Ranaviruses are large dsDNA viruses that are considered emerging pathogens, and they are known to cause mortality events in amphibian and fish populations. This research utilizes experimental and genomic data to elucidate the mechanisms driving the evolution and spread of ranaviruses, with a focus on host switching within the genus. In Chapter 1, we utilize virus challenge assays to examine potential transfer of ranaviruses between cultured juvenile largemouth bass (M. salmoides) and bullfrog tadpoles (Rana catesbeiana). Additionally, a commonly used antiparasitic treatment containing malachite green and formalin (MGF) was utilized to suppress the immune system of largemouth bass to assess the susceptibility of immunocompromised fish to ranaviruses. The results indicate that tadpoles are not susceptible to Largemouth Bass Virus (LMBV), but that bass are susceptible to ranaviruses isolated from amphibians. Furthermore, immunocompromised fish were more susceptible to both LMBV and FV3 infections than immunocompetent fish. In Chapter 2, we used eight sequenced ranavirus genomes and two selection-detection methods (site-based and branch-based) to identify genes that exhibited signatures of positive selection, potentially due to the selective pressures at play during host switching. We found evidence of positive selection acting on four genes via the site-based method, three of which are newly-acquired genes unique to ranavirus genomes. Our results suggest that the group of newly acquired genes in the ranavirus genome may have undergone recent adaptive changes that have facilitated interspecies and interclass host switching. In Chapter 3, we annotated and analyzed the nearly complete genomic sequence of LMBV to determine its taxonomic classification. The available genomic content and phylogenetic evidence suggests that LMBV is more closely related to amphibian-like ranaviruses (ALRVs) than grouper ranaviruses, and this is further supported by greater genomic collinearity between LMBV and ALRVs. This data suggests that the classification of LMBV as a ranavirus is warranted. The results presented here will help to clarify the taxonomic relationships of ranaviruses, and will also be useful in developing management strategies to limit interspecific and intraspecific viral spread. The information garnered from this research will have far-reaching implications in studies of amphibian conservation, disease evolution, and virology. / text

How specialist bees distinguish their host plants from co-blooming non-hosts is not well understood, but it is thought that they may be physiologically limited in their ability to recognize the majority of visual and olfactory cues presented by flowering plants. Species in the genus Diadasia collect pollen from just one of five plant families: Cactaceae, Malvaceae, Onagraceae, Asteraceae, and Convolvulaceae. Assuming a common ancestor for all Diadasia specialized on just one plant, this indicates host-switches in the past, and an ability to recognize more than one flowering plant. I hypothesized that host plants of Diadasia share the same visual and olfactory cues, thus enabling past host-switching to novel plant families, and that co-blooming non-hosts are highly dissimilar in terms of scent and visual appearance. I determined the compounds associated with the scent of Malvaceae and Cactaceae host flowers of North American Diadasia. I also evaluated visual cues for these flowers by measuring their full spectrum of reflected wavelengths, as well as select morphometric characters. I determined whether host flowers, regardless of taxa, were more similar to each other than non-hosts that were co-blooming and attractive to other bee species. Finally, I performed electroantennographic and behavioral experiments to assess the relative importance of these cues (visual and olfactory) in natural settings. Diadasia host plants share some chemical and visual characteristics that may in part explain the radiation of this group onto these particular hosts. First, host plants share a suite of scent compounds that are among the least variable across species. Many of these elicited antennal responses and did not decrease visitation when applied to host flowers. In contrast, some compounds produced by non-host flowers are detectable to Diadasia, but Diadasia are repelled by them when they are applied to host flowers. Diadasia host flowers do not share a color profile in common, but there is more reflectance in the bee-uv range as compared to other regions of the light spectrum. Also, Diadasia host flowers have a more contrasting central area that is relatively larger than in non-hosts.

