Project Team

Marcus Buggert, PhD: Assistant Professor, Karolinska Institutet, Sweden

Soo Aleman, MD, PhD: Adjunct Professor, Department of Infectious Diseases, Karolinska Institutet, Sweden

Yu Gao, PhD: Senior Researcher, Karolinska Institutet, Sweden

Takuya Sekine, PhD: Assistantant Professor, Karolinska Institutet, Sweden

Project summary:

The project will determine if persistence of SARS-CoV-2 or its proteins in the gut or tonsils can drive immune activation and microbiome changes in Long COVID. More specifically, the team is using a range of advanced technologies to analyze a unique collection of biopsy samples obtained from multiple gut regions of Long COVID patients. They are also analyzing tonsil samples collected from over 200 people (from both COVID-19-affected and unaffected individuals). These tonsil samples are part of the body’s lymphatic system. Experiments on these tissue samples will determine 1) if the SARS-CoV-2 virus can persist in distinct cell subsets of the gut and lymphatic system and 2) if persistence is associated with local and systemic immune, microbiome, or gene expression changes that might lead to Long COVID symptoms.

Project background:

A growing body of research shows that the SARS-CoV-2 virus or its proteins can persist in tissues of the gut and lymphatic system. For example, one study identified prolonged immune responses to SARS-CoV-2 and viral RNA in the tonsils of children months or years after initial infection. Another study found elevated T cell activation in tonsil tissues of patient with Long COVID.

These findings align with research generated by the current project’s team. In acute COVID-19, the team identified the presence of SARS-CoV-2 responsive CD8+ T cells in human tonsil tissue that were not detectable in blood samples collected from the same individuals. This distinction is crucial. The tonsil T cells are categorized as tissue-resident memory CD8+ T cells, which serve as sentinel defenders within tissues.  It suggests that if major lymphoid organs like the tonsils harbor SARS-CoV-2 RNA for extended periods of time, they could initiate pronounced tissue-resident memory cell activation within tissues – an immune process that might go unnoticed if analyses were restricted solely to blood testing.

Flow cytometry data from the project team showing the absence of SARS- CoV-2-reactive CD8+ T cells in blood compared to tonsils (left and middle graphs), due to TRM features of these cells (right).

The current project draws on this research to study similar trends in Long COVID. Specifically, the team is working to:

1. Investigate the presence of persistent SARS-CoV-2 in gut cell subsets and its association with inflammatory, microbial, and gene expression changes: Blood samples and tissue biopsies are being obtained from the colon, cecum, and ileum of patients with Long COVID and healthy people who had previous COVID-19. The team is employing an integrated single-cell analysis framework to determine:

  • SARS-CoV-2 cellular tropism: They will identify which cells and gut sites contain SARS-CoV-2 RNA or protein and determine if certain cell types demonstrate a propensity for persistence.
  • SARS-CoV-2 circulating proteins: They use sensitive methods to determine if SARS-CoV-2 proteins in blood reflect those identified in gut tissue.
  • T cell recognition of SARS-CoV-2: They will investigate if SARS-CoV-2-specific CD8+ T cells possess the capacity to kill virus-infected cells in the gut.
  • Transcriptome alterations: They will determine if SARS-CoV-2 reservoirs alters the local gene expression patterns of immune cells, the cell extracellular matrix, and epithelial cells in Long COVID.
  • Microbiome correlations: They will examine any link between SARS-CoV-2 reservoirs and microbiome disruptions – determining if such disruptions are a cause or consequence of Long COVID.
  • Gut barrier dysfunction: They will explore the association between SARS-CoV-2 reservoirs and epithelial barrier dysfunction in Long COVID – focusing on the potential for viral proteins or other organisms to penetrate the bloodstream.

2. Determine if persistent SARS-CoV-2 proteins in lymphoid tissue (tonsil) lead to sustained immune activation of T cells: The team is analyzing tonsil samples collected from individuals with and without a history of COVID-19 to determine the presence of lingering viral particles in lymphoid organs could contribute to heightened T cell activation, potentially leading to Long COVID symptoms. As part of this research they are characterizing many T cell properties including memory differentiation, cellular exhaustion, transcription factors, and activation markers. The team has also developed a lymphoid organoid model that is being used to determine if persistent SARS-CoV-2 particles can provoke T cell activation and proliferation in the laboratory.