Please read the abstract in the secton 00front of this document. / Thesis (PhD (Zoology))--University of Pretoria, 2007. / Zoology and Entomology / unrestricted

Newcastle disease (ND), caused by Newcastle disease virus, is an acute, contagious and pathogenic infection of pet, free living and domestic birds. ND is an epidemic disease and it is responsible for high economic losses due to up to 100 % mortality. The control of ND in the intensive commercial poultry farms is largely dependent on prophylactic immunisation using conventional vaccines. The ND V4 vaccine and its derivative ND V4-HR vaccine were selected originally for use in village chickens, due to their immunogenicity, thermostability, transmissibility and ease of administration. The efficacy of V4 and V4HR vaccines have been established in many Asian and African countries in their ability to challenge a wide range of recognised and local velogenic NDV. Therefore, ND V4 was tested for efficacy against B1172 challenge NDV isolated in south Africa in 1993. Twenty-eight one day-old replacement pullets were vaccinated by eye-drop route at 21 and 49 days old. Chickens vaccinated by eye-drop route were left to mingle with the unvaccinated in-contact chickens. At 63 days all chickens including the unvaccinated control group were individually challenged with B1172 NDV. Serological monitoring of NDV antibody response was done using HI and ELISA tests. The ND V4 vaccine induced full protection against B1172 NDV in chickens vaccinated by eye-drop vaccination and in 55 % of chickens vaccinated by the in-contact method. No association was seen between NDV antibody titer at pre-challenge and the ability to withstand B1172 challenge NDV. A fair to good agreement was seen between the HI and ELISA test in monitoring NDV antibody response during the experiment. Although, the ELISA showed a higher sensitivity and specificity than the HI test, further studies are required using this method of comparison. / Dissertation (MMed Vet (Poultry diseases))--University of Pretoria, 2002. / Production Animal Studies / unrestricted

Since 2002, the South African poultry industry has experienced outbreaks of Newcastle disease (ND) caused by a newly introduced virus (NDV) strain belonging to lineage 5d/VIId (“goose paramyxovirus” - GPMV). Control of the disease has proved difficult with commercially available vaccines appearing ineffective. In the first of two studies, broilers chicks were vaccinated with VG-GA vaccine (lineage II), then challenged with both GPMV and a “classic” challenge virus (RCV) of lineage 3d/VIII to compare the efficacy of the vaccine against both strains. In the second study, commercial and SPF hens in lay were vaccinated with La Sota vaccine and challenged with GPMV isolate, and immunohistochemistry staining used to determine the distribution pattern of viral antigen in the oviduct of the hens. The second study also compared the efficacy of cloacal and ocular routes of vaccination. The first study did not detect any statistically significant difference in protection offered by the vaccine against the GPMV strain in comparison to the RCV strain. The protection offered by the vaccine against challenge with both viruses was found to be dosedependant with 106.0 EID50 producing a 100% protection and 94.44% and 13.89% for 104.5 EID50 and 103.0 EID50 vaccination doses respectively. Protected birds did not manifest clinical signs, but still had macropathological lesions in some organs at necropsy. The computed protective doses (PD50 and PD90) for the VG-GA vaccine were 103.51 and 104.38 for GPMV and 103.79 and 104.43 for RCV. Results from the second study showed no clear difference in the protection of the oviduct from challenge with GPMV by either the cloacal and ocular routes of vaccination. Vaccinated birds were fully protected (100%) against challenge by La Sota vaccine, but not against infection and replication of the virus, as birds showed varying degrees of macropathology with numerous stained viral antigens in the oviducts demonstrated by immunohistochemistry. The susceptibility and colonisation of the oviduct of laying hens by both the lentogenic La Sota and the virulent NDV isolates was confirmed, with the uterus being more susceptible than magnum and isthmus. Necrosis and apoptosis of cells of the oviduct were not detected but cellular infiltration, gland dilatation and interstitial oedema were observed. / Dissertation (MSc (Veterinary Science))--University of Pretoria, 2009. / Production Animal Studies / unrestricted

