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A distribuição de anticorpos neutralizantes para o herpesvirus bovino 1 (HVB 1) foi estudada em quatro faixas etárias de bovinos, em 21 rebanhos de leite e de corte. Os resultados da sorologia foram analisados e relacionados com as respostas de questionários aplicados aos responsáveis pelos rebanhos. As taxas de freqüência de anticorpos neutralizantes para o HVB 1 foram comparadas segundo a aptidão e a faixa etária. Fatores como tipo de manejo e idade dos animais influenciaram na distribuição de anticorpos para o HVB 1. The distribution of neutralizing antibodies to bovine herpesvirus 1 (BHV 1) was studied in 21 beef and dairy cattle herds, in four different age groups. The results were analyzed and related with answers given to a questionnaire proposed to the farmers. The frequency rates of neutralizing antibodies for the BHV 1 were compared according to the dairy or beef herds and age of the animals. The animal husbandry and age influenced the distribution of neutralizing antibodies to this virus.

A distribuição de anticorpos neutralizantes para o herpesvirus bovino 1 (HVB 1) foi estudada em quatro faixas etárias de bovinos, em 21 rebanhos de leite e de corte. Os resultados da sorologia foram analisados e relacionados com as respostas de questionários aplicados aos responsáveis pelos rebanhos. As taxas de freqüência de anticorpos neutralizantes para o HVB 1 foram comparadas segundo a aptidão e a faixa etária. Fatores como tipo de manejo e idade dos animais influenciaram na distribuição de anticorpos para o HVB 1.The distribution of neutralizing antibodies to bovine herpesvirus 1 (BHV 1) was studied in 21 beef and dairy cattle herds, in four different age groups. The results were analyzed and related with answers given to a questionnaire proposed to the farmers. The frequency rates of neutralizing antibodies for the BHV 1 were compared according to the dairy or beef herds and age of the animals. The animal husbandry and age influenced the distribution of neutralizing antibodies to this virus.

Bovine abortions cause major financial loss to the farming industry of Ireland each year. The aim of this thesis is to evaluate the prevalence and trends of common infectious bovine abortifacients in Ireland in 2013. It will also examine in detail the aetiology of the identified abortifacients, the pathophysiology and in some cases the available preventative and curative measures which can be taken. This thesis has used quantitative data extracted from yearly reports compiled by the Department of Agriculture, Fisheries and the Marine and the Agri-Food Biosciences Institute of Ireland. The findings from this thesis demonstrate that while there were many common causes of abortion in cattle in Ireland in 2013 such as Salmonella dublin and Neospora caninum, the most common cause in 2013 was Trueperella pyogenes, detected in 208 out of the 3517 foetuses or foetal samples submitted.

There was almost a 20% decrease in the amount of positive cases of BVD being reported in new-born animals from 2013 to 2014. The number of positive BVD herds also decreased but herds with one or more PI’s increased. This indicates that farmers are not implanting the removal of PI’s firmly enough. The policing of this programme is not strict enough and with the absence of penalties, Ireland will struggle to gain a BVD free status. IBR is seasonal and spreads during the winter months when animals are being housed. High frequency of herds exposed are to BoHV-1 in Ireland. IBR primarily causes respiratory infections in cattle, which then may lead to abortion. It is difficult to diagnose if IBR is responsible for this abortion. Therefore, the number of abortions due to IBR may be higher than previously documented. However, BoHV-1 remains the leading cause of respiratory infections in cattle in Ireland.

