Scent is thought to be the most important mode of communication for many mammalian species, including otters. Spraint surveys, used widely to monitor otter populations, describe distribution only. Discrimination between identity parameters through chemical analysis of scent marks has been conducted for many species but the feasibility of using this to enhance population monitoring has not been fully tested. Scent presentation experiments were conducted on captive otters and provided some evidence that otters can determine the sex and reproductive status of the scent depositor. These experiments provide the first evidence of a potential role of both volatile and non-volatile compounds in otter communication, and revealed that otters learn where scent signals are likely to be presented. A series of experiments were conducted to optimise methods for the chemical analysis of otter scent. A combination of scent sample collection from otter carcasses and captive otters enabled the description of the chemical nature of otter scent. Scent was associated with age, sex and reproductive status, but not with diet. Scent was associated with individual identity and in females, progesterone concentrations, suggesting a role in reproductive behaviours. Spatial analysis at a national scale revealed differences in scent between genetically distinct subpopulations of otters. On a catchment scale there was no association between scent similarity and spatial proximity. Although scent differed significantly with individual identity, differences were not at sufficient resolution to allow discrimination between unknown samples collected in the wild. This thesis makes several major steps towards unravelling the complexity of otter scent communication and adds to the knowledge of otter biology and behaviour.

In sub-Saharan Africa, 90% of malaria cases are the result of transmission by the Anopheles gambiae species complex, causing 600,000 deaths annually. Increasingly, An. gambiae demonstrate behavioural and physiological resistance to control interventions and this, coupled with inadequate sampling methods, necessitates urgent development of new, efficient monitoring and control tools for malarial mosquitoes. The aim of this project was to examine the host-seeking behaviour of female An. gambiae to identify behavioural attributes that could be exploited in the design of novel trapping systems. To facilitate this, a wind tunnel arena with three-dimensional video-tracking was developed to quantify host-seeking flight of An. gambiae when presented with host-associated stimuli. In a constant flow of carbon dioxide and human-derived volatiles, mosquitoes were most active early in the night, suggesting a periodic responsiveness to olfactory stimuli, priming them to respond to potential hosts early in the night. Later spontaneous activity may increase the likelihood of encountering host odour plumes. Mosquitoes exhibited smooth and tortuous flight in up, down and crosswind directions in flows of clean and host odour-laden air, demonstrating a flexible suite of host-seeking behaviours. It is proposed that ‘dipping’ flight, consisting of high frequency vertical oscillations, may represent an alternative strategy to optomotor-guided anemotaxis in very low-light levels. When presented with black and clear targets in a flow of host odour-laden air, mosquitoes closely approached both targets more frequently than in a flow of clean air. Black targets were approached more frequently and collision avoidance was characterised by a rapid change to steep vertical flight. That mosquitoes avoided colliding with clear targets suggests involvement of an un-described sensory mechanism for detecting surfaces. Based on these findings, a prototype sticky trap incorporating a visual cue was tested in a malaria endemic region of Burkina Faso. The visually conspicuous trap caught more An. gambiae than a control trap, although both were equally efficient; additional design features could further optimise the visual trap. Overall, project results indicate that female An. gambiae exhibit a variety of integrated stimuli-response mechanisms that control navigation through the environment and towards potential hosts. Furthermore, they validate the approach of using quantified behaviours to improve the efficacy of monitoring tools.

No description available.

Genetic analysis of animals involved in captive breeding and reintroduction programmes can provide valuable information to aid in maintaining wild type genotypes and genetic variability. Hylobates moloch, also referred to as the silvery gibbon, is an Endangered primate species endemic to the Indonesian island of Java. As part of an overall conservation programme, a captive breeding and reintroduction programme is being organised. In order to aid both the management decisions within the breeding programme and success rates of re-introductions analyses at three genetic regions were carried out, with DNA extracted from non-invasively collated faecal samples. In order to assay if captively bred individuals were representative of their wild conspecifics, the population was split into two groups representative of wild born and captive born individuals. Genetic analyses at mitochondrial DNA hypervariable region-I (mtDNA HV-I), 15 microsatellite loci and the second exon of the major histocompatibility complex (MHC) DRB region, were carried out to ascertain genetic variability levels, levels of inbreeding, signs of selection and confirm the pedigree. Captive born individuals had markedly lower levels of variability at mtDNA HV-I, which was significant versus the wild group. The second neutral marker of microsatellites revealed no differentiation between wild and captive-born; moreover measures of standardised heterozygosity demonstrateda fairly high level of genomic variability overall. Pedigree analysis using the microsatellites produced information that differed from studbook entries. This was further supported by haplotypic data compiled from the MHC DRB exon 2 analysis. The MHC study revealed a total of 14 DRB alleles, 10 of which are from unknown lineages when compared to human and chimpanzees. As with microsatellites, no group differentiation between wild and captive has occurred but there are more rare alleles present within wild individuals. In conclusion, whilst genetic variation is both high and shows no deviation from wild-born to captive-born at neutral microsatellite loci, care should be taken to maintain rare mtDNA haplotypes and MHC DRB alleles in future generations.

