What is the Epstein-Barr virus?

The Epstein-Barr virus (EBV) is one of eight viruses in the herpes family. Also known as human herpesvirus 4 (HHV-4), it is one of the most common viruses in humans, with between 90 and 95 per cent of adults infected.

While it is best known as the cause of glandular fever, it is also associated with a wide range of more serious illnesses, including

• Cancers—such as Hodgkin’s lymphoma, Burkitt’s lymphoma, nasopharyngeal carcinoma, some stomach cancers and possibly breast cancer
• Conditions associated with Human Immunodeficiency Virus (HIV)—including hairy leukoplakia and central nervous system lymphomas
• Auto-immune diseases—dermatomyositis, systemic lupus erythematosus, rheumatoid arthritis, and multiple sclerosis

It is estimated that every year, around 200,000 cancers are directly attributable to EBV.  

At present there are two known strains of the virus, EBV-1 and EBV-2. EBV-2 is less effective at causing B-cell growth and proliferation than EBV-1. Scientists don’t know why the virus causes a relatively mild disease like glandular fever in some people and malignant tumours in others. Some evidence suggests that genetic factors and reduced immunity GLOSSARY immunityA body's reaction to the introduction of foreign substances, through the production of defensive substances such as antibodies. may play a role.

How it was discovered

In 1961, a surgeon working in Uganda, Denis Burkitt, presented the results of his research to staff at the Middlesex Hospital Medical School in Britain. He reported that the incidence of a certain tumour in African children had a geographic distribution corresponding to rainfall and temperature patterns.

The disease, which affects about 8 in every 100,000 children in parts of Africa and Papua New Guinea, quickly became known as Burkitt’s lymphoma. The influence of climate on its incidence seemed to suggest that some biological factor was involved. Three researchers, M.A. Epstein, Y.M. Barr and B.G. Achong, immediately began looking for possible cancer-causing viruses in samples of the tumour sent from Uganda to Britain.

In 1964, they identified the culprit using an electron microscope: a previously unknown member of the herpes family of viruses. Epstein and Barr were awarded the dubious honour of having the pathogen named after them. It was the first virus to be directly associated with human cancer.    

How the virus spreads

EBV is spread mainly via the transfer of saliva between individuals, which is the reason that glandular fever has been dubbed the ‘kissing disease’. It can also be spread by sharing utensils/toothbrushes etc, through blood (transfusions, organ transplants) and rarely via semen from sexual contact. People infected with the Epstein-Barr virus will retain it for life, but it may not make them sick. In fact, the virus infects almost everyone in developing countries and more than 95 per cent of people in developed countries.

Most people are infected with the virus during childhood, probably by their mothers, and are usually not noticeably affected. On the other hand, people infected for the first time during or after adolescence (10‑20 per cent of people living in developed countries) have a 50 per cent chance of contracting glandular fever. They may be ill for several weeks or months before their immune system kicks in to action.

Symptoms of EBV

The symptoms of EBV can include:

• fever/chills
• fatigue
• inflamed throat
• swollen lymph nodes in the neck
• swollen liver or spleen
• rash
• loss of appetite
• minor aches and pains

Often the symptoms of EBV are not distinguishable from other general colds and flu. Those who do experience symptoms generally recover within 2 to 4 weeks, however full recovery can take months.  It is important to note however that while your symptoms may have disappeared, the virus has not. Instead it becomes inactive—essentially hiding away inside the body’s B-cells. It is possible for the EBV to ‘reactivate’ later in life, though this is rare. ­­

Diagnosing EBV

It can be difficult for doctors to diagnose EBV because its symptoms so closely resemble a range of other illnesses. However, a blood test can confirm the presence of EBV antibodies in the system, whether someone is susceptible to the virus, or has had a recent or past infection. Find out more about the laboratory testing of EBV.

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A photomicrograph showing cells containing the Epstein-Barr virus. Image source: CDC/Dr Paul M. Feorino / Wikimedia Commons.

How does the virus work?

EBV produces about 100 different antigens GLOSSARY antigens A toxin or other foreign substance, usually a protein, which induces an immune response in the body, especially the production of antibodies. (large protein molecules) during the active phase of the viral cycle. In contrast, only about 10 antigens are produced during the inactive phase: these include the EBV nuclear antigens (EBVNAs 1–6), and the latent membrane proteins (LMPs 1–3).

