Habitat Composition, Sexual Conflict & Life History Evolution in Coelopa frigida. Dominic Alexander Edward, MSc Thesis submitted to the University of Stirling for the degree of Doctor of Philosophy January 2008 Table of Contents Table of Contents i Abstract viii Chapter 1 Introduction 1 1.1 Sexual conflict 1 1.2 Empirical Examples of Sexual Conflict 3 1.3 C. frigida – a model for the study of sexual conflict 13 1.4 Chromosomal inversion system 15 1.5 Reproduction in C. frigida 18 1.5.1 Mating behaviour 18 1.5.2 Large male mating advantage 21 1.5.3 Male mate choice 23 1.6 Sexual selection in C. frigida 25 1.6.1 Good genes 25 1.6.2 Fisherian selection 27 1.6.3 Convenience polyandry 29 1.6.4 Multiple selection processes acting in C. frigida 31 1.7 The water striders – a related mating system 32 1.8 Interactions between C. frigida & C. pilipes 34 1.9 Coelopids and seaweed 36 1.9.1 Nutritional value of different algae 37 1.10 Aims of this thesis 40 Chapter 2 General materials & methods 42 2.1 General procedures 42 i 2.1.1 Introduction 42 2.1.2 Collection of wild coelopids 42 2.1.3 Collection of algae 43 2.1.4 Culturing coelopids 43 2.1.5 Storage and manipulation of coelopids 44 2.1.6 Determining body size 45 2.2 Mating trials 45 2.3 Determination of αβ chromosomal inversion karyotype 46 2.3.1 Introduction 46 2.3.2 Gel preparation 47 2.3.3 Homogenisation of flies 48 2.3.4 Electrophoresis 49 2.3.5 Staining 49 Chapter 3 Climate change and coelopids 51 3.1 Introduction 51 3.2 Methods 56 3.2.1 Experimental Procedure 56 3.2.2 Statistical analysis 57 3.3 Results 57 3.3.1 Mainland populations 57 3.3.2 Scottish island populations 59 3.3.3 Swedish populations 59 3.3.4 Norwegian populations 60 3.4 Discussion 62 3.4.1 Current coelopid distribution 62 3.4.2 Effects of temperature change on coelopids 63 ii 3.4.3 Effects of temperature change on seaweed 64 3.4.4 Physical effects of climate change 66 3.4.5 Coelopids and the wrack bed community 67 Chapter 4 Species of algae and male mating behaviour 70 4.1 Introduction 70 4.2 Methods 71 4.2.1 Preparation 71 4.2.2 Experimental procedure 72 4.2.3 Statistical analysis 72 4.3 Results 72 4.3.1 Male harassment 72 4.3.2Mating success 75 4.4 Discussion 77 4.4.1 Summary of results 77 4.4.2 Environmentally determined sexual conflict 78 4.4.3 Manipulation of mating behaviour 80 Chapter 5 A changing environment & male mating behaviour 82 5.1 Introduction 82 5.2 Methods 84 5.2.1 Preparation 84 5.2.2 Experimental procedure 84 5.2.3 Statistical analysis 85 5.3 Results 86 5.3.1 Sucrose treatments 86 5.3.2 Algal treatments 86 iii 5.4 Discussion 89 Chapter 6 Condition dependent sexual conflict and the culture of C. frigida 92 in different environments 6.1 Introduction 92 6.2 Objectives 95 6.3 Methods – Part I 97 6.3.1 Preparation 97 6.3.2 Mating trials 97 6.3.3 Statistical analyses 97 6.4 Results – Part I 98 6.4.1 Summary of results 98 6.4.2 Male harassment 98 6.4.3 Time preceding a mount 100 6.4.4 Struggle duration and copulation success 100 6.4.5 Male persistence traits 100 6.5 Discussion – Part I 110 6.5.1 Harassment by small males 110 6.5.2 Females cultured on Laminaria mounted sooner 111 6.5.3 Struggle duration 113 6.5.4 Sexual conflict in different environments 113 6.5.5 Condition dependent sexual conflict 114 6.6 Methods – Part II 117 6.6.1 Controlled density cultures 117 6.6.2 Statistical analysis of controlled density cultures 119 6.6.3 Mating and oviposition trials 120 6.6.4 Statistical analysis of mating trials 120 iv 6.6.5 Statistical analysis of oviposition trials 121 6.7 Results – Part II 121 6.7.1 Controlled density cultures 121 6.7.2 Mating trials 133 6.7.3 Oviposition trials 144 6.8 Discussion – Part II 146 6.8.1 Oviposition trials 146 6.8.2 Controlled density cultures 147 6.8.3 Mating trials 150 Chapter 7 Analysing multivariate selection in C. frigida 154 7.1 Introduction 154 7.2 Methods 156 7.3 Results 157 7.3.1 Dataset A 157 7.3.2 Dataset B 162 7.4 Discussion 169 Chapter 8 Longevity of C. frigida 172 8.1 Introduction 172 8.2 Methods 177 8.2.1 Experimental procedure 177 8.2.2 Statistical analysis 177 8.3 Results 178 8.3.1 Summary of results 178 8.3.2 Effect of algal exposure 178 8.3.3 Effect of sex 179 v 8.4 Discussion 182 Chapter 9 Stable isotopes and larval diets 186 9.1 Introduction 186 9.2 Methods 189 9.2.1 Collection of algae and coelopids 189 9.2.2 Laboratory cultures 189 9.2.3 Stable isotope analysis 190 9.2.4 Statistical analysis 191 9.3 Results 191 9.4 