Development of Non-Antibiotic-Resistant, Chromosomally Based, Constitutive and Inducible Expression Systems for aroA-Attenuated Salmonella enterica Serovar Typhimurium

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Title Development of Non-Antibiotic-Resistant, Chromosomally Based, Constitutive and Inducible Expression Systems for aroA-Attenuated Salmonella enterica Serovar Typhimurium
Author Matic, Jake N.; Terry, Tamsin D.; Bockel, David Van; Maddocks, Tracy; Tinworth, David; Jennings, Michael Paul; Djordjevic, Steven P.; Walker, Mark J.
Journal Name Infection and Immunity
Year Published 2009
Place of publication United States
Publisher American Society for Microbiology
Abstract Live-vaccine delivery systems expressing two model antigens from Mycoplasma hyopneumoniae, F2P97 (Adh) and NrdF, were constructed using Salmonella enterica serovar Typhimurium aroA (STM-1), and immunogenicity in mice was evaluated. Recombinant plasmid-based expression (PBE) and chromosomally based expression (CBE) systems were constructed. The PBE system was formed by cloning both antigen genes into pJLA507 to create an operon downstream of temperature-inducible promoters. Constitutive CBE was achieved using a promoter-trapping technique whereby the promoterless operon was stably integrated into the chromosome of STM-1, and the expression of antigens was assessed. The chromosomal position of the operon was mapped in four clones. Inducible CBE was obtained by using the in vivo-induced sspA promoter and recombining the expression construct into aroD. Dual expression of the antigens was detected in all systems, with PBE producing much larger quantities of both antigens. The stability of antigen expression after in vivo passage was 100% for all CBE strains recovered. PBE and CBE strains were selected for comparison in a vaccination trial. The vaccine strains were delivered orally into mice, and significant systemic immunoglobulin M (IgM) and IgG responses against both antigens were detected among all CBE groups. No significant immune response was detected using PBE strains. Expression of recombinant antigens in S. enterica serovar Typhimurium aroA from chromosomally located strong promoters without the use of antibiotic resistance markers is a reliable and effective method of inducing a significant immune response.
Peer Reviewed Yes
Published Yes
Alternative URI http://dx.doi.org/10.1128/IAI.01301-08
Volume 77
Issue Number 5
Page from 1817
Page to 1826
ISSN 0019-9567
Date Accessioned 2010-02-18
Language en_AU
Research Centre Institute for Glycomics
Faculty Faculty of Science, Environment, Engineering and Technology
Subject Infectious Agents
URI http://hdl.handle.net/10072/33885
Publication Type Journal Articles (Refereed Article)
Publication Type Code c1x

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