The Japanese Crested Murrelet Synthliboramphus wumizusume occupies a limited range in Southern Korea and Japan and is considered vulnerable by the IUCN. There is strong indication of a colony of Japanese Murrelets located in Kaminoseki, Japan; however, no nests or individuals have yet been found. The is also evidence that murrelets make use of this habitat during their vulnerable autumnal molting season during which they cannot fly. This habitat is threatened by the construction of a nuclear power plant in Tanoura Bay. Construction of this plant would result in loss of nesting sites, food supply, and other components vital to the survival of the colony. This study attempts to detect the presence of Japanese Murrelets in Kaminoseki using bioacoustic monitoring of songmeters placed around Tanoura Bay. Preliminary sonograms created using the R package “Bioacoustics” did not yield conclusive results regarding the presence of Japanese Murrelets as the program captured background noise but did not pick up on bird calls heard during manual playback of the WAV files. Further research must be completed to refine the settings used in the program in order to conduct a more definitive analysis of the dataset.

Thesis (M. Phil.)--University of Hong Kong, 2006. / Title proper from title frame. Also available in printed format.

Thesis (M. P. A.)--University of Hong Kong, 2006. / Title proper from title frame. Also available in printed format.

<p>Avian Influenza (AI) is a contagious viral zoonotic disease with great public health implications and negative socioeconomic impact (WHO, 2006a). The highly pathogenic avian influenza (HPAI) infection is transmitted from birds to man mostly through contact with contaminated poultry and objects (INFOSAN, 2005), hence people who come in contact with birds such as live bird sellers (LBS) are the more vulnerable population (WHO, 2006a). Inadequate knowledge of AI health risks and poor practice of AI preventive measures amongst LBS increases the risk of spread of the infection in both humans and animals.The aim of this study was to describe and quantify the knowledge and practice of LBS with regards to avian influenza health risks and preventive activities in Agege, an urban area in Lagos State, Nigeria.</p>

Many physiological responses to dioxin-like compounds (DLCs), including polychlorinated dibenzodioxins (PCDDs) and polychlorinated dibenzofurans (PCDFs) are mediated by the aryl-hydrocarbon receptor (AhR). In birds, activation of the AhR stimulates the transcription of cytochrome P4501A (CYP1A) genes, including CYP1A4 and CYP1A5, and ultimately leads to expression of biotransformation enzymes, including ethoxyresorufin-O-deethylase (EROD). It is well established that potencies of different DLCs range over several orders of magnitude. There is also a wide variation among birds in their responsiveness to DLCs both in efficacy and threshold for effects. A molecular basis for this differential sensitivity has been suggested. Specifically, a comparison of the AhR ligand-binding domain (LBD) indicated that key amino acid residues are predictive of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) sensitivity. Based on sequencing of the AhR LBD from numerous avian species a sensitive classification scheme has been proposed (in order of decreasing sensitivity, chicken (type I; sensitive) > Common pheasant (type II; moderately sensitive) > Japanese quail (type III; insensitive)). A series of egg injection studies with White-leghorn chicken (<i>Gallus gallus domesticus</i>), Common pheasant (<i>Phasianus colchicus</i>) and Japanese quail (<i>Coturnix japonica</i>) were performed to determine whether molecular and biochemical markers of exposure to DLCs are predictive of the proposed classification scheme. In addition, I was interested in determining whether this classification scheme applies to other DLCs, specifically dibenzofurans. Determining which species are "chicken- like", "pheasant-like" and "quail-like" in their responses to DLCs should allow more refined risk assessments to be conducted as there would be less uncertainty about the potential effects of DLCs in those species for which population-level studies do not exist.<p> Several concentrations of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), 2,3,4,7,8-pentachlorodibenzofuran (PeCDF), or 2,3,7,8-tetrachlorodibenzofuran (TCDF) (triolein vehicle) were injected into the air cells of Japanese quail, Common pheasant and chicken eggs. Liver from 14 d post-hatch chicks was harvested for analysis of CYP1A4 and CYP1A5 mRNA abundance by quantitative real-time PCR (Q-PCR), and EROD activity. Lowest observed effective concentration (LOEC) and relative potency (ReP) values for CYP1A mRNA abundance and EROD activity were determined and used to make comparisons of sensitivity between each species and DLC potency within each species.<p> The TCDD is widely considered to be the most potent DLC and this is supported by the rank order of LOEC values for CYP1A5 mRNA abundance in White-leghorn chicken (TCDD > PeCDF > TCDF). CYP1A4 mRNA abundance and EROD activity in White-leghorn chicken were significantly increased in the lowest dose exposure groups of each of the three DLCs, so the potency of these compounds based on these endpoints was not established. Interestingly, TCDD was not the most potent DLC in Common pheasant and Japanese quail. In Common pheasant, PeCDF is the most potent as a CYP1A4 mRNA inducer, followed by TCDD and TCDF. However, TCDF was the most potent EROD activity inducer for Common pheasant, followed by PeCDF, and then TCDD. No significant increases were found in CYP1A5 mRNA abundance in pheasant within the tested dose ranges for all the three DLCs. No significant increases in either CYP1A5 mRNA abundance or EROD activity were found in Japanese quail. In addition, PeCDF and TCDF, but not TCDD, significantly increased CYP1A4 mRNA abundance.<p> According to the predicted relative sensitivity by comparing the AhR LBD amino acid sequences, the White-leghorn chicken is more responsive to DLCs than the Common pheasant which is more responsive than the Japanese quail. By comparing the relative sensitivity calculated based on the LOEC values from my study, the sensitivity order to TCDD and TCDF support the proposed molecular based species sensitivity classification scheme (chicken > pheasant > quail), while pheasant is almost as sensitive as chicken to PeCDF ( pheasant ¡Ý chicken > quail).<p> Taken together, the data suggest that TCDD is the most potent DLC in White-leghorn chicken, but not in Common pheasant, or or Japanese quail. The data suggest that in type II avian species PeCDF may be more potent than TCDD. In addition, I found in my study that different biomarkers have different responses, which depends on species and chemicals as well. These data provide further insight into avian sensitivities to DLCs.</p>

