Intra-specific variation in avian malaria : Linking infection dynamics to haplotypes

Avian Malaria infects thousands of species of birds across the Aves class. The most widespread (geographically and phylogenetically)morphological species is Plasmodium relictum, and of that, the mitochondrial lineage of SGS1 is the most common and invasive. It infects146 bird species, with large differences in infection outcome, from low intensity chronic disease to shorter, highly virulent infections. InPaper 1, I studied the gene expression of SGS1 in experimentally infected birds pre and post maximum parasitemia. The parasitemiavaried widely between birds and especially over the course of the infection. These differences were significantly linked to genes relatedto cell replication and cellular movement in high parasitemia infections and cellular metabolism in low parasitemia infections. We foundthat over time, variation in gene expression increased between samples, possibly illustrating individual responses of the parasites to theirhosts, and a desynchronisation in their lifecycles. Paper 2 explored the phylogeography of SGS1, and its related lineage GRW11, in thepalearctic region. Because the lineage system is defined by a highly conserved single mitochondrial gene, the nuclear polymorphic cellinvasion gene, merozoite surface protein 1 (msp1), was selected to study the genetic variation present in infected resident and migrant hostpopulations. We found extremely little variation, suggesting SGS1and GRW11 in Europe have an epidemic population structure, or thereis strong purifying selection pressure on the msp1 gene despite the wide host range. Paper 3 developed a genomic sequence capture methodusing 1035 probes designed for SGS1, and tested it on a range of SGS1, GRW11, and GRW4 samples. The probes effectively isolatedDNA from all three lineages, but sequencing success was low for samples with less than 1% parasitemia. We selected 25 genes to describethe higher-than-expected variation within SGS1 and with GRW11 and GRW4 samples. In Paper 4, two different host sources of SGS1infected blood were used to infect two groups of canaries. The groups differed in parasitemia and mortality, and from each group the threebirds with largest differences in infection outcome were selected for RNA sequencing to survey the underlying genomic variation. Thesource of the infection reliably separated the samples phylogenetically, with relatively less variation observed within the groups. Thissuggests that an infection is made up of a population of genetically diverse parasites. Paper 5 expanded on this idea by using the genomicsequence capture method from Paper 3 and refined bioinformatic methods from Paper 4 on some of the same samples from Paper 1.Samples collected at the same time points (8 and 20 days post infection) were sequenced. This allowed analyses of how the predominanthaplotypes change during an infection, and then link those haplotypes to the disease severity. We found that the least suppressed/mostvirulent haplotypes had genetic variants in genes related to cell invasion and immune evasion. The combined results of my thesis havefar-reaching implications that extend beyond the particular organism under investigation. The notion of genetic diversity within a singleinfections and the resulting parasite population dynamics offers exciting prospects for future research

Impact of the interactions between hosts vectors and pathogens on the transmission of avian malaria and flavivirus by mosquitoes.

