EFFECTS OF TEMPERATURE ON THE DEWLOPMENT OF LOM4 SALMONAE AND RESISTANCE TO RE-INFECTION IN RAINBOW TROUT (ONCORHYnCHUS IMYKlSs) A Thesis Subrnitted to the Graduate Faculty in Partial Fulfilment of the Requirements of the Degree of Master of Science in the Department of Pathology and Microbiology Faculty of Veterinary Medicine University of Prince Edward Island Holly J. Beaman Charlottetown, PEI September 1998 01998. Holly J. Bearnan National Library Bibliothèque nationale u.1 of Canada du Canada Acquisitions and Acquisitions et Bibliographie Services services bibliographiques 395 Wellington Street 395. rue Wellington Ottawa ON K1A ON4 OttawaON K1AON4 Canada Canada The author has granted a non- L'auteur a accordé une licence non exclusive licence allowing the exclusive permettant à la National Library of Canada to Bibliothèque nationale du Canada de reproduce, loan, distribute or sell reproduire, prêter, distribuer ou copies of this thesis in microform, vendre des copies de cette thèse sous paper or electronic formats. la forme de microfiche/nlm, de reproduction sur papier ou sur format électronique. The author retains ownership of the L'auteur conserve la proprieté du copy~@t in this thesis. Neither the droit d'auteur qui protège cette thèse. thesis nor substantial extracts from it Ni la thèse ni des extraits substantiels may be printed or otherwise de celle-ci ne doivent être imprimes - reproduced without the author's ou autrement reproduits sans son permission. autorisation. CONDITION OF USE The author has agreed that the Library, University of Prince Edward Island, may make this thesis fieely available for inspection. Moreover, the author has agreed that permission for extensive copying of this thesis for scholarly purposes may be granted by the professor or professors who supervised the thesis work recorded herein or, in their absence, by the Chairman of the Department or the Dean of the Faculty in which the thesis work was done. It is undentood that due recognition will be given to the author of this thesis and to the University of Prince Edward Island in any use of the material in this thesis. Copying or publication or any other use of the thesis for financiai gain without approval by the University of Prince Edward Island and the author's written permission is prohibited. Requests for permission to copy or to make any other use of material in this thesis in whole or in part should be addressed to: Chairman of the Department of Pathology and Micro Faculty of Vetennary Medicine University of Prince Edward Island Charlottetown, PEI Canada C 1A 4P3 Loma salmonne, an economically important microsporidian parasite of farm-reared Pacific salrnon, causes severe branchial infections. No treatment currently exists to control L. salmonae infections. Consequently, other mechanisms of control are essentid. Water temperature has a regdatory effect on the life cycle and development of many pathogens and has been used to control some infectious diseases. It may be used as a means of controlling parasitic infections with Loma salmonae; however, Iittle is known about the relevant pathobiology of this parasite. Objectives of this research were to determine the permissive developmental range for the life cycle of L. sulmonae, the effect of temperature on xenoma development rate and the abundance of branchial xenomas that develop, and to examine resistance induction for controlling parasitic infections. Juvenile rainbow trout reared in freshwater were exposed to infective tissue by either intubation or feeding of infective gill material. They were then reared at water temperatures of SO,7", 9", 1O*, 1 1O, 1 3 O, 1 5O, 17", 19", 20" or 21°C. Fish developed branchial xenomas within the range of 9°C to 20°C, indicating the permissive temperature range for L. sulmonae. Additionaily? rate of development was significantly faster at warmer water temperatures. Since exposure time was known, a polynomial regression model and a thermal unit summation model (OC- days) were useful for predicting time of xenoma onset. Due to variation in the length of the xenomas remained in the gills, no mode1 could be developed for predicting clearance time of the parasite fiom the gills. Quantification of branchial xenomas was conducted at water temperatures of 11 O, 13 O, 15", 17O and 19°C. The number of xenomas per gill filament was assessed to determine if temperature affected intensity of infection after al1 fish received the same initial dose via gastric intubation. The number of xenomas that developed in fish at al1 temperatures varied considerably among fish; there appears to be no effect of temperature. Upon reinfection of fish at 15"C, few to no xenomas were detected, regardless of water temperature during initial exposure. This suggests that