The specific objectives of this study were to determine the Newcastle disease virus (NDV) antibody titres from the chicken sera collected from various districts and provinces of Zambia and to determine the seroprevalence of ND in Zambian backyard chickens. Results showed that 73.9 % of the birds sampled tested positive for Newcastle disease (ND) antibodies. The seroprevalence of Newcastle disease virus (NDV) in Zambian backyard chicken flocks varied among the five provinces sampled, ranging from 82.6 % in Eastern Province to 48.3 % in Luapula Province. The seroprevalence of the virus also varied among the 11 districts sampled, ranging from 91.3 % in Monze District of Southern Province to 22.8 % in Mufulira District of the Copperbelt Province. The results indicated that the seroprevalence of ND in Zambia has increased since the last survey conducted in 1994. The data generated is expected to contribute towards a more clear understanding of the epidemiology of NDV that would ultimately contribute towards an improved ND control programme to benefit all stakeholders in Zambia. An improved ND control programme is expected to enhance flock numbers and ultimately improve the dietary requirements and income needs of many poor households in the country. / Dissertation (MSc)--University of Pretoria, 2012. / Veterinary Tropical Diseases / unrestricted

Although Newcastle disease is reported to be endemic in Nigeria, little information exists on the molecular epidemiology and the lineage distribution of the Newcastle disease viruses (NDVs) in the country, especially in the live bird markets. Recent studies reported the identification of three distinct sub-lineages namely; 5f, 5g and 5h in West Africa, particularly sub-lineages 5f and 5g were identified in Nigeria. In this study a total of 41 NDV isolates were analysed. Thirty six NDVs were recovered from a variety of poultry species from live bird markets in the six geo-political zones of Nigeria during active surveillance from 2007 to 2008. Five NDVs recovered from outbreaks in backyard and commercial poultry farms within the same period were also genetically characterized. A commonly used region of the virus genome that spans nucleotide 61 to nucleotide 374 of the Fusion protein, including the cleavage site was targeted. Based on sequence analysis, 39 of the isolates were classified as virulent. Of these, 20 were classified as sub-lineage 5g and 17 were classified as sub-lineage 5f. One isolate differ markedly from all other strains included in the phylogeny. Interestingly, 13 strains from the 5f group formed a distinct cluster that was not identified by other groups in similar studies. Phylogenetic analysis, amino acid sequence determination of the F0 cleavage site sequence analysis, pair wise distance analysis of the partial fusion protein gene sequences and Geographic Information System (GIS) mapping was done. Results showed close genetic similarities and provided evidence for the first time of the epidemiological link between the viruses circulating in the LBMs and those identified in outbreaks in backyard and commercial farms in Nigeria between 2007 and 2008. The emergence and identification of new sub-lineages gives an insight in to the high rate of genetic drift occurring in NDV strains in Nigeria, and raises concerns about the efficacy of current NDV control measures in the country. Thus there is need for continuous surveillance and characterization of NDV from Nigeria to monitor the emergence of new lineages and sub-lineages in the Nigerian poultry industry. Copyright / Dissertation (MSc)--University of Pretoria, 2011. / Production Animal Studies / unrestricted

Avian paramyxoviruses (APMVs) consist of twelve distinct serotypes (APMV-1 to -12) isolated from a wide variety of domestic and wild birds. APMV-1/Newcastle disease virus (NDV) is the most characterized and globally important avian pathogen, because of the huge economic loss associated with the disease. However, very little information is known about the pathogenicity of APMV 2-12. APMV expresses six structural and two accessory proteins. The functions of APMV accessory proteins (V and W) are not fully established. Only the function of V protein in NDV is studied so far. V protein was found to be an IFN antgonist and a major virulent determinant of NDV. In this study, we tested for the potential functions of W protein in NDV and fuctions of V protein in other APMV serotypes. Vaccination failure is a major cause for NDV outbreak in developing and tropical countries, because of thermolabile nature of vaccine strains. Thermostable and thermolabile NDV strains exhibit difference in W protein length. In the first part of our study, we mutated the genome of a thermolabile NDV strain to express W protein of different lengths, rescued recombinant viruses by reverse genetics system and tested for thermostability. Our results showed that W protein does not confer thermostability to NDV. In the second part of study, we constructed plasmids expressing APMV -2, -3 and -6V proteins and tested for IFN antagonism by a dual luciferase reporter assay. Our results showed that APMV-3V acts as IFN antagonist by blocking IFN induction and thereby may play an important role in the evasion of innate immunity. / Master of Science

