Preprint Single-cell profiling of innate and adaptive immune dysregulation in Long COVID, 2026, Satpathy et al.

[fresh_air]

Abstract​

The COVID-19 pandemic has infected more than 778 million people worldwide. Roughly 7% of these patients progress to Long COVID (LC), suffering from persistent symptoms and cognitive impairment well past the acute stage. As the mechanisms of LC remain elusive, we utilized single-cell profiling (SCP) on 156,478 peripheral blood mononuclear cells (PBMCs) from 20 LC patients and 18 recovered controls (RC) to characterize the disease-associated immune dysregulation. Comparative analysis of LC and RC profiles revealed cellular heterogeneity along with differential abundances across B, T, and myeloid cell compartments.

The focused analysis on the B-cell compartment showed that naïve B cells in the LC exhibit elevated IL4R expression and BCR signaling, indicative of sustained antigen exposure and aberrant chronic activation. Concurrently, monocytes adopted heightened interferon signaling and enhanced migratory states, culminating into impaired myeloid differentiation. Furthermore, the T-cell compartment exhibited a functional dichotomy, maintaining sustained quiescence in the central memory compartment while displaying chronic exhaustion within effector memory populations. This dysregulation of effector immunity extended to the NK compartment, where terminally differentiated cells exhibited increased cytotoxicity yet compromised regulatory function, potentially contributing to poor viral clearance. Cellular communication analysis further supports this NK cell dysfunction that is likely driven by galectin and prostaglandin signaling involving monocytes and B cells.

We stratified LC patients into mild and severe groups based on symptom and cognitive severity, identifying a distinct immune signature where severe disease is linked to chronic AP-1-mediated inflammation in NK cells and CD14+ monocytes. In contrast, patients with mild symptoms retain functionally competent NK cells with significantly lower exhaustion and apoptosis scores.

Collectively, these insights into persistent immune remodeling provide a crucial framework for future biomarker discovery and the development of targeted therapeutic strategies.


Link:

https://www.biorxiv.org/content/10.64898/2026.06.04.730206v1

DOI:

https://doi.org/10.64898/2026.06.04.730206

 

[SNT Gatchaman]

A few quotes —

we assessed IG gene usage diversity. Individuals in the recovered COVID-19 group exhibited higher immunoglobulin (Ig) isotype utilization, suggesting a more robust B cell compartment. Pathway analysis revealed Rho GTPase–mediated activation of NADPH oxidase and antigen driven B cell receptor signaling, culminating in secondary messenger generation within Long COVID enriched clusters. Taken together, these observations along with differential abundance, motivate a comparison between IL4+ B naïve cluster (enriched in Long COVID) and ZEB1+B naïve cluster (enriched in recovered COVID) to elucidate their transcriptional, and functional differences.

Fc Receptor-Like 6 (FCRL) 1 plays a role in building the BCR signalosome, influencing B cell responsiveness and antibody production. Along similar lines, B cell scaffold protein with ankyrin repeats (BANK1) participates in B cell signaling, specifically BCR and Toll like receptor (TLR) pathways, suggesting a role in signaling and may contribute to active defense against latent infection in the Long COVID group. Elevated IL-4 expression in naive cells further supports a role for antigen exposure and subsequent memory B-cell generation in the Long COVID group. These findings suggest that B cells in Long COVID patients are chronically activated, potentially leading to aberrant immune signaling well after the acute COVID phase has ended.

Among the non-classical monocytes, the CX3CR1⁺ subset was notably enriched in individuals with Long COVID and exhibited elevated expression of genes such as SKAP2, CX3CR1, HDAC9 (average log2FC > 1.5, P<0.001

Exploring the interferon response phenotype, we observed distinct interferon signaling activity in both the IL1B+ RETN Lo CD14+ and DDIT4+ PDIA3+ CD14+ monocyte clusters. While both clusters exhibited upregulation of interferon-stimulated genes, the transcriptional drivers differed markedly. The IL1B+ RETNLo CD14+ (Long COVID) cluster was predominantly characterized by elevated expression of genes CCR1, CCR2, CXCL8, IL1B, STAT2, whereas the DDIT4+ PDIA3+ CD14+ (recovered COVID) cluster showed enrichment of genes STAT1 and STAT3. This divergence highlights the presence of at least two distinct interferon response programs within the monocyte compartment in Long COVID, suggesting functional heterogeneity in antiviral and inflammatory signaling.

Beyond the T-cell compartment, our investigation into NK cell populations revealed two differentially abundant subsets with distinct transcriptional profiles in Long COVID and recovered COVID groups. […] This expression pattern suggests that NK cells in recovered individuals maintain a homeostatic yet alert state, balancing activation with regulation to facilitate coordinated immune resolution.

A notable finding was galectin mediated signaling originating from monocytes and targeting NK cells, observed exclusively in the Long COVID patients). This pathway likely promotes NK cell dysfunction, potentially through the expression of the ligand LGALS9 on monocytes and its corresponding receptor P4HB on FCER1G⁺ NK cells.

Collectively, cellular communication analysis highlighted a reprogrammed immune signaling network in Long COVID, defined by selective crosstalk among monocytes, B cells, and NK cells. The emergence of disease-specific ligand-receptor interactions suggests a unique immunoregulatory axis that may contribute to persistent inflammation and impaired immune resolution.

Uniquely, we evaluated immune dysregulation in a population of African Americans with high social vulnerability, a demographic underrepresented in Long COVID pathobiological research. COVID-19 has disproportionally affected African American communities with significantly higher morbidity and mortality rates, and recent evidence suggests that social risk factors and systemic barriers increase the risk of Long COVID development and persistence. Long COVID profiles may vary by race and ethnicity, indicating potential variation in the underlying drivers of disease versus variation in the symptom manifestation of those drivers. Our overarching findings are consistent with the latter and support current evidence of immune activation and fatiguability in response to persistent antigen exposure.