scholarly journals Persistence and Reactivation of Bovine Herpesvirus 1 in the Tonsils of Latently Infected Calves

2000 ◽  
Vol 74 (11) ◽  
pp. 5337-5346 ◽  
Author(s):  
M. T. C. Winkler ◽  
A. Doster ◽  
C. Jones
Keyword(s):  
Ribonucleotide Reductase ◽  
Virus Transmission ◽  
Bovine Herpesvirus ◽  
Rt Pcr ◽  
Dexamethasone Treatment ◽  
Viral Dna ◽  
Bovine Herpesvirus 1 ◽  
Lymphoid Follicles ◽  
Latently Infected ◽  
Herpesvirus 1

ABSTRACT Bovine herpesvirus 1 (BHV-1), like other members of theAlphaherpesvirinae subfamily, establishes latent infection in sensory neurons. Reactivation from latency can occur after natural or corticosteroid-induced stress culminating in recurrent disease and/or virus transmission to uninfected animals. Our previous results concluded that CD4+ T cells in the tonsil and other adjacent lymph nodes are infected and undergo apoptosis during acute infection (M. T. Winkler, A. Doster, and C. Jones, J. Virol. 73:8657–8668, 1999). To test whether BHV-1 persisted in lymphoreticular tissue, we analyzed tonsils of latently infected calves for the presence of viral DNA and gene expression. BHV-1 DNA was consistently detected in the tonsils of latently infected calves. Detection of the latency-related transcript (LRT) in tonsils of latently infected calves required nested reverse transcription-PCR (RT-PCR) suggesting that only a few cells contained viral DNA or that LRT is not an abundant transcript. bICP0 (immediate-early and early transcripts), ribonucleotide reductase (early transcript), and glycoprotein C (late transcript) were not detected by RT-PCR in latently infected calves. When reactivation was initiated by dexamethasone, bICP0 and ribonucleotide reductase transcripts were detected. Following dexamethasone treatment, viral nucleic acid was detected simultaneously in trigeminal ganglionic neurons and lymphoid follicles of tonsil. LRT was detected at 6 and 24 h after dexamethasone treatment but not at 48 h. Dexamethasone-induced reactivation led to apoptosis that was localized to tonsillar lymphoid follicles. Taken together, these findings suggest that the tonsil is a site for persistence or latency from which virus can be reactivated by dexamethasone. We further hypothesize that the shedding of virus from the tonsil during reactivation plays a role in virus transmission.

scholarly journals Infection of Cattle with a Bovine Herpesvirus 1 Strain That Contains a Mutation in the Latency-Related Gene Leads to Increased Apoptosis in Trigeminal Ganglia during the Transition from Acute Infection to Latency

2003 ◽  
Vol 77 (8) ◽  
pp. 4848-4857 ◽  
Author(s):  
Luciane Lovato ◽  
Melissa Inman ◽  
Gail Henderson ◽  
Alan Doster ◽  
Clinton Jones
Keyword(s):  
Nasal Cavity ◽  
Acute Infection ◽  
Bovine Herpesvirus ◽  
Pcr Analysis ◽  
Viral Gene ◽  
Trigeminal Ganglia ◽  
Dexamethasone Treatment ◽  
Bovine Herpesvirus 1 ◽  
Latently Infected ◽  
Herpesvirus 1

