Coronavirus Research Tracking - 26 June

Antibody testing.

In this week’s Research Tracker the focus is on research into immune responses to Covid-19. But first for context, a bit about the PCR tests used to detect the virus and some immune system basics.

The Research Tracker is prepared by Dr Robert Hickson for the Science Media Centre. As this is a new service, please don’t hesitate to provide feedback.

PCR Tests for SARS-CoV-2

PCR tests are used to determine if a person is currently infected with the virus. Swabs from the nose and/or throat are used to collect a sample. These are sites where the virus is known to occur. Genetic material from the swabs is then isolated for testing. 

There are a variety of PCR-based tests for the virus. They target different parts of the viral genome. Since the virus is made from RNA rather than DNA, a “reverse transcriptase PCR” (RT-PCR) test is used. This involves creating DNA copies of the viral RNA so the PCR test can be run. PCR tests usually take several hours, after sample preparation.

A comparison of seven of PCR tests found that all were suitable for routine diagnostic testing. These all require lab-based testing. More rapid home or office based tests are being developed.

In New Zealand ESR undertakes the PCR testing for SARS-CoV-2. They follow the testing protocol developed by the Institute of Virology at the Charité UniversitaetsMedizin Berlin. 

This test involves an initial RT-PCR test to detect if the coronavirus envelope protein gene is present. This is a general test to detect coronaviruses. If that test is positive then a second PCR test specific for the SARS-CoV-2 RdRP (RNA-dependent RNA polymerase) gene is used to confirm infection. A double test provides confidence that the SARS-CoV-2 virus is present.

Immunity 101

The immune system is very complex, so with apologies to immunologists, here’s a very simplified overview of some immune system responses. There are two general types of immunity. The first, “humoral immunity”, is also called antibody-mediated immunity. This identifies pathogens that are not in cells but circulating in the body. Immune system B cells when they come into contact with an antigen (usually part of a virus or other pathogen) produce antibodies against it.

These antibodies bind to the pathogen, preventing it from infecting other cells, and mark it for destruction by other immune cells (“neutralisation” in immunological speak). These antibodies are the focus of most initial immunological tests for infections. An antibody response can take two or more weeks to develop.

Antibodies come in various forms. Immunoglobulin M (IgM) are produced first, and increase shortly after infection. They are a generic response to infection. Antibodies that recognise and target a specific pathogen are called immunoglobulin G (IgG). A third type of antibody, called IgA, is associated with respiratory infections such as influenza. So it is also important in Covid-19 infections. 

The second type of immunity is “cellular immunity.” This involves T lymphocytes (or T cells) and mostly happens inside infected cells (which have proteins from the pathogen on their cell surfaces). Helper T cells release cytokines that help activated T cells bind to the infected cell, which then allows the T cell to break down the infected cell. Some infections can be successfully eliminated through cellular immunity without also showing a strong humoral immune response.

Scientific American provides a nice visual guide to virus infection and immune processes.

This week’s highlight focuses on humoral immunity.

Antibody tests have varying accuracy

Antibody tests are designed to detect if a person has generated an immune response to the infection - typically measured by quantifying the amount and type of antibodies that react to specific viral proteins.  

A study from April (as yet not published) found that 12 serological tests for the virus gave variable results. Accuracy improved when at least 2 weeks had passed since symptoms appeared. Antibody detection was greater for samples from hospitalised patients (compared to people with milder symptoms). While specificity was greater than 95% for the majority of tests, not all tests gave similar results when tested on the same samples. 

A systematic review of over 50 studies of antibody tests has also just been published by the Cochrane Library. This noted the usefulness of antibody testing between 15 and 35 days after symptom onset. However it also reports differences in accuracy, both for the type of test and the circumstances in which it is used (eg, for research or for clinical use). It suggests that the design, execution and reporting of studies of the accuracy of COVID‐19 tests requires considerable improvement.

The British Medical Journal provides an overview of antibody testing.

Improving antibody tests for Covid-19

A research team in the UK is developing a more sensitive Enzyme-Linked Immunosorbent Assay to detect antibodies that target SARS-CoV-2. Their recent paper (not yet peer reviewed) found that testing for antibodies that react to the complete spike protein were the most accurate (compared to parts of the spike protein and the nucleocapsid protein) in distinguishing infected from uninfected patients. The spike protein could also be detected in saliva samples as well as from serum, whereas the N-protein was not detected in saliva.

Testing serum and saliva samples from the same individual didn’t always give the same result, so the authors suggest testing both serum and saliva to confirm the presence of antibodies.

However, another research team developed a test and found that IgG antibody results from serum and saliva were consistent (study also not yet peer reviewed). They suggest that a saliva test will make antibody testing easier.

These conflicting results reinforce the Cochrane Library’s conclusion about room for improvement.

Most infected people seem to develop an antibody response

Evidence to date indicates that most people, but not everyone, infected produces antibodies against SARS-CoV-2. At least if tested two weeks after symptoms appear.

