Nature of early reproductive failure caused by bovine viral diarrhea virus

A 2-part study was undertaken to determine the effect of bovine viral diarrhea (BVD) virus on fertilization and early development of embryos. In experiment 1, 10 seronegative cows were superovulated and artificially inseminated twice during estrus. After the second insemination, 5 of the cows received intrauterine infusion of BVD virus suspension. The other 5 cows received suspending medium only and served as controls. All 10 cows were slaughtered on day 3, and ova and embryos were collected for morphologic evaluation. A total of 49 and 52 ova and embryos were collected from the control and virus-treated cows, respectively. Among the ova and embryos collected from control cows, 81.6% were fertilized, whereas only 52% were fertilized in the virus-treated group. The statistically significant difference (P less than 0.01) indicated that the virus interferes with fertilization. In experiment 2, the protocol was identical except for slaughter on day 13. Seventy-nine ova and embryos were collected from the 6 control cows, and the 6 virus-treated cows yielded 59 ova and embryos. Of the total ova and embryos recovered on day 13, 88.6% and 50.8% were hatched and developing normally in the control and virus-treated groups, respectively. The difference was highly significant (P less than 0.001). Unfertilized ova and degenerating embryos could not be differentiated on the basis of morphologic appearance. The nearly identical percentages of unfertilized ova in experiment 1 and unhatched ova and embryos in experiment 2 strongly suggested that fertilization failure is the principal manifestation of the observed adverse effect of BVD virus infection.     

Sensitivity and specificity of an enzyme-linked immunosorbent assay for the detection of bovine viral diarrhea virus antibody in cattle.

A reliable bovine viral diarrhea (BVD) viral antigen was prepared from BVD virus grown on Madin Darby bovine kidney (MDBK) cells by solubilizing the virus with detergent MEGA-10 (decanoyl-N-methylglucamide) followed by removal of hydrophobic proteins with Triton X-100 treatment. By these treatments, problems of high background associated with BVD viral antigen in the enzyme-linked immunosorbent assay (ELISA) were eliminated. With this new antigen, an ELISA was adapted to detect bovine serum antibody against BVD virus. The diagnostic specificity of the assay in 403 bovine sera collected from a BVD virus-free herd was 100%; in 296 bovine sera with serum neutralizing antibody titers of greater than or equal to 1:2, 289 sera were ELISA positive (relative sensitivity of 97.6%), two sera gave false negative reactions (0.7%) and five sera gave suspicious reactions (1.7%). These interpretations were based on positive/negative (P/N) ratio readings, i.e. a P/N ratio of less than 1.50, 1.50-1.99 and greater than or equal to 2.00 were interpreted as negative, suspicious and positive reactions, respectively. The ELISA results gave excellent agreement with serum neutralization in detecting both seropositive and seronegative animals (Kappa = 0.994). The ELISA assay was considered to be technically superior to the serum neutralization test for the routine detection of BVD viral antibody in bovine sera.     

Bovine viral diarrhoea virus infection in cattle,sheep and goats in northern Tanzania

Virological and serological investigations were carried out in cattle, sheep and goats raised in the northern part of Tanzania in order to explore the possibility of bovine viral diarrhoea (BVD) virus cycling within these animal species. Two noncytopathogenic BVD virus isolates (A4/5/Tan86 and A4/10/Tan86) were obtained from sera sampled from inapparently infected indigenous (zebu) cattle originating from Kiteto district in Arusha region. No BVD virus was isolated from any of the sheep or goat sera. Seroepidemiological investigations revealed widespread prevalence of neutralising antibodies to BVD virus not only in cattle but also in sheep and goats. The seropositive rates are discussed in relation to previous observations in Tanzania and other parts of the world and to the livestock husbandry practices in northern Tanzania.     

Frequency of persistent bovine viral diarrhea virus infection in selected cattle herds

Sera and blood buffy coat samples were obtained from 3,157 cattle in 66 selected herds. Antibodies to bovine viral diarrhea (BVD) virus were detected in 89% of the serum samples by immunoprecipitation or virus-neutralization tests. Cytopathic or noncytopathic BVD viruses were isolated from blood buffy coat samples from 60 cattle in 6 herds. A second blood buffy coat sample was obtained from 54 of the 60 cattle 2 months after the initial sampling, and BVD virus was isolated again from each cow. The 54 cattle were considered persistently infected with BVD virus. The frequency of persistent infection was 1.7%.     

