Bovine viral diarrhea virus (BVDV) is a pestivirus that suppresses the innate and adaptive host immune responses. Each of the two classified genotypes (BVDV1 and BVDV2) has two distinct biotypes – cytopathic (cp) and non-cytopathic (ncp) – and evidence has suggested that cytopathic strains may disrupt host interferon (IFN) synthesis and IFN-mediated responses. However, inconsistent results examining ncpBVDV strains have generated controversy regarding whether they also exhibit this capability. The purpose for this study was to determine the occurrence and functionality of IFN-induced responses within the serum cattle infected with ncpBVDV2-1373. Specifically, this involved analysing the changes in both the serum levels of IFN-α and IFN-γ and the expression of genes that are classically regulated by these cytokines. Serum analysis showed that the infected cattle induced both serum IFN-α and IFN-γ during BVDV infection while PBMC analysis showed increased expression of genes that classically respond to IFN-α – Mx-1, OAS-1, and STAT-1 – and IFN-γ – SOCS-1 and SOCS-3. These findings are supported by temporal kinome analysis, which verified activation of the JAK-STAT signalling network within the PBMCs of the virus-infected animals. In addition to establishing evidence for its synthesis, results from this challenge identified IFN-γ as a possible indicator of animal mortality as analysis of its change within the non-surviving, infected animals was statistically greater than the levels of the surviving, infected animals. Collectively, these results demonstrate 1373-mediated induction of, and host cell response to, both IFN-α and IFN–γ, and the potential for IFN-γ to be a predictive marker for mortality during BVDV infection.

Doctor of Philosophy / Department of Diagnostic Medicine/Pathobiology / Robert L. Larson / Bradley J. White / Bovine viral diarrhea virus (BVDV) is an endemic pathogen in the U.S. cow herd. The virus can cross the placental barrier and infect the unborn fetus. If infection occurs between 45 – 125 days of gestation, persistent infection (PI) in the unborn fetus is likely. Upon parturition, the PI calf is a lifelong shedder of BVDV significantly elevating the risk of viral exposure to non-PI cattle. Despite reports of significant production loss, many BVDV infections are subclinical and in some cases inconsequential. Our data has highlighted various factors potentially causing disparity in clinical outcomes following BVDV exposure including: variation of BVDV serum concentration among PI cattle which may influence the quantity of virus shed into the environment, preexisting BVDV immune (i.e. antibody) status among non-PI cattle, and the degree of stress experienced by non-PI cattle all may influence the susceptibility of disease. Additionally, cattle transiently infected (TI) with BVDV may temporarily shed BVDV thereby offering another source of exposure to non-PI cattle. Programs focusing on BVDV control and prevention consist of diagnostic tests to identify PI cattle, BVDV vaccines to reduce fetal infection and increase herd immunity, and biosecurity programs intended to prevent BVDV exposure to the resident herd. Survey work performed in Montana suggest that educating beef producers with regard to BVDV has significantly increased the implementation of these tools in order to reduce the risk of introducing BVDV to their resident herd. Despite the risk of production loss, the economic benefit of instituting whole-herd BVDV tests may vary due to herd prevalence. By utilizing Monte Carlo simulation, the current BVDV herd prevalence within the U.S. does not economically justify a nationwide BVDV eradication campaign. However, known BVDV positive herds and herds with an elevated likelihood (47%) of being BVDV positive displayed a positive economic outcome when whole-herd BVDV testing strategies were implemented across herd sizes of 50, 100, and 500 cows. The value of testing various testing modalities was dependent upon herd prevalence and herd size. These data suggest that veterinarians must critically evaluate the value of implementing whole herd testing protocols in U.S. beef herds.

Dissertation (Ph.D.)--Auburn University, 2007. / Abstract. Vita. Includes bibliographic references (p.281-340).

Thesis(M.S.)--Auburn University, 2005. / Abstract. Vita. Includes bibliographic references.

