Yijing Zhang

Comparison of three commercial vaccines for preventing persistent infection with bovine viral diarrhea virus.

Eighty crossbred beef heifers were randomly allocated to four groups to evaluate the efficacy of vaccination in preventing development of calves persistently infected with bovine viral diarrhea virus (BVDV). Group 1 (n=11) was non-vaccinated controls, whereas three groups were vaccinated with commercially available multivalent BVDV vaccines at weaning (approximately 7 mo of age), 28 d post-weaning, approximately 1 y of age, and 28 d later. Groups 2 (n=23) and 3 (n=23) were given a modified-live BVDV vaccine, whereas Group 4 was given an inactivated BVDV vaccine. Heifers were bred by AI and subsequently exposed to two bulls. At 61 d after AI, 70 heifers were pregnant (n=10 for Group 1 and n=20/group for Groups 2, 3, and 4). Three cattle persistently infected with BVDV were commingled with the pregnant heifers (in an isolated pasture) from 68 to 126 d after AI. Thereafter, viremias were detected in pregnant heifers from Groups 1, 3, and 4 (10/10, 1/20, and 10/20, respectively), but not in pregnant heifers from Group 2 (0/20). Resulting calves were assessed for persistent infection using serum PCR, ear notch antigen capture-ELISA, and immunohistochemistry. Persistently infected calves were only produced in Group 1 (10/10) and Group 4 (2/18). In conclusion, commercial vaccines provided effective fetal protection despite prolonged natural exposure to BVDV. Given that viremias were detected in 11 vaccinated heifers after BVDV exposure, and two vaccinated heifers gave birth to persistently infected calves, there is continued need for biosecurity and diagnostic surveillance, in addition to vaccination, to ensure effective BVDV control.

Protective effects against abortion and fetal infection following exposure to bovine viral diarrhea virus and bovine herpesvirus 1 during pregnancy in beef heifers that received two doses of a multivalent modified-live virus vaccine prior to breeding.

OBJECTIVE To determine whether administration of 2 doses of a multivalent, modified-live virus vaccine prior to breeding of heifers would provide protection against abortion and fetal infection following exposure of pregnant heifers to cattle persistently infected (PI) with bovine viral diarrhea virus (BVDV) and cattle with acute bovine herpesvirus 1 (BHV1) infection. DESIGN Randomized controlled clinical trial. ANIMALS 33 crossbred beef heifers, 3 steers, 6 bulls, and 25 calves. PROCEDURES 20 of 22 vaccinated and 10 of 11 unvaccinated heifers became pregnant and were commingled with 3 steers PI with BVDV type 1a, 1b, or 2 for 56 days beginning 102 days after the second vaccination (administered 30 days after the first vaccination). Eighty days following removal of BVDV-PI steers, heifers were commingled with 3 bulls with acute BHV1 infection for 14 days. RESULTS After BVDV exposure, 1 fetus (not evaluated) was aborted by a vaccinated heifer; BVDV was detected in 0 of 19 calves from vaccinated heifers and in all 4 fetuses (aborted after BHV1 exposure) and 6 calves from unvaccinated heifers. Bovine herpesvirus 1 was not detected in any fetus or calf and associated fetal membranes in either treatment group. Vaccinated heifers had longer gestation periods and calves with greater birth weights, weaning weights, average daily gains, and market value at weaning, compared with those for calves born to unvaccinated heifers. CONCLUSIONS AND CLINICAL RELEVANCE Prebreeding administration of a modified-live virus vaccine to heifers resulted in fewer abortions and BVDV-PI offspring and improved growth and increased market value of weaned calves.

Epidemiology of prolonged testicular infections with bovine viral diarrhea virus.

