scholarly journals A bovine herpesvirus 1 pUL51 deletion mutant shows impaired viral growth in vitro and reduced virulence in rabbits

Oncotarget ◽  
2016 ◽  
Vol 7 (11) ◽  
pp. 12235-12253 ◽  
Author(s):  
Sohail Raza ◽  
Mingliang Deng ◽  
Farzana Shahin ◽  
Kui Yang ◽  
Changmin Hu ◽  
...  
Keyword(s):  
Deletion Mutant ◽  
Bovine Herpesvirus ◽  
Bovine Herpesvirus 1 ◽  
Viral Growth ◽  
Herpesvirus 1

scholarly journals Bovine Herpesvirus 1 Tegument Protein VP22 Interacts with Histones, and the Carboxyl Terminus of VP22 Is Required for Nuclear Localization

2001 ◽  
Vol 75 (17) ◽  
pp. 8251-8258 ◽  
Author(s):  
Xiaodi Ren ◽  
Jerome S. Harms ◽  
Gary A. Splitter
Keyword(s):  
Nuclear Localization ◽  
Deletion Mutant ◽  
Bovine Herpesvirus ◽  
Mutant Virus ◽  
Tegument Protein ◽  
Carboxyl Terminus ◽  
Slight Reduction ◽  
Bovine Herpesvirus 1 ◽  
Herpesvirus 1

ABSTRACT The bovine herpesvirus 1 (BHV-1) UL49 gene encodes a viral tegument protein termed VP22. UL49 homologs are conserved among alphaherpesviruses. Interestingly, the BHV-1 VP22 deletion mutant virus is asymptomatic and avirulent in infected cattle but produces only a slight reduction in titer in vitro. Attenuation of the BHV-1 VP22 deletion mutant virus in vivo suggests that VP22 plays a functional role in BHV-1 replication. In herpes simplex virus type 1, the VP22 homolog was previously shown to interact with another tegument protein,VP16, the α-transinducing factor in vitro. In this report, we show that (i) the nuclear targeting of VP22 is independent of other viral factors, (ii) the carboxyl terminus of VP22 is required for its nuclear localization, (iii) VP22 associates with histones and nucleosomes, (iv) an antihistone monoclonal antibody cross-reacts with VP22, and (v) acetylation of histone H4 is decreased in VP22-expressing cells as well as virus-infected cells. Our data suggest that VP22 may have a modulatory function during BHV-1 infection.

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.

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.

scholarly journals Bovine Herpesvirus 1 UL3.5 Interacts with Bovine Herpesvirus 1 α-Transinducing Factor

2000 ◽  
Vol 74 (6) ◽  
pp. 2876-2884 ◽  
Author(s):  
Ngan Lam ◽  
Geoffrey J. Letchworth
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Bovine Herpesvirus ◽  
Mass Spectrometry Analysis ◽  
Bovine Herpesvirus 1 ◽  
Subcellular Compartment ◽  
Transfected Cells ◽  
Infected Cells ◽  
Herpesvirus 1

ABSTRACT The bovine herpesvirus 1 (BHV-1) UL3.5 gene encodes a 126-amino-acid tegument protein. Homologs of UL3.5 are present in some alphaherpesviruses and have 20 to 30% overall amino acid homology that is concentrated in the N-terminal 50 amino acids. Mutant pseudorabies virus lacking UL3.5 is deficient in viral egress but can be complemented by BHV-1 UL3.5 (W. Fuchs, H. Granzow, and T. C. Mettenleiter, J. Virol. 71:8886–8892, 1997). The function of BHV-1 UL3.5 in BHV-1 replication is not known. To get a better understanding of its function, we sought to identify the proteins that interact with the BHV-1 UL3.5 protein. By using an in vitro pull-down assay and matrix-assisted laser desorption ionization mass spectrometry analysis, we identified BHV-1 α-transinducing factor (αBTIF) as a BHV-1 UL3.5-interacting protein. The interaction was verified by coimmunoprecipitation from virus-infected cells using an antibody to either protein, by indirect immunofluorescence colocalization in both virus-infected and transfected cells, and by the binding of in vitro-translated proteins. In virus-infected cells, UL3.5 and αBTIF colocalized in a Golgi-like subcellular compartment late in infection. In transfected cells, they colocalized in the nucleus. Deletion of 20 amino acids from the N terminus of UL3.5, but not 40 amino acids from the C terminus, abolished the UL3.5-αBTIF interaction both in vitro and in vivo. The interaction between UL3.5 and αBTIF may be important for BHV-1 maturation and regulation of αBTIF transactivation activity.