ABSTRACT Bovine herpesvirus 1 (BHV-1) is an important pathogen of cattle and infection is usually initiated via the ocular or nasal cavity. After acute infection, the primary site for BHV-1 latency is sensory neurons in the trigeminal ganglia (TG). Reactivation from latency occurs sporadically, resulting in virus shedding and transmission to uninfected cattle. The only abundant viral transcript expressed during latency is the latency-related (LR) RNA. An LR mutant was constructed by inserting three stop codons near the beginning of the LR RNA. This mutant grows to wild-type (wt) efficiency in bovine kidney cells and in the nasal cavity of acutely infected calves. However, shedding of infectious virus from the eye and TG was dramatically reduced in calves infected with the LR mutant. Calves latently infected with the LR mutant do not reactivate after dexamethasone treatment. In contrast, all calves latently infected with wt BHV-1 or the LR rescued mutant reactivate from latency after dexamethasone treatment. In the present study, we compared the frequency of apoptosis in calves infected with the LR mutant to calves infected with wt BHV-1 because LR gene products inhibit apoptosis in transiently transfected cells. A sensitive TUNEL (terminal deoxynucleotidyltransferase-mediated dUTP-biotin nick end labeling) assay and an antibody that detects cleaved caspase-3 were used to identify apoptotic cells in TG. Both assays demonstrated that calves infected with the LR mutant for 14 days had higher levels of apoptosis in TG compared to calves infected with wt BHV-1 or to mock-infected calves. Viral gene expression, except for the LR gene, is extinguished by 14 days after infection, and thus this time frame is operationally defined as the establishment of latency. Real-time PCR analysis indicated that lower levels of viral DNA were present in the TG of calves infected with the LR mutant throughout acute infection. Taken together, these results suggest that the antiapoptotic properties of the LR gene play an important role during the establishment of latency.

scholarly journals Sequence Analysis of the UL39, UL38, and UL37 Homologues of Bovine Herpesvirus 1 and Expression Studies of UL40 and UL39, the Subunits of Ribonucleotide Reductase

Virology ◽  
1995 ◽  
Vol 212 (2) ◽  
pp. 734-740 ◽  
Author(s):  
Claire Simard ◽  
Isabelle Langlois ◽  
Dominik Styger ◽  
Bernd Vogt ◽  
Cestmir Vlcek ◽  
...  
Keyword(s):  
Sequence Analysis ◽  
Ribonucleotide Reductase ◽  
Bovine Herpesvirus ◽  
Bovine Herpesvirus 1 ◽  
Expression Studies ◽  
Herpesvirus 1

scholarly journals Phosphorylation of Bovine Herpesvirus 1 VP8 Plays a Role in Viral DNA Encapsidation and Is Essential for Its Cytoplasmic Localization and Optimal Virion Incorporation

2016 ◽  
Vol 90 (9) ◽  
pp. 4427-4440 ◽  
Author(s):  
Kuan Zhang ◽  
Robert Brownlie ◽  
Marlene Snider ◽  
Sylvia van Drunen Littel-van den Hurk
Keyword(s):  
Golgi Apparatus ◽  
Mammalian Cells ◽  
Cellular Localization ◽  
Bovine Herpesvirus ◽  
Wild Type ◽  
Viral Dna ◽  
Bovine Herpesvirus 1 ◽  
Virion Incorporation ◽  
Herpesvirus 1 ◽  
Two Stages

ABSTRACTVP8 is a major tegument protein of bovine herpesvirus 1 (BoHV-1) and is essential for viral replication in cattle. The protein undergoes phosphorylation after transcription through cellular casein kinase 2 (CK2) and a viral kinase, US3. In this study, a virus containing a mutated VP8 protein that is not phosphorylated by CK2 and US3 (BoHV-1-YmVP8) was constructed by homologous recombination in mammalian cells. When BoHV-1-YmVP8-infected cells were observed by transmission electron microscopy, blocking phosphorylation of VP8 was found to impair viral DNA encapsidation, resulting in release of incomplete viral particles to the extracellular environment. Consequently, less infectious virus was produced by the mutant virus than by wild-type (WT) virus. A comparison of mutant and WT VP8 by confocal microscopy revealed that mutant VP8 is nuclear throughout infection while WT VP8 is nuclear early during infection and is associated with the Golgi apparatus at later stages. This, together with the observation that mutant VP8 is present in virions, albeit in smaller amounts, suggests that the incorporation of VP8 may occur at two stages. The first takes place without the need for phosphorylation and before or during nuclear egress of capsids, whereas the second occurs in the Golgi apparatus and requires phosphorylation of VP8. The results indicate that phosphorylated VP8 plays a role in viral DNA encapsidation and in the secondary virion incorporation of VP8. To perform these functions, the cellular localization of VP8 is adjusted based on the phosphorylation status.IMPORTANCEIn this study, phosphorylation of VP8 was shown to have a function in BoHV-1 replication. A virus containing a mutated VP8 protein that is not phosphorylated by CK2 and US3 (BoHV-1-YmVP8) produced smaller numbers of infectious virions than wild-type (WT) virus. The maturation and egress of WT and mutant BoHV-1 were studied, showing a process similar to that reported for other alphaherpesviruses. Interestingly, lack of phosphorylation of VP8 by CK2 and US3 resulted in reduced incorporation of viral DNA into capsids during mutant BoHV-1 infection, as well as lower numbers of extracellular virions. Furthermore, mutant VP8 remained nuclear throughout infection, in contrast to WT VP8, which is nuclear at early stages and Golgi apparatus associated late during infection. This correlates with smaller amounts of mutant VP8 in virions and suggests for the first time that VP8 may be assembled into the virions at two stages, with the latter dependent on phosphorylation.

