Genomic organization of the major histocompatibility complex (MHC) can profoundly influence gene function and multigene family evolution. Situated at the interface of individual genetic variation and the adaptive immune response, MHC class I and II loci are intensively studied for disease associations and used as markers of adaptive genetic variation in evolutionary ecology research. Genomic sequence of MHC-containing cosmid clones from the red-billed gull (Larus scopulinus, Charadriiformes: shorebirds, gulls, and allies) was obtained for comparative analysis of avian MHC evolution. MHCI polymorphism was further investigated using cDNA library screening and locus-specific genotyping protocols. This first information regarding MHC organization and MHCI variation in charadriiforms suggests a complex evolutionary history to MHC architecture in birds. Duplication of MHCIIα loci in tandem MHCIIα/β pairs and their proximity to MHC-region gene COL11A2 are similar to arrangements in nonavian vertebrates, and contrast with the “minimal essential” MHC of the chicken (Gallus gallus, Galliformes: gamebirds). MHCI–TAP2 organization is shared with Galloanserae (gamebirds + waterfowl), as is a proposed major classical function for this MHCI gene. In contrast, the placement of MHCI genes adjacent to sequence from chromosomes 3, 5, and 22 of the chicken and zebra finch (Taeniopygia guttata, Passeriformes: perching birds) indicates interchromosomal rearrangements in birds and the possible genomic dispersal of nonclassical MHCI genes in the red-billed gull. Screening for avian malaria, genetic parentage tests, and field data from red-billed gulls at Kaikoura Peninsula, New Zealand were combined with MHCI genotypes to investigate relationships with disease and reproduction. Plasmodium infection was confirmed in red-billed gulls, and breeding condition was negatively associated with malarial infection and positively related to variation at the putative major MHCI locus. A low rate of extrapair paternity was identified across thirteen breeding seasons. Partners without extrapair young (EPY) had greater MHCI dissimilarity than was expected by chance, whereas lower individual MHCI variation and elevated hatching failure existed for pairs with EPY. In addition to contributing to studies of MHC evolution, sexual selection, and disease dynamics in the New Zealand avifauna, this research will facilitate studies of MHC genes in related charadriiforms, many of which are of conservation concern.

The species–area relationship (SAR) has been applied to predict species richness declines as area is converted to human-dominated land covers.In many areas of the world, however, many species persist in human-dominated areas, including threatened species. Because SARs are decelerating nonlinear, small extents of natural habitat can be converted to human use with little expected loss of associated species, but with the addition of more species that are associated with human land uses. Decelerating SARs suggest that, as area is converted to human-dominated forms, more species will be added to the rare habitat than are lost from the common one. This should lead to a peaked relationship between richness and natural area. I found that the effect of natural area on avian richness across Ontario was consistent with the sum of SARs for natural habitat species and human-dominated habitat species, suggesting that almost half the natural area can be converted to human-dominated forms before richness declines. However, I found that this spatial relationship did not remain consistent through time: bird richness increased when natural cover was removed (up to 4%), irrespective of its original extent. The inclusion of metapopulation processes in predictive models of species presence improves predictions of diversity change through time dramatically. Variability in site occupancy was common among bird species evaluated in this study, likely resulting from local extinction-colonization dynamics. Likelihood of species presence declined when few neighbouring sites were previously occupied by the species. Site occupancy was also less likely when little suitable habitat was present. Consistent with expectations that larger habitats are easier targets for colonists, habitat area was more important for more isolated sites. Accounting for the effect of metapopulation dynamics on site occupancy predicted change in richness better than land cover change and increased the strength of the regional richness–natural area relationship to levels observed for continental richness–environment relationships suggesting that these metapopulation processes “scale up” to modify regional species richness patterns making them more difficult to predict. It is the existence of absences in otherwise suitable habitat within species’ ranges that appears to weaken regional richness–environment relationships.

