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Comparision of Known Infected Fresh and Frozen Meat Samples for the Recovery of Trichinella Larvae Using the Magnetic Stirrer Digestion Method

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Comparision of Known Infected Fresh and Frozen Meat Samples for the Recovery of Trichinella Larvae Using the Magnetic Stirrer Digestion Method ITM – MSTAH thesis, No 100, 2008 COMPARISION OF KNOWN INFECTED FRESH AND FROZEN MEAT SAMPLES FOR THE RECOVERY OF TRICHINELLA LARVAE USING THE MAGNETIC STIRRER DIGESTION METHOD Dr. Vu Thi Nga “Thesis presented in partial fulfillment of the requirements for the degree of Mater of Science in Tropical Animal Health” Prince Leopold Institute of Tropical Medicine Department of Animal Health Antwerpen (Antwerp), Belgium Thesis of Master of Science in Tropical Animal Health Presented and defended on 8th of July 2008 at the Prince Leopold Institute of Tropical Medicine Antwerpen (Antwerp), Belgium Members of the jury: Dr J.Van Den Abbeele, Department of Parasitology - ITM, Chairman Dr T.Geurden, University of Ghent, ITM, External Member Prof. Dr. P.Dorny, Department of Animal Health - ITM, Promoter Prof. P.Van den Bossche, Department of Animal Health - ITM, Member Dr S.Gabriel, Department of Animal Health - ITM, Member Dr V.Delespaux, Department of Animal Health - ITM, Secretary ii Dedication This thesis is dedicated to my family, my husband Van & my son Ha Acknowledgements I am grateful to Prof. Dr. Pierre Dorny for his scholastic guidance, valuable suggestions, constructive criticisms, precious time in guiding and advising me to write the present work. I am indebted to Dr. Leen Claes for her patient, technical guidance and useful advises on the writing of this work. Special thanks go to Dr. Nicolas Praet whom assisted in the analysis data. Thanks also extend to all laboratory technicians in the department of animal health, ITM, especially to Mr. Bjorn Victor and Miss. Anke Van Hul for their technical support during work. I would like to express my special thank to the organizers of this course: Dr. Redgi De Deken, Dr. Eric Thys and all teachers from ITM. This course has definitely broadened my knowledge and given me the essential tools for my job. I would like to thank the Department of Animal Health, Institute of Tropical Medicine for the support to finance this work and to give me the opportunity to study the technique at the Trichinella Reference Centre, The French Food Safety Agency, France. Finally, I am thankful to all those who in any other way have contributed to the success of this work, which has had the financial support of the Directorate General of Development Co- operation, Belgium. iii Table of Contents Dedication ................................................................................................................................iii Acknowledgements..................................................................................................................iii Table of Contents .................................................................................................................... iv List of tables............................................................................................................................. vi List of figures ........................................................................................................................... vi List of abbreviations and acronyms .....................................................................................vii Summary................................................................................................................................viii Chapter I: Introduction........................................................................................................... 1 Chapter II: Literature review ................................................................................................. 3 2.1. Generalities.................................................................................................................. 3 2.2. Taxonomy.................................................................................................................... 3 2.3. Morphological features................................................................................................ 3 2.4. Freeze resistance.......................................................................................................... 4 2.5. Life cycle ..................................................................................................................... 4 2.6. Clinical signs ............................................................................................................... 4 2.7. Geographical distribution ............................................................................................ 5 2.8. Epidemiology............................................................................................................... 5 2.9. Diagnostic methods ..................................................................................................... 6 2.9.1. Direct detection .................................................................................................... 6 2.9.1.1. Trichinoscopy (Microscopic examination)................................................... 6 2.9.1.2. Artificial digestion ........................................................................................ 6 2.9.1.3. PCR............................................................................................................... 9 2.9.2. Indirect detection (Serology)................................................................................ 9 2.10. Situation of trichinellosis in the EU......................................................................... 9 iv 2.11. Trends and challenges of trichinellosis control in EU........................................... 10 2.12. Situation in Belgium .............................................................................................. 10 Chapter III: Materials and methods .................................................................................... 11 3.1. Materials .................................................................................................................... 11 3.1.1. Propagation of T. spiralis in the rat.................................................................... 11 3.1.2. Experimental samples ........................................................................................ 11 3.2. Methods ..................................................................................................................... 11 3.2.1. The incomplete artificial digestion method........................................................ 11 3.2.1.1. The incomplete artificial digestion method for collecting encapsulated larvae………………………………………………………………………………….11 3.2.1.2. Preparation of proficiency samples............................................................. 12 3.2.2. The artificial digestion method for recovery T. spiralis larvae.......................... 13 3.2.3. Validation method.............................................................................................. 13 Chapter IV: Results ............................................................................................................... 15 4.1. The incomplete digestion method to collect encapsulated larvae to prepare proficiency samples .............................................................................................................. 15 4.2. The artificial digestion method for recovery larvae .................................................. 15 4.2.1. Recovery of larvae from proficiency samples.................................................... 15 4.2.2. Comparison of recovered larvae between fresh and frozen meat samples ........ 19 4.2.2.1. Mean ........................................................................................................... 19 4.2.2.2. Coefficient of variation (CV %) ................................................................. 20 Chapter V: Discussion and conclusions ............................................................................... 21 Chapter VI: Recommendations ............................................................................................ 23 Bibliographical references..................................................................................................... 24 v List of tables Table I: Number of countries with documented Trichinella spp. infections in animals and human...............................................................................................................................................5 Table II: Methods used for meat inspection for Trichinella spp. in the EU ....................................8 Table III: Experimental samples ....................................................................................................11 Table IV: The results of recovered larvae from proficiency samples ............................................16 Table V: Mean of recovered larvae from proficiency samples......................................................19 Table VI: Coefficient of variation of recovered larvae from proficiency samples ........................20 List of figures Figure I: Encapsulated larvae.........................................................................................................15 Figure II: Preparation of proficiency samples................................................................................15 Figure III: Coiled larvae recovered from fresh meat samples........................................................15 Figure IV: Uncoiled larvae recovered from frozen meat samples .................................................15 Figure V: Graph of recovered larvae from fresh and frozen meat samples containing 3 larvae
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