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  • 1 Department of Microbiology, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, , Iran
  • | 2 Pediatric Infections Research Center, Mofid Children's Hospital, Shahid Beheshti, University of Medical Sciences, Tehran, , Iran
  • | 3 Department of Pathobiology, School of Public Health, Tehran University of Medical Sciences, Tehran, , Iran
  • | 4 Department of Biostatistics, School of Paramedical Sciences, Shahid Beheshti University of Medical Sciences, Tehran, , Iran
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Abstract

Antibiotic resistance and especially multiresistance in Enterococci, is a serious public health issue especially in infections of immunocompromised patients. EfrAB is a heterodimeric multidrug ATP-binding cassette (ABC) transporter that causes endogenous resistance to antimicrobials including fluoroquinolones in Enterococcus spp. The aim of this study was to seek the gene expression rate and role of efrAB efflux pump in ciprofloxacin resistant Enterococcus faecalis and Multilocus Sequence Typing (MLST) of multiresistant isolates. Phenotypic and genotyping identification of 80 E. faecalis isolates were performed. Minimum inhibitory concentrations (MICs) to ciprofloxacin (CIP) were measured with and without carbonylcyanide 3-chlorophenylhydrazone (CCCP) by broth microdilution. After DNA extraction and sequencing for detection of efrA and efrB genes, the efrAB efflux positive isolates that were resistant to ciprofloxacin and showed decrease of ciprofloxacin MIC range were identified. Isolates that exhibited decrease in ciprofloxacin MIC range from two to ten folds were assessed for biofilm formation and finally, the expression levels of efrB, efrA genes were measured by quantitative Real-Time PCR (qRT-PCR). High rates of resistance to tetracycline and minocycline and low rates of resistance to the most antibiotics used in this study were detected. The results in this study indicated that the incidence of Multiple drug resistance (MDR) was 23.7% and all isolates that were resistant to ciprofloxacin revealed several degrees of overexpression in efrA and efrB genes. Our study found two ST480 and one ST847 in E. faecalis isolates. In conclusion, despite of low frequency of resistance to the most antibiotics and MDRs in our region, we found one ST480 isolate with resistance to eight antibiotics that also exists in other parts of the world.

  • [1]

    Kitagawa K, Shigemura K, Yamamichi F, Alimsardjono L, Rahardjo D, Kuntaman K, et al.. International comparison of causative bacteria and antimicrobial susceptibilities of urinary tract infections between Kobe, Japan and Surabaya, Indonesia. JJID 2017; 71: 813.

    • Search Google Scholar
    • Export Citation
  • [2]

    Sharma D, Preston SE, Hage R. Emerging antibiotic resistance to bacterial isolates from human urinary tract infections in Grenada. Cureus 2019; 11: e5752.

    • Search Google Scholar
    • Export Citation
  • [3]

    Defoirdt T, Sorgeloos P, Bossier P. Alternatives to antibiotics for the control of bacterial disease in aquaculture. Curr Opin Microbiol 2011; 14: 2518.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [4]

    Lee EW, Huda MN, Kuroda T, Mizushima T, Tsuchiya T. EfrAB, an ABC multidrug efflux pump in Enterococcus faecalis. Antimicrob Agents Chemother 2003; 47: 37338.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [5]

    Shepard BD, Gilmore MS. Antibiotic-resistant enterococci: the mechanisms and dynamics of drug introduction and resistance. Microb Infect 2002; 4: 21524.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [6]

    Pesavento G, Calonico C, Ducci B, Magnanini A, Nostro AL. Prevalence and antibiotic resistance of Enterococcus spp. isolated from retail cheese, ready-to-eat salads, ham, and raw meat. Food Microbiol 2014; 41: 17.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [7]

    Conceição N, Oliveira CdCHB, Silva PRd, Ávila BGM, Oliveir AGd. Trends in antimicrobial resistance among clinical isolates of enterococci in a Brazilian tertiary hospital: a 4-year study. Rev Soc Bras Med Tro 2011; 44: 17781.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [8]

    Yasufuku T, Shigemura K, Shirakawa T, Matsumoto M, Nakano Y, Tanaka K, et al. Mechanisms of and risk factors for fluoroquinolone resistance in clinical Enterococcus faecalis isolates from patients with urinary tract infections. J Clin Microbiol 2011; 49: 391216.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [9]

    Jones ME, Draghi DC, Thornsberry C, Karlowsky JA, Sahm DF, Wenzel RP. Emerging resistance among bacterial pathogens in the intensive care unit – a European and North American Surveillance study (2000–2002). Ann Clin Microbiol Antimicrob 2004; 3: 14.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [10]

    Giannakopoulos X, Sakkas H, Ragos V, Tsiambas E, Bozidis P, Evangelou AM, et al.. Impact of enterococcal urinary tract infections in immuno-compromised–neoplastic patients. J BUON 2019; 24: 176875.