Los virus de influenza aviar de alta patogenicidad (IAAP) causan una enfermedad muy severa en pollos, los cuales frecuentemente inducen lesiones en el sistema nervioso central (SNC). Esta tesis recoge los resultados de tres estudios que se llevaron a cabo para determinar el mecanismo de patogénesis y neurotropismo, así como establecer la ruta de entrada al SNC para un virus H7N1 IAAP. En el primer estudio se estableció un modelo animal de infección en pollos libres de patógenos específicos, que consistía en la inoculación intranasal con el virus H7N1 IAAP. Para establecer este modelo, se utilizaron tres diferentes dosis del virus, obteniendo que las dosis más altas producen una enfermedad similar a la reportada para otros virus de IAAP. Además, se observó que las dosis más bajas causan infección demostrada porque con las dosis más bajas, el virus es hallado en muestras de tejido, muestras de heces y secreciones respiratorias. Adicionalmente, se pudo comprobar el alto neurotropismo del virus, ya que aún en pollos inoculados con bajas dosis el RNA viral es hallado en el CNS. La viremia fue detectada a un día post infección (dpi), sugiriendo que está podría ser la vía de diseminación al SNC. En el segundo estudio, se determinó la distribución topográfica del antígeno viral en el SNC durante las primeras horas post infección, lo cual permitió determinar que el virus se disemina de forma simétrica y bilateral en núcleos neurales del diencéfalo, mesencéfalo y rombencéfalo. La distribución del antígeno viral indica que el bulbo olfatorio y los nervios periféricos están involucrados en el proceso de invasión del SNC. El hallazgo de receptores aviares y humanos en las células endoteliales explica porque estas células son tan sensibles a la infección. El RNA viral fue hallado en el líquido cerebro espinal el primer dpi, lo que indica que el virus atraviesa la barrera hemato-encefálica (BHE). En el tercer estudio, la alteración de la BHE inducido por el virus H7N1 IAAP fue demostrado usando tres diferentes métodos que incluye la perfusión intracardial de Azul de Evans, la detección de la extravasación de la proteína del suero IgY, y evaluación del patrón de tinción con el marcador de las uniones fuertes de la BHE, ZO-1 y claudin-1. El antígeno viral fue observado a las 24 hpi en las células endoteliales, mientras que el daño de la BHE fue observado a las 36 hpi y 48 hpi. En resumen, se puede afirmar que el virus H7N1 IAAP se disemina por la vía hematógena durante las primeras horas pi, posiblemente favorecido por la presencia de receptores en las células endoteliales del sistema nervioso central, y poco después daña la BHE durante las primeras horas de infección como se demuestra por la presencia de extravasación del azul de Evans and IgY del suero. / Highly pathogenic avian influenza viruses (HPAIV) cause a very severe systemic disease in chickens, in which is also frequent to find central nervous system (CNS) lesions. In this thesis, three studies were undertaken in order to determine the mechanism of pathogenesis, the neurotropism and establish the route of entry into the CNS use for a H7N1 HPAI virus. In the first study, an animal model was set up that consisted of SPF chickens inoculated intranasally with the H7N1 HPAI virus. To do that, three different doses were used, obtaining that the highest dose induced a disease similar to the produce by other HPAI viruses, moreover, it was also observed that very low doses also cause infection demonstrated because viral RNA was found in tissues samples, faeces and respiratory secretions. Besides, the high neurotropism of this virus was demonstrated because still in chickens inoculated with low doses, viral RNA is found in the brain. Viremia was detected at one dpi, which indicated that the bloodstream is the pathway of viral spreading to the brain. In the second study, the topographical distribution study of the viral antigen during the first dpi was determined, which allow to determine that the virus disseminates showing a symmetrical and bilateral pattern in the diencephalon, mesencephalon and rhombencephalon, whereas in the telencephalon and cerebellum it was multifocal and random. Viral antigen distribution indicates that the olfactory bulb (OB) and peripheral nerves are not involved in the process of virus invasion into the brain. Avian and human influenza receptors were found in endothelial cells which explain why these cells are so sensitive to the infection. Viral RNA was found in cerebrospinal fluid (CSF) at one dpi, indicating that the virus was able to cross blood brain barrier (BBB). In the third study, the disruption of the BBB induce by the H7N1 HPAI was demonstrated using three different methods that include the intracardial perfusion of the tracer Evans blue (EB), detection of the extravasation serum IgY, and evaluation of the pattern of staining of the tight junction proteins ZO-1 and claudin-1. Viral antigen can be observed as early as 24 hpi in the endothelial cells, whereas disruption was detected at 36 and 48 hpi. In summary, it can be asserted that this H7N1 HPAIV disseminates via the haematogenous route early during the infection, favored by the presence of abundant receptors on the CNS endothelial cells, and soon after it disrupts the BBB during the first hours of infection as demonstrated by the presence of EB and serum IgY extravasation.

