Recombinant pseudorabies virus with gI/gE deletion generated by overlapping polymerase chain reaction and homologous recombination technology induces protection against the PRV variant PRV-GD2013

Abstract Background Since 2011, numerous highly virulent and antigenic variant viral strains have been reported in pigs that were vaccinated against the swine pseudorabies virus. These infections have led to substantial economic losses in the Chinese swine industry. Results This study, constructed a novel recombinant vaccine strain with gI/gE deletion (PRV-GD2013-ΔgI/gE) by overlapping PCR and homologous recombination technology. The growth curves and plaque morphology of the recombinant virus were similar to those of the parental strain. However, PRV-GD2013-ΔgI/gE infection was significantly attenuated in mice compared with that of PRV-GD2013. Two-week-old piglets had normal rectal temperatures and displayed no clinical symptoms after being inoculated with 105 TCID50 PRV-GD2013-ΔgI/gE, indicating that the recombinant virus was avirulent in piglets. Piglets were immunized with different doses of PRV-GD2013-ΔgI/gE, or a single dose of Bartha-K61 or DMEM, and infected with PRV-GD2013 at 14 days post-vaccination. Piglets given high doses of PRV-GD2013-ΔgI/gE showed no obvious clinical symptoms, and their antibody levels were higher than those of other groups, indicating that the piglets were completely protected from PRV-GD2013. Conclusions The PRV-GD2013-ΔgI/gE vaccine strain could be effective for immunizing Chinese swine herds against the pseudorabies virus (PRV) strain.

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Meclizine Inhibits Pseudorabies Virus Replication by Interfering With Virus Entry and Release

Frontiers in Microbiology ◽

10.3389/fmicb.2021.795593 ◽

2021 ◽

Vol 12 ◽

Author(s):

Panrao Liu ◽

Danhe Hu ◽

Lili Yuan ◽

Zhengmin Lian ◽

Xiaohui Yao ◽

...

Keyword(s):

Viral Entry ◽

Pseudorabies Virus ◽

Molecular Mechanisms ◽

Clinical Symptoms ◽

Antiviral Effect ◽

Inhibitory Effect ◽

Economic Losses ◽

Swine Industry ◽

Viral Attachment ◽

Viral Loads

Pseudorabies virus (PRV) is a pathogen that causes substantial economic losses to the swine industry. With the emergence and widespread of PRV variants since 2011 in China, current commercial vaccines cannot provide complete protection against PRV infection. Therefore, antiviral drugs may work as an alternative way to control and prevent PRV. In this study, the inhibitory effects and underlying molecular mechanisms of meclizine against PRV were studied. Meclizine displayed a significant inhibitory effect against PRV when it was added before, simultaneously with, or after virus infection. The inhibitory effect of meclizine occurred during viral entry and cell-to-cell spreading but not at viral attachment into PK-15 cells. Meclizine also inhibited viral particle release at the late stage of infection. The antiviral effect of meclizine was tested in mice, and the results showed that meclizine reduced the severity of clinical symptoms and the viral loads in tissues, and delayed the death, after PRV challenge. The above results indicated that meclizine had an inhibitory effect on PRV. Our findings will contribute to the development of potential therapeutic drugs against PRV infection.

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Development of Dual Fluorescent Microsphere Immunological Assay for Detection of Pseudorabies Virus gE and gB IgG Antibodies

10.21203/rs.3.rs-24925/v1 ◽

2020 ◽

Author(s):

chihai ji ◽

Jingyu Wang ◽

Yuchen Zeng ◽

Haoming Pan ◽

Yingfang Wei ◽

...

Keyword(s):