El herpesvirus bovino-1 (BHV-1) es un virus de genoma DNA perteneciente a la familia Herpesviridae, el cual afecta al bovino en el que provoca un amplio espectro de manifestaciones clínicas y pérdidas económicas. El principal componente inmunogénico de su envoltura es la glicoproteína D (gD), la cual ha sido caracterizada y utilizada como inmunógeno en distintos sistemas de expresión. El objetivo de este trabajo fue generar un poxvirus recombinante (Raccoonpox [RCN]) que expresara una versión truncada de la gD del BHV-1 para ser usado como inmunógeno. Para ello, se amplificó el gen que codifica para la versión truncada de la gD, la cual se clonó en el plásmido de transferencia pTK/ IRES/tpa que posee sitios de homología a la timidinakinasa del poxvirus, un sitio interno de entrada al ribosoma (IRES) y una señal secretoria (tPA), generando el constructo pTK/gD/IRES/tpa. Para generar el RCN recombinante, se tomaron células BSC-1, se infectaron con una cepa Silvestre del RCN (CDC/V71-I-85A) a un índice de multiplicidad de infección de 0,05 y se transfectaron con el constructo pTK/gD/IRES/tpa; generándose diferentes poblaciones virales con y sin el gen de interés. Para seleccionar los virus recombinantes que expresaban el gen de interés, se realizó una selección de recombinantes negativos para timidina kinasa y positivos para la gD por tres rondas de purificación de placas en monocapas de células RAT-2 las cuales son mutantes para timidina kinasa y en presencia de bromodeoxiuridina. Los virus recombinantes se confirmaron por PCR y secuenciación de nucleótidos y se denominaron RCN-gD. Bovine Herpesvirus-1 is a DNA virus belonging to the family Herpesviridae, which affects cattle, causing a wide spectrum of clinical manifestations and economic losses. The main immunogenic component is its envelope glycoprotein D (gD), which has been characterized and used as immunogen in different expression systems. The aim of this work was to generate a recombinant poxvirus (Raccoonpox [RCN]) expressing a truncated version of BHV-1 gD to be used as a vaccine. To do this, it was amplified the gene for a truncated version of gD which subsequently was cloned in transfer plasmid PTK/IRES/tpa which has homology to sites of poxvirus thymidine kinase, an internal site of ribosome entry (IRES) and a secretory signal (tPA), generating the construct PTK/gD/IRES/tpa. To generate the recombinant RCN, we took BSC-1 cells and we infected with a wild type RCN (CDC/V71-I-85A) at a multiplicity of infection of 0.05, then cells were transfected with the construct PTK/gD/IRES/tpa, generating different viral populations with and without the gene of interest. To select recombinant viruses expressing the gene of interest, we performed a selection of recombinant thymidine kinase negative and positive for gD by three rounds of plaque purification on RAT-2 cells monolayers which are thymidine kinase null and using bromodeoxyuridine. Recombinant viruses were recovered and confirmed by PCR and nucleotide sequencing and so called RCN-gD.

Objective—To assess long-term effects and risk factors for the efficacy of hyperimmunization protocols against infectious bovine rhinotracheitis (IBR) during a longitudinal field study of dairy and dairy-beef mixed farms. Animals—Approximately 7,700 cows from 72 farms. Procedures—Farms were assigned to 3 treatment groups (hyperimmunization groups [HIGs] 1 and 2, which were hyperimmunized with glycoprotein E [gE]–deleted marker vaccines, and a nonintervention group [NIG]). Cattle in HIG 1 were initially vaccinated with an attenuated vaccine, whereas cattle in HIG 2 were initially vaccinated with an inactivated-virus vaccine. Cattle in both HIGs received booster inoculations with inactivated-virus vaccines at 6-month intervals. The risk for gE seroconversion was compared among experimental groups via a shared frailty model with a piecewise constant baseline risk to correct for seasonal and secular effects. Results—Risk for gE seroconversion significantly decreased over time for the HIGs, compared with the NIG. Seasonal changes in the risk of gE seroconversion were detected, with a higher risk during winter periods, compared with grazing periods. No significant difference was detected between HIGs 1 and 2. The only significant risk factor was the number of buildings for cattle on a farm; the higher the number of buildings, the lower the risk for gE seroconversion. Prevalence of IBR decreased over time in both HIGs but remained constant or increased in the NIG. Conclusions and Clinical Relevance—Hyperimmunization via repeated administration of attenuated and inactivated-virus gE-deleted marker vaccines as well as inactivated-virus vaccines may provide a method for control of IBR.

