Nucleocytoplasmic shuttling of BEFV M protein-modulated by lamin A/C and chromosome maintenance region 1 through a transcription-, carrier- and energy-dependent pathway.

This study demonstrates for the first time that the matrix (M) protein of BEFV is a nuclear targeting protein that shuttles between the nucleus and the cytoplasm in a transcription-, carrier-, and energy-dependent manner. Experiments performed in both intact cells and digitonin-permeabilized cells revealed that M protein targets the nucleolus and requires carrier, cytosolic factors or energy input. By employing sequence and mutagenesis analyses, we have determined both nuclear localization signal (NLS) 6KKGKSK11 and nuclear export signal (NES) 98LIITSYL TI106 of M protein that are important for the nucleocytoplasmic shuttling of M protein. Furthermore, we found that both lamin A/C and chromosome maintenance region 1 (CRM-1) proteins could be coimmunoprecipitated and colocalized with the BEFV M protein. Knockdown of lamin A/C by shRNA and inhibition of CRM-1 by leptomycin B significantly reduced virus yield. Collectively, this study provides novel insights into nucleocytoplasmic shuttling of the BEFV M protein modulated by lamin A/C and CRM-1 and by a transcription- and carrier- and energy-dependent pathway.

Evaluation of antibody response to inactivated bovine ephemeral fever virus vaccine for Japanese Black calves in the field.

The purpose of this study was to investigate the antibody response to the bovine ephemeral fever virus (BEFV) vaccine in Japanese Black calves. Twenty-eight Japanese Black calves, which were raised on an ordinal farm, were divided into two groups. Fifteen calves received the inactivated BEFV vaccine at 12 and 16 weeks of age (vaccination group), and 13 calves did not receive the vaccine (non-vaccination group). Blood samples were obtained at 0, 4, 8, 12, 16, 20, 24, 28, and 32 weeks of age. As the results, in the vaccination group, the antibody titers at 16, 20, 24, 28, and 32 weeks of age were significantly higher than those at 0, 4, 8, and 12 weeks of age (p < 0.01). Additionally, antibody titer in the vaccination group increased after 16 weeks of age and showed a significantly higher level than that in the non-vaccination group throughout the remaining experimental period (p < 0.01). These results might be helpful in establishing a vaccination program against BEFV in calves.

Stretching the wings further- susceptibility of Culex pipiens Linnaeus to bovine ephemeral fever virus infection under experimental conditions.

Bovine ephemeral fever (BEF) is a significant viral disease of cattle in the tropical, subtropical, and temperate climatic zones. This disease is also known as three-day sickness due to the spontaneous recovery of the cattle within a short period (usually 3-5 days). Despite its short duration, the disease may have a considerable impact. It can cause heavy economic losses, primarily due to decreased milk production, lowered fertility in bulls, and even fatality in severe cases. The virus is suspected to be transmitted by haematophagous insects (mainly mosquitoes and Culicoides biting midges); however, the identity of a competent vector for BEFV remains a mystery. Here, we investigated whether BEFV may replicate for a short duration in Culex pipiens Linnaeus, 1758, the most prevalent mosquito species in Israel and a potential vector of this virus to Israeli cattle. We applied nested- qPCR to test BEFV abundance in Cx. pipiens every 24 h for 14 consecutive days post-infection. Additionally, we collected eggs laid by BEFV-infected females and investigated BEFV abundance in the different developmental stages of F1 mosquitos. Our results suggest that Cx. pipiens mosquitoes have the potential to act as a vector of BEFV and also indicate that BEFV may be vertically transmitted from Cx. pipiens female parent to her female offspring.

Effectiveness of the strain 919 bovine ephemeral fever virus vaccine in the face of a real-world outbreak: A field study in Israeli dairy herds.