A previous study in chickens revealed that myosin light chain kinase (MLCK), f actin, and myosin are found on the crystalline lens. Their polygonal arrangement at the posterior surface resembles a muscle tissue, which suggests that these proteins may have a contractile role in accommodation. The ciliary muscle in chickens is skeletal in nature and, therefore, chickens were used to test the hypothesis that contractile microfilaments play a role in accommodation. Ciliary nerve-induced accommodation was measured in the presence of an MLCK inhibitor 1-(5-Iodonaphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine hydrochloride (ML-7). Eyes of 6-day old white Leghorn chickens (gallus gallus domesticus) were enucleated in Tyrode’s saline solution while keeping the ciliary nerve intact. One eye was treated with ML-7 and the other eye was treated with vehicle only. Three concentrations of ML-7 were used: 1 µM, 10 µM, and 100 µM. Two experiments were carried out, one including a (3×10 min) wash and one without. Focal lengths of the vehicle- and ML-7-treated eyes were measured before, during and after accommodation. Immunoblots were used to detect the amount of phosphorylated myosin with and without the inhibitor. Focal lengths for accommodation were shorter than those at rest (p<0.001). In the wash experiment, vehicle-treated eyes had higher accommodative amplitudes compared to ML-7-treated eyes for all three dosage groups. In the no-wash experiment, only the 1 µM group demonstrated the same trend as the wash experiment. For the 10 µM and 100 µM groups, ML-7-treated eyes had higher accommodative amplitudes compared to vehicle-treated eyes. Immunoblots revealed varying amounts of inhibition within pairs of eyes as well as between birds for both experiments. Results from this experiment indicate that ML-7 was not effective at determining whether contractile microfilaments found on the lens contribute to accommodation.

A previous study in chickens revealed that myosin light chain kinase (MLCK), f actin, and myosin are found on the crystalline lens. Their polygonal arrangement at the posterior surface resembles a muscle tissue, which suggests that these proteins may have a contractile role in accommodation. The ciliary muscle in chickens is skeletal in nature and, therefore, chickens were used to test the hypothesis that contractile microfilaments play a role in accommodation. Ciliary nerve-induced accommodation was measured in the presence of an MLCK inhibitor 1-(5-Iodonaphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine hydrochloride (ML-7). Eyes of 6-day old white Leghorn chickens (gallus gallus domesticus) were enucleated in Tyrode’s saline solution while keeping the ciliary nerve intact. One eye was treated with ML-7 and the other eye was treated with vehicle only. Three concentrations of ML-7 were used: 1 µM, 10 µM, and 100 µM. Two experiments were carried out, one including a (3×10 min) wash and one without. Focal lengths of the vehicle- and ML-7-treated eyes were measured before, during and after accommodation. Immunoblots were used to detect the amount of phosphorylated myosin with and without the inhibitor. Focal lengths for accommodation were shorter than those at rest (p<0.001). In the wash experiment, vehicle-treated eyes had higher accommodative amplitudes compared to ML-7-treated eyes for all three dosage groups. In the no-wash experiment, only the 1 µM group demonstrated the same trend as the wash experiment. For the 10 µM and 100 µM groups, ML-7-treated eyes had higher accommodative amplitudes compared to vehicle-treated eyes. Immunoblots revealed varying amounts of inhibition within pairs of eyes as well as between birds for both experiments. Results from this experiment indicate that ML-7 was not effective at determining whether contractile microfilaments found on the lens contribute to accommodation.