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.

scholarly journals Bovine Herpesvirus 1 Glycoprotein M Forms a Disulfide-Linked Heterodimer with the UL49.5 Protein

1998 ◽  
Vol 72 (4) ◽  
pp. 3029-3036 ◽  
Author(s):  
S. X. Wu ◽  
X. P. Zhu ◽  
G. J. Letchworth
Keyword(s):  
Disulfide Bonds ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Bovine Herpesvirus ◽  
In Vitro Translation ◽  
Bovine Herpesvirus 1 ◽  
Western Blots ◽  
Sds Page ◽  
Herpesvirus 1

ABSTRACT Nine glycoproteins (gB, gC, gD, gE, gG, gH, gI, gK, and gL) have been identified in bovine herpesvirus 1 (BHV-1). gM has been identified in many other alpha-, beta-, and gammaherpesviruses, in which it appears to play a role in membrane penetration and cell-to-cell fusion. We sought to express BHV-1 open reading frame UL10, which encodes gM, and specifically identify the glycoprotein. We corrected a frameshift error in the published sequence and used the corrected sequence to design coterminal peptides from the C terminus. These were expressed as glutathione S-transferase fusion proteins inEscherichia coli. The fusion protein containing the 63 C-terminal amino acids from the corrected gM sequence engendered antibodies that immunoprecipitated a 30-kDa protein from in vitro translation reactions programmed with the UL10 gene. Proteins immunoprecipitated by this antibody from virus-infected cells ran at 36 and 43 kDa in reducing sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and 43 and 48 kDa in nonreducing SDS-PAGE. Only the larger of the pair was present in virions. A 7-kDa protein was released from gM by reducing agents. The 7-kDa protein was not recognized in Western blots probed with the anti-gM antibody but reacted specifically with antibodies prepared against BHV-1 UL49.5, previously reported to be a 9-kDa protein associated with an unidentified 39-kDa protein (X. Liang, B. Chow, C. Raggo, and L. A. Babiuk, J. Virol. 70:1448–1454, 1996). This is the first report of a small protein covalently bound to any herpesvirus gM. Similar patterns of hydrophobic domains and cysteines in all known gM and UL49.5 homologs suggest that these two proteins may be linked by disulfide bonds in all herpesviruses.

238 EFFICACY OF RECOMBINANT TRYPSIN AGAINST BOVINE HERPESVIRUS-1 ASSOCIATED WITH IN VITRO-DERIVED PORCINE EMBRYOS

2007 ◽  
Vol 19 (1) ◽  
pp. 234 ◽  
Author(s):  
M. S. D. Marley ◽  
M. D. Givens ◽  
P. K. Galik ◽  
K. P. Riddell ◽  
D. A. Stringfellow
Keyword(s):  
Virus Isolation ◽  
Bovine Herpesvirus ◽  
Positive Control ◽  
Negative Control ◽  
Animal Origin ◽  
Bovine Herpesvirus 1 ◽  
Herpesvirus 1 ◽  
Porcine Origin

TrypLETM (Invitrogen, Carlsbad, CA, USA) is a recombinant, fungal, trypsin-like protease that is used as a substitute for porcine-origin trypsin in cell culture procedures. It is stable at room temperature and does not present the same risk of contamination as animal-origin trypsin. Previously, TrypLE SelectTM (10X) was shown to remove bovine herpesvirus-1 (BHV-1) from Day 7 in vivo-derived embryos (Marley et al. 2006 Reprod. Fert. Dev. 18, 213–214). The objective of this study was to determine if the same treatment would effectively remove BHV-1 from Day 7 zona pellucida-intact, in vitro-derived porcine embryos. Day 7 in vitro-derived morulae and blastocysts and non-fertile or degenerate embryos (NFD) were washed according to the International Embryo Transfer Society protocol. One group of 10 NFD was not exposed to virus and served as the negative control. The remaining embryos and 10 NFD were exposed to 106–108 PFU/mL BHV-1 (Colorado strain) for 1 h. Following exposure, one group of 10 NFD was washed and served as the positive control. The remaining developed embryos were divided into groups of 10 and washed and treated as described in Table 1. Following treatment, the embryos were sonicated in groups of 5 and assayed by virus isolation. The negative control embryos, as well as the embryos treated with porcine-origin trypsin, TrypLE Select (10X) for 7 min, and TrypLE Select (10X) diluted 1 : 2 for 10 min, were negative for virus. The positive control embryos in addition to the other treatments were positive on virus isolation (Table 1). Although, TrypLE Select (10X) does have some antiviral effect when used for 10 min, it was not completely effective, as shown by the positive virus isolation results of one group of 10 embryos. The groups treated with TrypLE Select (10X) diluted 1 : 2 for 10 min were negative for virus; however, if a larger sample size had been tested, positive groups might have occurred. Though using a recombinant trypsin product would be beneficial over using an animal-origin product, it is not known if TrypLE Select (10X) would render a single IVF embryo free of infectious virus. Further research would also need to be performed to assess the viability of embryos following treatment with TrypLE Select (10X). In addition, other recombinant trypsin products need to be evaluated to determine their efficacy against BHV-1 associated with IVF embryos. Table 1.Effect of recombinant trypsin-like proteases on BHV-1 virus in porcine embryos

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