Bovine ephemeral fever virus (BEFV) is a globally spread arthropod-borne RNA virus that has significant economic impacts on the cattle industry. A live attenuated commercial BEF vaccine, based on the Australian BEFV strain 919, is widely used in Israel and other countries. A previous study has suggested the high effectiveness of this vaccine (ULTRAVAC BEF VACCINE™ from Zoetis®), but anecdotal reports of high BEF morbidity among vaccinated dairy herds in Israel casted doubt on these findings. To resolve this uncertainty, a randomized controlled field vaccine effectiveness study was conducted in Israel during a BEF outbreak which occurred in 2021. Eleven dairy herds were enrolled and monitored for BEF-associated morbidity and rumination alteration patterns using electronic monitoring tags (HR Tags, SCR® Dairy, Netanya, Israel). Four of the herds were naturally infected with BEFV during the outbreak, resulting in a total of 120 vaccinated and 311 unvaccinated subjects that were included in the effectiveness study. A mixed-effect Cox proportional hazard regression model was used to calculate the overall hazard ratio between vaccinated and unvaccinated cattle. This analysis demonstrated an average vaccine effectiveness of 60 % (95 % CI = 38 %-77 %) for preventing clinical disease. In addition, a non-statistically significant trend (p = 0.1) towards protection from mortality was observed, with no observation of mortality among the vaccinated groups compared to 2.61 % mortality (7/311) among the unvaccinated subjects. One hundred and thirty vaccinated and unvaccinated calves from affected and non-affected herds and with different status of morbidity were sampled and analysed by serum-neutralization test. The highest titers of BEFV-neutralizing antibodies were found in subjects that were both vaccinated and clinically affected, indicating a booster effect after vaccination. The results of the study provide evidence for the moderate effectiveness of the ULTRAVAC BEF VACCINE™ for the prevention of BEF.

Whole genome characterization and evolutionary analysis of bovine ephemeral fever virus isolated in Iran.

Bovine ephemeral fever virus (BEFV) is an economically important arthropod-borne virus of cattle and water buffaloes which is enzootic in Africa, Australia, and Asia. We characterized the entire length of BEFV BA/RZ/IR strain genome isolated in Iran and compared to the all BEFV full genomes available in the GenBank. The BEFV genomes were phylogenetically classified as 4 lineages including the Middle Eastern, East Asian, Australian, and South African lineages. The Iranian BA/RZ/IR strain, which displayed maximum sequence identity (96.72%) to the Chinese JT02L strain was clustered as a separate branch in the East Asian lineage of the virus. Using Shannon entropy analysis, amino acid variations were detected in the all proteins encoded by BEFV genomes. Particularly, the polymerase L and the accessory proteins Gns, α2 and ß exhibited the highest amino acid variations suggesting their significance in the viral replication efficiency. Our bioinformatics analyses also predict the occurrence of recombination event within the East Asian lineage of BEFV genomes. Our data show that the Chinese Henan 1 may be a hybrid strain constructed of the Chinese JT02L and Iranian BA/RZ/IR BEFV strains as the major and minor parents, respectively. These computational analyses suggest that the homologous recombination may be an evolutionary mechanism for BEFV as a member of the Rhabdoviridae family.

Role of cervids in the epidemiology of bovine ephemeral fever virus infection in the Republic of Korea: A cross-sectional retrospective study.

BACKGROUND: Bovine ephemeral fever (BEF) is a viral disease in cattle and buffaloes, with subclinical involvement in various ruminant species. OBJECTIVES: This study aimed to investigate bovine ephemeral fever virus (BEFV) transmission in deer in the Republic of Korea (ROK) and the potential risk factors associated with seropositivity. METHODS: We conducted a retrospective cross-sectional serological survey of neutralising antibodies against BEFV in cervid sera collected from the ROK. RESULTS: The seroprevalence of BEFV was estimated to be 10.8% (95% confidence interval [CI] = 8.5-14.1), demonstrating that exposure to this virus is prevalent among farmed and free-ranging cervids in the ROK. The results revealed that age class and geographic location affected seroprevalence. Older age and the presence of neighbouring ruminant farms were significant risk factors (odds ratio [OR] = 2.394, 95% CI = 1.195-4.796) and (OR = 1.533, 95% CI = 1.026-2.288), respectively. We also observed that the individual likelihood of positivity in the southern provinces was significantly higher than that in the northern provinces (OR = 1.744, 95% CI = 1.001-3.037). There were also significant differences in the seroprevalence of cervids between the western and eastern provinces (OR = 2.021, 95% CI = 1.047-3.900). Factors that were not significantly associated with BEFV antibody prevalence included herd size and species (p > 0.05). CONCLUSIONS: These results suggest that cervid species may serve as important reservoirs for the transmission of BEFV, highlighting the need for closer monitoring of BEFV infections in cervids in the ROK.

