Antibacterial and Antibiofilm Activity of Grape Seed Extract Against Carbapenem Resistant and Biofilm Producer Enterobacteriaceae

Document Type : Original paper


1 Immunology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.

2 Faculty of Pharmacy, Drug Applied Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.

3 Department of Microbiology, School of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran. Students’ Research Committee, Tabriz University of Medical Sciences, Tabriz, Iran.

4 Immunology Research Center, Tabriz University of Medical Sciences, Tabriz, Iran. Department of Microbiology, School of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran.


Background and objectives: Carbapenem-resistant and biofilm producing Enterobacteriaceaeare a major health problem. This study was carried to determine the antibacterial and antibiofilm activity of grape seed extract (GSE) against carbapenem-resistant and biofilm producing Enterobacteriaceae isolates. Methods: Antibiotics susceptibility patterns were detected by the disk diffusion method. carbapenem-resistant Enterobacteriaceae (CRE) isolates were screened by carbapenems disks and imipenem minimum inhibitory concentrations (MIC). The biofilm formation was detected by the microplate method. The carbapenemase genes were detected by PCR. The total polyphenolic content of GSE was determinate by Folin Ciocalteu technique. The antibacterial and antibiofilm effects of GSE were tested by the MIC and biofilm inhibitory concentration (BIC), respectively. Results: In this study, total phenolic content of extracted 1 gram of GSE was equivalent to 700 mg gallic acid. Eighteen non-duplicated CRE isolates were selected. All isolates were fosfomycin susceptible. Variable frequency of resistance to the other tested antibiotics was observed. The blaOXA-48 was the most common carbapenemase type. Among 18 isolates, 13 were biofilm producer while GSE inhibited CRE growth at 1024 µg/mL for 15 isolates and 2048 µg/mL for three isolates. Biofilm production was inhibited by GSE in 2000 µg/mL, 4000 µg/mL and 8000 µg/mL after 72 h incubation. Conclusion: The significant antibacterial and antibiofilm effects of GSE suggested GSE as a promising candidate for treatment of infections caused by these organisms.