Viral and viral protein specificity of antibodies induced in cows persistently infected with noncytopathic bovine viral diarrhea virus after vaccination with cytopathic bovine viral diarrhea virus

Neutralizing and nonneutralizing antibodies to bovine viral diarrhea (BVD) virus were detected in 3 cows persistently infected with noncytopathic BVD virus after vaccination with modified-live cytopathic BVD virus. Neutralizing antibodies detected in serum samples from each persistently infected cow at 3 weeks after vaccination were highly specific for certain isolates of cytopathic BVD virus and reacted only with a viral protein with a molecular weight of 53,000. Neutralizing antibodies to 1 of 3 isolates of noncytopathic BVD virus were detected in a serum sample obtained at 12 weeks after vaccination from 1 of 3 persistently infected cows. Nonneutralizing antibodies were detected in all cows at 7 to 12 weeks after vaccination. The nonneutralizing antibodies were less specific for isolates of BVD virus and reacted with viral proteins with molecular weights of 115,000, 80,000, 53,000, and 47,000.     

Fluorogenic RT-PCR assay (TaqMan) for detection and classification of bovine viral diarrhea virus

A single tube fluorogenic RT-PCR-based 'TaqMan' assay was developed for detection and classification of bovine viral diarrhea virus (BVDV). TaqMan-PCR was optimized to quantify BVD virus using the ABI PRISM 7700 sequence detection system and dual-labeled fluorogenic probes. Two different gene specific labeled fluorogenic probes for the 5' untranslated region (5' UTR) were used to differentiate between BVD types I and II. Sensitivity of the single tube TaqMan assay was compared with two-tube TaqMan assay and standard RT-PCR using 10-fold dilutions of RNA. Single tube TaqMan assay was 10-100-fold more sensitive than the two-tube TaqMan assay and the standardized single tube RT-PCR. Specificity of the assay was evaluated by testing different BVD virus strains and other bovine viruses. A total of 106 BVD positive and negative pooled or single serum samples, field isolates and reference strains were tested. Quantitation of cRNA from types I and II BVD virus was accomplished by a standard curve plotting cycle threshold values (C(T)) versus copy number. Single tube TaqMan-PCR assay was sensitive, specific and rapid for detection, quantitation and classification of BVD virus.     

Seroepidemiologic survey for antibodies to selected viruses in the respiratory tract of lambs

A serologic study was conducted to determine the prevalence of antibodies to, and infection rate of, Mastadenovirus ovi 5, M ovi 6, parainfluenza-3 (PI-3) virus, bovine herpesvirus-1 (BHV-1), respiratory syncytial virus (RSV), bovine viral diarrhea (BVD) virus, and ovine progressive pneumonia (OPP) virus in lambs at a ram lamb growth-rate test station. For 2 consecutive years, serum samples were prepared from blood collected from 1- to 2-month-old ram lambs as they entered the test station (1st sample) and again 2 months later (2nd sample). The 1st year, 59 producers submitted 237 lambs; the 2nd year, 65 producers submitted 253 lambs. Microtitration serum virus-neutralization tests were used to determine antibody titers for M ovi 5, M ovi 6, PI-3 virus, BHV-1, and BVD virus. Antibodies to RSV and OPP virus were determined, using indirect hemagglutination and agar-gel immunodiffusion, respectively. Based on results of the 1st blood samples collected, the mean prevalence for both years was as follows: 95% of the lambs were seropositive for M ovi 5; 87.2% for PI-3 virus; 84.5% for RSV; 41.7% for M ovi 6; 8.7% for BVD virus; 5.4% for BHV-1; and 3.3% for OPP virus. Based on the 2-year mean, M ovi 6 had the highest infection rate (207 of 484 [42.8%]) as determined by the number of lambs evaluated having a greater than or equal to 4-fold increase in serum antibody titer from the 1st to the 2nd sampling. Infection rates of the other viruses were: 31.0% for M ovi 5; 15.3% for PI-3 virus; 5.6% for RSV; 0.6% for BVD virus; and 0.4% for BHV-1. One lamb became seropositive for OPP virus the 2nd year.     

