phenanthridine Alkaloids on Swine-pathogen Interaction and the Epidemiology of Salmonella ...

216 Pages·2015·1.39 MB·English

The Role of Benzo(c)phenanthridine Alkaloids on Swine-pathogen Interaction and the
Epidemiology of Salmonella enterica

DISSERTATION

Presented in Partial Fulfillment of the Requirements for the Degree Doctor of Philosophy
in the Graduate School of The Ohio State University

By
Valeria Artuso-Ponte, DVM
Graduate Program in Comparative and Veterinary Medicine

The Ohio State University
2015

Dissertation Committee:
Dr. Wondwossen Gebreyes, Advisor
Dr. Prosper Boyaka
Dr. Steven Moeller
Dr. Paivi Rajala-Schultz
Copyrighted by
Valeria Artuso-Ponte
2015
ABSTRACT

Salmonella enterica is one of the leading foodborne pathogens worldwide and
consumption of contaminated pork has been associated with human salmonellosis. Pork
can be contaminated with Salmonella at any point in the food chain; however, infected
pigs carrying Salmonella have been identified as the most important source of Salmonella
for the carcasses. Furthermore, several factors including stress and the composition of the
intestinal microbiome have shown to influence the dynamics of the Salmonella infection
in swine, which may explain at least partially the great variability in Salmonella shedding
among pigs under natural and experimental conditions. Pre-harvest control of Salmonella
is paramount to decrease the risk of carcass contamination and improve the safety of the
food products. This dissertation includes 2 overarching goals containing a series of five
experiments (Chapters), aimed to evaluate the effect of a novel intervention strategy to
decrease Salmonella shedding by regulating stress response due to transportation. The
first goal evaluated the effectiveness of Quaternary Benzo(c)phenanthridine Alkaloids
(QBA) supplementation on Salmonella shedding and carcass contamination. We first
developed and validated a qPCR assay to optimize the detection limit, which allowed for
the detection and quantification of low numbers of Salmonella from swine feces and
carcass swabs (Chapter 3). The qPCR was implemented to accurately quantify
Salmonella on samples collected from pigs supplemented with QBA and non-
supplemented (Chapter 4). The results showed that QBA supplementation in the feed and
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the drinking water resulted in a reduction in the proportion of Salmonella-infected
finishing pigs as well as in the numbers of Salmonella shed through the feces after
transportation to the slaughterhouse. Moreover, supplementing pigs with QBA resulted in
a reduction in the level of carcass contamination with Salmonella. This first goal also
showed that stress due transportation influence the dynamics of the Salmonella infection,
increasing Salmonella shedding and the risk of carcass contamination.
The second overarching goal (Chapters 5, 6 and 7) evaluated the effect of QBA
supplementation on factors related to the host that may influence the dynamic of
Salmonella in swine. The first experiment (Chapter 5) was designed to assess the effect
of QBA supplementation on salivary cortisol concentrations, as a measure of stress
response. Findings from this study revealed that QBA supplementation was effective in
reducing the impact of stress due to transportation, as lower salivary cortisol
concentrations were observed in QBA-supplemented pigs. These results served as the
basis for the subsequent studies (Chapter 6 and 7) designed to evaluate the effect of QBA
supplementation on the intestinal barrier function and the intestinal microbiome
respectively. We hypothesized that if QBA supplementation regulates stress due to
transportation, the integrity and permeability of the intestinal barrier would be improved
and the intestinal microbial communities would be more stable. Accordingly, the study
presented in Chapter 6 was performed to evaluate whether QBA supplementation would
result in the increase of the transepithelial electrical resistance (TER) and the decrease in
the permeability of the intestinal epithelium to molecular markers as compared to non-
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supplemented pigs. Findings showed that TER and intestinal permeability coefficients
were not different between supplemented and non-supplemented pigs. However, these
measurements were taken at a single point (after the pigs were slaughtered) and did not
provide conclusive results as to whether stress due to transportation affected the integrity
of the intestinal barrier or the potential role of QBA supplementation in ameliorating its
negative impact on the intestinal barrier function. The study presented in Chapter 7 was
designed to examine the intestinal microbial profiles in finishing pigs after transportation
to the slaughterhouse in response to QBA supplementation. The results showed an
increased microbial diversity in QBA-supplemented pigs, represented by a more even
distribution of the classified phylotypes. However, based on the findings we could not
prove that stress due to transportation affected the diversity of intestinal microbial
populations, thus the higher microbial diversity observed in supplemented pigs are likely
due to the direct effect of QBA on the microbiome or on other factors that may influence
the intestinal microbiome.