Pseudorabies Virus ◽

High Specificity ◽

Antibody Detection ◽

Economic Losses ◽

Igg Antibodies ◽

Wild Type ◽

Swine Industry ◽

Igg Antibody ◽

Fluorescent Microsphere ◽

Specificity And Sensitivity

Abstract Background Pseudorabies, also known as Aujezsky’s disease, is an acute viral infection caused by pseudorabies virus (PRV). Swine are one of the natural hosts of pseudorabies, therefore, the disease brings huge economic losses to the swine industry. Establishment of a differential diagnosis technique that can distinguish between wild-type infected and vaccinated pigs, and monitor vaccine-induced IgG is crucial for eventual eradication of pseudorabies.Results The aim of this study was to develop a rapid dual detection method for PRV gE and gB protein IgG antibodies with high specificity and sensitivity. PRV gE codons at amino acid residues (aa) 52–238 and gB codons at aa 539–741 were expressed to obtain recombinant PRV gE and gB proteins by pMAL-c5x vector. After purification with Qiagen Ni–NTA agarose affinity, the two proteins were analyzed by SDS-PAGE and immunoblotting assay. Two single fluorescent-microsphere immunoassays (FMIA) were established by coupling two recombinant proteins (gE and gB) with two magnetic microbeads and an effective dual FMIA was developed by integrating the two single assays. Optimal serum dilution for each assay, correlation with other common swine virus-positive sera and comparison with ELISA for two PRV antigens were tested for validation. Compared with ELISA, the specificity and sensitivity were 99.26% and 92.3% for gE IgG antibody detection and 95.74% and 96.3% for gB IgG antibody detection by dual-FMIA.Conclusion We provide a new method for monitoring PRV protective antibody in vaccinated pigs and differentiating wild-type-PRV-infected from vaccinated pigs

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The Attenuated Pseudorabies Virus Vaccine Strain Bartha K61: A Brief Review on the Knowledge Gathered during 60 Years of Research

Pathogens ◽

10.3390/pathogens9110897 ◽

2020 ◽

Vol 9 (11) ◽

pp. 897

Author(s):

Jonas L. Delva ◽

Hans J. Nauwynck ◽

Thomas C. Mettenleiter ◽

Herman W. Favoreel

Keyword(s):

Vaccine Strain ◽

Pseudorabies Virus ◽

Economic Losses ◽

Vaccination Programs ◽

Live Vaccines ◽

Vector Vaccine ◽

Short Summary ◽

Aujeszky's Disease ◽

Aujeszky’S Disease ◽

Disease Epidemics

Pseudorabies virus (PRV) is a member of the alphaherpesvirus subfamily of the herpesviruses and is the causative agent of Aujeszky’s disease in pigs, causing respiratory, neurological, and reproductive symptoms. Given the heavy economic losses associated with Aujeszky’s disease epidemics, great efforts were made to develop efficacious vaccines. One of the best modified live vaccines to this day is the attenuated Bartha K61 strain. The use of this vaccine in extensive vaccination programs worldwide has assisted considerably in the eradication of PRV from the domesticated pig population in numerous countries. The Bartha K61 strain was described in 1961 by Adorján Bartha in Budapest and was obtained by serial passaging in different cell cultures. Ever since, it has been intensively studied by several research groups, for example, to explore its efficacy as a vaccine strain, to molecularly and mechanistically explain its attenuation, and to use it as a retrograde neuronal tracer and as a vector vaccine. Given that the Bartha K61 vaccine strain celebrates its 60th birthday in 2021 with no sign of retirement, this review provides a short summary of the knowledge on its origin, characteristics, and use as a molecular tool and as a vaccine.

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The use of tylvalosin (Aivlosin®) in the successful elimination of swine dysentery on a farrow-to-finish herd

Vlaams Diergeneeskundig Tijdschrift ◽

10.21825/vdt.v81i4.18333 ◽

2012 ◽

Vol 81 (4) ◽

pp. 205-210

Author(s):

P. Vyt ◽

L. Vandepitte ◽

A. Dereu ◽

M. Roozen

Keyword(s):

Management Practices ◽

Clinical Symptoms ◽

Monitoring Program ◽

Single Site ◽

Economic Losses ◽

Antimicrobial Use ◽

Rodent Control ◽

Feed Conversion ◽

Swine Industry ◽

Swine Dysentery

Swine dysentery causes severe economic losses in swine industry. Eliminating the disease at farm level can become problematic when resistance of the causative agent, Brachyspira hyodysenteriae, against pleuromutilins is reported. In this study, the use of tylvalosin (Aivlosin®) in eliminating the disease in a single-site, farrow-to-finish herd was evaluated. In addition, productivity parameters and antimicrobial use were compared prior to and after implementing the elimination protocol. On a mixed farm of 200 sows and 1500 finishers with a history of chronic dysentery, the B. hyodysenteriae isolate was resistant to pleuromutilins but had a low minimum inhibitory concentration for tylvalosin (2 μg/ml). Combined with a strict program for rodent control and hygiene, sows were treated with tylvalosin at a dose of 4.25 mg/kg BW daily for four weeks. The sows were washed one week after the start of the treatment before entering a clean stable. Piglets born from sows that had received this treatment, were considered free from dysentery and were kept separated from infected, untreated animals on the farm. A monitoring program with monthly sampling of sows and fatteners was installed to evaluate the absence of B. hyodysenteriae on the farm. After treatment, the clinical symptoms in the treated sows disappeared and remained absent in the offspring born after the procedure. Fecal samples examined by PCR remained negative for the whole testing period (14 months after the end of the treatment) and no clinical outbreaks were reported afterwards. The feed conversion ratio improved by 12%, the mortality rate with 37% and the antimicrobial use decreased by 71.5%. We conclude that the elimination of swine dysentery on a single-site, farrow-to-finish herd is possible using tylvalosin (AivlosinR) combined with strict hygiene and rodent control. The improvements of the technical parameters in this study are not only the result of the elimination of B. hyodysenteriae, but also changes in management practices influenced these parameters.

