A possible case of caprine-associated malignant catarrhal fever in a domestic water buffalo (Bubalus bubalis) in Switzerland

Background

Interspecific recombinant viruses R1ΔgC and R2ΔgI were isolated after in vitro co-infection with BoHV-1 and BoHV-5, two closely related alphaherpesviruses that infect cattle. The genetic characterization of R1ΔgC and R2ΔgI showed that they are composed of different sections of the parental genomes. The aim of this study was the characterization of the in vivo behavior of these recombinants in the natural host.

Results

Four groups of four 3-month-old calves of both genders were intranasally inoculated with either the recombinant or parental viruses. A control group of two animals was also included. Viral excretion and clinical signs were monitored after infection. Histopathological examination of the central nervous system (CNS) was performed and the establishment of latency in trigeminal ganglia was analyzed by PCR. The humoral response was also evaluated using ELISA tests. Three out of four animals from the BoHV-5 infected group excreted virus for 4-10 days. Two calves shed R1ΔgC virus for one day. In R2ΔgI and BoHV-1.2ΔgCΔgI groups, infectious virus was isolated only after two or three blind passages. None of the infected animals developed neurological signs, although those infected with BoHV-5 showed histopathological evidence of viral infection. Latent viral DNA was detected in at least one calf from each infected group. Serum and/or mucosal antibodies were detected in all groups.

Conclusion

Both BoHV-1/-5 recombinants and the BoHV-1 parental strain are attenuated in calves, although they are able to replicate in animals at low rates and to establish latent infections.     

Rise and survival of bovine herpesvirus 1 recombinants after primary infection and reactivation from latency

Recombination is thought to be an important source of genetic variation in herpesviruses. Several studies, performed in vitro or in vivo, detected recombinant viruses after the coinoculation of two distinguishable strains of the same herpesvirus species. However, none of these studies investigated the evolution of the relative proportions of parental versus recombinant progeny populations after coinoculation of the natural host, both during the excretion and the reexcretion period. In the present study, we address this by studying the infection of cattle with bovine herpesvirus 1 (BoHV-1). The recombination of two BoHV-1 mutants lacking either glycoprotein C (gC(-)/gE(+)) or E (gC(+)/gE(-)) was investigated after inoculation of cattle by the natural route of infection. The results demonstrated that (i) recombination is a frequent event in vivo since recombinants (gC(+)/gE(+) and gC(-)/gE(-)) were detected in all coinoculated calves, (ii) relative proportions of progeny populations evolved during the excretion period toward a situation where two populations (gC(+)/gE(+) and gC(-)/gE(+)) predominated without fully outcompeting the presence of the two other detected populations (gC(+)/gE(-) and gC(-)/gE(-)), and (iii) after reactivation from latency, no gC(+)/gE(-) and gC(-)/gE(-) progeny viruses were detected, although gC(+)/gE(-) mutants, when inoculated alone, were detected after reactivation treatment. In view of these data, the importance of gE in the biology of BoHV-1 infection and the role of recombination in herpesvirus evolution are discussed.     

PCR detection of bovine herpesviruses from nonbovine ruminants in Hungary

Polymerase chain reaction (PCR) was used to test six different nonbovine ruminant species for five bovine herpesviruses including infectious bovine rhinotracheitis virus (BoHV-1), bovine herpes mammillitis virus (BoHV-2), Movar-type herpesvirus (BoHV-4), bovine herpesvirus type 5 (BoHV-5), and alcelaphine herpesvirus 1 (AlHV-1). Species tested included 56 roe deer (Capreolus capreolus), 66 red deer (Cervus elaphus), 20 fallow deer (Dama dama), 16 mouflon (Ovis musimon), 34 domestic sheep, and 44 domestic goats, which were sampled in Hungary in 2003. Tracheal and popliteal lymph nodes collected from these animals were tested for the presence of the five bovine herpesviruses using three nested (two of which were duplex) PCR assays. Three bovine herpesviruses (BoHV-1, -2, and -4) were detected, whereas no evidence of AlHV-1 or BoHV-5 was observed. Prevalence of BoHV-1 ranged from 12% to 47%, and PCR-positive results were observed in all species tested. BoHV-2 was detected from roe deer, red deer, fallow deer, mouflon, and domestic sheep, and the prevalence in these species ranged from 3% to 50%. BoHV-4 was detected in all species, with prevalence ranging from 12% to 69%. Sequenced PCR products were 99-100% identical to bovine herpesviral sequences deposited in the GenBank.     