Previously, bovine viral diarrhea virus (BVDV) had been found in prolonged testicular infections following acute infection of immunocompetent bulls. The primary purpose of this research was to evaluate the production and maintenance of prolonged testicular infections after exposure to BVDV of seronegative bulls in varying circumstances. The secondary objective was to initiate assessment of the potential for transmission of BVDV via semen of bulls exhibiting a prolonged testicular infection. In total, 10 research trials were conducted. The first trial examined the duration of detectable virus in semen after intranasal inoculation of peri-pubertal bulls. The second to fifth trials examined the potential for prolonged testicular infections resulting from natural exposure of seronegative bulls to persistently infected heifers. In the last five trials, the potential for viral transmission from bulls exhibiting prolonged testicular infections to a small number of exposed animals (n=28) was evaluated. Results of this research demonstrated that prolonged testicular infections could result in detection of viral RNA in semen for 2.75 years with infectious virus grown from testicular tissue 12.5 months after viral exposure. A type 1b strain of BVDV caused prolonged testicular infection after natural exposure of seronegative bulls to a persistently infected heifer. However, transmission of BVDV to susceptible animals was not detected in the final five trials of this research. In conclusion, BVDV can persist in testicular tissue after acute infection for several years, but the potential for viral transmission from these prolonged testicular infections appears to be low.

Bovine viral diarrhea virus (BVDV) associated with single in vivo-derived and in vitro-produced preimplantation bovine embryos following artificial exposure

The objective was to determine the average amount of bovine viral diarrhea virus (BVDV) associated with single in vivo-derived and in vitro-produced bovine embryos following recommended processing procedures for embryos. In vivo-derived and in vitro-produced bovine embryos at 7d post-fertilization were exposed (for 2h) to 2 x 10(5-7) cell culture infective dose (CCID(50))/mL of SD-1 (a noncytopathic, Type 1a strain of BVDV), and then washed according to International Embryo Transfer Society (IETS) guidelines prior to testing. Of the 87 in vivo-derived embryos tested, 27% were positive for virus by quantitative polymerase chain reaction (qPCR). The range in amount of virus associated with 99% of the contaminated embryos was <or=6.62+/-1.57 copies/5 microL; 90% of the contaminated embryos had <or=4.64+/-1.57 viral copies/5 microL of embryo-associated virus, using tolerance intervals (P<0.05). The SEM was 0.33 and the mean of averages was 1.12/5 microL. Of the 87 in vitro-produced embryos, 42% were positive for virus. The range in amount of virus associated with 99% of the contaminated embryos was <or=3.44+/-0.89 copies/5 microL; 90% of the contaminated embryos had <or=2.40+/-0.89 viral copies/5 microL of embryo-associated virus using tolerance intervals (P<0.05; S.E.M. was 0.14 and the mean of averages was 0.55/5 microL). Therefore, although many embryos were positive for virus, there were limited numbers of copies, thereby posing doubt regarding their potential for contamination following embryo transfer.

Antiviral Treatment of Calves Persistently Infected with Bovine Viral Diarrhoea Virus

Background: Animals persistently infected (PI) with bovine viral diarrhoea virus (BVDV) are a key source of viral propagation within and among herds. Currently, no specific therapy exists to treat PI animals. The purpose of this research was to initiate evaluation of the pharmacokinetic and safety data of a novel antiviral agent in BVDV-free calves and to assess the antiviral efficacy of the same agent in PI calves. Methods: One BVDV-free calf was treated with 2-(2-benzimidazolyl)-5-[4-(2-imidazolino)phenyl]furan dihydrochloride (DB772) once at a dose of 1.6 mg/kg intravenously and one BVDV-free calf was treated three times a day for 6 days at 9.5 mg/kg intravenously. Subsequently, four PI calves were treated intravenously with 12 mg/kg DB772 three times a day for 6 days and two PI control calves were treated with an equivalent volume of diluent only. Results: Prior to antiviral treatment, the virus isolated from each calf was susceptible to DB772 in vitro. The antiviral treatment effectively inhibited virus for 14 days in one calf and at least 3 days in three calves. Subsequent virus isolated from the three calves was resistant to DB772 in vitro. No adverse effects of DB772 administration were detected. Conclusions: Results demonstrate that DB772 administration is safe and exhibits antiviral properties in PI calves while facilitating the rapid development of viral resistance to this novel therapeutic agent.

