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Background: Foot-and-mouth disease (FMD) is a devastating disease affecting cloven-hoofed animals, that leads to significant economic losses in affected countries and regions. Currently, there is an evident inclination towards the utilization of nanoparticles as powerful platforms for innovative vaccine development. Therefore, this study developed a ferritin-based nanoparticle (FNP) vaccine that displays a neutralizing epitope of foot-and-mouth disease virus (FMDV) VP1 (aa 140-158) on the surface of FNP, and evaluated the immunogenicity and protective efficacy of these FNPs in mouse and guinea pig models to provide a strategy for developing potential FMD vaccines., Results: This study expressed the recombinant proteins Hpf, HPF-NE and HPF-T34E via an E. coli expression system. The results showed that the recombinant proteins Hpf, Hpf-NE and Hpf-T34E could be effectively assembled into nanoparticles. Subsequently, we evaluated the immunogenicity of the Hpf, Hpf-NE and Hpf-T34E proteins in mice, as well as the immunogenicity and protectiveness of the Hpf-T34E protein in guinea pigs. The results of the mouse experiment showed that the immune efficacy in the Hpf-T34E group was greater than the Hpf-NE group. The results from guinea pigs immunized with Hpf-T34E showed that the immune efficacy was largely consistent with the immunogenicity of the FMD inactivated vaccine (IV) and could confer partial protection against FMDV challenge in guinea pigs., Conclusions: The Hpf-T34E nanoparticles stand out as a superior choice for a subunit vaccine candidate against FMD, offering effective protection in FMDV-infected model animals. FNP-based vaccines exhibit excellent safety and immunogenicity, thus representing a promising strategy for the continued development of highly efficient and safe FMD vaccines., (© 2024. The Author(s).)

The aim of this study was to produce E rns protein of bovine viral diarrhea virus (BVDV) by using suspensively cultured CHO cells expression system and to analyze the immunogenicity of the purified E rns protein. In this study, the recombinant eukaryotic expression plasmid pcDNA3.1-BVDV-E rns was constructed based on the gene sequence of BVDV-1 NADL strain. The E rns protein was secreted and expressed in cells supernatant after transfecting the recombinant expression plasmid pcDNA3.1-BVDV-E rns into CHO cells. The expression and purification of the E rns protein was analyzed by SDS-PAGE, the reactivity was determined with anti-His monoclonal antibodies and BVDV positive serum with Western blotting. Immunogenicity analysis of the E rns protein was determined after immunizing New Zealand white rabbits, and the serum antibodies were tested by indirect ELISA (iELISA) and indirect immunofluorescence (IFA). The serum neutralizing titer of the immunized rabbits was determined by virus neutralization test. The concentration of the purified E rns protein was up to 0.886 mg/mL by BCA protein quantification kit. The results showed that the E rns protein could be detected with anti-His monoclonal antibodies and anti-BVDV sera. Serum antibodies could be detected by iELISA on the 7th day post-prime immunization, and the antibody level was maintained at a high titer until the 28th day post-immunization. The antibody titer was 1:128 000. Furthermore, the expression of the E rns protein in BVDV-infected MDBK cells could be detected with immunized rabbits sera by IFA. Moreover, antigen-specific neutralizing antibodies of 2.71 log 10 was induced in rabbits. In this study, purified BVDV E rns protein was successfully produced using CHO suspension culture system, and the recombinant protein was proved to have a good immunogenicity, which may facilitate the development of BVD diagnosis method and novel subunit vaccine.

The African swine fever virus (ASFV) has caused a devastating pandemic in domestic and wild swine, causing economic losses to the global swine industry. Recombinant live attenuated vaccines are an attractive option for ASFV treatment. However, safe and effective vaccines against ASFV are still scarce, and more high-quality experimental vaccine strains need to be developed. In this study, we revealed that deletion of the ASFV genes DP148R , DP71L , and DP96R from the highly virulent isolate ASFV CN/GS/2018 (ASFV-GS) substantially attenuated virulence in swine. Pigs infected with 10 4 50% hemadsorbing doses of the virus with these gene deletions remained healthy during the 19-day observation period. No ASFV infection was detected in contact pigs under the experimental conditions. Importantly, the inoculated pigs were protected against homologous challenges. Additionally, RNA sequence analysis showed that deletion of these viral genes induced significant upregulation of the host histone H3.1 gene ( H3.1 ) and downregulation of the ASFV MGF110-7L gene. Knocking down the expression of H3.1 resulted in high levels of ASFV replication in primary porcine macrophages in vitro . These findings indicate that the deletion mutant virus ASFV-GS-Δ18R/NL/UK is a novel potential live attenuated vaccine candidate and one of the few experimental vaccine strains reported to induce full protection against the highly virulent ASFV-GS virus strain. IMPORTANCE Ongoing outbreaks of African swine fever (ASF) have considerably damaged the pig industry in affected countries. Thus, a safe and effective vaccine is important to control African swine fever spread. Here, an ASFV strain with three gene deletions was developed by knocking out the viral genes DP148R ( MGF360-18R ), NL ( DP71L ), and UK ( DP96R ). The results showed that the recombinant virus was completely attenuated in pigs and provided strong protection against parental virus challenge. Additionally, no viral genomes were detected in the sera of pigs housed with animals infected with the deletion mutant. Furthermore, transcriptome sequencing (RNA-seq) analysis revealed significant upregulation of histone H3.1 in virus-infected macrophage cultures and downregulation of the ASFV MGF110-7L gene after viral DP148R , UK , and NL deletion. Our study provides a valuable live attenuated vaccine candidate and potential gene targets for developing strategies for anti-ASFV treatment.