RESUMEN: El herpesvirus bovino-1 (BHV-1) es un virus de genoma DNA perteneciente a la familia Herpesviridae, el cual afecta al bovino en el que provoca un amplio espectro de mani festaciones clínicas y pérdidas económicas. El principal componente inmunogénico de su envoltura es la glicoproteína D (gD), la cual ha sido caracterizada y utilizada como inmunógeno en distintossistemas de expresión. El objetivo de este trabajo fue generar un poxvirusrecombinante (Raccoonpox[RCN]) que expresara una versión truncada de la gD del BHV-1 para ser usado como inmunógeno. Para ello, se amplificó el gen que codifica para la versión truncada de la gD, la cual se clonó en el plásmido de transferencia pTK/ IRES/tpa que posee sitios de homología a la timidinakinasa del poxvirus, un sitio interno de entrada al ribosoma (IRES) y una señal secretoria (tPA), generando el constructo pTK/gD/IRES/tpa. Para generar el RCN recombinante, se tomaron células BSC-1, se in fectaron con una cepa Silvestre del RCN (CDC/V71-I-85A) a un índice de multiplicidad de infección de 0,05 y se transfectaron con el constructo pTK/gD/IRES/tpa; generándose diferentes poblaciones virales con y sin el gen de interés. Para seleccionar los virus recombinantes que expresaban el gen de interés,se realizó una selección de recombinantes negativos para timidina kinasa y positivos para la gD por tres rondas de purificación de placas en monocapas de células RAT-2 las cuales son mutantes para timidina kinasa y en presencia de bromodeoxiuridina. Los virus recombinantes se confirmaron por PCR y secuenciación de nucleótidos y se denominaron RCN-gD ABSTRACT: Bovine Herpesvirus-1 is a DNA virus belonging to the family Herpesviridae, which affects cattle, causing a wide spectrum of clinical manifestations and economic losses. The main immunogenic component is its envelope glycoprotein D (gD), which has been characterized and used as immunogen in different expression systems. The aim of this work wasto generate a recombinant poxvirus(Raccoonpox[RCN]) expressing a truncated version of BHV-1 gD to be used as a vaccine. To do this, it was amplified the gene for a truncated version of gD which subsequently was cloned in transfer plasmid PTK/IRES/tpa which has homology to sites of poxvirus thymidine kinase, an internal site of ribosome entry (IRES) and a secretory signal (tPA), generating the construct PTK/gD/IRES/tpa. To generate the recombinant RCN, we took BSC-1 cells and we infected with a wild type RCN (CDC/V71-I-85A) at a multiplicity of infection of 0.05, then cells were transfected with the construct PTK/gD/IRES/tpa, generating different viral populations with and without the gene of interest. To select recombinant viruses expressing the gene of interest, we performed a selection of recombinant thymidine kinase negative and positive for gD by three rounds of plaque purification on RAT-2 cells monolayers which are thymidine kinase null and using bromodeoxyuridine. Recombinant viruses were recovered and confirmed by PCR and nucleotide sequencing and so called RCN-gD COL0001262

The IBR situation varies between the different European countries. Scandinavia, Austria and Switzerland are considered free of IBR, while Germany, the Netherlands, Belgium, France, Hungary, the Czech Republic and Slovakia have national eradication programs in place. To ensure the health statuses of the different nations, different directives are in place to regulate the moving of animals across borders. One of the main directives is the European Union Council Directive 64/432/EEC. This is a directive for both swine and bovines that are destined for breeding, production and slaughter. It also sets the standards for hygiene and states the type of transport vehicle approved for this kind of transportation. Norway as a member of the EEA has also implemented this directive. In addition to rules for transport of live animals there are also separate regulations for the transport of different genetic material. For sperm collection and storing, the Council Direc-tive 88/407/EECdictates how this should be handled and it also describes the demands for the donor bulls’ health to be documented appropriately. The trading of sperm with third countries is under the same directive. The embryo export and import follows Council Directive 89/556/EECwhich also includes the handling of the donor animal and the teams responsible for the collection and production. Since there is no intra-community border control, special care must be taken to make sure that animal transport is in accordance with European law. Random spot-checks need to be performed according Council Directive 90/425/EEC,this directive also describes the TRACES system which harmonizes the veterinarian health certificates and makes it easier to certify that all requirements are fulfilled. At entry into the EU, additional inspections are to be carried out as stated in Council Directive 91/496/EEC.Various measures must be taken if animals do not comply with regulations. Specific bio-security measures towards IBR are found described in Council Directive 2004/558/EECthat is implemented from Council Directive 64/432/EEC.It acknowledges eradication programs presented by certain Member States and lists regulations for the rest of the EU countries in case of clinical signs being discovered. The conditions for ac-quiring an IBR free-status are given in the Directive 2007/584/ECand include documenta-tion of absence of clinical signs and negative tests. The duty to notify in case of disease is regulated by Council Directive 82/894/EECand the codified forms and the codes for the notification of animal diseases pursuant toCouncil Directive 82/894/. When Norway took part in the EEA-agreement in 1994, the bovine import to Norway also commenced. Norway has had little problems with IBR/IPV with only two positive IPV out-breaks in the early 1960’s and a positive bulk milk sample in 1993. According to the Food Act, IBR is classified as a B-disease in Norway and it is notifiable. If there is suspicion of IBR, measures must be taken as listed in Regulation No 732where both the owner and the Nor-wegian Food Safety Authority have responsibilities in order to prevent spread. Since Norway is free of IBR, strict rules for import and export must be in place to prevent infection. Regulation No 305 dictates regulations for countries with a lower IBR health sta-tus. A procedure that includes identity check, clinical examination must be performed and no signs of IBR must be revealed. Also an isolation period must be finished before ship-ment. The transportation is also under regulation by Regulation No 384which only allows animals fit to finish the journey to travel. During the transit, the animals must be accom-panied by a health certificate according to Regulation No 1484. At import to Norway, there must be a registration with the Norwegian Food Safety Authority and the delivery must be reported using a CVED. For sperm, Regulation No 1489includes both members of the EEA and third countries, and it describes the demands for sperm collection procedure, approval of the bull station and the donor bull. Embryos must be collected and produced by approved groups according to Regulation No 1486 and there must be no findings of IBR. The embryo collection work must be indexed properly and saved. Norway has committed to surveillance and control the IBR-free situation and to produce a report documenting the findings. In return, Norway gets additional guarantees from their trading partner-countries listed in EFTA Surveillance Agency Regulation No 74/94/COL which ensures that there shall be no risk of IBR infection among imported bovines.