Vector-borne pathogens cause important diseases such as malaria and are nowadays a major public health concern, because they cause human –and animal- fatalities worldwide and have a significant impact on local economies. Factors associated to global change, such as habitat alteration and introduction of invasive species, have largely contributed to the spread of potential insect vectors and the pathogens they are able to transmit, thus creating novel epidemiological scenarios. Therefore, it becomes essential to study the factors that modulate the transmission risk of these disease agents involving the interactions between vertebrate hosts (humans and other animals), pathogens and insect vectors in natural ecosystems. In this thesis, I used a multidisciplinary approach combining molecular tools, experimental bioassays and statistical analyses to assess the ecological and evolutionary factors that affect the transmission success of two mosquito-borne pathogens. In particular, I considered the interactions between insect vectors, vertebrate hosts, the avian malaria parasites and the flavivirus Zika virus. I focused on two major steps directly influencing the pathogen transmission success: i) the contact rate between mosquitoes and infected/susceptible vertebrate hosts and ii) the development of the pathogen in the mosquito and its consequences on the pathogen transmission risk. To do that, first I tested the potential causes underlying differences in the biting patterns of mosquito species. I exposed two bird species to two mosquito species to determine the role of mosquito species identity and effect of three host-related factors on host-vector contact rates (i.e. body mass, gender, and infection status by avian malaria). I found clear interspecific differences in the biting rates of mosquitoes, which were also influenced by variation in hosts’ traits, although these effects differed depending on the particular mosquito-host assemblage. Therefore, the biting patterns of mosquitoes are far from being generalizable. Secondly, I assessed the vector competence of different mosquito species for the transmission of Zika virus and avian malaria using mosquito saliva. I found that the ability of mosquito-borne pathogens to develop in mosquitoes differed between insect species, which may be the result of complex co-evolutionary processes. In addition, I assessed the consequences of parasite development in the mosquito vectors and their implications for the pathogen transmission risk. I found that host parasite load and parasite identity play affect the impact of parasites on mosquito longevity finally determining the transmission risk of the parasites. With this information mostly derived from studies under controlled conditions, I assessed the importance of environmental conditions affecting the host-parasite-vector assemblages in the wild. I found that habitat characteristics, which determine the existence and abundance of insect vectors, and host related factors (i.e. immune-competence) determines the prevalence of avian malaria parasites in insular ecosystems. Altogether, in this thesis I identified key factors affecting the transmission success of vector-borne pathogen affecting humans or wildlife allowing a better understanding the complex transmission dynamics of vector-borne pathogens.Los patógenos transmitidos por vectores que causan enfermedades importantes como la malaria son a día de hoy un importante problema de salud pública, ya que causan numerosas muertes humanas y animales en todo el mundo teniendo un gran impacto en la economía local. Factores asociados al cambio global, así como la alteración del hábitat y la introducción de especies invasoras, han contribuido en gran medida a la expansión de posibles insectos vectores y de patógenos que pueden transmitir, creándose nuevos escenarios epidemiológicos. Por lo tanto, es esencial estudiar aquellos factores que influyen en el riesgo de transmisión de estos patógenos considerando las interacciones entre los huéspedes vertebrados (humanos y otros animales), los patógenos insectos vectores en los ecosistemas naturales. En esta tesis, mediante un enfoque multidisciplinario que combina herramientas moleculares, bioensayos experimentales y análisis estadísticos, se evalúan los factores ecológicos y evolutivos que afectan al éxito de transmisión de dos patógenos transmitidos por mosquitos. En particular, se consideraron las interacciones entre los parásitos de la malaria aviar y el virus (flavivirus) del Zika.y sus insectos vectores y hospedadores vertebrados, En concreto, me centro en dos procesos principales que influyen directamente en el éxito de la transmisión de patógenos: i) la tasa de contacto entre mosquitos y hospedadores vertebrados infectados/susceptibles y ii) el desarrollo del patógeno en el mosquito y sus consecuencias sobre el riesgo de transmisión de estos. Para ello, primero estudié las posibles causas que subyacen a las diferencias en los patrones de alimentación de las especies de mosquitos. Se expusieron dos especies de pájaros a dos especies de mosquitos para determinar el papel de la especie de mosquito y el efecto de tres factores relacionados con el hospedador vertebrado sobre las tasas de contacto entre el hospedador y el vector (masa corporal, género e infección por parásito de la malaria aviar). Encontré claras diferencias inter-específicas en las tasas de picadura de los mosquitos, que también se vieron influenciadas por la variación en los rasgos de los hospedadores, aunque estos efectos difirieron dependiendo de la asociación mosquito hospedador. Por lo tanto, los patrones de alimentación de los mosquitos parecen estar lejos de ser generalizables. En segundo lugar, estudié la competencia vectorial de diferentes especies de mosquitos para la transmisión del virus del Zika y de parásitos de la malaria aviar utilizando la saliva del mosquito. Descubrí que la capacidad de desarrollo de los patógenos difería entre las especies de mosquitos, pudiendo esto ser el resultado de complejos procesos coevolutivos. Además, evalué las consecuencias que tiene el desarrollo de los parásitos en los mosquitos en el riesgo de transmisión de estos. También encontré que tanto la carga parasitaria del hospedador vertebrado como la identidad del parásito afectan a la longevidad de los mosquitos y consecuentemente determina el riesgo de transmisión de los parásitos. Con esta información, derivada principalmente de estudios en laboratorio, con condiciones controladas, evalué como afectan las condiciones ambientales a las asociaciones entre hospedador-parásito-vector en la naturaleza. Descubrí que las características del hábitat, las cuales van a determinan la existencia y abundancia de insectos vectores, así como los factores relacionados con el huésped (inmunocompetencia), determinan la prevalencia de parásitos de la malaria aviar en los ecosistemas insulares.Premio Extraordinario de Doctorado U

