Surveys and vegetation analyses were used to determine the known distribution and the characteristics of the habitat utilised by Anolis sagrei in Taiwan. Sampled lizards were used for morphological comparisons, and to investigate the reproductive biology of this species in Taiwan. The results of this study indicate that the distribution of A. sagrei in Taiwan is extensive (≥237 ha) in Chiayi City and County (southwestern study site) and scattered (≥8 ha) in Hualien City and County (eastern study site). These lizards were mostly found in open sunny degraded man-made habitats. Although some variations were noted in the comparisons between the A. sagrei collected from the two study sites, it was concluded that the two populations likely have the same founder population. The reproductive biology study indicated that photoperiod and the associated temperatures determine the reproductive cycles in A. sagrei. It also demonstrated that reproduction in this species is energetically demanding. / Nature Conservation / MSc.(Nature Conservation)

Vertebrate genomes exhibit global methylation of cytosine residues where they occur in a cytosine-guanine dinucleotide (CpG) context and this epigenetic mark is generally thought to be repressive to transcription. Punctuating this pervasive DNA methylation landscape are short, contiguous regions of non-methylated DNA which are found at two thirds of mammalian gene promoters. These non-methylated regions exhibit CpG content close to expected levels as they escape the depletion of CpGs observed across the methylated fraction of the genome. The unique nucleotide properties of these CpG island (CGI) regions enable their identification by computational prediction in mammalian genomes. Owing to a lack of high-resolution genome-wide DNA methylation profiles in non-mammalian species, these CGI predictions have often been used as a proxy for non-methylated DNA in these organisms. In contrast to mammals, CGI predictions in cold-blooded vertebrates rarely coincide with gene promoters, leading to the belief that CGls are significantly divergent between vertebrate species, and that unique promoter-associated features may have been acquired during warmblooded vertebrate evolution. This thesis is primarily concerned with the location, establishment and biological function of non-methylated islands of DNA in vertebrate genomes. To experimentally determine genome-wide profiles of non-methylated DNA, a novel biochemical technique was established called biotinylated ZF-CxxC affinity purification (Bio-CAP), and development of this method is discussed in Chapter 3. Experimental analysis of non-methylated DNA profiles in this thesis initially addresses two main questions: (1) 'How does the non-methylated DNA landscape compare genome-wide for seven vertebrates considering distinct tissue types and developmental stages?' (2) 'How are vertebrate non-methylated regions of DNA defined and interpreted in the nuclear environment?' To address the first question, non-methylated DNA was profiled by Bio-CAP sequencing across the genomes of seven diverse vertebrate species, representing all major branch points of vertebrate evolution, and the results are discussed in Chapters 4 and S. Contrary to previously held dogma, experimentally determined nonmethylated islands of DNA (NMls) constitute an ancient epigenetic feature of vertebrate gene regulatory elements. However, despite having numerous high-resolution maps of vertebrate non-methylated DNA, the means by which NMls are identified and maintained in the nuclear environment remains poorly understood. To address the second question and identify features which determine the methylation state of DNA, exogenous DNA sequences were introduced into mouse embryonic stem (ES) c~.II~. Non-methylated DNA was profiled by Bio-CAP sequencing to investigate how different features, such as sequence-specific binding motifs, chromatin architecture and nucleotide composition of a given DNA sequence impact local DNA methylation patterns. Interestingly, the majority of exogenous promoters were appropriately non-methylated in mouse ES cells, germline and somatic cells suggesting that gene promoters have retained strong signals for the nonmethylated state across millions of years of evolution (discussed in Chapter 6). During mouse embryogenesis, genome-scale DNA demethylation and remethylation events occur to remodel the epigenetic landscape and loss of DNA methylation during this time leads to embryonic lethality. To investigate the biological function of non-methylated DNA, the third question addressed in this thesis is (3) 'What is the developmental importance of non-methylated islands of DNA during vertebrate embryogenesis?' To investigate this, members of the ZF-CxxC domain-containing family of chromatin modifiers were ablated in zebrafish embryos to perturb the chromatin landscape at NMls, and therefore interfere with their function during early development (Chapter 7). Early embryonic development and patterning was disrupted in knockdown embryos, suggesting that interpretation of non-methylated DNA and placement of chromatin modifications at NMls is essential for normal zebrafish embryogenesis. Together this work sheds light on the evolutionary origins of NMls, the mechanisms involved in the recognition and establishment of nonmethylated loci and provides an insight into the function of non-methylated DNA during early embryonic development.