Detection of bovine herpesvirus 1 and 5 in semen from Brazilian bulls

Bovine herpesvirus 1 (BoHV-1) and 5 (BoHV-5) are important pathogens of the respiratory and genital tract of cattle and may also affect the central nervous system and cause meningoencephalitis. Both virus types are estimated to be widely distributed in Southern Brazil. In the present study, BoHV-1 and/or BoHV-5 DNA were detected in bovine semen samples from two states of Brazil by two species-specific nested polymerase chain reactions (nPCRs). These nPCRs were used to assay 53 samples of fresh semen and 23 samples of frozen semen from breeding bulls. Viral DNA was detected in all 76 semen samples: all were positive for BoHV-5, whereas 34 of these were positive for BoHV-1 as well. Moreover, in five fresh and in 13 frozen semen samples—of a total number of 40 samples suitable for virus isolation—infectious BoHV-1 and/or BoHV-5 virus were detected. In conclusion, that both BoHV-1 and BoHV-5 were detected in bovine semen in Brazil highlighted the importance of examining bull semen in search for both agents to reduce the risk of transmitting these viruses.     

Superinfection prevents recombination of the alphaherpesvirus bovine herpesvirus 1

Homologous recombination between strains of the same alphaherpesvirus species occurs frequently both in vitro and in vivo. This process has been described between strains of herpes simplex virus type 1, herpes simplex virus type 2, pseudorabies virus, feline herpesvirus 1, varicella-zoster virus, and bovine herpesvirus 1 (BoHV-1). In vivo, the rise of recombinant viruses can be modulated by different factors, such as the dose of the inoculated viruses, the distance between inoculation sites, the time interval between inoculation of the first and the second virus, and the genes in which the mutations are located. The effect of the time interval between infections with two distinguishable BoHV-1 on recombination was studied in three ways: (i) recombination at the level of progeny viruses, (ii) interference induced by the first virus infection on β-galactosidase gene expression of a superinfecting virus, and (iii) recombination at the level of concatemeric DNA. A time interval of 2 to 8 h between two successive infections allows the establishment of a barrier, which reduces or prevents any successful superinfection needed to generate recombinant viruses. The dramatic effect of the time interval on the rise of recombinant viruses is particularly important for the risk assessment of recombination between glycoprotein E-negative marker vaccine and field strains that could threaten BoHV-1 control and eradication programs.     

Virologic investigations of free-living European bison (Bison bonasus) from the Bialowieza Primeval Forest,Poland

We conducted virologic investigations on postmortem specimens from 261 free-living European bison (Bison bonasus) from the Bialowieza Primeval Forest, Poland collected between 1990 and 2000. Fifty-four of 94 males had balanoposthitis; none of the 167 female bison examined had reproductive tract lesions. Peripheral blood, swabs, and various tissues were analyzed for bovine viruses as well as for viral DNA by bovine herpesvirus 1 (BoHV-1) and bovine herpesvirus 4 (BoHV-4) specific polymerase chain reaction (PCR) technique. An infectious bovine rhinotracheitis like BoHV-1 strain was isolated from the spleen of a female bison calf and additionally was detected by nested PCR from splenic tissue. None of the bison had significant antibody titers against BoHV-1, bovine herpesvirus 2, BoHV-4, caprine herpesvirus 1, cervid herpesvirus 1, or bovine viral diarrhea (BVD) virus-1. However, low antibody titers in two animals indicate that this European bison population has been exposed to BVD virus or BVD-like viruses and BoHV-2.     

Evaluation of three commercial bovine ELISA kits for detection of antibodies against Alphaherpesviruses in reindeer (<Emphasis Type="Italic">Rangifer tarandus tarandus</Emphasis>)

Background  

The genus Varicellovirus (family Herpesviridae subfamily Alphaherpesvirinae) includes a group of viruses genetically and antigenically related to bovine herpesvirus 1 (BoHV-1) among which cervid herpesvirus 2 (CvHV-2) can be of importance in reindeer. These viruses are known to be responsible for different diseases in both wild and domestic animals. Reindeer are a keystone in the indigenous Saami culture and previous studies have reported the presence of antibodies against alphaherpesviruses in semi-domesticated reindeer in northern Norway. Mortality rates, especially in calves, can be very high in some herds and the abortion potential of alphaherpesvirus in reindeer, unlike in bovines, remains unknown.     