Many oncolytic virotherapies have shown great advantages for rapid, rational design through recombinant DNA technology to facilitate the targeting of a broad spectrum of malignancies. Newcastle disease virus (NDV), an avian paramyxovirus, is naturally tumor-selective and inherently oncolytic. Our approach is to develop NDV-based nanoparticles (VBNP) for oncolytic virotherapy. VBNPs are non-infectious and non-replicating and are relatively safe. We obtained VBNPs by co-expressing matrix (M), hemagglutinin (HN), and fusion (F) proteins of NDV in avian/ mammalian cells. The budding characteristics, size and morphology of VBNPs were similar to authentic virions. As a proof of concept, we engineered the apoptin (VP3) gene of chicken anemia virus in VBNPs and specifically targeted them to folate-receptor bearing tumor cells by surface conjugation to folate. The VBNPs killed tumor cells by apoptosis and induced proinflammatory and chemotactic cytokines. The VBNPs, although not curative, were able to limit the progression of xenotransplanted fibrosarcoma and malignant glioma tumors and provided a survival advantage in nude mice. We also engineered NDV M based particles with nipah virus surface glycorporteins to target ephrin B receptors. NDV based nipah Virus BNPs (NiV-ndBNP) were morphologically similar to authentic NiV virions. NiV glycoproteins were incorporated into the NDV M based particles, despite poor sequence homology in the transmembrane domain and cytoplasmic tails of glycoproteins. Our results suggest that VBNPs could be used to deliver small molecules, tumor antigens, anti-tumor/ reporter genes and also aid in generating tumor specific immunity by rational design. / Master of Science

Chickens from lines selected for high (HA) and low (LA) antibody response to sheep red blood cell (SRBC) antigen were Used to study the role of genetic factors involved in resistance to Eimeria tenella and Newcastle disease virus (NDV). Routes of administration were intravenous for SRBC antigen and intranasal for NDV. Chicks were exposed to E. tenella either through a natural challenge via the litter or a controlled oral administration. Differences between lines were observed in resistance to cecal coccidiosis, with line HA chicks being more resistant than those from line LA. Results were similar. for both natural and controlled exposures, Differences in resistance to E. tenella were found among alleles for the I alloystern with degree of additivity from the I⁴ allele on resistance depending on the background genome. Chicks from line HA exhibited higher antibody titers to SRBC and lower titers to NDV than did those from line LA. This pattern was the same regardless of whether antigens were given together or alone. Correlations within lines for birds which received both antigens were positive and significant in line HA and not different from zero in line LA. When NDV was given at the time birds received a booster dose of SRBC, antibody titers for NDV and for primary and booster SRBC were higher in. line HA than line LA. Correlations between all titers were positive and significant in both lines. The data suggest that relationships between titers for SRBC and NDV are influenced by both genetic and nongenetic factors. / M.S.