The effect of a free-living stage in host-parasite coevolution: a skink mite phylogenetic study in New Zealand. During the last decade, phylogenetic trees have even been used to compare ecologically related taxa such as parasites and their hosts, and are used to determine their level of coevolution or reciprocal adaptation in time. Diverse coevolutionary events have been detected for this ecological association, where generally the parasite has been regarded as one that feeds exclusively on the host and is likely to cospeciate with it. A different coevolutionary pattern might occur when the parasite has a free-living stage in its life cycle, in which the parasite may have the opportunity to abandon its host and successfully colonise a new species (host-switching) making cospeciation less likely. Many New Zealand skinks are infested with a parasitic mite, Odontacarus sp. (Prostigmata: Leeuwenhoekiidae), which becomes free-living as an adult. The genetic variation of these mites found on four hosts was analyzed for host- parasite coevolutionary events. The hosts were the McCann’s skink and the common skink in coastal Birdling Flat, Canterbury, plus these species and the Grand and Otago skinks in Macraes Flat, Central Otago, South Island, New Zealand. The genetic variation of fast evolving nuclear Internal Transcribed Spacers 2 and mitochondrial Cytochrome c Oxidase I in Odontacarus mites found on these hosts was determined by PCR and DNA sequencing and phylogenetic trees were built using the computer programs PAUP*4 and MrBayes 3. The results show that mite haplotypes only had a significant geographical division and no host-related differences. In Birdling Flat, the COI haplotypes were represented in two groups that infested both regional hosts and had 5.7 % divergence. The same individual mites belonged to a single ITS 2 haplotype, thus indicating a historical geographical division between two populations that now interbreed successfully. The Macraes Flat mites were divided into two COI haplotypes with 2.4% divergence and internal nodes, which showed greater genetic variability than the Birdling Flat populations. The Macraes Flat mites formed two ITS 2 haplotypes with 6% divergence. This greater geographical structure of the Otago mites is probably due to the older age of the mainland area compared to the recently exposed coastal locality of Birdling Flat. The COI haplotypes from the two different regions had a mean distance of 15.5%, with an earlier divergence time than that known for the hosts. For both genes, the haplotypes from different regions had 100% bootstrap support and the parasite showed no host specificity. Mites of the different COI and ITS haplotypes were found on most of the host species that were sampled in Canterbury and Otago. The results of this study suggest that a free-living stage in a parasite’s life cycle can favour coevolutionary events such as inertia (failure to speciate) and host-switching, probably as a result of resource-tracking of the parasite. NB: Electronic files contained on CD to accompany print copy are not included with this version of the thesis.

ABSTRACT Social relationships, habitat utilisation and life history characteristics provide a framework which enables the survival of populations in fluctuating ecological conditions. An understanding of behavioural ecology is critical to the implementation of Natural Resource Management strategies if they are to succeed in their conservation efforts during the emergence of climate change. Egernia whitii from Wedge Island in the Spencer Gulf of South Australia were used as a model system to investigate the interaction of life history traits, scat piling behaviour and chemosensory communication in social lizards. Juveniles typically took ¡Ý 3 years to reach sexual maturity and the results of skeletochronological studies suggested longevity of ¡Ý 13 years. Combined with a mean litter size of 2.2, a pregnancy rate estimated at 75% of eligible females during short-term studies, and highly stable groups, this information suggests several life history features. Prolonged juvenile development and adult longevity may be prerequisite to the development of parental care. Parental care may, in turn, be the determining factor that facilitates the formation of small family groups. In E. whitii parental care takes the form of foetal and neonatal provisioning and tolerance of juveniles by small family or social groups within established resource areas. Presumably, resident juveniles also benefit from adult territorialism. Research on birds suggests that low adult mortality predisposes cooperative breeding or social grouping in birds, and life history traits and ecological factors appear to act together to facilitate cooperative systems. E. whitii practice scat piling both individually and in small groups. Social benefits arising from signalling could confer both cooperative and competitive benefits. Permanent territorial markers have the potential to benefit conspecifics, congenerics and other species. The high incidence of a skink species (E. whitii) refuging with a gecko species (N. milii) on Wedge Island provides an example of interspecific cooperation. The diurnal refuge of the nocturnal gecko is a useful transient shelter for the diurnal skink. Scat piling may release a species ¡®signature¡¯ for each group that allows mutual recognition. Scat piling also facilitates intraspecific scent marking by individual members, which has the potential to indicate relatedness, or social or sexual status within the group. The discovery of cloacal scent marking activity is new to the Egernia genus. E. Whitii differentiate between their own scats, and conspecific and congeneric scats. They scent mark at the site of conspecific scats, and males and females differ in their response to scent cues over time. Scat piling has the potential to make information concerning the social environment available to dispersing transient and potential immigrant conspecifics, enabling settlement choices to be made. This thesis explores some of the behavioural strategies employed by E. whitii to reduce risks to individuals within groups and between groups. Scents eliciting a range of behavioural responses relevant to the formation of adaptive social groupings, reproductive activity, and juvenile protection until maturity and dispersal are likely to be present in this species. Tests confirming chemosensory cues that differentiate sex, kin and age would be an interesting addition to current knowledge. The interaction of delayed maturity, parental care, sociality, chemosensory communication and scat piling highlights the sophistication of this species¡¯ behaviour. An alternative method for permanently marking lizards was developed. Persistence, reliability and individual discrimination were demonstrated using photographic identification and the method was shown to be reliable for broad-scale application by researchers. Naturally occurring toe loss in the field provided a context against which to examine this alternative identification method and revealed the need to further investigate the consequences of routine toe clipping, as this practice appears to diminish survivorship.

Surveys and vegetation analyses were used to determine the known distribution and the characteristics of the habitat utilised by Anolis sagrei in Taiwan. Sampled lizards were used for morphological comparisons, and to investigate the reproductive biology of this species in Taiwan. The results of this study indicate that the distribution of A. sagrei in Taiwan is extensive (≥237 ha) in Chiayi City and County (southwestern study site) and scattered (≥8 ha) in Hualien City and County (eastern study site). These lizards were mostly found in open sunny degraded man-made habitats. Although some variations were noted in the comparisons between the A. sagrei collected from the two study sites, it was concluded that the two populations likely have the same founder population. The reproductive biology study indicated that photoperiod and the associated temperatures determine the reproductive cycles in A. sagrei. It also demonstrated that reproduction in this species is energetically demanding. / Nature Conservation / MSc.(Nature Conservation)