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Multi-drug resistance pump gene deletion strains cannot survive in egg white because of ovotransferrin-derived antimicrobial activity, and are safe and effective oral live attenuated vaccines

Published: July 20, 2020
By: K. Vermeulen, F.D. Meyer, R. Raspoet, V. Eeckhaut, S. Kilroy, R. Ducatelle & F.V. Immerseel / Ghent University, Faculty of Veterinary Medicine, Department of Pathology, Bacteriology and Avian Diseases.

Multi-drug resistance (MDR) pumps are of vital importance for microbial survival in a variety of environments as they can export antimicrobial molecules, including host-derived antimicrobial peptides. In this way bacteria can withstand the antimicrobial peptides at mucosal surfaces, and colonization in specific host niches can be achieved. MDR pumps thus constitute a virulence trait as a protection mechanism against innate host defenses. The outer membrane channel TolC, used by Salmonella MDR pumps to export antimicrobial compounds, was shown to be crucial for survival of Salmonella in egg white, using deletion mutants and agar spot and egg white survival assays. MDR pump mutants had an identical phenotype, indicating that strategies of Salmonella to survive in egg white are likely based on protection against egg white antimicrobial components. The tolC gene promoter was shown to be activated by egg white, pointing to an active upregulation of defense mechanisms when encountering harsh environments. Testing of egg white fractions, derived by chromatographic methods, showed that ovotransferrin was the main driver of the antimicrobial activity against the TolC deletion mutant, and not against the wild type. Either the TolC channel thus aids in pumping out siderophores to compete with ovotransferrin for iron, or TolC pumps out ovotransferrin-derived antimicrobial molecules. Triple oral vaccination of layer pullets (day 1, week 6, week 16) with a ΔTolC outer membrane channel mutant or a ΔacrABacrEFmdtABC MDR pump mutant was shown to protect against organ colonization and egg contamination by the wild type challenge strain. In the challenge control group, intravenous challenge with Salmonella Enteritidis at week 24 resulted in high colonization levels in the gut and internal organs, including the reproductive tract, and in high egg contamination levels, while vaccinated animals had significantly lower challenge strain bacteria in their organs. Egg contamination was completely prevented. These data show that the deletion specific genes, based on knowledge of the pathogenesis of the infection, can generate potential safe and effective vaccine strains for Salmonella in poultry.
Keywords: Salmonella, vaccines, MDR pumps.


Abstract presented at the 3rd International Symposium on Alternatives to Antibiotics 2019.

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Karen Vermeulen
Ghent University
Fien De Meyer
Ghent University
Ruth Raspoet
Phileo by Lesaffre
Venessa Eeckhaut
Ghent University
Richard Ducatelle
Ghent University
Filip Van Immerseel
Ghent University
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