Discussion 195 Chapter 10 Amplified Fragment Length Polymorphism 200 10.1 Introduction 200 10.2 Methods 203 10.2.1 Preparation 203 10.2.2 DNA extraction 204 10.2.3 Restriction & ligation of adapters 205 10.2.4 Amplification of fragments 205 10.2.5 Fragment analysis 209 10.3 Results 209 10.4 Discussion 214 Chapter 11 General discussion 218 vi Acknowledgements 222 References 223 Appendix 1 - Chapter 3 264 Appendix 2 – Chapter 4 267 Appendix 3 – Chapter 5 273 Appendix 4 – Chapter 6 281 Appendix 5 – Chapter 7 312 Appendix 6 – Chapter 8 328 Appendix 7 – Chapter 9 333 Appendix 8 – Chapter 10 337 vii Abstract This thesis describes an investigation into the effects of habitat composition, principally the composition of algae in a wrack bed, on the life history of the seaweed fly, Coelopa frigida. The mating system of C. frigida is dominated by intense sexual conflict characterised by frequent harassment by males leading to a vigorous pre-mating struggle. This response leads to sexual selection for large male size and sexual dimorphism. The mating behaviour of C. frigida is affected by their environment, with exposure to brown algae inducing harassment in males and oviposition in females. Despite more than two decades of research into coelopid reproduction little is known about how habitat composition alters the patterns and processes of sexual conflict. Studies contained in this thesis consider environmental influences that both directly and indirectly influence sexual conflict. Direct effects of the environment are measured by conducting mating trials following culture of C. frigida on different species of algae and by exposing males to different species of algae. This work is accompanied by studies of larval development and adult survival on different algae. In recent years it has been observed that the distribution of European coelopids has undergone a northward range shift. An investigation into the current distribution of European coelopid species and a discussion of the effects of climate change that may have caused this change is included. Finally, the use of stable isotope analysis to determine the diet of wild coelopids and alternative statistical methods to analyse mating trials are described. viii Chapter 1 - Introduction 1.1 Sexual conflict The meeting of two unrelated individuals to share genetic information in the production of future generations is a natural phenomenon whose intricacies may too easily be taken for granted. Interactions between the sexes can provide a fascinating insight into the perpetual challenge of self preservation; giving purpose and meaning to a wide diversity of behaviours and structures seen in nature. Gone are the days of assumed cooperation between the sexes (Arnqvist & Rowe 2005). Sexual reproduction can instead be considered the acts of a single individual to maximise fitness that just happen to depend on another. As two individuals meet with independent interests, roles and resources to contribute to reproduction; mutual assistance is only one possible strategy that may satisfy individual needs. Sexual conflict results from a divergence in the evolutionary interests of the sexes, driven by the maximisation of individual, rather than group, fitness (Parker 1979). It is now widely understood that males and females are not naturally cooperative during reproduction and conflict arises over the relative investment of each individual in the production of progeny (Trivers 1972). There is often little cost to one individual if the other can be coerced into undertaking the bulk of responsibility and investment in shared offspring. A conflict of interests is often present even before contact is made as females frequently invest more resources in the production of gametes (Bateman 1948). Asymmetrical investment at this stage of reproduction can limit the reproductive capacity of females relative to males. However, anisogamy is by no means a pre- requisite for sexual conflict, which can occur at any stage of a reproductive encounter. 1 Conflicts of interest may arise in a range of traits related to courtship, mating, fertilisation and parental care (Chapman 2006). At a genetic level, sexual conflict may occur over the same or different loci in males and females. Intra-locus sexual conflict is defined as conflict over a trait encoded by the same gene in males and females, inter- locus conflict being conflict over traits encoded by different genes in each sex (Chapman 2006).