African swine fever virus (ASFV) causes significant morbidity and mortality in pigs worldwide. The lack of vaccines or therapeutic options warrants urgent further investigation. To this aim, we developed a rationally designed live attenuated ASFV-Δ110-9L/505-7R mutant based on the highly pathogenic Genotype II ASFV CN/GS/2018 backbone by deleting 2 well-characterized interferon inhibitors MGF110-9L and MGF505-7R. The mutant was slightly attenuated in vitro compared to parental ASFV but highly tolerant to genetic modifications even after 30 successive passages in vitro . Groups of 5 pigs were intramuscularly inoculated with increasing doses of the mutant, ranging from 10 3 to 10 6 hemadsorption units (HAD 50 ). Thirty-five days later, all groups were challenged with 10 2 HAD 50 of virulent parental ASFV. All the animals were clinically normal and devoid of clinical signs consistent with ASFV at the period of inoculation. In the virulent challenge, 2 animals from 10 3 HAD 50 -inoculated group and 1 animal from 10 4 HAD 50 -inoculated group were unprotected with severe postmortem and histological lesions. The rest of animals survived and manifested with relatively normal clinical appearance accompanied by tangible histological improvements in the extent of tissue damage. Meanwhile, antibody response, as represented by p30-specific antibody titers was positively correlated to protective efficacy, potentializing its usage as an indicator of protection. Moreover, compared to 1 dose, 2 doses provided additional protection, proving that 2 doses were better than 1 dose. The sufficiency in effectiveness supports the claim that our attenuated mutant may be a viable vaccine option with which to fight ASF. IMPORTANCE African swine fever virus (ASFV) is a causative agent of acute viral hemorrhagic disease of domestic swine which is associated with significant economic losses in the pig industry. The lack of vaccines or treatment options requires urgent further investigation. ASFV MGF110-9L and MGF505-7R, 2 well-characterized interferon inhibitors, were associated with viral virulence, host range, and immune modulation. In this study, a recombinant two-gene deletion ASFV mutant with deletion of MGF110-9L and MGF505-7R was constructed. The result showed that the mutant was safe, and also highly resistant to genetic modification even after 30 successive passages. High doses of our mutant (10 5 and 10 6 HAD 50 ) provided sterile immunity and complete protection in a virulent challenge. Two doses were superior to 1 dose and provided additional protection. This study develops a new ASFV-specific live attenuated vaccine and may be a viable vaccine option against ASF.

Multigene family (MGF) gene products are increasingly reported to be implicated in African swine fever virus (ASFV) virulence and attenuation of host defenses, among which the MGF360-9L and MGF505-7R gene products are characterized by convergent but distinct mechanisms of immune evasion. Herein, a recombinant ASFV mutant, ASFV-Δ9L/Δ7R, bearing combinational deletions of MGF360-9L and MGF505-7R, was constructed from the highly virulent ASFV strain CN/GS/2018 of genotype II that is currently circulating in China. Pigs inoculated intramuscularly with 10 4 50% hemadsorption doses (HAD 50 ) of the mutant remained clinically healthy without any serious side effects. Importantly, in a virulence challenge, all four within-pen contact pigs demonstrated clinical signs and pathological findings consistent with ASF. In contrast, vaccinated pigs (5/6) were protected and clinical indicators tended to be normal, accompanied by extensive tissue repairs. Similar to most viral infections, innate immunity and both humoral and cellular immune responses appeared to be vital for protection. Notably, transcriptome sequencing (RNA-seq) and quantitative PCR (qPCR) analysis revealed a regulatory function of the mutant in dramatic and sustained expression of type I/III interferons and inflammatory and innate immune genes in vitro . Furthermore, infection with the mutant elicited an early and robust p30-specific IgG response, which coincided and was strongly correlated with the protective efficacy. Analysis of the cellular response revealed a strong ASFV-specific interferon gamma (IFN-γ) response and immunostaining of CD4 + T cells coupled with a high level of CD163 + macrophage infiltration in spleens of vaccinated pigs. Our study identifies a new mechanism of immunological regulation by ASFV MGFs that rationalizes the design of live attenuated vaccine for implementation of improved control strategies to eradicate ASFV. IMPORTANCE Currently, the deficiency in commercially available vaccines or therapeutic options against African swine fever constitutes a matter of major concern in the swine industry globally. Here, we report the design and construction of a recombinant ASFV mutant harboring combinational deletions of interferon inhibitors MGF360-9L and MGF505-7R based on a genotype II ASFV CN/GS/2018 strain currently circulating in China. The mutant was completely attenuated when inoculated at a high dose of 10 4 HAD 50 . In the virulence challenge with homologous virus, sterile immunity was achieved, demonstrating the mutant's potential as a promising vaccine candidate. This sufficiency of effectiveness supports the claim that this live attenuated virus may be a viable vaccine option with which to fight ASF.