Main Subjects

[1] Paterson DL. Resistance in gram-negative bacteria: Enterobacteriaceae. Am J Med. 2006; 119(6): 20-28.
[2] Eryılmaz M, Bozkurt ME, Yildiz MM, Akin A. Antimicrobial resistance of urinary Escherichia coli isolates. Trop J Pharm Res. 2010; 9(2): 205-209.
[3] Vardakas KZ, Tansarli GS, Rafailidis PI, Falagas ME. Carbapenems versus alternative antibiotics for the treatment of bacteraemia due to Enterobacteriaceae producing extended-spectrum β-lactamases: a systematic review and meta-analysis. J Antimicrob Chemoth. 2012: 67(12): 2793-2803.
[4] Gupta N, Limbago BM, Patel JB, Kallen AJ. Carbapenem-resistant Enterobacteriaceae: epidemiology and prevention. J Clin Microbiol. 2011; 53(1): 60-67.
[5] Schwaber MJ, Carmeli Y. Carbapenem-resistant Enterobacteriaceae: a potential threat. Jama. 2008; 300(24): 2911-2913.
[6] Van Duin D, Kaye KS, Neuner EA, Bonomo RA. Carbapenem-resistant Enterobacteriaceae: a review of treatment and outcomes. Diagn Micr Infec Dis. 2013; 75(2): 115-120.
[7] Ghotaslou R, Salahi B. Effects of oxygen on in-vitro biofilm formation and antimicrobial resistance of Pseudomonas aeruginosae. Pharma Sci. 2013;19(3):96-99.
[8] Ghotaslou R, Bahari Z, Aliloo A, Gholizadeh P, Eshlaghi BS. The in vitro effects of silver nanoparticles on bacterial biofilms. J Microbiol Biotech Food Sci. 2017; 6(4): 1077-1080.
[9] Basri DF, Xian LW, Abdul Shukor NI, Latip J. Bacteriostatic antimicrobial combination: antagonistic interaction between epsilon-viniferin and vancomycin against methicillin-resistant Staphylococcus aureus. Biomed Res Int. 2014; Article ID 461756.
[10] Baydar NG, Sagdic O, Ozkan G, Cetin S. Determination of antibacterial effects and total phenolic contents of grape (Vitis vinifera L.) seed extracts. Int J Food Sci Technol. 2006; 41(7): 799-804.
[11] Jayaprakasha G, Selvi T, Sakariah K. Antibacterial and antioxidant activities of grape (Vitis vinifera) seed extracts. Food Res Int. 2003; 36(2): 117-122.
[12] Nirmala JG, Narendhirakannan R. In vitro antioxidant and antimicrobial activities of grapes (Vitis vinifera L.) seed and skin extracts-Muscat variety. Int J Pharm Pharm Sci. 2011; 3(4): 242-249.
[13] Clinical and Laboratory Standards Institute (CLSI). Performance standards for antimicrobial susceptibility testing; twenty-second informational supplements; Wayne: Clinical and Laboratory Standards Institute (CLSI), 2012.
[14] Andrews JM. Determination of minimum inhibitory concentrations. J Antimicrob Chemother. 2001; 48(S1): 5-16.
[15] Clinical and Laboratory Standards Institute (CLSI). Performance standards for antimicrobial susceptibility testing: twenty-first informational supplement: Wayne: Clinical and Laboratory Standards Institute (CLSI), 2011.
[16] Dallenne C, Da Costa A, Decré D, Favier C, Arlet G. Development of a set of multiplex PCR assays for the detection of genes encoding important β-lactamases in Enterobacteriaceae. J Antimicrob Chemother. 2010: 65(3): 490-495.
[17] NCBI. Basic local alignment search tool (BLAST). [Accessed 2017]. Available from: 
[18] Lahay clinic. ß-Lactamase classification and amino acid sequences. [Accessed 2017]. Available from:
[19] Burton E, Yakandawala N, LoVetri K, Madhyastha M. A microplate spectrofluorometric assay for bacterial biofilms. J Ind Microbiol Biot. 2007; 34(1): 1-4.
[20] Moskowitz SM, Foster JM, Emerson J, Burns JL. Clinically feasible biofilm susceptibility assay for isolates of Pseudomonas aeruginosa from patients with cystic fibrosis. J Clin Microbiol. 2004; 42(5): 1915-1922.
[21] Delgado-Valverde M, Sojo-Dorado J, Pascual Á, Rodríguez-Baño J. Clinical management of infections caused by multidrug-resistant Enterobacteriaceae. Ther Adv Infect Dis. 2013; 1(2): 49-69.
[22] Sadeghi MR, Ghotaslou R, Akhi MT, Asgharzadeh M, Hasani A. Molecular characterization of extended-spectrum β-lactamase, plasmid-mediated AmpC cephalosporinase and carbapenemase genes among Enterobacteriaceae isolates in five medical centres of East and West Azerbaijan, Iran. J Med Microbiol. 2016; 65(11): 1322-1331.
[23] Morrill HJ, Pogue JM, Kaye KS, La Plante KL. Treatment options for carbapenem-resistant Enterobacteriaceae infections. Open Forum Infect Dis. 2015; Article ID PMC4462593.
[24] Peng X, Ma J, Cheng KW, Jiang Y, Chen F, Wang M. The effects of grape seed extract fortification on the antioxidant activity and quality attributes of bread. Food Chem. 2010; 119(1): 49-53.
[25] Furiga A, Lonvaud-Funel A, Badet C. In vitro study of antioxidant capacity and antibacterial activity on oral anaerobes of a grape seed extract. Food Chem. 2009; 113(4): 1037-1040.
[26] Han Y. Synergic effect of grape seed extract with amphotericin B against disseminated candidiasis due to Candida albicans. Phytomedicine. 2007; 14(11): 733-738.
[27] Bruno G, Sparapano L. Effects of three esca-associated fungi on Vitis vinifera L.: V. Changes in the chemical and biological profile of xylem sap from diseased cv. Sangiovese vines. Physiol Mol Plant Pathol. 2007; 71(4): 210-229.
[28] Montealegre RR, Peces RR, Vozmediano JC, Gascueña JM, Romero EG. Phenolic compounds in skins and seeds of ten grape Vitis vinifera varieties grown in a warm climate. J Food Compos Analysis. 2006; 19(6): 687-693.
[29] Al-Habib A, Al-Saleh E, Safer AM, Afzal M. Bactericidal effect of grape seed extract on methicillin-resistant Staphylococcus aureus (MRSA). J Toxicol Sci. 2010; 35(3): 357-364.
[30] Brown JC, Huang G, Haley-Zitlin V, Jiang X. Antibacterial effects of grape extracts on Helicobacter pylori. Appl Environ Microbiol. 2009; 75(3): 848-852.
[31] Mirkarimi M, Amin-Marashi SM, Bargrizan M, Abtahi A, Fooladi I, Ali A. The antimicrobial activity of grape seed extract against two important oral pathogens. Zahedan J Res Med Sci. 2013; 15(1): 43-46.
[32] Zhao W, Xie Q, Bedran-Russo AK, Pan S, Ling J, Wu CD. The preventive effect of grape seed extract on artificial enamel caries progression in a microbial biofilm-induced caries model. J Dent. 2014; 42(8): 1010-1018.
[33] Ugartondo V, Mitjans M, Lozano C, Torres JL, Vinardell MP. Comparative study of the cytotoxicity induced by antioxidant epicatechin conjugates obtained from grape. J Agr Food Chem. 2006; 54(18): 6945-6950.
[34] Fan P, Lou H. Effects of polyphenols from grape seeds on oxidative damage to cellular DNA. Mol Cell Biochem. 2004; 267(1-2): 67-74.