Made available in DSpace on 2014-06-11T19:28:22Z (GMT). No. of bitstreams: 0 Previous issue date: 2012-02-07Bitstream added on 2014-06-13T19:16:12Z : No. of bitstreams: 1 pereira_mhc_me_botfmvz.pdf: 261210 bytes, checksum: 2050ad8bcfea0fcd04c28e04b116b34b (MD5) / Universidade Estadual Paulista (UNESP) / O objetivo deste estudo foi avaliar se vacinação contra doenças da reprodução melhora o desempenho reprodutivo de vacas de leite em lactação. Foram realizados quatro experimentos. O experimento 01 foi realizado em 38 fazendas que não utilizavam vacina contra Rinotraqueíte Infecciosa Bovina (IBR), Diarreia Viral Bovina (BVD) e Leptospirose, foram utilizadas 853 vacas Girolando que foram vacinadas (grupo tratado) ou não (grupo controle) no dia -11 (inicio do protocolo de IATF). A segunda dose da vacina foi realizada no dia 30 (diagnóstico de gestação). A vacina que foi utilizada é composta de amostras atenuadas quimicamente alteradas do vírus da IBR associada a amostras citopáticas e não citopáticas do vírus da BVD e culturas inativadas contra cinco sorotipos da Leptospira spp (canicola, grippotyphosa, hardjo, icterohaemorrhagiae e pomona). O experimento 02 foi realizado em 28 fazendas que não utilizavam vacina contra IBR, BVD e Leptospirose, foram utilizadas 287 vacas Girolando que foram pré vacinadas (grupo tratado) ou não (grupo controle) entre os dias -41 a -32, e a segunda dose da vacina foi realizada no dia -11 (inicio do protocolo de IATF). O experimento 03 foi realizado em 17 fazendas que não utilizavam vacina contra IBR/BVDV e Leptospirose, foram utilizadas 1680 vacas holandesas, sendo que vacas com mais de 28 dias em lactação foram vacinadas (grupo tratado) ou não (grupo controle), e a segunda dose da vacina foi realizada 14 dias após a primeira dose. As inseminações foram realizadas entre 15 a 135 dias após a segunda dose da vacina e as perdas de gestação foram avaliadas até 60 dias após a última IA. O experimento 04 foi realizado em 15 fazendas que utilizavam vacina contra IBR, BVD e Leptospirose, foram utilizadas 820 vacas Girolando que foram re-vacinadas... / The aim of this study was to evaluate the effect of vaccination against reproductive diseases on the reproductive performance of lactating dairy cows. The experiment 01 was performed in 38 farms that did not utilize vaccine against IBR/BVD and Leptospirosis, 853 Girolando cows were vaccinated (treated group) or not (control group) on day -11 (beginning of the TAI protocol). The second dose of vaccine was held on day 30 (preganancy diagnosis). The vaccine that was used consists of chemically altered live samples of the IBR virus associated with samples of cytpathic and non cytopathic BVD virus and inactivated cultures against five serotypes of Leptospira spp. (canicola, grippotyphosa, hardjo, icterohaemorrhagiae e pomona). The experiment 02 was performed in 28 farms that did not utilize vaccine against IBR/BVD and Leptospirosis, 287 Girolando cows were pre - vaccinated (treated group) or not (control group), between the days -41 to – 31, the second dose of vaccine was held on day -11 (beginning of the TAI protocol). The experiment 03 was performed in 17 farms that did not utilize vaccine against IBR/BVD and Leptospirosis, 1680 Holstein cows were used in this study, cows with more than 28 days in milk were vaccinated (treated group) or not (control group), the second dose of vaccine was performed 14 days after de first dose. The AI were performed between 15 and 135 days after the second vaccine dose and the pregnancy loss at 60 days after the last AI. The experiment 04 was performed in 15 farms that utilize vaccine against IBR/BVD and Leptospirosis, 820 Girolando cows were vaccinated (treated group) or not (control group) on day -11 (beginning of the TAI protocol). The pregnancy rate was evaluated in the experiments 1, 2 and 4 at 30 and 71 days after TAI, in experiment 03 the pregnancy...(Complete abstract click electronic access below)

