Antigenic and functional profiles of a Lawsonia intracellularis protein that shows a flagellin-like trait and its immuno-stimulatory assessment

Abstract

The obligate intracellular Lawsonia intracellularis (LI), the etiological agent of proliferative enteropathy (PE), is an economically important disease in the swine industry. Due to extreme difficulty of in vitro culture of the pathogen, molecular characterization of protein components of LI that are targets of the immune system, is difficult; thus, the scientific evidence to drive the development of preventive measures is lacking. In this work, we investigated the antigenic and functional characteristics of a putative flagellar-associated protein, LI0570, using in silico computational approaches for epitope prediction and an in vitro protein-based molecular assay. The amino acid sequence of LI0570 exhibited similarities to flagellar-associated proteins in four different bacterial strains. The presence of B cell linear confirmative epitopes of the protein predicted by a bioinformatics tool was validated by western blot analysis using anti-LI mouse hyperimmune serum, which implied that LI0570 induced production of antigen-specific antibodies in vivo. Further, TLR5-stimulating activity and IL-8 cytokine expression produced via downstream signaling were observed in HEK-Blue™-hTLR5 cells stimulated with LI0570. This result indicates that the LI0570 protein can trigger an innate immune response followed by a T-cell-related adaptive immune response in an infected host. Collectively, the data presented here support that the LI0570 protein which shows the antigenic potential could be a useful component of a recombinant vaccine against PE, providing progress toward an effective prevention strategy.

Potent O-antigen-deficient (rough) mutants of Salmonella Typhimurium secreting Lawsonia intracellularis antigens enhance immunogenicity and provide single-immunization protection against proliferative enteropathy and salmonellosis in a murine model

“…In the initial stage of vaccine development, LI antigen proteins (OptA, OptB, LfliC, and Lhly) were chosen based on their antigenic profiles. These proteins are also putative virulence factors (e.g., outer membrane, flagellin, and toxin proteins) of LI [ 7 , 10 , 12 ]. For construction of the Salmonella vector system, the attenuated rough ST delivery strain JOL1800 was generated from a parental lon - cpxR - asd mutant ST strain, JOL912, via rfaL gene deletion.…”

“…Our latest study showed that a putative flagellar-associated protein, LI0570, has flagellin-like traits and stimulates Toll-like receptor (TLR) 5-expressing immune cells. This leads to the production of proinflammatory cytokines and the subsequent activation of T cell-related adaptive immune responses [ 10 ]. Hemolytic and cytolytic activities are known to occur during natural LI infection in pigs [ 11 ].…”

Immunization of chicken with flagellin adjuvanted Salmonella enteritidis bacterial ghosts confers complete protection against chicken salmonellosis

“…The level of Toll-like receptor-5 ( TLR5 ) expression was assessed by labeling with an anti-human TLR5 antibody (Abcam. Cambridge, UK) as previously described ( Won and Lee, 2018 ). Further chicken PBMC was obtained from 3 chickens and seeded at 1 Χ 10 6 cell/ well in a 24-well plate.…”

Salmonella vector induces protective immunity against Lawsonia and Salmonella in murine model using prokaryotic expression system

“…The disease, characterized by weight loss and two clinical forms, poses a significant threat to the porcine industry [ 2 ]. Antibiotics have successfully managed LI, but antibiotic-resistant strains are emerging, requiring attention [ 3 4 5 6 7 ]. Current vaccination strategies include live-attenuated and inactivated [ 8 ] vaccines, but they have deficiencies, including inadequate immunity [ 9 ], safety concerns [ 10 ], and exorbitant production costs.…”