Keyla Santos Guedes de Sa 1,2,*, Julio Silva 1,2,*, Rafael Bayarri-Olmos 1,2,*, Ryan Brinda 1,2, Robert Alec Rath Constable 1,2, Patricia A Colom Diaz 1,2, Dong-il Kwon 1,2,3, Gisele Rodrigues 1,2, Li Wenxue 2,4, Christopher Baker 1,2, Bornali Bhattacharjee 1,2, Jamie Wood 5, Laura Tabacof 5, Yansheng Liu 2,4, David Putrino 5,†, Tamas L Horvath 6,†, Akiko Iwasaki 1,2,3,†
Author information Copyright and License information
PMCID: PMC11213106 PMID: 38947091
The complete version history of this preprint is available at medRxiv.
Summary
Acute SARS-CoV-2 infection triggers1 the generation of diverse and functional autoantibodies (AABs), even after mild cases. Persistently elevated autoantibodies have been found in some individuals with long COVID (LC). Using a >21,000 human protein array, we identified diverse AAB targets in LC patients that correlated with their symptoms. Elevated AABs to proteins in the nervous system were found in LC patients with neurocognitive and neurological symptoms. Purified Immunoglobulin G (IgG) samples from these individuals reacted with human pons tissue and were cross-reactive with mouse sciatic nerves, spinal cord, and meninges. Antibody reactivity to sciatic nerves and meninges correlated with patient-reported headache and disorientation. Passive transfer of IgG from patients to mice led to increased sensitivity and pain, mirroring patient-reported symptoms. Similarly, mice injected with IgG showed loss of balance and coordination, reflecting donor-reported dizziness. Our findings suggest that targeting AABs could benefit some LC patients.
Keywords: Autoantibodies, Long COVID, Chronic Pain, SARS-CoV-2
_________________________________________________________________________
Published: 22 April 2025
Unequal access to diagnosis of myalgic encephalomyelitis in England
Gemma Louise Samms & Chris P. Ponting
BMC Public Health volume 25, Article number: 1417 (2025)
Abstract
Background
People with Myalgic Encephalomyelitis (ME/CFS; sometimes referred to as chronic fatigue syndrome) experience poor health-related quality of life and only rarely recover. ME/CFS has no curative treatment, and no single diagnostic test. Public health and policy decisions relevant to ME/CFS require knowledge of its prevalence and barriers to diagnosis. However, people with ME/CFS report lengthy diagnostic delays and prevalence estimates vary greatly due to uneven diagnosis and misdiagnosis. Factors that influence diagnosis could be revealed by stratifying a single population by gender, age and ethnicity.
Methods
Hospital Episode Statistics data, routinely collected by the NHS in England, was downloaded from the Feasibility Self-Service of NHS DigiTrials. This was used to stratify individuals with the ICD-10 code that best reflects ME/CFS symptoms (G93.3) according to age, self-reported gender and ethnicity, General Practice and NHS England Integrated Care Board (ICB).
Results
In all, 100,055 people in England had been diagnosed with ME/CFS (ICD-10:G93.3) between April 1 1989 and October 7 2023, 0.16% of all registered patients. Of these, 79,445 were females and 20,590 males, a female-to-male ratio of 3.88:1. Female relative to male prevalence peaked at about 6-to-1 in individuals’ fourth and fifth decades of life. Prevalence varied widely across the 42 ICBs: 0.086%-0.82% for females and 0.024%-0.21% for males. White individuals were approximately fivefold more likely to be diagnosed with ME/CFS than others; Black, Asian or Chinese ethnicities are associated with particularly low rates of ME/CFS diagnoses. This ethnicity bias is stronger than for other common diseases. Among active English GP practices, 176 (3%) had no registered ME/CFS patients. Eight ICBs (19%) each contained fewer than 8 other-than-white individuals with a G93.3 code despite their registers containing a total of 293,770 other-than-white patients.
Conclusion
Other-than-white ethnic groups, older females (> 60y), older males (> 80y), and people living in areas of multiple deprivation are disproportionately undiagnosed with ME/CFS. Lifetime prevalence of ME/CFS for English females and males may be as high as 0.92% and 0.25%, respectively, or approximately 404,000 UK individuals overall (0.6%). This improved estimate of ME/CFS prevalence allows more accurate assessment of the socioeconomic and disease burden imposed by ME/CFS.
