Chembili Vipin ,Kanekar Saptami ,Fathima Fida,Musliyarakath Mujeeburahiman,Sneha S. Rao,Athmika,Ananthapadmanabha Bhagwath Arun,Punchappady Devasya Rekha
Development of drug resistance in opportunistic pathogens is one of the major healthcare challenges associated with infection management. Combination therapy has many advantages due to the simultaneous action of two drugs on two separate cellular targets. However, selection of the drugs should offer safety and synergistic interaction against most of the strains. Here, the efficacy of antibiotics in combination with quercetin, a natural flavonoid capable of targeting quorum sensing was tested against biofilm-forming Pseudomonas aeruginosa strains previously isolated from catheter associated urinary tract infection. Based on the antibiotic susceptibility pattern, synergistic effect of quercetin with selected antibiotics (levofloxacin, ceftriaxone, gentamycin, tobramycin and amikacin) was tested at the fractional concentrations of MIC by the checkerboard method and the fractional inhibitory concentration index (FICi) was calculated to estimate the synergistic effect. Effect of the synergistic combinations were further tested using time-kill assay, and against biofilm formation and biofilm cell viability. Cytotoxicity assays were performed using Human Embryonic Kidney 293T cells (HEK-293T) using the effective drug combinations with respective controls. The biofilm formation and biofilm cell viability were drastically affected with quercetin and selected antibiotics combinations with ≥80% inhibition. In vitro infection studies showed that all the strains could exert significant cell killing (68 to 85%) and the drug combinations decreased the infection rate significantly by reducing the cell killing effect of P. aeruginosa (p<0.05). The synergistic effect of quercetin is attributed to its quorum sensing inhibitory properties. These findings indicate that quercetin along with existing antibiotics can potentiate the treatment against P. aeruginosa infection and may reduce the selection pressure due to antibiotic overuse.
Resistance to antibiotics among the biofilm-forming Pseudomonas aeruginosa (P. aeruginosa) depict a formidable challenge to the healthcare sector. Infection with antibiotic resistant P. aeruginosa complicates treatment of various conditions ranging from the non-healing of skin wounds to chronic respiratory conditions. Expansion of antibiotic use facilitating the influx of antibiotic traces into the environment has contributed to the high rates of antibiotic resistance. The global changes in gene expression, enhancing virulence, and the acquisition of antibiotic resistance occurs also due to the protection of the bacteria within the biofilm architecture. In P. aeruginosa, quorum sensing is a predominant phenomenon regulating many virulence factors including biofilm formation.
The study was approved by the institutional ethics committee of Yenepoya (Deemed to be University) with protocol number: YUEC386/2016. Informed consent was obtained from all individual participants before collecting the urine samples.
In this study, the antibiotic with quercetin combinations against P. aeruginosa strains isolated from the catheter associated urinary tract infection showed encouraging results. Use of quercetin as a drug in combination enabled to reduce the antibiotic dose while, giving similar or better results compared to monotherapy. Among the antibiotics tested in combination, the synergistic effect shown by different strains are varied and it may be due to the complex genetic mechanisms including the resistance mechanisms. All the selected antibiotic combinations with quercetin have exhibited synergy however, the number of combinations showing synergy was different for different antibiotics and strains. Compared to other antibiotics, the quercetin–tobramycin and quercetin–amikacin combination showed synergy in more fractional doses against all the strains compared to other antibiotics. The synergistic activity of the two drug molecules are achieved if the two individual components have different modes of action and such action can bring about higher killing effect on the bacteria. Here, quercetin being a quorum sensing inhibitor, iwill act to control the bacterial population density, while the antibiotics exert the cell killing effect.
The authors thank Dr. Ashwini Prabhu for the technical support provided during in vitro infection studies.
Citation: Vipin C, Saptami K, Fida F, Mujeeburahiman M, Rao SS, Athmika, et al. (2020) Potential synergistic activity of quercetin with antibiotics against multidrug-resistant clinical strains of Pseudomonas aeruginosa. PLoS ONE 15(11): e0241304. https://doi.org/10.1371/journal.pone.0241304
Editor: Yu-Wei Lin, Monash University, AUSTRALIA
Received: May 11, 2020; Accepted: October 12, 2020; Published: November 6, 2020
Copyright: © 2020 Vipin et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
All relevant data are within the paper and its Supporting Information files.
CV and KS are recipients of Indian Council of Medical Research-Senior Research Fellowship (OMI-Fellowship/32/2018-ECD-I).
The authors have declared that no competing interests exist.