Speciation and diversification of the beetle tribe Cratopini on the islands of the South West Indian Ocean area. James J.N. Kitson A thesis submitted for the degree of Doctor of Philosophy. School of Biology University of East Anglia, UK September 2012 © This copy of the thesis has been supplied on condition that anyone who consults it is understood to recognise that its copyright rests with the author and that use of any information derived there- from must be in accordance with current UK Copyright Law. In addition, any quotation or extract must include full attribution. Thesis abstract The aim of this thesis is to examine a spectacular radiation of phytophagous beetles (Curculionidae: Entiminae: Cratopini) that inhabit the islands of the South- Western Indian Ocean. As there are no previous molecular studies of these beetles, it was necessary to take a varied approach that employed both population genetic methods and phylogenetic methods to gain a broad understanding of the evolution of the group. Questions addressed range from aspects of the ecology of individual species to the relationships within islands amongst morphospecies and finally to the biogeography of the group as a whole. Throughout, where appropriate, reference is made to the current understanding of Indian Ocean geology and how this effects the interpretation of the patterns seen. The relationships amongst morphospecies are complex with many found to be paraphyletic within much larger monophyletic clades of multiple morphospecies. Coalescent model based species delimitation techniques are used to define the boundaries of putative species for future examination. The overall biogeography of the group is found to be highly complex with multiple colonisation events across islands and archipelagos. Flight is found to have been lost on multiple occasions and the taxonomy of the group is discussed. Finally many Cratopine lineages are found to display strong population structure despite being flighted, possible reasons for this are discussed. i Supervisor details Main supervisor: Dr Brent Emerson Island Ecology and Evolution Research Group, Instituto de Productos Naturales y Agrobiología (IPNA-CSIC), C/Astrofísico Francisco Sánchez 3, La Laguna, Tenerife (Canary Islands, 38206), Spain Secondary supervisor: Dr David S Richardson School of Biological Sciences, University of East Anglia, Norwich Research Park, Norwich, Norfolk NR4 7TJ United Kingdom ii List of Contents Thesis abstract ......................................................................................................................... i Supervisor details .................................................................................................................. ii List of Contents ...................................................................................................................... iii List of Tables and Figures ................................................................................................. vii Acknowledgements ............................................................................................................xiv Chapter 1: Introduction ....................................................................................................... 1 1.1 Evolutionary Biology and Islands ......................................................................... 2 1.2 The study system and thesis structure ............................................................... 3 1.3 The geology of the South-West Indian Ocean ................................................... 5 1.4 References .................................................................................................................. 10 Chapter 2: Molecular characterisation of a trophic shift within an island radiation of insect herbivores (Curculionidae: Entiminae: Cratopus). ............ 14 2.1 Abstract ....................................................................................................................... 15 2.2 Introduction .............................................................................................................. 15 2.3 Methods ....................................................................................................................... 18 2.3.1 Beetle sampling .................................................................................................................... 18 2.3.2 Plant sampling ...................................................................................................................... 19 2.3.3 DNA extraction, PCR amplification and sequencing ............................................ 19 2.3.4 Phylogenetic, network and population genetic analyses ................................... 21 2.3.5 Divergence time estimation ............................................................................................ 22 2.3.6 Species assignment of trnL sequences ........................................................................ 23 2.3.7 Statistical analyses.............................................................................................................. 24 2.4 Results ......................................................................................................................... 25 2.4.1 MtDNA and nuclear gene sequencing ......................................................................... 25 2.4.2 Phylogenetic analyses and haplotype networks..................................................... 26 2.4.3 Population genetic analyses and cryptic species assessment ........................... 30 2.4.4 Divergence times .................................................................................................................. 32 iii 2.4.5 Plant analyses ....................................................................................................................... 32 2.5 Discussion .................................................................................................................. 36 2.5.1 Geographic structuring of genetic variation within Cratopus murinus and Cratopus ovalis ................................................................................................................................ 36 2.5.2 Trophic ecology of Cratopus murinus and Cratopus ovalis from dry forest sites ....................................................................................................................................................... 38 2.5.3 Variation in Cratopus murinus diet across dry forest sites ............................... 39 2.5.4 Trophic ecology of coralline islet populations of Cratopus murinus ............. 39 2.6 Conclusions ................................................................................................................ 40 2.7 References .................................................................................................................. 41 Chapter 3: Baseline molecular data for poorly understood diversity: a case study of Mauritian Cratopine weevils. ......................................................................... 45 3.1 Abstract ....................................................................................................................... 46 3.2 Introduction .............................................................................................................. 46 3.3 Methods ....................................................................................................................... 49 3.3.1 Beetle sampling .................................................................................................................... 49 3.3.2 DNA extraction, PCR amplification and sequencing ............................................ 51 3.3.3 Phylogenetic analyses ........................................................................................................ 53 3.3.4 Divergence time estimation ............................................................................................ 53 3.3.5 GMYC analyses ...................................................................................................................... 53 3.3.6 Population structure .......................................................................................................... 54 3.3.7 Phylogenetic diversity ........................................................................................................ 54 3.4 Results ......................................................................................................................... 55 3.4.1 MtDNA sequencing .............................................................................................................. 55 3.4.2 Phylogenetic analyses ........................................................................................................ 56 3.4.3 Divergence times and GMYC analyses ........................................................................ 59 3.4.4 Population structure .......................................................................................................... 67 3.4.5 Phylogenetic diversity ........................................................................................................ 68 iv 3.5 Discussion .................................................................................................................. 73 3.5.1 The Phylogenetics of Mauritian Cratopine weevils ............................................... 73 3.5.2 GMYC analyses .....................................................................................................................