Ecologie évolutive de la malaria aviaire (effets des caractéristiques de l'hôte et de l'environnement)

L étude des interactions hôtes-parasites est devenue un thème de recherche incontournable pour les sciences de l évolution. Cette coévolution complexe dépend de nombreux compromis évolutifs et peut être grandement influencée par les facteurs environnementaux. Nous nous proposons ici d étudier les interactions hôtes-parasites à plusieurs échelles, à travers des approches expérimentales et des études en populations naturelles, en étudiant les parasites de la malaria aviaire. Dans un premier temps, nous nous sommes intéressés à l influence des caractéristiques de l hôte et notamment au système immunitaire. Le système immunitaire est bénéfique pour l hôte dans sa lutte contre le parasite, mais peut également engendrer des coûts immunopathologiques. Des traits d histoire de vie, comme l âge ou le statut social peuvent modifier la parasitémie au sein des hôtes, sans toutefois avoir d effet sur la prévalence. Dans un second temps, l effet de certains facteurs environnementaux a été évalué au sein des interactions hôtes-parasites. La température et la contamination en métaux lourds ont un effet sur la prévalence dans les populations, mais n affectent pas la parasitémie. Au cours de cette thèse, nous avons également montré l influence directe des parasites sanguins sur la structure génétique des populations hôtes, notamment au niveau des gènes du CMH.Host-parasite interactions are one of the main topics in evolutionary sciences. This complex coevolution depends on several trade-offs and can be influenced by environmental factors. Here, we propose to study host-parasite interactions with a multi-level approach, using experimental and natural population studies, focusing on avian malaria parasites. First, we studied the effect of host characteristics, and more precisely the immune system. The immune system confers benefits in terms of protection against the parasite, but can also generated immunopathological costs. Life history traits, like age or social status, appear to modify parasitemia but not prevalence. In a second part, we evaluated the effect of environmental factors on host-parasite interactions. We found that temperature and heavy metal contamination had an effect on population prevalence, but not on host parasitemia. We also showed the direct parasite influence on host population genetic structure, and more precisely on MHC genes.DIJON-BU Doc.électronique (212319901) / SudocSudocFranceF

Avian malaria parasites and their vectors

Parazity způsobující ptačí malárii řadíme mezi Haemosporida, která představují monofyletickou skupinu dixenních protist patřících do kmene Apicomplexa. Jejich nepohlavní množení probíhá v obratlovčím mezihostiteli a ke vzniku gamet a sporogonii dochází v krevsajícím dvoukřídlém hmyzu, který je tudíž konečným hostitelem těchto parazitů. U ptáku jako mezihostitelů se setkáváme s následujícími třemi rody těchto parazitů: ​Plasmodium​, ​Haemoproteus ​a ​Leucocytozoon​. V disertační práci jsem se soustředil na Haemosporida volně žijících ptáků a na způsob jejich přenosu hmyzími vektory v přírodních populacích, což byla prozatím opomíjená oblast výzkumu zaměřeného na původce ptačí malárii. Výsledky byly získány jak tradičními metodami (vyšetření infekcí pomocí mikroskopování krevních roztěrů), tak především molekulárně biologickými metodami (detekcí parazitů pomocí nested PCR) založenými na práci s unikátními haplotypy jednotlivých linií haemosporid. Cílem studie bylo určit okruh možných přenašečů ptačích haemosporid na území České republiky s přihlédnutím ke specificitě ptačích haemosporid v rámci těchto vektorů a popsat diverzitu ptačích haemosporid v populacích jejich ptačích mezihostitelů. K tomu jsme si vybrali čtyři různé druhy ptáků, patřících do čtyř řádů (pěvci - Passeriformes, sovy -...Parasites causing avian malaria belong to the group Haemosporida, which represents a monophyletic group of dixenic protists within Apicomplexa. Their asexual reproduction takes place in a vertebrate intermediate host, and the formation of gametes and sporogony occur in blood-sucking dipteran insects, which are the definitive hosts of these parasites. Three main genera (​Plasmodium​, Haemoproteus ​and ​Leucocytozoon​) are found mostly in their avian hosts. We focused on the Haemosporida of wild birds and their transmission by insect vectors in natural populations, which had previously been a neglected area. Our results were obtained both by traditional methods (investigation of infections by microscopy of blood smears) and mainly by molecular methods (e.g. nested PCR) centered around work with unique haplotypes of the haemosporid lineages. The aim of our work was to determine the range of possible insect vectors of avian haemosporidians in the territory of the Czech Republic, taking into account the specificity of the parasites within these vectors, and to describe the diversity of haemosporidians in the populations of their bird intermediate hosts. We chose four different species of birds from four orders (Passeriformes, Strigiformes, Accipitriformes, and Galliformes). As potential vectors of avian...Katedra zoologieDepartment of ZoologyPřírodovědecká fakultaFaculty of Scienc

