In vitro Cytokine Responses to Virulent PRRS Virus Strains.

Porcine reproductive and respiratory syndrome (PRRS) affects farmed swine causing heavy direct and indirect losses. The infections sustained by PRRS viruses (PRRSV-1 and PRRSV-2) may give rise to severe clinical cases. This highlights the issue of PRRSV pathogenicity and relevant markers thereof. Since PRRSV strains can be discriminated in terms of immunotypes, we aimed to detect possible correlates of virulence in vitro based on the profile of innate immune responses induced by strains of diverse virulence. To this purpose, 10 field PRRSV isolates were investigated in assays of innate immune response to detect possible features associated with virulence. Tumor necrosis factor-α, interleukin (IL)-1beta, IL-8, IL-10, and caspase-1 were measured in cultures of PRRSV-treated peripheral blood mononuclear cells of PRRS-naive pigs, unable to support PRRSV replication. Two reference PRRSV strains (highly pathogenic and attenuated, respectively), were included in the screening. The PRRSV strains isolated from field cases were shown to vary widely in terms of inflammatory cytokine responses in vitro, which were substantially lacking with some strains including the reference, highly pathogenic one. In particular, neither the field PRRSV isolates nor the reference highly pathogenic strain gave rise to an IL-1beta response, which was consistently induced by the attenuated strain, only. This pattern of response was reversed in an inflammatory environment, in which the attenuated strain reduced the ongoing IL-1beta response. Results indicate that some pathogenic PRRSV strains can prevent a primary inflammatory response of PBMCs, associated with reduced permissiveness of mature macrophages for PRRSV replication in later phases.

Characterization of the blastogenic response to LPS of bovine peripheral blood mononuclear cells.

Mitogens are diverse compounds of plant and microbial origin, widely employed to test immunocompetence in animals. The blastogenic response of bovine Peripheral Blood Mononuclear Cells (PBMC) to lypopolysaccharides (LPS) has been investigated in our laboratories for a long time. In particular, a possible correlation between blastogenic response to LPS and disease resistance of periparturient dairy cows had been observed in previous studies. Most important, low responder cows presented a higher frequency of disease cases after calving, compared with high responder animals. Owing to the above, different aspects of the blastogenic response to LPS were investigated on PBMC of healthy Friesian cows, using a 72-hour Bromodeoxyuridin (BrDU) cell proliferation assay. Stimulation with LPS induced little if any replication of bovine PBMC over 72 hours despite consistent BrDU detection in all the PBMC samples under study. Poor replication of LPS-stimulated PBMC was confirmed by cell cycle and cell growth flow cytometry analyses. In particular, LPS stimulation gave rise to very low percentages of S phase cells, sometimes lower than in control, unstimulated cells, as opposed to Concanavalin A-stimulated PBMC. Magnetic separation and analysis of BrDU-treated bovine PBMC after exposure to LPS showed that both B and CD4 T cells are involved in the blastogenic response to LPS, in contrast with current data based on human and murine models. Finally, LPS caused an early, specific up-regulation of TNF-α and TLR4 genes in bovine PBMC, and significant correlations were shown between the expression of inflammatory cytokine and Indoleamine-pyrrole 2,3-dioxygenase (IDO1) genes. On the whole, our data indicate that differences in the blastogenic response to LPS could be partly accounted for by heterogenicity of responding cells (B and T lymphocytes), which might also have an impact on induction and regulation of inflammatory responses and endotoxin tolerance.

Attenuated Salmonella enterica serovar Typhimurium lacking the ZnuABC transporter: an efficacious orally-administered mucosal vaccine against salmonellosis in pigs.

We have recently demonstrated that an attenuated strain of Salmonella enterica serovar Typhimurium unable to synthesize the zinc transporter ZnuABC (S. Typhimurium ΔznuABC), is able to protect mice against systemic and enteric salmonellosis and is safe in pigs. Here, we have tested the protective effects of S. Typhimurium ΔznuABC in pigs. Resistance to challenge with the fully virulent strain S. Typhimurium ATCC 14028 was assessed in animals vaccinated with S. Typhimurium ΔznuABC (two dosages tested), in controls vaccinated with a formalin-inactivated virulent strain and in unvaccinated controls. Clinical signs of salmonellosis, faecal shedding and bacterial colonization of organs were used to assess vaccine-induced protection. After the challenge, pigs vaccinated with the attenuated S. Typhimurium ΔznuABC strain did not display clinical signs of salmonellosis (fever or diarrhoea). The vaccine also reduced intestinal tract colonization and faecal shedding of the fully virulent Salmonella strain, as compared to control groups. S. Typhimurium ΔznuABC represents a promising candidate vaccine against salmonellosis in pigs.