Many physiological responses to dioxin-like compounds (DLCs), including polychlorinated dibenzodioxins (PCDDs) and polychlorinated dibenzofurans (PCDFs) are mediated by the aryl-hydrocarbon receptor (AhR). In birds, activation of the AhR stimulates the transcription of cytochrome P4501A (CYP1A) genes, including CYP1A4 and CYP1A5, and ultimately leads to expression of biotransformation enzymes, including ethoxyresorufin-O-deethylase (EROD). It is well established that potencies of different DLCs range over several orders of magnitude. There is also a wide variation among birds in their responsiveness to DLCs both in efficacy and threshold for effects. A molecular basis for this differential sensitivity has been suggested. Specifically, a comparison of the AhR ligand-binding domain (LBD) indicated that key amino acid residues are predictive of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) sensitivity. Based on sequencing of the AhR LBD from numerous avian species a sensitive classification scheme has been proposed (in order of decreasing sensitivity, chicken (type I; sensitive) > Common pheasant (type II; moderately sensitive) > Japanese quail (type III; insensitive)). A series of egg injection studies with White-leghorn chicken (<i>Gallus gallus domesticus</i>), Common pheasant (<i>Phasianus colchicus</i>) and Japanese quail (<i>Coturnix japonica</i>) were performed to determine whether molecular and biochemical markers of exposure to DLCs are predictive of the proposed classification scheme. In addition, I was interested in determining whether this classification scheme applies to other DLCs, specifically dibenzofurans. Determining which species are "chicken- like", "pheasant-like" and "quail-like" in their responses to DLCs should allow more refined risk assessments to be conducted as there would be less uncertainty about the potential effects of DLCs in those species for which population-level studies do not exist.<p> Several concentrations of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), 2,3,4,7,8-pentachlorodibenzofuran (PeCDF), or 2,3,7,8-tetrachlorodibenzofuran (TCDF) (triolein vehicle) were injected into the air cells of Japanese quail, Common pheasant and chicken eggs. Liver from 14 d post-hatch chicks was harvested for analysis of CYP1A4 and CYP1A5 mRNA abundance by quantitative real-time PCR (Q-PCR), and EROD activity. Lowest observed effective concentration (LOEC) and relative potency (ReP) values for CYP1A mRNA abundance and EROD activity were determined and used to make comparisons of sensitivity between each species and DLC potency within each species.<p> The TCDD is widely considered to be the most potent DLC and this is supported by the rank order of LOEC values for CYP1A5 mRNA abundance in White-leghorn chicken (TCDD > PeCDF > TCDF). CYP1A4 mRNA abundance and EROD activity in White-leghorn chicken were significantly increased in the lowest dose exposure groups of each of the three DLCs, so the potency of these compounds based on these endpoints was not established. Interestingly, TCDD was not the most potent DLC in Common pheasant and Japanese quail. In Common pheasant, PeCDF is the most potent as a CYP1A4 mRNA inducer, followed by TCDD and TCDF. However, TCDF was the most potent EROD activity inducer for Common pheasant, followed by PeCDF, and then TCDD. No significant increases were found in CYP1A5 mRNA abundance in pheasant within the tested dose ranges for all the three DLCs. No significant increases in either CYP1A5 mRNA abundance or EROD activity were found in Japanese quail. In addition, PeCDF and TCDF, but not TCDD, significantly increased CYP1A4 mRNA abundance.<p> According to the predicted relative sensitivity by comparing the AhR LBD amino acid sequences, the White-leghorn chicken is more responsive to DLCs than the Common pheasant which is more responsive than the Japanese quail. By comparing the relative sensitivity calculated based on the LOEC values from my study, the sensitivity order to TCDD and TCDF support the proposed molecular based species sensitivity classification scheme (chicken > pheasant > quail), while pheasant is almost as sensitive as chicken to PeCDF ( pheasant ¡Ý chicken > quail).<p> Taken together, the data suggest that TCDD is the most potent DLC in White-leghorn chicken, but not in Common pheasant, or or Japanese quail. The data suggest that in type II avian species PeCDF may be more potent than TCDD. In addition, I found in my study that different biomarkers have different responses, which depends on species and chemicals as well. These data provide further insight into avian sensitivities to DLCs.</p>

