomes and Their Impact on Viral Infections is a non-credit course comprised of two sessions. The first, “Introduction to Microbiomes,” consists of 11 modules while the second, “Symbiotic Evolutions in the Microbiome World,” comprises nine modules and is available to students for up to eight weeks after the start date. With a transdisciplinary approach, students will have access to lectures and complementary material, and will receive a certificate and a digital badge upon course completion.

GVN awarded four course scholarships to investigators working in various stages of viral infection prevention, including, Joseph Osega, a Kenya-based technical advisor and national HIV recency coordinator, who has extensive knowledge of HIV, malaria and TB diagnostics to build capacity and develop public health infrastructure in Kenya; Nanma Cosmas, a lecturer and a doctoral candidate at the University of Jos, Nigeria, who focuses on prevention of HPV and other sexually transmitted infectious diseases among adolescent and young adults through studies of microbiome in various parts of the body; Onyekachukwu Okeke, a doctoral candidate at the University of Jos, Nigeria, who works at a medical laboratory and has been on the front line during the COVID-19 crisis; and, Sophia Osawe, a doctoral candidate at the University of Jos, who researches the effects of maternal HIV infection and prenatal immunization on the immune responses and growth of infants.

“The learning modules are designed by GVN virologists from 33 nations, 57 Centers of Excellence (CoEs), and 11 affiliated laboratories that are at the heart of GVN’s strength,” said Dr. Ramesh Akkina, a director with a GVN CoE at Colorado State University where he is a professor, and an instructor of the microbiomes course. “Besides furthering research, GVN members are focused on training virologists to help identify, research, and combat pandemics of the future.”

“Since the beginning of the COVID-19 pandemic more than 55 million cases have been reported, and this partnership provides a critical balance between creating new knowledge and making that knowledge available to researchers in the field,” Dr. Brechot added. “I am happy that GVN and USF have come together to partner on important initiatives to advance the transfer of knowledge.  The GVN is pleased to provide necessary training opportunities for tomorrow’s leaders.”

As the only coalition of its kind, GVN leads with scientific, evidence-based solutions to function as an essential global resource for researchers, medical practitioners and policymakers as well as students considering the field of virology as a career choice.

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About the Global Virus Network (GVN)

The Global Virus Network (GVN) is essential and critical in the preparedness, defense and first research response to emerging, exiting and unidentified viruses that pose a clear and present threat to public health, working in close coordination with established national and international institutions. It is a coalition comprised of eminent human and animal virologists from 57 Centers of Excellence and 11 Affiliates in 33 countries worldwide, working collaboratively to train the next generation, advance knowledge about how to identify and diagnose pandemic viruses, mitigate and control how such viruses spread and make us sick, as well as develop drugs, vaccines and treatments to combat them. No single institution in the world has expertise in all viral areas other than the GVN, which brings together the finest medical virologists to leverage their individual expertise and coalesce global teams of specialists on the scientific challenges, issues and problems posed by pandemic viruses. The GVN is a non-profit 501(c)(3) organization. For more information, please visit www.gvn.org. Follow us on Twitter @GlobalVirusNews

About USF Health

USF Health’s mission is to envision and implement the future of health. It is the partnership of the USF Health Morsani College of Medicine, the College of Nursing, the College of Public Health, the Taneja College of Pharmacy, the School of Physical Therapy and Rehabilitation Sciences, the Biomedical Sciences Graduate and Postdoctoral Programs, and USF Health’s multispecialty physicians group. The University of South Florida is a high-impact global research university dedicated to student success. Over the past 10 years, no other public university in the country has risen faster in U.S. News and World Report’s national university rankings than USF. For more information, visit health.usf.edu.

GVN MEDIA CONTACTS:
Chandrani Raysarkar
Phone: 240-535-1574; Email: [email protected]
Nora Samaranayake
Phone: 410-706-1966; Email: [email protected]

Rapid and Frequent Testing for COVID-19

The cases of new SARS-CoV-2 infections are currently increasing at an alarming rate, suggesting that winter might bring a large healthcare crisis. Diagnostics have become a critical tool in curbing COVID-19. It is important that accurate results be obtained rapidly and that collection of samples be as fast, simple, and convenient as possible.

What tests are becoming available, which promise better accuracy and greater speed and ease of use?  Thus far, testing has been based on a real-time reverse transcription polymerase chain reaction (RT-PCR) for SARS-CoV-2 RNA detection and serology on enzyme-linked immunosorbent assay (ELISA) for antibody tests. Yet, PCR can show false positives due to detection of fragmented RNA of virus. This has been an obstacle in determining the presence of infectious virus particles in COVID-19 patients. A recent study showed that the presence of infectious virus may be predictable by evaluating the threshold cycle (Ct value, the number of cycles of PCR amplification) (1).  Now, there are also a variety of novel tests for SARS-CoV-2 RNA detection in development that rely on CRISPR12 or 13 cleavage of SARS-CoV-2 amplified RNA, which releases a visually readable fluorescent chromophore. These rely on isothermal amplification, either LAMP(7) or recombinase polymerase amplification (RPA)(8-10), which does not require a thermal cycler. These test all have rapid turnaround times (on the order of an hour) and sensitivities that can be well comparable with other tests. One of these tests can use a cheap chemical hand warmer as the thermal source(9). Another similar CRISPR based assay can simultaneously detect and identify 169 different human viruses(11), illustrating the potential utility and efficiency of this system. These tests all have advantages of speedy detection and user-friendly application over RT-PCR.

Serological testing is also necessary. The presence of antibodies marks an ongoing or past infection, and anti-spike protein and neutralizing antibodies are considered a possible indication of immunity, although T cell activities are also contributing to protective immunity. Antibody tests can indicate past infections but are prone to be false positive in low frequency settings and have provided a difficulty with uneven results because of wide variations in test kit quality. What serological tests are likely to be most useful? As with RNA tests, this requires sensitivity, specificity, speed, and user-friendly application.

One consideration for serological tests is that there is some degree of serologic cross-reactivity of SARS-CoV-2 proteins with those of the other coronaviruses, especially SARS-CoV-1(12). It is also possible that the S2 spike domain of SARS-CoV-2  may also have some degree of cross-reactivity with that of other human coronavirus (13). Interestingly, some of these cross-reactive antibodies also appear to cross-neutralize, suggesting that there may exist conserved epitopes for the development of a broadly effective vaccine.

Most point-of care serologic assays rely in lateral flow-type assays, in which the results are read as bands on a strip and indicate the presence of IgG and/or IgM antibodies. The regulatory processes for these tests has perhaps lacked some of the usual rigor, and their accuracy tends to vary. This can be a problem in low incidence settings, where false positives are an issue. Fortunately, a recently published  study carefully compared five different test kits to ELISA and viral neutralization tests(14). As might be expected, antibodies were more readily detected when sera were collected more than 14 days after symptom onset. Using samples collected from this time period, three lateral flow tests for IgG were comparable to ELISA, but the other two kits were inferior. Prior to 14 days post symptoms, sensitivity was only on the order of 50-60% by any test. It is clear that serological tests are mostly retrospective in nature. They are useful for diagnosing past infections and viral prevalence in populations, and will be very important in evaluating vaccine durability.