scholarly journals Identification of a Novel Bovine Herpesvirus 1 Transcript Containing a Small Open Reading Frame That Is Expressed in Trigeminal Ganglia of Latently Infected Cattle

2004 ◽  
Vol 78 (10) ◽  
pp. 5438-5447 ◽  
Author(s):  
Melissa Inman ◽  
Joe Zhou ◽  
Heather Webb ◽  
Clinton Jones
Keyword(s):  
Fusion Protein ◽  
Bovine Herpesvirus ◽  
Open Reading Frame ◽  
Trigeminal Ganglia ◽  
Bovine Herpesvirus 1 ◽  
Human Neuroblastoma ◽  
Reading Frame ◽  
Gfp Fusion Protein ◽  
Latently Infected ◽  
Herpesvirus 1

ABSTRACT Bovine herpesvirus 1 (BHV-1), like other Alphaherpesvirinae subfamily members, establishes latency in sensory neurons. The latency-related (LR) RNA is abundantly expressed during latency, and expression of an LR protein is required for the latency reactivation cycle in cattle. Within LR promoter sequences, a 135-amino-acid open reading frame (ORF) was identified, ORF-E, that is antisense to the LR RNA. ORF-E is also downstream of the gene encoding the major viral transcriptional activator, bICP0. Strand-specific reverse transcription-PCR demonstrated that a transcript containing ORF-E was consistently expressed in trigeminal ganglia (TG) of latently infected calves, productively infected cultured cells, and acutely infected calves. As expected, a late transcript encoding glycoprotein C was not detected in TG of latently infected calves. The ORF-E transcript is polyadenylated and is expressed early when cultured bovine cells are productively infected. Protein coding sequences containing ORF-E were fused to green fluorescent protein (GFP) to examine the cellular localization of the putative protein. In transiently transfected mouse neuroblastoma (neuro-2A) and human neuroblastoma (SK-N-SH) cells, the ORF-E/GFP fusion protein was detected in discreet domains within the nucleus. In contrast, the ORF-E/GFP fusion protein was detected in the cytoplasm and nucleus of rabbit skin cells and bovine kidney cells. As expected, the GFP protein was expressed in the cytoplasm and nucleus of transfected cells. These studies indicate that the ORF-E transcript is consistently expressed during latency. We suggest that the ORF-E gene regulates some aspect of the latency reactivation cycle.

scholarly journals Dexamethasone Treatment of Calves Latently Infected with Bovine Herpesvirus 1 Leads to Activation of the bICP0 Early Promoter, in Part by the Cellular Transcription Factor C/EBP-Alpha

2009 ◽  
Vol 83 (17) ◽  
pp. 8800-8809 ◽  
Author(s):  
Aspen Workman ◽  
Sandra Perez ◽  
Alan Doster ◽  
Clinton Jones
Keyword(s):  
Promoter Activity ◽  
Bovine Herpesvirus ◽  
Mutant Virus ◽  
Bovine Herpesvirus 1 ◽  
Cellular Transcription Factor ◽  
Early Promoter ◽  
Latently Infected ◽  
Herpesvirus 1 ◽  
Factor C ◽  
Cellular Transcription