Molecular characterization and phylogenetic analysis of bovine ephemeral fever viruses in Khuzestan province of Iran in 2018 and 2020.

BACKGROUND: Bovine ephemeral fever (BEF) is an arthropod-borne viral disease caused by the BEF virus (BEFV). This single-stranded RNA virus that affects cattle and water buffalo is endemic in tropical and subtropical regions including Iran. While BEF is a major disease of cattle in Iran, information regarding its agent, molecular characterization, and circulating viruses are highly limited. The current study aimed to, firstly, determine the genetic and antigenic characteristics of BEFV strains in Khuzestan province in Southwest of Iran in 2018 and 2020 and, secondly, to compare them with strains obtained from other areas. RESULTS: By phylogenetic analysis based on the Glycoprotein gene, BEFV strains were divided into four clusters of Middle East, East Asia, South Africa, and Australia; in which the 2018 and 2020 Iranian BEFV strains were grouped in the Middle East cluster with the Turkish, Indian, and Israeli strains. Depending on the chronology and geographical area, the outbreaks of Turkey (2020), Iran (2018 and 2020), and India (2018 and 2019) are proposed to be related. These BEFVs had the highest identity matrix and the lowest evolutionary distance among the studied strains. Multiple sequence alignment of G1, G2, and G3 antigenic sites showed that these neutralizing epitopes are highly conserved among the strains of the Middle East cluster; however, the strains previously identified in Iran differed in three amino acids placed in G1 and G2 epitopes. CONCLUSION: The findings revealed that BEFVs circulating in the Middle East are closely related phylogenetically and geographically. They also have similar antigenic structures; therefore, developing a vaccine based on these strains can be effective for controlling BEF in the Middle East.

Identification of Cytotoxic T lymphocyte (CTL) Epitope and design of an immunogenic multi-epitope of Bovine Ephemeral Fever Virus (BEFV) Glycoprotein G for Vaccine Development.

Bovine ephemeral fever (BEF), a vector-borne disease of cattle and water buffalo, is enzootic in tropical and subtropical zones of Asia, Australia, and Africa. Since cytotoxic T lymphocytes (CTL) responses may play a key role in the control of bovine ephemeral fever virus (BEFV) infection, it is important to identify and characterize the CTL target epitopes of BEFV antigens. The current study has been designed to identify and characterize the potential CTL epitopes using the Immuno-informatics tools, and it helped find the potent vaccine candidates against BEF. Antigenicity, toxicity, allergenicity, and immunogenicity testing of predicted CTL epitopes was done. Total four CTL epitopes for BEFV G protein, have been identified as potential epitopes. Prediction of the 3D structure of multi-epitope (final structure) was performed using I-TASSER server. Model 1 was selected as the best model with C-Score: -3.71. The modeled G protein structure and multi-epitope structure were validated by the Ramachandran plots Prosa and Verify 3D server. Epitopic regions of 3D protein structure were identified by Chimera UCSF software. Physicochemical properties of the Multi epitope were evaluated using ProtParam server. This is the first report of CTL epitope in the G protein of BEFV. In this manner, they would play an important role in evoking the immune response as well as vaccine development. However, in vitro and in vivo experimental studies are required for suggested epitopes verification. The multi-epitope was designed from regions of the G protein sequence that lacked mutation and genomic diversity. Therefore, it can be introduced as a protein vaccine from all strains of BEFV as a vaccine candidate for design.

Seroprevalence of Bovine ephemeral fever virus in Gujarat State of India.

