Enterohaemorrhagic Escherichia coli produces outer membrane vesicles as an active defence system against antimicrobial peptide LL-37

Abstract

Antimicrobial peptides (AMPs) are important components of the innate immune system. Enterohaemorrhagic Escherichia coli (EHEC), a food-borne pathogen causing serious diarrheal diseases, must overcome attack by AMPs. Here, we show that resistance of EHEC against human cathelicidin LL-37, a primary AMP, was enhanced by butyrate, which has been shown to act as a stimulant for the expression of virulence genes. The increase of resistance depended on the activation of the ompT gene, which encodes the outer membrane protease OmpT for LL-37. The expression of the ompT gene was enhanced through the activation system for virulence genes. The increase in ompT expression did not result in an increase in OmpT protease in bacteria but in enhancement of the production of OmpT-loaded outer membrane vesicles (OMVs), which primarily contributed to the increase in LL-37-resistance. Furthermore, a sublethal dosage of LL-37 stimulated the production of OMVs. Finally, we showed that OMVs produced by OmpT-positive strains protect the OmpT-negative strain, which is susceptible to LL-37 by itself more efficiently than OMVs from the ompT mutant. These results indicate that EHEC enhances the secretion of OmpT-loaded OMVs in coordination with the activation of virulence genes during infection and blocks bacterial cell attack by LL-37.

Immunization of mice with chimeric antigens displaying selected epitopes confers protection against intestinal colonization and renal damage caused by Shiga toxin-producing Escherichia coli

“…Interestingly, the antigens selected for our vaccine design are different from those used in most trials [38][39][40] ; however, they are widely distributed in STEC and involved in several of its pathogenicity mechanisms 10,13 . While Cah and Hes are related to the bacterial−host interaction 9,41 , OmpT participates in the biogenesis of bacterial outer membrane vesicles (OMVs) 42 and the degradation of antimicrobial peptides like LL-37 43,44 . To our knowledge, this is the first formulation based on recombinant chimeric proteins that includes a virulence factor exclusive of LEE-negative STEC strains in a vaccine design.…”

Bacterial outer membrane vesicles provide an alternative pathway for trafficking of type III secreted effectors into epithelial cells

“…EHEC OMVs are sufficient to cause hemolytic uremic syndrome-like symptoms (39) and are a key activator of pro-inflammatory responses to bacterial infections in vivo (31). OMVs additionally play important roles during infection, including as a decoy against host-secreted antimicrobial peptides (24). OMVs are constantly produced, but the transition to the intra-host milieu and exposure to cues such as colonic medium and mucin significantly increases vesiculation.…”

“…OMVs are proteo-liposomes of 10-200 nm diameter formed by budding of the outer membrane, and can contain membrane-associated and soluble proteins, nucleic acids and small molecules. OMV production is a constitutive process, but is also a protective response and increases in the presence of environmental stresses and during infection (24-27). Often, this increase in OMV production is co-regulated with other virulence mechanisms (24, 28).…”

Secretion and Delivery of Intestinal Pathogenic Escherichia coli Virulence Factors via Outer Membrane Vesicles

“…In the presence of antimicrobial peptides or bacteriophages, increased production of membrane vesicles correlates with improved fitness and increased survival (Manning and Kuehn, 2011;Duperthuy et al, 2013). For example, EHEC enhances the secretion of outer membrane protease OmpTloaded OMVs during infection and thereby blocks bacterial cell attack by human antibacterial peptide cathelicidin LL-37 (Urashima et al, 2017).…”

“…OMVs can take on both defensive and offensive tasks during infection (MacDonald and Kuehn, 2012). Defensively, they can be used to sequester antibiotics, bacteriophages, and antibodies, bind or degrade antimicrobial peptides, as well as bait antigens to distract the immune system (Manning and Kuehn, 2011;MacDonald and Kuehn, 2012;Duperthuy et al, 2013;O'Donoghue and Krachler, 2016;Urashima et al, 2017;Reyes-Robles et al, 2018). The potential of OMVs as offensive weapons is evident in their ability to deliver virulence factors into host cells ( Figure 1BIII) Tan et al, 2007;Bomberger et al, 2009;Amano et al, 2010;Ellis and Kuehn, 2010;Kulp and Kuehn, 2010;Schaar et al, 2011;Rompikuntal et al, 2012;Bielaszewska et al, 2013Bielaszewska et al, , 2017Kunsmann et al, 2015;O'Donoghue and Krachler, 2016;Rüter et al, 2018) as well as to induce sepsis, sepsis-associated cardiomyopathy or disseminated intravascular coagulation in the absence of intact bacterial cells (Park et al, 2010;Shah et al, 2012;Svennerholm et al, 2017;Wang et al, 2019).…”