Habitat selection theory presumes that organisms are not distributed randomly in their environments because of habitat-specific differences in reproductive success and survival; unfortunately, many previous studies were either unable or failed to look for evidence of processes shaping nest site selection patterns. Furthermore, little is known about adaptive nest site selection in northern environments where habitats often have little vegetation and time and climatic constraints may be pronounced. Therefore, I investigated patterns of nest site selection by common eider ducks (<i>Somateria mollissima</i>) at an island colony in Canadas Eastern Arctic, and looked for evidence of selective processes underlying these patterns by employing experimental and observational techniques.<p>I characterized physical features of (a) non-nest sites (b) active nest sites and (c) unoccupied nest sites that had been used in previous years. Habitat features that distinguished non-nest sites from unoccupied nest sites were also important in distinguishing between active and unoccupied nest sites during the breeding season. Active nest sites were closer to herring gull (<i>Larus argentatus</i>) nests, farther from the ocean and had organic substrates. In general, habitat features associated with nest use were not strongly associated with success after the onset of incubation. Nests near fresh water ponds were more successful in one study year, but in the other two study years successful nests were initiated earlier and more synchronously than were unsuccessful nests. Common eiders settled to nest first near the geographic centre of the colony, whereas sites near the largest fresh water pond were occupied later; distance to ocean had no observable effect on timing of nesting. Nest density was greater farther from the ocean, but timing of nest establishment did not differ between high and low density plots. <p>I tested whether moss or duck down placed in nest bowls could increase nest establishment, or advance laying date. I placed this extraneous material in bowls before nesting and found no difference in likelihood of nest establishment; however, bowls containing duck down were initiated earlier (or had higher survival) than those containing no nesting material. To investigate the role of nest shelter and microclimate in nest site choices and female body condition, I placed plywood nest shelters over established nests. Temperature probes indicated that artificially-sheltered females experienced more moderate thermal environments and maintained higher body weight during late incubation than did unsheltered females. However, few eiders nested at naturally-sheltered sites, possibly because nest concealment increases susceptibility to mammalian predators. My results suggest that eider nest choices likely reflect trade-offs among selective pressures that involve the local predator community, egg concealment, nest microclimate and energy use.

Cystatin C, a 13 kDa low molecular weight protein is an inhibitor of cysteine proteases. Due to its low molecular weight and positive charge at physiological pH, it is freely filtered by the glomerulus and catabolized after reabsorption by proximal tubular cells with a low concentration (0.03-0.3 mg/L) in urine amongst healthy subjects. Urinary cystatin C is a potential biomarker detection of acute kidney injury (AKI) in the acute phase when patients are submitted to the intensive care unit. The aim in this report was to perform a full method validation of urinary analysis of cystatin C on a high throughput chemical analyzer by particle-enhanced turbidimetric immunoassay (PETIA) at the University Hospital in Uppsala, Sweden. The antigen excess, linearity, lower limit of quantification (LoQ), recovery, assay precision, stability and interference caused by haemoglobin was evaluated. No hook effect was observed, the assay was linear over the studied interval &lt;0.001-0.950 mg/L with a regression of R2=0.9994. The LoQ was calculated to 0.020 mg/L with a coefficient of variation (CV) ≤10% which was considered acceptable. The assay had a recovery between 93-100% and the assay precision had a total CV &lt;3.5%. Cystatin C is stable for 3 days in room temperature and 14 days in +4C. The assay did not show any major interference with haemoglobin. The urinary cystatin C showed good precision and performance characteristics by measurements using PETIA all of which is a necessary qualification for a biomarker at a 24-h running routine laboratory.