ABSTRACT Sensory neurons within trigeminal ganglia (TG) are the primary site for bovine herpesvirus 1 (BHV-1) latency. During latency, viral gene expression is restricted to the latency-related (LR) gene and the open reading frame ORF-E. We previously constructed an LR mutant virus that expresses LR RNA but not any of the known LR proteins. In contrast to calves latently infected with wild-type (wt) BHV-1 or the LR rescued virus, the LR mutant virus does not reactivate from latency following dexamethasone (DEX) treatment. In this study, we demonstrated that bICP0, but not bICP4, transcripts were consistently detected in TG of calves infected with the LR mutant or LR rescued virus following DEX treatment. Calves latently infected with the LR rescued virus but not the LR mutant virus expressed late transcripts, which correlated with shedding of infectious virus following DEX treatment. The bICP4 and bICP0 genes share a common immediate-early promoter, suggesting that this promoter was not consistently activated during DEX-induced reactivation from latency. The bICP0 gene also contains a novel early promoter that was activated by DEX in mouse neuroblastoma cells. Expression of a cellular transcription factor, C/EBP-alpha, was stimulated by DEX, and C/EBP-alpha expression was necessary for DEX induction of bICP0 early promoter activity. C/EBP-alpha directly interacted with bICP0 early promoter sequences that were necessary for trans activation by C/EBP-alpha. In summary, DEX treatment of latently infected calves induced cellular factors that stimulated bICP0 early promoter activity. Activation of bICP0 early promoter activity does not necessarily lead to late gene expression and virus shedding.

scholarly journals Impairment on nuclear maturation rate in oocytes from cows naturally infected by bovine herpesvirus 1 (BoHV-1)

2018 ◽  
Vol 38 (12) ◽  
pp. 2207-2212
Author(s):  
Vívian R.A. Mendes ◽  
Eduardo P. Costa ◽  
Vanessa L.D. Queiroz ◽  
Abelardo Silva Júnior ◽  
Saullo V.P. Alves ◽  
...  
Keyword(s):  
Bovine Herpesvirus ◽  
Cumulus Cells ◽  
Control Group ◽  
Viral Dna ◽  
Bovine Herpesvirus 1 ◽  
Nuclear Maturation ◽  
Maturation Rate ◽  
Developmental Capacity ◽  
Herpesvirus 1

ABSTRACT Bovine herpesvirus 1 (BoHV-1) is an important bovine pathogen that is responsible for causing respiratory diseases and reproductive failures. The presence of BoHV-1 in an in vitro embryo production system affects fertilization, maturation, and embryonic development. The objective of this study was to evaluate the developmental capacity of oocytes from naturally infected cows with no reproductive history. Moreover, this study investigated the presence of viral DNA in cumulus oophorus complexes (COCs). Experimental groups were differentiated by titrating the antibodies detected through seroneutralization assays, establishing three groups: seronegative animals (titer lower than 2), low titer (2 to 8), and animals with a titer above or equal to 16. COCs were obtained from 15 donors during 22 sessions of ultrasound-guided follicular aspiration. DNA was extracted from a pool of COCs obtained from all aspirations from the same donor as well as from whole blood and nested PCR reactions were performed. Only COCs with a compact layer of cumulus cells, an intact zona pellucida, and homogeneous cytoplasm were selected for in vitro culture and evaluation of nuclear maturation rate. After culturing for 24 hours, the oocytes were fixed and stained to evaluate the meiotic cell cycle stage. Oocytes that showed a chromosomal configuration in metaphase II were considered to have reached nuclear maturation. Compared with the other groups, the oocyte nuclear maturation rate in animals with a titer greater than or equal to 16 (50%) was compromised (P< 0.05). However, the viral titer did not influence the maturation rate of bovine oocytes in animals exhibiting low titration (62.2%) when compared with the control group (76.7%). Viral DNA was not observed in the blood samples but was detected in the COC pool from three seropositive donors. In view of the results obtained, we conclude that natural infections by the BoHV-1 virus can compromise the nuclear maturation rate in cows, depending on the titration levels of antibodies against the virus. Moreover, viral DNA could be present in COCs, contradicting the hypothesis that seropositive animals with no history of clinical symptomatology pose a negligible risk of transmitting BoHV-1 by COCs.

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