Creation of a reliable and easy to use serologic test would greatly improve ante mortem diagnosis of Mycobacterium avium subsp. avium and aid in the control of avian mycobacteriosis, particularly in captive birds. In order to determine whether serodiagnostics could be of value in testing ring-neck doves (Streptopelia risoria) for M. a. avium infection, Western blot analysis was used to assess the humoral response of ring-neck doves exposed to M. a. avium, and to evaluate whether an association could be made between humoral response and necropsy findings, histopathology, culture, and PCR testing. Western blot results were examined for reactivity patterns associating the humoral response with infection status, severity and type of lesions (diffuse vs multifocal granulomatous inflammation) and phenotype (white vs non-white). A sensitivity of 88.24% and a specificity of 100% were achieved utilizing Western blot analysis to detect M. a. avium infection in ring-neck doves, offering a negative predictive value of 93% and a positive predictive value of 100%. While Western blot analysis results did not reflect lesion severity, lesion type did partially correspond with the humoral response. The findings of the present study indicate that serologic testing can be used as a valuable ante mortem screening tool for identifying ring-neck doves infected with M. a. avium.

Avian circadian clocks are composed of a distributed network of neural and peripheral oscillators. Three neural pacemakers, located in the pineal, the eyes, and the hypothalamus, control circadian rhythms of many biological processes through complex interactions with slave oscillators located throughout the body. This system, an astonishing reflection of the life history of this diverse class of vertebrates, allows birds to coordinate biochemical and physiological processes and harmonize them with a dynamic environment. Much work has been done to understand what roles these pacemakers have in avian biology, how they function, and how they interact to generate overt circadian rhythms. The experimental work presented in this dissertation uses the domestic chicken, Gallus domesticus, as a model to address these questions and carry forward current understanding about circadian biology in this species. To do so, we utilized a custom DNA microarray to investigate rhythmic transcription in cultured chick pineal cells. We then sought to identify genes which might be a component of the pineal clock by screening for rhythmic transcripts that are sensitive to a phase-shifting light stimulus. Finally, we surgically removed the eyes or pineal from chickens to examine the roles of these extra-SCN pacemakers in regulating central and peripheral rhythms in metabolism and clock gene expression. Using these methods, we show that the oscillating transcriptome is diminished in the chick pineal ex vivo, while the functional clustering of clock controlled genes is similar. This distribution reveals multiple conserved circadian regulated pathways, and supports an endogenous role for the pineal as an immune organ. Moreover, the robustness of rhythmic melatonin biosysnthesis is maintained in vitro, demonstrating that a functional circadian clock is preserved in the reduced subset of the rhythmic pineal transcriptome. In addition, our genomic screen has yielded a list of 28 genes that are candidates for functional screening. These should be evaluated to determine any potential role they may have as a component of the pineal circadian clock. Finally, we report that the eyes and pineal similarly function to reinforce rhythms in brain and peripheral tissue, but that metabolism and clock gene expression are differentially regulated in chick.