Bovine ephemeral fever (BEF) virus (BEFV) is an arthropod borne virus that causes bovine ephemeral fever or three­day sickness in cattle and buffaloes. This is the first report on seroprevalence of BEF in cattle and buffaloes in Gujarat, India. Total of 92 animals, 78 cattle and 14 buffaloes from three regions (districts) of Gujarat state of India, were screened for the presence of anti­BEF antibodies. A total of 27 out of 92 animals were found positive and overall seroprevalence detected was 29.34% (95% CI 20.0­38.6%). A total of 19 out of 78 cattle and 8 out of 14 buffalo's samples were found positive BEFV antibodies. Species­wise seroprevalence in cattle and buffaloes was 24.35% (95% CI 14.8­33.8%) and 57.1% (95% CI 31.2­83.0%), respectively. There was a statistically significant (p < 0.05) species effect based on the seroprevalence. In cattle, location­wise seroprevalence was observed to be 26.82% (95% CI 13.2­40.3%) and 21.62% (95% CI 8.3­34.8%) in Navsari and Banaskantha districts, respectively. The effect of location is not statistically significant (p < 0.05). Cytopathic effect of Vero cells was characterized by rounding, granulation of the cytoplasm within 48­72 hrs of post infection. This was the first report demonstrating the presence of BEFV in Gujarat state.

Finding potential inhibitors against RNA-dependent RNA polymerase (RdRp) of bovine ephemeral fever virus (BEFV): an in-silico study.

The bovine ephemeral fever virus (BEFV) is an enzootic agent that affects millions of bovines and causes major economic losses. Though the virus is seasonally reported with a very high morbidity rate (80-100%) from African, Australian, and Asiatic continents, it remains a neglected pathogen in many of its endemic areas, with no proper therapeutic drugs or vaccines presently available for treatment. The RNA-dependent RNA polymerase (RdRp) catalyzes the viral RNA synthesis and is an appropriate candidate for antiviral drug developments. We utilized integrated computational tools to build the 3D model of BEFV-RdRp and then predicted its probable active binding sites. The virtual screening and optimization against these active sites, using several small-molecule inhibitors from a different category of Life Chemical database and FDA-approved drugs from the ZINC database, was performed. We found nine molecules that have docking scores varying between -6.84 to -10.43 kcal/mol. Furthermore, these complexes were analyzed for their conformational dynamics and thermodynamic stability using molecular dynamics simulations in conjunction with the molecular mechanics generalized Born surface area (MM-GBSA) scheme. The binding free energy calculations depict that the electrostatic interactions play a dominant role in the RdRp-inhibitor binding. The hot spot residues, such as Arg565, Asp631, Glu633, Asp740, and Glu707, were found to control the RdRp-inhibitor interaction. The ADMET analysis strongly suggests favorable pharmacokinetics of these compounds that may prove useful for treating the BEFV ailment. Overall, we anticipate that these findings would help explore and develop a wide range of anti-BEFV therapy.Communicated by Ramaswamy H. Sarma.

Short-Term Humoral Immune Response of the pcDNA4-G Plasmid Expressing the Bovine Ephemeral Fever Virus G Gene in BALB/c Mice.

Bovine ephemeral fever (BEF) is an arthropod­borne viral disease characterised by a short­term clinical expression that can lead to significant losses in high­yielding cattle and water buffaloes. In this study, we aimed to generate a recombinant plasmid expressing the glycoprotein (G) of the BEF virus (BEFV) and to stimulate a humoral immune response to this protein in BALB / c mice immunised with the recombinant plasmid. Expression of the encoded protein was demonstrated by western blotting and immunoperoxidase tests. The suitable plasmids were intramuscularly administered to BALB/c mice on days 0, 14 and 21. The antibody response in the immunised mice was measured by a plaque reduction neutralization test (PRNT) and enzyme­linked immunosorbent assay (ELISA). According to BEFV ELISA, only two of the seven animals in these groups exceeded the cut­off value. A significant difference was observed in the mean OD values at 450 nm absorbance in the pcDNA4­G­immunised group when compared with those in the plasmid control group at 30 days (p < 0.05). According to PRNT50 results, a 1:20 (p < 0.05) antibody response was obtained at 30 days in pcDNA4­G (100 µg)­immunised mice, whereas this ratio was 1:80 (p < 0.001) in BEFV­immunised mice (1,000 PFU/0.5 ml). We conclude that the humoral immune response was stimulated in experimental mice immunised with the recombinant plasmid. However, disappointingly, the antibody response was markedly low in pcDNA4­G­immunised mice.

