α-Chymotrypsin Immobilized on a Low-Density Polyethylene Surface Successfully Weakens Escherichia coli Biofilm Formation

Abstract

The protease α-chymotrypsin (α-CT) was covalently immobilized on a low-density polyethylene (LDPE) surface, providing a new non-leaching material (LDPE-α-CT) able to preserve surfaces from biofilm growth over a long working timescale. The immobilized enzyme showed a transesterification activity of 1.24 nmol/h, confirming that the immobilization protocol did not negatively affect α-CT activity. Plate count viability assays, as well as confocal laser scanner microscopy (CLSM) analysis, showed that LDPE-α-CT significantly impacts Escherichia coli biofilm formation by (i) reducing the number of adhered cells (−70.7 ± 5.0%); (ii) significantly affecting biofilm thickness (−81.8 ± 16.7%), roughness (−13.8 ± 2.8%), substratum coverage (−63.1 ± 1.8%), and surface to bio-volume ratio (+7.1 ± 0.2-fold); and (iii) decreasing the matrix polysaccharide bio-volume (80.2 ± 23.2%). Additionally, CLSM images showed a destabilized biofilm with many cells dispersing from it. Notably, biofilm stained for live and dead cells confirmed that the reduction in the biomass was achieved by a mechanism that did not affect bacterial viability, reducing the chances for the evolution of resistant strains.

Urinary Catheter Coating Modifications: The Race against Catheter-Associated Infections

“…There are several anti-fouling catheters including those coated with hydrogels (most popular) [27,28], poly(tetrafluoroethylene) [34][35][36][37][38], polyzwitterions [39][40][41], and poly(ethylene glycol) [42][43][44][45][46]. Furthermore, surface topography modification [47][48][49][50] and enzymes-immobilized coatings [51][52][53][54][55][56][57][58] have been explored for prevention of microbial colonization and biofilm formation. These polymers and modifications are discussed in detail in this section.…”

“…α-CT is a serine endopeptidase that cleaves peptide bonds by attacking the unreactive carbonyl group [52]. Based on the properties of α-CT, Catto and collaborators assessed whether α-CT will disrupt biofilm formation [55] since biofilm matrix consists of proteins, extracellular DNA, and polysaccharides [67]. In this in vitro study, α-CT was covalently immobilized on a low-density polyethylene (LDPE) pieces and incubated with LB media containing E. coli MG1655 in a CDC biofilm reactor with continuous stirring.…”

“…Additionally, the bio-volume of the polysaccharide matrix decreased. The authors conclude that the anti-biofilm properties of LDPE-α-CT may potentiate the activity of antimicrobials since the majority of the population would be in a planktonic form [55].…”

Testing Anti-Biofilm Polymeric Surfaces: Where to Start?

“…Thus, selection pressure decreases, limiting resistant-drug development, and potentially reinstating the efficacy of traditional antimicrobials [69]. Several natural and synthetic compounds, as well as matrix-targeting enzymes based on the previous biocide-free anti-biofilm mechanisms of action, have been coated or immobilized on polymeric surfaces, providing promising, eco-friendly, bio-inspired, anti-biofilm materials able to replace, or integrate with, presently dominating biocide-based approaches [69,70,71,72,73,74].…”

“…Sajeevan et al [77] impregnated silicon catheter tubes with anacardic acids that efficiently inhibited Staphylococcus aureus colonization and biofilm formation on its surface both in vitro and in vivo. Spadoni-Andreani et al [73] demonstrated that polypropylene surfaces coated with proteases weakened adhesion and increased the dispersion of Candida albicans biofilm cells and Cattò et al [74] proved that the proteases α-chymotrypsin prevented E. coli biofilm formation on polyethylene materials…”