EBV primarily infects the oropharynx (the part of the throat at the back of the mouth), specifically the salivary glands, as well as the oral mucosal membrane (the mucous membrane lining the inside of the mouth) and the nasopharyngeal epithelial tissue (above your soft palate). The virus replicates quickly in these areas, enabling it to spread easily through saliva from one person to another.

But EBV doesn’t stop there. After the initial infection it begins to infect white blood cells known as B lymphocytes (B-cells). Infection of B-cells with the virus transforms them into B-cells with unlimited growth potential, causing them to rapidly multiply and increase in number. This proliferation is combated by the immune system; it activates Cytotoxic T lymphocytes (T-cells) and Natural Killer Cells in defence against EBV, which combine with certain antigens produced by the virus and kill cells that harbour it.

Unfortunately, when the virus is associated with Burkitt’s lymphoma and nasopharyngeal carcinoma it appears to produce only one antigen—EBNA1. T-cells are unable to combine with this particular antigen and will not attack the infected cell.

• Antibodies and T-cells

  The two potent weapons of acquired immunity are the antibody GLOSSARY antibodyA protein produced by the body's immune system in response to a foreign substance (antigen). An antibody reacts specifically with the antigen that induced its formation and inactivates the antigen. Our bodies fight off an infection by producing antibodies. and the T-cell. They operate at different levels: antibodies at the molecular level and T-cells at the cellular level.

  Antibodies

  Antibodies are protein molecules that move freely in the bloodstream. They are manufactured by white blood cells in response to the presence of molecules on the surface of the disease-causing microorganism. These molecules are known as antigens. A particular antibody will combine best with the antigen that caused the production of that antibody: thus, the antibody is said to be specific for its antigen. Antibodies react chemically with these antigens in such a way that the microorganism or toxin is neutralised and destroyed before a cell is infected.

  T-cells

  At the cellular level, the body manufactures what are known as cytotoxic T lymphocytes—T-cells. These white blood cells recognise certain antigens produced by microorganisms and kill cells that harbour them.

  If an invading microorganism evades the antibodies and infects a cell, the T-cells will recognise the infected cell and kill it. Thus the two weapons of acquired immunity, the antibody and the T-cell, complement each other.

  Thanks for the memory

  Once an infection has been overcome, antibodies and T-cells remain in the body, often for years and sometimes for life, acting as a kind of ‘memory’. If the microorganism attacks again, these antibodies ‘remember’ it. They combine with the antigens on the surface of the microorganism and immobilise it so it is no longer able to damage cells in the body. If, despite the efforts of the antibodies, some cells in the body become infected, the memory T-cells are mobilised rapidly to kill the infected cells. Thus, the body has acquired immunity against that particular disease.

  Active immunisation

  Vaccination works by introducing harmless versions of a disease-causing microorganism or certain antigens of the microorganism to the immune systems of uninfected individuals. This ‘tricks’ the immune system into producing antibodies and, when required, T-cells specific to the microorganism. The body is thus equipped with the right armory should infection by the real microorganism occur. The vaccines used in vaccination are of several types:

  • live, attenuated microorganisms;
  • killed microorganisms; and
  • only one, or a few, antigens of the microorganisms, such as a toxin (toxic molecule) which is first rendered safe (detoxified). This type is called a subunit vaccine and is a type of acellular vaccine.

  Passive immunisation

  Protection against some infections may be achieved by passive immunisation. With this type of immunisation, antibodies produced in one person are introduced into another. These antibodies are injected into the host shortly before the expected exposure to a disease-causing microorganism. The antibodies are obtained from the blood plasma of people who have had the disease (or have been immunised against it). Injection of the antibodies confers immediate protection against infection for a short time (weeks). It may also be effective if given shortly after such exposure; for example, after being bitten by a snake or rabid dog.

Although our immune system may seem to have beaten EBV, in reality it has been duped. When our immune system begins to attack EBV, the virus admits defeat and retreats into hiding, lurking dormant inside the body’s B cells. In this way it continues to live ‘under the radar’. It is no longer under threat from the immune system, but is able to keep spreading from person to person without our knowledge that it’s even there. Occasionally if the correct conditions exist in the future (such as reduced immunity through infection), the virus can sometimes reactivate.