Understanding avian Plasmodium distribution: the role of vector and host

Malaria parasites have a complex life cycle involving sexual reproduction in the mosquito vector and asexual proliferation in the vertebrate host. Mosquito vectors are therefore the definitive host of the malaria parasite. The literature on avian malaria parasites remains biased towards bird-parasite associations, and avian malaria vectors are not well studied in this system. My dissertation fills a gap in the current body of avian malaria research by using molecular techniques to document 1) patterns in phylogeographic structure of avian Plasmodium across geographic regions; 2) absence of vector specificity in two common mosquito species in Ithaca, New York; and 3) evidence that local vectors amplify a local avian Plasmodium lineage. In my first chapter, I review our current understanding about the associations between avian malaria vectors and avian Plasmodium. I synthesize this with literature on human malaria-mosquito interactions and mosquito feeding preferences to argue that it is crucial to study vector ecology to understand host-parasite dynamics. Variation in mosquito ecology could help explain patterns observed in avian malaria parasite infection. In chapter 2, I document patterns of phylogeographic structure in Plasmodium parasites sampled across the range of a single bird species. I demonstrate that geographic patterns in parasite lineage distribution are not solely attributable to differences in the parasites found in different bird species. In chapter 3, I describe the avian Plasmodium lineages found in two abundant, local ornithophilic mosquito species and show that parasites sampled from mosquitoes are represented by many diverse cytochrome b haplotypes. The most common of these haplotypes are shared between mosquito species, and overlap only slightly with those previously isolated directly from numerous bird species. In chapter 4, I report results from a feeding experiment using laboratory-reared mosquitoes and wild-caught birds naturally infected with Plasmodium. I examine the variation within Cx. pipiens in its ability to be invaded by the parasite by conducting PCR on individual mosquitoes following incubation with sufficient time to allow the complete digestion of the blood meal and development of sporozoites in the salivary glands

Clocks in the wild: biological rhythms of great tits and the environment

Biological clocks play a fundamental role in the physiological and behavioural processes of organisms. Internal timekeepers evolved to anticipate environmental changes, the most important of these being the geophysical light-dark cycle, to coordinate external changes with the timing of internal processes. Research into functions of the biological clock during captive studies has provided valuable insight into mechanisms by which clocks function, and how small environmental changes can affect the clock and its outputs. However, biological clocks have so far been understudied in ecology. In this thesis, this gap in knowledge was addressed by placing studies of chronobiology into the context of the natural environment. A model species in avian ecology, the great tit (Parus major) was used to investigate biological rhythms in the wild at three levels; behaviour, transcripts and life histories. This thesis investigated how features of the natural environment shapes rhythms of behaviour and physiology in a wild animal, using experimental and observational approaches. Differences in timing of individual rhythms, or chronotype, may provide wild animals with different consequences for fitness. In this thesis, individual behavioural rhythms of incubating great tits were quantified for birds in city and forest environments. There were strong effects of both the number of days to hatching and site on timing of incubation activities, where city birds rose earlier, and stayed out later, than forest birds. Maternal chronotype was then linked to fitness traits. City birds face a number of new challenges in the urban habitat. The impacts of one feature of the urban habitat, artificial light at night, was tested using a forest nest box system. Nestling great tits were experimentally exposed to low-level artificial light at night, and aspects of condition and clock and immune gene transcripts were compared for nestlings under light at night and dark-night control. Nestlings under light at night treatment weighed less than control birds, and suppressive effects of light at night treatment were found for genes involved in the core pathways of the circadian clock and immune system. Time of day differences were also observed in transcript levels of genes. Parasitic infections can cause consequences for fitness and reproductive success of wild birds. In this study, effects of infection with avian malaria parasites on nestling condition and immune system were investigated, at city and forest sites. The prevalence of Leucocytozoon parasites was higher at forest sites than city sites and increased with the season. Infection had no suppressive effects on immune genes of nestlings, and no negative effects on condition were found. In mammals, malaria is otherwise known as the “circadian disease” due to rhythmic development of parasites during their life cycle. In this study, host-parasite interactions with avian malaria parasites were investigated in the context of biological rhythms in wild great tits. Transcript levels of nestlings were determined by field sampling across a temporal profile and linked to infections with Leucocytozoon parasites. Leucocytozoon infection reduced overall transcript levels for circadian clock and immune gene targets, but did not alter the timing of expression. This study ultimately demonstrated the importance of biological clocks for the ecology of great tits and provided important advances for studies of clocks in the wild