Resumo: O Vírus da Diarréia Viral Bovina (BVDV) é um dos patógenos mais importantes na pecuária bovina em todo mundo, principalmente por desencadear manifestações clínicas relacionadas à esfera reprodutiva. A infecção em fêmeas gestantes pode resultar em abortamentos, reabsorções embrionárias, mumificações fetais, má formações e nascimento de bezerros fracos além do aparecimento de animais persistentemente infectados e imunotolerantes ao vírus, que são a principal fonte de infecção e disseminação da doença nos rebanhos. Atualmente, a complexidade do diagnóstico e consequentemente a patogenia, estão relacionados às diferenças genotípicas do agente. Por isso, a presente pesquisa teve como objetivo verificar a ocorrência dos genótipos BVDV-1 (Singer) e BVDV-2 (VS-253) em vacas, e respectivos fetos, abatidas em um frigorífico no Estado de São Paulo por meio da análise do soro sanguineo por meio da técnica de virusneutralização. No contexto geral, 52,51% (115/219) das vacas testadas foram reagentes, mas nenhum feto (0/219) reagiu na virusneutralização. Pela análise cruzada conforme a estirpe viral, observou-se que 42% (92/219) das vacas foram reagentes tanto para o genótipo BVDV-1 como para o genótipo do BVDV-2. Por outro lado 4,10% (9/219) reagiram apenas para o genótipo BVDV-1 e 6,39% (14/219) reagiram apenas para o genótipo do BVDV 2. Notou-se portanto que ambas as estirpes estão disseminadas nas regiões estudadas, fato que justifica o emprego de antígenos diferentes para evitar diagnóstico falso-negativo. Por fim, não foi observado qualquer alteração nos fetos que pudessem ser caracterizada como patologia da enfermidade. / Abstract: The Bovine viral diarrhea virus (BVDV) is one of the pathogens in bovine livestock worldwide most important mainly triggered by clinical manifestations related to the reproductive sphere. The infection in pregnant females may result in abortions, embryonic resorptions, fetal mummification, poor training, birth of weak calves in addition to persistently infected and virus immunotolerant animals, which are the main source of infection and spread of the disease. Currently, the complexity to diagnosis and consequently to the pathogenesis are related genotypic differences that he presents. Therefore, this research aimed to verify the occurrence of BVDV- 1 (Singer) and BVDV-2 (VS-253) genotypes in cows and their respective fetuses slaughtered in a abattoir at the state of São Paulo by analyzing the blood serum using virusneutralization technique. In the general context, 52.51% (115/219) of cows were reagents, but no fetus (0/219) reacted in virusneutralization. After a cross-examination we observed that 42% (92/219) of cows reacted for both BVDV-1 and BVDV-2 genotype. Furthermore 4,10% (9/219) reacted only to the genotype BVDV-1 and 6,39% (14/219) responded only to the genotype 2 of BVDV. It was noted therefore that both strains are widespread in the regions studied, which also justifies the use of different antigens to avoid false-negative diagnosis. Finally, there was no change in fetuses that could be characterized as a pathology of the disease. / Orientador: Samir Issa Samara / Coorientador: Fabio Carvalho Dias / Banca: José Gabriel Amoril / Banca: Sandra Possebon Gatti / Mestre