There are many pathogens that infect birds and perhaps many more that researchers have not yet identified. Of all potential pathogens, the research presented in this manuscript focuses on two avian RNA viruses. First, a serodiagnostic test for newly described Avian Borna Virus (ABV), which has been recently identified as the etiological agent of Proventricular Dilatation Disease (PDD), was developed. PDD is a deadly disease which affects many birds, but to this point, has mainly been a concern of psittacines. The need for a diagnostic test is imperative. An antigen associated with PDD was identified from the brains of affected birds by use of the Western blot assay. This antigen was subsequently purified using various protein purification protocols, including a modification of reverse-phase chromatography. The antigen was then identified as the ABV nucleoprotein according to tandem mass spectroscopy analysis and protein database search. A serodiagnostic assay was developed and standardized using infected cell culture as an antigen source. Over 100 avian serum samples were submitted by veterinarians to test for the presence of antibodies against ABV nucleoprotein. This serodiagnostic assay was found to have 90% sensitivity and 82% specificity for the diagnosis of ABV in infected birds. Second, the ability of a carbohydrate epitope to enhance the humoral immune response to an influenza vaccine was tested in chickens. Influenza is a serious infection that causes 36,000 deaths annually in the United States. The need for a more efficacious is addressed by incorporating a carbohydrate antigen targeted by natural antibodies that are produced by chickens as well as humans. Therefore, chickens may be a suitable animal model to test this hypothesis. Influenza vaccines with alpha-gal antigen are prepared from cell culture. The antigen is then enzymatically removed from some vaccines and the nature of the ensuing humoral immune response to these vaccines in chickens is attempted. Though ABV is not known to be zoonotic at this time, zoonotic infections pose the highest risk as new and emerging infectious diseases in the human population. The following research contains applications relative to challenges faced by researchers and clinicians in infectious disease containment.

A proper vaccination program can play a critical role in prevention and control of avian influenza (AI) in commercial poultry. Low pathogenic avian influenza viruses (LPAIV) of H5 and H7 AI subtypes cause serious economic losses to the poultry industry and have the potential to mutate to highly pathogenic AI (HPAI) strains. Due to trade implications, differentiation of infected from vaccinated animals (DIVA) is an important issue in the control of AI. Therefore, the development and characterization of vaccine candidates with DIVA properties is critical in improving vaccination programs. Keeping these aspects in mind, we investigated the role of an NS1 mutant virus as a potential live attenuated DIVA vaccine. The NS1 protein of influenza virus plays a major role in blocking the host's antiviral response. Using an eight-plasmid reverse genetics system, we recovered the low pathogenic parental (H5N3) and NS1 mutant (H5N3/NS1/144) viruses. H5N3/NS1/144 expresses only the first 144 amino acids of the NS1 protein compared to the 230 of the parental H5N3. The growth properties of H5N3 and H5N3/NS1/144 were compared in cell culture and in different age embryonated chicken eggs. Our results confirmed that NS1 is involved in down regulation of interferon as shown by IFN-beta mRNA expression analysis and by the inability of H5N3/NS1-144 to efficiently grow in older age, interferon competent, chicken embryos. However with regards to safety the virus reverted to virulence within five back passages in chickens and was therefore not a safe vaccine candidate. However the killed form of H5N3/NS1-144 was a safer alternative and it also induced antibody titers and protection not significantly different from the parental H5N3 as vaccine. To further understand the reversion of H5N3/NS1/144 to virulence, we carried out 3 independent serial passages of H5N3/NS1/144 in increasing age of embryonated chicken eggs and examined the NS1 gene for presence of mutations. RT-PCR and sequence analysis of the NS gene in all three lineages showed the presence of a 54 amino acid deletion resulting in the generation of a 87 amino acids long NS1 ORF with a point mutation (L80V) at the site of deletion. In addition, the NS1 ORF in lineages L2 and L3 presented two additional point mutations in the RNA binding domain (Q40R and T73M). To determine if these mutations played a role in increased virulence, recombinant viruses expressing these mutant NS1 proteins in the background of parental virus were generated by reverse genetics and their replication properties and pathogenicity was examined in vitro, in ovo and in vivo systems. Our results showed that the 87 amino acid long NS1 protein clearly increased virus replication and virulence specifically in interferon competent systems. In addition, the two point mutations in the RNA binding domain of NS1 ORF expressing 87 a protein slightly increased the virus virulence. Overall this study reinforces the role of NS1 in influenza virus pathogenicity and supports the use of killed inactivated NS1 mutant virus vaccines as potential DIVA vaccines.