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Scripps Research Institute: SARS-CoV-2 and the D614G Mutation

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  • Scripps Research Institute: SARS-CoV-2 and the D614G Mutation

    Scripps Research Institute: SARS-CoV-2 and the D614G Mutation




    #15,320

    All viruses mutate over time, and for the most part, these mutations do little or nothing to affect the function or biological fitness of the virus. It's part of the evolutionary process, and despite Hollywood tropes, mutations are not necessarily ominous or bad.

    Single-stranded RNA viruses - like influenza and coronaviruses - are particularly prone to `duplication errors' during replication, and are constantly introducing mutations (see Mechanisms of Viral Mutation).


    Most of these mutations end up being of little consequence, and do nothing to affect the transmissibility, replication, host range, or virulence of the virus. Some even prove detrimental, making the virus less `fit' than its predecessors, or attenuating its virulence.

    But the reality is, we are in a pandemic because a (likely) bat-borne coronavirus hit the jackpot with the `right' combination of mutations to allow it to jump species, much in the way that a swine-origin H1N1 virus jumped to humans in 2009 to spark the last pandemic.


    And as we saw with H1N1pdm, once that virus began to circulate in humans, a small number of worrisome mutations began to appear (see EID Journal: Emergence of D225G Variant A/H1N1, 2013–14 Flu Season, Florida) that affected its virulence.

    Luckily, none of the mutations became `fixed', and so they were relatively rare events.


    Over the past couple of months we've been watching a curious, and concerning, difference in the spread of COVID-19 in Europe, the United States and South America when compared to China, Vietnam, and other Asian countries.

    This has led to speculation that a more transmissible `European' strain had emerged, and was then spread to North and South America.


    While many scientists (and public officials) have downplayed the idea, we've seen a number of pre-print studies (see More COVID-19 (SARS-CoV-2) Mutation Reports) that identified a specific mutation (D614G) in the spike protein that appears to enhance its transmissibility.

    Among them, this one from the Los Alamos National Laboratory, which describes a new, now dominant `European' strain of COVID-19.
    B Korber, WM Fischer, S Gnanakaran, H Yoon, J Theiler, W Abfalterer, B Foley, EE Giorgi, T Bhattacharya, MD Parker, DG Partridge, CM Evans, TI de Silva, on behalf of the Sheffield COVID-19 Genomics Group, CC LaBranche, DC Montefiori


    Another study from the University College London, called Emergence of genomic diversity and recurrent mutations in SARS-CoV-2, `. . . . identified close to 200 recurrent genetic mutations in the virus, highlighting how it may be adapting and evolving to its human hosts.'

    Yesterday the Scripps Research Institute published a press release on recent findings that seem to support the notion of a `more transmissible' strain of SARS-CoV-2 - originating in Europe - and now also dominant in North and South America.

    Their study - still under peer review - can be viewed at:
    Lizhou Zhang1#, Cody B Jackson1#, Huihui Mou1#, Amrita Ojha1, Erumbi S Rangarajan2, Tina Izard2, Michael Farzan1*, Hyeryun Choe1*


    While this study adds further weight to the idea of a `more transmissible strain' SARS-CoV-2, more work will be needed to confirm the theory. The press release follows:
    COVID-19-causing viral variant taking over in the United States and Europe now carries more functional, cell-binding spikes.
    June 12, 2020
    Additional Resources
    JUPITER, FL — A tiny genetic mutation in the SARS coronavirus 2 variant circulating throughout Europe and the United States significantly increases the virus’ ability to infect cells, lab experiments performed at Scripps Research show.
    “Viruses with this mutation were much more infectious than those without the mutation in the cell culture system we used,” says Scripps
    Research virologist
    Hyeryun Choe, PhD, senior author of the study.

    The mutation had the effect of markedly increasing the number of functional spikes on the viral surface, she adds. Those spikes are what allow the virus to bind to and infect cells.