it is not necessary for L. salmonae to complete its life cycle to invoke an immune response in the fish host. The relative percent protection model developed for the re-infection tnals indicated a high level of protection in fish which were previously exposed to L. salmonae. A potential procedure for controlling L. salmonae infections in farm-reared salmonids has resulted fiom the triaIs conducted herein. No prior studies have modelled development rates of fish pathogens, dthough the model developed in this thesis has significant potential for many fish parasites whose infective stages and development are dependent on water temperature. Fish can be exposed to the parasite at non-permissive temperatures which do not allow completion of the life cycle but induce significant protection against MerL. salmonae infections. ACKNOWLEDGMENTS 1 would like to extend my deepest gratitude to my supervisor, Dr. David Speare, for his constant enthusiastic support and guidance throughout my program. Your words of wisdom are greatly appreciated. 1 would also like to thank the members of my supervisory cornmittee, Drs. F. Markham, R. Cawthom, B. Horney, S. Jones and M. Brimacombe, who have dlmade helpful contributions dong the way. The technical support of Garth Arsenault was especially appreciated as 1was getting started in my program. Also, th& to Joanne Daley and others in the aquatic facilities who have lent a hand when needed. Audiovisual SeMces, Computer SeMces and the departmental secretaries were also very helpful and are highty appreciated. I value every fiiendship 1have gained at AVC, dl mean a great deal to me. Special thanks to Ryan Dickinson, Karen Harris, Crystal Trevors, Joanne Bowers, Huub Brouwers, Chris and Pat Campbell, Sandie Momson, Genaro Sanchez, John Ficele, Carolyn Baglole, Linda MacLean, Norma Guy, Jean Lavallée, Tanja Bihr, Gabino Pefialver, Judy Macintyre. Debby LeBlanc, and probably others 1 am forgetting. You have al1 played a part in making my experience here a great one. 1 owe specid appreciation to both my parents, Doug and Dawn for encouraging me to continue my education and lem a great deal more about the aquatic world. Finally. thanks to Tim for being incredibly supportive and patient. 1 acknowledge the financial support 1 received through an Natural Sciences and Engineering Research Council Strategic Grant. TABLE OF CONTENTS TitieofThesis .......................................................... i .. ConditionsofUse ...................................................... ri ... Permission to Use Postgraduate Thesis ...................................... III Certification of Thesis Work .............................................. iv Abstract ............................................................... v Tableofcontents ...................................................... vii ListofTables ......................................................... xii List of Abbreviations ................................................... xiv 1. GENERALINTRODUCTION ...................................... I 1.1 Aquaculture ............................................... 1 11History ............................................... 1 1.1.2 Current Status ........................................ -2 1.1.3 Sahonid Culture ...................................... 4 1.1 .4 Biological Constraints ................................... 5 1.2 Fish Health and Disease ....................................... 6 1.2.1 Factors Influencing Disease Onset in Cultured Fish ............ 7 1.2.2 Infectious Diseases of Salmonids .......................... 8 1.2.3 Protozoan Parasites ..................................... 9 1.3 Microsporidian Parasites ..................................... 10 1.3.1 Aquaculture Facilities .................................. L 1 1.3.2 Microsporidian Life Cycle .............................. 12 1.3.3 Diagnosis ............................................ 16 1.3.4 Treatrnent ........................................... 17 1.3-5 Tissue Reactions to Xenomas ............................1 9 1.3-6 Phagocytosis of Spores ................................. 20 1.4 Loma salmonae ........................................... -21 1.4.1 Distribution ....................................... 22 vii 1.4.2 EEects on Lnfected Fish ............................... -24 1.4.3 Other Loma spp ...................................... -25 1.5 Thermal Regulation ........................................ -26 1.5 .1 Effect of Temperature on Fish Development ............... -26 1S.2 EEect of Temperature on Immune Function ................-26 1S.3 EEect ofTemperature on Parasite Development ............ -27 1.6 Disease Resistance ......................................... -30 1.7 Research Objectives ........................................ -31