Multiple barriers to recombination between divergent HIV-1 variants revealed by a dual-marker recombination assay

Recombination is a major force for generating human immunodeficiency virus type 1 (HIV-1) diversity and produces numerous recombinants circulating in the human population. We previously established a cell-based system using green fluorescent protein gene (gfp) as a reporter to study the mechanisms of HIV-1 recombination. We now report an improved system capable of detecting recombination using authentic viral sequences. Frameshift mutations were introduced into the gag gene so that parental viruses do not express full-length Gag; however, recombination can generate a progeny virus that expresses a functional Gag. We demonstrate that this Gag reconstitution assay can be used to detect recombination between two group M HIV-1 variants of the same or of different subtypes. Using both gfp and gag assays, we found that, similar to group M viruses, group O viruses also recombine frequently. When recombination between a group M virus and a group O virus was examined, we found three distinct barriers for intergroup recombination. First, similar to recombination within group M viruses, intergroup recombination is affected by the identity of the dimerization initiation signal (DIS); variants with the same DIS recombined at a higher rate than those with different DIS. Second, using the gfp recombination assay, we showed that intergroup recombination occurs much less frequently than intragroup recombination, even though the gfp target sequence is identical in all viruses. Finally, Gag reconstitution between variants from different groups is further reduced compared with green fluorescent protein, indicating that sequence divergence interferes with recombination efficiency in the gag gene. Compared with identical sequences, we estimate that recombination rates are reduced by 3-fold and by 10- to 13-fold when the target regions in gag contain 91% and 72-73% sequence identities, respectively. These results show that there are at least three distinct mechanisms preventing exchange of genetic information between divergent HIV-1 variants from different groups.     

Productive Homologous and Non-homologous Recombination of Hepatitis C Virus in Cell Culture

Genetic recombination is an important mechanism for increasing diversity of RNA viruses, and constitutes a viral escape mechanism to host immune responses and to treatment with antiviral compounds. Although rare, epidemiologically important hepatitis C virus (HCV) recombinants have been reported. In addition, recombination is an important regulatory mechanism of cytopathogenicity for the related pestiviruses. Here we describe recombination of HCV RNA in cell culture leading to production of infectious virus. Initially, hepatoma cells were co-transfected with a replicating JFH1ΔE1E2 genome (genotype 2a) lacking functional envelope genes and strain J6 (2a), which has functional envelope genes but does not replicate in culture. After an initial decrease in the number of HCV positive cells, infection spread after 13–36 days. Sequencing of recovered viruses revealed non-homologous recombinants with J6 sequence from the 5′ end to the NS2–NS3 region followed by JFH1 sequence from Core to the 3′ end. These recombinants carried duplicated sequence of up to 2400 nucleotides. HCV replication was not required for recombination, as recombinants were observed in most experiments even when two replication incompetent genomes were co-transfected. Reverse genetic studies verified the viability of representative recombinants. After serial passage, subsequent recombination events reducing or eliminating the duplicated region were observed for some but not all recombinants. Furthermore, we found that inter-genotypic recombination could occur, but at a lower frequency than intra-genotypic recombination. Productive recombination of attenuated HCV genomes depended on expression of all HCV proteins and tolerated duplicated sequence. In general, no strong site specificity was observed. Non-homologous recombination was observed in most cases, while few homologous events were identified. A better understanding of HCV recombination could help identification of natural recombinants and thereby lead to improved therapy. Our findings suggest mechanisms for occurrence of recombinants observed in patients.     

Glycoprotein D of Bovine Herpesvirus 5 (BoHV-5) Confers an Extended Host Range to BoHV-1 but Does Not Contribute to Invasion of the Brain