    “The number—or density—of functional spikes on the virus is 4 or 5 times greater due to this mutation,” Choe says.
    The spikes give the coronavirus its crown-like appearance and enable it to latch onto target cell receptors called ACE2. The mutation, called D614G, provides greater flexibility to the spike’s “backbone,” explains co-author Michael Farzan, PhD, co-chairman of the Scripps Research Department of Immunology and Microbiology.

    More flexible spikes allow newly made viral particles to navigate the journey from producer cell to target cell fully intact, with less tendency to fall apart prematurely, he explains.

    “Our data are very clear, the virus becomes much more stable with the mutation,” Choe says.

    There has been much debate about why COVID-19 outbreaks in Italy and New York have so quickly overwhelmed health systems, while early outbreaks in places like San Francisco and Washington state proved more readily managed, at least initially. Was it something about those communities and their response, or had the virus somehow changed?

    All viruses acquire minute genetic changes as they reproduce and spread. Those changes rarely impact fitness or ability to compete. The SARS-CoV-2 variant that circulated in the earliest regional outbreaks lacked the D614G mutation now dominating in much of the world.

    But was that because of the so-called “founder effect,” seen when a small number of variants fan out into a wide population, by chance? Choe and Farzan believe their biochemical experiments settle the question.

    There have been at least a dozen scientific papers talking about the predominance of this mutation,” Farzan says. “Are we just seeing a ‘founder effect?’ Our data nails it. It is not the founder effect.”

    Choe and Farzan’s paper is titled “The D614G mutation in the SARS-CoV-2 spike protein reduces S1 shedding and increases infectivity.” Now undergoing peer review, it is being posted prior to publication to the pre-print site bioRxiv, and released early, amid news reports of its findings.

    Choe and Farzan note that their research was performed using harmless viruses engineered to produce key coronavirus proteins. Whether the changes they observed also translate to increased transmissibility in the real world requires additional epidemiological studies, they note.

    Encouragingly, the duo found that immune factors from the serum of infected people work equally well against engineered viruses both with and without the D614G mutation. That’s a hopeful sign that vaccine candidates in development will work against variants with or without that mutation, Choe says.

    Choe and Farzan have studied coronaviruses for nearly 20 years, since the first outbreak of SARS, a similar virus. They were the first to discover in 2003 that SARS bound to the ACE2 receptor on cells. Others’ experiments have shown the SARS-CoV-2 virus binds the same ACE2 receptor.

    But Farzan and Choe note a key structural difference between spike proteins on the first SARS virus and this new pandemic strain. With both, under an electron microscope, the spike has tripod shape, with its three segments bound together at a backbone-like scaffold. But SARS-CoV-2 is different. Its tripod is divided in two discreet segments, S1 and S2.

    Initially, this unusual feature produced unstable spikes, Farzan says. Only about a quarter of the hundreds of spikes on each SARS-CoV-2 virus maintain the structure they need to successfully infect a target cell. With the mutation, the tripod breaks much less frequently, meaning more of its spikes are fully functional, he says.

    The addition of the D614G mutation means that the amino acid at that location is switched from aspartic acid to glycine. That renders it more bendable, Farzan says.

    Evidence of its success can be seen in the sequenced strains that scientists globally are contributing to databases including GenBank, the duo reports. In February, no sequences deposited to the GenBank database showed the D614G mutation. But by March, it appeared in 1 out of 4 samples. In May, it appeared in 70 percent of samples, Farzan says.

    Over time, it has figured out how to hold on better and not fall apart until it needs to,” Farzan says. “The virus has, under selection pressure, made itself more stable.”

    It is still unknown whether this small mutation affects the severity of symptoms of infected people, or increases mortality, the scientists say. While ICU data from New York and elsewhere reports a preponderance of the new D614G variant, much more data, ideally under controlled studies, are needed, Choe says.

    In addition to senior authors Choe and Farzan, the authors include first authors Lizhou Zhang, Cody Jackson and Huihui Mou, plus co-authors Amrita Ojha, Erumbi Rangarajan and Tina Izard, all of Scripps Research.

