Antimicrobial activity and mode of action of 1,8-cineol against carbapenemase-producing Klebsiella pneumoniae

Chew-Li Moo,Saila Ismail,Chou-Min Chong,Wai-Sum Yap,Kok-Song Lai,Mohd Azuraidi Osman,Shun-Kai Yang,Swee-Hua-Erin Lim

doi:10.1038/s41598-021-00249-y

Abstract

Antimicrobial resistance remains one of the most challenging issues that threatens the health of people around the world. Plant-derived natural compounds have received considerable attention for their potential role to mitigate antibiotic resistance. This study was carried out to assess the antimicrobial activity and mode of action of a monoterpene, 1,8-cineol (CN) against carbapenemase-producing Klebsiella pneumoniae (KPC-KP). Results showed that resazurin microplate assay and time-kill analysis revealed bactericidal effects of CN at 28.83 mg/mL. Zeta potential showed that CN increased the surface charge of bacteria and an increase of outer membrane permeability was also detected. CN was able to cause leakage of proteins and nucleic acids in KPC-KP cells upon exposure to CN and ethidium bromide influx/efflux experiment showed the uptake of ethidium bromide into the cell; this was attributed to membrane damage. CN was also found to induce oxidative stress in CN-treated KPC-KP cells through generation of reactive oxygen species which initiated lipid peroxidation and thus damaging the bacterial cell membrane. Scanning and transmission electron microscopies further confirmed the disruption of bacterial cell membrane and loss of intracellular materials. In this study, we demonstrated that CN induced oxidative stress and membrane damage resulting in KPC-KP cell death.

Highlights

Klebsiella pneumoniae, one of the Gram-negative opportunistic pathogens is often the cause of hospital-acquired infections; these include bloodstream infections, respiratory and urinary tract infections, especially in immunocompromised patients with underlying diseases such as chronic pulmonary obstruction, diabetes mellitus or cancer[1,2]
Despite the known anti-inflammatory property and antibacterial activity of CN against a few types of mentioned bacteria, scientific data regarding the antibacterial mechanism of CN against bacteria is limited, especially with reference to antibiotic resistant bacteria to assess the antimicrobial activity of CN against K. pneumoniae carbapenemase (KPC)-KP cells
This study aimed to elucidate the mechanism of action of CN against KPC-KP cells via membrane-related assays, and oxidative stress assessment which further confirmed with scanning electron microscopy and transmission electron microscopy

Summary

Introduction

Klebsiella pneumoniae, one of the Gram-negative opportunistic pathogens is often the cause of hospital-acquired infections; these include bloodstream infections, respiratory and urinary tract infections, especially in immunocompromised patients with underlying diseases such as chronic pulmonary obstruction, diabetes mellitus or cancer[1,2]. Several studies reported that CN exhibits antibacterial activity against K. pneumoniae, E. coli, Salmonella enteritidis, S. aureus[21,24]. According to Sokovicxet al.[25], CN was effective against human pathogenic bacteria strains, namely Bacillus subtilis, E. coli O157:H7, Enterobacter cloacae, Micrococcus flavus, P. aeruginosa, Proteus mirabilis, S. aureus, S. enteritidis, Staphylococcus epidermidis, and Salmonella typhimurium with the MIC value ranging from 4.0 to 7.0 μg/mL. Despite the known anti-inflammatory property and antibacterial activity of CN against a few types of mentioned bacteria, scientific data regarding the antibacterial mechanism of CN against bacteria is limited, especially with reference to antibiotic resistant bacteria to assess the antimicrobial activity of CN against KPC-KP cells. This study aimed to elucidate the mechanism of action of CN against KPC-KP cells via membrane-related assays, and oxidative stress assessment which further confirmed with scanning electron microscopy and transmission electron microscopy

Objectives

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Results

Conclusion