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Thesis (Ph. D.)--University of Wisconsin--Madison, 1950. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references.

Marek's disease is a naturally occurring lymphoproliferative disease of chickens which is caused by a herpesvirus (Marek's disease virus, MDV). To determine whether the cells in Marek's disease lymphomas are homogeneous or consist of several lymphocytic subpopulations, lymphoid cells from a MDV infected group of chickens were compared with lymphoid cells from vaccinated chickens and uninfected chickens using computer assisted microphotometric analysis (CAMA). CAMA is a quantitative technique capable of differentiating cell populations based upon slight differences in the chromatin and cytoplasmic textures of stained cells. Lymphoma cells, splenocytes and peripheral blood lymphocytes (PBL) were stained with Papanicolaou stain and scanned on a Leitz MPVII microscope photometer operated on line to a PDP 11/45 computer. Two populations of lymphoid cell were detected in MD lesions using CAMA. One of these populations had features which were different from lymphoid cells from uninfected chickens and were termed "altered" cells. The other population had features which were similar to uninfected lymphocytes. Features were identified and selected that would classify cells as "altered" or "unaltered" and used to derive a decision rule to classify cells from other lymphoid populations. The decision rule classified up to 75% of cells from different lymphomas as "altered" cells. CAMA results were compared with clinical data and immunological data to quantitate T lymphocytes, B lymphocytes and MATSA (Marek's Disease Tumor Specific Antigen) positive cells. Results showed that there was a high correlation (r = 0.79) between the percentage of "altered" cells detected using CAMA and the percentage of MATSA positive cells. The majority of splenocytes and the PBL from the MDV group were classified as "unaltered".

Thesis (M.Sc.). / Written for the Dept. of Animal Science, Macdonald College of McGill University. Title from title page of PDF (viewed 2008/07/28). Includes bibliographical references.

Marek’s disease virus (MDV), the causative agent of Marek’s disease in chicken, is an important oncogenic avian pathogen which leads to world-wide economic losses in the poultry industry. It targets the chicken's immune system by initially causing a lytic infection in B-lymphocytes in lymphoid organs (spleen, bursa of Fabricius and thymus), followed by a latent infection of T-lymphocytes, which may lead to tumour formation. Despite the presence of well-established vaccination programs against MDV, it is still a major concern for the poultry industry due to the emergence of more virulent strains. As MDV is also considered an excellent model for herpesvirusinduced oncogenicity and immunosuppression, a better understanding of its pathogenesis, including the functional roles of individual MDV proteins, is of both biomedical as well as economical importance. All open reading frames (ORFs) of the CVI988 vaccine strain and the RB1B virulent strain were PCR-amplified from BAC DNA and cloned into the pDONR 207 entry vector by recombinatorial cloning (Gateway® system). Subsequently, all ORFs were subcloned into the yeast-two-hybrid (Y2H) vectors pGBKT7–DEST (bait) and pGADT7-DEST (prey), as well as other expression vectors. The Y2H bait and prey vector clone collections were transformed into the yeast strains AH109 and Y187, respectively. More than 140 ORFs, or ORF fragments, were analysed against each other in a comprehensive Y2H assay. Of > 20.000 interactions tested, 435 positive interactions between 115 ORFs were observed. Several of these interactions have previously been reported in other species of herpesvirus indicating that they may be conserved within the family. A subset of the positive interactions were confirmed using co-immunoprecipitation and LUMIER pull-down assays as a second independent assay. In the second part of the project all MDV proteins were tested for their ability to inhibit the chicken interferon-alpha (chIFN-α)-induced immune response. In functional luciferase reporter assay with a chicken Mx1 promoter containing an interferon stimulated responsive element (ISRE), four MDV-encoded chIFN-α inhibitors were identified, including UL12, UL26, UL50, and Meq, the main MDV oncoprotein. Both isoforms Meq and L-Meq derived from the oncogenic and the non-oncogenic vaccine strain, respectively, similarly inhibited the interferon response in a dose-dependent way, and Meq deletion mutants revealed that the Cterminal, proline-rich transactivating domain is not required for this inhibitory effect. In transient transfection experiments, Meq induced a dose-dependent proteasomal degradation of the chicken interferon regulatory factor 7 (chIRF7), which is required for chIFN-α- induced activation of ISRE. Over-expression of chIRF7 lead to a dosedependent degradation of Meq and its accumulation in the cytoplasm, suggesting that proteasomal degradation of both Meq and chIRF-7 is linked. Consistent with these findings, MDV deletion mutant lacking both copies of the Meq gene was more sensitive to chIFN-α treatment compared to wild-type virus.

Marek’s disease (MD) is a lymphoproliferative disease of chickens caused by an oncogenic herpesvirus, Marek’s disease virus (MDV). Despite the availability of MD vaccines, little is known about the underlying immunological mechanisms that mediate vaccine-induced immunity. The objective of this research was to elucidate these mechanisms. To characterize host responses in the lungs, chickens were vaccinated with herpesvirus of turkeys (HVT) and infected with MDV-RB1B. Vaccinated MDV-infected chickens had a higher accumulation of viral genome in the lungs, associated with T cell infiltration in lung tissue and an up-regulation of interferon (IFN) - and interleukin (IL) -10. This finding led us to conclude that IFN-γ has a role in immunity; hence, we further investigated the role of this cytokine. The hypothesis tested was that the protective efficacy of HVT against MDV-RB1B would be enhanced when combined with recombinant chicken IFN-γ (rChIFN-). Chicken IFN-γ coding sequence was cloned into an expression plasmid, and the bioactivity of rChIFN- was confirmed. Administration of this plasmid led to a significant reduction in tumour occurrence in HVT vaccinated MDV-infected chickens, suggesting enhanced vaccine-induced immunity. To shed more light on the relevance of IFN-γ to immunity against MD, studies were designed to down-regulate the expression of IFN-γ in chicken tissues. Three small interfering (si)RNAs specific for chicken IFN- were selected which significantly inhibited expression of IFN-γ by up to 80% in cultured cells. These three siRNAs and a non-target control were cloned and expressed as short hairpin RNA (shRNA) using an avian adeno-associated virus (rAAAV) vector system. An MDV challenge trial was conducted once shRNA expression by the rAAAV was confirmed in vitro. It was reasoned that down-regulation of IFN- would lead to abrogation of immunity conferred by HVT. There was an increase in the number of chickens with tumours that received HVT and rAAAV + IFN-γ shRNA compared to the control group, though not statistically significant. However, no difference in MDV genome load in feathers was detected between vaccinated MDV-infected birds with or without rAAAV. In summary, we have demonstrated here that cytokines are induced in the course of vaccination against Marek’s disease and that IFN-γ plays a role in vaccine-induced immunity against MD.

Thesis (Ph. D.) Veterinary Science --University of Wisconsin, 1970. / eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 51-57).

Thesis (M.S.)--University of Delaware, 2007. / Principal faculty advisor: Marlene G. Emara, Dept. of Animal & Food Sciences. Includes bibliographical references.

Call number: LD2668 .T4 1958 R63