scholarly journals High-level expression of biologically active bovine alpha interferon by Bovine herpesvirus 1 interferes only marginally with recombinant virus replication in vitro

2005 ◽  
Vol 86 (10) ◽  
pp. 2685-2695 ◽  
Author(s):  
Constanze Höhle ◽  
Axel Karger ◽  
Patricia König ◽  
Katrin Giesow ◽  
Günther M. Keil
Keyword(s):  
Cell Culture ◽  
Bovine Herpesvirus ◽  
Biologically Active ◽  
Expression Cassette ◽  
Alpha Interferon ◽  
Cell Culture Supernatant ◽  
Bovine Herpesvirus 1 ◽  
Glycoprotein E ◽  
Herpesvirus 1

An artificial open reading frame (ORF) for bovine alpha interferon (boIFN-α) with the codon preference of Bovine herpesvirus 1 (BHV-1) glycoprotein B was constructed to assess the effect of expression of boIFN-α by BHV-1 from an expression cassette. Transient expression of the ORF revealed that transfected cells secreted substantial amounts of biologically active boIFN-α, which moderately inhibited replication of BHV-1 after stimulation of bovine cells with 104 U ml−1. The boIFN-α-encoding expression cassette was recombined into the glycoprotein E locus of the glycoprotein E-negative BHV-1 vaccine strain GKD. Cells infected with the resulting recombinant BHV-1/boIFN-α secreted up to 107 U boIFN-α per ml cell culture supernatant, which is about 40- to more than 100-fold the activity reached with other virus expression systems. Bioassays demonstrated that the BHV-1-expressed interferon induced a rapid and sustained antiviral state in stimulated bovine cells. Analysis of the in vitro growth properties of the recombinant revealed, depending on the cell line used, no or only slight inhibition in direct spreading from cell to cell and a modest delay in virus egress from infected cells. Final titres, however, were comparable to those reached by the parent strain. Penetration into cells was not affected. The results from this study demonstrate that BHV-1/boIFN-α expresses high levels of boIFN-α, grows to high titres in cell culture and thus represents a potential alternative means to deliver endogenously produced boIFN-α in situ for a period of time.

211 LACTOFERRIN INHIBITS BOVINE HERPESVIRUS-1 IN CELL CULTURE AND ALLOWS NORMAL DEVELOPMENT OF IN VITRO-PRODUCED EMBRYOS

2006 ◽  
Vol 18 (2) ◽  
pp. 213 ◽  
Author(s):  
M. Givens ◽  
M. Marley ◽  
P. Galik ◽  
K. Riddell ◽  
D. Stringfellow
Keyword(s):  
Cell Culture ◽  
Cell Count ◽  
Bovine Milk ◽  
Control Sample ◽  
Bovine Herpesvirus ◽  
Blastocyst Development ◽  
Bovine Herpesvirus 1 ◽  
Herpesvirus 1 ◽  
In Vitro Embryo Production

Lactoferrin is an iron-binding glycoprotein found in milk, saliva, tears, and other exocrine secretions. It is known to have in vitro antiviral effects against human, feline, and canine herpesviruses. In addition, lactoferrin is known to be safe in cell culture. Bovine herpesvirus-1 (BHV-1) is a likely contaminant of in vitro embryo production. Further, trypsin treatment is not completely effective in removing the virus from these embryos. We hypothesized that a nontoxic concentration of lactoferrin might prevent replication of BHV-1 within in vitro embryo production systems. Thus, the specific objectives of this research were to determine if lactoferrin from bovine milk would inhibit BHV-1 in cell culture and to determine if in vitro-produced embryos could develop normally when cultured in lactoferrin. Two-fold dilutions of lactoferrin (from 10 to 0.625 mg/mL) were added to Madin Darby bovine kidney cells, followed in 15 min by the addition 104 PFU/mL of BHV-1 (Colorado strain). Samples of cell lysate were taken at Day 2 and virus was quantified by plaque assay. The percent of virus inhibited by the antiviral agent at each concentration was determined by comparison to equivalent samples from temporal control cultures in which no compound was added before or after inoculation (Percentage of virus inhibited = [Quantity of virus in the control sample - Quantity of virus in the compound sample]/Quantity of virus in the control sample � 100). Next, the effect of lactoferrin was determined on in vitro-produced embryos. Cumulus oocyte complexes were received from an abattoir, matured in transit, placed in fertilization drops for 6 h, and then placed in culture drops containing lactoferrin (10, 5, and 2.5 mg/mL). At Day 3.5, embryos > 4 cell stage were placed into fresh culture drops containing lactoferrin. On Day 7.5, blastocyst development was noted and the developed embryos were stained to count viable cells. Blastocyst development rate and nucleated cell count of the treated embryos were compared to those of the controls using Chi square test, and ANOVA and Tukey-Kramer HSD, respectively. Lactoferrin (10 mg/mL) inhibited 2 to 5 logs of virus. At concentrations of 5 and 2.5 mg/mL, 1 to 3 logs of virus were inhibited, and concentrations of 1.25 and 0.625 mg/mL inhibited 0 to 2 logs of virus. Lactoferrin did not affect the nucleated cell count of the treated embryos. In addition, unlike 10 and 5 mg/mL, 2.5 mg/mL of lactoferrin did not affect blastocyst development. These preliminary results indicate that lactoferrin from bovine milk can significantly inhibit BHV-1 in cell culture. Furthermore, supplementation of in vitro culture with 2.5 mg/mL of lactoferrin does not affect blastocyst development or cell count of in vitro-produced embryos.

