Lactococcus lactis is an immunomodulator and candidate live mucosal delivery vehicle for vaccine antigens and for
biologically active molecules, including immunoregulatory cytokines such as interferon-gamma (IFN-g). To provide a tool for investigating downregulation of allergic predisposition of pigs to experimental food allergy, porcine IFN-g was cloned and expressed as a fusion protein with the usp45 secretion signal. Immunoblot analysis with polyclonal anti-pIFN-g-antibody demonstrated that the recombinant porcine IFN-g (rpIFN-g) protein was expressed in the L. lactis transformants as a secreted product. Activity of rpIFN-g was confirmed by ability to upregulate class II major histocompatibility complex (MHC) on cells of the porcine monocytic cell line 3D4/31. The L. lactis producing biologically active rpIFN-g will be used to investigate its possible ability to modulate the allergic immune response phenotype of pigs.  Biologically active rpIFN-g was produced in L. lactis and upregulated MHC class II expression on porcine monocytic cells suggesting that the modified L. lactis might be able to enhance type 1 immune response and reduce susceptibility to allergy in animals/humans. The researchers have engineered and expressed afunctional rpIFN-g gene in L. lactis that would allow delivery to the pig’s intestinal mucosa.  Although use of this construct is limited by lack of control over quantity and conditional expression of recombinant protein production, containment of live genetically modified L. lactis secreting human IL-10 in pigs is possible using an auxotrophic thymidine mutant (Steidler et al., 2003). However, the presently described construct will be used as a heat-killed bacterial vehicle expressing the recombinant porcine IFN-g protein, for delivery both orally and systemically.