Phytochemical Composition and Anti-Efflux Pump Activity of Hydroalcoholic, Aqueous, and Hexane Extracts of Artemisia tournefortiana in Ciprofloxacin-Resistant Strains of Salmonella enterica Serotype Enteritidis
Background: The AcrB efflux pump in Salmonella species plays a significant role in the development of antibiotic resistance in ciprofloxacin-resistant Salmonella enteritidis. This study aimed to investigate the anti-efflux pump activity of Artemisia tournefortiana extracts among S. Enteritidis strains.
Methods: The hydroalcoholic, aqueous, and hexanolic extracts of A. tournefortiana were prepared and phytochemical composition of extract was determined using gas chromatography/mass spectrometry (GC/MS) method. After antibiogram, the AcrB efflux pump was detected in ciprofloxacin intermediate and resistant S. enteritidis strains using cartwheel and Polymerase chain reaction (PCR) methods. Finally, minimum inhibitory concentrations (MIC) of extracts against S. enteritidis strains were evaluated. After treatment of S. enteritidis strains with sub-MIC concentrations of extracts, the expression level of AcrB efflux pump gene was evaluated using Real-Time PCR.
Results: Phytochemical analysis of extracts using GC/MS method showed that hexadecanoic acid, ethyl ester (30.7%), and cyclopropane,1-(1-hydroxy-1-heptyl)-2-methylene-3-pentyl (17.8%) were the most dominant volatile components volatile compounds in the extract. The results of antibiogram, cartwheel and PCR methods showed that among 20 strains of S. enteritidis that were resistant and intermediate to ciprofloxacin, 16 strains had AcrB efflux pumps. Finally, Real-Time PCR results showed a significant down-regulation of acrB gene in S. enteritidis strains.
Conclusion: A. tournefortiana had anti-efflux activity and this plant can potentially be used as a natural efflux inhibitor for S. enteritidis strains.
2. García-Fernández A, Gallina S, Owczarek S et al (2015). Emergence of Ciprofloxacin-Resistant Salmonella enterica Serovar Typhi in Italy. PLoS One, 10(6):e0132065.
3. Rahman BA, Wasfy MO, Maksoud MA et al (2014). Multi-drug resistance and reduced susceptibility to ciprofloxacin among Sal-monella enterica serovar Typhi isolates from the Middle East and Central Asia. New Microbes New Infect,2(4):88-92.
4. Sharma V, Dahiya S, Jangra P et al (2013). Study of the role of efflux pump in ciprofloxacin resistance in Salmonella enter-ica serotype Typhi. Indian J Med Microbiol, 31(4):374-8.
5. Buckner MM, Blair JM, La Ragione RM et al (2016). Beyond Antimicrobial Resistance: Evidence for a Distinct Role of the AcrD Efflux Pump in Salmonella Biology. MBio, 7(6): e01916-16.
6. Blanco P, Hernando-Amado S, Reales-Calderon JA et al (2016). Bacterial Multi-drug Efflux Pumps: Much More Than Antibiotic Resistance Determinants. Mi-croorganisms, 4(1): E14.
7. Alcalde-RicoM, Hernando-Amado S, Blanco P, Martínez JL (2016). Multidrug efflux pumps at the crossroad between antibi-otic resistance and bacterial virulence. Front Microbiol, 7:1483.
8. Quinn T, O'Mahony R, Baird AW et al (2006). Multi-drug resistance in Salmonel-la enterica: efflux mechanisms and their relationships with the development of chromosomal resistance gene clusters. Curr Drug Targets, 7(7):849-60.
9. Bora KS, Sharma A (2011). The genus Ar-temisia: a comprehensive review. Pharm Biol,49(1):101-9.
10. Abad MJ, Bedoya LM, Apaza L, Bermejo P (2012). The artemisia L. Genus: a review of bioactive essential oils. Molecules, 17(3):2542-66.
11. Benabdelkader T, Zitouni A, Guitton Y, et al (2011). Essential oils from wild popu-lations of Algerian Lavandula stoechas L.: composition, chemical variability, and in vitro biological properties. Chem Biodivers, 8(5):937-53.
12. Clinical and laboratory standards institute (CLSI), 2015. Performance standards for antimicrobial susceptibility testing; 16th informational supplement. CLSI, Wayne, Pa. M100-S16, 26, no. 3.
13. Ferrari RG, Galiana A, Cremades R et al (2013). Expression of the marA, soxS, acrB and ramA genes related to the AcrAB/TolC efflux pump in Salmonella enterica strains with and without quino-lone resistance-determining regions gyrA gene mutations. Braz J Infect Dis, 17(2):125-30.
14. Gislene G. F. Nascimento. Juliana Locatelli. Paulo C. Freitas. Giuliana L. Silva. (2000). Antibacterial activity of plant extract and phytochemicals on antibiotic resistence Bacterial. Braz J Microbiol, 31(4):247-256.
15. Grillon A, Schramm F, Kleinberg M, Jehl F (2016). Comparative Activity of Ciprof-loxacin, Levofloxacin and Moxifloxacin against Klebsiella pneumoniae, Pseudo-monas aeruginosa and Stenotrophomo-nas maltophilia Assessed by Minimum Inhibitory Concentrations and Time-Kill Studies. PLoS One, 11(6):e0156690.
16. Zhong HQ, Zhang S, Pan H, Cai T (2013). Influence of induced ciprofloxacin re-sistance on efflux pump activity of Klebsiella pneumonia. J Zhejiang Univ Sci B, 14(9): 837–843.
17. Huang CB, Alimova Y, Myers TM, Eber-sole JL (2011). Short- and medium-chain fatty acids exhibit antimicrobial activity for oral microorganisms. Arch Oral Biol, 56(7):650-4.
18. Kazemi M, Mozaffarian V, Rustaiyan A et al (2010). Constituents of Artemisia tournefor-tiana Rchb. essential oil from Iran. J Essen-tial Oil Bearing Plants, 13(2): 185-190.
19. McKeegan KS, Borges-Walmsley MI, Walmsley AR (2003). The structure and function of drug pumps: an update. Trends Microbiol, 11(1):21-9.
20. MehtA J, Jandaik S (2016). Evaluation of phytochemical and synergistic interaction between plant extract and antibiotic for efflux pump inhibitory activity against Salmonella enterica serovar thyphimurium strains. Int J Pharm Pharm Sci, 8(10):217-223.
21. Mahmood HY, Jamshidi S, Sutton JM, Rahman KM (2016). Current advances in developing inhibitors of bacterial multi-drug efflux pumps. Curr Med Chem, 23(10):1062-1081.