Epstein-Barr virus (EBV) is a common virus that causes mononucleosis, or mono for short, and is associated with some types of cancer and autoimmune diseases. Despite EBV’s known effects and potential to cause disease, there are few therapeutic options and no licensed vaccines targeting the virus. Looking for ways to counter EBV, NIAID researchers are examining how the virus recognizes and interacts with cells at the molecular level. New research published in Immunity reveals the high-resolution crystal structure of a protein on the surface of EBV in complex with the receptor it binds to on the surface of human immune cells, called B cells. The researchers also discovered antibodies that potently neutralize EBV and found that they recognize the viral surface protein using interactions similar to those between EBV and its receptor on host cells. This research identifies a vulnerable site on EBV that could lead to the design of much-needed interventions against the virus.

EBV, also known as human herpesvirus 4, is one of the most common human viruses—nine out of ten people have or will have EBV in their lifetime. After being infected with EBV, many people experience no symptoms, but some experience symptoms of mononucleosis, such as fever, sore throat and fatigue. These symptoms are often mild but can be more severe in teens or adults. After the early stages of infection, the virus hides in the body and can emerge later in life or when the immune system is weakened. Recent studies have also found that EBV is linked to several types of cancer, autoimmune diseases including lupus, and other disorders.

A key step in EBV infection is for the virus to enter a cell in the body, which begins with the virus binding to a protein on the cell’s surface. The researchers, led by Dr. Masaru Kanekiyo, chief of the Molecular Immunoengineering Section at NIAID’s Vaccine Research Center, examined the atomic-level structure of an EBV surface protein called gp350 when bound to a protein on the surface of B cells called complement receptor type 2 (CR2). Usually, CR2 binds to a protein fragment, or ligand, called complement component C3d as a part of the immune response following a viral infection. The researchers found that the EBV protein precisely bound to the cell surface protein CR2 at the region where its natural ligand C3d binds, revealing that there is structural similarity between EBV and C3d in recognizing CR2 and how the virus exploits this interaction to enter and infect a cell.

The researchers also isolated neutralizing antibodies (nAbs)—immune proteins that neutralize EBV—from animals immunized against EBV and EBV-infected people. They found that the antibodies neutralized the virus in laboratory tests by binding to the EBV gp350 protein. They further determined the atomic-level structure of three of the nAbs when bound to EBV gp350. All three nAbs bound to gp350 at the same region of the protein—the region where it also binds to the cell protein CR2, demonstrating that this binding site is an important target on the virus for neutralization.

The way the CR2 cell surface protein binds its natural ligand C3d can be likened to a key fitting a lock. In this case, the key is a negatively charged pocket on the surface of C3d, while the lock is an arrangement of positively charged arginine residues on the surface of CR2. The researchers observed a remarkable molecular mimicry that occurred in duplicate. On one side, EBV gp350 mimics the characteristics of C3d, pretending to be the natural key that fits CR2 on the cell surface, unlocking the cell for the virus to infect it. On the other side, the anti-EBV nAbs mimic CR2, where they act as a lock to block the EBV gp350 protein from binding to a cell for the virus to infect. The mimicry existing on both sides of this lock-and-key set indicates that this interaction is an important step for EBV infection—and represents a major point of viral vulnerability, according to the researchers.

The findings define critical molecular interactions between EBV and its host cells. The researchers noted that more work is needed to apply these findings to the development of interventions, including examining whether the newly discovered nAbs can provide protection from EBV infection in animal models and people. This research may reveal new avenues to treat and prevent disease caused by this widespread pathogen.

Reference:

• M Gorden Joyce. Structural basis for complement receptor engagement and virus neutralization through Epstein-Barr virus gp350. Immunology [10.1016/j.immuni.2025.01.010] (2025).

