Effects of rearing conditions on growth, development and moulting in European lobster (Homarus gammarus) Submitted by Karen Lewanne Middlemiss to the University of Exeter as a thesis for the degree of Master of Science by Research in Biosciences, August 2014. This thesis is available for Library use on the understanding that it is copyright material and that no quotation from the thesis may be published without proper acknowledgement. I certify that all material in this thesis which is not my own work has been identified and that no material has previously been submitted and approved for the award of a degree by this or any other University. (Signature) ……………………………………………………………………………… 1 Acknowledgements Firstly, to my academic supervisor Associate Professor Dr. Rod Wilson for his guidance and exacting standards, which will hold me in good stead throughout my career. This has been a hugely rewarding project and I’m grateful for all your support, encouragement, and to have benefited from your wealth of experience and ideas throughout. It’s been a pleasure working with you. To Dr. Mauricio Urbina for helping to shape my scientific thought processes during our many enjoyable conversations. For your time and advice in the lab (especially statistical analysis and on all things crustacea), and for making me think about how to best write with the reader in mind. You have helped make this such an enjoyable experience. I am forever grateful that our career paths crossed and hope they will do so again in the future. To Dr. Esme Robinson, whom without, all of this would have remained in my imagination. From our time spent involved with research in Antarctica where you helped to plant the seed of exciting possibilities that a new career in science could offer, and your continued encouragement long after. You are a true friend and respected colleague. Thank you. I would also thank the National Lobster Hatchery, Padstow, Cornwall, UK, for provision of all animals used and for providing the facilities, staff assistance and the opportunity to undertake the research completed in Chapter 2 at the hatchery. I learned a great deal about both the aquaculture industry and crustacean larviculture during the 4 months spent working at your research facility, and enjoyed the experience greatly. And finally, to my late mother Lois, who is with me in spirit and whose love and encouragement to ‘never stop learning’ guided me here and stays with me always. I dedicate this work to you. 2 Table of Contents Acknowledgements....................................................................................................................2 Contents......................................................................................................................................3 List of table and figures..............................................................................................................5 Authors Declaration..............................................................................................................6 Chapter One - General Introduction....................................................................................7 Chapter Two – UV irradiation, ozonation and probiotic water treatments Abstract................................................................................................................................21 1. Introduction.........................................................................................................................22 2. Materials and Methods.....................................................................................................25 2.1 Experimental animals................................................................................................25 2.2 Rearing systems.........................................................................................................26 2.2.1 Experimental design.........................................................................................26 2.2.2 System management.......................................................................................27 2.3 Water treatment factors.............................................................................................27 2.3.1 Ozone..................................................................................................................27 2.3.2 Probiotic..............................................................................................................28 2.3.3 UV........................................................................................................................28 2.4 Bacteriology................................................................................................................28 2.5 Growth and survival...................................................................................................28 2.6 Osmoregulatory challenge test.................................................................................29 2.7 Statistical analysis......................................................................................................30 3. Results................................................................................................................................30 3.1 Growth and survival...................................................................................................30 3.2 Osmoregulatory challenge test.................................................................................33 3.3 Bacteriology................................................................................................................34 4. Discussion..........................................................................................................................35 5. Conclusion.........................................................................................................................41 6. Acknowledgements...........................................................................................................42 7. References.........................................................................................................................42 Chapter Three – Branchial grooming setae and microbial control Abstract .....................................................................................................................................50 1. Introduction........................................................................................................................51 2. Materials and methods ....................................................................................................54 2.1 Animals and experimental design............................................................................54 2.2 Scanning electron microscopy.................................................................................54 3 2.3 Statistical analysis......................................................................................................55 3. Results................................................................................................................................56 3.1 Branchial chamber structures and setal cleaning appendages...........................56 3.2 Microbial proliferation on gills...................................................................................59 4. Discussion..........................................................................................................................61 4.1 Gill grooming setae....................................................................................................61 4.2 Branchial cleaning mechanisms...............................................................................62 4.3 Microbial proliferation on gills...................................................................................64 5. Conclusions.......................................................................................................................66 6. Acknowledgements...........................................................................................................67 7. References ........................................................................................................................67 Chapter Four – Calcium and acid-base fluxes during a moult cycle Abstract......................................................................................................................................72 1. Introduction.......................................................................................................................72 2. Materials and methods.....................................................................................................76 2.1 Animals........................................................................................................................76 2.2 Experimental setup....................................................................................................76 2.3 Ion analysis.................................................................................................................77 2.4 Calculations.................................................................................................................78 2.5 Statistical analysis......................................................................................................78 3. Results................................................................................................................................79 3.1 Total body calcium.....................................................................................................79