Thesis (PhD)--Stellenbosch University, 2013. / ENGLISH ABSTRACT: Historical vicariant processes due to glaciations, resulting from the large-scale environmental changes during the Pleistocene (0.012-2.6 million years ago, Mya), have had significant impacts on the geographic distribution of species, especially also in marine systems. The motivation for this study was to provide novel information that would enhance ongoing efforts to understand the patterns of biodiversity on the South African coast and to infer the abiotic processes that played a role in shaping the evolution of taxa confined to this region. The principal objective of this study was to explore the effect of Pleistocene climate changes on South Africa′s marine biodiversity using five intertidal gastropods (comprising four rocky shore species Turbo sarmaticus, Oxystele sinensis, Oxystele tigrina, Oxystele variegata, and one sandy shore species Bullia rhodostoma) as indicator species. Sequence data obtained from partial segments of the mitochondrial cytochrome oxidase c subunit 1 (COI), and the nuclear ribosomal DNA (encompassing part of 5.8S, second Internal Transcribed Spacer and part of 28S, hereinafter called ITS2; or comprising part of the first Internal Transcribed Spacer, 5.8S, second Internal Transcribed Spacer and part of 28S, hereinafter called ITS), were used as genetic markers to construct phylogeographic patterns and to investigate demographic histories of the taxa. Population structure was investigated using haplotype network analyses, pairwise ΦST statistics, analysis of molecular variance (AMOVA), isolation by distance analyses, Bayesian analysis of population structure (BAPS) and coalescent analysis of gene flow. Demographic history was analysed through Fu′s Fs tests, mismatch distributions, and Bayesian skyline plots. Demographic analyses suggest that all five intertidal gastropods studied experienced demographic expansions dating to the late Pleistocene. The sandy shore direct developer B. rhodostoma began expansion after the LGM (c. 15 kya) whereas for the four rocky shore broadcast spawners (T. sarmaticus, O. sinensis, O. tigrina, and O. variegata) the onset of expansion coincided with or preceded the LGM (c. 25, 60, 50, 40 kya, respectively). Consistent with recent range expansions and gene flow patterns, the population genetic structure in all species was characterised by shallow or a lack of population differentiation. Oxystele variegata was an exception as it showed a deep disjunction, of late Pleistocene origin, between individuals in the west coast Namaqua Bioregion and those in the south coast Agulhas Bioregion. These results provide strong evidence of the vital role that Pleistocene climatic changes and current regimes played in shaping the nature and distribution of biodiversity on the South African coast. In addition, gene flow in all species, except O. tigrina, was remarkably asymmetrical with the regions around Cape Infanta and Port Elizabeth acting as source populations. Considering the generally weak population genetic structure and gene flow patterns detected for most gastropod species studied here, it is recommended that T. sarmaticus, O. sinensis, O. tigrina and B. rhodostoma be managed as panmictic populations, and that the region encompassing Cape Infanta, and Port Elizabeth should be prioritised for conservation as it appears to harbour source populations. Oxystele variegata was the only species showing distinct population structure and in this instance, species specific conservation efforts should recognize this divergence by treating the two genetic assemblages as distinct management units. / AFRIKAANSE OPSOMMING: Historiese vikariante prosesse kan toegeskryf word aan glasiasie, en het tot gevolg gehad dat grootskaalse veranderinge in die omgewing plaasgevind het tydens die Pleistoseen (,012 - 2.6 miljoen jaar gelede, Mjg). Dit het 'n beduidende impak gehad op die geografiese verspreiding van spesies, veral ook in die mariene stelsels. Die motivering vir hierdie studie was om nuwe data te voorsien wat sal bydrae tot die voortgesette pogings om die patrone van biodiversiteit langs die Suid-Afrikaanse kus te verstaan. Dit sou ook help om die abiotiese prosesse af te lei wat 'n rol gespeel het in die evolusie van taksa wat in hierdie streek voorkom. Die hoofdoel van hierdie studie was om die effek van die Pleistoseen klimaatsveranderinge op Suid-Afrika se mariene biodiversiteit te bepaal deur gebruik te maak van vyf intergety slak spesies as indikatore (vier wat in rotsagtige gebiede voorkom: Turbo sarmaticus, Oxystele sinensis, Oxystele tigrina, Oxystele variegata en 'n sanderige strand spesies: Bullia rhodostoma). Volgorde data verkry vanaf gedeeltelike segmente van die mitochondriale sitochroom oksidase c subeenheid 1 (COI), en die kern ribosomale DNA (bestaande uit 'n deel van 5.8S, tweede interne getranskribeerde spasieërders en 'n deel van 28S), hierna genoem ITS2 is gebruik as genetiese merkers om filogeografiese patrone te dokumenteer en ook om die