Virus World

SARS-CoV-2 can mutate to evade immunity, with consequences for the efficacy of emerging vaccines and antibody therapeutics. Herein we demonstrate that the immunodominant SARS-CoV-2 spike (S) receptor binding motif (RBM) is the most divergent region of S, and provide epidemiological, clinical, and molecular characterization of a prevalent RBM variant, N439K. We demonstrate that N439K S protein has enhanced binding affinity to the hACE2 receptor, and that N439K virus has similar clinical outcomes and in vitro replication fitness as compared to wild-type.

We observed that the N439K mutation resulted in immune escape from a panel of neutralizing monoclonal antibodies, including one in clinical trials, as well as from polyclonal sera from a sizeable fraction of persons recovered from infection. Immune evasion mutations that maintain virulence and fitness such as N439K can emerge within SARS-CoV-2 S, highlighting the need for ongoing molecular surveillance to guide development and usage of vaccines and therapeutics.

Preprint available in bioRxiv (Nov. 5, 2020):

https://doi.org/10.1101/2020.11.04.355842

A spike protein mutation D614G became dominant in SARS-CoV-2 during the COVID-19 pandemic.  However, the impact on viral spread and vaccine efficacy remains to be defined. Here, we engineer the D614G mutation in the USA-WA1/2020 strain and characterize its effect. D614G enhances replication on human lung epithelial cells and primary human airway tissues through an improved infectivity of virions. Hamsters infected with the G614 variant produced higher infectious titers in the nasal washes and trachea, but not lungs, confirming clinical evidence that the D614G mutation enhances viral loads in the upper respiratory tract of COVID-19 patients and may increases transmission. Sera from D614-infected hamsters exhibit modestly higher neutralization titers against G614 virus than against D614 virus, indicating that (i) the mutation may not reduce the ability of vaccines in clinical trials to protect against COVID-19 and (ii) therapeutic antibodies should be tested against the circulating G614 virus. Together with clinical findings, our work underscores the importance of this mutation in viral spread, vaccine efficacy, and antibody therapy.

Published in Nature (Oct. 26, 2020):

https://doi.org/10.1038/s41586-020-2895-3

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.

The SARS-CoV-2 virus has a low mutation rate overall (much lower than the viruses that cause influenza and HIV-AIDS). The D614G variant appears as part of a set of four linked mutations that appear to have arisen once and then moved together around the world as a consistent set of variations. "It's remarkable to me," commented Will Fischer of Los Alamos, an author on the study, "both that this increase in infectivity was detected by careful observation of sequence data alone, and that our experimental colleagues could confirm it with live virus in such a short time."....

Original Study Published in Cell:

https://doi.org/10.1016/j.cell.2020.06.043

A study involving more than 5,000 COVID-19 patients in Houston finds that the virus that causes the disease is accumulating genetic mutations, one of which may have made it more contagious. According to the paper published in the peer-reviewed journal mBIO, that mutation, called D614G, is located in the spike protein that pries open our cells for viral entry. It’s the largest peer-reviewed study of SARS-CoV-2 genome sequences in one metropolitan region of the U.S. to date.  The paper shows “the virus is mutating due to a combination of neutral drift — which just means random genetic changes that don’t help or hurt the virus — and pressure from our immune systems,” said Ilya Finkelstein, associate professor of molecular biosciences at The University of Texas at Austin and co-author of the study. The study was carried out by scientists at Houston Methodist Hospital, UT Austin and elsewhere.  During the initial wave of the pandemic, 71% of the novel coronaviruses identified in patients in Houston had this mutation. When the second wave of the outbreak hit Houston during the summer, this variant had leaped to 99.9% prevalence. This mirrors a trend observed around the world. A study published in July based on more than 28,000 genome sequences found that variants carrying the D614G mutation became the globally dominant form of SARS-CoV-2 in about a month. SARS-CoV-2 is the coronavirus that causes COVID-19.

So why did strains containing this mutation outcompete those that didn’t have it? Perhaps they’re more contagious. A study of more than 25,000 genome sequences in the U.K. found that viruses with the mutation tended to transmit slightly faster than those without it and caused larger clusters of infections. Natural selection would favor strains of the virus that transmit more easily. But not all scientists are convinced. Some have suggested another explanation, called “founder’s effects.” In that scenario, the D614G mutation might have been more common in the first viruses to arrive in Europe and North America, essentially giving them a head start on other strains. The spike protein is also continuing to accumulate additional mutations of unknown significance. The Houston Methodist-UT Austin team also showed in lab experiments that at least one such mutation allows spike to evade a neutralizing antibody that humans naturally produce to fight SARS-CoV-2 infections. This may allow that variant of the virus to more easily slip past our immune systems. Although it is not clear yet whether that translates into it also being more easily transmitted between individuals.

The good news is that this mutation is rare and does not appear to make the disease more severe for infected patients. According to Finkelstein, the group did not see viruses that have learned to evade first-generation vaccines and therapeutic antibody formulations. “The virus continues to mutate as it rips through the world,” Finkelstein said. “Real-time surveillance efforts like our study will ensure that global vaccines and therapeutics are always one step ahead.” The scientists noted a total of 285 mutations across thousands of infections, although most don’t appear to have a significant effect on how severe the disease is. Ongoing studies are continuing to surveil the third wave of COVID-19 patients and to characterize how the virus is adapting to neutralizing antibodies that are produced by our immune systems. Each new infection is a roll of the dice, an additional chance to develop more dangerous mutations.