Le pou de tête et le pou de corps sont deux écotypes indiscernables occupant chacun une niche écologique différente. Le pou de corps représente une menace réelle pour l'Homme en raison de son rôle de vecteur dans la transmission de trois maladies graves pour l'Homme à savoir: le typhus épidémique, la fièvre des tranchées et la fièvre récurrente. Dans cette thèse, nous avons obtenu des résultats concrets dans chacune des thématiques abordées. En effet, nous avons (i) mis en place un outil moléculaire qui permet de différencier pour la première fois entre le pou de tête et le pou de corps qui a montré efficacité sur le terrain, (ii) mis en évidence l'existence d'un nouveau clade mitochondrial (Clade D) renfermant des poux de tête et des poux de corps susceptible de vectoriser Bartonella quintana et Yersinia pestis, (iii) retracé les migrations humaines à travers l'analyse de poux anciens provenant de différentes périodes et localisations, (iv) démontré pour la première fois que Pediculus mjobergi est génétiquement proche du pou humain et confirmé l'hypothèse qu'à l'origine Pediculus mjobergi était un pou humain qui a été transféré aux singes du Nouveau Monde par les premiers Hommes à avoir atteint le continent américain il y a des milliers d'années et (v) mis en place un outil de détection et de contrôle de la résistance moléculaire des poux à la perméthrine. Cet outil fut particulièrement utile dans l'étude clinique que nous avons menée pour déterminer si l'utilisation de sous-vêtements imprégnés d'insecticide offrait une protection efficace à long terme contre les poux de corps infestant les personnes sans-abri. / Head louse (Pediculus humanus capitis) and body louse (Pediculus humanus humanus) are two indistinguishable ecotypes each occupying an ecological niche: hair for head louse and clothing for the body louse. Body louse represents a real threat to humans because of its role as vector for the transmission of three deleterious diseases namely epidemic typhus, trench fever and relapsing fever.In this thesis, we obtained concrete results that have led to scientific publications. Indeed, we (i) implemented a molecular tool to differentiate for the first time between head and body louse, (ii) we highlighted the existence of a fourth mitochondrial clade (Clade D) comprising head and body lice that can vectorize Bartonella quintana and Yersinia pestis, (iii) we traced human migration through the analysis of ancient lice from different periods and different area, (iv) we demonstrated for the first time that Pediculus mjobergi is genetically close to human louse and confirmed the hypothesis that initially Pediculus mjobergi was a human louse has been transferred to New World monkeys by the first humans who have reached the American continent thousands of years ago and (v) we have implemented a tool for detecting and monitoring the molecular resistance to permethrin of body lice that parasite sheltered homeless persons in Marseille. This tool was particularly useful in the clinical study we conducted to determine whether the use of long-lasting insecticide-treated underwear provides effective long-term protection against body lice in homeless persons.

Terrapins are one of the most endangered vertebrate groups, with almost half of the more than 300 extant species threatened with extinction. This study was conducted to investigate parasite host-switching from the invasive American Red-eared slider, Trachemys scripta elegans, to the native Mediterranean pond terrapin, Mauremys leprosa in natural environments in southern France and Spain. The study also included a risk assessment approach that was developed to assess the viability of the native M. leprosa terrapins in a small river of southern France. The diversity of haemogregarine parasites within South African terrapins was also explored. The thesis is structured as follows: CHAPTER 1 gives a broad introduction to reptiles and chelonians as well as the importance of chelonians in ecosystems with emphasis to the threats that are driving terrapins to decline. The various terrapin species examined during this study as well as the parasite groups of interest (Monogenea and Apicomplexa) are also discussed in detail. CHAPTER 2 documents the extent of platyhelminth invasions from T. s. elegans to natural M. leprosa populations in northern Spain and southern France. From DNA barcoding analysis based on the sequencing of the Cytochrome c Oxidase I gene, the inferred Minimum Evolution tree and p-distance comparisons of closely related haplotypes revealed a greater polystome richness within M. leprosa than expected, suggesting that host switching may take place in natural environments. T. s. elegans would serve as a carrier for a variety of polystomes that usually infest American turtles in their home range. These are transmitted to M. leprosa throughout the south of France, also suggesting that turtle polystomes are not strictly host-specific. CHAPTER 3 investigates polystome egg production under changing environmental conditions. The experimental procedure that was conducted on M. leprosa showed that environmental temperature has an effect on the egg laying rhythm of its parasite, i.e., Neopolystoma sp., with a two day lag of egg production in response to environmental temperature change. Results suggest the adaptability of the parasite to the physiology of their chelonian hosts which are ecthothermic animals. They also show that eggs production may be attributable to the release of host factors like hormones that once secreted may act and stimulate parasite reproduction. CHAPTER 4 relates risk assessment for the viability of the native Mediterranean pond terrapin (M. leprosa) in a natural environment by using the Relative Risk Model (RRM) method, taking into consideration various threats and environmental conditions that may impact this species. CHAPTER 5 examines the diversity of South African terrapin haemogregarines (Protozoa: Apicomplexa: Haemogregarinidae) as well as their phylogenetic placement among haemogregarines based on molecular and morphological evidences. CHAPTER 6 summarizes all findings of the study and explores new ways of Research. / PhD (Zoology), North-West University, Potchefstroom Campus, 2014

