Protective ability of subcellular extracts from Salmonella Enteritidis and from a rough isogenic mutant against salmonellosis in mice

We evaluated the efficacy of surface components enriched hot saline extracts (HE) from parental and two isogenic rough mutant strains of Salmonella Enteritidis as subcellular vaccine candidates. By a randomized mutagenesis approach from a clinical isolate of S. Enteritidis there were selected two rough mutants defective in LPS synthesis (R1 and R2 mutants). The mutations mapped to the wcaI gene and gmd gene, respectively, of the O-antigen gene cluster involved in O-antigen synthesis. BALB/c mice received intraperitoneally one single dose of 30 μg of HE from parental and mutant strains, and the protection against a lethal infection with S. Enteritidis was determined. In contrast to the wild type extract, immunization with rough extracts did not induce any distress symptoms in the mice. HE extract from wild type and R1 strains induced the highest immunogenic response with respect IFN-γ eliciting splenic cells, in contrast with HE-R2. These results correlated with the obtained levels of protection. Thus, at day 63 post-infection, HE from parental strain rendered an 80% level of protection; HE-R1 conferred a 60% level of protection, whereas HE-R2 did not protect the mice. Any of the antigenic extracts elicited systemic IgG1 and IgG2a responses, although these antibodies did not, however, correlate with protection. These results put forward the importance of cellular immune response mediated by IFN-γ in protection against salmonellosis. The significantly different protective capacity between HE extracts from both rough mutants suggest that other factors independent of the O-chain, like outer membrane proteins and fimbrial antigens, may be involved in protection. In summary, the HE is a good candidate acellular extract for evaluation of its protective ability against salmonellosis following vaccination in poultry.