With fewer than 5000 black rhinoceros remaining, ex situ populations play a vital role in the conservation of this species. To reinforce in-situ conservation efforts, captive populations must be self-sustaining, both demographically and genetically, to maximise future viability. The aim of this thesis was to determine the sustainability of the European captive population of eastern black rhinoceros, and investigate factors that may influence population performance in this species. Population viability analysis of demographic data from in situ managed black rhinoceros populations were used to establish the variability in population growth rates across reserves. Secondly, these data were used to calculate fecundity, mortality and population structure performance indicators, as a reference for how this species can perform under natural conditions. The same analyses were then applied to the European ex situ population, to determine whether the population is demographically and genetically self-sustaining, and identify areas for potential improvement. The European captive population of eastern black rhinoceros, although currently self-sustaining, is performing sub-optimally both with respect to their in situ counterparts, and to a proposed target of 5% growth per annum. Population performance is primarily limited by sub-optimal reproduction, both in terms of individuals producing fewer calves per annum, and due to a high degree of reproductive skew across the population, leading to a large proportion of individuals failing to produce offspring. A multi-institutional study was conducted on 90% of the European population, to investigate intrinsic differences in faecal reproductive hormone metabolites between breeding and non-breeding individuals. In females, irregular oestrous cyclicity was observed, with longer than average cycles observed more frequently in females that had never bred, and periods of acyclicity more common in females that had not bred for at least seven years. Non-proven females also had higher body condition scores, and were less likely to exhibit regular signs of oestrus. In males, non-breeding males had reduced faecal testosterone compared to males that had previously sired offspring. Extrinsic factors were also investigated, to determine whether differences in reproductive success could be attributed to aspects of the social or physical environment. However, no consistent relationships were observed between breeding and non-breeding males or females. Furthermore, adrenal activity and testosterone concentration were not correlated with environmental factors, indicating that extrinsic factors alone may not explain differences in reproductive success. However, within females, differences in hormone concentration were associated with irregular oestrous cyclicity. Long cycles were associated with increased glucocorticoid metabolite concentration, and oestradiol metabolites were lower during periods of acyclicity. The duration of the preceding luteal phase varied between cycle types, indicating that the occurrence of regular and irregular oestrous cyclicity may be influenced by hormone exposure during the preceding oestrous cycle. In summary, a number of intrinsic differences in reproductive hormones in both males and females have been identified, which may be related to differential reproductive success. A better understanding of the causes of these differences would be beneficial to maximise growth rates and overall population performance of this ex situ population.