Preprint of  Study available at medRxiv (Sept. 29, 2020):

https://doi.org/10.1101/2020.09.22.20199125

Researchers in the United States and Japan have conducted a study showing a common mutation in the spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) - the agent that causes coronavirus disease 2019 (COVD-19) - enhances the infectivity, replication, and early transmission of the virus. The team’s study of SARS-CoV-2 engineered to harbor the D614G mutation found that this strain was replicated more efficiently in primary human proximal airway epithelial cells than the wildtype virus did. In a hamster model of infection, the D614G strain also showed much faster respiratory droplet transmissibility than the wildtype virus shortly following infection. Ralph Baric (the University of North Carolina at Chapel Hill) and colleagues from the University of Wisconsin, University of Tokyo, and the National Institute of Infectious Diseases, Tokyo, say the findings support the need to periodically review SARS-CoV-2 contemporary isolates and identify any new variants with increased transmissibility and pathogenesis that may have emerged. A pre-print version of the paper is available on the server bioRxiv*, while the article undergoes peer review.

To investigate the function of the D614G substitution in SARS-CoV-2 replication and transmissibility, the researchers engineered variants containing the D614G mutation in the spike protein, as well as a second variant containing the gene for the bioluminescent reporter nanoLuciferease (nLuc). The team compared the growth of wildtype SARS-CoV-2 and the D614G variant in primary human nasal epithelia (HNE), large (proximal) airway epithelia (LAE), and distal lung small airway epithelia (SAE). The D614G-infected HNE and LAE cultures, but not the SAE cultures, exhibited significantly higher viral titers than the wildtype-infected cultures. Competitive co-infection assays performed in LAE cultures simultaneously infected with both viruses showed that the D614G variant became dominant in the cultures, whether the wildtype virus was originally present at a 1:1 or 10:1 ratio over the D614G mutant. Next, the team performed scanning and transmission electron microscopy to visualize virions present on the surface of primary human airway cell cultures. No significant differences in virion morphology or the number of spike proteins were observed between the two viruses.

Further analysis revealed more differences between the viruses

The researchers used the nLuc-expressing recombinant SARS-CoV-2 encoding either wildtype or D614G spike to measure antibody neutralization activity in serum samples taken from mice vaccinated with D614 (wildtype) spike. This revealed that the samples half-maximal inhibitory dilution values against the D614G virus were between 0.8 and 5.1 times higher than against the wildtype virus, indicating that the D614G variant makes SARS-CoV-2 more sensitive to neutralizing antibodies.

Evaluating respiratory droplet transmissibility

To evaluate the role of the D614G variant in SARS-CoV-2 respiratory droplet transmissibility, the researchers set up eight pairs of hamsters, each comprising a naïve hamster alongside an infected animal 1 day following infection.  Both the wildtype and D614G viruses were efficiently transmitted to naïve hamsters. At 4 and 6 days following infection, the infected hamsters and the exposed hamsters exhibited similar viral titers, regardless of which virus they had been infected with.  However, five of eight hamsters exposed to the D614G-infected group showed infection and had detectable viral shedding on day 2, while those exposed to the wildtype-infected group showed no infection or viral shedding. This suggests that the D614G variant is transmitted much more quickly between hamsters via droplets and aerosols than the wild type virus is.

Study available at preprint in bioRxiv (Sept. 29, 2020):

https://doi.org/10.1101/2020.09.28.317685

A vaccine candidate developed by scientists in the United States performed well in an experiment that pitched it against the most common mutation of the coronavirus, according to a new study. The product, manufactured by Moderna, a biotechnology company based in Massachusetts, was tested on a modified strain that simulated the D614G mutation of the coronavirus. Research has shown this is present in more than 70 per cent of the confirmed infections around the world, and in close to 100 per cent in some European countries. Compared to other strains, D614G produces more viral copies in the respiratory tract and spreads more efficiently from person to person. What the US researchers, led by Professor Drew Weissman from the University of Pennsylvania, sought to find out was if the mutation could evade a vaccine-induced immune response.

Their methodology involved injecting the Moderna vaccine into humans, monkeys and mice, then collecting blood samples a few weeks later, after antibodies had been produced. What they found was that the antibodies were up to four times more likely to bind to the pseudovirus if it had the D614G gene than if it did not. “This is not an escape mutation that would impede current vaccines,” the researchers in a paper published on Friday on the preprint website Medrxiv.org, which means it has not been peer-reviewed. But they said they were not surprised by the results. That is because, according to the study, the mutation makes a subtle change to the virus’ spike protein, which makes it easier to bind with a receptor on the host cell. But that also increases the chances of antibodies binding with the virus. “The gain in infectivity provided by D614G comes at the cost [to the pathogen] of making the virus more vulnerable to neutralising antibodies,” Weissman said. His collaborators were from Duke University, Harvard Medical School and Los Alamos National Laboratory.

Preprint available at medRxiv (July 24, 2020):

https://doi.org/10.1101/2020.07.22.20159905