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Application of Bacteriophage Cocktail in Leafy Green Wash Water to Control Salmonella Enterica

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Application of Bacteriophage Cocktail in Leafy Green Wash Water to Control Salmonella Enterica University of Massachusetts Amherst ScholarWorks@UMass Amherst Masters Theses Dissertations and Theses November 2015 Application of Bacteriophage Cocktail in Leafy Green Wash Water to Control Salmonella Enterica Andrea W. Lo University of Massachusetts Amherst Follow this and additional works at: https://scholarworks.umass.edu/masters_theses_2 Part of the Food Microbiology Commons Recommended Citation Lo, Andrea W., "Application of Bacteriophage Cocktail in Leafy Green Wash Water to Control Salmonella Enterica" (2015). Masters Theses. 282. https://doi.org/10.7275/7517344 https://scholarworks.umass.edu/masters_theses_2/282 This Open Access Thesis is brought to you for free and open access by the Dissertations and Theses at ScholarWorks@UMass Amherst. It has been accepted for inclusion in Masters Theses by an authorized administrator of ScholarWorks@UMass Amherst. For more information, please contact [email protected]. APPLICATION OF BACTERIOPHAGE COCKTAIL IN LEAFY GREEN WASH WATER TO CONTROL SALMONELLA ENTERICA A Thesis Presented by ANDREA WEN-YUN LO Submitted to the Graduate School of the University of Massachusetts Amherst in partial fulfillment of the requirements for the degree of MASTER OF SCIENCE September 2015 Department of Food Science © Copyright by Andrea Wen-Yun Lo 2015 All Rights Reserved APPLICATION OF BACTERIOPHAGE COCKTAIL IN LEAFY GREEN WASH WATER TO CONTROL SALMONELLA ENTERICA A Thesis Presented By ANDREA WEN-YUN LO Approved as to style and content by: _____________________________________ Amanda Kinchla, Chair _____________________________________ Lynne McLandsborough, Member _____________________________________ David Sela, Member ____________________________________ Eric Decker, Department Head Food Science ACKNOWLEDGEMENTS I would like to thank God for carrying me through this entire journey. I would also like to thank my advisor Amanda Kinchla for her guidance and oversight in this project. This experience has taught me a great deal and helped me develop important technical skills. I am also grateful for direction from Dr. Lynne McLandsborough and Dr. David Sela. The experience I gained and opportunities I had have taught me valuable life lessons. This work has been supported by the National Institute of Food and Agriculture, U.S. Department of Agriculture, the Massachusetts Agricultural Experiment Station and the Food Science Department of the University of Massachusetts Amherst, under project number MAS00440. A special thanks to my family for their support, and my lab-mates Thomas Carlisle, Kristin Wong, Sam Alcaine, Juhong Chen, Ziyuan Wang, Danhui Wang, Troy Hinkley, and Angelyca Jackson for their encouragement and advice. iv ABSTRACT APPLICATION OF BACTERIOPHAGE COCKTAIL IN LEAFY GREEN WASH WATER TO CONTROL SALMONELLA ENTERICA SEPTEMBER 2015 ANDREA WEN-YUN LO, B.S. UNIVERSITY OF CALIFORNIA IRVINE M.S. UNIVERSITY OF MASSACHUSETTS AMHERST Directed by: Professor Amanda Kinchla Produce is responsible for 46% of all foodborne illnesses in the USA. Salmonella enterica causes 19,000 hospitalizations each year, and has been associated with produce. Presently, chlorine based sanitizers are most often used, however organic matter reduces its antimicrobial activity. Bacteriophage treatments are an all-natural, alternative method for pathogen inactivation. The objective of this study was to determine the efficacy of a five-strain bacteriophage treatment against a S. enterica cocktail in simulated wash waters at different temperatures. Bacteriophage and S. enterica were enumerated in simulated wash water solutions. One set of experiments studied bacteriophage and S. enterica growth in TSB+vegetable solutions. Bacteriophage behavior was not statistically different (p < 0.05) in spinach, romaine, or iceberg lettuce across different concentrations of organic matter. S. enterica reduction was approximately 2 log over 135 minutes for vegetable solutions and for the TSB control. S. enterica reduction was only 0.5 log in water solutions. The next set of experiments studied bacteriophage and S. enterica growth in vegetable solutions. Spinach wash water and tryptone soy broth solutions (TSB) at 20 v °C and 37 °C. S. enterica was not reduced in spinach solution studies at 20 °C and 37 °C or at broth solutions at 20 °C. However, S. enterica was effectively reduced 4 log in broth solutions at 37 °C up to 7.5 hours, but grew to high levels after 24 hours. These results indicate that bacteriophage could not effectively control bacteria levels in produce wash water, and may need to be optimized. vi TABLE OF CONTENTS Page ACKNOWLEDGEMENTS ............................................................................................... iv ABSTRACT .........................................................................................................................v LIST OF TABLES ............................................................................................................ ix LIST OF FIGURES ............................................................................................................x LIST OF ABBREVIATIONS ............................................................................................ xi CHAPTER 1. INTRODUCTION ...........................................................................................................1 1.1 Objectives ..................................................................................................................2 2. BACTERIOPHAGE APPLICATIONS IN FOOD SYSTEMS: OPPORTUNITIES AND OBSTACLES .................................................................................................3 2.1 Introduction ................................................................................................................3 2.2 Bacteriophage infection cycle ....................................................................................3 2.3 History of bacteriophage in medical applications ......................................................5 2.4 Bacteriophage application of surfaces .......................................................................7 2.5 Bacteriophage applications in meat and dairy products ............................................9 2.6 Bacteriophage applications in produce safety..........................................................10 2.6.1 Background information of produce safety ......................................................10 2.6.2 Bacteriophage applications in produce ............................................................13 2.6.3 Recommendations on optimum bacteriophage applications in produce .........15 2.7 Conclusion ..............................................................................................................18 3. S. ENTERICA GROWTH IN WASH WATER SOLUTIONS .....................................19 3.1 Experimental background .......................................................................................19 vii 3.2 Materials and Methods ..........................................................................................19 3.2.1 S. enterica strain preparation ............................................................................19 3.2.2 Simulated wash water solution preparation ......................................................20 3.2.3 S. enterica growth curve studies in TSB+100 NTU vegetable solutions .........20 3.2.4 S. enterica growth curve studies in 100 NTU vegetable solutions ...................20 3.2.5 Statistical Analysis ............................................................................................21 3.3 Discussion ...............................................................................................................21 3.3.1 Growth curves of S. enterica in simulated wash water solutions with TSB ....21 3.3.2 Growth curves of S. enterica in simulated wash water solutions ...................23 3.3.3 Conclusion ......................................................................................................25 4. BACTERIOPHAGE AND S. ENTERICA BEHAVIOR IN SIMULATED WASH WATER SOLUTIONS WITH TRYPTONE SOY BROTH AND VEGETABLES ..........................................................................................................26 4.1 Experimental background ......................................................................................26 4.2 Materials and Methods .........................................................................................26 4.2.1 Simulated wash water solution preparation .....................................................26 4.2.2 Bacteriophage activity in wash water solution studies ....................................27 4.2.3 Challenge studies simulated wash water with broth and S. enterica studies ...27 4.2.4 Statistical Analysis ...........................................................................................27 4.3 Discussion of results .............................................................................................28 4.3.1 Incubation studies of bacteriophage and organic matter .................................28 4.3.2 Bacteriophage and S. enterica in simulated wash water solutions with TSB ..29 4.4 Conclusion ............................................................................................................33 viii 5. BACTERIOPHAGE AND S. ENTERICA BEHAVIOR IN SIMULATED WASH WATER SYSTEMS IN ORGANIC MATTER DERIVED FROM VEGETABLES ......................................................................................................34 5.1 Experimental
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