Abstract
Abstract Background Atrocious use of antibiotics has led to the emergence of MDR in uropathogenic Escherichia coli (UPEC) that poses a serious challenge in the management of urinary tract infections (UTIs). The WHO has described antibiotic resistance in uropathogens as a key pressure point in the burgeoning global antimicrobial resistance crisis. Given this grim situation, we are exploring phage-encoded lysins as plausible alternatives. Objectives Our study involved an in silico strategy for the discovery and characterization of lysin sequences (seq) targeting E. coli cell wall and evaluating the bactericidal activity of these recombinant lysins using in-vitro assays. Methods Novel lysin sequences were searched by BLAST homology and by screening E. coli prophages in the database (using PHASTER). Lysozyme-like domain was observed in 9 out of 16 lysins. Their characterization depicted modular or globular structure. Based on the physicochemical properties, 7 out of 16 lysins were selected for cloning, expression, and purification as recombinant proteins for evaluating the bactericidal activity. Results Among the several lysins screened, lysin seq 5 demonstrated the highest activity using in vitro assays. Using static biofilm assay, lysin seq 5 (180 μg) showed efficient reduction (>50%) in the biofilm formed by ATCC UPEC 700928 strain. As per turbidity reduction method, lysin seq 5 (50 μg) showed 37% drop in OD600nm on UPEC 700928 strain after 3 h of incubation at 37°C. Using spot on lawn assay, lysin seq 5 (20 μg) exhibited lytic activity (zone of inhibition) on five drug-resistant clinical UTI isolates, which were pretreated with an outer membrane permeabilizer (OMP), viz., EDTA (0.3 mM). Conclusions Lysin seq 5 exhibited antibiofilm activity against UPEC 700928 strain as well as lytic activity against drug-resistant clinical UTI isolates. Screening of additional drug-resistant clinical isolates from UTI patients is underway.
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