A study in New York City (not yet peer reviewed) found nearly everyone who was confirmed to have the virus tested positive (strongly or weakly) for IgG antibodies after one or two tests. Only 3 of 514 people (0.6%) didn’t appear to produce IgG antibodies. Forty nine other people who initially had negative or weak antibody results did not come back for further testing. 

Another study (also not yet peer reviewed) of 177 mostly hospitalised Covid-19 patients who were tested  at least twice found that a small percentage (2 to 8.5%, depending on how long patients were followed up) did not appear to develop IgG antibodies.

On the other hand, a small study of seven families (pre-print paper) found that family members who became infected from another person in the house did not produce antibodies. They did, however, develop a weak T cell response to the infection. The authors note that a T cell response without antibody production has been can be found previously for other types of infection.

However, antibodies can be lower in people with mild or no Covid-19 symptoms

A study of 160 hospital workers in France (a pre-print paper) found that for people with milder symptoms the immune response appeared to strengthen over time. Detection of neutralising antibodies was more common if testing occurred 28-41 days after symptoms appeared. 

In a study of 500 hospital workers in England (also a pre-print) people who self-reported Covid-19 symptoms were more likely to have antibodies than those who did not report symptoms (35% vs 17%). Levels of antibodies were also greater in those who reported symptoms. A limitation of this study is that most of the subjects did not have earlier PCR tests that confirmed that they had been infected.

Research recently published in Nature found that asymptomatic subjects had lower levels of IgG than people displaying symptoms. They also report that 40% of the asymptomatic and 12.9% of those showing symptoms lost the antibodies over the next few weeks or months. However, this was a small study, with only 37 infected people.

Immunity may be short-lived

Antibody testing of thousands of people has occurred in Wuhan, where most people were assumed to have become infected. The authors of a pre-print paper report that more than 10% of confirmed COVID-19 cases had no detectable serum levels of IgG antibodies to the virus 21 days after symptoms began. Few healthcare workers (who the authors assume were exposed to the virus) were found to have IgG antibodies, leading them to conclude that immunity is short-lived.

A study (not yet peer reviewed) that has examined immune responses to human coronavirus infections over 35 years also concludes that immunity is short lived. They found antibody levels fell substantially six months after infection, and that people often became re-infected the next year. Whether that holds for the novel SARS-CoV-2 is uncertain.

How effective are the antibodies in controlling infection?

Antibodies are a good sign that someone has been infected by the virus, but they do not necessarily indicate how effective they are at controlling infection.

A paper published in Nature reports that sera from 149 infected people varied in their ability to neutralise the virus in laboratory tests. However, they also discovered that all the subjects had rare antibodies that target the receptor binding domain of the spike protein  and these were very effective in neutralising the virus. Treatments that could bolster this antibody may provide a stronger immune response.

There may be some protection from previous coronavirus infections

There are also some reports (in pre-print) that T cells generated in response to previous human coronavirus infections may also bind to SARS-CoV-2, although less effectively. The extent to which they provide some cross-immunity is uncertain at this stage. Given the large numbers being infected by SARS-CoV-2 in some countries, any effect may be small.

On the other hand, previous coronavirus infections may inhibit a strong immune response

The Hoskins Effect (also called “original antigenic sin”) is an immune response previously seen in influenza and dengue fever infections where children (who have less experienced immune systems) can mount more targeted immune responses to novel infections. 

This is also reported to have been observed in a laboratory study where proteins from different coronaviruses were tested against sera collected before the pandemic. In an as yet to be peer-reviewed paper antibodies from young people (under 19 years) were more effective at targeting the SARS-CoV-2 proteins compared with antibodies taken from healthy people over 65. The over 65’s were more effective in targeting a common human coronavirus. Healthy adults between 20 and 65 overlapped the results of the other two groups, which doesn’t appear to make the results deterministic.

These researchers propose that multiple exposures to coronaviruses over time may slow the immune response to novel coronaviruses. This is a hypothesis that needs further testing.

Caveats on current antibody testing

As some of these examples show, there are still apparent contradictions or knowledge gaps about immune responses to Covid-19. Better antibody tests are likely to appear over the coming months, which will improve knowledge about who has been infected, and how long antibodies to SARS-CoV-2 may persist.

What is also required is more longitudinal studies that follow the same people over several months to better understand the dynamics of immune responses.

Research on cellular immune responses to Covid-19 is also starting to provide a better understanding of how effective immune responses are in different groups of patients.

Call for a global system to monitor immune systems

The pandemic has prompted some researchers to call for the development of a Global Immunological Observatory. This would help improve serological testing, and improve preparedness for future pandemics.

Such an Observatory would involve routine collection of seasonal serum samples to develop a baseline immunological picture; repeated sampling from different age groups to understand immune correlates with susceptibility or protection; and regular immune-surveillance of other species (such as bats) to provide early warnings of potential emerging diseases.

The Observatory would also stimulate development of improved surveillance tools. The cost isn’t assessed, but would be considerable. International scientific and political collaboration would be essential.

Other SMC resources