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The objective of this study was to compare reproductive protection in cattle against bovine viral diarrhea virus (BVDV) and bovine herpesvirus 1 (BoHV-1) provided by annual revaccination with multivalent modified-live viral (MLV) vaccine or multivalent combination viral (CV) vaccine containing temperature-sensitive modified-live BoHV-1 and killed BVDV when MLV vaccines were given pre-breeding to nulliparous heifers. Seventy-five beef heifers were allocated into treatment groups A (n=30; two MLV doses pre-breeding, annual revaccination with MLV vaccine), B (n=30; two MLV doses pre-breeding, annual revaccination with CV vaccine) and C (n=15; saline in lieu of vaccine). Heifers were administered treatments on days 0 (weaning), 183 (pre-breeding), 366 (first gestation), and 738 (second gestation). After first calving, primiparous cows were bred, with pregnancy assessment on day 715. At that time, 24 group A heifers (23 pregnancies), 23 group B heifers (22 pregnancies), and 15 group C heifers (15 pregnancies) were commingled with six persistently infected (PI) cattle for 16days. Ninety-nine days after PI removal, cows were intravenously inoculated with BoHV-1. All fetuses and live offspring were assessed for BVDV and BoHV-1. Abortions occurred in 3/23 group A cows, 1/22 group B cows, and 11/15 group C cows. Fetal infection with BVDV or BoHV-1 occurred in 4/23 group A offspring, 0/22 group B offspring, and 15/15 group C offspring. This research demonstrates efficacy of administering two pre-breeding doses of MLV vaccine with annual revaccination using CV vaccine to prevent fetal loss due to exposure to BVDV and BoHV-1.

Bovine herpesvirus type 1 (BoHV-1) is a widespread respiratory infection that significantly impacts cattle health worldwide. To address this issue in China, we previously developed a novel double gene-deleted vaccine targeting gG and tk. In this study, we further evaluated the efficacy of this vaccine by challenging vaccinated cattle with a prevalent wild-type BoHV-1 strain and comparing its effectiveness against a commercially available inactivated BoHV-1 vaccine. Post-immunization, all cattle maintained normal rectal temperatures and exhibited no respiratory symptoms. Cattle receiving the gene-deleted vaccine showed a significant increase in the expression of immune markers IFN-γ and TNF-α. Following exposure to wild-type BoHV-1, all immunized groups produced high levels of neutralizing antibodies and specific gB antibodies. Notably, virus shedding was significantly lower in the vaccinated groups compared to the non-immune challenge group. Histological analysis of lung tissues revealed that vaccinated calves had more intact lung structure than their unimmunized counterparts after the challenge. Additionally, the gG−/tk- gene-deleted vaccine demonstrated a higher protective rate based on the average scores of clinical symptoms and lung lesions. Overall, the BoHV-1 gG−/tk- gene-deleted vaccine outperformed the other vaccines tested. This study confirms that the gene-deleted vaccine provides robust protection and superior immunogenicity compared to existing inactivated vaccines, underscoring its potential for future market application. •The immunogenicity of a BoHV-1 double-gene deletion vaccine was compared with that of a commercial inactivated vaccine.•Cattle in the double-gene deletion vaccine group effectively produced cellular and humoral immune responses comparable to that of the inactivated vaccine group.•Virus shedding was significantly lower in the vaccinated groups compared to the non-immune challenge group, especially in double-gene deletion group.•After challenge, calves vaccinated with the double-gene deletion vaccine had more intact lung structures and showed a higher rate of protention.

