This is a preview and has not been published.

Estimation of the In-Vitro Synergistic Effect of Vancomycin with Bacteriophage and its Endolysin on Iraqi Local Isolates of Enterococcus faecalis


  • Rand R. Hafidh Department of Microbiology, College of Medicine, University of Baghdad, Baghdad, Iraq.
  • Zainab Oday Hamed Department of pharmacy, Baghdad College of Medical Sciences, Baghdad, Iraq.
  • Abdullah A. Awni Ministry of Health, Baghdad Health Directorate -Al-Karkh, Al-Furat General Hospital, Baghdad, Iraq.
  • Manal N. Jassas Microbiology Division, Laboratory Department, Ghazi Hariri Hospital for Specialized Surgery, Baghdad, Iraq.



Bacteriophages, Enterococcus faecalis, Endolysin, Multidrug resistance bacteria, Phage therapy, Phage cocktail


Enterococcus faecalis is known to cause many serious infections with an upsurge in the development of antibiotic resistant strains, especially vancomycin. Lytic bacteriophage and its endolysin have the potential to affect and lyse specific bacterial cells making them a possible alternative therapy to antibiotic resistant strains. The study aimed to investigate the synergistic effect of phage/ phage cocktail/ endolysin with vancomycin on E. faecalis isolated from urine samples. The antibioresistant profile 17 E. faecalis was done with an emphasis on the sensitivity to vancomycin. The lytic phages were isolated from the local environment. The infectivity and the coverage rate for the phage and phage cocktail were calculated. The enzyme of the lytic phage was isolated by chromatography. The synergistic effect was tested by turbidity reduction assay to evaluate the growth reduction level. It had been found that 6/17 E. faecalis isolates were resistant to vancomycin, 7/17 isolates were intermediate, and 4/17 were susceptible. Three lytic phages were identified with an infectivity rate of 100%, 76.4%, and 88.2%. The phage cocktail yielded 100% rate of infectivity and 91.6 % coverage rate. The reduction in the growth levels of the 17 E.faecalis after treatment with phage / phage cocktail/ vancomycin/ mixture of vancomycin and phage cocktail was effective. The synergistic effect of vancomcyin together with the phage cocktail significantly reduced the growth of 70.6% of the isolates. The antibacterial activity of vancomycin plus endolysin treatment was revealed in a time dependent manner. Thus, the synergistic effect of vancomycin with phage/ endoloysin illustrated an effective method to treat the antibiotic-resistant strains of E. faecalis to overcome this emerging clinical problem.


Download data is not yet available.


Bolocan AS, Upadrasta A, Bettio PHA, Clooney AG, Draper LA, Ross RP, et al. Evaluation of Phage Therapy in the Context of Enterococcus faecalis and Its Associated Diseases. Viruses. 2019; 11(4): 366.

Fiore E, Van Tyne D, Gilmore MS. Pathogenicity of Enterococci. Microbiol Spectr. 2019; 7(4): 10.

Sirichoat A, Flórez AB, Vázquez L, Buppasiri P, Panya M, Lulitanond V, et al. Antibiotic Resistance-Susceptibility Profiles of Enterococcus faecalis and Streptococcus spp. From the Human Vagina, and Genome Analysis of the Genetic Basis of Intrinsic and Acquired Resistances. Front Microbiol. 2020; 11: 1438.

Tagliabue A, Rappuoli R. Changing Priorities in Vaccinology: Antibiotic Resistance Moving to the Top. Front Immunol. 2018; 9: 1068.

Mahmood BSh. Silver Nanoparticles and their Role in Gene Expression of Motility Gene motB and Repression of AI-2-Controlled Gene. Baghdad Sci J. 2020; 17(3): 916-923.

Abdul Alaameri SK, Al-Hayanni HSA. Antibacterial and anti-biofilm effects of Sumac (Rhus coriaria L) fruits extracts against some multidrug-resistant pathogenic bacteria. J Fac Med Baghdad. 2022; 64(3): 183-188.

Simmonds P, Aiewsakun P. Virus classification - where do you draw the line? Arch Virol. 2018; 163(8): 2037-2046.

García R, Latz S, Romero J, Higuera G, García K, Bastías R. Bacteriophage Production Models: An Overview. Front Microbiol. 2019; 10: 1187.

Abedon ST, Danis-Wlodarczyk KM, Wozniak DJ. Phage Cocktail Development for Bacteriophage Therapy: Toward Improving Spectrum of Activity Breadth and Depth. Pharmaceuticals (Basel). 2021; 14(10): 1019.

Khalifa L, Gelman D, Shlezinger M, Dessal AL, Coppenhagen-Glazer S, Beyth N, et al. Defeating Antibiotic- and Phage-Resistant Enterococcus faecalis Using a Phage Cocktail in Vitro and in a Clot Model. Front Microbiol. 2018; 9: 326.

Ahmed ShH, Hafidh RR. The Isolation of specifically lytic phages along with their extracted endolysins as antibacterial agents to MDR Enterococcus faecalis. Res J Pharm Technol. 2021; 14(9): 4547-4.

Jassas MN, Hafidh RR. Characterization of the Antibacterial, Antibiofilm Activities, and Genetic Structure of Endolysin Extracted from Enterococcus faecalis Phage. Int J Drug Deliv Technol. 2021; 11(3): 817-826.