_________________________________________________________________________
Indistinguishable mitochondrial phenotypes after exposure of healthy myoblasts to myalgic encephalomyelitis or control serum
Audrey A. Ryback, Charles Hillier, Camila M Loureiro, Chris P Ponting, Caroline F Dalton
doi: https://doi.org/10.1101/2025.06.03.657595
This article is a preprint and has not been certified by peer review
Abstract
Myalgic Encephalomyelitis (ME)/Chronic Fatigue Syndrome is a disease of uncertain aetiology that affects up to 400,000 individuals in the UK. Exposure of cultured cells to the sera of people with ME has been proposed to cause phenotypic changes in these cells in vitro when compared to sera from healthy controls. ME serum factors causing these changes could inform the development of diagnostic tests. In this study, we performed a large-scale, pre-registered replication of an experiment from Fluge et al (2016) that reported an increase in maximal respiratory capacity in healthy myoblasts after treatment with serum from people with ME compared to serum from healthy controls. We replicated the original experiment with a larger sample size, using sera from 67 people with ME and 53 controls to treat healthy cultured myoblasts, and generated results from over 1,700 mitochondrial stress tests performed with a Seahorse Bioanalyser. We observed no significant differences between treatment with ME or healthy control sera for our primary outcome of interest, oxygen consumption rate at maximal respiratory capacity. Results from our study provide strong evidence against the hypothesis that ME blood factors differentially affect healthy myoblast mitochondrial phenotypes in vitro.
Competing Interest Statement
The authors have declared no competing interest.
Funder Information Declared
Action for ME, https://ror.org/0569v7v35, Clare Francis Research Fellowship
bioRxiv and medRxiv thank the following for their generous financial support:
The Chan Zuckerberg Initiative, Cold Spring Harbor Laboratory, the Sergey Brin Family Foundation, California Institute of Technology, Centre National de la Recherche Scientifique, Fred Hutchinson Cancer Center, Imperial College London, Massachusetts Institute of Technology, Stanford University, The University of Edinburgh, University of Washington, and Vrije Universiteit Amsterdam.
Posted June 06, 2025.
_________________________________________________________________________
Precision phenotyping for curating research cohorts of patients with unexplained post-acute sequelae of COVID-19
Authors: Azhir A, Hügel J, Tian J, Cheng J, Basset IV, Bell DS, … Estiri H (Massachusetts General Hospital, USA)
Name and Date of Publication: Med. 14 March 2025
Link: https://doi.org/10.1016/j.medj.2024.10.009
Easy Read Overview: Scientists studied health records to create a better tool for finding and predicting long COVID (PASC) symptoms, which current methods often miss or get wrong. Their new algorithm works more accurately across different ages, genders, and ethnic groups, even spotting rare symptoms like hearing loss and diabetic issues. It can help doctors diagnose long COVID more fairly and correctly, and the tool is free for use in any healthcare system.
Current diagnostic codes for post-acute sequelae of SARS-CoV-2 (PASC) are often inaccurate, biased across demographics, and underestimate their true prevalence. Similarly, the criteria to predict those most at risk of developing post-acute sequelae of SARS-CoV-2 (PASC) are inadequate. The authors conducted a retrospective longitudinal case-control study to develop a more accurate precision phenotyping algorithm. This tool aims to not only improve diagnosis but also predict the onset, affected systems, and duration of PASC across different demographic groups.
The study analysed the electronic health records (EHR) of three groups: 85,364 confirmed COVID-19 patients (at least one year post-infection), 170,497 post-pandemic individuals without COVID-19, and 39,817 pre-pandemic controls. A Sequential Pattern Mining algorithm, based on WHO’s PASC definition and Clinical Classifications Software, was used to sift through this data. The outputs were independently reviewed and refined.
Compared to the International Classification of Diseases U09.9 diagnosis code, the new algorithm achieved 79.9% diagnostic precision—an improvement of 2.7%—and more accurately estimated community prevalence. It effectively excluded false positives (due to pre-existing conditions) and identified false negatives (conditions misdiagnosed, but linked to prior COVID-19). The breadth of data allowed the algorithm to detect rarer sequelae beyond common symptoms like fatigue or sleep issues, including vision and hearing loss, diabetic complications, and sexual dysfunction.
The algorithm also provided a more balanced demographic analysis, which allowed for better analysis of which sequelae are more common based on age, gender, and ethnicity. For example, those under 45 were more likely to develop gynaecological, dermatological, or psychiatric sequelae; women had higher odds of PASC in certain organs; Asian and Hispanic patients had lower PASC risk; while Black patients faced higher risk regardless of age or health status.