Bovine herpesvirus type 1 (BoHV-1) establishes lifelong latency in trigeminal ganglionic (TG) neurons following intranasal and ocular infection in cattle. Periodically, the latent virus reactivates in the TG due to stress and is transported anterogradely to nerve endings in the nasal epithelium, where the virus replicates and sheds. Consequently, BoHV-1 is transmitted to susceptible animals and maintained in the cattle population. Modified live BoHV-1 vaccine strains (BoHV-1 MLV) also have a similar latency reactivation. Therefore, they circulate and are maintained in cattle herds. Additionally, they can regain virulence and cause vaccine outbreaks because they mutate and recombine with other circulating field wild-type (wt) strains. Recently, we constructed a BoHV-1 quadruple mutant virus (BoHV-1qmv) that lacks immune evasive properties due to UL49.5 and glycoprotein G (gG) deletions. In addition, it also lacks the gE cytoplasmic tail (gE CT) and Us9 gene sequences designed to make it safe, increase its vaccine efficacy against BoHV-1, and restrict its anterograde neuronal transport noted above. Further, we engineered the BoHV-1qmv-vector to serve as a subunit vaccine against the Rift Valley fever virus (BoHV-1qmv Sub-RVFV) (doi: 10.3390/v15112183). In this study, we determined the latency reactivation and nasal virus shedding properties of BoHV-1qmv (vector) and BoHV-1qmv-vectored subunit RVFV (BoHV-1qmv sub-RVFV) vaccine virus in calves in comparison to the BoHV-1 wild-type (wt) following intranasal inoculation. The real-time PCR results showed that BoHV-1 wt- but not the BoHV-1qmv vector- and BoHV-1qmv Sub-RVFV-inoculated calves shed virus in the nose following dexamethasone-induced latency reactivation; however, like the BoHV-1 wt, both the BoHV-1qmv vector and BoHV-1qmv Sub-RVFV viruses established latency, were reactivated, and replicated in the TG neurons. These results are consistent with the anterograde neurotransport function of the gE CT and Us9 sequences, which are deleted in the BoHV-1qmv and BoHV-1qmv Sub-RVFV.

Recombinant herpesvirus vaccine vectors offer distinct advantages in next-generation vaccine development, primarily due to the ability to establish persistent infections to provide sustainable antigen responses in the host. Recombinant bovine herpesvirus-4 (BoHV-4) has been previously shown to elicit protective immunity in model laboratory animal species against a variety of pathogens. For the first time, we describe the induction of antigen-specific immune responses to two delivered antigens in the host species after intranasal nebulization of recombinant BoHV-4 expressing the chimeric peptide containing the bovine viral diarrhea virus (BVDV) glycoprotein E2 and the bovine herpesvirus 1 (BoHV-1) glycoprotein D (BoHV-4-A-CMV-IgK-gE2gD-TM). In this study, four cattle were immunized via intranasal nebulization with the recombinant BoHV-4 construct. Two of the cattle were previously infected with wild-type BoHV-4, and both developed detectable serologic responses to BVDV and BoHV-1. All four immunized cattle developed detectable viral neutralizing antibody responses to BVDV, and one steer developed a transient viral neutralizing response to BoHV-1. Approximately one year after immunization, immunosuppressive doses of the glucocorticoid dexamethasone were administered intravenously to all four cattle. Within two weeks of immunosuppression, all animals developed viral neutralizing antibody responses to BoHV-1, and all animals maintained BVDV viral neutralizing capacity. Overall, nebulization of BoHV-4-A-CMV-IgK-gE2gD-TM persistently infects cattle, is capable of eliciting antigen-specific immunity following immunization, including in the presence of pre-existing BoHV-4 immunity, and recrudescence of the virus boosts the immune response to BoHV-4-vectored antigens. These results indicate that BoHV-4 is a viable and attractive vaccine delivery platform for use in cattle.