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I dedicate this thesis to my husband Santiago and to my son Pedro.
I also dedicate this thesis to my parents Raul and Cristina, to my sister Ana Paula and my
brothers Alejandro and Julian.

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ACKNOWLEDGMENTS

I would like to express my gratitude to Dr. Gebreyes for providing me with the
opportunity to pursue my Doctorate degree. I extend my sincere appreciation to the
members of my committee, Dr. Moeller, Dr. Boyaka and Dr. Rajala-Schultz for their
advice, guidance, support and commitment throughout this learning process.
I would also like to acknowledge the faculty and staff from the Department of
Veterinary Preventive Medicine. I’m especially grateful to Dr. Shuenemann for his support
and guidance. I’m grateful to my friends and colleagues Joany Van Balen and Luciana da
Costa for their help, the great times shared and their unconditional support. The appreciation
is extended to Phytobiotics for their financial support and collaboration.
Thanks to Walter, Silvia, Jesus, Evelyn, Sophia, for making this time in
Columbus more enjoyable and thank you for this lifetime friendship. Thank you to all my
Columbus friends: Jason, Natalia, Larry, Claudia, Gaby, Joel, Fernanda, Leandro,
Tatiana, Marcos, Isabel, Shuenemann family, Hoet family, Da Costa family, for all the
great moments.
Lastly, and most importantly, I would like to acknowledge Santiago Bas, my
husband, for all his support and encouragement during these years. Thanks Pedro, our
son, for filling our lives with love, joy and happiness. I would have never got this done
without their love. I am grateful to my parents Raul and Cristina for tought me that
everything is possible with hard work, dedication, sacrifice and honesty; thanks for
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always being there and for your unconditional love and support during these years.
Thanks to my sister Pauli and my brothers Ale y Juli for being always there, for making us
feel loved and close to your hearts. Thanks to Gennaro and Martiniano for making me laugh
and brighten my days. Thanks to Martin, Yoi, Gabriel, Graciela, Male, Luli, Pato, Francisco
and Clara for your support and encouragement.

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VITA

November 5, 1981……………………………………….... Born – Laguna Larga,
Argentina.
2005... ...………..…………………………………………. D.V.M Catholic University
of Córdoba, Cordoba,
Argentina.
2010 to present....…………………………………………. Graduate Research
Associate, Department of
Veterinary Preventive
Medicine, The Ohio State
University.

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PUBLICATIONS

1. Bayleyegn Molla, Allyson Sterman, Jennifer Mathews, Valeria Artuso-Ponte,
Melanie Abley, William Farmer, Paivi Rajala-Schultz, W. E. Morgan Morrow,
and Wondwossen A. Gebreyes. Salmonella enterica in commercial swine feed
and subsequent isolation of phenotypically and genotypically related strains from
fecal samples. Applied and Environmental Microbiology. 2010;76(21):7188–
7193.
2. Daniel A. Tadesse, Peter B. Bahnson, Julie A. Funk, W.E. Morgan Morrow,
Melanie J. Abley, Valeria A. Ponte, Siddhartha Thakur, Thomas Wittum, Fred J.
DeGraves, Paivi J. Rajala-Schultz, and Wondwossen A. Gebreyes. Yersinia
enterocolitica of Porcine Origin: Carriage of Virulence Genes and Genotypic
Diversity. Foodborne Pathogens and Disease. 2013;10(1):80-86.
3. Robbins RC, Artuso-Ponte VC, Moeser AJ, Morrow WE, Spears JW, Gebreyes
WA. Effects of quaternary benzo(c)phenanthridine alkaloids on growth
performance, shedding of organisms, and gastrointestinal tract integrity in pigs
inoculated with multidrug-resistant Salmonella spp. American Journal of
Veterinary Research. 2013;74(12):1530-1535.
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