    The work was supported by the National Institutes of Health through an administrative supplement to RO1 AI129868 for coronavirus research.

    https://afludiary.blogspot.com/2020/...ars-cov-2.html
    All medical discussions are for educational purposes. I am not a doctor, just a retired paramedic. Nothing I post should be construed as specific medical advice. If you have a medical problem, see your physician.

  • #3
    I do not think S D614G is of any importance. It is hard to be sure but I have been watching the sequences and its spread seems to be due to random chance. D614G occurred right at the the beginning of the outbreak and was holding its own in China but it was one case from the G branch that first showed up in Germany and went on to dominate in Europe and then seed the US East Coast. The West Coast saw a separate introduction from China this time of the D strain and both lineages now co exist in the US without either displacing the other. The picture is further muddied by wildly different levels of sequence generation from samples between countries, for example Switzerland has submitted about the same number of full sequences as Spain and Italy combined and Europe, as a whole, have generated far more sequences per case than others. Over all both strains are holding their own. ORF1a L3606F is much more interesting as it keeps occurring, and then holding its own, and is showing tentative signs of host adaption, much less sexy though as it codes for a non structural protein which suppresses the intra-cellular immune response not the Spike whose function - and importance - is much easier to grasp.

    Comment


    • #4
      Source: https://www.sciencedaily.com/release...0702144054.htm

      Newer variant of COVID-19-causing virus dominates global infections
      Virus with D614G change in Spike out-competes original strain, but may not make patients sicker

      Date:
      July 2, 2020
      Source:
      DOE/Los Alamos National Laboratory
      Summary:
      New research shows that a specific change in the SARS-CoV-2 coronavirus virus genome, previously associated with increased viral transmission and the spread of COVID-19, is more infectious in cell culture.

      Research out today in the journal Cell shows that a specific change in the SARS-CoV-2 coronavirus virus genome, previously associated with increased viral transmission and the spread of COVID-19, is more infectious in cell culture. The variant in question, D614G, makes a small but effective change in the virus's 'Spike' protein, which the virus uses to enter human cells.

      Bette Korber, a theoretical biologist at Los Alamos National Laboratory and lead author of the study, noted, "The D614G variant first came to our attention in early April, as we had observed a strikingly repetitive pattern. All over the world, even when local epidemics had many cases of the original form circulating, soon after the D614G variant was introduced into a region it became the prevalent form."

      Geographic information from samples from the GISAID COVID-19 viral sequence database enabled tracking of this highly recurrent pattern, a shift in the viral population from the original form to the D614G variant. This occurred at every geographic level: country, subcountry, county, and city.

      Two independent lines of experimental evidence that support these initial results are included in today's paper. These additional experiments, led by Professor Erica Ollmann Saphire, Ph.D., at the La Jolla Institute, and by Professor David Montefiori, Ph.D., at Duke University, showed that the D614G change increases the virus's infectivity in the laboratory. These new experiments, as well as more extensive sequence and clinical data and improved statistical models, are presented in the Cell paper. More in vivo work remains to be done to determine the full implications of the change...

      Comment


      • #5
        A word of caution in assigning to much significance to theses cell culture experiments, as they are dealing with a tiny part of the infection process and immune response, and should be treated with extreme caution if extrapolating to behaviour in a host organism.

        Comment


        • #6
          Journal Pre-proof

          Making sense of mutation: what D614G means for the COVID-19 pandemic remains
          unclear


          Nathan D. Grubaugh, William P. Hanage, Angela L. Rasmussen
          PII: S0092-8674(20)30817-5
          DOI: https://doi.org/10.1016/j.cell.2020.06.040
          Reference: CELL 11499

          To appear in: Cell

          Please cite this article as: Grubaugh, N.D., Hanage, W.P., Rasmussen, A.L., Making sense of
          mutation: what D614G means for the COVID-19 pandemic remains unclear, Cell (2020), doi: https://
          doi.org/10.1016/j.cell.2020.06.040.

