Cloning, Characterization, and Antibacterial Properties of Endolysin LysE Against Planktonic Cells and Biofilms of Aeromonas hydrophila.

Multidrug-resistant bacteria are emerging as a major global threat to public health. Bacteriophages are an important source of antimicrobial enzymes and could be developed as an alternative antibiotic candidate. This study investigates the antibacterial capacity of the endolysin LysE against Aeromonas hydrophila. The endolysin LysE gene was cloned and expressed in Escherichia coli BL21 (DE3) cells. Purified recombinant LysE protein was tested for its antimicrobial activity against A. hydrophila. The study reveals that recombinant LysE protein was highly effective against Gram-negative bacteria when combined with antimicrobials that alter the permeability of the outer membrane. Specifically, the enzyme had the highest muralytic activity at pH 4, and maintained over 50% of the activity at pH 10. Moreover, endolysin displayed more than 50% activity even after 30 min of incubation at 100 °C. Also, endolysin LysE resulted in one log reduction in CFU/mL in 30 min and demonstrated antibiofilm capabilities when combined with EDTA. Interestingly, checkerboard assay showed its synergistic effects in combination with lower concentrations of colistin against A. hydrophila. Additionally, in vitro tests with Channa striatus kidney (CSK) cell lines do not show cytotoxic effects. Taken together, these findings suggest that LysE can be employed with outer membrane permeabilizers to expand the arsenal repertoire against Gram-negative bacteria in the aquaculture, food, and medical industries.