Ruminant alphaherpesviruses related to bovine herpesvirus 1 (BoHV-1) are a cluster of viruses antigenically and genetically closely related. The prototype of this cluster, BoHV-1, is a major pathogen of cattle associated with various clinical manifestations including infectious bovine rhinotracheitis (IBR) and infectious pustular vulvovaginitis (IPV). IBR is a disease of major economic concern in many parts of the world and especially in Europe, both in countries where this infection has been eradicated and in those where the control of IBR is currently or will be undertaken. The massive use of vaccination allowed a significant reduction of the number of IBR clinical cases. However, the existence of closely related viruses to BoHV-1 is a threat for IBR eradication programmes. Consequently, the main objective of the present work is dedicated to afford a better knowledge of the interaction between alphaherpesviruses and their ruminant hosts in order to contribute to improve the control of IBR. To meet the objective, two approaches have been developed: the study of the viral diversity aiming to extend both epidemiological and virological data about ruminant alphaherpesviruses related to BoHV-1 and the study of the heterologous protection aiming to protect minor ruminant species by the concept of the cascade vaccination. Illustrating the problematic of the cluster of ruminant alphaherpesviruses related to BoHV-1, an original situation has been described recently in Belgium. During 2001 and 2002 hunting seasons, 28.9% of red deer were detected seropositive to BoHV-1. Due to an apparent lack of contact between cattle and red deer, it was suggested that a BoHV-1 related virus was spreading in the Belgian red deer population. Thus, the first isolation of cervid herpesvirus 1 (CvHV-1) in wild fauna is reported, which brings the opportunity to deeper analyse the antigenic, genomic and genetic relationship between BoHV-1 and its related ruminant alphaherpesviruses. This isolation demonstrates that a ruminant can be strongly identified as BoHV-1 positive while in actual fact it is infected with a related but distinct alphaherpesvirus and this ruminant will be declared as false positive. The problem is even greater when these viruses become latent allowing their possible reactivation and persistence for a very long time in their ecological niches. It is necessary to have tests which can differentiate related alphaherpesviruses that infect different ruminant species. The control of IBR relies on the use of BoHV-1 gB and gE blocking enzyme linked immunosorbent assays (ELISA) in order to differentiate infected and gE-negative vaccinated animals. Knowing that CpHV-1 is the most distant virus from BoHV-1, it can be hypothesised that a BoHV-1 gB blocking ELISA detects CpHV-1 but that CpHV-1 infection could be discriminated by a BoHV-1 gE blocking ELISA. CpHV-1 being mainly distributed in the Mediterranean part of Europe as Greece, Spain and Italy, the analysis was performed with field serums collected in France with the aim to update the epidemiological situation of the infection in Europe. Besides BoHV-1, CpHV-1 is the most relevant infection in Europe but is sadly neglected. The first reason is that economic losses are restricted to a herd level in contrast with IBR that brings an economical impact at a country level. The second reason is that goat is considered as a minor species. In this context, the problem is still not big enough for commercial interest towards vaccine development. The European Union has recently pointed out the problem of minor uses and minor species and allowed off label use of veterinary medicinal products or the use of a product licensed for a major species when an authorised veterinary medicinal product is not available (cascade principle). Goat being a minor species and CpHV-1 sharing close antigenic and genetic properties with BoHV-1, a live attenuated gE-negative BoHV-1 vaccine has been assessed in goats to protect against either a nasal or a genital CpHV-1 infection.

Resumo: Nove fêmeas bovinas canuladas no rúmen foram utilizadas para avaliar um preparado de anticorpos policlonais (PAP) de origem aviária contra as bactérias ruminais Streptococcus bovis, Fusobacterium necrophorum, Clostridium aminophilum, Peptostreptococcus anacrobius e Clostridium sticklandii. O delineamento experimental foi o quadrado latino 3 X 3 replicado 3 vezes, em arranjo fatorial de tratamentos 3 X 3 referente a 2 modificadores ruminais, monensina (MON) e PAP mais o grupo controle (CON) e 3 fontes energéticas, milho seco moído (MSM), silagem de grão úmido de milho (SGUM) e polpa cítrica (PC). Cada subperíodo experimental foi composto de 21 dias, onde a coleta de líquido ruminal foi realizada às 0, 2, 4, 6, 8, 10 e 12 h após a alimentação no último dia do período. Foram avaliados o consumo de matéria seca e os parâmetros de fermentação ruminal pH, concentração total de ácidos graxos de cadeia curta (AGCCt), lactato total e nitrogênio amoniacal (N-NH3). O PAP mostrou-se tão eficaz como a monensina em elevar o pH ruminal de bovinos alimentados com dietas ricas em concentrados, às 4 h após a alimentação, sem contudo, alterar (P>0,05) o consumo de matéria seca, a concentração total de ácidos graxos de cadeia curta, a proporção molar de acetato ou as concentrações