scholarly journals Intraspecific bovine herpesvirus 1 recombinants carrying glycoprotein E deletion as a vaccine marker are virulent in cattle

2006 ◽  
Vol 87 (8) ◽  
pp. 2149-2154 ◽  
Author(s):  
Benoît Muylkens ◽  
François Meurens ◽  
Frédéric Schynts ◽  
Frédéric Farnir ◽  
Aldo Pourchet ◽  
...  
Keyword(s):  
High Frequency ◽  
Bovine Herpesvirus ◽  
Clinical Score ◽  
Natural Host ◽  
Field Strain ◽  
Bovine Herpesvirus 1 ◽  
Glycoprotein E ◽  
Control Programmes ◽  
Herpesvirus 1

Vaccines used in control programmes of Bovine herpesvirus 1 (BoHV-1) utilize highly attenuated BoHV-1 strains marked by a deletion of the glycoprotein E (gE) gene. Since BoHV-1 recombinants are obtained at high frequency in experimentally coinfected cattle, the consequences of recombination on the virulence of gE-negative BoHV-1 were investigated. Thus, gE-negative BoHV-1 recombinants were generated in vitro from several virulent BoHV-1 and one mutant BoHV-1 deleted in the gC and gE genes. Four gE-negative recombinants were tested in the natural host. All the recombinants were more virulent than the gE-negative BoHV-1 vaccine and the gC- and gE-negative parental BoHV-1. The gE-negative recombinant isolated from a BoHV-1 field strain induced the highest severe clinical score. Latency and reactivation studies showed that three of the recombinants were reexcreted. Recombination can therefore restore virulence of gE-negative BoHV-1 by introducing the gE deletion into a different virulence background.

EvaGreen-based Multiplex Real-time PCR Assay for Rapid Differentiation of Wild-Type and Glycoprotein E-Deleted Bovine Herpesvirus-1 Strains

2017 ◽  
Vol 28 (4) ◽  
pp. 248-252 ◽  
Author(s):  
Sachin S. Pawar ◽  
Chetan D. Meshram ◽  
Niraj K. Singh ◽  
Mohini Saini ◽  
B. P. Mishra ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Bovine Herpesvirus ◽  
Wild Type ◽  
Pcr Assay ◽  
Bovine Herpesvirus 1 ◽  
Glycoprotein E ◽  
Herpesvirus 1

scholarly journals Safety and immunogenicity of a glycoprotein E gene-deleted bovine herpesvirus 1 strain as a candidate vaccine strain

2016 ◽  
Vol 36 (11) ◽  
pp. 1067-1074
Author(s):  
Marcelo Weiss ◽  
◽  
Deniz Anziliero ◽  
Mathias Martins ◽  
Rudi Weiblen ◽  
...  
Keyword(s):  
Acute Infection ◽  
Clinical Signs ◽  
Bovine Herpesvirus ◽  
Elisa Test ◽  
Bovine Herpesvirus 1 ◽  
Glycoprotein E ◽  
Virus Shedding ◽  
Group I ◽  
Virus Dose ◽  
Herpesvirus 1

