Finding an accepted Long COVID biomarker will be complicated. This blood test could give us answers.

It seems like every week a potential biomarker for Long COVID emerges from the towering stacks of scientific studies on the disease. Various biochemical, vascular, and neurological markers have shown potential as diagnostic biomarkers —  but nothing has been widely accepted. At least, not yet.

Like other diseases that lack accepted biomarkers including Myalgic Encephalomyelitis (ME), Long COVID is more difficult to study as a result. But finding one will deepen our understanding of the disease and speed up drug development. One thing we know: biomarkers, which are measurable traits in the body, can take time to develop and must be validated in a thorough scientific process.

Still, despite limited global government funding and the many challenges of studying complex diseases like Long COVID, scientists may be closing in on biomarkers for the disease.

Last year, immunologist Johan Van Weyenbergh published a study in The Lancet: Microbe that identified a potential blood-based biomarker for Long COVID with 94% sensitivity. Van Weyenbergh’s team discovered SARS-CoV-2 RNA transcripts in the blood cells of people with Long COVID. They also found a specific type of RNA, called ORF1ab RNA, which suggested there could also be ongoing viral replication of SARS-C0V-2 in people with the disease.

“I can see a number of ways where this [potential biomarker] might be useful and warrants further investigation,” said Annie Antar, an assistant professor of medicine at the Johns Hopkins University School of Medicine who researches HIV and Long COVID. 

“A more full characterization of the biomarker will be really important,” said Antar, who is not affiliated with Van Weyenbergh’s research.

For a biomarker to be widely accepted, it must undergo an extensive process in which scientists validate their findings with larger groups of patients. Then, other scientists replicate those results in their own independent cohorts.

Even when validated through this process, biomarkers can exclude some people who may have a disease. For example, if the biomarker from Van Weyenbergh’s study continues to have 94% sensitivity, that means it will correctly identify Long COVID in 94% of people with the disease — but will miss the other 6%.

Despite Van Weyenbergh’s promising research, his team ran out of funding to validate the finding in a larger cohort last fall. His encouraging study could lead to a biomarker for a disabling disease affecting over 400 million people globally, but he couldn’t find support from European governments, many of which have notoriously underfunded research into Long COVID. 

“A lot of long haulers are going overlooked or being misdiagnosed because we don’t have tests [for Long COVID],” said Devin Russell, the CEO of Long COVID Foundation. When Russell came across Van Weyenbergh’s study last year, he knew it was something he wanted his non-profit to support.

Together with activist group Long COVID Action Project, they raised over $34,000, primarily from people with the disease. Van Weyenbergh’s research also caught the attention of the leading Long COVID research group the Polybio Research Foundation, which he said further supported his research with a $159,000 grant.*

Now, Van Weyenbergh is working with his team at the Rega Institute for Medical Research in Leuva, Belgium on confirming their original findings. 

“To get a biomarker, we have to validate it in [an] independent cohort,” he said.

The potential RNA biomarker

In the original study, published in April, Van Weyenbergh’s team discovered SARS-CoV-2 RNA transcripts in the blood cells of people with Long COVID. They also found antisense ORF1ab RNA. The researchers wrote that the upregulation of this strand of RNA suggests there could also be ongoing viral replication of SARS-C0V-2 in people with the disease.

A growing body of research has found SARS-C0V-2 in many reservoirs in the body including the skull bone marrow, gut, and various other tissues. The presence of a persistent SARS-CoV-2 virus, especially a replicating one, could impact every system of the body, driving a downstream effect that may contribute to symptoms — a theory that researchers call viral persistence.  It’s one of the leading hypotheses of how the chronic disease occurs.

But it’s often difficult and expensive for researchers to study viral reservoirs. Since SARS-CoV-2 can hide in host cells, scientists must find it through invasive biopsies in places like the lymph nodes and gut, as researchers at the University of California, San Francisco have done in the Long COVID LIINC study.

While procedures like this have led to important discoveries that advance our understanding of Long COVID, they aren’t always the best biomarkers. They’re often too expensive and timely, not to mention uncomfortable for people with Long COVID.

Ideally, biomarkers should be “non-invasive and affordable,” Van Weyenbergh said, such as a blood test or saliva swab.

To find the presence of SARS-CoV-2 in people with Long COVID, his team used digital transcriptomics, a process of studying collections of RNA transcripts in a cell or tissue. By assessing the copies of these genes, transcriptomics can help researchers understand why some diseases develop and identify potential diagnostic markers, among other uses. 

“Transcriptomics is a snapshot of the immune system,” Van Weyenbergh said. 

In the 2024 study, using whole blood from 48 people with Long COVID and 12 matched controls, Van Weyenbergh’s team found differences in how 212 genes were expressed between the two groups.  Many coronavirus RNAs were upregulated, including nucleocapsid, ORF7a, ORF3a, Mpro — a target for the antiviral drug Paxlovid — and antisense ORF1ab RNA, the marker suggesting viral replication. 

“This isn’t meaningless fragments at all,” said Amy Proal, president of Polybio Research Foundation. “The virus is there and it’s doing something.”

Polybio and its Long COVID Research Consortium — an international network of prominent Long COVID researchers — focus their attention on viral reservoirs. Studies funded by Polybio have found persistent SARS-C0V-2 virus in the gut, lungs, and lymph nodes. Another research team supported by Polybio found persistent virus in platelets associated with bone marrow cells, which could cause fibrinoid microclots in people with Long COVID. When Proal came across Van Weyenbergh’s study, she thought it was important for the field and knew her foundation needed to support it.