The authors emphasise that their algorithm aligns with the National Academies of Sciences, Engineering, and Medicine’s (NASEM) newly proposed broad definition of PASC as an infection-associated chronic condition (IACC). They have made their tools and data publicly available for integration into any healthcare system.
_________________________________________________________________________
Comparative Study Between Cognitive Phenotypes of Myalgic Encephalomyelitis /Chronic Fatigue Syndrome and Multiple Sclerosis
Authors: Sebaiti MA, Oubaya N, Gounden Y, Samson C, Lechapt E, Wahab A … & Authier F-J (Paris-East Creteil University, France)
Name and Date of Publication: Diagnostics. 17 February, 2025
Link: https://pmc.ncbi.nlm.nih.gov/articles/PMC11854609/pdf/diagnostics-15-00487.pdf
Easy Read Overview: This study compared thinking and memory problems in people with ME/CFS and multiple sclerosis (MS). It found that both groups had trouble with memory and attention, but people with ME/CFS had more difficulty remembering new information. The results suggest that special brain training could help people with ME/CFS improve their thinking skills.
ME/CFS involves a range of disabling symptoms, with cognitive impairments being one of the most commonly reported. The aim of this study was to identify a specific cognitive profile unique to ME/CFS. Multiple sclerosis (MS) was used as a comparison group due to the overlapping symptoms of pain, cognitive impairment and fatigue.
A retrospective analysis of patient data from the Henri Mondor Hospital in France was undertaken. Data from 40 participants with ME/CFS (Fukuda criteria), and 40 participants with MS (relapsing-remitting) were included in this study. All participants had completed a screening of cognitive function and neuropsychological tests. Neuropsychological tests assessed short and long-term memory, visual and auditory processing, processing speed, executive function, working memory, and planning ability. All participants completed the tests in the same order.
The authors found that both ME/CFS and MS participant groups showed deficits in episodic memory retrieval, visual selective attention, and reading speed. ME/CFS participants were also found to have a lower performance in consolidation processes than MS participants. In both groups, performance on the cognitive tests was not related to levels of fatigue, pain and depression.
The authors conclude that this study highlights the similarities and differences in cognitive profiles of ME/CFS and MS patients. The impairment in consolidation processes shown by ME/CFS participants helps define a specific cognitive phenotype for ME/CFS, especially as this was not correlated with pain, fatigue or depression. The authors suggest that future research, especially with functional imaging, may identify the neurobiological mechanism causing this. The authors also propose that this research can be used to assist the management of ME/CFS, by considering implementing cognitive training with a focus on verbal learning strategies and working memory exercises.
_________________________________________________________________________
Untargeted metabolomics and quantitative analysis of tryptophan metabolites in myalgic encephalomyelitis patients and healthy volunteers: A comparative study using high-resolution mass spectrometry
Authors: Abujrais S, Vallianatou T, and Bergquist J (Uppsala University, Sweden)
Name and Date of Publication: ACS Chemical Neuroscience. September 20, 2024
Link: https://doi.org/10.1021/acschemneuro.4c00444
Easy Read Overview: Researchers studied people with ME/CFS and found changes in how their bodies process tryptophan, a substance important for energy, mood, and the immune system. They found that different patients had different changes, but some common chemical differences were seen in all ME/CFS patients. These results may help doctors find better ways to diagnose and treat ME/CFS in the future.
The tryptophan metabolic pathway has received attention in ME/CFS research due to its involvement in immune function, neurotransmission and energy production. These authors used untargeted metabolomics – scanning a biological sample for a wide range of chemical compounds, without choosing them in advance – as well as targeted analysis of tryptophan and its metabolites, to gain a better understanding of the altered metabolic pathways underlying ME/CFS.
Plasma samples were obtained from 19 patients from Stora Sköndal in Stockholm (ME-SK), and 19 patients from Gottfries Clinic (ME-GC), as well as 24 healthy controls (HC). The 38 ME/CFS patients met three diagnosis criteria – Canadian Consensus Criteria, International Consensus Criteria, and Institute of Medicine (IOM) criteria. A pooled plasma sample was created for quality control (QC). Liquid chromatography-high-resolution mass spectrometry (LC-HRMS) was used for both targeted and untargeted analysis.