Molecular data reveal a cryptic species within the Culex pipiens mosquito complex

This is the peer reviewed version of the following article: Dumas, E., Atyame, C. M., Malcolm, C. A., Le Goff, G., Unal, S., Makoundou, P., Pasteur, N., Weill, M. and Duron, O. (2016), Molecular data reveal a cryptic species within the Culex pipiens mosquito complex. Insect Mol Biol, 25: 800–809,which has been published in final form at doi:10.1111/imb.12264. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Self-Archiving. © 2016 The Royal Entomological Society.The Culex pipiens mosquito complex is a group of evolutionarily closely related species including the common house mosquito, Cx. pipiens, and the southern house mosquito, Cx. quinquefasciatus, which both are infected by the cytoplasmically inherited Wolbachia symbiont. A Wolbachia-uninfected population of Cx. pipiens was however described in South Africa and was recently proposed to represent a cryptic species where Wolbachia spread has been prevented by reproductive isolation. In this study, we reconsider the existence of this novel species by undertaking an extensive screening for the presence of Wolbachia-uninfected Cx. pipiens specimens and by characterizing their genetic relatedness with known members of the complex. We first reported on the presence of Wolbachia-uninfected specimens in several Cx. pipiens breeding sites in Europe and North Africa. Using a multi-locus typing scheme, we next confirm that these uninfected specimens unambiguously belong to the Cx. pipiens complex. While, uninfected specimens shared ancestral nuclear DNA polymorphism with infected Cx. pipiens specimens, they also harbor novel mitochondrial haplotypes which are closely related, but different, to those found in all other Cx. pipiens complex members. Overall, these results corroborate the presence of a cryptic species within the Cx. pipiens complex where ancestral levels of mitochondrial diversity have been maintained. We further evidence a geographic distribution far wider than previously suspected, ranging from the North of Europe to the South of Africa.Peer reviewe

Urbanisation and avian fitness: an investigation of avian malaria prevalence and feather corticosterone level of blue tit Cyanistes caeruleus nestlings across two breeding seasons

