Isolation, characterization and evaluation of virulent bacteriophages against Listeria monocytogenes

Mastura Akhtar; Stelios Viazis; Kyle Christensen; Phillip Kraemer; Francisco Diez-Gonzalez

doi:10.1016/j.foodcont.2016.12.035

Isolation, characterization and evaluation of virulent bacteriophages against Listeria monocytogenes

Mastura Akhtar, Stelios Viazis, Kyle Christensen, Phillip Kraemer, Francisco Diez-Gonzalez

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17 Scopus citations

Abstract

Bacteriophages are considered a potential antimicrobial food intervention due to their ability to inactivate target bacteria in different environments. This study was undertaken to isolate, characterize, and identify a mixture of phages that could be used for control of Listeria monocytogenes. A total of 37 phages were isolated from raw sewage sludge using L. monocytogenes serotypes 4b, 1/2a and 1/2b strains. The host range of these phages was determined by spot-testing. Ten lytic phages were selected for further evaluation with efficiency of plating (EOP) analysis and characterization by pulsed field gel electrophoresis (PFGE). According to the spot tests and EOP results, the isolated phages against L. monocytogenes 4b strains were found to be capable of lysing a wider range of ribotypes. Also, three phages specific for 1/2a and 1/2b serotypes had similar lysis capabilities over a range of target Listeria with high EOP values between 0.5 and 1.0. PFGE analysis identified 5 distinct restriction profiles of the 10 selected phages whose DNA was digested with either EcoRI or HinDIII. Transmission electron microscopy (TEM) pictures revealed that five selected phages belonged to the Myoviridae and one phage to the Podoviridae family. Based on screening results, a phage cocktail (LP1) comprised of six lytic phages was used to inactivate a mixed culture of five L. monocytogenes strains. LP1 was able to lyse bacterial cells within 30–60 min at multiplicity of infection values (MOI, PFU/CFU) of 1000 and maintained the target cells under detection levels for 24 to 48 h at 4, 10, and 30 °C. These results suggested the LP1 phage collection contained effective and strongly lytic phages that may serve as antibacterial preparation to control L. monocytogenes of different serotypes.

Original language	English (US)
Pages (from-to)	108-115
Number of pages	8
Journal	Food Control
Volume	75
DOIs	https://doi.org/10.1016/j.foodcont.2016.12.035
State	Published - May 1 2017

Bibliographical note

Funding Information:
We are grateful to Prof. Dr. Martin J. Loessner and Dr. Jochen Klumpp, Institute of Food Science and Nutrition, Zürich, Switzerland, for their help and guidance with the methodology of phage isolation. We would like to thank I. Scott Joseph, assistant business unit manager, and Angella Gardner, program supervisor, Seneca Wastewater Treatment Plant, Eagan, MN for their help to provide sewage samples for this study. We also appreciate the support of Dr. David Smith, professor, Food Science and Nutrition, University of Minnesota with sample collection from the wastewater treatment plants in MN. This study was funded by the U. S. Department of Agriculture's Integrated Organic Program under award No. 2007-51300-03796.

Publisher Copyright:
© 2016 Elsevier Ltd

Keywords

Antibacterial
Bacteriophages
Food safety
Listeria control
Listeria phages
Lysis

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abstract = "Bacteriophages are considered a potential antimicrobial food intervention due to their ability to inactivate target bacteria in different environments. This study was undertaken to isolate, characterize, and identify a mixture of phages that could be used for control of Listeria monocytogenes. A total of 37 phages were isolated from raw sewage sludge using L. monocytogenes serotypes 4b, 1/2a and 1/2b strains. The host range of these phages was determined by spot-testing. Ten lytic phages were selected for further evaluation with efficiency of plating (EOP) analysis and characterization by pulsed field gel electrophoresis (PFGE). According to the spot tests and EOP results, the isolated phages against L. monocytogenes 4b strains were found to be capable of lysing a wider range of ribotypes. Also, three phages specific for 1/2a and 1/2b serotypes had similar lysis capabilities over a range of target Listeria with high EOP values between 0.5 and 1.0. PFGE analysis identified 5 distinct restriction profiles of the 10 selected phages whose DNA was digested with either EcoRI or HinDIII. Transmission electron microscopy (TEM) pictures revealed that five selected phages belonged to the Myoviridae and one phage to the Podoviridae family. Based on screening results, a phage cocktail (LP1) comprised of six lytic phages was used to inactivate a mixed culture of five L. monocytogenes strains. LP1 was able to lyse bacterial cells within 30–60 min at multiplicity of infection values (MOI, PFU/CFU) of 1000 and maintained the target cells under detection levels for 24 to 48 h at 4, 10, and 30 °C. These results suggested the LP1 phage collection contained effective and strongly lytic phages that may serve as antibacterial preparation to control L. monocytogenes of different serotypes.",

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Isolation, characterization and evaluation of virulent bacteriophages against Listeria monocytogenes

Abstract

Bibliographical note

Keywords

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