Laboratory diagnosis and transmissibility of bovine viral diarrhea virus from a bull with a persistent testicular infection.

Recently, in the United States, a dairy bull was diagnosed as the second confirmed case of persistent testicular infection (PTI) with bovine viral diarrhea virus (BVDV). The first objective of this study was to evaluate the testing methodologies currently used by the artificial insemination industry in order to improve the detection of bulls with PTI. This study evaluated the impact of multiple factors ([1] sample tested, [2] sample handling, [3] assay used, and [4] assay methodology) on the sensitivity of detection of BVDV. The second objective of this study was to evaluate the transmissibility of BVDV from the bull through casual or sexual contact. Results from this study indicate that straws of semen should be transported to the diagnostic laboratory in liquid nitrogen dry shippers. PCR proved to be a more sensitive assay than virus isolation; however, certain PCR protocols exhibited greater diagnostic sensitivity than others. Insemination with cryopreserved semen from this infected bull caused viral transmission to a seronegative heifer resulting in viremia and seroconversion. After 42 months of age, the bull appeared to clear the infection. In conclusion, this bull validates that natural exposure to a 1a strain of BVDV can result in a unique PTI causing contamination of semen with detectable infectious virus. Appropriate handling and testing of samples is necessary in order to detect bulls exhibiting PTI. Additionally, PTI with BVDV may potentially be cleared after an extended duration.

Bovine viral diarrhea virus is inactivated when whole milk from persistently infected cows is heated to prepare semen extender

Bovine viral diarrhea virus (BVDV) can be present in cryopreserved bovine semen and be transmitted through artificial insemination. Because BVDV can be shed in milk, the virus might also be introduced as a contaminant of milk-based semen extenders. Thus, the purpose of this study was to evaluate the epidemiologic risk of using heated, BVDV-contaminated milk to prepare semen extender. Milk was obtained from cows free of and persistently infected (PI) with BVDV. Six replicates of milk samples were processed by heating (85-92.2 degrees C, 10min). Samples of milk collected before and after heating were assayed for BVDV. Additionally, milk was injected intravenously into eight BVDV seronegative calves to monitor for seroconversion and viral infection. Virus was not detected in any milk samples from negative animals. Virus was consistently isolated from unheated milk samples from PI cows by passage of somatic cells, ultracentrifugation, and animal inoculation. Virus was usually detected in these samples by RT-nPCR (reverse transcription nested polymerase chain reaction). In heated milk samples from PI cows, no infectious BVDV was detected using any technique, but viral RNA was detected using RT-nPCR in four of six replicates. Bovine viral diarrhea virus in milk from PI cows was inactivated by heating. Therefore, properly heated milk used in semen extenders will not result in transmission of infectious BVDV. Although RT-nPCR detected the presence of viral RNA in milk samples after heating, the virus was not infectious as demonstrated by lack of replication despite using multiple sensitive techniques.

Safety and efficacy of vaccination of seronegative bulls with modified-live, cytopathic bovine viral diarrhea viruses

The objectives were to vaccinate peri-pubertal bulls with a modified-live vaccine consisting of cytopathic BVDV strains Singer and 296 and evaluate the resulting: (a) transient shed of modified-live, cytopathic BVDV in semen; (b) risk of prolonged testicular infection; and (c) protection against subsequent testicular infection due to viral challenge. Seronegative, peri-pubertal bulls were vaccinated subcutaneously with a standard dose of vaccine (n=11) or were maintained as unvaccinated controls (n=11). Forty-nine days after vaccination, all bulls were intranasally inoculated with a noncytopathic field strain of BVDV. Semen and testicular biopsies collected after vaccination and challenge were assayed for BVDV using virus isolation, reverse transcription-nested PCR, or immunohistochemistry, and the identity of viral strains was determined by nucleotide sequencing of PCR products. Vaccination of peri-pubertal bulls with this vaccine caused a short-term, transient shed of only the type 1a strain of modified-live, cytopathic BVDV in semen for up to 10d after vaccination. The vaccine did not cause prolonged testicular infection. Vaccination with this product prevented development of prolonged testicular infections after subsequent exposure to a field strain of BVDV.