Terrapins are one of the most endangered vertebrate groups, with almost half of the more than 300 extant species threatened with extinction. This study was conducted to investigate parasite host-switching from the invasive American Red-eared slider, Trachemys scripta elegans, to the native Mediterranean pond terrapin, Mauremys leprosa in natural environments in southern France and Spain. The study also included a risk assessment approach that was developed to assess the viability of the native M. leprosa terrapins in a small river of southern France. The diversity of haemogregarine parasites within South African terrapins was also explored. The thesis is structured as follows: CHAPTER 1 gives a broad introduction to reptiles and chelonians as well as the importance of chelonians in ecosystems with emphasis to the threats that are driving terrapins to decline. The various terrapin species examined during this study as well as the parasite groups of interest (Monogenea and Apicomplexa) are also discussed in detail. CHAPTER 2 documents the extent of platyhelminth invasions from T. s. elegans to natural M. leprosa populations in northern Spain and southern France. From DNA barcoding analysis based on the sequencing of the Cytochrome c Oxidase I gene, the inferred Minimum Evolution tree and p-distance comparisons of closely related haplotypes revealed a greater polystome richness within M. leprosa than expected, suggesting that host switching may take place in natural environments. T. s. elegans would serve as a carrier for a variety of polystomes that usually infest American turtles in their home range. These are transmitted to M. leprosa throughout the south of France, also suggesting that turtle polystomes are not strictly host-specific. CHAPTER 3 investigates polystome egg production under changing environmental conditions. The experimental procedure that was conducted on M. leprosa showed that environmental temperature has an effect on the egg laying rhythm of its parasite, i.e., Neopolystoma sp., with a two day lag of egg production in response to environmental temperature change. Results suggest the adaptability of the parasite to the physiology of their chelonian hosts which are ecthothermic animals. They also show that eggs production may be attributable to the release of host factors like hormones that once secreted may act and stimulate parasite reproduction. CHAPTER 4 relates risk assessment for the viability of the native Mediterranean pond terrapin (M. leprosa) in a natural environment by using the Relative Risk Model (RRM) method, taking into consideration various threats and environmental conditions that may impact this species. CHAPTER 5 examines the diversity of South African terrapin haemogregarines (Protozoa: Apicomplexa: Haemogregarinidae) as well as their phylogenetic placement among haemogregarines based on molecular and morphological evidences. CHAPTER 6 summarizes all findings of the study and explores new ways of Research. / PhD (Zoology), North-West University, Potchefstroom Campus, 2014

