Laboratory Findings and Biomarkers in Long COVID: What Do We Know So Far? Insights Into Epidemiology, Pathogenesis, Therapeutic Perspectives and Challenges

Abstract:

Long COVID (LC) encompasses a constellation of long-term symptoms experienced by at least 10% of people after the initial SARS-CoV-2 infection, and so far has affected about 65 million people. The etiology of LC remains unclear; however, many pathophysiological pathways may be involved, including viral persistence; chronic, low grade inflammatory response; immune dysregulation and defective immune response; reactivation of latent viruses; autoimmunity; persistent endothelial dysfunction and coagulopathy; gut dysbiosis; hormonal dysregulation, mitochondrial dysfunction; and autonomic nervous system dysfunction.

There are no specific tests for the diagnosis of LC, and clinical features including laboratory findings and biomarkers may not specifically relate to LC. Therefore, it is of paramount importance to develop and validate biomarkers that can be employed for the prediction, diagnosis and prognosis of LC and its therapeutic response. Promising candidate biomarkers that are found in some patients are markers of systemic inflammation including acute phase proteins, cytokines and chemokines; biomarkers reflecting SARS-CoV-2 persistence, reactivation of herpesviruses and immune dysregulation; biomarkers of endotheliopathy, coagulation and fibrinolysis; microbiota alterations; diverse proteins and metabolites; hormonal and metabolic biomarkers; as well as cerebrospinal fluid biomarkers. At present, there are only two reviews summarizing relevant biomarkers; however, they do not cover the entire umbrella of current biomarkers or their link to etiopathogenetic mechanisms, and the diagnostic work-up in a comprehensive manner.

Herein, we aim to appraise and synopsize the available evidence on the typical laboratory manifestations and candidate biomarkers of LC, their classification based on main LC symptomatology in the frame of the epidemiological and pathogenetic aspects of the syndrome, and furthermore assess limitations and challenges as well as potential implications in candidate therapeutic interventions.

Source: Tsilingiris, D.; Vallianou, N.G.; Karampela, I.; Christodoulatos, G.S.; Papavasileiou, G.; Petropoulou, D.; Magkos, F.; Dalamaga, M. Laboratory Findings and Biomarkers in Long COVID: What Do We Know So Far? Insights Into Epidemiology, Pathogenesis, Therapeutic Perspectives and Challenges. Preprints.org 2023, 2023051487. https://doi.org/10.20944/preprints202305.1487.v1 (Full text available as PDF file)

Musculoskeletal involvement: COVID-19 and post COVID 19

Abstract:

The worldwide pandemic of coronavirus disease 2019 (COVID-19) was known to predominantly affect the lungs, but it was realized that COVID-19 had a large variety of clinical involvement. Cardiovascular, gastrointestinal, neurological, and musculoskeletal systems are involved by direct or indirect mechanisms with various manifestations.

The musculoskeletal involvement can manifest during COVID-19 infection, due to medications used for the treatment of COVID-19, and in the post/long COVID-19 syndrome. The major symptoms are fatigue, myalgia/arthralgia, back pain, low back pain, and chest pain. During the last two years, musculoskeletal involvement increased, but no clear consensus was obtained about the pathogenesis. However, there is valuable data that supports the hypothesis of angiotensinconverting enzyme 2, inflammation, hypoxia, and muscle catabolism. Additionally, medications that were used for treatment also have musculoskeletal adverse effects, such as corticosteroid-induced myopathy and osteoporosis. Therefore, while deciding the drugs, priorities and benefits should be taken into consideration.

Symptoms that begin three months from the onset of the COVID-19 infection, continue for at least two months, and cannot be explained by another diagnosis is accepted as post/long COVID-19 syndrome. Prior symptoms may persist and fluctuate, or new symptoms may manifest. In addition, there must be at least one symptom of infection. Most common musculoskeletal symptoms are myalgia, arthralgia, fatigue, back pain, muscle weakness, sarcopenia, impaired exercise capacity, and physical performance. In addition, the female sex, obesity, elderly patients, hospitalization, prolonged immobility, having mechanical ventilation, not having vaccination, and comorbid disorders can be accepted as clinical predictors for post/long COVID-19 syndrome.