Urbanised landscapes vary significantly from natural habitats because of their different and unique ecological features. These features can affect the ability of both animals and plants to occupy urban habitats. Ecological studies on avian species have often reported a reduction in several breeding parameters of urban populations compared to their rural counterparts. However, how the urban environment can influence the breeding success of these species remains largely unknown. One hypothesis is that urban-specific factors alter key physiological traits modulated by stress hormone levels in birds. Indeed, long-term exposure to high levels of stress can lead to sustained elevation of basal glucocorticoid levels and consequent detrimental effects, such as impaired immunity, inhibited growth and reduced survival. For instance, alterations to the microclimate, pollution, and limitations of food resources may act as strong stressors, resulting in an increased likelihood of parasite infection and related fitness costs, affecting the capacity of hosts to occupy urban areas. Environmental conditions can fluctuate from year to year, affecting birds directly or indirectly by limiting food availability, especially for carnivorous birds of which the main food (insects, e.g. caterpillars) is highly dependent on weather conditions. As the urban conditions are already harsh, fluctuations in the other environmental factors, like weather conditions, have the potential to severely affect urban birds. Conversely, the more favourable conditions in rural habitats may allow the birds to buffer against other potentially negative environmental factors. Therefore, in order to understand the impact of urbanization on bird stress and fitness across fluctuating environmental conditions, the first aim of my thesis was to explore and compare the fitness of an urban and a rural population of blue tit Cyanistes caeruleus across two breeding seasons (2016 and 2017) in relation to the stress levels they experience. Previous studies have suggested the use of corticosterone (CORT, the main avian glucocorticoid) levels as a biomarker of stress experienced by animals. In birds, long term or chronic exposure to stressors can be measured from feather corticosterone (fCORT). This non-invasive method can measure the level of CORT that has been metabolised and deposited in feathers during feather growth. Thus, I compared two populations of blue tits in relation to the level of stress experienced by nestlings, throughout their first thirteen days of life, that may influence their fitness in urban and rural habitats. CORT levels in nestlings can also be directly influenced by parents before oviposition (e.g. by maternal deposition of CORT in yolk). Therefore, I also tested the role of origin of the bird on their fCORT levels, experimentally, to assess whether the pre-oviposition environment has a role in nestling’s fitness (Chapter 2). Then, as parasite infections can play a major role in fitness differences between urban and rural birds, I decided to measure the prevalence of avian malaria (haemosporidian parasites of the genera Plasmodium, Haemoproteus and Leucocytozoon), which are widespread parasites in birds, infecting blood cells. Specifically, I compared avian malaria prevalence in nestlings from the two populations across the two breeding seasons, as well as tested the role of parental origin (based on a cross-fostering experiment) on their susceptibility to infection (Chapter 4). However, to detect haemosporidian parasite acute infection and identify the parasite genera in the studied blue tit populations, I developed a new molecular method, as current tools did not allow these investigations (Chapter 3). Using this new approach, I explored my next aim, where I tested the effect of Leucocytozoon infection prevalence on fitness-related traits (body weight and survival) of nestling blue tits from the two populations during the two breeding seasons. I also tested the potential synergistic impact of infection and urban-related stress on blue tit fitness by examining the relationship between the two factors – fCORT level and Leucocytozoon infection prevalence (Chapter 5). I accomplished these aims using both experimental and correlational approaches, the former involving a cross-fostering experiment and a vector-manipulation experiment. In 2016, I cross-fostered some clutches between and within sites to test for any effects that may be derived from inherited or maternal traits from parents to their offspring. Additionally, as both populations showed high malaria prevalence in 2016, in the 2017 breeding season, I conducted a vector-repellent experiment to experimentally reduce infections in nestlings; this was done with the goal of better understanding the impact of parasites on bird’s fitness. The key findings of my thesis are as follows: First, monitoring the two populations of blue tit over two breeding seasons revealed that most breeding parameters are significantly different between the urban and rural blue tits. In both seasons, urban birds showed a significantly lower clutch size, hatching and fledging success compared to the rural ones. Additionally, fledging success at both sites was considerably lower in one breeding season (2017), during which nestling body weight was significantly lower in urban than rural birds; however, in 2016, when fledging success was higher, the two populations barely differed in their weight. Second, mirroring nestling body weight, during the 2017 breeding season, fCORT levels in nestlings were significantly higher in the urban nestlings compared to their rural counterparts, but not in 2016, further suggesting that 2017 was a more challenging year. However, I found no association between the reduction in fitness-related traits (body weight and fledging success) and fCORT. This lack of association between fCORT levels and nestlings’ fitness-related traits could have been missed as I pooled the feather samples per nest and did not measure individual nestlings for fCORT level, thus losing the inter-individual variation in fCORT levels that could be associated with fitness-related traits. Third, mirroring the fCORT variations, Leucocytozoon parasite infections varied in the two populations across seasons, showing that urban populations had lower or higher prevalence compared to rural birds depending on the year (2016 and 2017, respectively). I found a strong association between infection with Leucocytozoon prevalence and lower weight of urban nestlings just before fledging (day 13 of age) as well as a reduction in urban nestling survival, which instead was not observed in rural birds. I found no association between infection prevalence and fCORT level, measured per nest, not individuals. Finally, experimentally tested in the field, the origin of the bird did not influence the infection susceptibility to Leucocytozoon, nor did it influence fCORT levels. In summary, my thesis highlighted the importance of year to year variation between the two populations (i.e. the urban and the rural populations of blue tits), that could be influenced by fluctuating environmental factors such as weather and food availability. Urban and rural populations that show similarities in certain traits during one year of study may be different during another year (e.g. fCORT level and body weight). Prevalence of vector-borne pathogens like Leucocytozoon parasites in a given population may also differ between populations, which can vary from year to year. The extent of the fitness effect of parasite infection also depends on various factors fluctuating from one year to another. This emphasises the need for longitudinal studies monitoring individuals and populations over multiple years and across a wide range of habitats that differ in quality and features

