The research team, led by Emory University and funded by the National Institute of Health’s (NIH) Rapid Acceleration of Diagnostics (RADx) Tech program, developed a novel method for evaluating how mutations to SARS-CoV-2 can affect recognition by antibodies used in rapid antigen tests. Because most rapid antigen tests detect the SARS-CoV-2 nucleocapsid protein (N protein), the team directly measured how mutations to the N protein impacted diagnostic antibodies’ ability to recognize their target.

“Based on our findings, none of the major past and present SARS-CoV-2 variants of concern contain mutations that would affect the capability of current rapid antigen tests to detect antibodies,” says first study author Filipp Frank, Ph.D., an assistant professor in the department of biochemistry at Emory University. “Further, these data allow us to look one step ahead and predict test performance against almost any variant that may arise.”

The study used a method called deep mutational scanning to evaluate all possible mutations in the N protein in a single, high-throughput experiment. Researchers then measured the impact of the mutations on their interaction with antibodies used in 11 commercially available rapid antigen tests and identified mutations that may allow for antibody escape.

“Accurate and efficient identification of infected individuals remains a critically important strategy for COVID-19 mitigation, and our study provides information about future SARS-CoV-2 mutations that may interfere with detection,” says senior study author Eric Ortlund, Ph.D., a professor in the department of biochemistry at Emory University. “The results outlined here can allow us to quickly adapt to the virus as new variants continue to emerge, representing an immediate clinical and public health impact.”

Findings show that it’s relatively rare for variants to have mutations to the N protein that allow them to evade diagnostic tests, but there are a small proportion of sequences that could impact detection. Researchers, public health officials, and test manufacturers can use these data to determine if a diagnostic test needs to be evaluated for its ability to detect these mutations or to inform future test design.

“Considering the endless cycle of new variants, the data from this study will be useful for years to come,” says Bruce J. Tromberg, Ph.D., director of the National Institute of Biomedical Imaging and Bioengineering (NIBIB) and lead for the RADx® Tech program at NIH.

While many variants of concern contain multiple mutations to the N protein, the study authors note that their method does not evaluate how multiple mutations could affect diagnostic antibody recognition, representing a limitation of the study.

The project was supported by NIBIB under award numbers 75N92019P00328, U54EB015408, and U54EB027690 as part of the RADx initiative, launched to speed innovation in the development, commercialization and implementation of technologies for COVID-19 testing.

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“This is essential, because inflammation releases free radicals against which nerve fibers cannot protect themselves. Therefore, the inflammation must be cleared quickly, which is precisely what Schwann cells do,” explains Dr. Sabine Taschner-Mandl, who designed the study and heads a research group at St. Anna CCRI. The new results, in which the Medical University of Vienna is also significantly involved, were published in the journal Glia.

Do Schwann cells protect against malignancy?

How are these results related to tumor growth? After nerve injury, Schwann cells adopt a “repair” mode that is also found in benign infantile nerve tumors. There, it causes the tumor cells to mature and thus reach a stage where they lose their aggressive properties and no longer divide unchecked (Weiss T., Taschner-Mandl S., et al., Nat Commun 2021).

“Based on the current results, we now suspect that the immune cell functions of Schwann cells also become effective in childhood nerve tumors. This is because in cancer, there is always a kind of inflammation bubbling away that never comes to a halt. In benign nerve tumors, ganglioneuromas, the accompanying chronic inflammation could be stopped by Schwann cells similar to nerve healing, because unlike malignancies, benign nerve tumors have many Schwann cells in their microenvironment. We also see that a lot of immune cells migrate into these tumors, for which the Schwann cells could also be responsible,” says Sabine Taschner-Mandl.

Healthy Inflammation: First Activate, Then Shut Down

In particular, the current study shows that Schwann cells can influence certain immune cells, so-called T cells, which play an important role in the defense against cancer. Schwann cells — both those in nerve regeneration and those in benign tumors — carry MHC-I and MHC-II molecules on their surface that are important for T-cell regulation. Via these molecules, Schwann cells present recognition features of material they have previously taken up from their environment.

We mimicked an inflammatory response in the laboratory and detected a whole range of additional stimulatory and inhibitory surface molecules that are also necessary for T cell activation,” explains Jakob Berner, MSc, co-first author of the study and interim PhD student in Kaan Boztug’s group at St. Anna CCRI. “Our experiments show that Schwann cells are able to take up large amounts of material via phagocytosis.”

As the first immune response to a nerve cut, Schwann cells secrete substances that attract T cells, macrophages and other immune cells. Now it turned out that not only a reaction between the classical immune cells takes place, but also between Schwann cells and T cells.