Musculoskeletal pain is also a major problem and tends to be in chronic form. There is no consensus on the mechanism, but inflammation and angiotensin-converting enzyme 2 seem to play an important role. Localized and generalized pain may occur after COVID-19, and general pain is at least as common as localized pain. An accurate diagnosis allows physicians to initiate pain management and proper rehabilitation programs.

Source: Evcik D. Musculoskeletal involvement: COVID-19 and post COVID 19. Turk J Phys Med Rehabil. 2023 Feb 28;69(1):1-7. doi: 10.5606/tftrd.2023.12521. PMID: 37201006; PMCID: PMC10186015.

Influence of Chronic Fatigue Syndrome Codiagnosis on the Relationship between Perceived and Objective Psychoneuro-Immunoendocrine Disorders in Women with Fibromyalgia

Abstract:

Although the predominant symptom in fibromyalgia (FM) is muscle pain, and fatigue in chronic fatigue syndrome (CFS), differential diagnosis is very difficult. This research investigates the psychoneuroimmunoendocrine disorders of FM patients and ascertains whether a previous CFS diagnosis affected them.

Through accelerometry objective parameters, physical activity/sedentarism levels in relation to fatigue are studied, as well as whether perceived levels of stress, anxiety, and pain correspond to objective biomarkers, all of these with respect to a reference group (RG) of women without FM.

FM patients have a worse psychological state and perceived quality of life than those with RG. These perceived outcomes are consistent with impaired objective levels of a sedentary lifestyle, higher systemic levels of cortisol and noradrenaline, and lower levels of serotonin.

However, FM patients with a previous CFS diagnosis had lower systemic levels of IL-8, cortisol, oxytocin, and higher levels of adrenaline and serotonin than FM patients without diagnosed CFS.

In conclusion, while perceived health parameters do not detect differences, when objective neuroimmunoendocrine parameters related to stress, inflammation, pain, and fatigue are used, people with CFS could be overdiagnosed with FM. This reinforces the need for objective biomarker assessment of these patients for better diagnostic discrimination between both syndromes.

Source: Otero E, Gálvez I, Ortega E, Hinchado MD. Influence of Chronic Fatigue Syndrome Codiagnosis on the Relationship between Perceived and Objective Psychoneuro-Immunoendocrine Disorders in Women with Fibromyalgia. Biomedicines. 2023; 11(5):1488. https://doi.org/10.3390/biomedicines11051488 https://www.mdpi.com/2227-9059/11/5/1488 (Full text)

Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) and Comorbidities: Linked by Vascular Pathomechanisms and Vasoactive Mediators?

Abstract:

Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) is often associated with various other syndromes or conditions including mast cell activation (MCA), dysmenorrhea and endometriosis, postural tachycardia (POTS) and small fiber neuropathy (SFN). The causes of these syndromes and the reason for their frequent association are not yet fully understood.

We previously published a comprehensive hypothesis of the ME/CFS pathophysiology that explains the majority of symptoms, findings and chronicity of the disease. We wondered whether some of the identified key pathomechanisms in ME/CFS are also operative in MCA, endometriosis and dysmenorrhea, POTS, decreased cerebral blood flow and SFN, and possibly may provide clues on their causes and frequent co-occurrence.

Our analysis indeed provides strong arguments in favor of this assumption, and we conclude that the main pathomechanisms responsible for this association are excessive generation and spillover into the systemic circulation of inflammatory and vasoactive tissue mediators, dysfunctional β2AdR, and the mutual triggering of symptomatology and disease initiation. Overall, vascular dysfunction appears to be a strong common denominator in these linkages.

Source: Wirth KJ, Löhn M. Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) and Comorbidities: Linked by Vascular Pathomechanisms and Vasoactive Mediators? Medicina. 2023; 59(5):978. https://doi.org/10.3390/medicina59050978  https://www.mdpi.com/1648-9144/59/5/978 (Full text)

Investigating the factors associated with meaningful improvement on the SF-36-PFS and exploring the appropriateness of this measure for young people with ME/CFS accessing an NHS specialist service: a prospective cohort study

Abstract:

Objectives: Paediatric myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is relatively common and disabling, but little is known about the factors associated with outcome. We aimed to describe the number and characteristics of young people reaching the 10-point minimal clinically important difference (MCID) of SF-36-Physical Function Subscale (SF-36-PFS), and to investigate factors associated with reaching the MCID.

