Evaluation of classical swine fever E2 (CSF-E2) subunit vaccine efficacy in the prevention of virus transmission and impact of maternal derived antibody interference in field farm applications.

BACKGROUND: Classical swine fever (CSF) is one of the most devastating pig diseases that affect the swine industry worldwide. Besides stamping out policy for eradication, immunization with vaccines of live attenuated CSF or the CSF-E2 subunit is an efficacious measure of disease control. However, after decades of efforts, it is still hard to eliminate CSF from endemically affected regions and reemerging areas. Most of previous studies demonstrated the efficacy of different CSF vaccines in laboratories under high containment conditions, which may not represent the practical performance in field farms. The inadequate vaccine efficacy induced by unrestrained factors may lead to chronic or persistent CSF infection in animals that develop a major source for virus shedding among pig populations. In this study, a vaccination-challenge-cohabitation trial on specific-pathogen-free (SPF) pigs and long-term monitoring of conventional sows and their offspring were used to evaluate the efficacy and the impact of maternally derived antibody (MDA) interference on CSF vaccines in farm applications. RESULTS: The trials demonstrated higher neutralizing antibody (NA) titers with no clinical symptoms and significant pathological changes in the CSF-E2 subunit vaccine immunized group after CSFV challenge. Additionally, none of the sentinel pigs were infected during cohabitation indicating that the CSF-E2 subunit vaccine could provoke adequately acquired immunity to prevent horizontal transmission. In field farm applications, sows immunized with CSF-E2 subunit vaccine revealed an average of higher and consistent antibody level with significant reduction of CSF viral RNA detection via saliva monitoring in contrast to those of live attenuated CSF vaccine immunized sows possessing diverse antibody titer distributions and higher viral loads. Furthermore, early application of the CSF-E2 subunit vaccine in 3-week-old piglets illustrated no MDA interference on primary immunization and could elicit consistent and long-lasting adequate antibody response suggesting the flexibility of CSF-E2 subunit vaccine on vaccination program determination. CONCLUSIONS: The CSF-E2 subunit vaccine demonstrated significant efficacy and no MDA interference for immunization in both pregnant sows and piglets. These advantages provide a novel approach to avoid possible virus shedding in sow population and MDA interference in piglets for control of CSF in field farm applications.

The impact of porcine circovirus associated diseases on live attenuated classical swine fever vaccine in field farm applications.

Porcine circovirus associated diseases (PCVADs) are among the most important diseases affecting the worldwide swine industry. Vaccination against porcine circovirus type 2 (PCV2) infection has been utilized for disease control and effectively reduces clinical signs of PCVADs. To evaluate the efficacy of the PCV2 vaccine in field farms, we conducted a trial using conventional pigs immunized with the subunit PCV2 vaccine followed by PCV2 challenge. Immunized pigs demonstrated lower serum viral loads, less viral antigen staining in lymph nodes, and higher average daily weight gain, confirming the protective efficacy of the vaccine. However, low levels of PCV2 infection were still detected in vaccinated pigs after challenge, suggesting that the PCV2 vaccine was unable to eradicate the virus, which could lead to asymptomatic PCV2 subclinical infection (PCV2-SI) in pig farms. Additionally, PCV2 infection is a risk factor for impaired pig immune response development during the weaning to growth stages, which is a crucial period to receive vaccines against classical swine fever (CSF). Therefore, the impact of PCV2-SI or PCV2-systemic disease (PCV2-SD) on live attenuated CSF vaccine was investigated. After PCV2 challenge, there was no difference in levels of classical swine fever virus (CSFV) neutralizing antibodies (NA) between pigs with PCV2-SD and PCV2-SI, suggesting that the efficacy of CSF vaccine was compromised. Moreover, results of long-term monitoring of CSFV NA titers in PCV2-SI pigs with minimized interference by maternally-derived antibodies suggested that serum PCV2 viral loads greater than 102 copies/mL may compromise the efficacy of CSF vaccine. Overall, a conventional pig model was established to demonstrate the impaired efficacy of the subunit PCV2 vaccine and its impact on the CSF vaccine in vaccination-challenge trials. Additionally, the impaired efficacy of the PCV2 vaccine resulted in increased PCV2-SI, eventually leading to compromised the live attenuated CSF vaccine induced NA response in field farm applications.

Growth properties and vaccine efficacy of recombinant pseudorabies virus defective in glycoprotein E and thymidine kinase genes.

Pseudorabies virus (PRV) is an alphaherpesvirus that causes pseudorabies (PR), an economically important viral disease of pigs. Marker vaccines were widely used in PR prevention and eradication programs. The purpose of this study was to construct a novel recombinant virus with deletions at defined regions in the glycoprotein E (gE) and thymine kinase (TK) genes by homologous recombination. This study also evaluated the safety and efficacy of the virus for a live attenuated marker vaccine. No significant difference was observed in virus replication between gE gene-deleted (gE(-)), gE/TK double gene-deleted (gE(-)TK(-)), and wild-type PRV by growth curve analysis. However, gE(-)TK(-) PRV was completely attenuated in mice. To evaluate the immunogenicity of gE(-)TK(-) PRV, four 12-week-old specific-pathogen-free pigs per group were immunized intramuscularly with viral titers of 1×10(4), 1×10(5), or 1×10(6) TCID50, followed by intranasal challenge infection with virulent PRV (1×10(8) TCID50) at 3 weeks post vaccination. The gE(-)TK(-) PRV-vaccinated pigs displayed no general adverse effects after immunization and had protective immune responses after PRV challenge. Thus, gE(-)TK(-) PRV was safe and efficacious and might be a potential candidate for a live attenuated marker vaccine against PRV.

