Temperature effects on the development in the solitary bee Osmia bicornis (Hymenoptera, Megachilidae) titelblatt Development time, brain microstructure, and cognitive abilities Dissertation zur Erlangung des Doktorgrades der Naturwissenschaften (Dr. rer. nat.) der Fakultät für Biologie und vorklinische Medizin der Universität Regensburg vorgelegt von Sabine Radmacher aus Mechernich im Jahr 2011 Temperature effects on the development in the solitary bee Osmia bicornis (Hymenoptera, Megachilidae) Development time, brain microstructure, and cognitive abilities Dissertation zur Erlangung des Doktorgrades der Naturwissenschaften (Dr. rer. nat.) der Fakultät für Biologie und vorklinische Medizin der Universität Regensburg vorgelegt von Sabine Radmacher aus Mechernich im Jahr 2011 Das Promotionsgesuch wurde eingereicht am: 07.07.2011 Die Arbeit wurde angeleitet von: Prof. Dr. Erhard Strohm Unterschrift: „Oh glücklich! wer noch hoffen kann aus diesem Meer des Irrtums aufzutauchen. Was man nicht weiß das eben brauchte man, und was man weiß kann man nicht brauchen.“ Johann Wolfgang von Goethe, „Faust - Der Tragödie erster Teil“ TABLE OF CONTENTS TABLE OF CONTENTS LIST OF PUBLICATIONS ............................................................................................ 5 CHAPTER 1: GENERAL INTRODUCTION ............................................................. 6 1.1 Temperature – an important environmental factor for insects ........................... 6 1.2 Temperature and bee development .................................................................... 8 1.3 The study species: Osmia bicornis .................................................................. 11 1.4 Outline of this thesis ........................................................................................ 12 CHAPTER 2: EFFECTS OF CONSTANT AND FLUCTUATING TEMPERATURES ON THE DEVELOPMENT OF THE SOLITARY BEE OSMIA BICORNIS (HYMENOPTERA, MEGACHILIDAE) ................................. 15 2.1 Summary.......................................................................................................... 15 2.2 Introduction ..................................................................................................... 16 2.3 Materials and Methods .................................................................................... 17 2.3.1 Bees ..................................................................................................... 17 2.3.2 Provision weight ................................................................................. 18 2.3.3 Temperature treatments ...................................................................... 18 2.3.4 Observation cocoons ........................................................................... 19 2.3.5 Wintering ............................................................................................. 19 2.3.6 Data analyses ....................................................................................... 20 2.4 Results ............................................................................................................. 20 2.4.1 Prepupal weight................................................................................... 20 2.4.2 Mortality.............................................................................................. 22 2.4.3 Duration of development ................................................................... 22 2.5 Discussion........................................................................................................ 24 2.5.1 Prepupal weight................................................................................... 24 2.5.2 Mortality.............................................................................................. 25 2.5.3 Duration of development .................................................................... 26 2.6 Acknowledgements .......................................................................................... 27 CHAPTER 3: TEMPERATURE EFFECTS ON SOLITARY BEE DEVELOPMENT – DOES TEMPERATURE EXPERIENCED AS LARVA INFLUENCE THE DURATION OF THE PREPUPAL STAGE? ........................... 28 3.1 Summary.......................................................................................................... 28 3.2 Introduction ..................................................................................................... 29 1 TABLE OF CONTENTS 3.3 Materials and Methods .................................................................................... 30 3.3.1 Bees ..................................................................................................... 30 3.3.2 Temperature treatments ....................................................................... 31 3.3.3 Observation cocoons ........................................................................... 31 3.3.4 Data analyses ....................................................................................... 32 3.4 Results ............................................................................................................. 32 3.4.1 Development from egg to cocoon ....................................................... 32 3.4.2 Development inside the cocoon .......................................................... 33 3.4.3 Complete development ....................................................................... 34 3.5 Discussion........................................................................................................ 36 3.5.1 Development from egg to cocoon ....................................................... 36 3.5.2 Development inside the cocoon and complete development .............. 36 3.6 Acknowledgements .......................................................................................... 39 CHAPTER 4: EFFECTS OF TEMPERATURE DURING DEVELOPMENT ON THE SYNAPTIC ORGANIZATION IN THE BRAIN OF THE SOLITARY BEE OSMIA BICORNIS (HYMENOPTERA, MEGACHILIDAE) ................................. 40 4.1 Summary.......................................................................................................... 40 4.2 Introduction ..................................................................................................... 41 4.3 Materials and Methods .................................................................................... 43 4.3.1 Bees and temperature treatments ........................................................ 43 4.3.2 Brain preparation and immunocytochemistry ..................................... 43 4.3.3 Imaging, estimating neuropil sizes, and counting MG ....................... 44 4.3.4 Data analyses ....................................................................................... 45 4.4 Results ............................................................................................................. 46 4.4.1 Immunofluorescence labelling of neuropils ........................................ 46 4.4.2 Body size and calyx branch size ......................................................... 47 4.4.3 MG density and MG number .............................................................. 47 4.4.4 Comparison of relative temperature effects on honeybees and O. bicornis .................................................................................................... 49 4.5 Discussion........................................................................................................ 50 4.5.1 Labelling of neuropils ......................................................................... 50 4.5.2 Body size and neuropil size ................................................................ 51 4.5.3 MG density and MG number .............................................................. 51 4.5.4 Comparison of temperature effects on honeybees and O. bicornis .... 52 4.6 Acknowledgements .......................................................................................... 53 2 TABLE OF CONTENTS CHAPTER 5: DOES TEMPERATURE EXPERIENCED DURING METAMORPHOSIS AFFECT THE SYNAPTIC ORGANIZATION IN THE BRAIN OF A SOLITARY BEE? ................................................................................. 54 5.1 Summary.......................................................................................................... 54 5.2 Introduction ..................................................................................................... 55 5.3 Materials and Methods .................................................................................... 56 5.3.1 Bees and temperature treatments ........................................................ 56 5.3.2 Brain preparations and immunocytochemistry ................................... 58 5.3.3 Imaging, estimating neuropil sizes, and counting MG ....................... 58 5.3.4 Data analyses ....................................................................................... 60 5.4 Results ............................................................................................................. 61 5.4.1 Immunofluorescence labelling of neuropils ........................................ 61 5.4.2 Effects of T1 ........................................................................................ 61 5.4.3 Effects of T2 ........................................................................................ 62 5.4.4 Comparison of relative temperature effects ........................................ 65 5.5 Discussion.......................................................................................................