Fig 7. Chromosomal integrant (Ty21a-Gad) can survive extreme pH when pre-grown for 24 hours under appropriate inducing conditions.

A. Selected bacteria were grown in TSB-MES pH 5.5 with or without glucose for 18 or 24 hours prior to acid challenge. Although Ty21a-Gad did not express much detectable Gad protein at 18 hours in the presence of glucose, Gad-expression was easily observed at 24 hours. Interestingly, Gad proteins were expressed even in the absence of glucose after 18 hours. In contrast to single copy Ty21a-Gad, the plasmid construct (Ty21a pGad) existed at ~ 10 copies per cell [44] and expressed a higher level of Gad proteins under all conditions. An antibody-binding, non-specific band was used as the loading control. B. Consistent with level of Gad protein expression, Ty21a-Gad acid survival was comparable to that of Ty21a after 18 hours of growth in TSB-MES pH 5.5 with glucose, but acid survival improved significantly (~3 logs) when Ty21a-Gad were grown for 24 hours prior to acid challenge.