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The Efficacy of Bacteriocins to Control Listeria monocytogenes
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- Author / Creator
- Balay, Danielle R
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Listeria monocytogenes is a problem in ready-to-eat meat products and new methods of food preservation, such as bacteriocins, are being investigated to control this organism. Leucocin A and carnocyclin A, are class IIa and class IIc bacteriocins that have antilisterial activity. The aims of this research were to: a) improve the purification protocols for leucocin A and carnocyclin A to increase the yield of peptide obtained for downstream experiments and potential commercialization purposes; and b) evaluate the efficacy of leucocin A and an analogue of leucocin A to inhibit the growth of L. monocytogenes on wieners in the presence of spoilage organisms. The strains of L. monocytogenes used in this research were associated with foodborne illness and developed to use in challenge studies. The strains of L. monocytogenes were also grown in media supplemented with different carbohydrates (glucose, sucrose, fructose, mannose, cellobiose) with or without the addition of leucocin A or carnocyclin A and the growth curves were modelled to determine the growth kinetics. Lastly, RNA-Sequencing was employed to investigate the transcriptome of L. monocytogenes FSL C1-056 when cells were grown in mannose or cellobiose and treated with carnocyclin A. Additionally, the genome of L. monocytogenes FSL C1-056 was sequenced, annotated and used to assess the expressed genes and potential ncRNA regions that may relate to L. monocytogenes stress response to carnocyclin A. Minimum inhibitory concentrations could not be determined for L. monocytogenes exposed to leucocin A in broth; however, on agar the minimum inhibitory concentrations ranged from 11.7–62.5 M and 62.5–>500 M for leucocin A and leucocin A analogue, respectively. The inactivation profiles of L. monocytogenes in broth in the presence of leucocin A suggested each isolate had different levels of resistance to the bacteriocin as determined by the initial bactericidal effect. The formation of spontaneously resistance subpopulations were also observed for each strain of L. monocytogenes. In situ, leucocin A reduced the counts L. monocytogenes on wieners during storage, regardless of the presence of C. divergens. B. thermosphacta was unaffected by the presence of leucocin A on wieners over the duration of storage. Growth in media containing glucose, mannose, and fructose increased the sensitivity of some strains of L. monocytogenes to leucocin A and carnocyclin A, while growth in cellobiose and sucrose increased the resistance of L. monocytogenes to leucocin A and carnocyclin A, as evidenced by a shorter lag phase. Strains of L. monocytogenes developed resistance to both leucocin A and carnocyclin A, but the time to develop resistance was longer when strains were exposed to carnocyclin A. L. monocytogenes transcriptome revealed the expression of genes involved in the mannose phosphotransferase system were down-regulated when L. monocytogenes was grown with carnocyclin A in either mannose or cellobiose by 6.87 to 7.42- log2 fold and 2.14 to 2.22- log2 fold, respectively, as compared to cultures grown without carnocyclin A. The expression of genes involved in -glucoside and cellobiose specific phosphotransferase systems was upregulated by 1.64 to 1.77- log2 fold when L. monocytogenes was grown in cellobiose, as compared to cultures grown without carnocyclin A. There were a total of 366 ncRNAs predicted from the genome, yet only 7 of them were differentially expressed for all the treatment group comparisons, including 4 T-box riboswitches, Rli47, Rli53, and Rli24. These studies suggest that leucocin A may be beneficial to the industry as a surface application on wieners to help reduce L. monocytogenes counts due to post-processing contamination even in the presence of spoilage organisms. Carbohydrates influence the growth kinetics and stress response of L. monocytogenes strains in the presence of leucocin A and carnocyclin A. The stress response and the development of resistance to carnocyclin A in L. monocytogenes FSL C1-056 may be influenced by regulatory ncRNAs. The functional genes and ncRNAs expressed are informative in understanding the impact of bacteriocins on L. monocytogenes growth and resistance. All together the efficacy of bacteriocins to control L. monocytogenes can be influenced by application methods, product composition, and strain individuality. The findings presented in this thesis will allow for a more complete understanding of the broad stress response to bacteriocins in L. monocytogenes, which can lead to the development of resistance, and how the industry can mitigate this risk for effective control of L. monocytogenes in ready-to-eat meat products.
Supplemental material related to this thesis is available at https://era.library.ualberta.ca/collections/7p88ck40x
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- Subjects / Keywords
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- Graduation date
- Fall 2019
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- Type of Item
- Thesis
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- Degree
- Doctor of Philosophy
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- License
- Permission is hereby granted to the University of Alberta Libraries to reproduce single copies of this thesis and to lend or sell such copies for private, scholarly or scientific research purposes only. Where the thesis is converted to, or otherwise made available in digital form, the University of Alberta will advise potential users of the thesis of these terms. The author reserves all other publication and other rights in association with the copyright in the thesis and, except as herein before provided, neither the thesis nor any substantial portion thereof may be printed or otherwise reproduced in any material form whatsoever without the author's prior written permission.