Host-parasite dynamics in the endangered yellow-eyed penguin (Megadyptes antipodes) : investigations of Plasmodium and Eimeria in geographically distinct populations : a thesis presented in partial fulfilment of the requirements for the degree of Master of Veterinary Science in Wildlife Health at Massey University, Palmerston North, Manawatu, New Zealand

The yellow-eyed penguin (Megadyptes antipodes) is an endangered species endemic to New Zealand, facing ongoing threats to the persistence of the species. The yellow-eyed penguin occupies a restricted range including mainland New Zealand and adjacent islands, and the subantarctic Auckland Islands and Campbell Island. There has been no effective migration between the birds occupying mainland New Zealand, and the subantarctic islands, creating two population centres that are both geographically and genetically distinct. The two population centres of the yellow-eyed penguin, defined by the geographical range, occupy vastly different habitat ranges and face ongoing site-specific threats. The mainland population has declined by up to 65% in the last 20 years, while the sub-Antarctic population appears stable through limited monitoring. Examining host-pathogen dynamics of geographically distinct wildlife populations can inform predictions of a population’s response to major climate shifts or translocations. This study therefore explored possible differences in the host-pathogen dynamics between the two population centres of the yellow-eyed penguin. The study sampled wild penguins from the two major population centres, the mainland and sub-Antarctic population centres, to examine the prevalence and pathogen load differences of two known parasites Eimeria and Plasmodium. In addition, the study documented differences in the prevalence of Plasmodium infection between penguins in rehabilitation and those in the wild. Eimeria is a host-adapted parasite that principally causes disease in wild birds when the host is immunosuppressed through stress or intercurrent disease. Therefore, this study used Eimeria as an ecological biomarker to assess the distinct wild yellow-eyed penguin populations. The first morphological description of a novel species of Eimeria protozoa from a yellow-eyed penguin host is provided in this study. A high prevalence and pathogen load of Eimeria oocysts in faecal samples from wild yellow-eyed penguins was identified from the wild sub-Antarctic (apparent prevalence 76.6%, 95% CI 62.78-86.40% and mean pathogen load 9723 +/- 5831 oocysts/gram) and mainland populations surveyed (apparent prevalence 58.5%, 95% CI 43.37-72.24% and mean pathogen load 1050 +/- 398 oocysts/gram), with only weak evidence for a difference in the apparent prevalence and pathogen load between the two population centres. There was, however, a significant difference in body condition scores of infected penguins between the two populations, suggesting possible differences in the host-pathogen dynamics between the two distinct population centres. These results confirm the hypothesis that these coccidia are characteristic of a highly host-adapted endemic parasite in the yellow-eyed penguins. Penguins have a known susceptibility to Plasmodium sp., with infection capable of causing significant morbidity and mortality. This study provides the first report of a positive result for Plasmodium sp. from wild yellow-eyed penguins using LSU-RNA qPCR, and documents the estimated pathogen load (mean pathogen load 122 +/-29 Plasmodium DNA copies / 10,000 avian cells) of infected wild birds. There was good evidence for a difference in apparent prevalence between the two population centres, with a higher apparent prevalence of avian malaria in the wild mainland population (6.8%, 95% CI 2.96-15.05%), and an apparent absence of infection in the sub-Antarctic population surveyed (0%, 95% CI 0-5.58%). The study also documented a high apparent prevalence (65.9%, 95% CI 51.14-78.12%) of Plasmodium sp. infection in yellow-eyed penguins in rehabilitation, identifying three separate strains of avian malaria (Plasmodium sp. Lineage LINN1, Plasmodium relictum lineage SGS1 and Plasmodium elongatum lineage GRW06) via molecular sequencing. These results confirm the hypothesis that avian malaria is characteristic of a vector-borne parasite in the yellow-eyed penguins, and despite the presence of competent vectors in both habitats, only the northernmost population of yellow eyed penguins is currently infected. However, this disease is likely to emerge in the sub-Antarctic population in response to predicted climate shifts. The results of this work provide a platform for further research into the host-pathogen dynamics of the novel Eimeria species identified, and the potential host effects of this parasite during environmental stressors. In addition, the findings of this study suggest future monitoring of avian malaria prevalence and mortality rates in the yellow-eyed penguin are critical to understand the emerging risk of this pathogen in the context of ongoing climate shifts. Finally, the high prevalence of Plasmodium infection documented in yellow-eyed penguins in rehabilitation demonstrates the threat of this parasite to the success of ongoing rehabilitation efforts, and the need to investigate measures to mitigate infection risk for this species in the future