In this last aspect, it would be extremely convenient to have a neutralization assay that does not involve live virus, with its attendant requirement for biosafety level 3 containment. One of the studies is based upon inhibition of binding of the receptor binding domain (RBD) of the S protein to immobilized ACE2 protein (15). The results are read colorimetrically and are near 100% sensitive and specific. An advantage of this test is that it would measure neutralization by IgA, IgM, and IgG all at the same time, increase sensitivity. This type of test will obviously be highly useful as vaccines begin to be widely deployed.

Regarding the sampling procedures, we are still using taking nasal swabs; this is a sensitive method but is prone to be false negatives due to poor sampling. Saliva is clearly far simpler to collect than nasal swabs or blood. How do tests of saliva compare with those of nasal swabs and blood?  For RNA detection by RT-PCR, tests of saliva appear to have at least comparable sensitivity to those of nasal swabs(2-4), representing a considerable simplification of testing. Potentially better is RNA detection in saliva using loop-mediated isothermal amplification (LAMP), which avoids the necessity for thermal cycler instrumentation and for RNA purification. Results appear to be equally sensitive and specific to those from RT-PCR(5). Another isothermal method that is fully automated with less sensitivity, delivers results on a dipstick in 90 minutes(6). What about antibodies in saliva? Saliva is much easier to collect. It appears that antibodies are as detectable in saliva as in serum. It’s been shown that viral antibodies persist in saliva for months(16). Another study showed persistent IgG activity against both N protein and the RBD of the S protein(17). The anti-N antibody showed 100% of sensitivity and the RBD antibody 100% specificity. Thus, a multiplexed assay should be 100% accurate.

We are heading into a period of perhaps explosive increases in rates of infection. Widespread and frequent testing with contact tracing can be a practical way to actively prevent transmission of virus, particularly, in big groups of people (i.e., school settings). The tests described above are playing a role in curbing the pandemic.

 

  1. J. Bullard et al., Predicting infectious SARS-CoV-2 from diagnostic samples. Clin Infect Dis, (2020).
  2. L. Azzi et al., Saliva is a reliable tool to detect SARS-CoV-2. J Infect 81, e45-e50 (2020).
  3. M. Baghizadeh Fini, Oral saliva and COVID-19. Oral Oncol 108, 104821 (2020).
  4. K. K. To et al., Consistent detection of 2019 novel coronavirus in saliva. Clin Infect Dis, (2020).
  5. S. Wei et al., Field-deployable, rapid diagnostic testing of saliva samples for SARS-CoV-2. medRxiv, (2020).
  6. D. A. Collier et al., Point of Care Nucleic Acid Testing for SARS-CoV-2 in Hospitalized Patients: A Clinical Validation Trial and Implementation Study. Cell Rep Med 1, 100062 (2020).
  7. Z. Ali et al., iSCAN: An RT-LAMP-coupled CRISPR-Cas12 module for rapid, sensitive detection of SARS-CoV-2. Virus Res 288, 198129 (2020).
  8. J. Arizti-Sanz et al., Integrated sample inactivation, amplification, and Cas13-based detection of SARS-CoV-2. bioRxiv, (2020).
  9. X. Ding et al., Ultrasensitive and visual detection of SARS-CoV-2 using all-in-one dual CRISPR-Cas12a assay. Nat Commun 11, 4711 (2020).
  10. T. Hou et al., Development and evaluation of a rapid CRISPR-based diagnostic for COVID-19. PLoS Pathog 16, e1008705 (2020).
  11. C. M. Ackerman et al., Massively multiplexed nucleic acid detection with Cas13. Nature 582, 277-282 (2020).
  12. W. N. Chia et al., Serological differentiation between COVID-19 and SARS infections. Emerg Microbes Infect 9, 1497-1505 (2020).
  13. K. W. Ng et al., Preexisting and de novo humoral immunity to SARS-CoV-2 in humans. Science, (2020).
  14. K. Bond et al., Evaluation of Serological Tests for SARS-CoV-2: Implications for Serology Testing in a Low-Prevalence Setting. J Infect Dis 222, 1280-1288 (2020).
  15. C. W. Tan et al., A SARS-CoV-2 surrogate virus neutralization test based on antibody-mediated blockage of ACE2-spike protein-protein interaction. Nat Biotechnol 38, 1073-1078 (2020).
  16. B. Isho et al., Persistence of serum and saliva antibody responses to SARS-CoV-2 spike antigens in COVID-19 patients. Sci Immunol 5, (2020).
  17. N. Pisanic et al., COVID-19 serology at population scale: SARS-CoV-2-specific antibody responses in saliva. J Clin Microbiol, (2020).

GVN Statement on Pfizer and BioNTech Data from COVID-19 Vaccine Study

Baltimore, Maryland, USA, July 23, 2020:  The Global Virus Network (GVN), a coalition comprised of the world’s preeminent human and animal virologists from 57 Centers of Excellence and 10 Affiliates in 33 countries, said today that the results of the Phase 3 study by Pfizer and BioNTech on their mRNA-based vaccine candidate, BNT162b2, represents a major breakthrough in the global effort to develop an effective vaccine against COVID-19.  Pfizer and BioNTech announced earlier today that the Phase 3 study of BNT162b2 has thus far demonstrated 90% efficacy against COVID-19.  The GVN, which has brought together the world’s foremost virologists to collaboratively evaluate the immune response against SARS-CoV-2, congratulates Pfizer and BioNTech for their pioneering work and for achieving this milestone.

As emphasized by Pfizer, the GVN believes an important next step is to evaluate the duration of protection as well as the fine immune response characteristics.  GVN scientists will continue to contribute to these important milestones to curb, and ultimately control, the COVID-19 pandemic.

About the Global Virus Network (GVN)

The Global Virus Network (GVN) is essential and critical in the preparedness, defense and first research response to emerging, exiting and unidentified viruses that pose a clear and present threat to public health, working in close coordination with established national and international institutions. It is a coalition comprised of eminent human and animal virologists from 57 Centers of Excellence and 10 Affiliates in 33 countries worldwide, working collaboratively to train the next generation, advance knowledge about how to identify and diagnose pandemic viruses, mitigate and control how such viruses spread and make us sick, as well as develop drugs, vaccines and treatments to combat them. No single institution in the world has expertise in all viral areas other than the GVN, which brings together the finest medical virologists to leverage their individual expertise and coalesce global teams of specialists on the scientific challenges, issues and problems posed by pandemic viruses. The GVN is a non-profit 501(c)(3) organization. For more information, please visit www.gvn.org. Follow us on Twitter @GlobalVirusNews

Herd Immunity – Can We Get There?