Part of the success of the study is that Van Weyenbergh and his team used whole blood, or blood that has not been separated out into different components, Proal said. It’s more common for researchers to use “spun” blood or plasma. Using whole blood allowed Van Weyenbergh to capture all fractions of the virus from host cells with probes using digital transcriptomics. 

Validating a biomarker

Using Long COVID blood samples from Polybio Research Consortium partners in France and the U.K., Van Weyenbergh plans to validate his study with around 100 Long COVID samples and 100 controls. 

If his team is able to confirm and replicate the findings in this larger cohort, they’ll likely draw attention from other researchers who would attempt to independently verify the same results. If independent teams are also able to replicate the results, the blood test could head to a diagnostic development space.

“While this has potential to be a really important clinical test and [could be used in clinical trials] were it to replicate,” Proal explained, “it would be one of many tests we’ve run on people with Long COVID.” She said that biomarkers for diseases can leave people out, which is why it is important to compile a fleet of tests. 

Some Long COVID and ME patient advocates have also stated that biomarkers should assess disease severity, measured by independent, objective questionnaires. If the biomarker is able to identify who has and doesn’t have a disease — but not how severely affected patients are by it — they won’t be helpful in clinical drug trials, advocates argue.

In a project summary about the biomarker study, Van Weyenbergh’s team stated their goal is to develop a combination of tests, including digital transcriptomics, proteomics (the study of proteins), and metabolomics (the study of metabolites) that could eventually become a standard blood test for the disease.

A blood test of that caliber could be used in clinical trials. Not only could it help researchers recruit for specific trials, but also give an endpoint, or a measurement that would help scientists understand if a treatment was effective or not. 

While this potential diagnostic RNA biomarker might not pick up everyone with Long COVID, Antar, the HIV and Long COVID physician-scientist not affiliated with the study, explained that studying and identifying a subgroup that doesn’t test positive for a biomarker could lead to better understanding of the disease. 

The subgroup would demonstrate that “different pathways” can lead to Long COVID, Antar said.” Researchers already suspect this is the case, as studies support that other factors such as autoimmunity, reactivated viruses, and inflammation could contribute as drivers of the disease.

While this has potential to be a really important clinical test and [could be used in clinical trials] were it to replicate, it would be one of many tests we’ve run on people with Long COVID.

Amy Proal, Polybio Research Foundation

Great urgency

Validated biomarkers take time and can prove troublesome — especially when they aren’t funded with urgency from government agencies like the National Institutes of Health (NIH). A lack of accepted biomarkers can create a vicious cycle of underfunding and recognition — even with diseases as prevalent and serious as Long COVID and ME. It also can lead to stigma, misinformation, and a lack of safety nets for people with the diseases.

“There is so much stigma, underestimation, and disbelief,” Van Weyenbergh said about Long COVID. People with Long COVID are often relieved to learn he’s found persistent SARS-C0V-2 in their blood, which is different from the reaction he gets from people with other diagnoses, he said.

“Once there’s a test, the disease becomes easier to diagnose, to publicize, and to write about,” wrote Andrea Martell, who tracks and writes about the importance of biomarkers for Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS). She wrote to The Sick Times that biomarkers can turn a patient population into a traceable dataset. “It becomes easier to explain,” she stated. “Humans like things that are simple to understand.” 

Martell believes that accepted biomarkers could change the landscape for ME/CFS, including winning over allies, helping the Food and Drug Administration to approve treatments, and increasing funding to study the immunobiological basis of the disease. “We need to concentrate our funding and advocacy efforts on getting an accurate, sensitive, specific, safe, accessible diagnostic blood test,” she wrote. 

Martell also stated that identifying subtypes within Long COVID and ME/CFS is important in finding accurate biomarkers before performing various immunobiological and multi-omic analyses.

A recent NIH RECOVER paper added weight to the need for a diagnostic biomarker for Long COVID. The Annals of Medicine study of over 10,000 participants found that 25 routine lab tests commonly run in doctors’ offices could not identify Long COVID. The result wasn’t surprising to people with Long COVID, as many have experienced extreme medical gaslighting from providers when common tests come back “normal.”

The RECOVER authors stated in their conclusion that understanding the biological basis of Long COVID will “likely require a rigorous focus on investigations beyond routine clinical laboratory studies (for example, transcriptomics, proteomics, metabolomics) to identify novel biomarkers.” 

Russell, of the Long COVID Foundation, wants to know why something as important as this diagnostic test isn’t the focus of RECOVER. Since its launch in 2021, the large, $1.65 billion dollar NIH study has been criticized by the Long COVID community for largely prioritizing observational research. “We’ve seen this before with chronic Lyme, HIV, and other chronic illnesses, [the NIH] doesn’t seem to put the money in the right place and get the solutions that people need,” Russell said.

As a result, many have placed their hope into non-government funded research. Private researchers like Polybio and their collaborators often show the importance of “high risk, high reward” research that gets to the root cause of a disease. Similarly, other promising biomarker research for ME is coming from the Open Medicine Foundation. The foundation recently announced $800,000 in initial funding for Bioquest, a large-scale study to identify a blood test for the disease using a potential protein and metabolite biomarker.

Still, while many experts in this story expressed excitement about the potential for Van Weyenbergh’s RNA-based biomarker, all remained cautious of getting ahead of the research.

“There are lots of biomarkers out there that all have value,” Van Weyenbergh said. “The best biomarker would be the one that’s more extensively validated.”

[The NIH] doesn’t seem to put the money in the right place and get the solutions that people need.

Devin Russell, Long COVId Foundation

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*Editor’s note: The Polybio Research Foundation shares a funding source (the Balvi and Kanro funds) with The Sick Times. Our newsroom operates independently of financial supporters.

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