Analysis found significant differences between the two ME/CFS cohorts, suggesting the metabolic impact of ME/CFS can vary widely, however, 30 common significantly altered metabolites were identified. The untargeted analysis found dysregulation of quinolinic acid and indoleacetic acid (tryptophan metabolic products), L-adrenaline and S-adenosyl-L-homocysteine (SAH), and the vitamin B3, the arginine-proline and the aspartate-asparagine pathways.
The targeted analysis of tryptophan metabolites and related compounds also identified differences between ME-SK and ME-GC, as well as between ME/CFS and HC. The analysis found significantly altered levels of hypoxanthine, a biomarker for hypoxia, indicative of reduced oxygen extraction in ME/CFS; nicotinamide and riboflavin, which are important in energy production; and phenylalanine, involved in neurotransmission. Significant alteration were also observed in metabolites within the serotonin pathway, which is involved in mood and sleep regulation, and in the kynurenine pathway, which is associated with neurotransmission, inflammation and immune response (with alteration of 3-hydroxyanthranilic acid (3HAA) posing significant health implications). Differences in some metabolites between sex and/or age indicated potential age-related changes, as well as biological differences between male and female ME/CFS presentation.
This study identified several potential biomarkers, as well as potential targets for the treatment of ME/CFS. The authors recommend that further studies, particularly into the kynurenine pathway, use similar sample sizes and analytic methods to avoid discrepancies and to examine the effect of medications, as well as the menstrual cycle, on the metabolites.
_________________________________________________________________________
ME/Chronic fatigue syndrome: The mysterious illness trapping people in their bodies
Authors: Madden-Smith Z
Name and Date of Publication: Re: News. May 12, 2025
Link: https://youtu.be/DsOAq6cs564?si=1Z0kkbLvX9FjAO40
This New Zealand video news story explores the impact of severe ME/CFS on those who live with the condition as well as their carers. It highlights the difficulty getting access to sufficient care in New Zealand and one mother’s solution in setting up a shared care system.
“Someone who is so physically unwell is being treated so poorly, as if they can just snap out of it. I don’t think I can think of another condition that would be treated this way. It just blows my mind,” said researcher Dr Anna Brooks.
“I just don’t think we’ve got to grips with this situation. Why is this not an emergency?” says ME/CFS educator Rose Silvester.
The video runs for 16 minutes and includes some music that may be difficult for some to tolerate.
_________________________________________________________________________
My wife has largely been bedbound for decades. Then our child joined her in isolation
Authors: McCluskey P
Name and Date of Publication: SBS News. 20 May 2025
Link: https://www.sbs.com.au/news/insight/article/toll-of-being-bedbound-with-me-cfs-chronic-fatigue-syndrome/dew0m9ikt
Peter McCluskey is carer for his wife and daughter, who both live with ME/CFS. In this article, he writes about his experience.
“The hardest part of being an ME/CFS carer is witnessing relentless suffering.
The quality of life for those with severe ME/CFS is often compared to that of late-stage cancer. But with this syndrome, the suffering is unending.
I effectively have a front row seat to a misery show that just goes on and on.
Equally challenging is communicating with some health professionals, who dismiss or downplay the illness, leaving carers and patients feeling invalidated and hopeless.
This deepens the isolation of managing this illness alone, amplifying the emotional toll on carers who must constantly advocate against disbelief while supporting their loved ones.”
_________________________________________________________________________
Skeletal muscle properties in long COVID and ME/CFS differ from those induced by bed rest
Braeden T. Charlton , Anouk Slaghekke , Brent Appelman , Moritz Eggelbusch , Jelle Y. Huijts , Wendy Noort , Paul W. Hendrickse , Frank W. Bloemers , elle J. Posthuma , Paul van Amstel , Richie P. Goulding , Hans Degens , Richard T. Jaspers , Michele van Vugt , Rob CI Wüst
doi:https://doi.org/10.1101/2025.05.02.25326885
This article is a preprint and has not been peer-reviewed This is new medical research that has yet to be evaluated and so should not be used to guide clinical practice.
