Prime-boost vaccination with attenuated Salmonella Typhimurium ΔznuABC and inactivated Salmonella Choleraesuis is protective against Salmonella Choleraesuis challenge infection in piglets

Abstract

Background: Salmonella enterica serovar Choleraesuis (S. Choleraesuis) infection causes a systemic disease in pigs. Vaccination could represent a solution to reduce prevalence in farms. In this study, we aimed to assess the efficacy of an attenuated strain of Salmonella enterica serovar Typhimurium (S. Typhimurium ΔznuABC) against S. Choleraesuis infection. The vaccination protocol combined priming with attenuated S. Typhimurium ΔznuABC vaccine and boost with an inactivated S. Choleraesuis vaccine and we compared the protection conferred to that induced by an inactivated S. Choleraesuis vaccine.

Methods: The first group of piglets was orally vaccinated with S. Typhimurium ΔznuABC and boosted with inactivated S. Choleraesuis, the second one was intramuscularly vaccinated with S. Choleraesuis inactivated vaccine and the third group of piglets was unvaccinated. All groups of animals were challenged with a virulent S. Choleraesuis strain at day 35 post vaccination.

Results: The results showed that the vaccination protocol, priming with S. Typhimurium ΔznuABC and boosted with inactivated S. Choleraesuis, applied to group A was able to limit weight loss, fever and organs colonization, arising from infection with virulent S. Choleraesuis, more effectively, than the prime-boost vaccination with homologous S. Choleraesuis inactivated vaccine (group B).

Conclusion: In conclusion, these research findings extend the validity of attenuated S. Typhimurium ΔznuABC strain as a useful mucosal vaccine against S. Typhimurium and S. Choleraesuis pig infection. The development of combined vaccination protocols can have a diffuse administration in field conditions because animals are generally infected with different concomitant serovars.

Keywords: Attenuated and inactivated vaccines; IFN-gamma; Mucosal vaccine; Piglets; Salmonella Choleraesuis; Salmonellosis.

Fig. 1

Attenuated S. Typhimurium ΔznuABC is not detectable from day 18 after vaccination. In a, symbols depict the mean concentration of S. Typhimurium ΔznuABC (expressed as LOG₁₀ CFU/g) in group A at different time points (day 1, 5, 13, 18, 33 after vaccination). Bars represent the standard deviation. In b, columns represent the percentage of shedder piglets of group A at different time-points. S. Typhimurium ΔznuABC is not detectable from day 18 after vaccination. Group B and C are not shown because negative. Depicted microbiological results derived from the semi-quantitative approach

Fig. 2

Vaccination with attenuated S. Typhimurium ΔznuABC prevents fever. Body temperature of groups A, B and C is shown at different time points (day 3, 4, 5 and 7 after challenge). Symbols represent mean and bars standard deviation. Symbols (*) represent differences statistically significant among groups with p < 0.01

Fig. 3

Attenuated strain of S. Typhimurium does not retard the weight gain of piglets. Symbols represent the weight gain of eighteen animals, divided into groups A, B and C, while vertical bars represent the standard deviation from the beginning to the end of the study (day 0 and 47). No differences are recorded. The weight of animals vaccinated with the attenuated strain of S. Typhimurium is not different from other groups

Fig. 4

S. Typhimurium ΔznuABC vaccine reduces organ colonization. Amount of S. Choleraesuis in lymph nodes, ileum, cecum and colon of group A-C piglets was determined 12 days after challenge. Depicted microbiological results derived from the semi-quantitative approach. Each symbol represents microbiological results obtained from each animal and each bar represents mean concentration. Differences are statistically significant in cecum between group A and the other groups (p < 0.05) and in lymph nodes between group A and B

Fig. 5

S. Choleraesuis challenge infection induces IFN-ɤ production. Symbols and bars represent piglets and IFN-ɣ mean concentration, respectively. Concentration of IFN-ɤ was normalized in relation to total protein content. Difference was statistically significant between the unvaccinated group (C) and group A vaccinated with the attenuated strain ** (p < 0.05). LLD indicates Lower Limit of Detection (15.6 pg/ml); LS indicates Limit of Sensitivity (minimum detectable dose, 3.9 pg/ml)

Fig. 6

Attenuated S. Typhimurium ΔznuABC vaccine and inactivated S. Choleraesuis vaccine induce antibody production. Symbols and bars represent mean and standard deviation of s/p ratio in 3 groups, respectively. The pattern of humoral immunity is similar between group A (vaccinated with Attenuated S. Typhimurium ΔznuABC) and in group B (vaccinated with killed S. Choleraesuis). The X-axis is divided into two segments to differentiate between antibody response after vaccination (DAV) and challenge (DAC). Differences are statistically significant between vaccinated piglets with attenuated strain and unvaccinated piglets from day 18 after vaccination. On the contrary, differences are statistically different between vaccinated piglets with inactivated strain and unvaccinated piglets only at day 35 and 47 after vaccination. Gray star indicates significant difference between groups B and C, while black star indicates significant difference between group A and C