Tuberculosis (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 produced in
both E. coli and Lactococcus lactis displaying 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 TB vaccine in a mouse model. As a model organism, nonpathogenic
Mycob... More
Tuberculosis (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 produced in
both E. coli and Lactococcus lactis displaying 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 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-CoA reductase (PhaB), and PHA synthase
(PhaC), were engineered into E. coli−mycobacteria 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 a mycobacterial vector control (MVC) were used in a
mouse immunology trial, with comparison to PBS vaccinated and 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.