Prevalence of Bovine viral diarrhoea virus (BVDV) antibodies among sheep and goats in India

The aim of this study was to determine the prevalence of pestivirus antibodies in sheep and goats in India. A total of 2803 serum samples collected between 2004 and 2008 from 1777 sheep in 92 flocks and 1026 goats in 63 flocks belonging to 13 states were tested by competition ELISA for detection of pestivirus antibodies. In sheep, the true prevalence rate was 23.4% (95% confidence interval: 22.9%–27.0%) and in goats it was 16.9% (95% CI: 16.4%–21.3%). The flock level seroprevalence was 66.3% for sheep and 54.0% for goats. Geographical variation in individual and flock prevalence was highly significant. A significant association (p?<?0.05) was found between sheep and goat flocks having cattle contact and the flock level seroprevalence. The seroprevalence was lower in 6 months–1 year age group compared to the 1–2 year and >2 year age groups in both sheep and goats. Cross neutralization studies on 61 seropositive sheep and 34 seropositive goat samples representing all positive flocks, exhibited > four fold higher titre to bovine viral diarrhoea virus type 1 (BVDV-1) in 41 sheep and 23 goat samples and to BVDV-2 in one sheep and goat each. This study for the first time showed serological evidence of wide spread BVDV infections in Indian sheep and goats, with BVDV-1 predominating and BVDV-2 occasionally besides highlighting the potential risk of infection to other species, which needs to be considered whenever BVD control measures are initiated.     

Comparison of persistence of seven bovine viruses on bovine embryos following in vitro exposure

The ability of seven cytopathic strains of bovine viruses to adhere to the zona pellucida of six-to-eight day-old bovine embryos were compared. Embryos were exposed to virus by placing them either in virus suspensions or by culturing them on infected bovine turbinate cultures for 18-24 h. After exposure to bovine virus diarrhea virus (BVDV), infectious bovine rhinotracheitis virus (IBV), bluetongue virus (BTV), pseudorabies virus (PRV), vesicular stomatitis virus (VSV), parainfluenza 3 virus (PI3), or bovine enterovirus virus (BEV), the embryos were tested for virus by culture in bovine turbinate cells and by morphological examination using electron microscopy (EM). A special technique to minimize loss of embryos processed for EM was developed. More embryos had viral particles on the surface of the zona pellucida after exposure to 18-24 hour infected cell cultures than did embryos exposed to viral culture suspensions. The most dramatic finding was that BTV adhered in large numbers to the surface of the zona pellucida of exposed embryos. IBRV, PRV, and VSV comprised an intermediate group, with virions occasionally detected on the surface of exposed embryos after 5 washes. Therefore, extensive washing is required. The PI3 and BEV were easily removed from embryo-exposed virus by washing. BVD was difficult to identify morphologically, making assessment by EM unreliable. There was no evidence that any one of the seven viruses penetrated the intact zona pellucida. Using a micromanipulator, 42 embryos were also directly inoculated through the zona pellucida with +/- 50 picoliters of virus inoculum or medium.(ABSTRACT TRUNCATED AT 250 WORDS)     

Prevalence of serum antibodies to bovine herpesvirus-1 and bovine viral diarrhea virus in beef cattle in Uruguay

Our objective was to determine the prevalence of serum antibodies to bovine herpesvirus-1 (BHV-1) and bovine viral diarrhea (BVD) virus in beef cattle in Uruguay. A random sample of 230 herds selected with probability proportional to population size based on the number of cattle was chosen from a list frame of all registered livestock farms as of June 1999. Sera from up to 10 heifers, cows and bulls (up to 30 sera total per herd) were collected on selected farms between March 2000 and March 2001 and evaluated by means of enzyme-linked immunosorbent assays (ELISAs). Overall, 6358 serum samples were evaluated. We also collected data on previous diagnosis of BHV-1 or BVD infections and on the use of vaccines against these agents.

The estimated prevalence of exposure to BHV-1 and BVD at the herd level for the Uruguayan beef population was 99% and 100%, respectively. Approximately 37% of beef cattle in Uruguay have been exposed to BHV-1 and 69% to BVD virus. Only 3% of beef herds in Uruguay regularly (typically, annually) use vaccines against either of these agents.     

Experimental production of bovine respiratory tract disease with bovine viral diarrhea virus

Five 6-month-old calves were inoculated with bovine viral diarrhea (BVD) virus (n = 3) or Pasteurella haemolytica (n = 2) endobronchially with a fiberoptic bronchoscope. Five additional calves were inoculated sequentially with BVD virus followed by P haemolytica at a 5-day interval. Blood samples were collected daily from the calves for bacterial isolation. Clinical signs of respiratory tract disease in calves were recorded daily. If the calves survived, they were killed for necropsy 3 or 4 days after inoculation with P haemolytica (or 8 days after inoculation with BVD virus). The extent and nature of pulmonary lesions in the calves were determined, and the lower portion of the respiratory tract (lungs and trachea) was examined for both these organisms. The 3 calves, inoculated with BVD virus only, developed mild clinical signs mainly manifested as fever, nasal discharge, and occasional cough. Approximately 2% to 7% of the total lung capacity of these calves was pneumonic. Mild clinical signs and localized lesions involving about 15% of the lung volume developed in the 2 calves exposed to P haemolytica only. However, severe fibrinopurulent bronchopneumonia and pleuritis involving 40% to 75% of lung volume developed in the 5 calves inoculated sequentially with BVD virus and P haemolytica. The possible role BVD virus may have in bovine respiratory tract disease is discussed.     