In some people, however, the T-cell response is not strong enough, resulting in unrestricted growth of B-cell lymphocytes, which greatly increases the risk of the patient developing a form of cancer.

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A colourised electron micrograph image of a T-cell, one of the body's defences against EBV. Image source: NIAID / Flickr.

How does EBV induce the growth of cancers?

Since its initial discovery as the first human cancer virus, EBV has continued to be associated with the development of a wide range of tumours. Scientists have been working for years to determine why EBV causes cancer in only a small percentage of the vast number of people who carry the virus, how it develops into different types of cancers, and why it affects different populations and age groups across the world.  

A key piece in the puzzle was understanding how our immune systems respond to and fight EBV. In the 1970s doctors discovered that a large number of patients who had received transplants in the decade before had developed cancer—generally skin cancer but also lymphomas too. These became known as post-transplant lymphoma, but doctors could not work out what caused them. Samples of these tumours were tested and found to be positive for EBV.

With the outbreak of the AIDS virus in the 1980s, it was found that some patients of the disease developed certain types of cancer including lymphomas that had close similarities to post-transplant lymphomas. When analysed, almost all the samples tested positive for EBV too.

What was the link between people with HIIV and transplant patients? The answer lay in their immune systems—specifically its weakened state after either transplant or fighting the AIDS virus.

Further understanding of the virus and the way it worked came in 1984 when the complete DNA sequence of EBV was published. The key proteins made by the virus—those which allowed it to push B cells to multiply and become cancerous—were now known to scientists.

Researchers concluded that although there were a combination of variables that caused EBV cancers to develop, there was one consistent factor—a change in the balance between the immune system and the virus. Basically, the ability of the body and its immune system to stay in control of the virus was imperative to stopping cancers forming.

A weakened or poorly functioning immune system (due to illness, disease, drugs or even a genetic variation in genes that reduces the ability of immune cells to spot EBV) result in the normally shy and contained virus kicking up its heels and running rampant. The proteins made by the virus direct the cells to keep dividing and multiplying exponentially—the principal cause of cancer.  

• Epstein-Barr virus and related cancers

  Characterisation of EBV-associated malignancies

  Malignancy	Subtype	EBV gene expression pattern	% EBV positivity
  Burkitt’s lymphoma	Endemic Non-endemic	Latency GLOSSARY LatencyA time delay between the cause and effect of some physical change in the system being observed. I	>95% 15-30%
  Hodgkin’s disease	MC LD NS LP	Latency II	70% >95% 10-40% <5%
  Non-Hodgkin’s lymphoma	Nasal T/NK Angioimmunoblastic Lymphadenopathy	Latency II Latency II	>90% Unknown
  Nasopharyngeal carcinoma	Anaplastic	Latency II	>95%
  Breast cancer	Medullary carcinoma Adenocarcinoma	Not clear	0-51%
  Gastric cancer	Lymphoepithelioma-like Adenocarcinoma	Controversial Novel LMP-1 Negative Latency III	>90% 5-25%
  Post-transplant lymphoproliferative disorders		Latency III	>90%
  Aids-associated lymphomas	IP-CNS Other	Latency III	>95% 30-50%
  Leiomyosarcomas in immunosuppressed individuals	Leiomyosarcomas varies	Unclear	Frequent

The search for a vaccine

As with most viruses, the best chance of defence against EBV is vaccination. Yet a vaccine has been elusive, partly because the virus is so good at hiding.

To prevent Burkitt’s lymphoma or nasopharyngeal carcinoma, a vaccine would need to provide 100 per cent immunity or be capable of establishing a T-cell population that recognises EBNA1. Both tasks are extremely difficult. It may be possible, though, to produce a vaccine against glandular fever and post-transplant lymphoproliferative disease, since both produce antigens that are recognised by T-cells.

Australian scientists have produced such a vaccine. Based at the QIMR Berghofer Medical Research Institute, they have conducted a trial with a prototype vaccine using human volunteers.

