Sustainable control of gastrointestinal parasite infections in ruminants Word count: "< 28.533 >" Laurens Zwanenburg Student number: 01509101 Supervisor: Prof. dr. Edwin Claerebout Supervisor: Prof. dr. Stig Thamsborg A dissertation submitted to Ghent University in partial fulfilment of the requirements for the degree of Master of Veterinary Medicine Academic year: 2017 - 2018 Ghent University, its employees and/or students, give no warranty that the information provided in this thesis is accurate or exhaustive, nor that the content of this thesis will not constitute or result in any infringement of third-party rights. Ghent University, its employees and/or students do not accept any liability or responsibility for any use which may be made of the content or information given in the thesis, nor for any reliance which may be placed on any advice or information provided in this thesis. Acknowledgements First of all, many thanks to Prof. dr. E. Claerebout and Prof. dr. S. Thamsborg for their supervision and guidance with the development and implementation of this master thesis. Both supervisors allowed me to work independently within their departments and provided helpful feedback on the results of the study. Furthermore, I would like to thank Prof. dr. Peter Geldhof and O. Paerewijck for their technical support with the resazurin-reduction assay, which was conducted at the University of Ghent. For the larval- migration-inhibition assay, which was conducted at the University of Copenhagen, I would like to thank Prof. dr. A. Williams for his technical support. Finally, I would like to acknowledge both universities and the Erasmus+ Programme for making the international exchange possible. Content Overview of used abbreviations................................................................................................................................................... 5 Abstract ....................................................................................................................................................................................... 6 1. Literature study .................................................................................................................................................................. 7 1.1. Gastrointestinal nematodes ..................................................................................................................................... 7 1.1.1. Introduction .................................................................................................................................................... 7 1.1.2. Epidemiology ................................................................................................................................................. 9 1.1.3. Pathogenesis and clinical signs .................................................................................................................... 10 1.1.4. Diagnostics .................................................................................................................................................. 11 1.1.5. Current control and anthelmintic resistance .................................................................................................. 13 1.1.6. Alternative control ........................................................................................................................................ 15 1.1.6.1. Management systems ............................................................................................................................. 16 1.1.6.2. Biological control ..................................................................................................................................... 17 1.1.6.3. Dietary supplementation .......................................................................................................................... 18 1.2. Giardia duodenalis ................................................................................................................................................. 20 1.2.1. Introduction .................................................................................................................................................. 20 1.2.2. Epidemiology and prevalence ...................................................................................................................... 21 1.2.3. Pathogenesis and clinical signs .................................................................................................................... 22 1.2.4. Diagnostics .................................................................................................................................................. 23 1.2.5. Current treatment ......................................................................................................................................... 24 1.2.6. Alternative control ........................................................................................................................................ 25 2. Issue and purpose ............................................................................................................................................................ 26 3. Materials and methods ..................................................................................................................................................... 27 3.1. Larval-migration-inhibition assay (LMIA) ................................................................................................................ 27 3.1.1. Parasites ...................................................................................................................................................... 27 3.1.2. Chemicals and compounds .......................................................................................................................... 27 3.1.3. Protocol ........................................................................................................................................................ 28 3.1.4. LMIA on different DMSO levels .................................................................................................................... 30 3.1.5. LMIA on different bioactive forages .............................................................................................................. 30 3.2. Resazurin-reduction assay ..................................................................................................................................... 31 3.2.1. Chemicals and compounds .......................................................................................................................... 31 3.2.2. The protocol ................................................................................................................................................. 31 3.2.3. Evaluating different DMSO levels ................................................................................................................. 32 3.2.4. Resazurin-reduction assay with different compounds ................................................................................... 33 3.3. Statistical analysis ................................................................................................................................................. 33 4. Results ............................................................................................................................................................................. 34 4.1. Larval-migration-inhibition assay ............................................................................................................................ 34 4.2. Resazurin-reduction assay ..................................................................................................................................... 37 5. Discussion ........................................................................................................................................................................ 39 6. References ....................................................................................................................................................................... 43 Appendices ............................................................................................................................................................................... 60 Appendix 1: larval-migration-inhibition assay ........................................................................................................................ 60 Appendix 2: resazurin-reduction assay ................................................................................................................................. 64 4 Overview of used abbreviations AR Anthelmintic resistance BZ (Pro)benzimidazoles CPG Cysts per gram dH2O Demineralised sterile-filtered water ELISA Enzyme-linked immunosorbent assay EPG Eggs per gram FEC Faecal egg count GIN Gastrointestinal nematodes LMIA Larval-migration-inhibition assay MIC Minimum inhibitory concentration ML Macrocyclic lactones PGE Parasitic gastroenteritis PCR Polymerase chain reaction TST Targeted selective treatment TT Targeted treatment 5 Abstract – Gastrointestinal parasite infections, particularly caused by strongyle nematodes and Giardia, are considered to be a major threat regarding health, animal welfare and productivity in livestock. Although drugs for treatment against gastrointestinal nematodiosis are available, the use of those anthelmintics is