demografiese geskiedenis van die spesies te ondersoek. Bevolking struktuur is ondersoek deur gebruik te maak van haplotipe netwerk analise, paarsgewyse ΦST statistiek, analise van molekulêre variansie (AMOVA), isolasie deur afstand analise, Bayesiaanse analise van die bevolking struktuur (BAPS) en analise van gene vloei. Demografiese geskiedenis is ontleed deur Fu se Fs toetse, misparing verdelings, en Bayesiaanse luglyn kurwes. Demografiese ontleding dui daarop dat al vyf die intergety slakke wat ondersoek is demografiese uitbreidings ervaar het wat terugdateer tot die einde van die Pleistoseen. Die sanderige strand direkte ontwikkelaar, B. rhodostoma, het die uitbreiding begin na die LGM (c. 15 Kya), terwyl vir die vier rotsagtige kusbewoners wat eiers oor 'n uitgebreide gebiede versprei (T. sarmaticus, O. sinensis, O. tigrina, en O. variegata) het die aanvang van die bevolkings uitbreiding saamgeval met die laaste galsiasie of dit voorafgegaan (c. 25, 60, 50, 40 Kya, onderskeidelik). In ooreenstemming met die onlangse reeks bevolkings uitbreidings, is die bevolking genetiese struktuur in alle spesies gekenmerk deur weinig differensiasie. Oxystele variegata was 'n uitsondering en het 'n ontwrigting van laat Pleistoceen oorsprong getoon tussen individue langs die weskus Namaqua Biostreek en dié in die suid kus Agulhas biostreek. Hierdie resultate voorsien sterk bewyse van die belangrike rol wat die Pleistoseen klimaatsveranderinge gespeel het in die vorming en verspreiding van biodiversiteit langs die Suid-Afrikaanse kus. Daarbenewens, geen vloei in alle spesies, behalwe O. tigrina, was merkwaardig asimmetries. Kaap Infanta en Port Elizabeth verteenwoordig moontlik die bron bevolkings. Met inagneming van die geringe bevolking genetiese struktuur en geenvloei patrone wat waargeneem is vir die meeste slak spesies wat bestudeer is, word dit aanbeveel dat T. sarmaticus, O. sinensis, O. tigrina en B. rhodostoma bestuur word as 'n panmiktiese bevolking, en dat die streek wat Kaap Infanta en Port Elizabeth insluit geprioritiseer moet word vir bewaring. Oxystele variegata was die enigste spesie wat duidelike bevolking struktuur getoon het en in hierdie geval, moet spesie spesifieke bewaringspogings aangewend word.

The butterfly family Lycaenidae (including the Riodinidae) contains an estimated 30% of all butterfly species and exhibits a diverse array of life history strategies. The early stages of most lycaenids associate with ants to varying degrees, ranging from casual facultative coexistence through to obligate association where the long-term survival of the butterfly is dependent on the presence of its attendant ants. Attendant ants guard the butterflies against predators and parasites during their vulnerable period of larval growth and pupation. The caterpillars, in return, reward the ants by providing attractive secretions from specialized glands in their cuticle. The prevalence of caterpillar-ant associations in the species rich Lycaenidae is in contrast with other Lepidoptera, where ant association appears only as isolated cases in otherwise non ant-associated lineages. This has led to the proposal that ant association may have influenced lycaenid diversification or even enhanced the rates of speciation in the group. In contrast, facultative ant-associated butterflies exhibit high levels of host plant integrity, so it is reasonable to assume that host plants may have played a significant role in their diversification. Since the influence of ants (or plants) on diversification is independent of geographic speciation modes such as vicariance or peripheral isolates, there is an underlying inference of sympatric speciation. Certain prerequisites thought to be important for sympatric speciation, such as mating on the host plant (or in the presence of the appropriate ant) as well as ant dependent oviposition preferences are characteristic of many obligate myrmecophiles. Not surprisingly, it has been suggested that evidence for sympatric speciation is more likely to be found in the Insecta since this additional mode of diversification could account for the large numbers of insect species. This thesis tested the diversification processes in obligate and facultative ant associated lycaenids using comparative methodologies in hierarchical molecular phylogenetic analyses. First, several hypotheses relating to the influence of ants on diversification in obligately ant associated lycaenid butterflies were tested in a phylogeographic analysis of the Australian endemic Jalmenus evagoras. The phylogeographic analysis revealed that regional isolation of butterfly subpopulations coincident with locally adapted ant taxa could generate a phylogenetic pattern in which related lycaenids would be seen to associate with related or ecologically similar ants. Likewise, ecological shifts in habitat preferences by lycaenids could lead to co-diversification with habitat specialist ants, even though in both cases, the ants may play only an incidental role in the diversification process. A