Abstract
Patients with long COVID and myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) suffer from a reduced exercise capacity, skeletal muscle abnormalities and post-exertional malaise (PEM), where worse symptoms with cognitive or physical exertion. PEM often results in avoidance of physical activity, resulting in a lower aerobic fitness, which may contribute to skeletal muscle abnormalities. Here, we compared whole-body exercise responses and skeletal muscle adaptations after strict 60-day bed rest in healthy people with those in patients with long COVID and ME/CFS, and healthy age- and sex-matched controls. Bed rest altered the respiratory and cardiovascular responses to (sub)maximal exercise, while patients demonstrated respiratory alterations only at submaximal exercise. Bed rest caused muscle atrophy, and the reduced oxidative phosphorylation related to reductions in maximal oxygen uptake. Patients with long COVID and ME/CFS did not have muscle atrophy, but had fewer capillaries and a more glycolytic fibers, none of which were associated with maximal oxygen uptake. While the whole-body aerobic capacity is similar following bed rest compared to patients, the skeletal muscle characteristics differed, suggesting that physical inactivity alone does not explain the lower exercise capacity in long COVID and ME/CFS.
• Systrom at the CDC: The ME/CFS Mitochondrial Drug Trial Fails Plus More
- The failure of the mitochondrial drug trial was unfortunate, but the data gained during the trial could help explain a lot about key problems such as fatty acid metabolism in ME/CFS.
- While the Astellas drug failed, the recent one-off Mestinon study succeeded in increasing energy production (VO2 max) during exercise in ME/CFS patients, apparently by affecting blood flows. In fact, three of the five drugs Systrom reported he uses in ME/CFS affect blood flows in one way or the other.
- Preload failure – the inability to return normal amounts of blood back to the heart – is the signature invasive exercise finding in ME/CFS – and is also commonly found in POTS and people with small fiber neuropathy.
- Preload failure is also accompanied by problems with oxygen extraction by the mitochondria and/or right/left shunting of blood away from the muscles. In one, mitochondrial failure is the cause; in the other, blood vessel problems are – and the treatments for each are completely different.
- While it’s not clear that shunting is present in ME/CFS and long COVID, a fibromyalgia study found evidence that more blood than usual was being shunted into the hands of FM patients and was getting stuck there. Since the hands serve as reservoirs of blood for the rest of the body, getting blood stuck in the hands could negatively affect the blood flows needed for exercise, concentration, etc.
- While attempts to link small fiber neuropathy to the exercise problems in ME/CFS have thus far failed, more extensive assessments of the small nerve fibers are underway and could yield different results.
- A trend towards excessive activation of the TRAIL inflammatory pathway, and inflammatory activity, in general, was upregulated in ME/CFS during exercise. Interestingly, Nancy Klimas’s supercomputer studies plucked out a drug called etanercept that knocks down the end of the TRAIL pathway line – a powerful inflammatory cytokine called TNF-a. A case report of a severe long-COVID patient found that drug to be helpful.
medRxiv
[Preprint]. 2025 Apr 17:2025.04.16.25325949. [Version 1] doi: 10.1101/2025.04.16.25325949
Persistent immune dysregulation and metabolic alterations following SARS-CoV-2 infection
Silvia Lucena Lage 1,*,†, Katherine Bricker-Holt 1,†, Joseph M Rocco 1, Adam Rupert 2, Frank X Donovan 3, Yevgeniya A Abramzon 3, Settara C Chandrasekharappa 3, Colton McNinch 4, Logan Cook 1, Eduardo Pinheiro Amaral 5, Gabriel Rosenfeld 4, Thomas Dalhuisen 6, Avery Eun 6, Rebecca Hoh 6, Emily Fehrman 6, Jeffrey N Martin 7, Steven G Deeks 6, Timothy J Henrich 6, Michael J Peluso 6, Irini Sereti 1,*
Copyright and License information PMCID: PMC12047922 PMID: 40321289
The complete version history of this preprint is available at medRxiv.
Abstract
SARS-CoV-2 can cause a variety of post-acute sequelae including Long COVID19 (LC), a complex, multisystem disease characterized by a broad range of symptoms including fatigue, cognitive impairment, and post-exertional malaise. The pathogenesis of LC is incompletely understood. In this study, we performed comprehensive cellular and transcriptional immunometabolic profiling within a cohort that included SARS-CoV-2-naïve controls (NC, N=30) and individuals with prior COVID-19 (~4-months) who fully recovered (RC, N=38) or went on to experience Long COVID symptoms (N=58). Compared to the naïve controls, those with prior COVID-19 demonstrated profound metabolic and immune alterations at the proteomic, cellular, and epigenetic level. Specifically, there was an enrichment in immature monocytes with sustained inflammasome activation and oxidative stress, elevated arachidonic acid levels, decreased tryptophan, and variation in the frequency and phenotype of peripheral T-cells. Those with LC had increased CD8 T-cell senescence and a distinct transcriptional profile within CD4 and CD8 T-cells and monocytes by single cell RNA sequencing. Our findings support a profound and persistent immunometabolic dysfunction that follows SARS-CoV-2 which may form the pathophysiologic substrate for LC. Our findings suggest that trials of therapeutics that help restore immune and metabolic homeostasis may be warranted to prevent, reduce, or resolve LC symptoms.