Recent World Animal Health Organization (OIE) reports on Avian Influenza (AI) outbreaks in Asia, Europe and Canada suggest that there is a nonzero probability that an outbreak may occur anywhere in the world, including the US. To help evaluate possible policy in the face of such an event, this dissertation does an economic evaluation of the implications of using two mitigation strategies: one corresponding to the currently response strategy; and the other an OIE recommended one utilizing vaccination. To do this, the dissertation develops and uses an integrated economic-epidemic model. In this effort, I first estimate the cost of an AI outbreak under a deterministic disease spread assumption where a new vaccination strategy and the current strategy are compared. Subsequently, I introduce risk in the model and construct 95 percent confidence intervals for the outbreak costs, and I rank the outcomes of the alternative strategies using stochastic dominance criteria. In addition, during both phases, I develop and estimate the breakeven probability for an event where ex-ante fixed costs of vaccine stockpiling are justified by the reduction in disease event damages. Results under deterministic disease spread assumption suggest that the vaccination strategy lowers the cost of outbreaks as opposed to the current strategy. This happens because vaccination reduces the number of culled and quarantined flocks. The study is conducted in three locations, yielding the finding that the costs of an outbreak vary depending on the densities of poultry flocks. I also find that when consumer demand shifts due to the outbreak, the costs are much larger. Finally, I find that ex-ante vaccine stockpiling is justified for all the sub-regions if the probability of outbreak exceeds 0.07. The stochastic disease spread assumption results also show that the vaccination strategy dominates in first degree stochastic dominance sense. Consistent with stochastic dominance results, the 95 percent confidence intervals have narrower ranges under the vaccination strategy than without it. Finally, the distribution of the breakeven probability for vaccine stocking has a mode of 0.07 and that the probability is accurate with 82 percent likelihood. However, the threshold varies with the disease transmission parameters and could reach up to 0.32.

Habitat selection research has focused on the role of vegetative and geologic habitat characteristics or antagonistic behavioral interactions. Conspecifics can confer information about habitat quality and provide positive density-dependent effects that may result in improved fitness, resulting in positive behavioral responses to conspecifics as a habitat selection strategy. I conducted 3 replicated, manipulative experiments to investigate use of conspecific cues in habitat selection for the golden-cheeked warbler (Dendroica chrysoparia) using simulated conspecific vocalizations during pre-settlement and post-breeding periods, across a range of woodland canopy cover. I measured territory density, pairing, and fledging success in paired treatment and control units. Territory density was >2 times higher in treatment units across the range of canopy (P = 0.02). Pairing success was positively correlated with territory density (P = 0.008). Territory density response was higher for pre-settlement than post-breeding treatment (P = 0.004). I found pre-settlement and post-breeding conspecific cues influence golden-cheeked warbler habitat selection, inducing settlement in previously unoccupied areas, and producing aggregations within areas of similar vegetative characteristics. Better understanding of social information use in habitat selection can improve our understanding of species distributions, yielding more accurate predictive distribution models; improve our ability to predict impacts of habitat changes on habitat use, survival, reproduction, and ultimately fitness; and provide a potential tool for attracting individuals to restored or managed sites.

Man-made farm ponds are unique geographic features of the Taoyuan Tableland. Besides irrigation, they provide refuges for wintering birds. The issue at hand is that these features are disappearing and bring with it the loss of this refuge function. It is ecologically significant because one fifth of all the bird species in Taiwan find a home on these ponds. This study aims at characterizing the diversity of bird species associated with these ponds whose likelihood of survival was assessed along the gradient of land development intensities. Such characterization helps establish decision criteria needed for designating certain ponds for habitat preservation and developing their protection strategies. A holistic model was developed by incorporating logistic regression with error back-propagation into the paradigm of artificial neural networks (ANN). The model considers pond shape, size, neighboring farmlands, and developed areas in calculating parameters pertaining to their respective and interactive influences on avian diversity, among them the Shannon-Wiener diversity index (H’). Results indicate that ponds with regular shape or the ones with larger size possess a strong positive correlation with H’. Farm ponds adjacent to farmland benefited waterside bird diversity. On the other hand, urban development was shown to cause the reduction of farmland and pond numbers, which in turn reduced waterside bird diversity. By running the ANN model with four neurons, the resulting H’ index shows a good-fit prediction of bird diversity against pond size, shape, neighboring farmlands, and neighboring developed areas with a correlation coefficient (r) of 0.72, in contrast to the results from a linear regression model (r < 0.28). Analysis of historical pond occurrence to the present showed that ponds with larger size and a long perimeter were less likely to disappear. Smaller (< 0.1 ha) and more curvilinear ponds had a more drastic rate of disappearance. Based on this finding, a logistic regression was constructed to predict pond-loss likelihood in the future and to help identify ponds that should be protected. Overlaying results from ANN and form logistic regression enabled the creation of pond-diversity maps for these simulated scenarios of development intensities with respective to pond-loss trends and the corresponding dynamics of bird diversity.