• The future of vaccines

  New techniques for the development and delivery of vaccines are aiding the search for effective immunisation strategies.

  Gene guns and golden bullets

  One such technique uses DNA to produce what are known as nucleic acid GLOSSARY nucleic acidA large molecule made up of a sequence of phosphorylated nitrogen-containing bases. DNA and RNA are both nucleic acids. vaccines (sometimes called ‘naked DNA’ vaccines). Scientists isolate the genes from disease-causing bacteria or viruses that provide information to make specific antigens. These genes are then inserted into a plasmid GLOSSARY plasmidA small, circular DNA molecule. Bacteria can have plasmids in addition to the DNA of the main chromosome. Foreign DNA can be added to plasmids. The modified plasmid then transports the DNA into a new cell. (which is a genetic element capable of replicating independently of the chromosomal DNA) which, in turn, is injected into the patient. This can be done in the usual way with a hypodermic syringe into muscle tissue, or with a ‘gene gun’, which fires tiny gold particles coated with the DNA into the surface layers of the skin.

  Once inside the body the plasmids penetrate host cells, where they start manufacturing antigens. These antigens, released over a long period, induce an active immune response by the body, including the production of both antibodies and specialised white blood cells (cytotoxic T lymphocytes).

  The technique may be used to produce vaccines for both viral and bacterial infections; it shows such promise that clinical investigation of a possible AIDS vaccine has already begun.

  ‘Antigen factory’ vaccines

  A similar technique involves the insertion of selected genes from a disease into benign bacteria or viruses. These are then administered to the patient and serve as a sort of antigen factory, using the inserted genes to churn out antigens of the disease-causing organism. These antigens invoke an immune response that will help protect the patient from a subsequent infection. Several ‘antigen factory’ organisms, including the cowpox virus, are being tested for use in HIV vaccines.

  Making vaccines more effective

  Conventional vaccines often use weakened or killed cells of the disease itself. While this has proved effective against many diseases, some vaccines developed by this technique produce occasional side-effects in patients. New methods using recombinant DNA technology have led to the development of many vaccines that use only a small part of the disease-causing organism or that use a harmless version of the microbe. For example, by removing genes from the cholera bacterium it becomes safe to use in a vaccine.

  These methods produce extremely safe vaccines, but they are often less effective than whole-cell vaccines. A major area of vaccine research seeks ways of making such ‘subunit’ vaccines more effective in producing an immune response. This usually involves the use of what are called adjuvants, which are substances added to the vaccine to aid its operation. Conventional vaccines mostly use aluminium salt as an adjuvant, but recent work has tested oil-based emulsions that contain biodegradable material.

  Vaccines have been developed at the University of Oxford in the UK that use genetically engineered viruses as a vector to carry genes for both antigen and adjuvant proteins. This technology could potentially help in the fight against diseases such as malaria and HIV that have been traditionally difficult to vaccinate against.

  User-friendly vaccines

  Vaccine development agencies recognise the importance of increasing the rate of childhood immunisation. One way of achieving this would be to develop vaccines that could be taken orally or nasally, rather than by injection.

  With this in mind, researchers have investigated the use of what are called microcapsules. These consist of an inner reservoir of antigen surrounded by an outer, biodegradable polymer wall, through which the antigen is released slowly. Vaccines administered in this way have been shown to produce strong, sustained immune responses for some antigens. One advantage of microcapsules is that refrigeration is not required, making them suitable for remote regions.

  Scientists are currently investigating the possible safety implications of having microcapsules in the body for extended periods. If the method proves to be safe, it may become widely used for vaccine administration.

  Combining vaccines

  Another way of boosting the rate of childhood immunisation would be to combine vaccines so that patients could be vaccinated against several diseases at one time. Some combinations are already available (the diphtheria-tetanus-pertussis vaccine is one example), and researchers continue to seek ways of combining vaccines without reducing their effectiveness.

  Linking chains

  Armed with an understanding of the molecular structure of antigens for a particular disease, scientists are often able to replicate certain peptides of the antigen in the laboratory. These peptides show promise as vaccines because they can produce an immune response in patients. Indeed, the new Epstein-Barr virus vaccine currently under clinical trial is based on a peptide GLOSSARY peptidePeptide: a molecule consisting of a short chain of amino acids. Longer chains of amino acids are called proteins. found on one of the virus’s antigens.