An epidemiological and serological survey of bovine herpesvirus 1 (BHV1) infection was conducted in Andalusia from January to April of 2000. A total of 4,035 blood samples were collected from 164 herds. A questionnaire, which included variables potentially associated with infection, was filled out for each herd. Serum samples were obtained to identify specific BHV1 antibodies and were tested using a blocking ELISA test. The observed crude odds ratio (OR) (estimate of the chance of a particular event occurring in an exposed group in relation to its rate of occurrence in a nonexposed group) for vaccination is 9.8 (95% confidence interval: 8.3-11.7). The vaccinated group comprised large dairy farms. This study can only be considered as representative of unvaccinated, small to medium size dairy farms and beef farms in Andalusia. True seroprevalence of the BHV1 virus in nonvaccinated bovine populations in Andalusia reached 45.7% of individuals and 70.4% of herds. Risk factors for BHV1 infection in bovine Andalusian nonvaccinated herds are nonexistence of specific cattle infrastructure (OR: 3.07), beef crossbreeding (OR: 7.90), affiliation with Livestock Health Defence Associations (OR: 2.57), a history of reproductive disorders (OR: 8.39), external replacement (OR: 2.74), proximity to an urban area (OR: 6.11) and herd size (41.98). To control for confounding effects, a binomial logistic regression model was developed. From this regression, BHV1 infections are concentrated in large herds, with external replacement, located close to urban areas. This is the first published report on BHV1 prevalence in the South of Spain., Desde enero a abril de 2000 se ha realizado un estudio epidemiológico y serológico sobre la infección por el herpesvirus bovino tipo 1 (HVB1) en Andalucía, donde se tomaron un total de 4035 muestras de sangre procedentes de 164 rebaños, donde además se cumplimentó un cuestionario que incluye variables potencialmente asociadas a la infección. Mediante un kit ELISA de bloqueo se evaluó la presencia de anticuerpos frente al HVB1. El valor de odds ratio (OR) (probabilidad que tiene un suceso de ocurrir en un grupo expuesto a un factor en relación a la probabilidad en un grupo no expuesto) cruda para la vacunación frente al HVB1 es de 9,8 (IC 95%: 8,3-11,7). Al haberse vacunado las granjas grandes, que fueron eliminadas, el estudio es representativo sólo de granjas de pequeño y mediano tamaño. La prevalencia real en animales no vacunados es del 45,7%, mientras que la dispersión es del 70,4%. Los factores de riesgo detectados son: inexistencia de infraestructuras específicas (OR: 3,07), animales de razas no puras (OR: 7,90), pertenencia a asociaciones de defensa sanitaria (OR: 2,57), historial de trastornos reproductivos (OR: 8,39), reemplazo externo (OR: 2,74), proximidad a zonas urbanas (OR: 6,11) y tamaño del rebaño (41,98). Para limitar el impacto de variables confusoras, se generó un modelo mediante regresión logística, que señala que las infecciones se concentran en grandes rebaños con reemplazo externo en las proximidades de áreas urbanas. Este es el primer trabajo acerca de la prevalencia del HVB1 en el sur de España.

