DNA sequence analysis of glycoprotein G gene of bovine ephemeral fever virus and development of a double oil emulsion vaccine against bovine ephemeral fever.

The surface glycoprotein G is considered as the major neutralizing and protective antigen of bovine ephemeral fever virus (BEFV). Comparison of the deduced amino acid sequence of G protein of BEFV isolates during the period 1984-2004 outbreaks in Taiwan showed amino acid substitutions in the neutralizing epitopes. All the isolates differ markedly in the neutralizing epitope at the same amino acid positions compared to the currently available killed vaccine strain (Tn73). Tn88128 strain isolated in 1999 showed the maximum variability of 12 amino acids, 5 amino acid in the neutralization epitope and 7 apart from, respectively. Combinations of both Tn88128 (1999) and commercially available vaccine strain (Tn73) were developed and its safety was evaluated in mice, guinea pigs, calves, and pregnant cows. None of the animals showed any adverse effect or clinical signs. Calves were immunized with commercial vaccine (Tn73) and, combined vaccine (Tn73 and Tn88128), respectively, with adjuvants such as Al-gel and water-in-oil-in-water (w/o/w) oil and PBS alone and challenged with Tn88128 strains. Except PBS administered animals, all the vaccinated animals showed protective immune response. However, animals immunized with combined vaccine plus w/o/w adjuvant elicited stronger neutralization antibodies and long lasting immunity compared to other vaccines.

Heterologous protection in pigs induced by a plasmid-cured and crp gene-deleted Salmonella choleraesuis live vaccine.

In this study, we exploited a crp (cAMP receptor protein) gene-deleted, virulence plasmid-cured Salmonella choleraesuis mutant with decreased carbon source utilization, designated S.C.-Deltacrp/vpl(-), as a live vaccine strain. Normal weight gain with no clinical signs was observed in pigs immunized with high doses of S.C.-Deltacrp/vpl(-) live vaccine. Vaccination in pregnant sows induced high maternal antibodies, which could prevent piglets from Salmonella infection. Moreover, serial transmission of the vaccine strain in piglets produced no evidence of reversion to virulence. Furthermore, the peripheral blood mononuclear cells from immunized piglets also developed Salmonella specific T-cell proliferative response in vitro. Our results indicate that immunogenic antigens in S.C.-Deltacrp/vpl(-) can induce adequate immunity to protect pigs against challenge with a heterologous virulent strain. Thus, this mutant holds promise for the development of a new live S. choleraesuis vaccine.

Enhancement of humoral and cellular immune responses by an oral Salmonella choleraesuis vaccine expressing porcine prothymosin alpha.

Previously, we showed that murine prothymosin alpha (ProT) enhances the efficacy of a pseudorabies DNA vaccine delivered by bacterial vectors. In this study, we cloned and sequenced the cDNA for porcine ProT. The deduced amino acid sequence of porcine ProT exhibited high homology to ProT from other mammals. Oral Salmonella choleraesuis vaccine carrying the ProT eukaryotic expression plasmid protected mice against virulent S. choleraesuis challenge. The adjuvant effect of ProT on humoral and cellular immune responses enhanced protective efficacy of the vaccine. Furthermore, both humoral and cellular immune responses played roles in the protective immune responses induced by the vaccine. Collectively, our results show that delivery of the ProT gene carried by attenuated S. choleraesuis augmented the immunogenicity of oral S. choleraesuis vaccine.

A simple selection system for construction of recombinant gD-negative pseudorabies virus as a vaccine vector.

We describe a simple, efficient two-step method for construction of glycoprotein D (gD)-negative pseudorabies virus (PrV) carrying transgenes inserted in place of the gD gene. The first step was the use of the thymidine kinase (TK) gene of herpes simplex virus (HSV) for insertional inactivation of the gD gene in a PrV mutant deficient in both TK and glycoprotein E (gE). The gD-negative, HSV-TK-positive mutant could be selected in HAT medium. The second step was substitution of HSV-TK with other genes of interest. The resultant gD/gE/TK-negative mutant was easily isolated by acyclovir selection. The expression of the transgene was detectable in vivo and the antibody responses against both inserted antigens and PrV were induced. The protective efficacy of the gD/gE/TK-negative PrV against lethal PrV challenge was also demonstrated. This PrV mutant carrying immunogenic proteins from unrelated porcine pathogens may be tested as a multivalent vaccine candidate for swine.