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Construction of a Quadruple Gene-deleted Vaccine Confers Complete Protective Immunity Against Emerging PRV Variant Challenge in Piglets

10.21203/rs.3.rs-985687/v1 ◽

2021 ◽

Author(s):

Leqiang Sun ◽

Yajie Tang ◽

Keji Yan ◽

Huawei Zhang

Keyword(s):

Pseudorabies Virus ◽

Clinical Signs ◽

Economic Losses ◽

Swine Industry ◽

Live Attenuated Vaccine ◽

Specific Antibodies ◽

Variant Strain ◽

Attenuated Vaccine ◽

Suckling Piglets ◽

Full Protection

Abstract Background: Pseudorabies virus (PRV) causes Aujeszky’s disease or pseudorabies (PR) in pigs worldwide, which leads to heavy economic losses to the swine industry. Pigs are the natural host, meanwhile, animals such as dogs, cats, foxes, rabbits, cattle and sheep are susceptible to infection. In 2011, the emerging PRV variant led to the outbreak of PR in Bar-tha-K61-vaccinated pigs. The PR outbreaks demonstrated that the Bartha-K61 vaccine did not provide full protection against the emerging PRV variant. It is widely believed that PRV live-attenuated vaccines could control PRV infection. Methods: In this study, we developed a novel PRV live-attenuated vaccine by deleting its gI, gE, US9, and US2 genes through CRISPR/Cas9, which was named PRV GDFS-delgI/gE/US9/US2.Results: Safety experiments confirmed that PRV GDFS-delgI/gE/US9/US2 was safe for 5 to 7-day-old suckling piglets. Piglets immunized with the PRV GDFS-delgI/gE/US9/US2 vaccine did not produce PRV gE-specific antibodies but could generate PRV gB-specific antibodies and high neutralizing titers against the PRV GDFS strain (variant PRV strain) or PRV Ea strain (older PRV strain). After challenge with the emerging PRV GDFS variant, none of the piglets immunized with the PRV GDFS-delgI/gE/US9/US2 vaccine showed any clinical signs, and their rectal temperatures were normal. Moreover, the autopsy and histopathological analyses revealed that the piglets in the PRV GDFS-delgI/gE/US9/US2 vaccine group did not show apparent gross or pathological lesions. Furthermore, the piglets in the PRV GDFS-delgI/gE/US9/US2 vaccine groups did not present weight loss. According to the criteria of the OIE terrestrial manual, the results of the experiment confirmed that the PRV GDFS-delgI/gE/US9/US2 vaccine could provide full protection against the emerging PRV variant strain in piglets. Conclusions: The PRV GDFS-delgI/gE/US9/US2 strain is a potential new live-attenuated vaccine against emerging PRV variant strain infections in China.

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ASFV bearing an I226R gene-deletion elicits a robust immunity in pigs to African swine fever

Journal of Virology ◽

10.1128/jvi.01199-21 ◽

2021 ◽

Author(s):

Yanyan Zhang ◽

Junnan Ke ◽

Jingyuan Zhang ◽

Jinjin Yang ◽

Huixian Yue ◽

...