Expression and immunological studies of classical swine fever virus glycoprotein E2 in the bi-cistronic baculovirus/larvae expression system.

To develop an economical, easy technique for producing recombinant E2 glycoprotein (rE2) of classical swine fever virus (CSFV) as a candidate immunogen, a bi-cistronic baculovirus/larvae expression vector was constructed using p10 promoter, an internal ribosome entry site, and the gfp gene. Trichoplusia ni larvae were successfully infected with the occluded recombinant baculovirus via feed, and the characteristics of rE2 were confirmed by immunoblot and glycosylation stain. rE2 at a concentration of 0.6-0.8 mg/ml without degradation was obtained from hemolymphs of infected larvae that emitted high levels of green fluorescence. Immunization assays indicated that mice and piglets immunized with rE2-containing hemolymph elicited high titers of anti-CSFV E2 antibodies with virus-neutralizing activity. This is the first study to indicate that baculovirus/T. ni larvae-expressed rE2 can be served as a vaccine candidate. This system provides an economical alternative for the production of vaccine components in the veterinary industry.

Mutations in the Salmonella enterica serovar Choleraesuis cAMP-receptor protein gene lead to functional defects in the SPI-1 Type III secretion system.

Salmonella enterica serovar Choleraesuis (Salmonella Choleraesuis) causes a lethal systemic infection (salmonellosis) in swine. Live attenuated Salmonella Choleraesuis vaccines are effective in preventing the disease, and isolates of Salmonella Choleraesuis with mutations in the cAMP-receptor protein (CRP) gene (Salmonella Choleraesuis Deltacrp) are the most widely used, although the basis of the attenuation remains unclear. The objective of this study was to determine if the attenuated phenotype of Salmonella Choleraesuis Deltacrp was due to alterations in susceptibility to gastrointestinal factors such as pH and bile salts, ability to colonize or invade the intestine, or cytotoxicity for macrophages. Compared with the parental strain, the survival rate of Salmonella Choleraesuis Deltacrp at low pH or in the presence of bile salts was higher, while the ability of the mutant to invade intestinal epithelia was significantly decreased. In examining the role of CRP on the secretory function of the Salmonella pathogenicity island 1 (SPI-1) encoded type III secretion system (T3SS), it was shown that Salmonella Choleraesuis Deltacrp was unable to secrete the SPI-1 T3SS effector proteins, SopB and SipB, which play a role in Salmonella intestinal invasiveness and macrophage cytotoxicity, respectively. In addition, caspase-1 dependent cytotoxicity for macrophages was significantly reduced in Salmonella Choleraesuis Deltacrp. Collectively, this study demonstrates that the CRP affects the secretory function of SPI-1 T3SS and the resulting ability to invade the host intestinal epithelium, which is a critical element in the pathogenesis of Salmonella Choleraesuis.

Efficacy of a novel Pasteurella multocida vaccine against progressive atrophic rhinitis of swine.

The efficacy of a novel vaccine composed of three short recombinant subunit Pasteurella multocida toxin (PMT) proteins in combination with a bi-valent P. multocida whole-cell bacterin (rsPMT-PM) was evaluated in field studies for prevention and control of progressive atrophic rhinitis (PAR) of swine at 15 conventional farrow-to-finish farms. Experimental piglets that were immunized twice with the rsPMT-PM vaccine developed detectable titers of neutralizing antibodies (greater than 1:8) that prevented the growth retardation and pathological lesions typically observed following challenge with authentic PMT. A total of 542 sows were vaccinated once or twice prior to parturition and serum neutralizing antibody titers were evaluated. Both single and double vaccination protocols induced neutralizing antibody titers of 1:16 or higher in 62% and 74% of sows, respectively. Notably, neither sows nor piglets at a farm experiencing a severe outbreak of PAR at the time of the vaccination trial had detectable antibody titers, but antibody titers increased significantly to 1:16 or higher in 40% of sows following double vaccination. During the year after vaccination, clinical signs of PAR decreased in fattening pigs and growth performance improved sufficiently to reduce the rearing period until marketing by 2 weeks. Collectively, these results indicate that the rsPMT-PM vaccine could be used to provide protective immunity for controlling the prevalence and severity of PAR among farm-raised swine.

Immunogenicity and efficacy of three recombinant subunit Pasteurella multocida toxin vaccines against progressive atrophic rhinitis in pigs.

Three short fragments of recombinant subunit Pasteurella multocida toxin (rsPMT) were constructed for evaluation as candidate vaccines against progressive atrophic rhinitis (PAR) of swine. PMT-specific antibody secreting cells and evidence of cellular immunity were detected in rsPMT-immunized pigs following authentic PMT challenge or homologous antigen booster. Piglets immunized with rsPMT fragments containing either the N-terminal or the C-terminal portions of PMT developed high titers of neutralizing antibodies. Pregnant sows immunized with rsPMT had higher levels of maternal antibodies in their colostrum than did those immunized with a conventional PAR-toxoid vaccine. Offspring from rsPMT vaccinated sows had better survival after challenge with a five-fold lethal dose of authentic PMT and had better growth performance after challenge with a sublethal dose of toxin. Our findings indicate these non-toxic rsPMT proteins are attractive candidates for development of a subunit vaccine against PAR in pigs.