Immunity of pregnant ewes to infection with Salmonella Brandenburg following vaccination

Abstract

Salmonella Brandenburg is responsible for ovine abortion and death in New Zealand. The aims of the present investigation were, firstly, to develop an experimental model for S. Brandenburg infection in pregnant ewes. Secondly, to investigate the efficacy of three different types of vaccines - live attenuated S. Typhimurium mutant (Δcya Δcrp), subunit of S. Brandenburg and multivalent killed whole cell bacterin - on their ability to generate immune responses and provide protection against experimental infections in pregnant ewes and newborn lambs. Thirdly, to investigate the efficacy of a combined vaccination - a live attenuated S. Typhimurium mutant followed by a subunit of S. Brandenburg - on its ability to induce immunity against S. Brandenburg infection in pregnant ewes. An infection model was developed that mimicked natural field infections under a high challenge. When a range (2 x 10⁹ - 2 x 10¹⁰ cfu) of infective organisms (S. Brandenburg) were given by the oral and intraocular route to pregnant ewes followed by a period of starvation (36 - 40 h), a clinical syndrome resulted.

In the first trial, two-year-old ewes were immunised with either a live attenuated vaccine (eyedrop), a subunit vaccine or an inactivated vaccine (both administered subcutaneously), or served as unvaccinated controls. Ewes were boosted with an identical vaccine 4 weeks later. In the second trial, the ewes were primed with a live attenuated vaccine followed by the subunit vaccine 4 weeks later. All animals in both trials were challenged with virulent S.Brandenburg 4-6 weeks after the booster. In the first trial, new born lambs at age of 3 weeks were immunized with the live vaccine and grazed with their dams on pasture contaminated with S. Brandenburg. The presence and number of Salmonella spp in samples taken throughout the trials were determined using an enrichment selection procedure and confirmed by serology and pulsed field-gel electrophoresis (PFGE). Humoral immunity was determined by measurement of plasma total IgG, IgGl, IgM and salivary IgA antibodies reactive to either cell wall fractions (CWF), whole cell (both from S. Brandenburg) or lipopolysaccharide (LPS, from S. Typhimurium), using an indirect ELISA. The cell-mediated immunity was measured in vitro by either lymphocyte proliferation or IFNγ release in purified peripheral blood mononuc1ear cells and cultured whole blood, respectively.

There was no significant protection against mortality and abortion following vaccination with the live attenuated, subunit and inactivated vaccines. There was a significant but transient decrease in the number of ewes shedding S. Brandenburg and also in the quantity of these bacteria in the vaccine groups (p=0.03), as compared to the controls at 42 days after challenge. Lambs from the challenged ewes did not shed Salmonella spp after being vaccinated with the live attenuated vaccine, in contrast to some of the controls, when grazed on pasture contaminated with S. Brandenburg.

The subunit and inactivated vaccines induced high humoral immune responses after immunization. The live vaccine poorly induced antibody responses, but did stimulate a cellular immune response as determined by IFNγ production in vitro. The lymphocyte proliferative response to all vaccines was weak following immunization but strong after challenge. Protection from lethal infection was associated with IFNγ production in vitro following challenge. All lambs born from surviving ewes had high maternal antibody titers, irrespective of the maternal vaccination programme and use of the live vaccine did not induce a humoral immune response. Using a combined live - subunit vaccination protocol, ewes produced significantly higher antibody production (total IgG, IgG1 and IgM) than the control animals (p<0.01). Levels of IFNγ production was significantly enhanced following use of the live attenuated vaccine (p<0.01), but not the subunit vaccine.

In conclusion, the use of any of the three types of vaccine did not significantly protect sheep against lethal experimental challenge with S. Brandenburg despite high levels of antibody and cell mediated immunity following vaccination, but provided transient protection against bacterial shedding in faeces (p<0.05). Similarly the combined vaccination strategy did not protect animals from experimental challenge. Protection from lethal infection was associated with IFNγ production in vitro.