Molecular mechanism of antimicrobial activity of chlorhexidine against carbapenem-resistant Acinetobacter baumannii.

Acinetobacter baumannii causes hospital-acquired infections, especially in those with impaired immune function. Biocides or disinfectants are widely used antibacterial agents used to eradicate the effect of A. baumannii on inanimate objects and health care environments. In the current study, the antimicrobial activity of chlorhexidine has been investigated against carbapenem-resistant (RS-307, RS-7434, RS-6694, and RS-122), and sensitive (ATCC-19606 and RS-10953) strains of A. baumannii. We have determined growth kinetics, antimicrobial susceptibility, ROS production, lipid peroxidation, cell viability using flow cytometry assay (FACS), and membrane integrity by scanning electron microscope (SEM). The effect of chlorhexidine on the bacterial membrane has also been investigated using Fourier transform infrared (FTIR) spectroscopy. The present study showed that 32µg/ml chlorhexidine treatment results in the decreased bacterial growth, CFU count and cell viability. The antibacterial activity of chlorhexidine is due to the elevated ROS production and higher lipid peroxidation. These biochemical changes result in the membrane damage and alteration in the membrane proteins, phospholipids, carbohydrates, nucleic acids as evident from the FTIR and SEM data. Therefore, chlorhexidine has the potential to be used in the hospital setups to remove the spread of A. baumannii.

Comparative mechanism based study on disinfectants against multidrug-resistant Acinetobacter baumannii.

Acinetobacter baumannii has emerged as a hospital-acquired pathogen and has spread in the hospital settings, leading to enhanced nosocomial outbreaks associated with high death rates. Therefore, the aim of the current study is to determine the effective concentration of disinfectants like sodium hypochlorite, hydrogen peroxide, and chlorine dioxide, against multidrug-resistant (MDR) strains of A. baumannii. In this study, we have investigated the effect of disinfectants on different MDR strains i.e. RS307, RS6694, RS7434, RS10953, RS122, and sensitive strain ATCC-19606 of A. baumannii, via differential growth curves analysis, disc diffusion assay, estimation of reactive oxygen species (ROS), lipid peroxidation, and protein carbonylation. All the results collectively showed that 1% sodium hypochlorite (P value < 0.0027), 2.5% hydrogen peroxide (P value = 0.0032), and 10 mM (P value = 0.017) chlorine dioxide significantly inhibit the growth of MDR strains of A. baumannii. A significant increase in the ROS generations, altered lipid peroxidation, and a decrease in protein carbonylation was also observed after treatment with disinfectants, which confirmed its ROS-dependent damage mechanism. These disinfectants also enhance the membrane leakage of reducing sugar, protein, and DNA. The current study highlights and recommends the use of 2.5% hydrogen peroxide to control the MDR strains of A. baumannii in the hospital setup. Therefore, the present results will help in selecting concentrations of different disinfectants for regular use in hospital setups to eradicate the multidrug-resistant A. baumannii from the hospital setup.

Rationale and design of an inhibitor of RecA protein as an inhibitor of Acinetobacter baumannii.

Acinetobacter baumannii is one of the ESKAPE pathogen, which causes pneumonia, urinary tract infections, and is linked to high degree of morbidity and mortality. One-way antibiotic and disinfectant resistance is acquired by the activation of RecA-mediated DNA repair (SOS-response) that maintain ROS-dependent DNA damage caused by these anti-bacterial molecules. To increase the efficacy of different anti-microbial, there is a need to design an inhibitor against RecA of A. baumannii. We have performed homology modeling to generate the structure of RecA, followed by model refinement and validation. High-throughput virtual screening of 1,80,313 primary and secondary metabolites against RecA was performed in HTVS, SP, and XP docking modes. The selected 195 compounds were further analyzed for binding free energy by molecular mechanics approach. The selected top two molecules from molecular mechanics approach were further validated by molecular dynamics simulation (MDS). In-silico high-throughput virtual screening and MDS validation identified ZINC01530654 or  (+-)-2-((4-((7-Chloro-4-quinolyl)amino)pentyl)ethylamino)ethanol sulfate (or hydroxychloroquine sulfate) as a possible lead molecule binding to RecA protein. We have experimentally determined the mechanism of ZINC01530654 to RecA protein. These findings suggest a strategy to chemically inhibit the vital process controlled by RecA that could be helpful for the development of new antibacterial agents.