University of Kentucky UKnowledge Theses and Dissertations--Veterinary Science Veterinary Science 2021 AN ASSESSMENT OF THE CARRIER STATE AND A NOVEL MARKER OF LEPTOSPIRA AND ABORTION IN CENTRAL KENTUCKY HORSES Gloria Louise Gellin University of Kentucky, [email protected] Author ORCID Identifier: https://orcid.org/0000-0001-6458-7320 Digital Object Identifier: https://doi.org/10.13023/etd.2021.371 Right click to open a feedback form in a new tab to let us know how this document benefits ou.y Recommended Citation Gellin, Gloria Louise, "AN ASSESSMENT OF THE CARRIER STATE AND A NOVEL MARKER OF LEPTOSPIRA AND ABORTION IN CENTRAL KENTUCKY HORSES" (2021). Theses and Dissertations--Veterinary Science. 51. https://uknowledge.uky.edu/gluck_etds/51 This Doctoral Dissertation is brought to you for free and open access by the Veterinary Science at UKnowledge. It has been accepted for inclusion in Theses and Dissertations--Veterinary Science by an authorized administrator of UKnowledge. For more information, please contact [email protected]. STUDENT AGREEMENT: I represent that my thesis or dissertation and abstract are my original work. Proper attribution has been given to all outside sources. I understand that I am solely responsible for obtaining any needed copyright permissions. I have obtained needed written permission statement(s) from the owner(s) of each third-party copyrighted matter to be included in my work, allowing electronic distribution (if such use is not permitted by the fair use doctrine) which will be submitted to UKnowledge as Additional File. I hereby grant to The University of Kentucky and its agents the irrevocable, non-exclusive, and royalty-free license to archive and make accessible my work in whole or in part in all forms of media, now or hereafter known. I agree that the document mentioned above may be made available immediately for worldwide access unless an embargo applies. I retain all other ownership rights to the copyright of my work. I also retain the right to use in future works (such as articles or books) all or part of my work. I understand that I am free to register the copyright to my work. REVIEW, APPROVAL AND ACCEPTANCE The document mentioned above has been reviewed and accepted by the student’s advisor, on behalf of the advisory committee, and by the Director of Graduate Studies (DGS), on behalf of the program; we verify that this is the final, approved version of the student’s thesis including all changes required by the advisory committee. The undersigned agree to abide by the statements above. Gloria Louise Gellin, Student Dr. Craig N. Carter, Major Professor Dr. Daniel Howe, Director of Graduate Studies AN ASSESSMENT OF THE CARRIER STATE AND A NOVEL MARKER OF LEPTOSPIRA AND ABORTION IN CENTRAL KENTUCKY HORSES DISSERTATION A dissertation submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy in the College of Agriculture, Food and Environment at the University of Kentucky By Gloria Louise Gellin Lexington, Kentucky Director: Dr. Craig Carter, Professor of Veterinary Science Lexington, Kentucky 2021 Copyright © Gloria Louise Gellin 2021 https://orcid.org/0000-0001-6458-7320 ABSTRACT OF DISSERTATION AN ASSESSMENT OF THE CARRIER STATE AND A NOVEL MARKER OF LEPTOSPIRA AND ABORTION IN CENTRAL KENTUCKY HORSES Leptospirosis is a reemerging zoonotic infection of worldwide importance and affects all mammals. The bacterium is transmitted to animals and humans by urine, fetal membranes and body fluids. Leptospira shedding in the urine contaminates both soil and water, exposing both humans and animals to the bacterium. Leptospirosis in horses can cause abortion and is one of the etiologies of equine recurrent uveitis which can lead to blindness. Equine leptospiral abortion in Central Kentucky is primarily caused by serovar Pomona, with occasional cases attributed to serovar Grippotyphosa. There are a few reports in the literature attributing abortion to serovar Bratislava in the United States. Interestingly, Bratislava has the highest seropositivity in the horse in the United States. Two studies were conducted that are included in this dissertation. The first was to determine the prevalence of leptospirosis in horses located in Central Kentucky submitted for necropsy to the University of Kentucky Veterinary Diagnostic Laboratory. Heart, vitreous humor, kidney and urine samples were collected for microagglutination testing (MAT) and real-time PCR (qPCR). Heart blood and vitreous were tested using MAT for serovars Grippotyphosa, Pomona and Bratislava. Kidney, vitreous and urine samples were tested for pathogenic Leptospira by qPCR. MAT test results for heart blood indicated an increased seroprevalence for Bratislava as compared to Grippotyphosa and Pomona. Three horses had positive titers for serovar Bratislava in vitreous and heart blood samples. All urine samples tested negative by qPCR, and only one kidney sample had a weak positive result. Four vitreous samples tested positive for leptospirosis by qPCR, but all samples were negative upon MAT testing. No samples