Keywords: post-acute sequelae of SARS-CoV-2, COVID-19, Long COVID, inflammation, metabolism, immunosenescence
_________________________________________________________________________
Skeletal muscle properties in long COVID and ME/CFS differ from those induced by bed rest
View ORCID ProfileBraeden T. Charlton , View ORCID ProfileAnouk Slaghekke , View ORCID ProfileBrent Appelman , View ORCID ProfileMoritz Eggelbusch , Jelle Y. Huijts , View ORCID ProfileWendy Noort , View ORCID ProfilePaul W. Hendrickse , View ORCID ProfileFrank W. Bloemers , View ORCID ProfileJelle J. Posthuma , View ORCID ProfilePaul van Amstel , View ORCID ProfileRichie P. Goulding , View ORCID ProfileHans Degens , View ORCID ProfileRichard T. Jaspers , View ORCID ProfileMichele van Vugt , View ORCID ProfileRob CI Wüst
doi:https://doi.org/10.1101/2025.05.02.25326885
This article is a pre-print. It reports new medical research that has yet to be evaluated and so should not be used to guide clinical practice.
Abstract
Patients with long COVID and myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) suffer from a reduced exercise capacity, skeletal muscle abnormalities and post-exertional malaise (PEM), where worse symptoms with cognitive or physical exertion. PEM often results in avoidance of physical activity, resulting in a lower aerobic fitness, which may contribute to skeletal muscle abnormalities. Here, we compared whole-body exercise responses and skeletal muscle adaptations after strict 60-day bed rest in healthy people with those in patients with long COVID and ME/CFS, and healthy age- and sex-matched controls. Bed rest altered the respiratory and cardiovascular responses to (sub)maximal exercise, while patients demonstrated respiratory alterations only at submaximal exercise. Bed rest caused muscle atrophy, and the reduced oxidative phosphorylation related to reductions in maximal oxygen uptake. Patients with long COVID and ME/CFS did not have muscle atrophy, but had fewer capillaries and a more glycolytic fibers, none of which were associated with maximal oxygen uptake. While the whole-body aerobic capacity is similar following bed rest compared to patients, the skeletal muscle characteristics differed, suggesting that physical inactivity alone does not explain the lower exercise capacity in long COVID and ME/CFS.
_________________________________________________________________________
Resource 19 June 2025 Open access Transparent process
Replicated blood-based biomarkers for myalgic encephalomyelitis not explicable by inactivity
Sjoerd Viktor Beentjes https://orcid.org/0000-0002-7998-4262 [email protected], Artur Miralles Méharon https://orcid.org/0009-0006-7977-3955, Julia Kaczmarczyk, Amanda Cassar, Gemma Louise Samms, Nima S Hejazi, Ava Khamseh https://orcid.org/0000-0001-5203-2205 [email protected], and Chris P Ponting https://orcid.org/0000-0003-0202-7816 [email protected]Author Information
EMBO Mol Med (2025) https://doi.org/10.1038/s44321-025-00258-8
Abstract
Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a common female-biased disease. ME/CFS diagnosis is hindered by the absence of biomarkers that are unaffected by patients’ low physical activity level. Our analysis used semi-parametric efficient estimators, an initial Super Learner fit followed by a one-step correction, three mediators, and natural direct and indirect estimands, to decompose the average effect of ME/CFS status on molecular and cellular traits. For this, we used UK Biobank data for up to 1455 ME/CFS cases and 131,303 controls. Hundreds of traits differed significantly between cases and controls, including 116 significant for both female and male cohorts. These were indicative of chronic inflammation, insulin resistance and liver disease. Nine of 14 traits were replicated in the smaller All-of-Us cohort. Results cannot be explained by restricted activity: via an activity mediator, ME/CFS status significantly affected only 1 of 3237 traits. Individuals with post-exertional malaise show stronger biomarker differences. Single traits could not cleanly distinguish cases from controls. Nevertheless, these results keep alive the future ambition of a blood-based biomarker panel for accurate ME/CFS diagnosis.