The concept of herd immunity is frequently brought up as an alternative to lockdown and intervention-based strategies for curbing the COVID-19 pandemic, this sensitive issue being much influenced by policy makers. In this context it is important to first define herd immunity. This is obtained when enough of the population has immunity to an infectious pathogen, thus resulting in prevention of its transmission. The clearest examples of achieving herd immunity are provided by successful vaccination programs (i.e., polio, smallpox and measles). The question of whether acquisition of herd immunity to SARS-CoV-2 is possible depends upon important questions. How long will it take to get such herd immunity? What will be the cost in human deaths? Will there be a successful vaccine program? And does infection provide a lasting immunity for further prevention of reinfection?

Let’s first consider the possibility of obtaining herd immunity by vaccination. Ideally, strong herd immunity can be achieved if a vaccine provides a very significant (say around 80-90%) protection (sterilizing immunity) for extended periods, and if most of the population has been vaccinated. However, it is currently uncertain that these conditions will be met. A recent survey conducted in 19 countries showed that approximately, 70% of participants were likely to take a SARS-CoV-2 vaccine(1), indicating that public acceptance will by no means be universal. Assuming such a vaccine were 70% effective, this would leave 50% of the population able to be infected. Interestingly, if a vaccine were government mandated, acceptance dropped to 50 to 60%. Obviously if we are trying to reach herd immunity by vaccination, vaccine efficacy and acceptance will be critical.

Is it possible that natural infection of a large percentage of the population would result in widespread immunity? This could in theory lead to a reduction in further viral spread to the point where it would become relatively insignificant. This depends, however, upon several factors. First, is that a large proportion of the population would need to be infected. Given an estimated infection mortality rate of 0.3 to 0.5%, this would likely result in an unacceptably high number of deaths. Second, how long would it take? Indeed we know that beside some areas such as New York or Paris for example the percentage of infected individuals has apparently remained quite low (10-15%). Third, infection would need to lead to reasonably durable immunity, and we will consider this in more detail.

As to whether infection confers immunity, this is complicated by not knowing the levels of protective immunity to SARS-CoV-2. Inducing neutralizing antibodies have been a major target for development of many vaccines based on the evaluation of immunity to other coronaviruses. Importantly, evaluation of the rate of reinfection can predict a potential acquisition of protective immunity by natural infection. Reinfection is not necessarily proven by the second positive diagnostic to SARS-CoV-2; there is a question of whether the patient has truly become reinfected or the original infecting virus has simply rebounded.

The answer seems to be that reinfection does occur but is quite rare. Indeed, there are only a handful of well documented cases showing reinfection with SARS-CoV-2. In one well characterized instance, a patient was asymptomatically reinfected 4-5 months after a first symptomatic infection(2). Viruses from the first and second infections were phylogenetically distinct, indicating that they were independent infections. Interestingly, although neutralizing and IgG antibodies were not detectable at the onset of the second infection, they rapidly and robustly appeared (3). A handful of other cases have been reported.  However, their relative rarity suggests that reinfection is the exception rather than the rule. What, then, are the potential protective mechanisms and how long do they last?

The short answer is that we don’t have exact answers yet. Much attention has focused on antibody response and durability, particularly on antibodies to the spike protein and neutralizing antibodies (NA), which are a subset of the anti-spike antibodies. Both anti-spike and neutralizing IgG (but not IgM and IgA) antibodies decrease but persist at measurable levels for at least three to six months after the resolution of the infection. Indeed, anti-spike and NA correlate reasonably well with protective immunity ((4, 5) and these two studies). and. Moreover, as mentioned above, priming by memory cells can result in rapid and robust expression of antibodies. One study showed that a better correlation between NAs and spike antibodies could be obtained by measuring NAs and both anti-receptor binding domain (RBD) and S2 domain antibodies(5). However, it is unclear what level of NAs would be protective against SARS-CoV-2 infection.

It is likely that T cell activity will be necessary for significantly durable immunity. T cell activity is more difficult to assess than antibody response, and correspondingly less is known for its role in lasting protective immunity. We should point out that some patients who have recovered from mild COVID-19 and are PCR negative for SARS-CoV-2, show T cell activity against viral peptides even though they were antibody negative(6), suggesting the involvement of cell-mediated immunity. A strong T cell cytotoxic response is evident during disease, and in the convalescent phase, a memory T cell response is common even in the absence of antibodies(7). This suggests that immune responses will be durable, although this is not yet known for SARS-CoV-2.

Many immunologists are still evaluating a lasting immunity to SARS-CoV-2.  No matter what, infecting a substantial majority of the population to reach herd immunity would likely result in a massive death toll. For example, in the U.S., probably one to two million people would die. In addition, we need to consider serious medical and financial consequences for ill individuals with COVID-19. Further, many people who have recovered from the virus report lingering health effects. Earlier in the pandemic, Sweden was pursuing a herd immunity strategy by essentially relaxed containment; yet that strategy has led in June and July 2020 to the highest rate of death in Northern Europe.

Overall containing the pandemics will not be the result of a single intervention; it will combine efficient prevention measures, novel preventive and curative treatments as well as sufficiently effective and safe vaccine. In any case, waiting for spontaneous herd immunity does not appear as a reasonable option for the population.

  1. J. V. Lazarus et al., A global survey of potential acceptance of a COVID-19 vaccine. Nature Medicine, (2020).
  2. K. K. To et al., COVID-19 re-infection by a phylogenetically distinct SARS-coronavirus-2 strain confirmed by whole genome sequencing. Clin Infect Dis, (2020).
  3. K. K. To et al., Serum antibody profile of a patient with COVID-19 reinfection. Clin Infect Dis, (2020).
  4. A. S. Iyer et al., Persistence and decay of human antibody responses to the receptor binding domain of SARS-CoV-2 spike protein in COVID-19 patients. Sci Immunol 5, (2020).
  5. T. J. Ripperger et al., Detection, prevalence, and duration of humoral responses to SARS-CoV-2 under conditions of limited population exposure. medRxiv, (2020).
  6. S. Schwarzkopf et al., Cellular Immunity in COVID-19 Convalescents with PCR-Confirmed Infection but with Undetectable SARS-CoV-2-Specific IgG. Emerg Infect Dis 27, (2020).
  7. T. Sekine et al., Robust T Cell Immunity in Convalescent Individuals with Asymptomatic or Mild COVID-19. Cell, (2020).
  8. N. Le Bert et al., SARS-CoV-2-specific T cell immunity in cases of COVID-19 and SARS, and uninfected controls. Nature 584, 457-462 (2020).

How Long Is A SARS-CoV-2 Infected Person Contagious?