Complete genome sequencing and assessment of mutation-associated protein dynamics of the first Indian bovine ephemeral fever virus (BEFV) isolate.

BACKGROUND: Bovine ephemeral fever (BEF) is a re-emerging disease caused by bovine ephemeral fever virus (BEFV). Although it poses a huge economic threat to the livestock sector, complete viral genome information from any South Asian country, including India, lacks. AIM: Genome characterization of the first Indian BEFV isolate and to evaluate its genetic diversity by characterizing genomic mutations and their associated protein dynamics. MATERIALS AND METHODS: Of the nineteen positive blood samples collected from BEF symptomatic animals during the 2018-19 outbreaks in India, one random sample was used to amplify the entire viral genome by RT-PCR. Utilizing Sanger sequencing and NGS technology, a complete genome was determined. Genome characterization, genetic diversity and phylogenetic analyses were explored by comparing the results with available global isolates. Additionally, unique genomic mutations within the Indian isolate were investigated, followed by in-silico assessment of non-synonymous (NS) mutations impacts on corresponding proteins' secondary structure, solvent accessibility and dynamics. RESULTS: The complete genome of Indian BEFV has 14,903 nucleotides with 33% GC with considerable genetic diversity. Its sequence comparison and phylogenetic analysis revealed a close relatedness to the Middle Eastern lineage. Genome-wide scanning elucidated 30 unique mutations, including 10 NS mutations in the P, L and GNS proteins. The mutational impact evaluation confirmed alterations in protein structure and dynamics, with minimal effect on solvent accessibility. Additionally, alteration in the interatomic interactions was compared against the wild type. CONCLUSION: These findings extend our understanding of the BEFV epidemiological and pathogenic potential, aiding in developing better therapeutic and preventive interventions.

Carbonized Lysine-Nanogels Protect against Infectious Bronchitis Virus.

In this study, we demonstrate the synthesis of carbonized nanogels (CNGs) from an amino acid (lysine hydrochloride) using a simple pyrolysis method, resulting in effective viral inhibition properties against infectious bronchitis virus (IBV). The viral inhibition of CNGs was studied using both in vitro (bovine ephemeral fever virus (BEFV) and pseudorabies virus (PRV)) and in ovo (IBV) models, which indicated that the CNGs were able to prevent virus attachment on the cell membrane and penetration into the cell. A very low concentration of 30 µg mL-1 was found to be effective (>98% inhibition) in IBV-infected chicken embryos. The hatching rate and pathology of IBV-infected chicken embryos were greatly improved in the presence of CNGs. CNGs with distinctive virus-neutralizing activities show great potential as a virostatic agent to prevent the spread of avian viruses and to alleviate the pathology of infected avian species.

Bta-miR-101 suppresses BEFV replication via targeting NKRF.

MicroRNAs (miRNAs), as a kind of small noncoding RNAs, have been proved to play a regulatory role in virus infection. However, the role and mechanism of cellular miRNAs in bovine transient fever virus (BEFV) infection are largely unknown. In the present study, we found that bta-miR-101 was significantly up-regulated in the Madin-Darby Bovine Kidney (MDBK) cells upon BEFV infection. Notably, bta-miR-101 mimic dramatically inhibited BEFV replication, while bta-miR-101 inhibitor facilitated BEFV replication, suggesting that bta-miR-101 acted as an anti-viral host factor restraining BEFV replication. Subsequently, NF-κB repressing factor (NKRF) was identified as a target gene of bta-miR-101 by dual luciferase reporter assay, and bta-miR-101 mimic significantly down-regulated expression of NKRF, while bta-miR-101 inhibitor up-regulated its expression, respectively. Furthermore, NKRF could induce apoptosis, and favored the replication of BEFV. Finally, bta-miR-101 inhibited BEFV-induced apoptosis via targeting NKRF to suppress virus replication. In general, our study provides a novel mechanism for bta-miR-101 to exert its antiviral function, which provides a theoretical basis for the development of antiviral strategy.

