Cloning, Recombinant Production and Functional Analysis of Staphylococcal Phage Endolysins
Abstract
Staphylococci are pathogens that cause serious infections in humans and animals. Nosocomial infections caused by staphylococci, particularly methicillin-resistant Staphylococcus aureus (MRSA) strains, are mostly transmitted through healthcare workers, patients, or contaminated materials and food. In recent years, studies have been carried out to develop alternative antimicrobial strategies due to the inadequacy of existing antibiotics in the prevention of systemic, skin and implant-related biofilm infections caused by these multi-antibiotic resistant strains. One of these new approaches is the development of products containing the bacteriophage endolysin, which is particularly effective against multi-antibiotic-resistant bacteria. In this study, endolysin genes of bacteriophages (prophages) integrated into the chromosomes of Staphylococcus strains were amplified by polymerase chain reaction (PCR) and cloned into pET SUMO and pET-30b(+) vectors and produced recombinantly in E. coli. Anti-staphylococcal and antibiofilm activity of recombinant endolysins against S. aureus, S. epidermidis, and S. haemolyticus strains isolated from clinical specimens, were demonstrated using turbidity reduction, biofilm removal in microwell plates by crystal violet method, and capacity of endolysins to kill biofilm-forming bacteria by confocal microscopy imaging by live-dead staining. The combination of endolysin was shown to reduce bacterial culture turbidity by at least 50% at 60 minutes and biofilms by approximately 70% at 12 hours. These results show that endolysins have the potential to be used in the prevention of staphylococcal infections.
Keywords
Bacteriophage, Anti-biofilm, Endolysin, Confocal Microscopy, Staphylococcus
Supporting Institution
Karadeniz Teknik Üniversitesi
Project Number
TSA-2019-8344
Thanks
This work was supported by Office of Scientific Research Projects of Karadeniz Technical
University.
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