Immunologic and virologic findings in a bull chronically infected with noncytopathic bovine viral diarrhea virus

Depressed lymphocyte blastogenesis in response to mitogen stimulation, depressed iodination of protein by neutrophils, and enhanced ingestion of Staphylococcus aureus by neutrophils were detected in a bull with chronic bovine viral diarrhea (BVD). Before developing chronic BVD, the bull was vaccinated with a killed cytopathic BVD virus. Neutralizing antibodies specific for the vaccine virus were detected in serum specimens obtained from the bull immediately before death. A noncytopathic BVD virus was isolated from the spleen after death. The immunologic and virologic findings in this bull supported reported research findings on the pathogenetic mechanisms involved in chronic BVD and mucosal disease.     

Persistent infections after natural transmission of bovine viral diarrhoea virus from cattle to goats and among goats

Bovine viral diarrhoea virus (BVDV) is an economically important pathogen of cattle worldwide. Infection of a pregnant animal may lead to persistent infection of the foetus and birth of a persistently infected (PI) calf that sheds the virus throughout its life. However, BVD viruses are not strictly species specific. BVDV has been isolated from many domesticated and wild ruminants. This is of practical importance as virus reservoirs in non-bovine hosts may hamper BVDV control in cattle. A goat given as a social companion to a BVDV PI calf gave birth to a PI goat kid. In order to test if goat to goat infections were possible, seronegative pregnant goats were exposed to the PI goat. In parallel, seronegative pregnant goats were kept together with the PI calf. Only the goat to goat transmission resulted in the birth of a next generation of BVDV PI kids whereas all goats kept together with the PI calf aborted. To our knowledge, this is the first report which shows that a PI goat cannot only transmit BVD virus to other goats but that such transmission may indeed lead to the birth of a second generation of PI goats. Genetic analyses indicated that establishment in the new host species may be associated with step-wise adaptations in the viral genome. Thus, goats have the potential to be a reservoir for BVDV. However, the PI goats showed growth retardation and anaemia and their survival under natural conditions remains questionable.     

Safety of a temperature-sensitive vaccine strain of bovine viral diarrhea virus in pregnant cows

Safety tests were conducted in 78 pregnant cows vaccinated with a commercial preparation of a temperature-sensitive vaccine strain of bovine viral diarrhea (BVD) virus. After vaccination, seroconversion was detected in 33 (97%) of 34 cattle that did not have antibodies against BVD virus. Overall, 43 (91%) of 47 cows with prevaccination titers less than or equal to 4 seroconverted. During the test period, cows did not become naturally infected with BVD virus, and BVD-associated reactions to the vaccine were not observed in vaccinated cows. Calves born to vaccinated cows did not have clinical signs of fetal BVD. Precolostral blood samples collected from the progeny of cows that were seronegative at vaccination were free of antibody against BVD virus. Bovine viral diarrhea virus was not isolated from the cattle evaluated in the present study.     

Transplacental BVD virus transmission after experimental inoculation of goats in different pregnancy stages

Eight groups of altogether 25 goats without neutralizing antibodies against BVD virus, were inoculated either intranasally or intranasally and subcutaneously with two different BVD virus isolates during different stages of gestation. In all 18 goats inoculated within the first 78 days of gestation an abortion and foetal death rate of approximately 100% occurred. Only one goat gave birth to a clinically healthy kid. The other seven goats which were inoculated after the 78th day of gestation showed also a high foetal death rate. Only two of them gave birth to clinically healthy kids. Neutralizing antibodies against BVD virus could be detected in blood samples drawn from 14 kids born at normal term including stillborn and non-viable offsprings. BVD virus was reisolated from different organs taken from seven foetuses. It was not possible to isolate BVD virus from any of the normal offsprings.     