O uso de antibióticos promotores de crescimento de forma indiscriminada vem causando enorme preocupação a cada ano por causa de uma possível resistência cruzada. Essa é uma das razões pela quais pesquisas vêm sendo desenvolvidas para substituição destes agentes por produtos naturais e/ou alternativos. Desde 1973 o Conceito de Exclusão Competitiva (CE) vem sendo estudado e usado na avicultura mundial. Com esse objetivo, inúmeras pesquisas relacionadas com produtos de Exclusão Competitiva vêm demonstrando, ao passar dos anos, não só a eficiência desse método no controle de patógenos indesejáveis como Salmonella ssp, como também sua eficácia em desempenho zootécnico. Dessa forma, o presente estudo teve como objetivo verificar a influência de um produto de Exclusão Competitiva sobre os índices zootécnicos e parâmetros de imunidade de frangos de corte, tendo um antibiótico e um controle para comparação. A revisão bibliográfica também apresenta pesquisas sobre o controle de Salmonella spp com o uso da CE. Foram utilizados 420 pintos de corte, criados até 42 dias de idade em gaiolas. O delineamento utilizado foi o inteiramente casualizado, com 3 tratamentos: Dieta basal (DB) com Antibiótico; DB com produto de Exclusão Competitiva, e DB sem Antibiótico (controle); sendo 20 repetições/tratamento. Quanto ao desempenho zootécnico e, considerando-se o período total de criação, o produto de Exclusão Competitiva testado demonstrou efeito benéfico para os valores de ganho médio de peso (GMP) e peso final (PF), que foram melhores ou similares comparados ao antibiótico principalmente até a fase final de criação. Já os resultados de conversão alimentar e mortalidade mostraram-se melhores com o uso da CE quando foram comprados com os demais tratamentos. Com relação à imunidade, a Exclusão Competitiva não apresentou efeito sobre o parâmetro avaliado neste trabalho. Diante dos resultados aqui obtidos, conclui-se que os ensaios com aditivos alternativos devem continuar, já que houve evidências de seus possíveis efeitos benéficos na produção animal. / The use of growth-promoting antibiotics inappropriately has been addressing concerns every year for possible cross-resistance. This is one of the reasons why studies have been carried through to replace such agents by natural and/or alternative products. Since 1973, the concept of Competitive Exclusion (CE) has been studied and used in poultry raising worldwide. Aiming at reaching the said objective, several studies on Competitive Exclusion products have been showing, over the years, not only how effective the method is in controlling undesirable pathogens, such as Salmonella ssp, but also its efficacy for animal performance. Thus, the present paper aims at verifying the influence of a Competitive Exclusion product on zoological properties and immunity parameters for broilers, having one antibiotic group and one control group for comparison. In addition, the bibliography includes studies on the control of Salmonella ssp when CE is used. A total of 420 broilers, reared in cages for maximum 42 days of age, have been used. The study design consisted in a randomized study, with three treatment groups: Baseline diet (DB) and antibiotic; DB and Competitive Exclusion, DB without Antibiotic (control); with 20 repetitions/treatment. With regard to animal performance and, considering total breeding period, the investigational Competitive Exclusion product has showed beneficial effect on average weight gain (GMP) and final weight (PF) values, which have been found to be improved or similar to the antibiotic group, specially when used until the final breeding stage. In turn, the results for feed conversion and mortality have showed to be improved when CE was used in comparison to the other two treatments. In relation to immunity, Competitive Exclusion has not affected the immunity parameter assessed in this paper. According to the results found in this study, it can be concluded that the use of alternative additives should be further studied, as evidences of their possible beneficial effects on animal raising have been found.