Sperm competition, when sperm from more than one male compete to fertilise the same ova, has driven a diversity of adaptations. Increasingly, molecular techniques have been used to study the effect of post-copulatory sexual selection, including sperm competition, on proteins that are essential to reproduction. Genomic studies have revealed the rapid evolution of ejaculate proteins in polyandrous species. Additionally, there is evidence that gene expression can be altered plastically, in response to cues of sperm competition. Such studies are limited however, as the transcriptome does not always equal the proteome. Here, quantitative proteomics techniques are utilised to explore plasticity in reproduction, at a molecular level, in the house mouse (Mus musculus domesticus) (Chapters 2 – 4). In addition, adaptations to sperm competition are considered within mammalian testes and sperm proteins (Chapters 5 and 6). Contrary to predictions of sperm competition theory, within rodents, sperm production is suppressed in subordinate males. In addition, dominant males develop significantly larger seminal vesicles. Here, quantitative proteomic analysis reveals that the composition of the secretion within the major accessory sex gland in house mice differs according to social status. Subordinate males produce a more concentrated protein secretion, which contains a greater proportion of the protein SVS2. This protein is essential in copulatory plug formation, and increasing the proportion of SVS2 within their seminal fluid may allow subordinate males to produce copulatory plugs of an equivalent size to those produced by dominant. Within mammals, the oviduct is the site of fertilisation and can exert control over sperm storage and movement. As male house mice can plastically alter their ejaculate production and allocation according to local sperm competition risk, it is feasible that females may alter the environment within the oviduct in response to similar cues of multiple mating, to maintain control of paternity and reduce the risk of polyspermy. I found no evidence for plasticity in the oviduct proteins of female house mice following either a high or low level of interactions with male house mice. Indeed, the oviduct proteome was more similar within siblings, balanced across treatments, than within treatment groups. I did find evidence for plasticity in female behaviour, when interacting with a novel male, and in female ano-genital distance. Mothers in populations with high levels of sperm competition may benefit from producing more competitive male offspring. I analysed male offspring produced by female house mice that had encountered either a high or low level of male odour and interactions prior to pregnancy. Quantitative proteomic investigation revealed that ejaculate composition differed according to the level of male interaction their mother had encountered before pregnancy. In contrast, I found no evidence for variation in male mating behaviour, reproductive morphology or ejaculate size. This study reveals the potential for maternally driven, subtle alterations in ejaculate composition. Sperm competition has driven the evolution of testicular architecture. Comparative analyses reveal that relatively larger testes also produce sperm more efficiently. Improved spermatogenesis efficiency is primarily due to a faster rate of each seminiferous epithelial cycle, thereby increasing sperm production rate. This trait is linked to sperm competition, and so sperm competition is selecting for a greater sperm output, primarily by increasing the rate of production. Sexual conflict and post-copulatory sexual selection drive the rapid evolution of genes involved in reproduction. The reproductive proteomes of relatively closely related species may therefore be very different from each other. Comparative proteomic analysis of cauda epididymal samples from two groups of mammals, rodents and ungulates, reveals broad similarities in the sperm proteome. Closer analysis of proteins known to be involved in sperm – egg interactions suggests these proteins are very divergent, and exhibit a low level of sequence homology.

The reproduction of the two-spot ladybird, Adalia bipunctata, is inhibited by a sexually transmitted ectoparasitic mite Coccipolipus hippodamiae that sterilizes female hosts, and a range of heritable microbes that kill male hosts during embryogenesis for instance Spiroplasma. Historical sampling indicated these two parasites were present commonly in A. bipunctata in the south of Sweden but were absent or rare in northern populations. In this thesis, I first established that the distribution of the mite was still stable over time, with a boundary to mite presence at 61˚N, as found 10 years previously. The incidence of male-killing Spiroplasma, in contrast, showed a small northward shift in its presence. I then examined the causes of mite presence/absence, and concluded ladybirds from northern populations were competent to carry and transmit mite infection, but that host phenology prevented its persistence in natural populations. I then explored the hypothesis that mite presence might select for increased reluctance of females to mate by comparing mating behaviour between mite present/mite absent populations. I observed that whilst rejection behaviour was protective against mite transmission, it was not more commonly observed in ladybirds derived from mite present populations. Finally, I examined whether the presence of male-killing Spiroplasma might affect mite epidemiology, and whether the mite itself might impact on the dynamics of the male-killing Spiroplasma. I observed that Spiroplasma did not affect individual competence to acquire and transmit mite infection, and did not protect its host against mite induced sterility. However, Spiroplasma was observed to mildly prolong host longevity, which may enhance the capacity of the mite to pass from overwintered to new generation cohorts of its host. Further, population sex ratio biases induced by Spiroplasma were predicted to influence mite epidemiology, through altering mating rate and per contact mite transmission probability. I therefore conclude first that mite incidence can be explained through host phenological variation, and that sex ratio distorting symbionts may impact on the dynamics of the mite infection. Future studies should explore the causes of high prevalence Spiroplasma infection in A. bipunctata in southern Sweden, and the features that prevent the establishment of infection in the north of the country.