As the COVID-19 pandemic rages on, the global community has become accustomed to comprehensive, interventional strategies such as wearing a mask, social distancing, hand hygiene, and surface cleaning and disinfection. It is well known that transmission of SARS-CoV-2 occurs through direct, indirect, or close contact with infected people through infectious secretions, such as saliva and respiratory secretions or their respiratory droplets. In addition, the scientific community has confirmed that the airborne transmission of SARS-CoV-2 viruses in aerosols (smaller than 100 μm) can remain suspended in air for many seconds to hours and are highly concentrated near an infected person, thus infecting people most easily in close proximity (1). Furthermore, aerosols containing infectious virus can also travel more than 2 m and accumulate in poorly ventilated indoor air, leading to superspreading events. World Health Organization and Centers for Disease Control and Prevention have also acknowledged this under certain circumstances, such as enclosed spaces, prolonged exposure to respiratory particles (i.e., shouting, singing, and exercising), and inadequate ventilation or air (2, 3). Thus, effective control strategies and standardized guidance to the public are integral in mitigating COVID-19. In particular, understanding the duration of infectiousness in infected persons with SARS-CoV-2 is critical for developing evidence-based public health policies on isolation, contact tracing and returning to work. In general, the levels of viral RNA were determined by using quantitative reverse transcription-polymerase chain reaction. However, detection of viral RNA does not necessarily indicate that a person is infectious and able to transmit the virus to another person. Although it is critical to determine the levels of infectious virus particles in infected COVID-19 patients, requirement of Biosafety Level-3 laboratory for the virus titration has hindered this approach. In this perspective section, we will discuss currently available scientific data regarding the levels of infectious virus particles in asymptomatic individuals, in mild and severe COVID-19 patients and in children and young adults.

Asymptomatic and presymptomatic individuals represent a source of potentially transmissible virus (4). Asymptomatic infections have no specific incubation period due to no clinical signs. However, the viral loads detected in asymptomatic populations have been reported in several studies to be similar to those in symptomatic patients. In a nursing facility, quantitative SARS-CoV-2 viral loads detected in residents were similarly high in the four symptom groups (residents with typical symptoms, those with atypical symptoms, those who were presymptomatic, and those who remained asymptomatic). Notably, 17 of 24 specimens (71%) from presymptomatic persons had viable virus by culture 1 to 6 days before the development of symptoms. In a surveillance study, asymptomatic cases of samples were collected through swabbing of contacts or facility/family/household testing in the context of outbreak investigations. Despite the uncertainty of their date of exposure or start of infection, cultivable virus was isolated from samples collected from asymptomatic individuals (41% of tested samples).

Detection of infectious SARS-CoV-2 from upper respiratory tract of mild-to-moderate COVID-19 patients showed that infectious virus can persist for more than a week after symptom onset, declining over time (5). At 10 days after symptom onset, probability of culturing virus declines to 6%. This is in line with current WHO guidance on release from isolation. Similarly, shedding of the virus in mild COVID-19 patients was determined by measuring the levels of transcribed subgenomic mRNA and isolation of infectious viruses (6). Pharyngeal virus shedding was very high during the first week of symptoms, with a peak at 7.1 × 108 RNA copies per throat swab on day 4. In addition, infectious viruses were successfully isolated from these samples, confirming active virus replication in the upper respiratory tract. No viruses were isolated after day 7 onset. These findings suggest efficient transmission of SARS-CoV-2, through active pharyngeal viral shedding at a time at which symptoms are still mild and typical of infections of the upper respiratory tract. In a surveillance study in Manitoba, Canada, the presence of infectious viruses was determined by evaluating samples from day of symptom onset (day 0) up to 21 days after symptom onset (7). Within this range of samples, positive cultures were observed up to day 8 after symptom onset with the probability of obtaining peak titers on day 3. Similarly, in Hong Kong, virus was isolated from the samples of mild patients collected within the first 8 days of illness with median viral RNA load of 7.54 log10 genome copies/mL (8). From severe COVID patients, prolonged duration of cultivable virus was detected for up to 20 days after symptom onset, suggesting that prolonged excretion of infectious virus is associated to the severity of the disease (9).

Severity in most children is limited, and children do not seem to be major drivers of transmission. However, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infects children of all ages. In South Korea, large-scale testing, aggressive contact tracing and testing, and isolation/direct observation of asymptomatic or mildly symptomatic children have identified the presence of asymptomatic (20 of 91 [22%]), presymptomatic (18 of 91 [20%]), and symptomatic children (53 of 91 [58%]) (10). Presymptomatic children remained symptom free for a median (range) of 2.5 (1-25) days before exhibiting any symptoms. A minority of children (6 [7%]) were identified as infected; this highlights the concept that infected children may be more likely to go unnoticed either with or without symptoms and continue on with their usual activities, which may contribute to viral circulation in the community. In a separate study with 12 symptomatic children, infectious viruses were detected at a median of 2 days after symptom onset (11). Median viral RNA load at diagnosis was 3.0 × 106 copies/mL (mean 4.4 × 108 ranging from 6.9 × 103 to 4.4 × 108 copies/mL. A limitation of this study is the small number of children assessed. However, viral load at diagnosis is comparable to that of adults and symptomatic children of all ages shed infectious virus in early acute illness, a prerequisite for further transmission. Considering the relatively low frequency of infected children, even in severely affected areas, biological or other unknown factors could lead to the lower transmission in this population. Large serologic investigations and systematic surveillance for acute respiratory diseases and asymptomatic presentations are still needed to assess the role of children in this pandemic.

Although the spectrum of COVID-19 ranges from asymptomatic to severe infections, most patients experience mild disease (80%). Scientific data indicate that infectious virus in mild patients can persist for a week after symptom onset. Furthermore, infectious virus can be isolated from asymptomatic individuals. This clearly enforces the importance of wearing a mask, quarantine, and contact tracing to mitigate the transmission of SARS-CoV-2. Recent studies support that wearing masks can save lives not only by cutting down the chances of both transmitting and catching the coronavirus (12) but also by reducing the severity of infection in contracted individuals (13). It is well demonstrated that self-quarantine of close contacts exposed to COVID-19 prevents transmission to others. Contact tracing must be conducted for close contacts (any individual within 6 feet of an infected person for at least 15 minutes) of laboratory-confirmed or probable COVID-19 patients. However, relaxed social distancing and opposition of wearing masks are hindering the mitigation of SARS-CoV-2, resulting in continuous increase of COVID-19 cases. Even when the vaccine will be available, we would not immediately stop social distancing, wearing masks, and other interventional measures until reaching efficient levels of viral mitigation. The message is clear that a simple practice of wearing masks can protect ourselves and save other lives from circulating SARS-CoV-2.