ABSTRACT: A glycoprotein E-deleted Brazilian bovine herpesvirus 1 (BoHV-1gEΔ) was tested regarding to safety and immunogenicity. Intramuscular inoculation of young calves with a high virus dose did not result in clinical signs or virus shedding during acute infection or after dexamethasone administration. Calves vaccinated once IM (group I) or subcutaneously (group II) with live BoHV-1gEΔ or twice with inactivated virus plus aluminum hydroxide (group IV) or Montanide™ (group V) developed VN titers of 2 to 8 (GMT:2); 2 to 4 (GMT:1.65); 2 to 16 (GMT:2.45) and 2 to 128 (GMT:3.9), respectively. All BoHV-1gEΔ vaccinated calves remained negative in an anti-gE ELISA. Lastly, six young calves vaccinated with live BoHV-1gEΔ and subsequently challenged with a virulent BoHV-1 strain shed less virus and developed only mild and transient nasal signs comparing to unvaccinated calves. Thus, the recombinant BoHV-1gEΔ is safe and immunogenic for calves and allows for serological differentiation by a gE-ELISA test.

scholarly journals A glycoprotein E gene-deleted bovine herpesvirus 1 as a candidate vaccine strain

2015 ◽  
Vol 48 (9) ◽  
pp. 843-851 ◽  
Author(s):  
M. Weiss ◽  
M.C.S. Brum ◽  
D. Anziliero ◽  
R. Weiblen ◽  
E.F. Flores
Keyword(s):  
Vaccine Strain ◽  
Bovine Herpesvirus ◽  
Candidate Vaccine ◽  
Bovine Herpesvirus 1 ◽  
Glycoprotein E ◽  
E Gene ◽  
Herpesvirus 1

148 RISK OF TRANSMISSION OF BOVINE HERPESVIRUS (BHV-1) BY INFECTED SEMEN TO RECIPIENTS AND EMBRYOS

2009 ◽  
Vol 21 (1) ◽  
pp. 173 ◽  
Author(s):  
A. Bielanski ◽  
A. Lalonde ◽  
J. Algire
Keyword(s):  
Reproductive Tract ◽  
Bovine Herpesvirus ◽  
Corpora Lutea ◽  
Bovine Herpesvirus 1 ◽  
Bull Semen ◽  
Stage Embryo ◽  
Morula Stage ◽  
Herpesvirus 1 ◽  
Isolation Test

Bovine herpesvirus-1 (BHV-1) causes a variety of economically important respiratory and reproductive problems, the latter including vulvovaginitis, endometritis and infertility. For that reason, several countries have eradicated the disease and others have schemes in progress to achieve freedom. Although there is a considerable amount of information about the risk of BHV-1 transmission through contaminated semen used for artificial insemination, there is no available evidence to indicate whether the resulting embryos, when used for embryo transter (ET), can lead to the transmission of BHV-1 to recipients and offspring. For this experiment, bull semen contaminated in vitro with BHV-1 at 102 TCID50 mL–1 (Colorado strain) and then cryopreserved was used for insemination (2 times at estrus) of BHV-1 seronegative, superovulated heifers (N = 18). Embryos were collected postmortem 7 days post-insemination and were washed according to the IETS recommendations (however without trypsin treatment) or left unwashed. On 4 occasions, washed embryos were transferred to BHV-1 seronegative recipients. The remaining embryos and other samples collected from the reproductive tract were tested for BHV-1 presence using the standard virus isolation test. In total, out of 144 unfertilized oocytes and embryos collected, 9 were ET quality. Most of the embryos were degenerated (N = 79) or unfertilized (N = 56). The 4 heifers, which each received a single morula-stage embryo, maintained seronegative status, but did not become pregnant. BHV-1 was detected in 43% (23/53) unwashed and 0% (0/57) of washed embryos, 78% (14/18) of follicular fluid samples, 89% (16/18) of oviductal epithelial cells, 78% (14/18) of endometrium, and 89% (16/18) of corpora lutea tissues. Results herein suggest that BHV-1 can be transmitted by infected semen to embryo donors. The resulting unwashed embryos may remain infectious. However, whether BHV-1 uninfected offspring can be produced by ET of BHV-1 contaminated embryos that are washed according to the IETS guidelines, remains to be determined.

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