Keyword(s):

Vaccine Candidate ◽

Clinical Symptoms ◽

African Swine Fever Virus ◽

Clinical Signs ◽

African Swine Fever ◽

Parental Virus ◽

Economic Losses ◽

Egfp Expression ◽

Post Inoculation ◽

Swine Industry

African swine fever (ASF) is a severe hemorrhagic infectious disease in pigs caused by the African swine fever virus (ASFV), leading to devastating economic losses in the epidemic regions. Its control currently depends on thorough culling and clearance of the diseased and the surrounding suspected pigs. ASF vaccine has been extensively explored for years worldwide, especially in hog-intensive areas where it is highly desired, but it is still unavailable due to numerous reasons. Herein, we reported another ASF vaccine candidate named SY18ΔI226R bearing a deletion of the I226R gene in replacement of an eGFP expression cassette at the right end of the viral genome. This deletion results in complete loss of virulence of SY18 as the gene-deleted strain does not cause any clinical symptoms in all pigs inoculated with either a dosage of 10 4.0 TCID 50 or 10 7.0 TCID 50 . An apparent viremia with the gradual decline was monitored, while the virus shedding was only occasionally detected in oral- or anal swabs. ASFV specific antibody appeared at 9 days post-inoculation. After intramuscular challenge with its parental strain ASFV SY18 on 21 days post inoculation, all the challenged pigs survived without obvious febrile or abnormal clinical signs. No viral DNA could be detected on the dissection of any tissue when viremia disappeared. These indicated that SY18ΔI226R is safe in swine and elicits a robust immunity to the virulent ASFV infection. IMPORTANCE: Outbreaks of African swine fever have resulted in devastating losses to the swine industry worldwide, but there is currently no commercial vaccine available. Although several vaccine candidates have been reported, none has been approved for use due to several reasons, especially the ones concerning bio-safety. Here, we identified a new undescribed functional gene, I226R. When deleted from the ASFV genome, the virus completely loses its virulence in the swine. Importantly, pigs infected with this gene-deleted virus were resistant to infection by an intramuscular challenge of 10 2.5 or 10 4.0 TCID 50 of its virulent parental virus. Furthermore, rarely the nucleic acid of the gene-deleted virus and its virulent parental virus was detected from oral- or anal swabs. Viruses could not be detected in any tissues after necropsy when viremia became negative, indicating that robust immunity was achieved. Therefore, SY18ΔI226R is a novel, ideal and efficacious vaccine candidate for genotype II ASF.

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Detection of Pseudorabies Virus in Wild Boar Foetus

Animals ◽

10.3390/ani10020366 ◽

2020 ◽

Vol 10 (2) ◽

pp. 366 ◽

Cited By ~ 1

Author(s):

Maria Irene Pacini ◽

Mario Forzan ◽

Giovanni Cilia ◽

Lucrezia Bernardini ◽

Filippo Marzoli ◽

...

Keyword(s):

Wild Boar ◽

Pseudorabies Virus ◽

Economic Losses ◽

Limited Information ◽

Swine Industry ◽

Nasal Swabs ◽

Infected Area ◽

Reproductive Consequences ◽

Embryonic Death

Pseudorabies, or Aujeszky’s disease, is a notifiable worldwide infection of domestic and feral swine that causes economic losses for the swine industry. In domestic pigs, the virus is responsible for nervous and/or respiratory symptoms; in pregnant sows, it is one of the major causes of stillbirth, mummification, embryonic death, and infertility (SMEDI). It is known that PRV infection in wild boar is associated with low pathogenicity and attenuated or absent symptomatology, but limited information is available about the ability of the virus to infect the foetuses of infected wild boar pregnant sows. Due to scarce information about the reproductive consequences, we investigate the possible intrauterine vertical transmission of the virus in wild boar pregnant sow living in a highly infected area. A number of 54 hunted wild boar were sampled during 2018–2019, and blood, genital and nasal swabs, placenta, and fetuses were collected for serological and molecular investigations. A seroprevalence of 74% (40/54) was detected, while 1/24 pregnant sow and 1/24 pooled foetuses tested positive by PCR (gene gB). This is the first evidence of viral detection in foetuses from seropositive pregnant wild boar. This finding suggests the possible pathogenetic role of PRV on pregnancy in wild boar and the existence of an additional transmission route.

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Outbreak of Porcine Reproductive and Respiratory Syndrome Virus 1 in Taiwan

Viruses ◽

10.3390/v12030316 ◽

2020 ◽

Vol 12 (3) ◽

pp. 316 ◽

Cited By ~ 2

Author(s):

Wei-Hao Lin ◽

Kraijak Kaewprom ◽

Sheng-Yuan Wang ◽

Chuen-Fu Lin ◽

Cheng-Yao Yang ◽

...