RACK1 degrades MAVS to promote bovine ephemeral fever virus replication via upregulating E3 ubiquitin ligase STUB1.

Receptors for activated C kinase 1 (RACK1) could competitively combine with mitochondrial antiviral signaling protein (MAVS) to inhibit the type I interferon (IFN) signaling pathway during viral infection in vitro. However, whether RACK1 can degrade MAVS to enhance viral replication is still unknown. In this study, we found that bovine epidemic fever virus (BEFV) infection triggered the expression of RACK1. Overexpression of RACK1 promoted BEFV replication, while knockdown of RACK1 inhibited the replication of BEFV. Further research showed that RACK1 inhibited the type I IFN signaling pathway during BEFV infection by degrading MAVS, and RACK1 degraded MAVS via the ubiquitin-proteasome system. Mechanistically, RACK1 up-regulated the expression of E3 ubiquitin ligase STIP1 homology and U-box containing protein 1 (STUB1), thereby promoting the ubiquitination and degradation of MAVS. In addition, RACK1 degraded MAVS by enhancing the interaction between STUB1 and MAVS but not via its interaction with STUB1. Overall, our study reveals a novel mechanism by which RACK1 inhibits the type I IFN signaling pathway to BEFV infection through degradation of MAVS, thereby promoting viral infection. These findings provide a new perspective for the MAVS degradation regulated by RACK1.

Genome sequence of Bovine Ephemeral fever virus vaccine strain of South African origin.

Bovine Ephemeral fever virus (BEFV) is endemic in South Africa and has a negative economic impact on the meat and dairy industries. Bovine ephemeral fever or three-day stiff-sickness is controlled through annual vaccination with a live attenuated virus manufactured by Onderstepoort Biological Products (South Africa). We announce the genome sequences of two South African Bovine Ephemeral Virus strains; the live attenuated vaccine strain (14 876 nucleotides) and a field strain (14 883 nucleotides). A mutation in the alpha 3 open reading frame rendered the gene non-functional in both genomes. Phylogenetic analysis based on the glycoprotein gene showed that the two strains clustered with the South African lineage.

Isolation and genetic characterization of bovine ephemeral fever virus from epidemic-2020 in Turkey.

Bovine ephemeral fever virus (BEFV) infection occurs seasonally in many tropical and subtropical regions of Africa, Asia (including the Middle East), and Australia while it is exotic in Europe. In this study, the epidemiology of BEFV infection in Turkey that bridges southeastern Europe and Asia, geographically, was investigated according to the comparison of the nucleotide sequences of the virus caused the last epidemic in 2020 with those of the strains previously detected in Turkey as well as BEFV strains from other countries. In the phylogenetic analysis, based on an alignment of full-length G gene sequences, BEFVs from epidemic-2020 were located in Middle Eastern lineage and appear to represent most closely related BEFVs from India-2018 and 2019. The findings will contribute to a better understanding of BEFV epidemiology in Turkey.

Expression and immunogenicity of secreted forms of bovine ephemeral fever virus glycoproteins applied to subunit vaccine development.