While Schwann cells initially fuel the inflammatory response by releasing interferon-gamma, they can later shut it down by up-regulating the T-cell inhibitory PD-L1 molecule.

“First activate, then shut down — that’s the normal process of an inflammatory response. If this were also the case in cancer, then it could curb cancer growth,” comments Sabine Taschner-Mandl. Whether and how these findings can be used for potential cancer therapies is now being researched.

“What’s important about the tuatara is that it represents this enormous evolutionary story that we are lucky enough to catch in what is likely its closing act,” Carrano said. “Even though it looks like a relatively simple lizard, it embodies an entire evolutionary epic going back more than 200 million years.”

The discovery comes from a handful of specimens including an extraordinarily complete and well-preserved fossil skeleton excavated from a site centered around an Allosaurus nest in northern Wyoming’s Morrison Formation. Further study of the find could help reveal why this animal’s ancient order of reptiles were winnowed down from being diverse and numerous in the Jurassic to just New Zealand’s tuatara surviving today.

The tuatara looks a bit like a particularly stout iguana, but the tuatara and its newly discovered relative are in fact not lizards at all. They are actually rhynchocephalians, an order that diverged from lizards at least 230 million years ago, Carrano said.

In their Jurassic heyday, rhynchocephalians were found nearly worldwide, came in sizes large and small, and filled ecological roles ranging from aquatic fish hunters to bulky plant munchers. But for reasons that still are not fully understood, rhynchocephalians all but disappeared as lizards and snakes grew to be the more common and more diverse reptiles across the globe.

This evolutionary chasm between lizards and rhynchocephalians helps explain the tuatara’s odd features such as teeth fused to the jaw bone, a unique chewing motion that slides the lower jaw back and forth like a saw blade, a 100-year-plus lifespan and a tolerance for colder climates.

Following O. gregori‘s formal description, Carrano said the fossil has been added to the museum’s collections where it will remain available for future study, perhaps one day helping researchers figure out why the tuatara is all that remains of the rhynchocephalians, while lizards are now found across the globe.

“These animals may have disappeared partly because of competition from lizards but perhaps also due to global shifts in climate and changing habitats,” Carrano said. “It’s fascinating when you have the dominance of one group giving way to another group over evolutionary time, and we still need more evidence to explain exactly what happened, but fossils like this one are how we will put it together.”

The researchers named the new species after museum volunteer Joseph Gregor who spent hundreds of hours meticulously scraping and chiseling the bones from a block of stone that first caught museum fossil preparator Pete Kroehler’s eye back in 2010.

“Pete is one of those people who has a kind of X-ray vision for this sort of thing,” Carrano said. “He noticed two tiny specks of bone on the side of this block and marked it to be brought back with no real idea what was in it. As it turns out, he hit the jackpot.”

The fossil is almost entirely complete, with the exception of the tail and parts of the hind legs. Carrano said that such a complete skeleton is rare for small prehistoric creatures like this because their frail bones were often destroyed either before they fossilized or as they emerge from an eroding rock formation in the present day. As a result, rhynchocephalians are mostly known to paleontologists from small fragments of their jaws and teeth.

After Kroehler, Gregor and others had freed as much of the tiny fossil from the rock as was practical given its fragility, the team, led by DeMar, set about scanning the fossil with high-resolution computerized tomography (CT), a method that uses multiple X-ray images from different angles to create a 3D representation of the specimen. The team used three separate CT scanning facilities, including one housed at the National Museum of Natural History, to capture everything they possibly could about the fossil.

Once the fossil’s bones had been digitally rendered with accuracy smaller than a millimeter, DeMar set about reassembling the digitized bones of the skull, some of which were crushed, out of place or missing on one side, using software to eventually create a nearly complete 3D reconstruction. The reconstructed 3D skull now provides researchers an unprecedented look at this Jurassic-age reptile’s head.

Given Opisthiamimus‘s diminutive size, tooth shape and rigid skull, it likely ate insects, said DeMar, adding that prey with harder shells such as beetles or water bugs might have also been on the menu. Broadly speaking, the new species looks quite a bit like a miniaturized version of its only surviving relative (tuataras are about five times longer).

“Such a complete specimen has huge potential for making comparisons with fossils collected in the future and for identifying or reclassifying specimens already sitting in a museum drawer somewhere,” DeMar said. “With the 3D models we have, at some point we could also do studies that use software to look at this critter’s jaw mechanics.”

Funding and support for this research were provided by the Smithsonian and the Australian Research Council.