Bovine herpesvirus 1 (BoHV-1) and BoHV-5 are closely related pathogens of cattle, but only BoHV-5 is considered a neuropathogen. We engineered intertypic gD exchange mutants with BoHV-1 and BoHV-5 backbones in order to address their in vitro and in vivo host ranges, with particular interest in invasion of the brain. The new viruses replicated in cell culture with similar dynamics and to titers comparable to those of their wild-type parents. However, gD of BoHV-5 (gD5) was able to interact with a surprisingly broad range of nectins. In vivo, gD5 provided a virulent phenotype to BoHV-1 in AR129 mice, featuring a high incidence of neurological symptoms and early onset of disease. However, only virus with the BoHV-5 backbone, independent of the gD type, was detected in the brain by immunohistology. Thus, gD of BoHV-5 confers an extended cellular host range to BoHV-1 and may be considered a virulence factor but does not contribute to the invasion of the brain.Bovine herpesvirus 1 (BoHV-1) and BoHV-5 belong to the subfamily Alphaherpesvirinae and are closely related pathogens of cattle (22). The protein repertoire of the two viruses averages 82% amino acid identity (20). Both viruses are neurotropic, but only BoHV-5 can significantly replicate in the central nervous system (CNS) to cause encephalitis of either naturally infected cattle or experimentally inoculated laboratory animals (2, 5, 6, 12, 40, 41, 44). Glycoprotein D (gD) is accepted as the critical and essential receptor-binding protein of many alphaherpesviruses (reviewed in references 8 and 48). The main gD receptors identified to date include members of the tumor necrosis factor (TNF) receptor family (HveA) and the poliovirus receptor family (HveB or nectin 2 and HveC or nectin 1) (28, 42, 51). Furthermore, a modified form of heparan sulfate, 3-O-sulfated heparan sulfate, can mediate herpesvirus entry (46). J1.1-2 cells (J cells) represent a subpopulation of thymidine kinase-negative baby hamster kidney (BHK) cells selected for their property of being resistant to infection with herpes simplex virus type 1 (HSV-1), HSV-2, and BoHV-1. The expression of nectin 1 in those cells rendered them susceptible to BoHV-1 infection and replication, which suggests that nectin 1 can serve as a receptor for BoHV-1 gD (gD1) (16, 18, 28). Interestingly, we observed that BoHV-5 was able to productively replicate in J cells without the nectin 1 receptor.According to a previously reported sequence comparison of BoHV-1 and BoHV-5 (20), the highest divergence between the two viruses mapped to the latency-related region and the immediate-early proteins (less than 75% amino acid identity) BICP0, BICP4, and BICP22. Glycoprotein E (gE) was also listed in this category, with 74% amino acid identity between gE of BoHV-1 (gE1) and gE5. This fact also gave ample reason for attempts to map the neurovirulent phenotype of BoHV-5 to the gE5 molecule (3, 4, 13). In contrast, the highest sequence similarities between the two viruses were described for proteins involved in viral DNA replication and processing as well as certain virion proteins. Among others, the predicted amino acid sequences of gD1 and gD5 were listed as being 98% identical (20). However, our own analysis using the European Molecular Biology software suite (43) revealed only 79.9% amino acid identity. Obviously, the most extensive difference between gD1 and gD5 maps to a glycine-rich stretch located in the molecule''s ectodomain, between amino acids (aa) 280 and 330 of gD5, in close vicinity to the transmembrane region.Based on these considerations, we hypothesized that BoHV-5 was able to make use of a cellular receptor that is unavailable to BoHV-1. To test this hypothesis, the gD genes were removed from bacterial artificial chromosomes (BACs) harboring the genome of either BoHV-5 or BoHV-1 (27). In a second step, gD exchange viruses were created by the cotransfection of the gD-less BACs with appropriate plasmids carrying either the gD1 or gD5 gene and appropriate flanking sequences. The newly generated viruses included an intertypic BoHV-5 mutant carrying gD1 in the place of gD5 and a corresponding BoHV-1 carrying gD5. These mutants, together with appropriate revertant mutants, were then used to explore their ability to infect J cells in vitro and their ability to cause neurological disease and invade the brain in vivo, in a previously established mouse model (2). Our results indicate that gD5 confers an extended host range to BoHV-1 but is nonessential for the invasion of the brain.     

Mechanism of RNA recombination in carmo- and tombusviruses: evidence for template switching by the RNA-dependent RNA polymerase in vitro

RNA recombination occurs frequently during replication of tombusviruses and carmoviruses, which are related small plus-sense RNA viruses of plants. The most common recombinants generated by these viruses are either defective interfering (DI) RNAs or chimeric satellite RNAs, which are thought to be generated by template switching of the viral RNA-dependent RNA polymerase (RdRp) during the viral replication process. To test if RNA recombination is mediated by the viral RdRp, we used either a purified recombinant RdRp of Turnip crinkle carmovirus or a partially purified RdRp preparation of Cucumber necrosis tombusvirus. We demonstrated that these RdRp preparations generated RNA recombinants in vitro. The RdRp-driven template switching events occurred between either identical templates or two different RNA templates. The template containing a replication enhancer recombined more efficiently than templates containing artificial sequences. We also observed that AU-rich sequences promote recombination more efficiently than GC-rich sequences. Cloning and sequencing of the generated recombinants revealed that the junction sites were located frequently at the ends of the templates (end-to-end template switching). We also found several recombinants that were generated by template switching involving internal positions in the RNA templates. In contrast, RNA ligation-based RNA recombination was not detected in vitro. Demonstration of the ability of carmo- and tombusvirus RdRps to switch RNA templates in vitro supports the copy-choice models of RNA recombination and DI RNA formation for these viruses.     

