Arlehamn, C. L. | Search bioRxiv

bioRxiv2025

Peptide Driven Identification of TCRs (PDI-TCR) reveals dynamics and phenotypes of CD4 T cells in tuberculosis

Assigning antigen specificity to T cell receptor (TCR) sequences is challenging due to the TCR repertoires diversity and the complexity of TCR:antigen recognition. We developed the Peptide-Driven Identification of TCRs (PDI-TCR) assay that combines in vitro expansion of cells with peptide pools, bulk TCR sequencing, and statistical analysis to identify antigen-specific TCRs from human blood. A key feature of PDI-TCR is the ability to distinguish true antigen-specific TCR clonotypes from TCRs associated with unspecific bystander activation by comparing responses to non-overlapping peptide pools. We applied PDI-TCR to Tuberculosis (TB) patients, sampling blood at diagnosis and throughout treatment, and Mycobacterium tuberculosis (Mtb)-sensitized healthy individuals (IGRA+). We identified hundreds of Mtb-specific TCRs, as well as unspecific TCRs, and characterized their phenotype in each cohort by single-cell RNA sequencing ex vivo. Mtb-specific T cells were highly diverse, with short-lived effector phenotypes only present in TB at diagnosis, while memory phenotypes were maintained through treatment. In contrast, unspecific expanded T cells were more clonally restricted, had a cytotoxic phenotype, and were maintained throughout treatment. This showcases PDI-TCR as a powerful tool for identifying antigen-specific TCRs, which enables direct ex vivo identification and monitoring of antigen-specific T cells.

immunology↗

bioRxiv2022

Host genetic background is a barrier to vaccine-induced protection against infection

The heterogeneity of immune responses observed in humans is difficult to model in standard inbred laboratory mice. To capture the diversity inherent in mice and better understand how host variation affects BCG-induced immunity against Mycobacterium tuberculosis, 24 unique Collaborative Cross (CC) recombinant inbred mouse strains and the C57BL/6 reference strain were vaccinated with or without BCG, and then challenged with low-dose aerosolized virulent M. tuberculosis. In contrast to standard lab strains, BCG protected only half of the CC strains tested. Furthermore, BCG efficacy is dissociable from inherent susceptibility to TB. As these strains differed primarily in the genes and alleles they inherited from the CC founder strains, we conclude that the host genetic background has a major influence on whether BCG confers protection against M. tuberculosis infection and indicates that host genetics should be considered as an important barrier to vaccine-mediated protection. Importantly, we wished to identify the components of the immune response stimulated by BCG, which were subsequently recalled after Mtb infection and associated with protection. The T cell immune response following BCG vaccination and Mtb challenge was extensively characterized. Although considerable diversity was observed, BCG vaccination had little impact on the composition of T cells recruited and maintained in the lung after infection. Instead, the variability was largely shaped by the genetic background. We developed models to detect vaccine-induced differences, which identified immune signatures associated with BCG-elicited protection against TB. Importantly, even when categorized as susceptible vs. resistant, and protected vs. unprotected, many of the protected CC strains had unique flavors of immunity, indicating multiple paths to protection. Thus, CC mice can be used to define correlates of protection and to identify vaccine strategies that protect a larger fraction of genetically diverse individuals instead of optimizing protection for a single genotype.

immunology↗