In 2007, an outbreak of epizootic hemorrhagic disease (EHD) occurred in Turkey. On the basis of clinical investigation, 41 cattle were suspected to have EHD. Reverse transcription-PCR and sequence analyses indicated that the virus belonged to EHD virus serotype 6, thus confirming EHD virus infection of cattle in Turkey. UK Research & Innovation (UKRI) Biotechnology and Biological Sciences Research Council (BBSRC) (BBS/E/I/00001144) UK Research & Innovation (UKRI) Biotechnology and Biological Sciences Research Council (BBSRC) (BBS/E/I/00001159) UK Research & Innovation (UKRI) Biotechnology and Biological Sciences Research Council (BBSRC) (BBS/E/I/00001446)

An epidemiological and serological survey of bovine herpesvirus 1 (BHV1) infection was conducted in Andalusia from January to April of 2000. A total of 4,035 blood samples were collected from 164 herds. A questionnaire, which included variables potentially associated with infection, was filled out for each herd. Serum samples were obtained to identify specific BHV1 antibodies and were tested using a blocking ELISA test. The observed crude odds ratio (OR) (estimate of the chance of a particular event occurring in an exposed group in relation to its rate of occurrence in a nonexposed group) for vaccination is 9.8 (95 % confidence interval: 8.3-11.7). The vaccinated group comprised large dairy farms. This study can only be considered as representative of unvaccinated, small to medium size dairy farms and beef farms in Andalusia. True seroprevalence of the BHV1 virus in nonvaccinated bovine populations in Andalusia reached 45.7% of individuals and 70.4% of herds. Risk factors for BHV1 infection in bovine Andalusian nonvaccinated herds are nonexistence of specific cattle infrastructure (OR: 3.07), beef crossbreeding (OR: 7.90), affiliation with Livestock Health Defence Associations (OR: 2.57), a history of reproductive disorders (OR: 8.39), external replacement (OR: 2.74), proximity to an urban area (OR: 6.11) and herd size (41.98). To control for confounding effects, a binomial logistic regression model was developed. From this regression, BHV1 infections are concentrated in large herds, with external replacement, located close to urban areas. This is the first published report on BHV1 prevalence in the South of Spain. Desde enero a abril de 2000 se ha realizado un estudio epidemiológico y serológico sobre la infección por el herpesvirus bovino tipo 1 (HVB1) en Andalucía, donde se tomaron un total de 4035 muestras de sangre procedentes de 164 rebaños, donde además se cumplimentó un cuestionario que incluye variables potencialmente asociadas a la infección. Mediante un kit ELISA de bloqueo se evaluó la presencia de anticuerpos frente al HVB1. El valor de odds ratio (OR) (probabilidad que tiene un suceso de ocurrir en un grupo expuesto a un factor en relación a la probabilidad en un grupo no expuesto) cruda para la vacunación frente al HVB1 es de 9,8 (IC 95%: 8,3-11,7). Al haberse vacunado las granjas grandes, que fueron eliminadas, el estudio es representativo sólo de granjas de pequeño y mediano tamaño. La prevalencia real en animales no vacunados es del 45,7%, mientras que la dispersión es del 70,4%. Los factores de riesgo detectados son: inexistencia de infraestructuras específicas (OR: 3,07), animales de razas no puras (OR: 7,90), pertenencia a asociaciones de defensa sanitaria (OR: 2,57), historial de trastornos reproductivos (OR: 8,39), reemplazo externo (OR: 2,74), proximidad a zonas urbanas (OR: 6,11) y tamaño del rebaño (41,98). Para limitar el impacto de variables confusoras, se generó un modelo mediante regresión logística, que señala que las infecciones se concentran en grandes rebaños con reemplazo externo en las proximidades de áreas urbanas. Este es el primer trabajo acerca de la prevalencia del HVB1 en el sur de España

Sets of serum and milk samples were collected from various countries and prepared, lyophilised and distributed by 1 laboratory to 12 reference laboratories in Europe. The serum sets contained the three European bovine herpesvirus 1 (BHV1) reference serum samples (EU1, EU2 and EU3), serum samples from naturally and experimentally BHV1-infected cattle, from vaccinated, and vaccinated-challenged cattle, from uninfected cattle, and a series of serum dilutions. In addition, sets of milk samples were distributed. The samples were tested for antibodies against BHV1 in virus neutralisation tests, in gB-specific ELISAs, in indirect ELISAs and in gE-specific ELISAs. It was found that the virus neutralisation test and the gB-specific ELISAs were most sensitive for the detection of antibodies in serum, whereas for assaying milk samples the indirect ELISAs were the tests of choice. The results show that the quality of most laboratories appeared to be adequate, but that one laboratory performed considerably below an acceptable level of quality. Four samples from the panel have been proposed that might be selected as reference sera in addition to the three European reference samples.