Keyword(s):

Vaccine Strain ◽

Respiratory Symptoms ◽

Laboratory Tests ◽

Economic Losses ◽

Respiratory Syndrome Virus ◽

Swine Industry ◽

European Isolate ◽

Prrs Virus ◽

Suckling Piglets ◽

Central Taiwan

Porcine reproductive and respiratory syndrome (PRRS) causes significant economic losses in the swine industry worldwide. The PRRS virus (PRRSV) can be divided into two species, PRRSV 1 (European) and PRRSV 2 (North American). In Taiwan, PRRSV 2 isolates are dominant and cause respiratory symptoms in nursing pigs. From October to November 2018, in a pig herd in central Taiwan, pregnant sows had abortions and stillbirths, and piglets suffered from respiratory disorders. Laboratory tests identified the presence of PRRSV 1 in serum from sows and suckling piglets in this scenario. The complete genome of the identified PRRSV 1 strain was genetically closely related to that of a European PRRSV vaccine strain (98.2%). This local European isolate is designated as PRRSV/NPUST-2789-3W-2/TW/2018 (NPUST2789). This report is the first to indicate an outbreak in Taiwan of a PRRSV 1 strain that shares a common evolutionary ancestor with the European PRRSV vaccine strain.

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Isatis root polysaccharide promotes maturation and secretory function of monocyte-derived dendritic cells

BMC Complementary Medicine and Therapies ◽

10.1186/s12906-020-03103-2 ◽

2020 ◽

Vol 20 (1) ◽

Author(s):

Xuebing Wang ◽

Zewen Chen ◽

Tong Chen ◽

Xiao Li ◽

Shucheng Huang ◽

...

Keyword(s):

Dendritic Cells ◽

Pseudorabies Virus ◽

Molecular Mechanisms ◽

Peripheral Blood Monocyte ◽

Economic Losses ◽

Enzyme Linked Immunosorbent Assay ◽

Swine Industry ◽

Chinese Medicinal Herb ◽

Animal Virus ◽

Immature Dendritic Cells

Abstract Background Pseudorabies virus (PRV) is an animal virus that is globally responsible for the high economic losses in the swine industry. Isatis root is a traditional Chinese medicinal herb that possesses immune-enhancing and antiviral properties. However, the molecular mechanisms underlying the effects of the active component of the isatis root polysaccharide (IRPS) extract on immature dendritic cells remain elusive. Methods In this study, we investigated the molecular changes in primary porcine peripheral blood monocyte-derived dendritic cells (MoDCs) during PRV infection, using enzyme-linked immunosorbent assay (ELISA) and quantitative reverse transcription-polymerase chain reaction. Additionally, we studied the effect of IRPS on PRV-infected DCs. Results The results showed that IRPS stimulated the maturation of MoDCs, induced IL-12 secretion, and downregulated IL-6 expression. Conclusions Collectively, these results suggest that IRPS is a promising candidate for promoting maturation of DCs and enhancing their secretory potential after PRV infection.

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G-Quadruplexes Formation at the Upstream Region of Replication Origin (OriL) of the Pseudorabies Virus: Implications for Antiviral Targets

Viruses ◽

10.3390/v13112219 ◽

2021 ◽

Vol 13 (11) ◽

pp. 2219

Author(s):

Yance Zhu ◽

Wenhao Liu ◽

Chao Zhang

Keyword(s):

Pseudorabies Virus ◽

Resonance Energy Transfer ◽

Resonance Energy ◽

Cd Spectroscopy ◽

Host Cells ◽

Upstream Region ◽

Economic Losses ◽

Swine Industry ◽

Taq Polymerase

Pseudorabies virus (PRV) is the causative agent of Aujeszky’s disease, which still causes large economic losses for the swine industry. Therefore, it is urgent to find a new strategy to prevent and control PRV infection. Previous studies have proven that guanine (G)-rich DNA or RNA sequences in some other viruses’ genomes have the potential to form G-quadruplex (G4), which serve as promising antivirus targets. In this study, we identified two novel G4-forming sequences, OriL-A and OriL-S, which are located at the upstream origin of replication (OriL) in the PRV genome and conserved across 32 PRV strains. Circular dichroism (CD) spectroscopy and a gel electrophoresis assay showed that the two G-rich sequences can fold into parallel G4 structures in vitro. Moreover, fluorescence resonance energy transfer (FRET) melting and a Taq polymerase stop assay indicated that the G4 ligand PhenDC3 has the capacity to bind and stabilize the G4. Notably, the treatment of PRV-infected cells with G4-stabilizer PhenDC3 significantly inhibited PRV DNA replication in host cells but did not affect PRV’s attachment and entry. These results not only expand our knowledge about the G4 characteristics in the PRV genome but also suggest that G4 may serve as an innovative therapeutic target against PRV.

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