 

References

  1. PRATHER, K.A., MARR, L.C., SCHOOLEY, R.T. MCDIARMID, M.A., WILSON, M.E., MILTON, D.K. 2020. Airborne transmission of SARS-CoV-2. DOI: 10.1126/science.abf0521
  2. 2020. Transmission of SARS-CoV-2: implications for infection prevention precautions. https://www.who.int/news-room/commentaries/detail/transmission-of-sars-cov-2-implications-for-infection-prevention-precautions
  3. 2020. Scientific Brief: SARS-CoV-2 and Potential Airborne Transmission. https://www.cdc.gov/coronavirus/2019-ncov/more/scientific-brief-sars-cov-2.html
  4. Arons MM, Hatfield KM, Reddy SC, et al. 2020. Presymptomatic SARS-CoV-2 infections and transmission in a skilled nursing facility. N Engl J Med 382:2081-2090.
  5. Singanayagam Anika, Patel Monika , Charlett Andre , Lopez Bernal Jamie , Saliba Vanessa , Ellis Joanna , Ladhani Shamez , Zambon Maria , Gopal Robin. 2020. Duration of infectiousness and correlation with RT-PCR cycle threshold values in cases of COVID-19, England, January to May 2020. Euro Surveill. 25.
  6. Wölfel R, Corman  VM, Guggemos  W, Seilmaier  M, Zange  S, Müller  MA, et al. 2020. Virological assessment of hospitalized patients with COVID-2019. Nature. 581:465–469.
  7. Bullard J, Dust K, Funk D, Strong JE, Alexander D, Garnett L, et al. 2020. Predicting infectious SARS-CoV-2 from diagnostic samples. Clin Infect Dis. ciaa638.
  8. Perera RAPM, Tso E, Tsang OTY, Tsang DNC, Fung K, Leung YWY, et al. SARS-CoV-2 virus culture and subgenomic RNA for respiratory specimens from patients with mild coronavirus disease. Emerg Infect Dis. 2020;26(11).
  9. van Kampen JJA, van de Vijver DAMC, Fraaij PLA, Haagmans BL, Lamers MM, Okba N, et al. Shedding of infectious virus in hospitalized patients with coronavirus disease-2019 (COVID-19): duration and key determinants. medRxiv. 2020.06.08.20125310.
  10. Han MS, Choi EH, Chang  SH,  et al.  Clinical characteristics and viral RNA detection in children with coronavirus disease 2019 in the Republic of Korea. JAMA Pediatr. doi:10.1001/jamapediatrics.2020.3988.
  11. L’Huillier AG, Torriani G, Pigny F, Kaiser L, Eckerle I. Culture-Competent SARS-CoV-2 in Nasopharynx of Symptomatic Neonates, Children, and Adolescents. Emerg Infect Dis. 2020 Oct;26(10):2494-2497. doi: 10.3201/eid2610.202403. Epub 2020 Jun 30. PMID: 32603290; PMCID: PMC7510703.
  12. Leffler, C. T. et al.Preprint at medRxiv https://doi.org/10.1101/2020.05.22.20109231 (2020).
  13. Gandhi, M., Beyrer, C. & Goosby, E. Gen. Intern. Med. https://doi.org/10.1007/s11606-020-06067-8 (2020).

Effect of Suppressed Innate Immunity on Covid-19 Severity

One of the important determinants of severe Covid-19 appears to be an inappropriate response by the innate immune system. Specifically, this involves, on the one hand, insufficient or delayed expression and signaling by type 1 interferons (IFNs), which induce innate cell-mediated immunity. The interferon (IFN) response constitutes the major first line of defense against viruses. On the other hand, this involves an overly active inflammatory cytokine response, with its attendant tissue damage (this “cytokine storm” was explored in our previous Perspective at https://gvn.org/category/sars-cov-2/gvn-sars-cov-2-perspectives/). Let’s look at some of the lines of evidence for these contradictory events.

One way in which an aberrant immune response to SARS-CoV-2 is reflected at the cellular level is by a greatly increased ratio of neutrophils to lymphocytes(1). This is correlated with low expression of type I and III interferons and high expression of pro-inflammatory factors including IL-6 and a variety of chemokines(2) that act as attractants for neutrophils and monocyte/macrophages. These activities are the outcome of interactions between host factors that recognize pathogen associated molecular patterns (PAMPs), such as viral RNA in endosomes, and viral proteins that are antagonistic to these factors and their signaling pathways. Genetic differences in host factors can result in profound differences in host responses to pathogens (recent findings are described below). Different viruses also tend to have different or unique antagonists to host immune factors that can greatly influence the outcome of infection.

What are some of the molecular studies that point to defects in interferon activity in severe Covid-19? Recently, interest has been increasing in toll-like receptors (TLRs), especially TLR3 and TLR7, which recognize viral RNA and are important in interferon type I and inflammatory cytokine expression. TLRs play a key role in the recognition of PAMPs and trigger the activation of specific signaling pathways, thereby inducing the transcription of inflammatory and/or anti-inflammatory cytokine. One interesting report looked at two sets of two brothers(3) who, although young and otherwise healthy, had severe Covid-19 (one died). Whole exome sequencing revealed that both sets of brothers had mutations in TLR7, which serves as a sensor for viral RNA. One set had a missense mutation predicted to result in an inactive TLR7, while the other set had a frame shifting 4 nucleotide deletion, resulting in a nonsense protein. Stimulation of primary immune cells in vitro with the TLR agonist imiquimod resulted in defective expression of type I interferon-related genes normally regulated by TLR7. While the limited nature of the study does not permit a conclusion of causality, several factors make it likely that these loss of function mutations are significant. Exhibition of severe disease in young men is rare. Despite rare cases of loss of function mutations in TLR7, two different loss of function mutations in two young brother pairs with severe disease indicate its potential role in Covid-19 severity. It should be pointed out that TLR7 is located on the X chromosome, so a single mutant copy would cause loss of function. One of the mothers was heterozygous for wild type TLR7, making her a carrier. Thus, if problems with the TLR7 pathway exacerbate Covid-19, males might be likelier to have an insufficiency.

Another study analyzed the complete genomes or exomes of 659 patients with life threatening Covid-19 and compared them with those of 534 people with asymptomatic or benign infections(4). They characterized 13 genetic loci encoding factors in the TLR3-interferon regulatory factor 7 (IRF7) pathway, which also regulates type I interferon production and immunity to influenza virus. They found that 3.5% of the people with life threatening Covid-19 had loss of function variants at these loci. Moreover, when immune cells from patients with these variants were tested in vitro, they were found to be defective in type-I interferon immune activities, and further in vivo study confirmed impaired production of type I IFN during the course of SARS-CoV-2 infection. About half of these patients also had extremely low levels of serum interferon α, a type I interferon.

Yet, another study further implicates lack of appropriate interferon activity in severe Covid-19(5). In this study, 101 of 987 patients with life threatening Covid-19 had auto-antibodies against interferon α, interferon ω or both. These were not present in 663 patients with mild disease. The auto-antibodies were able to neutralize the antiviral effects of interferon in vitro (and likely in vivo). Interestingly, the auto-antibodies were about 5-fold more prevalent in men than in women.

None of these studies by themselves show a specific defect in interferon activity in a majority of cases. However, taken together, they certainly suggest that a great variety of different defects related to the antiviral activities of type 1 interferons may be surprisingly common. Probably, one of the factors explains why some people resist serious disease while, for others, it is life-threatening. Further, investigations in this area will be most interesting.