Master of Science / Department of Diagnostic Medicine/Pathobiology / Raymond R. R. Rowland / Pestiviruses such as classical swine fever virus (CSFV) and bovine viral diarrhea virus (BVDV) are some of the most economically important livestock diseases in the world. The antigenic similarities between members of the pestivirus genus allow for both BVDV and CSFV to infect swine. Infections with heterologous pestiviruses in swine can interfere with diagnostic tests for CSFV. The identification of cross-reactive and cross-neutralizing epitopes between CSFV and BVDV for the development of improved diagnostics and vaccines that allow for the differentiation of infected animals from vaccinated animals (DIVAs) are necessary to accurately detect and control CSFV. The overall goal of this research was to identify epitope regions recognized by antibodies that can differentiate between CSFV and BVDV. The approach was to use serum neutralization assays to confirm the presence of neutralizing antibodies to BVDV in swine serum collected from animals immunized with one of three separate Alphavirus vaccine constructs: BVDV-1b, CSFV E2, and CSFV E[superscript]rns. Results showed that animals immunized with the Alphavirus BVDV-1b construct had high neutralizing titers against BVDV-1a and animals immunized with Alphavirus CSFV E2 and E[superscript]rns constructs had low, but detectable, neutralizing activity. Polypeptide fragments of CSFV and BVDV E2 were then expressed in E. coli and purified using affinity chromatography. Serum from a pig immunized with the CSFV E2 Alphavirus construct was tested against two fragments of CSFV E2, 2/4 and 4/4, and four fragments BVDV E2, 1/4, 2/4, 3/4, and 4/4, using western blot analysis. Reactivity to fragments CSFV E2 2/4 and 4/4 and BVDV E2 1/4 and 4/4 was observed. The results of this study identified CSFV amino acid positions 774 through 857 and BVDV amino acid positions 783 through 872 as the regions that contain the epitopes recognized by cross-reactive antibodies between BVDV and CSFV E2. These results provide more specific sequence regions to improve CSFV diagnostic assays and DIVA vaccines.

Bovine viral diarrhea virus (BVDV) is an important viral pathogen affecting all ages of cattle, resulting in significant economic losses worldwide. BVDV infection is associated with a diverse array of symptoms including gastrointestinal disorder, respiratory distress, fetal malformation, stillbirth, abortions, and mucosal disease (MD). Transplacental infections of fetuses between 42 and 125 days of gestation can result in immune-tolerance and the surviving fetuses become persistently infected (PI). PI animals are major reservoir of BVDV and it becomes problematic to control the disease. The objectives of this dissertation were to: 1) develop a cost-effective testing scheme to detect BVDV PI animals from exposed herds, 2) characterize two virulent BVDV-2 Mississippi isolates associated with severe hemorrhagic diseases, and 3) perform phylogenetic analysis based on sequences of 5'UTR, E2, and NS5B regions. First, we developed a BVDV testing scheme by combining pooled real-time RT-PCR with antigen capture enzyme-linked immunosorbent assay (ACE) to screen cattle herds. From positive pools individual positives were identified using ACE. Data from a three year period indicated that 92.94% PI animals were infected with BVDV-1, 3.53% with BVDV-2, and 3.53% with both BVDV-1 and BVDV-2. Analysis of the 5'UTR of 22 isolates revealed the predominance of BVDV-1b followed by BVDV-2a. Second, two virulent BVDV isolates, M10-3432 and M10-5347, were successfully recovered from an adult beef breeding cow and feedlot calf respectively. When compared to the reference strain BVDV-2 125c, five and three unique amino acids in E2 regions were different from M10-5347 and M10-3432 respectively. Phylogenetic analysis of E2 region grouped both Mississippi isolates in BVDV-2a, a subtype containing high virulent strains. M10-3432 was clustered with high virulent strain 890 while M10-5347 was clustered with high virulent strain CD87. Third, we compared the phylogenetic analyses of BVDV based on the sequences of 5'UTR, E2, and NS5B at either nucleotides or amino acids level. Although slight differences were observed, the virulent BVDV isolates were consistently classified into BVDV-2a cluster regardless of region of sequences used. Furthermore, phylogenetic tree constructed using combined two or more regions had higher posterior probability and bootstrap value than phylogenetic trees constructed using a single region