de lactato total e nitrogênio amoniacal. Foi observada diminuição (P<0,05) na proporção molar de propionato com a utilização do PAP em relação a monensina, porém não em relação ao controle. / Abstract: Nine ruminally cannulated cows were used to test an avianderived polyclonal antibody preparation (PAP) against the specific ruminal bacterias Streptococcus bovis, Fusobacterium necrophorum, Clostridium aminophilum, Peptostreptococcus anacrobius and Clostridium sticklandii. The experimental design was a 3 X 3 latin square replicated 3 times with a factorial arrangement of treatments 3 X 3 regarding 2 rumen modifiers (monensin (MON) and PAP) plus control group (CON) and 3 energetic sources. The energetic sources utilized were dry-grounded corn grain (CG), high moisture corn silage (HMCS) and citrus pulp (CP). Each trial had 21 days; the ruminal fluid sampling was carried out on the last day of the period at 0, 2, 4, 6, 8, 10 and 12 h after morning meal. Dry matter intake and rumen fermentation parameters such as pH, total short chain fatty acids (tSCFA), lactic acid and ammoniacal nitrogen (NH3-N) concentration were analyzed in this trial. PAP was as effective as monensin in increasing ruminal pH of cattle fed high concentrate diets, 4 h after feeding, without interfering (P>0.05) on dry matter intake, total short chain fatty acids concentration, molar proportion of acetate, lactic acid and ammoniacal nitrogen (NH3-N) concentration. A decrease (P<0.05) in molar proportion of propionate was observed with PAP utilization compared to monensin but not in relation to control. / Orientador: Mário de Beni Arrigoni / Coorientador: Paulo Henrique Mazza Rodrigues / Banca: Rafael da Costa Cervieri / Banca: Renata Helena Branco Arnandes / Banca: Flávio Augusto Portela Santos / Banca: Telma Teresinha Berchiell / Doutor

Biological functions that use amino acids (AA) are limited by AA supply. This concept was likened to staves in a barrel, where the shortest stave determines the barrel's ability to hold water (Mitchell and Block, 1946). Inaccuracies in models that predict nutrient supply and requirements of dairy cows result in inefficient feeding, as under-prediction of requirements results in deficiency, and over-prediction results in excess. To avoid limitations in production due to AA deficiencies, protein is fed in quantities that likely exceed requirements. Overfeeding of AA results in increased expenses for producers and increased N excretion to the environment, providing economic and environmental incentives to increase N-efficiency. Work presented in the following chapters evaluated the impact of AA supplementation on milk production in dairy cattle, and evaluated the PREP10 and NRC (2001) nutrient requirement model predictions. In two feeding trials (Chapter 2 and Chapter 3), low protein diets did not result in decreased milk production, indicating that protein requirements were overestimated. Although supplementation of AA did not increase milk production, low protein diets resulted in greater N-efficiency, especially when supplemented with Histidine. Evaluation of the PREP10 and NRC (2001) models (Chapter 4) used production data from the literature to identify deficiencies in prediction equations, and found that correction of model bias would considerably reduce prediction errors. Model inaccuracies affect the inefficiency of dairy cow feeding, and must be evaluated to improve feed efficiency. Such improvements could act synergistically with AA supplementation to more closely match nutrient supply to requirements. / Master of Science / Biological functions that use amino acids (AA) are limited by AA supply. This concept was likened to staves in a barrel, where the shortest stave determines the barrel’s ability to hold water (Mitchell and Block, 1946). Inaccuracies in models that predict nutrient supply and requirements of dairy cows result in inefficient feeding, as under-prediction of requirements results in deficiency, and over-prediction results in excess. To avoid limitations in production due to AA deficiencies, protein is fed in quantities that likely exceed requirements. Overfeeding of AA results in increased expenses for producers and increased N excretion to the environment, providing economic and environmental incentives to increase N-efficiency. Work presented in the following chapters evaluated the impact of AA supplementation on milk production in dairy cattle, and evaluated the PREP10 and NRC (2001) nutrient requirement model predictions. In two feeding trials (Chapter 2 and Chapter 3), low protein diets did not result in decreased milk production, indicating that protein requirements were overestimated. Although supplementation of AA did not increase milk production, low protein diets resulted in greater N-efficiency, especially when supplemented with Histidine. Evaluation of the PREP10 and NRC (2001) models (Chapter 4) used production data from the literature to identify deficiencies in prediction equations, and found that correction of model bias would considerably reduce prediction errors. Model inaccuracies affect the inefficiency of dairy cow feeding, and must be evaluated to improve feed efficiency. Such improvements could act synergistically with AA supplementation to more closely match nutrient supply to requirements.

