Inhibition of the assembly of amyloid curli fibers, the main structural components of E. coli biofilms, by a B. subtilis secondary metabolite

SERRA, D.O.; CORDISCO, E.

Los Cocos - Córdoba

Congreso; XVII Congreso Argentino de Microbiología General; 2022

Sociedad Argentina de MIcrobiología General

Biofilms are surface-associated multicellular communities that bacteria build by embedding themselves in an extracellular matrix (ECM) composed of polymeric fibers. Due to their high antibiotic tolerance, bacterial biofilms are involved in more than 50% of all chronic infections. An example of that are the urinary tract infections (UTI) caused by Escherichia coli, which frequently associate with the formation of biofilms on catheters and the bladder. Recognizing the need for solutions to combat biofilm-based infections in general, and of E. coli in particular, we focused on the search for compounds that can interfere with the production of curli, which are amyloid protein fibers that constitute the major structural component of E. coli biofilms. To do so, we explored interactions of E. coli with distinct microorganisms in agar-grown macrocolonies biofilms as a platform for the search of curli inhibitors. We found that B. subtilis NCIB 3610 is able to inhibit the production of curli amyloid fibers in macrocolonies of E. coli strains that produce them as the main ECM element. Curli inhibition was detected by the loss of staining with amyloid-specific dyes and by the absence of curli-dependent morphology of E. coli macrocolonies when they grew in close proximity to B. subtilis NCIB 3610 or in the presence of extracts derived from cell-free culture supernatants of this strain. This inhibitory action on curli was found to be mediated by a PKS metabolite whose synthesis in B. subtilis requires activation by the 4'-phosphopantetheinyl transferase (PPTase) associated with secondary metabolism. Analyses of expression of the csgBAC operon -which encodes the curli structural subunits CsgB and CsgA- independently of its natural promoter in the presence of the metabolite showed that the inhibitory effect occurs at post-transcriptional level. Further experimental evidences such as the detection of unpolymerized CsgA subunits in the agar underlying the macrocolony biofilms treated with the PKS metabolite and the inhibition of in vitro polymerization of CsgA-His-tagged in the presence of the purified metabolite demonstrated that the B. subtilis compound acts impeding the assembly of curli subunits into amyloid fibers. In sum, this work reveals a novel microbial compound that targets the major structural component of E. coli biofilms and provides molecular insights into its mode of action.