  Nevertheless, despite considerable promise, there has been surprisingly little progress in the development of synthetic peptide vaccines. One reason for this might be that the peptides are too small and unstable to provoke an effective immune response.

  Australian scientists at the Cooperative Research Centre for Vaccine Technology are pioneering work to polymerise (join together) small peptides. Early results suggest that the polymerisation process aids the potency of the peptides as antigens, and may also allow peptides against more than one disease to be included in the same molecular structure.

  Vaccines don’t always target microbe-borne diseases

  Trials in the USA have been conducted to test whether drug addiction can be treated with vaccines. Nicotine, cocaine and methamphetamine addiction could all potentially be treated this way. Vaccination for addiction gradually stimulates the immune system to produce antibodies that bind to the drug. This then progressively prevents the drug from entering the brain, reducing the effect on the body with minimal withdrawal symptoms.

  The body’s own cancer cells may also be targeted by vaccines. By targeting the specific molecules that are found on the surface of tumour cells, vaccines can produce an immune response to cancerous cells. Cancer vaccines that show promise are those that target telomerase, an enzyme that makes cancer cells immortal.

One of the first steps in the development of the vaccine was to define the antigens of the virus that are important in the immune control of it. Antigens stimulate the manufacture of T-cells; if they—or their most important parts—could be introduced artificially to an individual, then the immune system might be ‘tricked’ into producing T-cells that would recognise the real antigen should infection occur.

The researchers were able to produce a peptide identical to part of the EBV antigen EBNA3. This peptide formed the basis of the vaccine: by injecting it into patients, researchers hoped to ‘arm’ the body with T-cells in readiness for an invasion of the real disease.

The trial has been completed and scientists are now confident that the vaccine does not have any harmful effects on patients. Recipients didn't develop infectious mononucleosis, but being a small trial the vaccine's effectiveness in preventing disease could not be accurately assessed.

Another vaccine developed in Europe that is based on a surface protein of the virus, gp350, has successfully reduced EBV infections in an 18-month trial. Attempts are also being made to grow and expand T-cells in the laboratory to help cure various forms of EBV-induced cancers. This approach should be particularly useful for treating patients suffering from post-transplant lymphoproliferative disease.

Recently, a group from Germany described an alternative strategy for EBV vaccine. This group produced non-infectious virus-like particles in which several EBV proteins critical for viral pathogenesis GLOSSARY were either functionally inactivated or deleted.  Immunisation of animals with these virus-like particles induced anti-EBV neutralising antibody and cellular immune responses. Human trials with this vaccine formulation are expected to start very soon.

Over the last decade, there has been considerable progress on the development of therapeutic vaccines for EBV-associated cancers. These diseases include nasopharyngeal carcinoma, Hodgkin lymphoma, non-Hodgkin lymphoma and T-cell lymphoma, which generally express limited number of EBV proteins.

Two different types of therapeutic vaccines have been tested in patients with nasopharyngeal carcinoma. One of these vaccines is based on a poxvirus vector (modified vaccinia Anakara) which expresses EBV encoded EBNA1 and LMP2 proteins.  Nasopharyngeal carcinoma patients in Hong Kong who were immunised with this vaccine showed and a three to fourfold increase in the magnitude of T-cell responses to the proteins. Similar results were also obtained from another follow-up study in the United Kingdom. Another vaccine is based on dendritic GLOSSARY dendritichaving a branched form resembling a tree cells sensitised with EBV peptides or infected with viral vectors encoding EBV proteins. This vaccine induced immune response against the EBV proteins and tumor regression in two of nine patients.  

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Trials for EBV vaccines are currently underway. Image source: buffaloboy / iStockPhoto.

Conclusion

Despite its prevalence in the human population, finding a cure for EBV has not been seen as a high priority. This may be because the majority of people do not suffer serious health effects, while those who do tend to live in developing countries. However, as new cancers are linked with the virus, the need to find a solution will become of more interest to researchers, drug companies and pharmaceuticals. And that would certainly be worth a celebratory kiss.

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Image source: mcvickerphotog / Flickr.