Proteomic Characterization of Bovine Herpesvirus 4 Extracellular Virions

Gammaherpesviruses are important pathogens in human and animal populations. During early events of infection, these viruses manipulate preexisting host cell signaling pathways to allow successful infection. The different proteins that compose viral particles are therefore likely to have critical functions not only in viral structures and in entry into target cell but also in evasion of the host''s antiviral response. In this study, we analyzed the protein composition of bovine herpesvirus 4 (BoHV-4), a close relative of the human Kaposi''s sarcoma-associated herpesvirus. Using mass spectrometry-based approaches, we identified 37 viral proteins associated with extracellular virions, among which 24 were resistant to proteinase K treatment of intact virions. Analysis of proteins associated with purified capsid-tegument preparations allowed us to define protein localization. In parallel, in order to identify some previously undefined open reading frames, we mapped peptides detected in whole virion lysates onto the six frames of the BoHV-4 genome to generate a proteogenomic map of BoHV-4 virions. Furthermore, we detected important glycosylation of three envelope proteins: gB, gH, and gp180. Finally, we identified 38 host proteins associated with BoHV-4 virions; 15 of these proteins were resistant to proteinase K treatment of intact virions. Many of these have important functions in different cellular pathways involved in virus infection. This study extends our knowledge of gammaherpesvirus virions composition and provides new insights for understanding the life cycle of these viruses.     

Genetic Recombination in Mycobacteria

Evidence for genetic recombination between Mycobacterium smegmatis strain Rabinowitchi (Rab) and strain Jucho or PM5 is presented. Backcrosses of recombinants by either parental strain indicated four different types of mating behavior, suggesting that the mycobacterial compatibilities are controlled by at least two different factors. No sex factor that transfers at a high frequency or that is sensitive to acridine dyes was detected. Analysis of segregation of unselected markers revealed that strain Jucho, or PM5, contributes the majority of alleles in almost all recombinants obtained from different selective media. Efforts to construct linkage maps for the markers employed failed because of ordering ambiguities. Mating medium containing streptomycin prevented genetic recombination when strain Rab was resistant to the antibiotic and Jucho, or PM5, was sensitive, but it did not prevent recombination when Rab was sensitive to streptomycin and Jucho, or PM5, was resistant. Very low frequency of recombinant formation was observed when Jucho, or PM5, had been treated with streptomycin, whereas recombinants were formed at fairly high frequencies when Rab had been treated with the antibiotic, suggesting that the roles of parental strains in zygote formation were not identical. The results suggest a polar transfer of genetic material from Rab to Jucho, or PM5, although an alternative possibility of cell fusion followed by exclusion could not be excluded.     

RNA recombination of murine coronaviruses: recombination between fusion-positive mouse hepatitis virus A59 and fusion-negative mouse hepatitis virus 2.

It has previously been shown that the murine coronavirus mouse hepatitis virus (MHV) undergoes RNA recombination at a relatively high frequency in both tissue culture and infected animals. Thus far, all of the recombination sites had been localized at the 5' half of the RNA genome. We have now performed a cross between MHV-2, a fusion-negative murine coronavirus, and a temperature-sensitive mutant of the A59 strain of MHV, which is fusion positive at the permissive temperature. By selecting fusion-positive viruses at the nonpermissive temperature, we isolated several recombinants containing multiple crossovers in a single genome. Some of the recombinants became fusion negative during the plaque purification. The fusion ability of the recombinants parallels the presence or absence of the A59 genomic sequences encoding peplomers. Several of the recombinants have crossovers within 3' end genes which encode viral structural proteins, N and E1. These recombination sites were not specifically selected with the selection markers used. This finding, together with results of previous recombination studies, indicates that RNA recombination can occur almost anywhere from the 5' end to the 3' end along the entire genome. The data also show that the replacement of A59 genetic sequences at the 5' end of gene C, which encodes the peplomer protein, with the fusion-negative MHV-2 sequences do not affect the fusion ability of the recombinant viruses. Thus, the crucial determinant for the fusion-inducing capability appears to reside in the more carboxyl portion of the peplomer protein.     