An experimental vaccine designed against the highly pathogenic avian influenza H5N1 (HPAI H5N1) virus circulating in U.S. cattle was fully protective in research mice in a new study published in Nature Communications. NIAID scientists at Rocky Mountain Laboratories (RML) in Hamilton, Montana, led the animal study with colleagues from HDT Bio in Seattle who developed the replicating RNA vaccine (repRNA) platform.

Along with confirming that a single immunization with the experimental vaccine was effective against the new flu type in cattle (HPAI A H5N1 clade 2.3.4.4b), the study also allowed scientists to evaluate the vaccine method for “cross protection.” Would it work against the new virus if designed with components used in stockpiled vaccines from an older H5N1 virus (A/Vietnam/1203/2004)? They found that when the test vaccine used a design from the older H5N1 virus, protection was diminished. The findings suggest that the HPAI H5N1 circulating in the U.S. may be able to evade immunity from older H5N1 viruses.

Scientists designed the repRNA vaccine to express the protective vaccine components, as well as the RNA replication machinery derived from an alphavirus. This allows for robust expression of the protective vaccine components upon delivery with LION™, a proprietary nanoparticle formulation. The repRNA/LION technology is the basis of a vaccine that received emergency use authorization in India for COVID-19. Additional applications of repRNA/LION are advancing toward clinical trials for other serious viral diseases after showing effectiveness against several different viruses in the lab.

Scientists at RML and HDT Bio are continuing to develop the vaccine platform, and evaluations in animal models developed at RML are ongoing.

Reference: D Hawman, et al. Clade 2.3.4.4b but not historical clade 1 HA replicating RNA vaccine protects against bovine H5N1 challenge in mice. Nature Communications DOI: https://doi.org/10.1038/s41467-024-55546-7 (2025).
 

Subclinical Disease in Monkeys Exposed to H5N1 by Mouth and Stomach

A new study published in Nature found that highly pathogenic H5N1 avian influenza virus (HPAI H5N1) administered directly into the mouth and stomach of research monkeys caused self-limiting infection with no recognizable clinical signs of disease. By comparison, other routes of transmission resulted in mild or severe disease. The findings suggest that drinking raw milk contaminated with H5N1 virus can result in infection but may be less likely to lead to severe illness. Nevertheless, exposure by raw milk – which is a source of several foodborne illnesses – should be avoided to prevent H5N1 infection and potential further spread.

The research team, from NIH’s National Institute of Allergy and Infectious Diseases (NIAID), exposed cynomolgus macaques to the same clade 2.3.4.4b HPAI H5N1 virus circulating in U.S. cattle. Transmission routes included via the nose, windpipe (trachea) or directly into the mouth and stomach to mimic infection routes in people. Animals exposed via the nose and windpipe became infected, developed pneumonia and had varying degrees of disease. Animals infected in a manner that mimicked drinking had a more limited infection with no obvious disease signs. To what extent this work mirrors human infection remains unclear.

The study does suggest that infection through contaminated liquids like raw milk represents a risk for HPAI H5N1 infection of primates. The work cites the “local environment” in the stomach as potentially inactivating the virus and thus, possibly reducing the exposure dose. Scientists at NIAID’s Rocky Mountain Laboratories in Hamilton, Montana, led the work.

They exposed six animals each via the nose to mimic an upper-respiratory tract infection; the windpipe to mimic a lower-respiratory tract infection; and in the mouth and stomach to mimic consuming contaminated products. They used a dose of virus close to what has been found in contaminated raw milk. Researchers regularly monitored and examined animals for up to 14 days.

Animals exposed in the mouth and stomach became infected but showed no signs of influenza illness throughout the study. Animals exposed in the nose showed mild respiratory disease, peaking at day 10. Animals exposed in the windpipe showed severe respiratory illness within a week.

Reference: K Rosenke, A Griffin, F Kaiser, et al. Pathogenesis of bovine H5N1 clade 2.3.4.4b infection in Macaques. Nature DOI: 10.1038/s41586-025-08609-8 (2025).