One other study identified a 3p21.31 gene cluster that conferred a risk of severe Covid-19. This region contains three chemokine receptor genes, all involved in innate immunity(6). It turns out to be a region derived from Neanderthals, and is present at variable incidence worldwide except in Africa. This provides yet another clue that the innate immune response to SARS-CoV-2 may be an important determinant of whether an infected individual will, or will not, develop critical Covid-19.

In this light, it has been proposed that some cross-protection could be afforded by administering live attenuated vaccines, such as measles-mumps-rubella, and oral polio vaccine(7) (non-specific effect of vaccination against SARS-CoV-2). The stimulation of innate immunity by these vaccines could provide temporary protection against Covid-19. If proven to be effective against Covid-19, emergency immunization with these vaccines could be used for protection against other unrelated emerging pathogens.

 

References

  1. Y. Liu et al., Neutrophil-to-lymphocyte ratio as an independent risk factor for mortality in hospitalized patients with COVID-19. J Infect 81, e6-e12 (2020).
  2. D. Blanco-Melo et al., Imbalanced Host Response to SARS-CoV-2 Drives Development of COVID-19. Cell 181, 1036-1045 e1039 (2020).
  3. C. I. van der Made et al., Presence of Genetic Variants Among Young Men With Severe COVID-19. JAMA 324, 663-673 (2020).
  4. Q. Zhang et al., Inborn errors of type I IFN immunity in patients with life-threatening COVID-19. Science, (2020).
  5. P. Bastard et al., Auto-antibodies against type I IFNs in patients with life-threatening COVID-19. Science, (2020).
  6. D. Ellinghaus et al., Genomewide Association Study of Severe Covid-19 with Respiratory Failure. N Engl J Med, (2020).
  7. K. Chumakov, C. S. Benn, P. Aaby, S. Kottilil, R. Gallo, Can existing live vaccines prevent COVID-19? Science 368, 1187-1188 (2020).

Transmission Dynamics of SARS-CoV-2: Superspreaders and Superspreading Events

The concept of Superspreaders and Superspreading events has recently attracted a lot of attention. In fact it is important to understand that most of SARS-CoV-2 infected persons are in fact not contagious! Thus, transmission is really dependent on a handful of individuals we call Superspreaders who nurture Superspreading events. Let’s distinguish between Superspreaders and Superspreading events. Superspreaders are individuals who infect a high number of persons; why?  This is not clear, and this is a most important issue to clarify in the future.  We know they yield high viral load and that they are generally, but not always, young people.  But, this cannot fully explain their massive contamination impact.  Moreover, they are frequently asymptomatic, thereby significantly increasing risk of dissemination. Superspreading events, which involve at least one Superspreader, are events which favor large scale transmission, such as close contacts in indoor situations. Secondary transmissions from infected people then result in a large number of further infections, and so on. Thus, by some estimations  only 20% of infected individuals cause 80% of infections(1).  Identification of Superspreading events depends upon contact tracing.  Furthermore, DNA sequencing of viral genomes adds a great deal to a clearer understanding of these phenomena, thus, confirming the substantial role of Superspreaders in the pandemic. Let’s look at some well-characterized superspreader events to try to better understand how the majority of SARS-CoV-2 infections occur, enabling us to gain an understanding of what might be done to prevent them.

One of the early recognized superspreading events occurred in mid-March in Skagit County, Washington(2) at a 2.5 hr-long choir practice in which 61 people sang in close proximity. Probable or confirmed infections occurred in 87% of the attendees. Given the low incidence of COVID-19 at the time of this event, it is likely that all the infections originated with a single individual, and that the act of singing vigorously launched many viral laden particles into the air. People were relatively closely positioned. Thus, it is likely that superspreading occurs in an unusually favorable environment. Looking at it another way, the virus got “lucky.”  Another study, unrelated to the choir event, analyzed the sequences of 453 viral genomes collected between February and March in Washington(3).  It is possible to infer the likelihood of how many people have been infected by a single person using viral genomic epidemiology, especially given the relative genetic stability of SARS-CoV-2. A phylogenetic analysis strongly suggested that 84% of the 453 viral genomes derived from a single introduction sometime in early February.  In the choir study, it is clear how virus was transmitted. In the genomic study, it is only clear that a single infected individual somehow infected a great number of people through primary, secondary, and other less direct routes.

Another well-studied superspreader event began with a single infected individual in a meat packing plant is Postville, Iowa. In this case, viral spread could be ascertained by both contact tracing and by genomic epidemiology. Fourteen independent viral introductions were identified in the region, but the only virus to spread widely was the one from the meat packing plant. The virus from this individual passed first to numerous other workers, then to family members, then to the community in Postville (87 cases), and finally to other locations in an area of 185 square miles in Iowa, Wisconsin and Minnesota. These conclusions were supported by genomic sequences from 27 different infected individuals. Again, it appears that prolonged close contact indoors facilitated transmission and suggests that this is a critical feature of superspreading events.

Perhaps, the best characterized superspreading event, originating from an international business meeting of Biogen in Boston(4) in which more than 90 individuals became infected.  This event recently received considerable attention. The large number of infections was indicative of a possible superspreading event. A recent study looked at this event and its consequences in detail, using genomic epidemiology(5). They were able to identify and track the virus in question by a single nucleotide polymorphism (SNP), C2416T. Among 80 separate introductions from four continents into the Boston area early in the pandemic, which they inferred from phylogenetic analyses, the C2416 SNP was unique to one virus. Comparing other viral genomes from various parts of the world with C2416T, the parental origin appears to be in Europe, perhaps France, with an estimated most recent common ancestor existing about two weeks prior to the conference aroundFebruary 26-27. Of all samples collected prior to March 10, the only instances of C2416T were from people who had attended the conference, indicating that a superspreading event had indeed occurred there.

Subsequent samples (744) from infected individuals in Boston and surrounding areas were collected over a period from February to June and genomes were sequenced. Remarkably, 35% of the samples had the C2416 SNP. Since no sample prior the March 10 had this SNP, it suggests that the superspreading event at the Biogen meeting February 26-27 resulted in virus from a single individual infecting more than a third of all infected people in the Boston area. Percentages of the C2416T SNP in regions around Boston ranged from 30-46% in four adjacent counties. In addition, a second SNP, G26233T, appears to have emerged during the event, enabling further tracking. Data from this SNP shows a likely export to other states and countries, with further community spread in Virginia, Michigan and Australia. Some caution is warranted, however, since genomic sampling is not generally done on a randomized basis.