Globally disseminated bacterial pathogens frequently cause epidemics that are of major importance in public health. Of particular significance is the capacity for some of these bacteria to switch into a new environment leading to the emergence of pathogenic clones. Understanding the evolution and epidemiology of such pathogens is essential for designing rational ways for prevention, diagnosis and treatment of the diseases they cause. Whole-genome sequencing of multiple isolates facilitating comparative genomics and phylogenomic analyses provides high-resolution insights, which are revolutionizing our understanding of infectious diseases. In this thesis, a range of population genomic analyses are employed to study the molecular mechanisms and the evolutionary dynamics of bacterial pathogen niche adaptation, specifically between humans, animals and the environment. A large-scale population genomic approach was used to provide a global perspective of the host-switching events that have defined the evolution of Staphylococcus aureus in the context of its host-species. To investigate the genetic basis of host-adaptation, we performed genome-wide association analysis, revealing an array of accessory genes linked to S. aureus host-specificity. In addition, positive selection analysis identified biological pathways encoded in the core genome that are under diversifying selection in different host-species, suggesting a role in host-adaptation. These findings provide a high-resolution view of the evolutionary landscape of a model multi-host pathogen and its capacity to undergo changes in host ecology by genetic adaptation. To further explore S. aureus host-adaptive evolution, we examined the population dynamics of this pathogen after a simulated host-switch event. S. aureus strains of human origin were used to infect the mammary glands of sheep, and bacteria were passaged in multiple animals to simulate onward transmission events. Comparative genomics of passaged isolates allowed us to characterize the genetic changes acquired during the early stages of evolution in a novel host-species. Co-infection experiments using progenitor and passaged strains indicated that accumulated mutations contributed to enhanced fitness, indicating adaptation. Within-host population genomic analysis revealed the existence of population bottlenecks associated with transmission and establishment of infection in new hosts. Computational simulations of evolving genomes under regular bottlenecks supported that the fitness gain of beneficial mutations is high enough to overcome genetic drift and sweep through the population. Overall, these data provide new information relating to the critical early events associated with adaptation to novel host-species. Finally, population genomics was used to study the total diversity of Legionella longbeachae from patient and environmental sources and to investigate the epidemiology of a L. longbeachae outbreak in Scotland. We analysed the genomes of isolates from a cluster of legionellosis cases linked to commercial growing media in Scotland and of non-outbreak-associated strains from this and other countries. Extensive genetic diversity across the L. longbeachae species was identified, associated with intraspecies and interspecies gene flow, and a wide geographic distribution of closely related genotypes. Of note, a highly diverse pool of L. longbeachae genotypes within compost samples that precluded the genetic establishment of an infection source was observed. These data represent a view of the genomic diversity of this pathogen that will inform strategies for investigating future outbreaks. Overall, our findings demonstrate the application of population genomics to understand the molecular mechanisms and the evolutionary dynamics of bacterial adaptation to different ecological niches, and provide new insights relevant to other major bacterial pathogens with the capacity to spread between environments.

The fungus-growing ants have long provided a spectacular example of co-evolutionary integration between distantly related taxa. Their ecological success has been thought to depend largely on the evolutionary alignment of reproductive interests between ants and fungi, following vertical transmission and the ancient suppression of fungal sexuality. In my dissertation I explored the role of lateral cultivar switching on the evolution of the fungus-gardening ant mutualism. First, I provided the first evidence for sexual reproduction in the attine cultivars, together with evidence of extensive independent long-distance horizontal transmission of fungal genes. In fact, fungi have greater gene flow relative to their host ants, crossing the Gulf of Mexico between Latin America and Cuba, over which the ants cannot readily disperse. Second, for the special case of leaf-cutting ants, I show that the cultivar population was largely unstructured with respect to host ant species, and leaf-cutting ants interact largely with a single species of fungus. Finally, I examined the effect of post-glacial expansion on the population structure of the northern fungus-gardening ant Trachymyrmex septentrionalis and compared it with that of its two microbial mutualists: a community of lepiotaceous fungal cultivars and associated antibiotic-producing Pseudonocardia bacteria. This comparison allowed me to examine the effect of historical biogeographic forces, such as climate-driven range shifts, on the population structure of the ants and their microbial symbionts. While neither the cultivar nor the Pseudonocardia genetic structure was correlated with that of the ants, they were significantly, though weakly, correlated with each other. These results suggest that biogeographic forces may act differently on macro- and microscopic organisms, even in the extreme case where some microbial mutualists may be vertically transmitted from generation to generation and share the same joint ecological niche. Thus, binding forces that appear to enforce host fidelity are relatively weak and pairwise associations between cultivar lineages and ant species have little opportunity for evolutionary persistence. Taken together, my studies suggest that mechanisms other than long-term pairwise interactions between ants and fungi (so-called partner fidelity feedback) govern the evolution of the mutualism over evolutionary time. / text

Host-parasite associations are born by cospeciations or by host switches. Feather lice have traditionally been used as model parasites for studying these events and underlying ecological factors. By now tens of analyses have addressed comparisons of host and parasite phylogenies to study cospeciations and host switches in lice, however, these analyses are strongly biased towards the temperate zone. Tropical environment could provide new insight into the origin of host-parasite interactions, because it is ecologically unique. This work aims to supplement the knowledge of host-parasite associations in lice using coevolution analyses of two feather lice genera and their passerine hosts in tropical rainforest in Cameroon. It shows that lice in the tropics cospeciate rarely. To assess whether host switches are non-random and occur preferentially between hosts with specific traits, this work also analyses relations between parasite genetic distances and hosts' trait similarities. No effect of host morphology and spatial distribution was found. However, genetic distances of the lice strongly correlate with genetic distances of their hosts.

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.