Ex vivo Platforms to Study the Primary and Recall Immune Responses to Intracellular Mycobacterial Pathogens and Peptide-Based Vaccines

William C. Davis, Asmaa H. Mahmoud, Gaber S. Abdellrazeq, Mahmoud M. Elnaggar, John L. Dahl, Victoria Hulubei, Lindsay M. Fry

Research output: Contribution to journalReview articlepeer-review


Progress in the study of the immune response to pathogens and candidate vaccines has been impeded by limitations in the methods to study the functional activity of T-cell subsets proliferating in response to antigens processed and presented by antigen presenting cells (APC). As described in this review, during our studies of the bovine immune response to a candidate peptide-based vaccine and candidate rel deletion mutants in Mycobacterium avium paratuberculosis (Map) and Mycbacterium bovis (BCG), we developed methods to study the primary and recall CD4 and CD8 T-cell responses using an ex vivo platform. An assay was developed to study intracellular killing of bacteria mediated by CD8 T cells using quantitative PCR to distinguish live bacteria from dead bacteria in a mixed population of live and dead bacteria. Through use of these assays, we were able to demonstrate vaccination with live rel Map and BCG deletion mutants and a Map peptide-based vaccine elicit development of CD8 cytotoxic T cells with the ability to kill intracellular bacteria using the perforin-granzyme B pathway. We also demonstrated tri-directional signaling between CD4 and CD8 T cells and antigen-primed APC is essential for eliciting CD8 cytotoxic T cells. Herein, we describe development of the assays and review progress made through their use in the study of the immune response to mycobacterial pathogens and candidate vaccines. The methods obviate some of the major difficulties encountered in characterizing the cell-mediated immune response to pathogens and development of attenuated and peptide-based vaccines.

Original languageEnglish (US)
Article number878347
JournalFrontiers in Veterinary Science
StatePublished - May 3 2022

Bibliographical note

Funding Information:
The studies reviewed in this manuscript were supported in part by the National Institute of Food and Agriculture-Agriculture and Food Research Initiative Competitive Grant 2018-67015-28744 (WD). Mention of trade names, proprietary products, or specified equipment do not constitute a guarantee or warranty by the USDA and does not imply approval to the exclusion of other products that may be suitable. USDA is an Equal Opportunity Employer. This review was also supported in part by the WSUMAC: http://vmp.vetmed.wsu.edu/resources/monoclonal-antibody-center .

Publisher Copyright:
Copyright © 2022 Davis, Mahmoud, Abdellrazeq, Elnaggar, Dahl, Hulubei and Fry.


  • Mycobacterium avium subspecies paratuberculosis
  • Mycobacterium bovis
  • Mycobacterium tuberculosis
  • cellular immune response
  • intracellular pathogens
  • peptide vaccines

PubMed: MeSH publication types

  • Journal Article
  • Review


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