Bovine Viral Diarrhea Virus (BVDV) is a significant disease causing agent with major economic impact on the cattle industry, causing both productive and reproductive losses. One reason for its widespread distribution is that the majority of all BVDV infections occur without clinical signs, leaving most cases of BVDV undetected in cow herds. BVDV occur as cytopathic (CP) or non-cytopathic (NCP) biotypes, classified according to whether or not they produce visible changes in cell culture. CP BVDV biotype but not NCP biotype is implicated in the induction of apoptosis in vivo. The interaction of BVDV with its host has several unique features, most notably the capacity to infect its host either transiently or persistently. The pathogenesis of the disease caused by BVDV is complicated and interaction between BVDV and the host are poorly understood. The overall goal of this research is to identify mechanistic pathways that govern the outcome of BVDV infection in susceptible host cells. Specific aspects of this goal is to understand BVDV biotypes-induced changes on cellular proteome, cell death and survival mechanisms used by BVDV biotypes in apoptosis pathway, interactions of BVDV NS3 viral protein with host cellular proteins and how BVDV cell entry and infection interfere with an early step of professional antigen presentation, antigen uptake. The results of this work showed, for the first time, the successful use of proteomics in studying BVDV-host interactions in a comprehensive approach. Using the Gene Ontology and systems biology analysis we identified biotype-related differences in significant biological pathways and functions. Also, using a proteomics approach, we identified multiple critical cellular proteins that interact with CP NS3 viral protein at multiple stages of CP BVDV replication cycle. This project provides insight into the cellular pathways and functions involved in the viral cytopathogenicity of CP BVDV biotype. In addition, our data not only confirmed the previous observations on the critical involvement of the intrinsic pathway of apoptosis in CP BVDV infection, it also identified multiple mitochondrial and antioxidant proteins contributing to this pathway. Finally, we show that BVDV exploit selective antigen uptake mechanisms in professional antigen presenting cells monocytes during viral entry.

Doctor of Philosophy / Department of Diagnostic Medicine/Pathobiology / Daniel U. Thomson / Bovine viral diarrhea virus (BVDV) has emerged as one of the most important infectious diseases in cattle. One particular important manifestation, after successfully establishing an in utero infection of the fetus during the first trimester, is the development of a persistently-infected BVDV (PI-BVDV) calf. Persistently infected BVDV animals are a continuous source of virus and can shed the virus in virtually all secretions and excretions, including nasal discharges, saliva, semen, urine, tears, milk, and, to a lesser extent, feces. The objectives of this research were to determine: 1) the effects of short term exposure (13 – 18 days on feed (DOF)) to PI-BVDV feeder cattle; 2) the outcome of testing and removing PI-BVDV feeder calves at time of feedlot arrival on health, performance, and carcass characteristics; 3) the survival of BVDV on materials associated with livestock production; and 4) characterization of testing and longitudinal prevalences for PI-BVDV beef cattle. Testing and removing PI-BVDV calves at 13 to 18 DOF was too late to remove a morbidity effect due to PI-BVDV exposure. However, mortality, performance, and carcass characteristics were not different in cattle exposed to PI-BVDV cattle. Additionally, there were no harmful outcomes when newly arrived feeder cattle were exposed to a PI-BVDV animal for one to two days following feedlot entry. A non-cytopathic, Type 1b, BVDV was capable of surviving after application to various materials used in livestock production. BVDV tended to survive longer on non-porous materials than porous materials. When in the presence of mucus, BVDV was protected from degradation for longer periods of time than when not in the presence of mucus. There was no difference in overall PI-BVDV prevalence within cattle sampled in 2006 and 2007. Cattle that weighed less than 300 lbs. had a greater likelihood of being PI-positive than cattle with increased weights. Several months of the year had a greater likelihood of having PI-positive animals. Based on operation, cow-calf and stocker operations had a greater likelihood of having PI-positive animals than did feedlot operations.