    • Search Google Scholar
    • Export Citation
  • [11]

    El Khoury J, Fishman J. Linezolid in the treatment of vancomycin‐resistant Enterococcus faecium in solid organ transplant recipients: report of a multicenter compassionate‐use trial. Transpl Infect Dis 2003; 5: 1215.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [12]

    Barbosa J, Ferreira V, Teixeira P. Antibiotic susceptibility of enterococci isolated from traditional fermented meat products. Food Microbiol 2009; 26: 52732.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [13]

    Kumar A, Schweizer HP. Bacterial resistance to antibiotics: active efflux and reduced uptake. Adv Drug Deliv Rev 2005; 57: 1486513.

  • [14]

    Higgins CF. ABC transporters: from microorganisms to man. Annu Rev Cell Dev Biol 1992; 8: 67113.

  • [15]

    Fath MJ, Kolter R. ABC transporters: bacterial exporters. Microbiol Rev 1993; 57: 9951017.

  • [16]

    Kristiansen JE, Hendricks O, Delvin T, Butterworth TS, Aagaard L, Christensen JB, et al.. Reversal of resistance in microorganisms by help of non-antibiotics. J Antimicrob Chemother 2007; 59: 121779.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [17]

    Levy SB. Active efflux mechanisms for antimicrobial resistance. Antimicrob Agents Chemother 1992; 36: 695.

  • [18]

    Lewis K. In search of natural substrates and inhibitors of MDR pumps. J Mol Microbiol Biotechnol 2001; 3: 24754.

  • [19]

    Saier M. Jr, Tam R, Reizer A, Reizer J. Two novel families of bacterial membrane proteins concerned with nodulation, cell division and transport. Mol Microbiol 1994; 11: 8417.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [20]

    Hürlimann LM, Corradi V, Hohl M, Bloemberg GV, Tieleman DP, Seeger MA. The heterodimeric ABC transporter EfrCD mediates multidrug efflux in Enterococcus faecalis. Antimicrob Agents Chemother 2016; 60: 540011.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [21]

    Lerma LL, Benomar N, Valenzuela AS, Muñoz MdCC, Gálvez A, Abriouel H. Role of EfrAB efflux pump in biocide tolerance and antibiotic resistance of Enterococcus faecalis and Enterococcus faecium isolated from traditional fermented foods and the effect of EDTA as EfrAB inhibitor. Food Microbiol 2014; 44: 24957.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [22]

    Levine DP. Vancomycin: a history. Clin Infect Dis 2006; 42: S512.

  • [23]

    Gutiérrez-Castrellón P, Díaz-García L, de Colsa-Ranero A, Cuevas-Alpuche J, Jiménez-Escobar I. Efficacy and safety of ciprofloxacin treatment in urinary tract infections (UTIs) in adults: a systematic review with meta-analysis. Gac Med Mexico 2015; 151: 21028.

    • Search Google Scholar
    • Export Citation
  • [24]

    Mandell L, Tillotson G. Safety of fluoroquinolones: an update. Can J Infect Dis Med 2002; 13: 5461.

  • [25]

    Huycke MM, Sahm DF, Gilmore MS. Multiple-drug resistant enterococci: the nature of the problem and an agenda for the future. Emerg Infect Dis 1998; 4: 239.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [26]

    Lee G. Ciprofloxacin resistance in Enterococcus faecalis strains isolated from male patients with complicated urinary tract infection. Korean J Urol 2013; 54: 38893.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [27]

    Matsumoto T, Hamasuna R, Ishikawa K, Takahashi S, Yasuda M, Hayami H, et al.. Nationwide survey of antibacterial activity against clinical isolates from urinary tract infections in Japan (2008). Int J Antimicrob Agents 2011; 37: 21018.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [28]

    Talebi M, Moghadam NA, Mamooii Z, Enayati M, Saifi M, Pourshafie MR. Antibiotic resistance and biofilm formation of Enterococcus faecalis in patient and environmental samples. Jundishapur J Microbiol 2015; 8: e23349.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [29]

    Wayne P. CLSI. Performance standards for antimicrobial susceptibility testing; Twenty-fifth informational supplement. CLSI document M100-S25. Clinical and Laboratory Standards Institute; 2015.

