Characterization and Evaluation of a Salmonella enterica Serotype Senftenberg Mutant Created by Deletion of Virulence-Related Genes for Use as a Live Attenuated Vaccine

Abstract

ABSTRACT Natural infections of chickens with Salmonella enterica subsp. enterica serovar Senftenberg ( S. Senftenberg) are characterized by low-level intestinal invasiveness and insignificant production of antibodies. In this study, we investigated the potential effects of lon and cpxR gene deletions on the invasiveness of S . Senftenberg into the intestinal epithelium of chickens and its ability to induce an immune response, conferring protection against S . Senftenberg infection. With the allelic exchange method, we developed JOL1596 (Δ lon ), JOL1571 (Δ cpxR ), and JOL1587 (Δ lon Δ cpxR ) deletion mutants from wild-type S . Senftenberg. Deletion of the lon gene from S . Senftenberg produced increased frequency of elongated cells, with significantly greater amounts of exopolysaccharide (EPS) than in the cpxR -deleted strain and the wild-type strain. The in vivo intestinal loop invasion assay showed a significant increase in epithelial invasiveness for JOL1596 (Δ lon ) and JOL1587 (Δ lon Δ cpxR ), compared to JOL1571 (Δ cpxR ) and the wild-type strain. Furthermore, the S . Senftenberg wild-type and mutant strains were internalized at high levels inside activated abdominal macrophages from chicken. The in vivo inoculation of JOL1587 (Δ lon Δ cpxR ) into chickens led to colonization of the liver, spleen, and cecum for a short time. Chickens inoculated with JOL1587 (Δ lon Δ cpxR ) showed significant increases in humoral, mucosal, and cellular immune responses specific to S . Senftenberg antigens. Postchallenge, compared to the control group, the JOL1587 (Δ lon Δ cpxR )-inoculated chickens showed not only lower persistence but also faster clearance of wild-type S . Senftenberg from the cecum. We conclude that the increased intestinal invasiveness and colonization of internal organs exhibited by JOL1587 (Δ lon Δ cpxR ) led to the establishment of immunogenicity and conferred protective efficacy against S . Senftenberg infections in chickens.