comparison of naturally-acquired and vaccine-acquired immunity to loma salmonae in rainbow trout

139 Pages·2013·6.6 MB·English

COMPARISON OF NATURALLY-ACQUIRED AND VACCINE-ACQUIRED
IMMUNITY TO LOMA SALMONAE IN RAINBOW TROUT (ONCORHYNCHUS
MYKISS)
BY
JENNIFER E. HARKNESS
A Thesis
Submitted to the Graduate Faculty
in Partial Fulfillment of the Requirements
for the Degree of
MASTER OF SCIENCE
Department of Pathology and Microbiology
Faculty of Veterinary Medicine
University of Prince Edward Island
©APRIL 2012. J.E. HARKNESS
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University of Prince Edward Island
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ABSTRACT
Microsporidial Gill Disease of Salmon (MGDS) is a result of infection with Loma salmonae and
has been described in fanned and wild Pacific salmon as causing severe branchitis, resulting in
significant economic losses in aquaculture, as fish succumbing to the disease are those nearing
market size. No treatment currently exists to control L. salmonae, although previous studies have
documented disease resistance upon initial recovery and secondary exposure to the parasite. The
objectives of this study were to further examine naturally acquired as well as vaccine-acquired
immunity to L. salmonae in rainbow trout (RBT). It is believed that farm reared salmon are
exposed to continual low doses of the parasite over time, and the naturally acquired immunity
studies focused on the protective response generated by recovery of a low dose cohabitation
exposure and subsequent high dose oral re-challenge. Having the ability to partially protect RBT
upon re-exposure to L. salmonae contradicted what is being seen in the netpen scenario, as
Chinook and Coho salmon are assumed to have no immunity to re-infection. The investigation
into vaccine-acquired immunity focused on the use of heterologous live vaccines, containing
Glugea anomala and Glugea hertwigi, respectively. Having common characteristics and being
phylogenetically related to L. salmonae, the intraperitoneal (IP) injection into RBT 6 weeks pre­
exposure to L. salmonae was able to reduce xenoma burden significantly as compared to naive
controls. Since live vaccines are not marketable, further studies investigated the use of killed
heterologous (G. anomala and G. hertwigi) vaccines in comparison with the widely published
killed homologous vaccine containing Loma sp. The superiority of both the killed heterologous
vaccines were noted, as they were able to reduce xenoma burden upon exposure to the parasite by
upwards of 15% compared to the killed homologous vaccine. As a final approach, naturally-
acquired immunity was revisited to determine if prophylactic oral treatment of immunostimulant,
Provale™, a 0-1,3/1,6 glucan in synergy with low dose recovery could significantly boost the
immune response of RBT to L. salmonae. Little evidence was found to support this hypothesis.
The immune responses and consequently protection to MGDS in RBT depends greatly upon the
intensity of the primary infection and subsequent recovery. Timing of vaccination is essential to
the protection of RBT to L. salmonae. Although RBT is a useful surrogate model for infection
with MGDS, it does not mimic the exact severity of infection seen in Pacific salmon, particularly
Chinook and Coho salmon. With this in mind, experimental studies mimicking those carried out
in the thesis in Chinook salmon would be beneficial to the control and treatment of L. salmonae.
v
ACKNOWLEDGEMENTS
I wish to thank my supervisor Dr. David Speare for his constant support, creativity, sincerity and
encouragement throughout my program. I greatly respect your enthusiasm as a research scientist
and supervisor and appreciate your honesty and personal integrity. I would also like to thank the
other members of my Supervisory Committee, Drs. Fred Markham, Spencer Greenwood, Collins
Kamunde, Fred Kibenge and the members of my Examination Committee, Drs. David Speare,
Gary Conboy, Collins Kamunde, Mark Fast and Spencer Greenwood for reviewing and correcting
my thesis.
I would still be wandering around the Aquatic Facility staring at empty tanks without the support
and encouragement of Dr. Nicole Guselle. As you can imagine, she has had a significant hand in
the initialization and continuation of all the experimental aspects of the thesis. In addition, many
thanks and gratitude to the AVC Aquatic Facility staff including Wayne Petley, Angela Hamish-
Driscoll and Lee Dawson, I couldn’t have achieved this without your tireless work ethics to
maintain an amazing facility.
Thanks to Diane MacLean and Rita Saunders for their administrative assistance and Kathy Jones
for her photography assistance.
I am extremely grateful to have met so many interesting people over the past few years and wish
to thank everyone who has had a hand in the completion of this thesis, whether it was through
guidance, having a shoulder to cry on, listening to weekly, if not daily rants of a graduate student,
or simply being there in a time of need. To each and every one of you, I thank you for your
kindness, support, friendship and love.