BoAHV1 is a significant cattle pathogen, resulting in substantial economic losses worldwide. B-cell epitopes within the viral proteins are not well understood. In this study, we screened a 12-mer phage display peptide library using a commercial BoAHV1 antibody. A total of 16 phage clones displaying individual 12-mer peptides were identified as reactive with this antibody using dot bot analysis. Through sequence alignment, 10 putative BoAHV1 B-cell epitopes, designated as PV109, PV108, PV116, PV59, PV50, PV130, PV113, PV49, PV62, and PV133, located within viral proteins gB, gC, gG, gM, UL36, UL37, and UL49, respectively, were recognized by both commercial BoAHV1 antibody andBoAHV1 IgG positive cattle serum through dot blot assay. Interestingly, immunization of mice with the synthesized peptide PV116 led to the production of antibodies suitable for Western blot analysis. Furthermore, an ELISA kit for the detection of BoAHV1 IgG antibodies in serum was developed, utilizing the identified epitope PV49, PV108, PV109, and PV116 as coating antigen. Collectively, we have identified 10 novel B-cell epitopes ofBoAHV1. Among them, PV116 is capable of inducing the production of antibodies suitable for Western blotting assay, which also shows potential for the development diagnostic tools.

Bovine herpesvirus type 1 (BHV-1) is a highly contagious DNA virus that causes a variety of diseases affecting the reproductive and respiratory tracts. These diseases can reduce the health and production performance of cattle, causing significant economic losses in the cattle industry. The current ELISA kits used to detect BHV-1 have long lead times and are expensive, and are not suitable for bulk testing on large farms. therefore, there is an urgent need to develop a rapid and cost-effective alternative to the BHV-1 test. In this study, recombinant gD protein was expressed by prokaryotic system, and then used as antigen to immunize New Zealand white rabbits to obtain polyclonal antibodies (pAb). An indirect enzyme-linked immunosorbent assay (iELISA) based on gD protein was established for the detection of BHV-1 antibodies in clinical samples. The optimal cutoff value was determined to be 0.6185 using 60 clinical serum samples. This method had no cross-reaction with other common bovine viruses. The developed iELISA method and commercially available kits were used to detect 60 bovine serum samples, with a concordance rate of 93.3%. In summary, we established a rapid and reliable iELISA method based on gD protein, which is suitable for epidemio-logical monitoring of BHV-1.

Background The reproductive problem is an animal health-related bottleneck that constrains livestock genetic improvement efforts in tropical countries such as Ethiopia. The infectious causes of reproductive disorders are one cause of decreased reproductive efficiency. This study aimed to determine the seroprevalence to Bovine Herpesvirus-1 (BHV1), Bovine Viral Diarrhea Virus ( BVDV), Neospora caninum ( N . caninum) and C. burnetii (C. burnetii) exposures in dairy cows with reproductive disorders in selected areas of Ethiopia. Overall, 164 serum samples were collected from October 2018 to May 2019 from animals with a history of reproductive disorders. The collected sera were tested for antibody titers to Brucella species, N. caninum , BVDV, BHV1, C. burnetii and Chlamydophila abortus (C. abortus ) using Rose Bengal and ELISA. Results The apparent seroprevalence of BHV1, BVDV, N. caninum and C. burnetii were 61%, 33.5%, 4.9% and 0.6%, respectively. Among the selected study areas, the mean apparent seroprevalence was significantly greater in Bishoftu (35.9%), Holeta (34.2%) and Adaberga (28.6%) than in Mekelle (9.9%) and Ambo (16.2%). Among the specific seroprevalence in specific areas, BHV1 was the most common in Adaberga, with an apparent seroprevalence of 92.9%. Similarly, the seroprevalence of BVDV was the highest in Holeta, with an apparent seroprevalence of 73.3%. On the other hand, no seropositive animal to Brucella spp. or C. abortus was found in these study areas. Conclusion BVDV and BHV1 seroprevalence was higher in dairy cattle with a history of reproductive disorder in Ethiopia as compared to the seroprevalence of N. caninum and C. burnetii .

Bovine herpes virus 1 (BHV-1) is primarily associated with clinical syndromes such as rhinotracheitis, pustular vulvovaginitis and balanoposthitis, abortion, infertility, conjunctivitis and encephalitis in bovine species. The main sources of infection are the nasal exudates and the respiratory droplets, genital secretions, semen, fetal fluids and tissues. The BHV-1 virus can become latent following a primary infection with a field isolate or vaccination with an attenuated strain. The viral genomic DNA has been demonstrated in the sensory ganglia of the trigeminal nerve in infectious bovine rhinotracheitis (IBR) and in sacral spinal ganglia in pustular vulvovaginitis and balanoposthitis cases. BHV-1 infections can be diagnosed by detection of virus or virus components and antibody by serological tests or by detection of genomic DNA by polymerase chain reaction (PCR), nucleic acid hybridization and sequencing. Inactivated vaccines and modified live virus vaccines are used for prevention of BHV-1 infections in cattle; subunit vaccines and marker vaccines are under investigation.