Design: Prospective observational cohort.

Setting: A specialist UK National Health Service (NHS) ME/CFS service, Southwest England. Recruitment between March 2014 and August 2015.

Participants: 193 eligible patients with ME/CFS aged 8 to 17 years reported baseline data. 124 (65%) and 121 (63%) with outcome data at six- and 12-months, respectively.

Outcome measures: SF-36-PFS (primary outcome). Chalder Fatigue Questionnaire, school attendance, visual analogue pain scale, Hospital Anxiety and Depression Scale, Spence Young people Anxiety Scale, Clinical Global Impression scale, and EQ-5D-Y (secondary)

Results: At six-months 48/120 (40%) had reached the MCID for SF-36-PFS. This had increased to 63/117 (54%) at 12-months. On the CGI, 77% and 79% reported feeling either a little better, much better, or very much better. Those with worse SF-36-PFS at baseline assessment were more likely to achieve the MCID for SF-36-PFS at six-months (OR 0.97, 95% CI 0.96 to 0.99, p-value 0.003), but there was no evidence of effect at 12-months (OR 0.98, 95% CI 0.97 to 1.00, p-value 0.038). No other factors at baseline were associated with the odds of reaching the MCID at six-months. However, at 12-months there was strong evidence of an effect of pain on MCID (OR 0.97, 95% CI 0.95 to 0.99, p-value 0.001), and SF-36-PFS on MCID (OR 0.96, 95% CI 0.94 to 0.98, p-value 0.001).

Conclusions: 40% and 54% of young people reached the MCID at 6 and 12 months respectively. No factors at assessment (other than SF-36-PFS at six-months, and pain and SF-36-PFS at 12-months) are associated with MCID of SF-36-PFS at either six-, or 12- months. Further work is needed to explore the most appropriate outcome measure for capturing clinical meaningful improvement for young people with ME/CFS.

Source: Gaunt, D. M., Brigden, A. L. C., Metcalfe, C., Loades, M., & Crawley, E. M. (Accepted/In press). Investigating the factors associated with meaningful improvement on the SF-36-PFS and exploring the appropriateness of this measure for young people with ME/CFS accessing an NHS specialist service: a prospective cohort study. BMJ Open. https://research-information.bris.ac.uk/en/publications/investigating-the-factors-associated-with-meaningful-improvement- 

Development of a Definition of Postacute Sequelae of SARS-CoV-2 Infection

Key Points:

Question  What symptoms are differentially present in SARS-CoV-2–infected individuals 6 months or more after infection compared with uninfected individuals, and what symptom-based criteria can be used to identify postacute sequelae of SARS-CoV-2 infection (PASC) cases?

Findings  In this analysis of data from 9764 participants in the RECOVER adult cohort, a prospective longitudinal cohort study, 37 symptoms across multiple pathophysiological domains were identified as present more often in SARS-CoV-2–infected participants at 6 months or more after infection compared with uninfected participants. A preliminary rule for identifying PASC was derived based on a composite symptom score.

Meaning  A framework for identifying PASC cases based on symptoms is a first step to defining PASC as a new condition. These findings require iterative refinement that further incorporates clinical features to arrive at actionable definitions of PASC.

Abstract:

Importance  SARS-CoV-2 infection is associated with persistent, relapsing, or new symptoms or other health effects occurring after acute infection, termed postacute sequelae of SARS-CoV-2 infection (PASC), also known as long COVID. Characterizing PASC requires analysis of prospectively and uniformly collected data from diverse uninfected and infected individuals.

Objective  To develop a definition of PASC using self-reported symptoms and describe PASC frequencies across cohorts, vaccination status, and number of infections.