          This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition
          of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of
          record. This version will undergo additional copyediting, typesetting and review before it is published
          in its final form, but we are providing this version to give early visibility of the article. Please note that,
          during the production process, errors may be discovered which could affect the content, and all legal
          disclaimers that apply to the journal pertain.
          ? 2020 Elsevier Inc.

          Making sense of mutation: what D614G means for the COVID-19 pandemic remains unclear
          Nathan D. Grubaugh1
          *, William P. Hanage2
          *, Angela L. Rasmussen3
          *
          1Department of Epidemiology of Microbial Diseases, Yale School of Public Health, New Haven, CT 06510, USA
          2Center for Communicable Disease Dynamics, Department of Epidemiology, Harvard T. H. Chan School of Public
          Health, Boston, MA 02115, USA
          3Center for Infection and Immunity, Columbia Mailman School of Public Health, New York, NY 10032, USA
          Correspondence: grubaughlab@gmail.com


          Abstract
          Korber et al. (2020) found that a SARS-CoV-2 variant in the spike protein, D614G, rapidly became dominant around the world. While clinical and in vitro data suggest that D614G changes the virus phenotype, the impact of the mutation on transmission, disease, and vaccine and therapeutic development are largely unknown.

          Introduction
          Following the emergence of SARS-CoV-2 in China in late 2019, and the rapid expansion of the COVID19 pandemic in 2020, questions about viral evolution have come tumbling after. Did SARS-CoV-2 evolve to become better adapted to humans? More infectious or transmissible? More deadly? Virus mutations can rise in frequency due to natural selection, random genetic drift, or features of recent epidemiology. As these forces can work in tandem, it’s often hard to differentiate when a virus mutation becomes common through fitness or by chance. It is even harder to determine if a single mutation will change the outcome of an infection, or a pandemic.

          The new study by Korber et al. (2020) sits at the heart of this debate. They present compelling data that an amino acid change in the virus’ spike protein, D614G, emerged early during the pandemic, and viruses containing G614 are now dominant in many places around the world. The crucial questions are whether this is the result of natural selection, and what it means for the COVID-19 pandemic. For viruses like SARS-CoV-2 transmission really is everything - if they don’t get into another host their lineage ends. Korber et al. (2020) hypothesized that the rapid spread of G614 was because it is more infectious than D614. In support of their hypothesis, the authors provided evidence that clinical samples from G614 infections have a higher levels of viral RNA, and produced higher titers in pseudoviruses from in vitro experiments; results that now seem to be corroborated by others [e.g. (Hu et al., 2020; LorenzoRedondo et al., 2020; Ozono et al., 2020; Wagner et al., 2020)].

          Still, these data do not prove that G614 is more infectious or transmissible than viruses containing D614. And because of that, many questions remain on the potential impacts, if any, that D614G has on the COVID-19 pandemic.

          Will D614G make outbreaks harder to control?
          To answer this question, we must first explore how G614 became the dominant genotype, and what impacts it may have on transmission. As an alternative hypothesis to the one described above, the increase in the frequency of G614 may be explained by chance, and the epidemiology of the pandemic.

          In February, the area with the most COVID-19 cases shifted from China to Europe, and then in March on to the US. As this and other work shows, the great majority of SARS-CoV-2 lineages in the US arrived from Europe, which is unsurprising considering the amounts of travel between the continents. Whether lineages become established in a region is a function not only of transmission, but also the number of times they are introduced. There is good evidence that for SARS-CoV-2, a minority of infections are responsible for the majority of transmission (Endo et al., 2020). Therefore, while most introductions go extinct, those that make it, make it big (Lloyd-Smith et al., 2005). Over the period that G614 became the global majority variant, the number of introductions from China where D614 was still dominant were declining, while those from Europe climbed. This alone might explain the apparent success of G614.

          Even if viruses containing G614 got “lucky” in escaping China, the variant may still provide a transmission boost. The clinical and in vitro data provided by Korber et al. (2020) certainly make this a plausible scenario. However, higher detection of SARS-CoV-2 RNA in oral and nasal swabs may not be a direct reflection of transmission potential. In addition, much transmission likely happens in the presymptomatic stage, and we don’t know how these differences during the symptomatic phase compare.