Serologic examination of aborted ovine and bovine fetal fluids for the diagnosis of border disease, bluetongue, bovine viral diarrhea, and leptospiral infections

Fetal serum from most of 994 bovine and 553 ovine aborted fetuses was tested serologically for antibodies to border disease (BD), bovine viral diarrhea (BVD), and bluetongue (BT) viruses, and to Leptospira sp., and the results were compared with the results of isolation procedures, fluorescent antibody tests (FAT), and histologic examinations of the same fetuses. Antibodies to BT virus were not found in any of the 994 bovine and 553 ovine fetuses. Antibody titers to BVD virus were present in 39 of 966 bovine fetuses tested, and BVD virus was detected in 4 of the 39. Four of 74 fetuses in which the BVD virus was detected by FAT or isolation had titers to BVD virus. Microagglutination (MAT) titers to 1 or more of 5 serovars of leptospires were present in 52 of 773 bovine fetal sera tested. Leptospires were not detected by FAT in any bovine fetuses that had leptospiral antibody titers. Leptospires were detected by FAT in 15 aborted calves, and none of these had MAT titers. Antibody titers to BD virus were present in 80 of 486 fetal lamb sera tested, and the virus was detected by FAT or isolation in 3 of the 80 fetuses. Border disease virus was detected in 14 of 486 fetal lambs tested. Twelve of the 14 were tested serologically and 3 had titers to BD virus. Leptospiral antibody titers were present in 27 of 326 ovine fetal sera tested. Leptospires were not detected in any of the 326 ovine fetuses tested by FAT.     

Antibodies to bovine viral diarrhoea virus in beef cattle

Infection of cattle with bovine viral diarrhoea virus (BVD virus) is common throughout the world(1) and the prevalence of neutralising antibodies to the virus reported from surveys ranges from about 40% to 90%(2)(3)(4). The first isolation of BVD virus in New Zealand was reported in 1967(5) and, since that time, evidence of widespread infection in dairy cattle has been presented(6). Whilst the diseases associated with BVD viral infection have been well recognised in dairy herds, there has been a belief that infection of beef herds is less common. Based on this belief has been the fear that the growth of the dairy beef industry could lead to the introduction of BVD virus into an essentially naive beef population with disastrous results such as those reported by MacNeil and van der Oord⁽⁷⁾. We decided therefore to sample beef cattle submitted to abattoirs throughout New Zealand for serological evidence of prior exposure to BVD virus.     

Specificity of neutralizing and precipitating antibodies induced in healthy calves by monovalent modified-live bovine viral diarrhea virus vaccines

Serum was obtained at weekly intervals after vaccination of 6 healthy calves with either of 2 commercially available monovalent modified-live bovine viral diarrhea (BVD) virus vaccines. Detectable neutralizing antibodies to each of 10 cytopathic and 10 noncytopathic isolates of BVD virus were produced by 1 or more of the calves by 14 days after vaccination, but no calf produced detectable neutralizing antibodies to all 20 BVD viruses. At that time, precipitating antibodies against viral-induced polypeptides of approximately 115,000; 80,000; 56,000; 48,000; 39,000; and 25,000 daltons were detected in sera from some calves. Also at that time, specificity of the antibodies for polypeptides of certain viruses was detected. At 21 days after vaccination, each calf produced neutralizing antibodies to all 20 BVD viruses. At that time, precipitating antibodies to each of the aforementioned viral induced polypeptides were detected in serum from each calf. Precipitating antibodies to viral induced polypeptides of 61,000 and 37,000 daltons were detected in samples of sera obtained from some calves at 42 days after vaccination.     

Neutralization kinetics of bovine viral diarrhea virus by hyperimmune serum: one or multi-hit mechanism

The kinetics of bovine viral diarrhea (BVD) virus neutralization (VN) by hyperimmune serum followed first order kinetics at low dilutions (1:8 and 1:16) of hyperimmune bovine serum. A lag phase in the VN curve occurred when the serum was further diluted. Addition of rabbit anti-bovine IgG, but not anti-IgM, significantly increased the degree of VN after BVD virus was reacted with further diluted (1:256 dilution) anti-BVD bovine hyperimmune serum. Incubation of virus-hyperimmune serum (1:64 dilution) at 4 degrees C for 0 to 60 min, followed by incubation at 37 degrees C indicated VN occurred as a two-stage process: a binding (temperature-independent) phase that was followed by a triggering (temperature-dependent) phase. The data favor the thesis that neutralization of BVD virus occurs by a multi-hit mechanism and requires combination of at least two molecules of antibody with each virus.