Design, Setting, and Participants  Prospective observational cohort study of adults with and without SARS-CoV-2 infection at 85 enrolling sites (hospitals, health centers, community organizations) located in 33 states plus Washington, DC, and Puerto Rico. Participants who were enrolled in the RECOVER adult cohort before April 10, 2023, completed a symptom survey 6 months or more after acute symptom onset or test date. Selection included population-based, volunteer, and convenience sampling.

Exposure  SARS-CoV-2 infection.

Main Outcomes and Measures  PASC and 44 participant-reported symptoms (with severity thresholds).

Results  A total of 9764 participants (89% SARS-CoV-2 infected; 71% female; 16% Hispanic/Latino; 15% non-Hispanic Black; median age, 47 years [IQR, 35-60]) met selection criteria. Adjusted odds ratios were 1.5 or greater (infected vs uninfected participants) for 37 symptoms. Symptoms contributing to PASC score included postexertional malaise, fatigue, brain fog, dizziness, gastrointestinal symptoms, palpitations, changes in sexual desire or capacity, loss of or change in smell or taste, thirst, chronic cough, chest pain, and abnormal movements. Among 2231 participants first infected on or after December 1, 2021, and enrolled within 30 days of infection, 224 (10% [95% CI, 8.8%-11%]) were PASC positive at 6 months.

Conclusions and Relevance  A definition of PASC was developed based on symptoms in a prospective cohort study. As a first step to providing a framework for other investigations, iterative refinement that further incorporates other clinical features is needed to support actionable definitions of PASC.

Source: Thaweethai T, Jolley SE, Karlson EW, et al. Development of a Definition of Postacute Sequelae of SARS-CoV-2 Infection. JAMA. Published online May 25, 2023. doi:10.1001/jama.2023.8823 https://jamanetwork.com/journals/jama/fullarticle/2805540 (Full text)

Biomarkers for myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS): a systematic review

Abstract:

Background: Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a multifaceted condition that affects most body systems. There is currently no known diagnostic biomarker; instead, diagnosis is dependent on application of symptom-based case criteria following exclusion of any other potential medical conditions. While there are some studies that report potential biomarkers for ME/CFS, their efficacy has not been validated. The aim of this systematic review is to collate and appraise literature pertaining to a potential biomarker(s) which may effectively differentiate ME/CFS patients from healthy controls.

Methods: This systematic review was conducted according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses and Cochrane review guidelines. PubMed, Embase and Scopus were systematically searched for articles containing “biomarker” and “ME/CFS” keywords in the abstract or title and if they included the following criteria: (1) were observational studies published between December 1994 and April 2022; (2) involved adult human participants; (3) full text is available in English (4) original research; (5) diagnosis of ME/CFS patients made according to the Fukuda criteria (1994), Canadian Consensus Criteria (2003), International Consensus Criteria (2011) or Institute of Medicine Criteria (2015); (6) study investigated potential biomarkers of ME/CFS compared to healthy controls. Quality and Bias were assessed using the Joanna Briggs Institute Critical Appraisal Checklist for Case Control Studies.

Results: A total of 101 publications were included in this systematic review. Potential biomarkers ranged from genetic/epigenetic (19.8%), immunological (29.7%), metabolomics/mitochondrial/microbiome (14.85%), endovascular/circulatory (17.82%), neurological (7.92%), ion channel (8.91%) and physical dysfunction biomarkers (8.91%). Most of the potential biomarkers reported were blood-based (79.2%). Use of lymphocytes as a model to investigate ME/CFS pathology was prominent among immune-based biomarkers. Most biomarkers had secondary (43.56%) or tertiary (54.47%) selectivity, which is the ability for the biomarker to identify a disease-causing agent, and a moderate (59.40%) to complex (39.60%) ease-of-detection, including the requirement of specialised equipment.

Conclusions: All potential ME/CFS biomarkers differed in efficiency, quality, and translatability as a diagnostic marker. Reproducibility of findings between the included publications were limited, however, several studies validated the involvement of immune dysfunction in the pathology of ME/CFS and the use of lymphocytes as a model to investigate the pathomechanism of illness. The heterogeneity shown across many of the included studies highlights the need for multidisciplinary research and uniform protocols in ME/CFS biomarker research.