Efficacy of four commercially available multivalent modified-live virus vaccines against clinical disease, viremia, and viral shedding in early-weaned beef calves exposed simultaneously to cattle persistently infected with bovine viral diarrhea virus and cattle acutely infected with bovine herpesvirus 1.

OBJECTIVE To evaluate the efficacy of 4 commercially available multivalent modified-live virus vaccines against clinical disease, viremia, and viral shedding caused by bovine viral diarrhea virus (BVDV) and bovine herpesvirus 1 (BHV1) in early-weaned beef calves. ANIMALS 54 early-weaned beef steers (median age, 95 days). PROCEDURES Calves were randomly assigned to 1 of 5 groups and administered PBSS (group A [control]; n = 11) or 1 of 4 commercially available modified-live virus vaccines that contained antigens against BHV1, BVDV types 1 (BVDV1) and 2 (BVDV2), parainfluenza type 3 virus, and bovine respiratory syncytial virus (groups B [11], C [10], D [11], and E [11]). Forty-five days after vaccination, calves were exposed simultaneously to 6 cattle persistently infected with BVDV and 8 calves acutely infected with BHV1 for 28 days (challenge exposure). For each calf, serum antibody titers against BVDV and BHV1 were determined before vaccination and before and after challenge exposure. Virus isolation was performed on nasal secretions, serum, and WBCs at predetermined times during the 28-day challenge exposure. RESULTS None of the calves developed severe clinical disease or died. Mean serum anti-BHV1 antibody titers did not differ significantly among the treatment groups at any time and gradually declined during the study. Mean serum anti-BVDV antibody titers appeared to be negatively associated with the incidence of viremia and BVDV shedding. The unvaccinated group (A) had the lowest mean serum anti-BVDV antibody titers. The mean serum anti-BVDV antibody titers for group D were generally lower than those for groups B, C, and E. CONCLUSIONS AND CLINICAL RELEVANCE Results indicated differences in vaccine efficacy for the prevention of BVDV viremia and shedding in early-weaned beef calves.

Inactivation at various temperatures of bovine viral diarrhea virus in beef derived from persistently infected cattle.

Bovine viral diarrhea virus (BVDV) is a pestivirus that is enzootic in most cattle populations throughout the world. This virus is present throughout the body of persistently infected (PI) cattle. Previous research has not assessed the cooking temperature at which BVDV in meat from PI cattle can be inactivated. Therefore, muscle tissue from 6 PI cattle was harvested, refrigerated, frozen, and heated to various internal temperatures. The concentration of virus present was determined by virus isolation. Average cell culture infective doses (50% endpoint; CCID(50)) of BVDV per gram of frozen, uncooked meat from PI cattle were 10(5.85) CCID(50)/g of whole cuts and 10(6.02) CCID(50)/g of ground meat. The virus in whole and ground meat was consistently inactivated when cooked to temperatures greater than or equal to 75°C. A second objective of this research was to thoroughly reassess if Vero cells were permissive to BVDV infection in our laboratory to provide further indication of whether primates, including humans, might be susceptible to BVDV. Vero cells were not permissive to infection with any of 43 different strains of BVDV that readily replicated in Madin Darby bovine kidney cells. In conclusion, this bovine pathogen, which is not considered to be a human pathogen, can be inactivated by cooking ground or whole cuts of meat to 75°C or higher. Care should be taken to ensure that susceptible hosts such as pigs are not fed improperly cooked meat, meat by-products, or waste food originating from PI cattle.