Identification and Characterization of Antimicrobial Peptides Isolated from Entophytic Bacteria and Their Activity against Multidrug-Resistance Gram-Negative Bacteria in South Korea

Authors: Maryam Beiranvand

Abstract:

Multi-drug resistance in various microorganisms has increased globally in many healthcare facilities. Less effective antimicrobial activity of drug therapies for infection control becomes trouble. Since 1980, no new type of antimicrobial drug has been identified, even though combinations of antibiotic drugs have been discovered almost every decade. Between 1981 and 2006, over 70% of novel pharmaceuticals and chemical agents came from natural sources. Microorganisms have yielded almost 22,000 natural compounds. The identification of antimicrobial components from endophytes bacteria could help overcome the threat posed by multi-drug resistant strains. The project aims to analyze and identify antimicrobial peptides isolated from entophytic bacteria and their activity against multidrug-resistant Gram-negative bacteria in South Korea. Endophytic Paenibacillus polymyxa. 4G3 isolated from the plant, Gynura procumbery exhibited considerable antimicrobial activity against Methicillin-resistant Staphylococcus aureus, and Escherichia coli. The Rapid Annotations using Subsystems Technology showed that the total size of the draft genome was 5,739,603bp, containing 5178 genes with 45.8% G+C content. Genome annotation using antiSMASH version 6.0.0 was performed, which predicted the most common types of non-ribosomal peptide synthetase (NRPS) and polyketide synthase (PKS). In this study, diethyl aminoethyl cellulose (DEAEC) resin was used as the first step in purifying for unknown peptides, and then the target protein was identified using hydrophilic and hydrophobic solutions, optimal pH, and step-by-step tests for antimicrobial activity. This crude was subjected to C18 chromatography and elution with 0, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, and 100% methanol, respectively. Only the fraction eluted with 20% -60% methanol demonstrated good antimicrobial activity against MDR E. coli. The concentration of the active fragment was measured by the Brad-ford test, and Protein A280 - Thermo Fisher Scientific at the end by examining the SDS PAGE Resolving Gel, 10% Acrylamide and purity were confirmed. Our study showed that, based on the combined results of the analysis and purification. P polymyxa. 4G3 has a high potential exists for producing novel functions of polymyxin E and bacitracin against bacterial pathogens.

Keywords: endophytic bacteria, antimicrobial activity, antimicrobial peptide, whole genome sequencing analysis, multi -drug resistance gram negative bacteria