Source: Maksoud R, Magawa C, Eaton-Fitch N, Thapaliya K, Marshall-Gradisnik S. Biomarkers for myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS): a systematic review. BMC Med. 2023 May 24;21(1):189. doi: 10.1186/s12916-023-02893-9. PMID: 37226227; PMCID: PMC10206551. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10206551/ (Full text)

Autoantibodies to Selenoprotein P in Patients with Chronic Fatigue Syndrome Suggest Selenium Transport Impairment and Acquired Resistance to Thyroid Hormone

Abstract:

Chronic Fatigue Syndrome (CFS) presents with symptoms similar to hypothyroidism, including mental and physical fatigue, poor sleep, depression, and anxiety. However, the typical thyroid hormone (TH) profile of elevated thyrotropin (TSH) and low thyroxine (T4) is not observed. Recently, autoantibodies to the selenium transporter SELENOP (SELENOP-aAb) have been identified in Hashimoto’s thyroiditis and shown to impair selenium transport and selenoprotein expression. We hypothesized that SELENOP-aAb are prevalent in CFS and impair TH metabolism.

Selenium status in CFS (n=167) was compared to that of healthy controls (n=545). Two additional small groups were included, namely patients with fibromyalgia (FM; n=39), a disease often comorbid with CFS, and patients with post-COVID condition (n=24). The serum/plasma Se biomarkers total Se, glutathione peroxidase (GPx3) and SELENOP levels showed linear correlations without reaching saturation, indicative of Se deficiency. TSH and total T4 levels fitted within normal ranges, but relative total T3 (%TT3) was low, and relative rT3 (%rT3) was elevated in CFS. SELENOP-aAb prevalence was 9.6-15.6% in CFS versus 0.9-2.0% in controls, depending on cut-off for positivity.

An impairment of Se transport in SELENOP-aAb positive CFS patients is suggested by the lack of correlation between total Se and GPx3 activity. The same patients present with disturbed TH parameters, including low deiodinase (DIO) activity (SPINA-GD index) and particularly low urinary iodine as compared to controls (43.2 (16.0) vs. 89.0 (54.9) µg/L, P<0.001), indicating that SELENOP-aAb affect TH deiodination and iodine excretion.

We conclude that a considerable subset of CFS patients express SELENOP-aAb that disturb Se transport and cause low GPx3 and DIO activities. Hereby, TH deiodination decreases as an acquired condition that is not readily reflected by TSH or T4 in blood. This hypothesis opens new explanations and therapeutic options for SELENOP-aAb positive CFS and, perhaps, post-COVID condition patients, but requires additional clinical evidence from intervention trials.

Source: Sun, Qian and Oltra, Elisa and Dijck-Brouwer, D. A. Janneke and Chillon, Thilo Samson and Seemann, Petra and Asaad, Sabrina and Demircan, Kamil and Espejo-Oltra, José Andrés and Sánchez-Fito, Teresa and Martin-Martinez, Eva and Minich, Waldemar B. and Muskiet, Frits A. J. and Schomburg, Lutz, Autoantibodies to Selenoprotein P in Patients with Chronic Fatigue Syndrome Suggest Selenium Transport Impairment and Acquired Resistance to Thyroid Hormone. Available at SSRN: https://ssrn.com/abstract=4332223 or http://dx.doi.org/10.2139/ssrn.4332223 (Full text available as PDF file)

Hypothalamus volumes in adolescent Myalgic Encephalomyelitis/Chronic Fatigue Syndrome: Impact of self-reported fatigue and illness duration

Abstract:

Adolescent Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) is a complex illness of unknown aetiology. Emerging theories suggest ME/CFS may reflect a progressive, aberrant state of homeostasis caused by disturbances within the hypothalamus, yet few studies have investigated this using magnetic resonance imaging in adolescents with ME/CFS.

