Nutritional Resources for Pollinators from Mass-Flowering Crop Cultivars Thesis submitted for the degree of Doctor of Philosophy Department of AgroEcology, Rothamsted Research & Institute of Neuroscience, Newcastle University Jonathan Carruthers April 2016 Abstract The scarcity of flowers to provide dietary nectar and pollen is a key driver of recent declines in pollinators in agricultural areas, but the planting of mass- flowering crops enhances resources available to pollinators during parts of the year. This thesis investigates the nutritional resources provided for insect pollinators from various cultivars of two mass-flowering crops: short rotation coppice willow (Salix species) and oilseed rape (Brassica napus L.). Willow cultivars vary in the numbers of flowers produced per plant, as well as in the quantity of nectar sugar secreted by those flowers. There were neither qualitative nor quantitative differences in pollen production between the cultivars. Foraging insect pollinators showed preferences for cultivars with more rewarding flowers. Oilseed rape flowers of different cultivars produced a mass of nectar sugar that varied by up to three fold in mass when grown in a glasshouse. Cultivars differed in the size of their flowers, but neither flower size nor the seed yields they produced in industry trials were correlated with their nectar yields. When plants were grown in field conditions, differences between oilseed rape cultivars in nectar production were also present, although less pronounced. The weights of bumble bee (Bombus terrestris L.) colonies diverged after two weeks when restricted to foraging on plots containing either a high or a low nectar yielding cultivar. The findings indicate that efforts to breed and to plant more widely the more rewarding cultivars of mass-flowering crops would enhance the resources available to pollinators in spring. As this is a critical time for pollinators, the extra resources could aid their survival and lead to more robust populations. i ii Acknowledgements For their advice and guidance over the last four years, I would like to thank my supervisors: Dr Alison Haughton and Dr Sam Cook at Rothamsted Research, Professor Geraldine Wright at Newcastle University, and Professor Juliet Osborne at the University of Exeter. Thank you for the energy and perspective that each of you brought to this project; you make a great advisory team. To my supervisors at Rothamsted especially, thank you for all the discussions and encouragement during the writing stage. This work was made possible by funding from the Biotechnology and Biological Sciences Research Council. For growing the oilseed rape plants used in my trials, my thanks go to the team at the Rothamsted farm, and the glasshouse support staff, particularly Jill Maple, Richard Parkinson and Jack Turner, who looked my plants so well, and looked after me a bit too. I wish to thank Mark Nightingale at Elsoms Seeds for helpful discussions about oilseed rape, and Will Macalpine for introducing me to the willows at Rothamsted and answering my many questions about them. A very big thank you goes to Suzanne Clark for advice on statistical techniques, and for the insightful conversations that helped me to make sense of them. Thanks are also due to Eileen Power, for introducing me to the delights of HPLC, and to Dan Stabler, for going well beyond the call of duty to help me use the technique, and for his unfailing cheerfulness that kept me from despairing about my peaks. The Bee Group at Rothamsted, including past and present members, has been a constant source of enthusiasm for the science, and of practical help. It was a pleasure to work alongside Trish Wells, Beth Nicholls, Stephan Wolf, Pete Kennedy, and Matthias Becher, as well as Jenny Swain and Jason Lim, who also got me unstuck in a literal sense on one heroic occasion. iii I want to thank my friends who saw me through the highs and lows of the last four years. At Rothamsted, Aislinn Pearson and Joe Helps have been brilliant and kept me going during the writing stage, while Helen Metcalfe, Hayley Jones, Laura Seamons, Tristan Eagling and Joanna Scales did a lot to make my first three years here fun, and were much missed afterwards. Special thanks also go to Jack Wall, Dafydd Launder, Reiss Reid, Alex Ball, Marcus Keatinge, Jasper van Heusden, Simon Walton, Richard Scott and Will Hauwert; you are all fantastic, and your friendship has kept me going. Lastly, thank you to my family: Stephen, Lindsey, Chris and Ben for admirably tolerating and encouraging my interest in bees, and my parents for always believing in me. iv Contents List of Figures .............................................................................................................................. x List of Tables ............................................................................................................................. xiii Chapter 1. General Introduction ......................................................................................... 1 1.1 Introduction to Pollination and Pollinators ............................................................ 1 1.1.1 The role of pollinators ....................................................................................... 1 1.1.2 Pollinator species ................................................................................................ 1 1.2 Pollinator Declines ..................................................................................................... 2 1.2.1 Evidence of declining richness and abundance among pollinator species 2 1.2.2 Causes of pollinator declines ............................................................................ 3 1.2.3 Evidence of limitation of pollinator populations by food availability ....... 5 1.3 Nectar ........................................................................................................................... 6 1.3.1 Components of nectar........................................................................................ 6 1.3.2 Secretion and reabsorption of nectar ............................................................... 8 1.3.3 Nectar properties ................................................................................................ 9 1.3.4 Variability in nectar ......................................................................................... 11 1.4 Pollen ......................................................................................................................... 12 1.4.1 The pollen grain ............................................................................................... 12 1.4.2 Pollen as a food reward ................................................................................... 13 1.4.3 Comparison of pollens as nutritional resources .......................................... 14 1.5 Mass-Flowering Crops as Sources of Nutrition for Pollinators ......................... 15 1.5.1 Effects on pollinators of mass-flowering crop availability ......................... 15 1.5.2 Differences in floral rewards between crop cultivars ................................. 16 1.6 Short Rotation Coppice Willow ............................................................................. 17 1.6.1 Cultivation of willow ....................................................................................... 17 1.6.2 Floral biology of willow .................................................................................. 18 1.6.3 Nutritional value of willow for pollinators .................................................. 19 1.7 Oilseed Rape ............................................................................................................. 21 1.7.1 Cultivation of oilseed rape .............................................................................. 21 1.7.2 Floral biology of oilseed rape ......................................................................... 22 1.7.3 Nutritional value of oilseed rape for pollinators ......................................... 25 1.8 Objectives .................................................................................................................. 26 v Chapter 2. General Materials and Methods ..................................................................... 27 2.1 Introduction ............................................................................................................... 27 2.2 Collection and Analysis of Nectar .......................................................................... 27 2.2.1 Sample collection .............................................................................................. 27 2.2.2 Measurement of nectar volumes .................................................................... 29 2.2.3 Analysis of nectar sugars ................................................................................. 29 2.3 Collection and Analysis of Pollen .......................................................................... 31 2.3.1 Sample collection .............................................................................................. 31 2.3.2 Preparation of samples for analysis of free amino acids ............................. 31 2.3.3 Preparation of samples for analysis of protein-bound amino acids .......... 32 2.3.4 Analysis of pollen free- and protein-bound amino acid content ............... 33 2.4 Bumble Bee Colonies ................................................................................................ 35 2.4.1 Bumble bee lifecycle