Volume 25, Issue 3 (August & September 2022)                   J Arak Uni Med Sci 2022, 25(3): 382-393 | Back to browse issues page


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sahabzamani A, sadrnia M, Akbari M, saki S. Efflux Inhibition in Clinical Isolates of Pseudomonas Aeruginosa Using Sertraline for Ciprofloxacin Resistance Breakdown. J Arak Uni Med Sci 2022; 25 (3) :382-393
URL: http://jams.arakmu.ac.ir/article-1-7351-en.html
1- Department of Biology, Arak Branch, Islamic Azad University, Arak, Iran.
2- Department of Biology, Arak Branch, Islamic Azad University, Arak, Iran. , Department of Biology, Payame Noor University (PNU), Tehran, Iran.
3- Department of Biology, Arak Branch, Islamic Azad University, Arak, Iran., Infectious Diseases Research Center (IDRC), Arak University of Medical Sciences, Arak, Iran. , majakbari@yahoo.com
4- Department of Medical Laboratory Sciences, Arak Branch, Islamic Azad University, Arak, Iran.
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Introduction
Pseudomonas aeruginosa plays a key role in various nosocomial infections worldwide. This bacterium is found in many habitats and grows in aerobic conditions. It causes disease with various virulence factors. Currently, P. aeruginosa is inherently resistant to many antibiotics. It might be due to the low permeability of the outer membrane and the efflux pump and the leakage of the enzymes inactivating the antimicrobial agent, making the treatment very difficult. Fluoroquinolones, especially ciprofloxacin, are widely used in the treatment of P. aeruginosa infections. The emergence of resistant strains has caused concerns in this regard. The spread of infectious diseases and consequently an increase in drug resistance have been one of the most serious threats to the successful treatment and health of the community.  The very high costs of processing an antibiotic and its long time to treat a bacterium have resulted in developing newer ways of using antibiotics in the treatment of bacteria. One way to reduce the bacterial stability is the inhibition of antibiotic disposal systems, such as the efflux pump in bacteria [1]..  Sertraline (with trade names of Zoloft and Acentra) is an antidepressant drug from the group of serotonin reuptake inhibitors. It is used to treat depression and is taken orally. Sertraline prevents the reuptake of serotonin by postsynaptic neuroreceptors and an increase in the concentration of serotonin in the central nervous system. Sertraline also slightly inhibits the reuptake of dopamine. Its chemical formula is C17H17Cl2N. Its half-life is between 13 and 45 hours. It has been used in some studies to reduce the function of efflux pump proteins of Gram-negative bacteria, such as E. coli. 
The present study aimed to investigate using this compound in inhibiting the efflux pump in P. aeruginosa .[7]

Materials and Methods
P. aeruginosa isolates were collected from clinical centers. Using biochemical reactions and differential diagnosis, P. aeruginosa isolates were evaluated in terms of sensitivity to the antibiotic ciprofloxacin from the 12-hour growth of the bacteria based on the CLSI2020 M100 standard guidelines in the form of Kirby-Bauer test by Disk diffusion method with five units of ciprofloxacin disk. The isolates with a zone of less than 18 mm were considered resistant strains. Then, for further examination and verification, MIC was evaluated using the CLSI2020M7-8 standard, and the minimum inhibitory concentration of ciprofloxacin for the growth of P. aeruginosa was determined. The resistant strains in terms of the presence of the efflux pump were identified by PCR method using the primer designed by studying the efflux pump gene. 
For resistance breakdown evaluation, 2 % of sertraline was added to the Mueller Hinton agar medium for determining the zone of inhibition of ciprofloxacin.
The microplate dilution method was used for determining the minimum inhibitory concentration. The minimum bactericidal concentration was determined by culturing and MTT methods. Resistance breakdown in broth media was evaluated by the reduction in the minimum inhibitory concentration of the isolates in Muller Hinton Broth and the determination of a significant reduction in bacterial resistance to ciprofloxacin.

Results 
The effect of sertraline on the disc diffusion method was evaluated. In this method, before the addition of sertraline, there was no zone of inhibition (Figure 3), but after the addition of sertraline, the zone of inhibition increased to 25 mm. It means that with the addition of sertraline, the completely drug-resistant bacteria became sensitive.
The results of minimum inhibitory concentration and minimum lethal concentration of ciprofloxacin for drug-resistant strains of P. aeruginosa and standard strain P. aeruginosa 25873 can be seen in Table 3. The results of the MTT assay were also consistent with the culture results in the case of MBC determination.
The results of the presence of the efflux pump gene showed that the resistant strains had efflux pumps.
The results showed that sertraline can prevent the assembling of the efflux pump in P. aeruginosa due to its formulation or interference with its function by forming a complex with the efflux pump proteins. Therefore, fluoroquinolone antibiotics, such as ciprofloxacin, released out of the cell by the efflux pump, prevent the division of the bacteria; thus, the protein-producing function is impaired and the effect of the antibiotic increases.

Discussion
Based on the results, it was concluded that sertraline could inhibit the efficiency of the efflux pump in resistant P. aeruginosa isolates and break the ciprofloxacin resistance in clinical isolates of P. aeruginosa. 
It is recommended to examine these materials for efflux pumps in vivo.

Ethical Considerations

Compliance with ethical guidelines

This research was approved by the Ethics Committee of the Faculty of Medical Sciences of Islamic Azad University, Brojerd Branch (Code: IR.IAU.B.REC.1401.011).

Funding
This article was extracted from the Ph.D. dissertation of the first author at the Department of Microbiology, Faculty of Basic Sciences, Islamic Azad University, Arak Branch.

Authors' contributions
All authors contributed to the research and preparation of this article.

Conflicts of interest
According to the authors, this article has no conflict of interest

 
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Type of Study: Original Atricle | Subject: Infection
Received: 2022/12/19 | Accepted: 2023/01/1

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