The same report looked at infection clusters at homeless shelters, nursing facilities, and a hospital to gain a better understanding of transmission dynamics. They analyzed 193 viral genomes collected from the Boston Health Care for the Homeless Program and identified 4 clusters of 20 or more highly similar genomes, including two clusters containing the C2416T SNP. They also investigated a superspreading event at a skilled nursing facility, in which 82/97 (84%) residents and 36/97 (37%) of staff were infected. In fact, 75% of viral genomes from different individuals had highly similar genomes, suggesting that they arose from a single recent introduction. This took place even though strict interventional measures were in place. Interestingly, two other clusters of three closely related genomes were detected. This represents independent introductions, but these failed to massively spread. In the case of two clusters of infection at Massachusetts General Hospital, highly similar genomes were not found, suggesting a lack of significant in-hospital spread.

What is our conclusion? First, it is now clear that a majority of transmissions result from superspreading events, facilitated by conducive conditions. These include indoor location, close contact, lengthy contact, indoor activity such as singing or talking, poor air ventilation, and lack of mitigation procedures (i.e., wearing masks and physical distancing). Tracing Superspreading events in Hong Kong also confirmed that the largest cluster (106 cases) was traced to four bars followed by a wedding (22 cases) and attendance at a temple (19 cases) (1). This study suggests that disease control efforts should focus on avoiding gathering events and mitigating their impact.  The rapid tracing and quarantine of confirmed contacts, along with the implementation of physical distancing policies including either closures or reduced capacity measures targeting high-risk social settings such as bars, weddings, religious sites and restaurants, should be efficient to prevent the occurrence of superspreading events. Overall, the issue of Superspreaders and Superspreading events illustrates the impact of molecular epidemiology for deciphering the patterns of COVID-19 dissemination. What we still clearly lack, however, is the understanding of the very early phases of the pandemics in China. This would be very useful for the whole appraisal of transmission dynamics.

  1. D. C. Adam et al., Clustering and superspreading potential of SARS-CoV-2 infections in Hong Kong. Nat Med, (2020).
  2. L. Hamner et al., High SARS-CoV-2 Attack Rate Following Exposure at a Choir Practice – Skagit County, Washington, March 2020. MMWR Morb Mortal Wkly Rep 69, 606-610 (2020).
  3. T. Bedford et al., Cryptic transmission of SARS-CoV-2 in Washington state. Science, (2020).
  4. A. Schuchat, C. C.-R. Team, Public Health Response to the Initiation and Spread of Pandemic COVID-19 in the United States, February 24-April 21, 2020. MMWR Morb Mortal Wkly Rep 69, 551-556 (2020).
  5. J. Lemieux et al., Phylogenetic analysis of SARS-CoV-2 in the Boston area highlights the role of recurrent importation and superspreading events. medRxiv, (2020).

GVN 2020 Special Annual Meeting Executive Summary

A New Era in the Fight Against COVID-19 Pandemic: Forging a “Viral Pandemic Readiness Alliance”

A September 22-23, 2020 Special Meeting of Top Global Experts Launches “Global Virus Network’s Vision for Future Pandemic Preparedness”

We are in the midst of a pandemic that has completely upended the world with major economic, social and psychological impacts. The major threat to public health is not only connected to COVID-19-related mortalities, but also to associated morbidity and, possibly, sequelae; moreover COVID-19 impacts overall population health due to the disorganization of health systems.

We must be very humble, as we cannot predict what the future could hold: seasonal variations? Long term persistence? Or regression? We need to keep in mind that the reason behind the SARS-CoV-1 epidemic’s regression has remained in part mysterious. In fact, eradication of the virus seems impossible, and herd immunity may be very difficult to achieve. Thus, we must learn to live with the virus.

It is increasingly clearer that we are not facing “another health crisis.” We are entering a new era where novel modes of organization must be designed. We cannot wait for the current crisis to conclude to prepare for the next—we must act now!

Despite significant progress in global health following previous epidemics and pandemics (including HIV, Influenza and Ebola), and although we were aware of the potential risk of such new pandemics, we were not sufficiently prepared. There are two immediate consequences for global health policies:

  • The importance of infectious diseases, global and “one health” are only further emphasized.
  • The divergence between politics and health in many countries has led to disastrous decisions. As such, we need to provide governing leaders with science-driven and independent strategies.

The Global Virus Network (GVN) is poised to be an important partner in achieving these objectives. This is a coalition of the foremost virologists worldwide, representing 57 research centers and 10 affiliates in 33 countries, and growing by the day. The GVN coordinates scientific projects and has organized task forces on specific viruses including Zika, Chikungunya, and HTLV-1, and now SARS-CoV-2. The organization also has a major focus on education, training and mentoring others in the field. Globally, there is a lack of critical mass in scientists, medical doctors and public health professionals working on infectious diseases. The GVN plays a significant role in advocacy and providing statements (in particular through its website https://gvn.org/). Science-driven and independent expertise are key drivers of meaningful public health strategies, and through its network of outstanding virologists worldwide, GVN offers national and international institutions, as well as industrial partners, a unique source of information and recommendations.

In this context, the GVN organized a two-day workshop dedicated to COVID-19 and future pandemic preparedness with the aim to evaluate what has been improperly and properly handled during these first eight months of the COVID-19 pandemic spreading. The workshop looked at precisely identifying the challenges ahead, the actions to take and how the GVN can collaborate with the many institutions to meet these needs. The goal of the workshop was not to revisit in detail all topics and known facts.  A video of the full post-meeting press conference, can be found here.

The following summarizes the major issues discussed:

1) Preparedness: We were not prepared, and we need to prepare now; This implies novel organizational modalities.

  • Cooperation and coordination, beyond goodwill and fashionable wordings; too many institutions are still working in silos with self-interest strategies.
  • Leverage technologic innovation and scientific progress to produce diagnostics, vaccines, and therapeutics.
  • Contemplate and implement novel modes of interactions between academics and industrials, and such partnerships have been at the heart of the GVN since its inception.
  • Multi- and transdisciplinary collaborations, including social and behavioral sciences, and perception of communication.
  • International collaborations: one country alone cannot solve the problem. While this seems obvious, most countries have reacted on an isolated basis. A global collaboration network for pandemic preparedness and prevention needs to be implemented immediately.

2) Prediction: Humans are the best sentinels. Is it feasible to predict future pandemics? How to sufficiently organize surveillance?

  • We must recognize that we cannot predict future pandemics, though we can improve our strategies. Yet, we have sufficient technologies and data analysis systems (including artificial intelligence), but we need to establish implementation and global data sharing mechanisms.
  • We know that animal viruses are major risk factors for the next epidemics and pandemics. This is even increasingly at stake. During the meeting scientists emphasized that five of the seven human coronaviruses identified (229E, NL63, OC43, HKU1, SARS-CoV-1, MERS-CoV and SARS-CoV-2) in the last 20 years have emerged from bats. Humans are modifying ecosystems and are in fact accelerating transmission events.
  • Comprehensive sequencing-based analysis of all viruses worldwide (“animal viromes”) provides useful knowledge but does not predict transmission to humans. GVN scientists point to the importance of focusing surveillance efforts to the human populations who interface with animals.

3) Origin: There is no scientific evidence that SARS-CoV-2 was disseminated by human manipulation.

