Recombinant Probiotic Expressing Listeria Adhesion Protein Attenuates Listeria monocytogenes Virulence In Vitro

Abstract

Background Listeria monocytogenes, an intracellular foodborne pathogen, infects immunocompromised hosts. The primary route of transmission is through contaminated food. In the gastrointestinal tract, it traverses the epithelial barrier through intracellular or paracellular routes. Strategies to prevent L. monocytogenes entry can potentially minimize infection in high-risk populations. Listeria adhesion protein (LAP) aids L. monocytogenes in crossing epithelial barriers via the paracellular route. The use of recombinant probiotic bacteria expressing LAP would aid targeted clearance of Listeria from the gut and protect high-risk populations from infection.Methodology/Principal FindingsThe objective was to investigate the ability of probiotic bacteria or LAP-expressing recombinant probiotic Lactobacillus paracasei (LbpLAP) to prevent L. monocytogenes adhesion, invasion, and transwell-based transepithelial translocation in a Caco-2 cell culture model. Several wild type probiotic bacteria showed strong adhesion to Caco-2 cells but none effectively prevented L. monocytogenes infection. Pre-exposure to LbpLAP for 1, 4, 15, or 24 h significantly (P<0.05) reduced adhesion, invasion, and transepithelial translocation of L. monocytogenes in Caco-2 cells, whereas pre-exposure to parental Lb. paracasei had no significant effect. Similarly, LbpLAP pre-exposure reduced L. monocytogenes translocation by as much as 46% after 24 h. LbpLAP also prevented L. monocytogenes-mediated cell damage and compromise of tight junction integrity. Furthermore, LbpLAP cells reduced L. monocytogenes-mediated cell cytotoxicity by 99.8% after 1 h and 79% after 24 h.Conclusions/SignificanceWild type probiotic bacteria were unable to prevent L. monocytogenes infection in vitro. In contrast, LbpLAP blocked adhesion, invasion, and translocation of L. monocytogenes by interacting with host cell receptor Hsp60, thereby protecting cells from infection. These data show promise for the use of recombinant probiotics in preventing L. monocytogenes infection in high-risk populations.

Synthetic biology platform technologies for antimicrobial applications

“…Such probiotics convey prophylactic effects by outcompeting pathogens for ecological niches and by employing conventional strategies for toxin inhibition [92,93].…”

Engineering commensal bacteria for prophylaxis against infection

“…Paton et al have engineered E. coli strains producing chimeric surface lipopolysaccharides that are capable of neutralizing, on contact, shiga-toxin secreted by pathogenic E. coli or cholera toxin secreted by V. cholerae in vivo [22,23]. Probiotic lactic acid bacteria (LAB) have been engineered to secrete either pathogenic adhesion proteins or flagellin in order to competitively inhibit adhesion of pathogens such as Listeria monocytogenes or Salmonella enterica [24,25]. These studies aimed to engineer commensal bacteria to outcompete for valuable surface space in the host s intestine, forcing pathogens to pass through the gastrointestinal tract without detrimental effects.…”

Bioengineered Probiotics – A Solution to Broaden Probiotics Efficacy!

“…Similar strategy was also used to control Helicobacter pylori infection by expressing urease, a colonization factor in host gastric cells, in probiotic bacteria, creating a competitive environment to exclude H. pylori colonization. Similarly, expression of Listeria adhesion protein on probiotics was able to reduce Listeria monocytogenes infection in vitro [7]. Salmonella infection was also controlled by expressing flagellar proteins which resulted in a protective immune response against salmonellosis.…”