with positive MAT titers were positive by qPCR for any of the samples tested. Females were more likely to have positive MAT titers and were considerably older than males. MAT titers in females were also significantly higher as compared males. Finally, there was widespread seroprevalence in horses, regardless of the reason for necropsy submission. This suggests that exposure to Leptospira on Central Kentucky horse farms is common and the risk of exposure to humans and other animals is possible. The second study evaluated the enzyme heme oxygenase-1 (HO-1) and its potential as a marker for abortion in pregnant mares with elevated MAT titers. HO-1 is the rate-limiting step in the breakdown and degradation of heme to carbon monoxide, free iron and biliverdin, which is converted to bilirubin. Increased levels of HO-1 and its’ by-products are upregulated during inflammation and sepsis. HO-1 together with its by-products are essential in protecting the body from both increased inflammation and in reducing the risk of sepsis in humans, mice and rats. HO-1 together with its by-products are also essential in helping maintain pregnancy in humans, mice and rats. HO-1 in the serum of horses has not been previously investigated. The presence of HO-1 in the serum of healthy non-pregnant mares (NPM), pregnant mares throughout pregnancy (PMOT), pregnant mares 7 months pregnant with and without placentitis (PM) and pregnant mares with high MAT titers (MATS) was investigated. HO-1 levels in both NPM≥ and PMOT were lower than PM, and significantly lower than MATS. Pregnancy alone increased HO-1, the same has been shown in other mammals and humans. Additionally, mares pregnant with elevated leptospiral titers had significantly higher HO-1 levels compared to other mares. This indicates that both pregnancy and high MAT titers increase the animals’ HO-1 response. Further investigation of HO-1 in horses is needed to ascertain its’ importance during infection and/or pregnancy and potential therapeutics. KEYWORDS: Leptospirosis, Horses, Serovars, Heme Oxygenase-1, Pregnancy Gloria Louise Gellin June 30, 2021 Date AN ASSESSMENT OF THE CARRIER STATE AND A NOVEL MARKER OF LEPTOSPIRA AND ABORTION IN CENTRAL KENTUCKY HORSES By Gloria Louise Gellin Dr. Craig N. Carter Director of Dissertation Dr. Daniel Howe Director of Graduate Studies June 30, 2021 Date ACKNOWLEDGMENTS The author wishes to thank the following people: Dr. Craig Carter for his willingness to ‘take on’ a very unconventional PhD student, who chose to work full-time during the majority of this degree. His encouragement, patience, guidance and immense knowledge help make this possible even though it took a very long time! My committee members: Dr. John Timoney, Dr. David Horohov and Dr. Wayne Sanderson for their guidance, support and not giving up on me. Dr. Mary Rossano for graciously accepting to be my outside committee member. The studies for my PhD could not have been completed without the help of many people including but not limited to: Meg Steinman-your immense knowledge and friendship helped me through this; Sarah Partin and Kim England for your many hours doing MATs; Sara Welsh for her organization and collection of samples; Eva Langlois and Susan Minnis for their help in collecting samples; Dr. Erdal Erol for his guidance and willingness and trust to allow me to work in his lab on the weekends alone; Jocelynn Morgan for her endless patience, guidance and friendship; Dr. Li Zeng and Noah Forrey for their help and their kindness in always answering my questions; Dr. Jackie Smith for your endless data extraction, suggestions and encouragement; Derrick Miles, Nathan McCurdy and James Mason for all your help in data extraction and allowing me to ‘entertain you’! To my friends (too many to mention) who continue to ask me to still do things, even though I have said no many times! Your support and iii encouragement helped me make it to this point. A special thank you to Dr. Janet Lensing who didn’t think I was crazy (ok maybe a little) when I told her I had this outrageous idea about going back to school! Finally, to my family who have always been there to support me, no matter how ‘cranky’ and stressed I have been while navigating through this degree. Especially my father (Dr. M. Gellin), his love, support and patience have made this possible. His motto has always been you should never stop learning-he told this to many dental students throughout his years teaching and he continues to learn even into his ‘golden years’; both my brother (Dr. R. Gellin) and I followed his advice through numerous college degrees! My mother would have been very proud. There is always room for one more doctor in the family!!! iv TABLE OF CONTENTS ACKNOWLEGEMENTS ........................................................................................ iii LIST OF TABLES ................................................................................................ viii LIST OF FIGURES ..............................................................................................