          The pseudovirus assays used in this study can demonstrate the ability to infect a cell in culture and the results are important, but it’s not clear what it means for the ability to productively transmit to a new host. These assays don’t account for the effect of other viral or host proteins, and the parade of biochemical host-pathogen interactions that must occur to support infection and transmission. Therefore, as prior experience with the 2013-2016 Ebola epidemic suggests (Marzi et al., 2018), it’s impossible to conclude that a single mutation alone would have a major impact in a large, diverse human population based on in vitro infectivity and fitness data.

          If G614 truly is more transmissible in equivalently mixing populations, then yes, the virus will be harder to control. But we cannot definitively answer this question at the moment
          .

          Will D614G make infections more severe?
          So far there is no evidence that infection with SARS-CoV-2 containing the G614 variant will lead to more severe disease.
          By examining clinical data from 999 COVID-19 cases diagnosed in the United Kingdom, Korber et al. (2020) found that patients infected with viruses containing G614 had higher levels of virus RNA, but not did not find a difference in hospitalization outcomes. These clinical observations are supported by two independent studies: 175 COVID-19 patients from Seattle, WA (Wagner et al., 2020) and 88 COVID-19 patients from Chicago, IL (Lorenzo-Redondo et al., 2020). Viral load and disease severity are not always correlated, particularly when viral RNA is used to estimate virus titer. The current evidence suggests that D614G is less important for COVID-19 than other risk factors, such as age or comorbidities.

          Will D614G impact therapeutic and vaccine designs?
          While the D614G mutation is located in the virus’ external spike protein that receives a lot of attention from the human immune system, and thus could have an influence on the ability of SARS-CoV-2 to evade vaccine-induced immunity, we think that it’s unlikely for these reasons. D614G is not in the receptor-binding domain (RBD) of the spike protein, but the interface between the individual spike protomers that stabilize its mature trimeric form on the virion surface through hydrogen bonding. Korber et al. (2020) propose that this may result in the loss of between-protomer hydrogen bonds, modulate interactions between spike protomers, or change glycosylation patterns. While any of these changes could alter infectivity, it is less likely that it would drastically alter the immunogenicity of RBD epitopes thought to be important for antibody neutralization. Furthermore, Korber et al. (2020) and others (Hu et al., 2020; Ozono et al., 2020) found that the antibodies generated from natural infection with viruses containing D614 or G614 could cross neutralize, suggesting that the locus is not critical for antibodymediated immunity. The D614G mutation is therefore unlikely to have a major impact on the efficacy of vaccines currently in the pipeline, some of which exclusively target the RBD.

          Because the specific effect of D614G on spike function in entry and fusion is unknown, the impact of this mutation on therapeutic entry inhibitors is unknown. There is no current evidence that it would interfere with therapeutic strategies such as monoclonal antibodies designed to disrupt spike binding with ACE2 or drugs that modulate downstream processes such as endosomal acidification. However, until we better understand the role of D614G during natural SARS-CoV-2 infection, the mutation should be taken into consideration for any vaccine or therapeutic design.

          Conclusions
          While there has already been much breathless commentary on what this mutation means for the COVID19 pandemic, the global expansion of G614 whether through natural selection or chance means that this variant now is the pandemic. As a result its properties matter. It is clear from the in vitro and clinical data that G614 has a distinct phenotype, but whether this is the result of bonafide adaptation to human ACE2, whether it increases transmissibility, or will have a notable effect, is not clear. The work by Korber et al. (2020) provides an early base for more extensive epidemiological, in vivo experimental, and diverse clinical investigations to fill in the many critical gaps in how D614G impacts the pandemic.

          ...
          "Safety and security don't just happen, they are the result of collective consensus and public investment. We owe our children, the most vulnerable citizens in our society, a life free of violence and fear."
          -Nelson Mandela

          Comment


          • JJackson
            JJackson commented
            Editing a comment
            Well balanced explanation, and no hype. Kudos to the authors and Pathfinder for posting it.
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