Wild animals are adapted to survive in different niches and therefore represent a great source for natural variation studies. Mapping of complex traits in model organisms has, for some time been constrained by low genetic variation of laboratory cultured strains. Although informative, this approach is fairly limiting. Recently, research in many model species has benefited from the creation of multi-parental crosses derived from wild-caught strains. Studying natural genetic variation using this approach allows for a better understanding of gene function as allelic interactions in divergent genetic backgrounds play important roles in determining complex traits. Such an approach was notably missing in Caenorhabditis elegans research. To remedy this, a new 4-parental recombination inbred line (RIL) panel that is representative of genotypically distinct groups of C. elegans isolates and distinct from the canonical N2 strain has been created. In this thesis I have used C. elegans, that has a short lifespan, high fecundity and wide array of genetic and genomic resources, to investigate variation in lifespan, its related traits and in the response to dietary restriction (DR). My specific aims were to further characterise a number of previously isolated lifespan QTLs, and to analyse lifespan and stress resistance in a new panel of 4-parent RILs. This work has discovered that the effect of DR on lifespan in C. elegans varies between genotypes and that such differences are seen in introgression lines (ILs), RILs and in wild isolates. A wide review of the literature on DR shows support for the view that genotype-specific effects on lifespan are widespread and that for some genotypes DR can be deleterious. I have also discovered that the newly created 4- parental panel of RILs contains significant, ecologically relevant, variation in lifespan and in stress resistance, that lifespan and stress resistance; are not correlated in these lines, and that this can be used to identify quantitative trait loci (QTLs) controlling this variation. Importantly, some of these QTLs cannot be explained by known lifespan regulating genes. Furthermore, the analyses revealed that the cold stress resistance in C. elegans is related to the control of translation, that the major QTLs detected in the RILs cannot be a consequence of genes known to be involved in cold stress resistance and that one may be a consequence of variation in eftu- 2 a part of the translation machinery. These results highlight the importance of exploring various genetic backgrounds in quantitative genetics and present a wider picture of the genetic interactions that are likely to be happening in the wild.

It has been widely acknowledged that a global change in energy production, from fossil fuels to renewable sources, is required in order to reduce carbon dioxide outputs and help mitigate anthropogenic climate change. The UK is recognised as having one of the largest practical marine energy resources in Europe, including ‘wet renewables’ energy sources; tidal-stream and wave energy. Scotland, as well as having some of the best marine energy resources, also holds internationally important numbers of breeding seabirds. Both wave energy and tidal stream devices have the potential to place a new anthropogenic pressure on already declining seabird populations. Wet renewables are predicted to impact seabird populations through collision, disturbance, habitat loss and changes to food availability. However, few devices have been deployed to enable monitoring of impacts and there have been few studies of the ecological implications and magnitude of any impacts to these populations. The aim of this thesis is to address key knowledge gaps relating to our understanding of seabird ecology in high-energy marine environments, specifically exploring spatial, temporal and habitat drivers for their use of these areas. This is to help expand the scientific basis used to assess the impacts of marine renewable energy devices on marine birds. This thesis also aims to increase our understanding of bird survey requirements for wet renewable energy schemes to help develop methods for environmental impact assessment. Chapters 2, 3 and 4 demonstrate that marine birds show differing spatial and temporal patterns of abundance and behaviour within high-energy marine environments. By using shore-based vantage point methods in three high-energy areas in the Northern Isles, I explore some variations in vantage point methods. Chapter 5 provides the most extensive and up-to-date review of diving and foraging behaviour for UK marine bird species. I identify knowledge gaps including the species and parameters for which there are a paucity of studies. I demonstrate a more robust approach to data collection in these high-energy tidal stream, which can shed insight on how they are being used by marine birds, while the diving and foraging behaviour synthesis provided many of the values that are required to undertake underwater collision risk modeling. It is hoped that developers can adopt the approaches identified here, which will improve the accuracy of their assessments and improve monitoring of any potential impacts.

Two cloned Eco RI fragments have been isolated from a Xenopus borealis genomic library in phage lambda, using an actin cDNA clone as a probe. Both fragments have been restriction-mapped and partially sequenced. As a result of this, one fragment, 2.6 kb in size, has been found to encode an amino acid sequence identical to the first 149 amino acids of the mammalian cardiac muscle actin, while the other, 3.6 kb in size, encodes a protein which is almost identical to the first 267 amino acids of the mammalian beta-cytoskeletal actin. The putative muscle actin gene contains introns at different positions to those of the cytoskeletal actin gene. These intron positions are identical to those of the analogous genes in mammals. These data suggest that vertebrate muscle and cytoskeletal actin genes became separate before the divergence of amphibians from the rest of the vertebrates. Other sequence studies performed include an examination of codon usage and sequence divergence in actin genes. 'Repetitive DNA' has been found in the 3.6 kb Eco Rl fragment and has been roughly localized. Primer extension has been used to show that the partial leader sequence of an oocyte mRNA species is identical to that found in the beta-like cytoskeletal actin clone. A large intron is present in the leader of this gene. Near the transcriptional start site, several small sequences have been found which exist in similar positions in the rat beta-cytoskeletal actin gene, and may possibly be involved in the control of gene expression. Another oocyte actin mRNA species encodes a protein similar to the N-terminal end of the mammalian gamma- cytoskeletal actin, and its leader sequence is surprisingly similar to that of the beta-like actin message. Primer extension has also been used to study the expression of actin genes in several laevis non-muscle tissues, as well as in JU borealis skeletal muscle.