Uma série de fatores como ambiente, nutrição e doenças podem ser consideradas como estressores em sistemas de produção. Frangos de corte expostos a estressores por períodos prolongados de tempo apresentam de forma geral, redução do ganho de peso e consumo de ração, aumento da conversão alimentar, aumento da média de mortalidade e predisposição a doenças. A vacinação é uma prática essencial para um adequado manejo da produção e o desenvolvimento da imunidade do frango de corte frente a desafios bacterianos e virais presentes no ambiente. O entendimento dos fatores que podem interferir com o sucesso da vacinação é essencial para a otimização da saúde e do bem-estar animal, permitindo a utilização de todo o seu potencial genético e nutricional. Poucos estudos têm enfatizado o efeito do estresse por calor na imunidade celular e humoral de frangos de corte. Este trabalho tem como objetivo estudar o efeito do estresse por calor sobre a imunidade celular (linfócitos B e T), humoral sérica (IgM e IgY) e o peso relativo de órgãos (baço, bursa e fígado) de frangos de corte submetidos a um protocolo de vacinação para a Doença de Newcastle (cepa LaSota). Para tanto, 96 frangos de corte Cobb machos foram aleatoriamente divididos em quarto grupos: grupo 1. Frangos de corte não vacinados expostos à temperatura termoneutra; grupo 2. Frangos de corte vacinados expostos à temperatura termoneutra; grupo 3. Frangos de corte não vacinados expostos ao protocolo de estresse térmico por calor (38±2°C); e grupo 4. Frangos de corte vacinados expostos ao protocolo de estresse térmico por calor (38±2°C). Todas as aves foram alojadas em isoladores com água e ração ad libitum. Foi utilizada uma vacina com o vírus atenuado para Doença de Newcastle (cepa LaSota) administrada em duas doses, aos 7 e aos 14 dias de vida. Os frangos submetidos ao protocolo de estresse foram expostos a uma temperatura de (38±2°C) do segundo até o sexto dia de vida. Os dados obtidos demonstraram que o estresse por calor de forma isolada reduziu o peso relativo do fígado e aumenta aquela do baço e da bursa e induziu uma alteração significativa do perfil de células imunes no sangue periférico das aves, como consequência, observou-se alteração no padrão de imunoglobulinas, o que influenciou diretamente a resposta da ave frente ao desafio ambiental (quando o estresse foi avaliado isoladamente) ou vacinal (quando o estresse foi avaliado juntamente ao desafio vacinal com o vírus da Doença de Newcastle). Dessa forma, sugerimos que o estresse por calor diminuiu a eficácia da resposta vacinal aos frangos de corte, em função de mudanças que induziu no perfil de linfócitos e produção de imunoglobulinas. / A number of factors, such as environment, nutritional status and diseases are stressful for animals during livestock production. It has been shown that broilers exposed to stressors for prolonged periods tend to present decreased weight gain and feed intake and to have an increased feed conversion ratio, increase mortality rate and predisposition to diseases caused by secondary agents. Vaccination is an essential practice for adequate management of the livestock and protection of poultry against bacterial and viral infectious agents. Understanding the factors that modify the success vaccination is essential for optimizing animal health and welfare, thus allowing them to use of all the genetic and nutritional potential they have. Few studies have emphasized the effects of heat stress on cellular and humoral broilers immunity. This study aimed at discussing the effects of heat stress on cellular immunity (B and T lymphocytes) and humoral immunity (IgM and IgY) of broilers undergoing a vaccination protocol for Newcastle disease (LaSota strain). For this purpose, 96 male broilers (Cobb) were random divided into four groups: 1. Unvaccinated broiler chickens exposed to thermoneutral temperature; 2. Vaccinated broiler chickens exposed to thermoneutral temperature; 3. Unvaccinated broiler chickens exposed to a heat stress protocol (38±2°C). 4. Vaccinated broiler chickens exposed to a heat stress protocol (38±2°C). All broilers were housed in isolators and provided water ad lib. We used a live Newcastle vaccine disease virus of LaSota strain given in two doses at 7 and 14 days. The broiler chickens were exposed to heat stress (38±2°C) from the 2nd to 6th day of life. It has been observed that when combined with the vaccination challenge, heat stress was able to change immune cells profile from a B to T cytotoxic and T helper immune cells, keeping this change pattern until the end of the study period (over 19 days). This change in the cellular response pattern modified the ability of the broiler chickens´ immune system to react while challenged by vaccination, reducing its efficiency and making them more susceptible to the disease agent. The data obtained suggest that heat stress by itself induced considerable modifications in the liver, spleen and bursa de Fabricius and induced a significant modification in the profile of immune cells in peripheral blood of birds, with responsive modifications in the pattern of immunoglobulins, directly influencing the response of the broilers against the environmental challenge (when the stress is evaluated separately) or the vaccination (when the stress is measured along the challenge vaccination with Newcastle Disease virus). Therefore, we suggest that the heat stress was capable of reduce the efficacy of the vaccinal response in broiler chickens, related to the modification of the lymphocytes and the production of immunoglobulins.