Rift Valley fever (RVF) is a vector-borne zoonotic viral disease that causes abortion storms, fetal malformations, and neonatal mortality in livestock ruminants. In humans, RVF can lead to hemorrhagic fever, encephalitis, retinitis, or blindness, and about 1% of patients die. Since there are no registered vaccines for human use, developing RVF vaccines for use in animals is crucial to protect animals and prevent the spread of the virus from infecting humans. We recently developed a live bovine herpesvirus type 1 quadruple gene-mutant vector (BoHV-1qmv) that lacks virulence and immunosuppressive properties. Further, we engineered a BoHV-1qmv-vectored subunit Rift Valley fever virus (RVFV) vaccine (BoHV-1qmv Sub-RVFV) for cattle, in which a chimeric polyprotein coding for the RVFV Gc, Gn, and bovine granulocyte–macrophage colony-stimulating factor (GMCSF) proteins is fused but cleaved proteolytically in infected cells into individual membrane-anchored Gc and secreted Gn-GMCSF proteins. Calves vaccinated with the BoHV-1qmv Sub-RVFV vaccine generated moderate levels of RVFV-specific serum-neutralizing (SN) antibodies and cellular immune responses. In the current study, we repurposed the BoHV-1qmv Sub-RVFV for sheep by replacing the RVFV Gc and Gn ORF sequences codon-optimized for bovines with the corresponding ovine-codon-optimized sequences and by fusing the sheep GM-CSF ORF sequences with the Gn ORF sequence. A combined primary intranasal-plus-subcutaneous primary immunization induced a moderate level of BoHV-1 (vector)- and vaccine strain MP12-specific SN antibodies and MP-12-specific cellular immune responses. Notably, an intranasal booster vaccination after 29 days triggered a rapid (within 7 days) rise in MP-12-specific SN antibody titers. Therefore, the BoHV-1qmv-vectored subunit RVFV vaccine is safe and highly immunogenic in sheep and can potentially be an efficient subunit vaccine for sheep against RVFV.

Infection of cattle by bovine herpesvirus type 1 (BHV-1) can lead to upper respiratory tract disorders, conjunctivitis, genital disorders and immune suppression. BHV-1-induced immune suppression initiates bovine respiratory disease complex (BRDC), which costs the US cattle industry approximately 3 billion dollars annually. BHV-1 encodes at least three proteins that can inhibit specific arms of the immune system: (i) bICP0 inhibits interferon-dependent transcription, (ii) the UL41.5 protein inhibits CD8+ T-cell recognition of infected cells by preventing trafficking of viral peptides to the surface of the cells and (iii) glycoprotein G is a chemokine-binding protein that prevents homing of lymphocytes to sights of infection. Following acute infection of calves, BHV-1 can also infect and induce high levels of apoptosis of CD4+ T-cells. Consequently, the ability of BHV-1 to impair the immune response can lead to BRDC. Following acute infection, BHV-1 establishes latency in sensory neurons of trigeminal ganglia (TG) and germinal centers of pharyngeal tonsil. Periodically BHV-1 reactivates from latency, virus is shed, and consequently virus transmission occurs. Two viral genes, the latency related gene and ORF-E are abundantly expressed during latency, suggesting that they regulate the latency-reactivation cycle. The ability of BHV-1 to enter permissive cells, infect sensory neurons and promote virus spread from sensory neurons to mucosal surfaces following reactivation from latency is also regulated by several viral glycoproteins. The focus of this review is to summarize the biology of BHV-1 and how this relates to BRDC.