Seneca Valley virus (SVV) infection in pigs is associated with porcine idiopathic vesicular disease (PIVD). Outbreaks of SVV infection in pig herds have been reported in several Asia and Americas countries. Recently, a series of outbreaks of SVV infection occurred in China, Canada, Thailand and the United States. However, no available vaccines have been developed to limit the transmission of SVV. The SVV CH-FJ-2017 from Fujian province in China is a representative of the epidemic strains, and shows 98.5-99.9% capsid protein amino acid identity with the recent SVV strains. In the present study, we developed a SVV CH-FJ-2017 inactivated vaccine. The SVV was produced by cultivation of BHK-21 cells in roller bottles, inactivated with binary ethylenimine, and mixed with oil adjuvant (Montanide ISA). The immunogenicity of the inactivated vaccine in pigs was evaluated by neutralizing test, and the immunized pigs were challenged with SVV CH-FJ-2017. The results showed that animals receiving one dose of the inactivated vaccine (2 μg/dose) with oil adjuvant developed high neutralizing antibody titers and showed no clinical signs after virus challenge comparing with the non-vaccinated animals, indicating a good protective efficacy of the produced vaccine against SVV infection. This is the first reported SVV vaccine that can be used for control of SVV infection in pigs., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Foot-and-mouth disease (FMD) is a highly contagious vesicular disease that affects domestic and wild cloven-hoofed animals worldwide. Recently, a series of outbreaks of type A FMDV occurred in Southeast Asian countries, China, the Russia Federation, Mongolia, Kazakhstan and South Korea. The FMD virus (A/GDMM/CHA/2013) from China's Guangdong province (2013) is representative of those responsible for the latest epidemic, and has low amino acid identity (93.9%) in VP1 protein with the epidemic strain A/WH/CHA/09 from Wuhan, China in 2009. Both of isolates belong to the Sea-97 genotype of ASIA topotype. Therefore, the application of a new vaccine strain with cross-protective efficacy is of fundamental importance to control the spread of the two described pandemic strains. A chimeric strain rA/P1-FMDV constructed by our lab previously through replacing the P1 gene in the vaccine strain O/CHA/99 with that from the epidemic stain A/WH/CHA/09, has been demonstrated to exhibit good growth characteristics in culture, and the rA/P1-FMDV inactivated vaccine can provide protection against epidemic strain A/WH/CHA/09 in cattle. However, it is still unclear whether the vaccine produces efficient protection against the new pandemic strain (A/GDMM/CHA/2013). Here, vaccine matching and pig 50% protective dose (PD50) tests were performed to assess the vaccine potency. The vaccine matching test showed cross-reactivity of sera from full dose vaccine vaccinated pigs with A/WH/CHA/09 and A/GDMM/CHA/2013 isolates, with average r1 values of 0.94±0.12 and 0.68±0.06 (r1≥0.3), which indicates that the rA/P1-FMDV vaccine is likely to confer good cross-protection against the two isolates. When challenged with two pandemic isolates A/WH/CHA/09 and A/GDMM/CHA/2013 strain, the vaccine achieved 12.51 PD50 and 10.05 PD50 per dose (2.8μg), respectively. The results indicated that the rA/P1-FMDV inactivated vaccine could protect pigs against both A/WH/CHA/09 and A/GDMM/CHA/2013 pandemic isolates., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

The foot-and-mouth disease virus (FMDV) is the causative agent of FMD, a highly infectious and devastating viral disease of domestic and wild cloven-hoofed animals. FMD affects livestock and animal products' national and international trade, causing severe economic losses and social consequences. Currently, inactivated vaccines play a vital role in FMD control, but they have several limitations. The genetic code expansion technology provides powerful strategies for generating premature termination codon (PTC)-harbouring virus as a live but replication-incompetent viral vaccine. However, this technology has not been explored for the design and development of new FMD vaccines. In this study, we first expanded the genetic code of the FMDV genome via a transgenic cell line containing an orthogonal translation machinery. We demonstrated that the transgenic cells stably integrated the orthogonal pyltRNA/pylRS pair into the genome and enabled efficient, homogeneous incorporation of unnatural amino acids into target proteins in mammalian cells. Next, we constructed 129 single-PTC FMDV mutants and four dual-PTC FMDV mutants after considering the tolerance, location, and potential functions of those mutated sites. Amber stop codons individually substituted the selected amino acid codons in four viral proteins (3D, L, VP1, and VP4) of FMDV. We successfully rescued PTC-FMDV mutants, but the amber codon unexpectedly showed a highly degree of mutation rate during PTC-FMDV packaging and replication. Our findings highlight that the genetic code expansion technology for the generation of PTC-FMD vaccines needs to be further improved and that the genetic stability of amber codons during the packaging and replication of FMDV is a concern. [ABSTRACT FROM AUTHOR]