Recombination between sequences of hepatitis B virus from different genotypes

A comparison of 25 hepatitis B virus (HBV) isolates for which complete genome sequences are available revealed two that occupied different positions in phylogenetic trees reconstructed from different open reading frames. Further analysis indicated that this incongruence was the result of recombination between viruses of different genomic and antigenic types. Both putative recombinants originated from geographic regions where multiple genotypes are known to cocirculate. A search of the sequence databases showed evidence of similar intergenotypic recombinants. These observations indicate that recombination between divergent strains may represent an important source of genetic variation in HBV.     

RNA recombination in a coronavirus: recombination between viral genomic RNA and transfected RNA fragments.

Mouse hepatitis virus (MHV), a coronavirus, has been shown to undergo a high frequency of RNA recombination both in tissue culture and in animal infection. So far, RNA recombination has been demonstrated only between genomic RNAs of two coinfecting viruses. To understand the mechanism of RNA recombination and to further explore the potential of RNA recombination, we studied whether recombination could occur between a replicating MHV RNA and transfected RNA fragments. We first used RNA fragments which represented the 5' end of genomic-sense sequences of MHV RNA for transfection. By using polymerase chain reaction amplification with two specific primers, we were able to detect recombinant RNAs which incorporated the transfected fragment into the 5' end of the viral RNA in the infected cells. Surprisingly, even the anti-genomic-sense RNA fragments complementary to the 5' end of MHV genomic RNA could also recombine with the MHV genomic RNAs. This observation suggests that RNA recombination can occur during both positive- and negative-strand RNA synthesis. Furthermore, the recombinant RNAs could be detected in the virion released from the infected cells even after several passages of virus in tissue culture cells, indicating that these recombinant RNAs represented functional virion RNAs. The crossover sites of these recombinants were detected throughout the transfected RNA fragments. However, when an RNA fragment with a nine-nucleotide (CUUUAUAAA) deletion immediately downstream of a pentanucleotide (UCUAA) repeat sequence in the leader RNA was transfected into MHV-infected cells, most of the recombinants between this RNA and the MHV genome contained crossover sites near this pentanucleotide repeat sequence. In contrast, when exogenous RNAs with the intact nine-nucleotide sequence were used in similar experiments, the crossover sites of recombinants in viral genomic RNA could be detected at more-downstream sites. This study demonstrated that recombination can occur between replicating MHV RNAs and RNA fragments which do not replicate, suggesting the potential of RNA recombination for genetic engineering.     

Natural Recombination among Plant Virus Genomes: Evidence from Tobraviruses

RNA recombination in plants was first identified by the repairin vivoof a deleted genomic RNA of brome mosaic virus. Subsequently, evidence of recombination has been detected not only in experimental systems but also among an increasing number of naturally occurring isolates of plant viruses. This article discusses the different recombinants that have been found among viruses in the genusTobravirusand describes other examples of recombination among plant viruses and between the genomes of viruses and their hosts.     

Apoptotic and developmental effects of bovine Herpesvirus type-5 infection on in vitro-produced bovine embryos

Bovine Herpesvirus type-5 (BoHV-5), which is potentially neuropathogenic, was recently described to be related with reproductive disorders in cows. The objective was to elucidate mechanisms involved in propagation of BoHV-5 in embryonic cells. For this purpose, bovine embryos produced in vitro were assayed for apoptotic markers after experimental infection of oocytes, in vitro fertilization, and development. Host DNA fragmentation was detected with a TUNEL assay, expression of annexin-V was measured with indirect immunofluorescence, and viral DNA was detected with in situ hybridization. Infective BoHV-5 virus was recovered from embryos derived from exposed oocytes after two consecutive passages on Madin-Darby bovine kidney (MDBK) cells. The viral DNA corresponding to US9 gene, localized between nucleotides 126243 to 126493, was detected in situ and amplified. There was no significant difference between the ratio of TUNEL stained nuclei and total cells in good quality blastocysts (0.87 ± 0.05, mean ± SD), but there were differences (P < 0.05) between infected (0.18 ± 0.05) and uninfected blastocysts (0.73 ± 0.07). The Annexin-V label was more intense in uninfected embryos (0.79 ± 0.04; P < 0.05). The quality of infected and uninfected embryos was considered equal, with no significant effect on embryonic development. In conclusion, we inferred that BoHV-5 infected bovine oocytes, replicated, and suppressed some apoptotic pathways, without significantly affecting embryonic development.