Abstract
ABSTRACTTuberculosis (TB) is a disease caused by Mycobacterium tuberculosis or Mycobacterium bovis and still remains one of the world's biggest global health burdens. Recently, engineered polyhydroxyalkanoate (PHA) biobeads that were produced in both Escherichia coli and Lactococcus lactis and displayed mycobacterial antigens were found to induce significant cell-mediated immune responses in mice. We observed that such PHA beads contained host cell proteins as impurities, which we hypothesized to have the potential to induce immunity. In this study, we aimed to develop PHA beads produced in mycobacteria (mycobacterial PHA biobeads [MBB]) and test their potential as a TB vaccine in a mouse model. As a model organism, nonpathogenic Mycobacterium smegmatis was engineered to produce MBB or MBB with immobilized mycobacterial antigens Ag85A and ESAT-6 on their surface (A:E-MBB). Three key enzymes involved in the poly(3-hydroxybutyric acid) pathway, namely, β-ketothiolase (PhaA), acetoacetyl-coenzyme A reductase (PhaB), and PHA synthase (PhaC), were engineered into E. coli-Mycobacterium shuttle plasmids and expressed in trans. Immobilization of specific antigens to the surface of the MBB was achieved by creating a fusion with the PHA synthase which remains covalently attached to the polyester core, resulting in PHA biobeads displaying covalently immobilized antigens. MBB, A:E-MBB, and an M. smegmatis vector control (MVC) were used in a mouse immunology trial, with comparison to phosphate-buffered saline (PBS)-vaccinated and Mycobacterium bovis BCG-vaccinated groups. We successfully produced MBB and A:E-MBB and used them as vaccines to induce a cellular immune response to mycobacterial antigens.IMPORTANCE Tuberculosis (TB) is a disease caused by Mycobacterium tuberculosis or Mycobacterium bovis and still remains one of the world's biggest global health burdens. In this study, we produced polyhydroxyalkanoate (PHA) biobeads in mycobacteria and used them as vaccines to induce a cellular immune response to mycobacterial antigens.
Highlights
IMPORTANCE Tuberculosis (TB) is a disease caused by Mycobacterium tuberculosis or Mycobacterium bovis and still remains one of the world’s biggest global health burdens
Adverse events linked to bacillus Calmette-Guérin (BCG) vaccination range from mild, localized complications to more serious, systemic, or disseminated BCG disease in which M. bovis BCG is confirmed in one or more anatomical sites far from either the site of injection or regional lymph nodes
A two-plasmid system utilizing compatible E. coli-Mycobacterium shuttle plasmids pMycVec1 and pMycVec2 [29] was used, as was a system based on the pMIND plasmid [30, 31]
Summary
IMPORTANCE Tuberculosis (TB) is a disease caused by Mycobacterium tuberculosis or Mycobacterium bovis and still remains one of the world’s biggest global health burdens. These strategies have been successful in reducing both the incidence and the prevalence of TB globally. Adverse events linked to BCG vaccination range from mild, localized complications to more serious, systemic, or disseminated BCG disease in which M. bovis BCG is confirmed in one or more anatomical sites far from either the site of injection or regional lymph nodes. Immunocompromised individuals such as those with HIV infection have a significantly higher risk of developing BCG-related diseases [8,9,10,11]. A limited understanding of immunity to M. tuberculosis is significantly hindering vaccine development [12]
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