comparative methodology was then applied in a molecular phylogenetic analysis of the genus Jalmenus to test for a signal of diversification consistent with shifts in ant partners, and to infer the processes by which ants could influence speciation. Several other specific hypotheses relating to monophyly and taxonomy were also examined. Comparative analysis of the Jalmenus phylogeny found that attendant ant shifts coincided with high levels of sympatry among sister species. This pattern could be explained by sympatric speciation; however, data suggested it was more likely that ant shifts occurred during butterfly population expansions as a result of vegetation and climate changes in the Pleistocene. Fragmentation of populations associating with novel ants could promote rapid ecological and behavioural changes and this could result in reproductive isolation of conspecifics when in secondary contact. Diversification would then continue in sympatry. In contrast, secondary contact of populations associating with the same ant species would result in homogenisation of the two lycaenid lineages or the extinction of one. A phylogeographic analysis of the facultative myrmecophiles, Theclinesthes albocincta/T. hesperia, was then undertaken to infer the evolutionary processes (such as the effects of host plant shifts) that could result in extant demographics. Species-specific questions of taxonomy, relative population ages and dispersal routes in arid Australia were also addressed. Results from the analysis suggested the two taxa were conspecific and had diversified in the late Pleistocene as a consequence of isolation in refugia in and around the arid areas of mainland Australia. However, as was the case in the J. evagoras population analysis in which attendant ant shifts were not detected, host plant shifts were not detected in the population analysis of T. albocincta/hesperia. Host plant or attendant ant shifts manifest more frequently at the species level, thus it was necessary to test the influence of host plant shifts at this higher level. The comparative methodology was then applied to a molecular phylogenetic analysis of the facultative ant-associated section Theclinesthes (comprising Theclinesthes, Sahulana and Neolucia) to test for modes of diversification consistent with host plant shifts. The relative importance of other influences on diversification was also assessed. Akin to the Jalmenus analysis, the prediction that sister species ranges should broadly overlap when a shift in host plants had taken place was upheld in the comparative analysis. Species in the genera Jalmenus and Theclinesthes were found to have diversified in the Pleistocene so were subject to the same climatic oscillations that influenced patterns of vegetation expansion and contraction across much of Australia. Thus, the similarity and predictability of relationships in the comparative analyses based on biological data suggested that host plant shifts have influenced diversification in facultative myrmecophiles by inhibiting gene flow in secondary contact in similar fashion to that of attendant ant shifts identified in the Jalmenus phylogeny. Interpretation of data in these analyses suggested that allopatric diversification was the most common mode of speciation. Isolation was inferred to be the result of fragmentation following long distance dispersal across wide expanses of marginal habitat, or vicariance following the closing of biogeographical barriers. However, attendant-ant and host-plant shifts clearly played an important role in the diversification process, and in the maintenance of species integrity among lycaenid butterflies. Furthermore, exceptions to the predicted patterns of range overlap and ecological shifts provided clues to additional modes of diversification including shifts in habitat preferences and an unusual temporal shift following changes in specific host plant phenology resulting in allochronic diversification. Inferring modes of diversification using comparative methods based on range overlap and biological traits in a phylogenetic context is not new; however, the interpretation presented in this thesis is in contrast with contemporary methods. It is clear that the patterns of species range overlap and the ecological preferences of sister taxa are intimately related among lycaenid species that diversified during the Pleistocene. As a result, different influences on diversification can be highlighted in phylogenies when applying existing comparative methodologies but without necessarily drawing the same conclusions about modes of diversification. A more inclusive explanation for patterns of range overlap among sister taxa is detailed, a consequence of which is a method for estimating rates of extinction in a phylogeny where comprehensive distributional, biological and taxonomic data are available. These patterns and predictions may be applicable to a range of taxa, especially those that have diversified in the Pleistocene. Plans for future studies are outlined.