  • GVN scientists all concur on this controversy.
  • The mission to find out the origins of the virus was a true international collaboration and transparent process featuring scientists from China, America, Australia, Japan, France, and the Philippines.
  • There is a 1,200-nucleotide difference between the closest backbone virus and SARS-CoV-2, representing 4 to the power of 1,200 possible combinations. Even if someone had unlimited research funding and all the best virologists in the world, no one could make this virus.
  • An extensive study will be conducted starting in China and through Southeast Asia to identify the origins of the virus and to allow much better surveillance and mitigation for future emerging viruses.

4) Transmission: “Super spreaders” and “super spreading” events are major drivers of pandemics.

  • COVID-19 is a highly contagious respiratory disease with very low mortality directly induced by the virus, thus the ideal condition for a virus to spread. The importance of masks, physical distancing and handwashing is well-known. GVN scientists also emphasize the importance of research on disinfectants, an underappreciated protective measure.
  • As re-emphasized in this workshop, only a handful of those infected seem highly contagious. Thus, transmission is driven by a limited number of individuals who behave as “super spreaders.” Why do some individuals (a.k.a. “super spreaders”) transmit viruses to so many others? Although we know that such individuals show high viral load and are generally, yet not always, younger, this cannot fully account for this spreading. What are the other factors? Thus far, research focused on such individuals is mandatory. The question remains whether we can identify novel biomarkers, though we would need to fully exclude stigmatization.
  • Also, this implies for obvious statistical reasons that large gatherings are major risk factors for being in contact with such rare “super spreaders” and thus contributors to rapid viral wide spreading. Therefore, we should not only speak of “super spreaders” but also of “super spreading” events.
  • Aerosol-related transmission is still a controversial issue. Yet, GVN scientists have emphasized that the impact of short-range aerosol-driven transmission contributes to the dissemination of the virus, particularly in the context of “super spreading” events. Masks are very efficient against large droplets but are unfortunately less efficient against such aerosols.

5) Diagnostic: Efficient and rapid diagnostic testing is the key for controlling an infectious disease, and we have not benefited enough from the huge technology progress in this area.

  • Nothing is needed more than rapid diagnostic tests. We need to trace and follow infected individuals and their contacts. We need to educate the general public. This is absolutely the foundation, and we cannot do anything without it.
  • There is now ample evidence that salivary sampling can be used instead of nasal swabs in both symptomatic and asymptomatic infected individuals. This can overhaul access to testing, in particular but not only in children. Rapid tests, whether molecular or immune-based, are now available at a low cost, and presentations have been made by GVN scientists demonstrating these points. Point-of-care rapid tests should also be available.
  • Important progress has been made regarding serological assays, offering major insights not only on the epidemiology but also defining the neutralization capacity of detected antibodies as novel correlates for protection. These are fully necessary for evaluating protective measures, novel therapies and vaccines. As an example, some presentations showed that the nature of the antibodies to SARS-CoV-2 significantly differs when comparing children and elderly, possibly accounting for variations in disease severity. Yet, we need standardized protocols for neutralizing assays. Also, the protective efficacy of antibodies needs to be further substantiated. GVN scientists have emphasized the need to get access to the cellular immune response for delineating such correlates of protection.
  • Discussions have been focused on how we should provide novel organizational schemes to favor rapid translation from technology-driven research to routine testing, and partnerships between academic and industrial partners should be reinforced in an international context. Institutions such as the Coalition for Epidemic Preparedness and Innovations created for vaccine development are interesting models to get such novel consortia moving faster.

6) Therapeutics: Despite a huge effort made on drug repurposing so far, we have achieved limited results.

  • Drug repurposing must continue to be at the heart of the therapeutic strategy, providing immediate access of well characterized molecules and allowing massive screening for antiviral activities. However, we do not yet have access to drugs that can prevent transmission in high-risk groups or treat early infections. In fact, we are left with combining steroids, Remdesivir (with some but limited efficacy) and anticoagulants for severe infections with pneumonia. Though, several ongoing studies offer hope for novel prophylactic and early treatment molecules.
  • In this context, GVN scientists have emphasized the need for research agencies to fund not only drug repurposing but also drug discovery. Drug discovery will take time to lead to novel accessible molecules – this is a long battle and not a single crisis.
  • Several presentations demonstrated the potential of novel therapeutic avenues, from immunomodulatory to direct antiviral approaches. Antivirals are only meaningful in the early phase of the infection.
  • The trend will be to use drugs targeting multiple pathways and to combine antivirals and immunomodulatory molecules. Additionally, GVN scientist are addressing the possibility of developing broad spectrum antivirals, which could be effective against coronaviruses, influenza and filoviruses (involved in hemorrhagic fevers such as Ebola, Zika etc.).

7) Vaccines: Safety, efficacy and durability are predominant concerns of COVID-19 vaccine development. Nonspecific immunization procedures must be considered along with COVID-19-specific vaccines.

  • Enormous parallel efforts are being made worldwide utilizing innovative approaches to shorten the vaccine development time. There is uncertainty as to when vaccines for COVID-19 will be readily available for mass vaccination and which formula will be the most efficient. Importantly, we need to ensure the safety of vaccines by testing proper animal models and complying with regulatory requirements – we simply cannot incur adverse reactions.
  • We also need second-generation vaccines that are more focused on the cell immune response.
  • Stimulation of the Innate immune response by non-specific immunization, for example: Bacille Calmette-Guérin (BCG), Oral Polio Virus, is extremely important. GVN scientists made important presentations on this topic, illustrating how BCG-based strategies have already allowed in different contexts to decrease the neonates’ overall mortality in Africa and the rate of respiratory infections in elderly. Mechanisms accounting for stimulation of innate immune response in COVID-19 were thoroughly discussed, and ongoing trials on the impact of BCG and Oral Polio Virus-based vaccines on COVID-19 were deliberated. This approach is complementary to specific vaccine development and might offer a bridge before getting an efficient and sufficiently characterized vaccine.

Conclusions:

It is not a crisis – it is a new era. We have major challenges ahead.  We need a new organization and we need it now.  This is where the GVN is very important, and complementary to national and international agencies. This workshop has led GVN to forge a unified and multidisciplinary pandemic response strategy, tentatively named the Viral Pandemic Readiness Alliance (VPRA) by collaborations with university, industry, government and communities to merge the efforts and find solutions together.

  • True international collaborations are essential and go beyond good and fashionable wordings. Global, One Health and VPRA strategy can support future pandemic preparedness with distribution of diagnostics, vaccine and therapeutics and other interventional measures.
  • In a surge of COVID-19 publications and news releases, we need reliable channels for dissemination of scientific knowledge and information sharing. GVN and VPRA can contribute to this global collaboration effort by assisting the UN, WHO, CEPI, Wellcome Trust, the Bill and Melinda Gates Foundation, and other organizations to serve this purpose.

 

GVN 2020 Meeting Press Release

GVN International Press Conference September 24, 2020