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Master of Science / Department of Animal Sciences and Industry / Barry J. Bradford / Two experiments were conducted to explore potential cellular and biochemical mechanisms by which zinc may impact milk production and mammary health of dairy cattle. For the first experiment, twelve lactating Holstein cows were enrolled in a replicated 3 x 3 Latin square, and provided a balanced diet supplemented with either 30 ppm zinc as ZnSO₄ (30-ZS), 60 ppm zinc as ZnSO₄ (60-ZS), or 60 ppm zinc as a mixture of ZnSO₄ and Zn methionine (60-ZM). Blood and milk were collected and analyzed for markers of blood-milk leak and dairy food properties of milk. Milk cells were also isolated and analyzed for RNA abundance of genes related to zinc-mediated apoptosis. Aside from a tendency for 60-ZS to increase feed intake and 60-ZM to increase milk fat compared to 30-ZS, no effects were observed on performance, markers of blood-milk leak, transcript abundance of selected genes, or dairy food properties of milk. A cell culture study was also conducted using immortalized bovine mammary epithelial (MAC-T) cells. Messenger RNA abundance of GPR39—a G protein-coupled receptor shown to regulate cellular activity in response to extracellular zinc concentrations in other epithelial cell lines—was knocked down using RNA interference. Following GPR39 knockdown, cells were treated with 0 or 100 μM zinc. Phosphorylation of the kinases ERK1/2 and AKT was measured in GPR39-knockdown and control cells treated with either 0 or 100 μM zinc. No effect of zinc treatment or GPR39 knockdown was detected on kinase phosphorylation. Interestingly, the 100 μM zinc treatment showed a tendency to increase GPR39 mRNA abundance in control cells. There remain many questions about the cellular mechanisms whereby zinc can impact milk production in dairy cattle.

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Made available in DSpace on 2014-06-11T19:32:58Z (GMT). No. of bitstreams: 0 Previous issue date: 2008-06-11Bitstream added on 2014-06-13T20:44:27Z : No. of bitstreams: 1 marino_ct_dr_botfmvz.pdf: 353504 bytes, checksum: b342669cd7175c9e70b7faf645d74384 (MD5) / Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) / Universidade Estadual Paulista (UNESP) / Nove fêmeas bovinas canuladas no rúmen foram utilizadas para avaliar um preparado de anticorpos policlonais (PAP) de origem aviária contra as bactérias ruminais Streptococcus bovis, Fusobacterium necrophorum, Clostridium aminophilum, Peptostreptococcus anacrobius e Clostridium sticklandii. O delineamento experimental foi o quadrado latino 3 X 3 replicado 3 vezes, em arranjo fatorial de tratamentos 3 X 3 referente a 2 modificadores ruminais, monensina (MON) e PAP mais o grupo controle (CON) e 3 fontes energéticas, milho seco moído (MSM), silagem de grão úmido de milho (SGUM) e polpa cítrica (PC). Cada subperíodo experimental foi composto de 21 dias, onde a coleta de líquido ruminal foi realizada às 0, 2, 4, 6, 8, 10 e 12 h após a alimentação no último dia do período. Foram avaliados o consumo de matéria seca e os parâmetros de fermentação ruminal pH, concentração total de ácidos graxos de cadeia curta (AGCCt), lactato total e nitrogênio amoniacal (N-NH3). O PAP mostrou-se tão eficaz como a monensina em elevar o pH ruminal de bovinos alimentados com dietas ricas em concentrados, às 4 h após a alimentação, sem contudo, alterar (P>0,05) o consumo de matéria seca, a concentração total de ácidos graxos de cadeia curta, a proporção molar de acetato ou as concentrações de lactato total e nitrogênio amoniacal. Foi observada diminuição (P<0,05) na proporção molar de propionato com a utilização do PAP em relação a monensina, porém não em relação ao controle. / Nine ruminally cannulated cows were used to test an avianderived polyclonal antibody preparation (PAP) against the specific ruminal bacterias Streptococcus bovis, Fusobacterium necrophorum, Clostridium aminophilum, Peptostreptococcus anacrobius and Clostridium sticklandii. The experimental design was a 3 X 3 latin square replicated 3 times with a factorial arrangement of treatments 3 X 3 regarding 2 rumen modifiers (monensin (MON) and PAP) plus control group (CON) and 3 energetic sources. The energetic sources utilized were dry-grounded corn grain (CG), high moisture corn silage (HMCS) and citrus pulp (CP). Each trial had 21 days; the ruminal fluid sampling was carried out on the last day of the period at 0, 2, 4, 6, 8, 10 and 12 h after morning meal. Dry matter intake and rumen fermentation parameters such as pH, total short chain fatty acids (tSCFA), lactic acid and ammoniacal nitrogen (NH3-N) concentration were analyzed in this trial. PAP was as effective as monensin in increasing ruminal pH of cattle fed high concentrate diets, 4 h after feeding, without interfering (P>0.05) on dry matter intake, total short chain fatty acids concentration, molar proportion of acetate, lactic acid and ammoniacal nitrogen (NH3-N) concentration. A decrease (P<0.05) in molar proportion of propionate was observed with PAP utilization compared to monensin but not in relation to control.