We conducted a volumetric analysis to investigate whether whole and regional hypothalamus volumes in adolescents with ME/CFS differed compared to healthy controls, and whether these volumes were associated with fatigue severity and illness duration. 48 adolescents (25 ME/CFS, 23 controls) were recruited. Lateralised whole and regional hypothalamus volumes, including the anterior superior, superior tubular, posterior, anterior inferior and inferior tubular subregions, were calculated from T1 weighted images.

When controlling for age, sex and intracranial volume, Bayesian linear regression revealed no evidence for differences in hypothalamus volumes between groups. However, in the ME/CFS group, a negative linear relationship between right anterior superior volumes and fatigue severity was identified, which was absent in controls. In addition, Bayesian ordinal regression revealed a likely-positive association between illness duration and right superior tubular volumes in the ME/CFS group.

While these findings suggest overall comparability in regional and whole hypothalamus volumes between adolescents with ME/CFS and controls, preliminary evidence was identified to suggest greater fatigue and longer illness duration were associated with greater right anterior superior and superior tubular volumes, respectively. These regions contain the anterior and superior divisions of the paraventricular nucleus, involved in the neuroendocrine response to stress, suggesting involvement in ME/CFS pathophysiology. However, replication in a larger, longitudinal cohort is required.

Source: Hollie Byrne, Elisha K Josev, Sarah J Knight, Adam Scheinberg, Katherine Rowe, Lionel Lubitz, Marc L Seal. Hypothalamus volumes in adolescent Myalgic Encephalomyelitis/Chronic Fatigue Syndrome: Impact of self-reported fatigue and illness duration. medRxiv 2023.05.16.23290031; doi: https://doi.org/10.1101/2023.05.16.23290031 (Full text available as PDF file)

Viable SARS-CoV-2 Omicron sub-variants isolated from autopsy tissues

Introduction: Pulmonary and extrapulmonary manifestations have been described after infection with SARS-CoV-2, the causative agent of coronavirus disease 2019 (COVID-19). The virus is known to persist in multiple organs due to its tropism for several tissues. However, previous reports were unable to provide definitive information about whether the virus is viable and transmissible. It has been hypothesized that the persisting reservoirs of SARS-CoV-2 in tissues could be one of the multiple potentially overlapping causes of long COVID.

Methods: In the present study, we investigated autopsy materials obtained from 21 cadaveric donors with documented first infection or reinfection at the time of death. The cases studied included recipients of different formulations of COVID-19 vaccines. The aim was to find the presence of SARS-CoV-2 in the lungs, heart, liver, kidneys, and intestines. We used two technical approaches: the detection and quantification of viral genomic RNA using RT-qPCR, and virus infectivity using permissive in vitro Vero E6 culture.

Results: All tissues analyzed showed the presence of SARS-CoV-2 genomic RNA but at dissimilar levels ranging from 1.01 × 102 copies/mL to 1.14 × 108 copies/mL, even among those cases who had been COVID-19 vaccinated. Importantly, different amounts of replication-competent virus were detected in the culture media from the studied tissues. The highest viral load were measured in the lung (≈1.4 × 106 copies/mL) and heart (≈1.9 × 106 copies/mL) samples. Additionally, based on partial Spike gene sequences, SARS-CoV-2 characterization revealed the presence of multiple Omicron sub-variants exhibiting a high level of nucleotide and amino acid identity among them.

Discussion: These findings highlight that SARS-CoV-2 can spread to multiple tissue locations such as the lungs, heart, liver, kidneys, and intestines, both after primary infection and after reinfections with the Omicron variant, contributing to extending knowledge about the pathogenesis of acute infection and understanding the sequelae of clinical manifestations that are observed during post-acute COVID-19.

Source: Santiago Maffia-Bizzozero, Cintia Cevallos, Federico Remes Lenicov, Rosa Nicole Freiberger, Cinthya Alicia Marcela Lopez, Alex Guano Toaquiza, Franco Sviercz, Patricio Jarmoluk, Cristina Bustos, Adriana Claudia D’Addario, Jorge Quarleri, and M. Victoria Delpino. Viable SARS-CoV-2 Omicron sub-variants isolated from autopsy tissues. Front. Microbiol., 22 May 2023. https://www.frontiersin.org/articles/10.3389/fmicb.2023.1192832/full (Full text)