Amrita Ray Mohapatra1, Kadirvelu Jeevaratnam2. 1. Department of Biochemistry and Molecular Biology, Pondicherry University, Puducherry, 605014, India. 2. Department of Biochemistry and Molecular Biology, Pondicherry University, Puducherry, 605014, India. Electronic address: jeevskj@gmail.com.
Abstract
BACKGROUND: Catheter-associated urinary tract infections are one of the most common types of hospital-acquired infections that start with bacterial adhesion and lead to biofilm formation. The antagonistic activity of lactic acid bacteria against pathogenic organisms makes them important for medical applications. OBJECTIVE: This study evaluated the precise method for purification of bacteriocin from Lactobacillus plantarum subsp. argentoratensis SJ33, and its characterization and effectiveness for biofilm inhibition on urinary catheters coated with bacteriocin. METHODS: Purification of bacteriocin was carried out using various methods such as cell adsorption-desorption, gel permeation chromatography, and hydrophobic interaction chromatography. Bacteriocin preparation was analysed using reverse-phase high-performance liquid chromatography (HPLC) and further characterised by Tricine SDS-PAGE and Q-TOF ESI MS. Antibacterial activity of bacteriocin was assessed against 16 different Gram-positive and Gram-negative bacterial strains, and their effect on morphology was observed under scanning electron microscopy (SEM). Biofilm adherence and inhibition were evaluated by crystal violet assay, fluorescence microscopy and SEM. RESULTS AND CONCLUSIONS: Bacteriocin preparation exhibited broad-spectrum activity against both Gram-positive and Gram-negative bacteria, and SEM analysis revealed membrane pore formation. On treating with various enzymes, bacteriocin was found to be sensitive to proteases, which confirmed its proteinaceous nature. Bacteriocin showed its applicability at acidic pH in the urinary tract. Antibiofilm activity of bacteriocin established its significance in catheter-associated biofilm inhibition against Pseudomonas aeruginosa and Staphylococcus aureus. Molecular weight of bacteriocins, namely Bac F1 and Bac F2 as resolved by RP-HPLC, was estimated to be 4039 Da and 1609 Da, respectively.
BACKGROUND: Catheter-associated urinary tract infections are one of the most common types of hospital-acquired infections that start with bacterial adhesion and lead to biofilm formation. The antagonistic activity of lactic acid bacteria against pathogenic organisms makes them important for medical applications. OBJECTIVE: This study evaluated the precise method for purification of bacteriocin from Lactobacillus plantarum subsp. argentoratensis SJ33, and its characterization and effectiveness for biofilm inhibition on urinary catheters coated with bacteriocin. METHODS: Purification of bacteriocin was carried out using various methods such as cell adsorption-desorption, gel permeation chromatography, and hydrophobic interaction chromatography. Bacteriocin preparation was analysed using reverse-phase high-performance liquid chromatography (HPLC) and further characterised by Tricine SDS-PAGE and Q-TOF ESI MS. Antibacterial activity of bacteriocin was assessed against 16 different Gram-positive and Gram-negative bacterial strains, and their effect on morphology was observed under scanning electron microscopy (SEM). Biofilm adherence and inhibition were evaluated by crystal violet assay, fluorescence microscopy and SEM. RESULTS AND CONCLUSIONS: Bacteriocin preparation exhibited broad-spectrum activity against both Gram-positive and Gram-negative bacteria, and SEM analysis revealed membrane pore formation. On treating with various enzymes, bacteriocin was found to be sensitive to proteases, which confirmed its proteinaceous nature. Bacteriocin showed its applicability at acidic pH in the urinary tract. Antibiofilm activity of bacteriocin established its significance in catheter-associated biofilm inhibition against Pseudomonas aeruginosa and Staphylococcus aureus. Molecular weight of bacteriocins, namely Bac F1 and Bac F2 as resolved by RP-HPLC, was estimated to be 4039 Da and 1609 Da, respectively.