Calcium, telomere length, and parasitism in passerines nesting at high elevation

2020 Spring.Includes bibliographical references.Most organisms are exposed to numerous environmental stressors at various points throughout life, and, through natural selection, organisms' responses to such stressors have been optimized by natural selection for the best fitness outcomes. During the breeding season, wild vertebrates often make a trade-off between current reproduction and self-maintenance when dealing with environmental stressors. The total cost of reproduction is made up of all of the resources and energy that go into activities related to reproduction (e.g., nest building, finding a mate, foraging for food and nutrients related to offspring production, parental care) that do not go into self-maintenance. The cost of these activities can vary depending on resource availability, where limited resources can increase the cost associated with breeding due to increased energy associated with foraging and competing for the resource. In birds, calcium is a critical resource due to its importance in egg production and offspring development, and low calcium availability often leads to decreased reproductive success. In my first chapter, I used an experimental approach to assess the effects of supplemental calcium on reproductive parameters of Tree Swallows (Tachycineta bicolor) in a high elevation environment. Calcium-supplemented birds in my study area laid more, larger eggs, and had higher hatching success compared to control females. These results provide evidence that calcium availability is a constraint on breeding Tree Swallows at high elevation, perhaps due to the harsh conditions and concomitantly higher metabolic costs that force a costlier and more intense trade-off between foraging for food or for calcium. The increase in reproductive parameters for calcium supplemented nests in Chapter 1 highlights a cost associated with calcium foraging that constrains reproduction. For my second chapter, I aimed to better understand how calcium availability affects the cost of reproduction in mother Tree Swallows and offspring by using telomere shortening as a proxy of life stress and lifespan. Telomeres are terminal features of chromosomes consisting of repetitive DNA sequences that shorten with age and stress, and whose length is positively correlated with survival. I used telomere shortening as a proxy for the costs associated with reproduction to better understand life history trade-offs of Tree Swallows at high elevation sites. Similar to Chapter 1, I found that Tree Swallows supplemented with calcium had higher reproductive success, although I also found that supplemented nests had more telomere shortening compared to birds at control nests. These results provide evidence that Tree Swallows supplemented with calcium experience higher reproductive output at the cost of lower expected survival in the form of more telomere shortening. While investing resources in reproduction may lead to higher reproductive output for the current breeding season, this increase in reproductive success can come with a cost to survivorship. One way that resource allocation can shape survivorship is through investment in immune function. In many systems, however, more species-level and individual-level research is needed on host-parasite relationships before trade-offs between immune function and reproduction can be assessed. For my third chapter, I conducted a survey of avian Haemosporida: blood parasites that include those that cause avian malaria. I surveyed an avian community for haemosporidian parasites in the Colorado Rocky Mountains in order to estimate prevalence and diversity of blood parasites and to find species-level and individual-level characteristics that influence infection prevalence. I found that open cup nesters have higher blood parasite prevalence than cavity or open cup nesters. Additionally, male Ruby-crowned Kinglets, White-crowned Sparrows, and Wilson's Warblers had a higher prevalence of haemosporidian parasites compared to the other species analyzed, as did Red-breasted Nuthatches, which, like Ruby-crowned Kinglets, have a high body condition index. This chapter presents baseline knowledge of avian blood parasite presence, prevalence, and diversity across avian species in the Colorado Rocky Mountains and adds to our knowledge of host-parasite relationships of blood parasites and their avian hosts