Thanks to my parents Donna McKinnon and Michael Poczynek, to my sister Sarah and to the rest
of my family for their continued support, encouragement, constructive criticism, and ultimately
love. You all hold a special place in my heart.
Special thanks goes to my dearest of friends, Jessica Fry and Angela Hamish-Driscoll. You have
both made me a better person and I love you both dearly.
A very large, and very deserving thanks goes to the hundreds of fish that were used in my
research, because without them, this thesis would have never been possible. Their sacrifice is
truly appreciated and will serve the greater good of the future of aquaculture.
I acknowledge the financial support I received through a Natural Sciences and Engineering
Research Council Strategic Grant, the Springboard Atlantic Fund and the Department of
Pathology and Microbiology at the Atlantic Veterinary College (UPEI).
DEDICATION
To my grandparents,
Della and Burleigh MacKinnon.
Grammy, I hope I’ve made you proud!
Grampy, your love of fish and wildlife resonates in me!
Thank you.
Love Always,
Jenny Penny
vii
TABLE OF CONTENTS
Title of Thesis i
Conditions of Use ii
Permission to Use Postgraduate Thesis iii
Certification of Thesis Work iv
Abstract v
Acknowledgements vi
Dedication vii
Table of Contents viii-x
List of Figures xi
List of Tables xii
List of Abbreviations xiii
1. GENERAL INTRODUCTION
1.1 Aquaculture 1
1.1.1 Global Production 1
1.1.2 Canadian Production 2
1.1.3 Salmonid Production 4
1.1.4 Chinook Salmon 5
1.1.5 Netpens 7
1.2 Disease in Netpens 9
1.2.1 Diseases of Chinook Salmon 10
1.2.2 Protistan Diseases of Salmon in Seawater 11
1.3 Microsporidian Parasites 13
1.3.1 Microsporidian Life Cycle 15
1.3.2 Microsporidians in Human and Animal Health 18
1.4 Loma salmonae 20
1.4.1 Transmission 21
1.4.2 Diagnosis and Clinical Signs 22
1.4.3 Immunological Resistance 26
1.4.4 Drug Treatment 29
1.4.5 Vaccines Against Loma salmonae 32
1.5 Research Objectives 33
1.6 REFERENCES 35
2. COMPARATIVE RESISTANCE TO LOMA SALMONAE IN RAINBOW TROUT
(ONCORHYNCHUS MYKISS) SUBSEQUENT TO RECOVERY FROM AN
INITIAL LOW DOSE COHABITATION EXPOSURE
2.1 ABSTRACT 48
2.2 INTRODUCTION 49
2.3 MATERIALS AND METHODS 52
2.3.1 Sample Population 52
2.3.2 Experimental Design 52
2.3.3 Methods of Infection 53
2.3.4 Challenge Dose Quantification 53
2.3.5 Sampling and Infection Assessment 53
2.3.6 Data Analysis 54
2.4 RESULTS 55
2.5 DISCUSSION 58
2.6 REFERENCES 64
FIRST DEMONSTRATION OF THE PARTIAL EFFICACY OF A
HETEROLOGOUS VACCINE AGAINST LOMA SALMONAE, USING SPORES
FROM TWO SPECIES OF GLUGEA
3.1 ABSTRACT 68
3.2 INTRODUCTION 69
3.3 MATERIALS AND METHODS 76
3.3.1 Sample Population 76
3.3.2 Experimental Design 77
3.3.3 Methods of Infection 79
3.3.4 Challenge Dose Quantification 79
3.3.5 Sampling and Infection Assessment 80
3.3.6 Data Analysis 80
3.4 RESULTS 80
3.5 DISCUSSION 83
3.6 REFERENCES 86
RESISTANCE TO AN INFECTION WITH LOMA SALMONAE IN RAINBOW
TROUT (ONCORHYNCHUS MYKISS) VACCINATED WITH HETEROLOGOUS
AND HOMOLOGOUS KILLED SPORES AND WITH OR WITHOUT
SUBSEQUENT ORAL ADMINISTRATION OF PROVALE™, A p-1,3/1,6
GLUCAN
4.1 ABSTRACT 91
4.2 INTRODUCTION 92
4.3 MATERIALS AND METHODS 96
4.3.1 Sample Population 96
4.3.2 Experimental Design 97
4.3.3 Methods of Infection 100
4.3.4 Challenge Dose Quantification 101
4.3.5 Sampling and Infection Assessment 101
4.3.6 Data Analysis 102
4.4 RESULTS 102
4.5 DISCUSSION 106
4.6 REFERENCES 110
GENERAL DISCUSSION 114
5.1 REFERENCES 119
APPENDIX A 122
APPENDIX B 123
APPENDIX C
APPENDIX D
APPENDIX E