AIMS: Vaccines for bovine ephemeral fever virus (BEFV) are available but are difficult to produce, expensive or suffer from genetic instability. Therefore, we designed constructs encoding C-terminally truncated forms (transmembrane anchoring region deleted) of glycoproteins G and GNS such that they were secreted from the cell into the media to achieve high-level antigen expression, correct glycosylation pattern and enable further simple purification with the V5 epitope tag. METHODS AND RESULTS: In this study, synthetic biology was employed to create membrane-bound and secreted forms of G and GNS glycoprotein. Mammalian cell culture was employed as an antigen expression platform, and the secreted forms of G and GNS protein were easily purified from media using a highly effective, single-step method. The V5 epitope tag was genetically fused to the C-termini of the proteins, enabling detection of the antigen through immunoblotting and immunomicroscopy. Our data demonstrated that the C-terminally truncated form of the G glycoprotein was efficiently secreted from cells into the cell media. Moreover the immunogenicity was confirmed in mice test. CONCLUSIONS: The immuno-dot blots showed that the truncated G glycoprotein was present in the total cell extract, and was clearly secreted into the media, consistent with the western blotting data and live-cell images. Our strategy presented the expression of secreted, epitope-tagged, forms of the BEFV glycoproteins such that appropriately glycosylated forms of BEFV G protein was secreted from the BHK-21 cells. This indicates that high-level expression of secreted G glycoprotein is a feasible strategy for large-scale production of vaccines and improving vaccine efficacy. SIGNIFICANCE AND IMPACT OF THE STUDY: The antigen expression strategy designed in this study can produce high-quality recombinant protein and reduce the amount of antigen used in the vaccine.

Genetically modified rabies virus vector-based bovine ephemeral fever virus vaccine induces protective immune responses against BEFV and RABV in mice.

Bovine ephemeral fever (BEF), caused by the bovine ephemeral fever virus (BEFV), is associated with an acute febrile infection in cattle and widespread in tropical and subtropical areas, leading to great economic losses to cattle and milk industry. However, no efficacious BEF vaccine is currently available in China. Herein, we generated a recombinant rabies virus (RABV) expressing BEFV glycoprotein (LBNSE-BG), utilizing a reverse genetics system based on the recombinant rabies virus strain LBNSE. It was found that mice immunized with LBNSE-BG produced robust neutralizing antibodies against both BEFV and RABV, and developed complete protection from lethal RABV challenge. Further studies showed that LBNSE-BG activated more dendritic cells (DCs), B cells and T cells in immunized mice than the parent virus LBNSE. Collectively, these findings demonstrate that the recombinant LBNSE-BG described here has the potential to be developed as a cost-effective and efficacious bivalent vaccine for cattle use in endemic areas of BEF and rabies.

Aspirin and 5-Aminoimidazole-4-carboxamide Riboside Attenuate Bovine Ephemeral Fever Virus Replication by Inhibiting BEFV-Induced Autophagy.

Recent study in our laboratory has demonstrated that BEFV-induced autophagy via activation of the PI3K/Akt/NF-κB and Src/JNK pathways and suppression of the PI3K-AKt-mTORC1 pathway is beneficial for virus replication. In the current study, we found that both aspirin and 5-aminoimidazole-4-carboxamide-1-ß-riboside (AICAR) siginificantly attenuated virus replication by inhibiting BEFV-induced autophagy via suppressing the BEFV-activated PI3K/Akt/NF-κB and Src/JNK pathways as well as inducing reversion of the BEFV-suppressed PI3K-Akt-mTORC1 pathway. AICAR reversed the BEFV-activated PI3K/Akt/NF-κB and Src/JNK pathways at the early to late stages of infection and induced reversion of the BEFV-suppressed PI3K-AKt-mTORC1 pathway at the late stage of infection. Our findings reveal that inhibition of BEFV-induced autophagy by AICAR is independent of AMPK. Furthermore, we found that AICAR transcriptionally downregulates the ATG related genes ULK1, Beclin 1, and LC3 and enhances Atg7 degradation by the proteasome pathway. Aspirin suppresses virus replication by inhibiting BEFV-induced autophagy. It directly suppressed the NF-κB pathway and reversed the BEFV-activated Src/JNK pathway at the early stage of infection and reversed the BEFV-suppressed PI3K/Akt/mTOR pathway at the late stage of infection. The current study provides mechanistic insights into the effects of aspirin and AICAR on BEFV replication through suppression of BEFV-induced autophagy.