Song M, Wu D, Hu Y, Luo H, Li G. Characterization of an Enterococcus faecalis Bacteriophage vB_EfaM_LG1 and Its Synergistic Effect With Antibiotic. Front Cell Infect Microbiol. 2021; 11: 698807.

De Almeida CV, Taddei A, Amedei A. The controversial role of Enterococcus faecalis in colorectal cancer. Ther Adv Gastroenterol. 2018; 11: 175628481878360.

Iseberg HD. Clinical microbiology procedures handbook. Washington DC: ASM Press; 2004.

Balouiri M, Sadiki M, Ibnsouda SK. Methods for in vitro evaluating antimicrobial activity: A review. J Pharm Anal. 2016; 6(2): 71-79.

Alsaadi LAS. Molecular detection of the mex efflux pumps genes in extensively drug-resistatn and pandrug-resistant Pseudomonas aeruginosa isolated from Iraqi patients in Diyala. World Bull Public Health. 2022; 10: 97-105.

Patel JB. Performance Standards for Antimicrobial Susceptibility Testing. 30th ed. M100, Wayne PA: Clinical and Laboratory Standards Institute; 2020.

Andrews JM. Determination of minimum inhibitory concentrations. J Antimicrob Chemother. 2001; 48(1): 5-16.

Jurczak-Kurek A, Gąsior T, Nejman-Faleńczyk B, Bloch S, Dydecka A, Topka G, et al. Biodiversity of bacteriophages: morphological and biological properties of a large group of phages isolated from urban sewage. Sci Rep. 2016; 6: 34338.

Ali RM, Abdulamir SA, Kadhim RS. Extraction, purification and therapeutic use of bacteriophage endolysin against multi-drug resistant Staphylococcus aureus : In vivo and in vitro study. J Contemp Med Sci. 2018; 4: 33-38.

Jasim HN, Hafidh RR, Abdulamir AS. Formation of therapeutic phage cocktail and endolysin to highly multi-drug resistant Acinetobacter baumannii: in vitro and in vivo study. Iran J Basic Med Sci 2018; 21: 1100-1108.

Mohammed AR. Modifying Plaque assay and Clearance test as tools in determination of phage typing for E. Coli bacterial interspecies. Baghdad Sci J. 2013; 10(1): 161-167.

Lin E, Bhusal Y, Horwitz D, Shelburne SA, Trautner BW. Overtreatment of Enterococcal Bacteriuria. Arch Intern Med. 2012; 172(1): 33–38.

Li M, Yang F, Lu Y, Huang W. Identification of Enterococcus faecalis in a patient with urinary-tract infection based on metagenomic next-generation sequencing: a case report. BMC Infect Dis. 2020; 20: 467.

Jafarzadeh SR, Tajbakhsh E, Momtaz H, Kabiri SM. Prevalence of Virulence Genes and Antibiotic Resistance Pattern in Enterococcus faecalis Isolated from Urinary Tract Infection in Shahrekord, Iran. Rep Biochem Mol Biol. 2021; 10(1): 50–59.

Ghalavand Z, Alebouyeh M, Ghanati K, Azimi L, Rashidan M. Genetic relatedness of the Enterococcus faecalis isolates in stool and urine samples of patients with community-acquired urinary tract infection. Gut pathogens. 2020; 12: 42.

Palmer KL, Kos VN, Gilmore MS. Horizontal Gene Transfer and the Genomics of Enterococcal Antibiotic Resistance. Curr Opin Microbiol. 2010; 13: 632–639.

Chatterjee A, Johnson CN, Luong P, Hullahalli K, McBride SW, Schubert AM, et al. Bacteriophage Resistance Alters Antibiotic-Mediated Intestinal Expansion of Enterococci. Infect Immun. 2019; 87 (6): e00085-19.

Shlezinger M, Coppenhagen-Glazer S, Gelman D, Beyth N, Hazan R. Eradication of Vancomycin-Resistant Enterococci by Combining Phage and Vancomycin. Viruses 2019; 11(10): 954.

Zaidan IA, AL-Kazaz AA, Mohammed AS. Effect the combination of antibiotics on clinical isolates of Staphylococcus aureus. Baghdad Sci J. 2009; 6 (4): 683-92.

Melconian AK, Al-Baldawi MS, Al-Falahi MA. Effect of Subinhibitory concentration of Antibiotic on Bacterial Adherence to Orthopedic Prosthetic Device. Baghdad Sci J. 2021; 4 (1): 28-34.

Zhao Y, Feng L, Zhou B, Zhang X, Yao Z, Wang L, et al. A newly isolated bacteriophage vB8388 and its synergistic effect with aminoglycosides against multi-drug resistant Klebsiella oxytoca strain FK-8388. Microb. Pathog . 2023; 174: 105906.

Manohar P, Madurantakam Royam M, Loh B, Bozdogan B, Nachimuthu R, Leptihn S. Synergistic Effects of Phage-Antibiotic Combinations against Citrobacter amalonaticus. ACS Infect Dis. 2022 14; 8(1): 59-65.

Hong HW, Kim YD, Jang J, Kim MS, Song M, Myung H. Combination Effect of Engineered Endolysin EC340 With Antibiotics. Front Microbiol. 2022 Feb 15; 13: 821936.