    • Search Google Scholar
    • Export Citation
  • [30]

    Lerma LL, Benomar N, Valenzuela AS, Munoz MdCC, Gálvez A, Abriouel H. Role of EfrAB efflux pump in biocide tolerance and antibiotic resistance of Enterococcus faecalis and Enterococcus faecium isolated from traditional fermented foods and the effect of EDTA as EfrAB inhibitor. Food Microbiol 2014; 44: 24957.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [31]

    Shiadeh SMJ, Hashemi A, Fallah F, Lak P, Azimi L, Rashidan M. First detection of efrAB, an ABC multidrug efflux pump in Enterococcus faecalis in Tehran, Iran. Acta Microbiol Immunol Hung 2019; 66: 5768.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [32]

    Ruiz-Garbajosa P, Bonten MJ, Robinson DA, Top J, Nallapareddy SR, Torres C, et al.. Multilocus sequence typing scheme for Enterococcus faecalis reveals hospital-adapted genetic complexes in a background of high rates of recombination. J Clin Microbiol 2006; 44: 222028.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [33]

    Tendolkar PM, Baghdayan AS, Gilmore MS, Shankar N. Enterococcal surface protein, Esp, enhances biofilm formation by Enterococcus faecalis. Infect Immun 2004; 72: 60329.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [34]

    Rutala WA, Kanamori H, Gergen MF, Knelson LP, Sickbert-Bennett EE, Chen LF, et al.. Enhanced disinfection leads to reduction of microbial contamination and a decrease in patient colonization and infection. Infect Cont Hosp Ep 2018; 39: 111821.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [35]

    Levitus M, Perera TB. Vancomycin-Resistant Enterococci (Vre). StatPearls [Internet]. StatPearls Publishing; 2018.

  • [36]

    Pazoles J, Talbot MK, Alder EA, White AC, Jonas BM, Murray BE, et al.. Enterococcus faecalis multi-drug resistance transporters: application for antibiotic discovery. J Mol Microbiol Biotechnol 2001; 3: 17984.

    • Search Google Scholar
    • Export Citation
  • [37]

    Al-Jarousha A-MK, Saed A, Afifi H. Prevalence of multidrug resistant enterococci in nosocomial infection in Gaza strip. J Al-Aqsa Unv 2008; 12: 1524.

    • Search Google Scholar
    • Export Citation
  • [38]

    Hasan AS, Nair D, Kaur J, Baweja G, Deb M, Aggarwal P. Resistance patterns of urinary isolates in a tertiary Indian hospital. J Ayub Med Coll Abbottabad 2007; 19: 3941.

    • Search Google Scholar
    • Export Citation
  • [39]

    Kuch A, Willems RJ, Werner G, Coque TM, Hammerum AM, Sundsfjord A, et al.. Insight into antimicrobial susceptibility and population structure of contemporary human Enterococcus faecalis isolates from Europe. J Antimicrob Chemother 2011; 67: 5518.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [40]

    Wang Q-Y, Li R-H, Shang X-H. Urinary tract infection caused by Enterococcus isolates: aetiology and antimicrobial resistance patterns. J Chemother 2015; 27: 11719.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [41]

    Yilema A, Moges F, Tadele S, Endris M, Kassu A, Abebe W, et al.. Isolation of enterococci, their antimicrobial susceptibility patterns and associated factors among patients attending at the University of Gondar Teaching Hospital. BMC Infect Dis 2017; 17: 276.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [42]

    Lubelski J, Konings WN, Driessen AJ. Distribution and physiology of ABC-type transporters contributing to multidrug resistance in bacteria. Microbiol Mol Biol Rev 2007; 71: 46376.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [43]

    Kang S, Lee S, Choi S. Distribution of multidrug efflux pump genes in Enterococci spp. isolated from bovine milk samples and their antibiotic resistance patterns. Korean J Microbiol 2013; 49: 12630.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [44]

    Sanchez Valenzuela A, Lavilla Lerma L, Benomar N, Gálvez A, Perez Pulido R, Abriouel H. Phenotypic and molecular antibiotic resistance profile of Enterococcus faecalis and Enterococcus faecium isolated from different traditional fermented foods. Foodborne Pathog Dis 2013; 10: 1439.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [45]

    Mendes RE, Deshpande L, Streit JM, Sader HS, Castanheira M, Hogan PA, et al.. ZAAPS programme results for 2016: an activity and spectrum analysis of linezolid using clinical isolates from medical centres in 42 countries. J Antimicrob Chemother 2018; 73: 18807.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [46]

    Said LB, Klibi N, Lozano C, Dziri R, Slama KB, Boudabous A, et al.. Diversity of enterococcal species and characterization of high-level aminoglycoside resistant enterococci of samples of wastewater and surface water in Tunisia. Sci Total Environ 2015; 530: 1117.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [47]

    Sassi M, Guérin F, Zouari A, Beyrouthy R, Auzou M, Fines-Guyon M, et al.. Emergence of optrA-mediated linezolid resistance in enterococci from France, 2006–16. J Antimicrob Chemother 2019; 74: 146972.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [48]

    Zhou W, Gao S, Xu H, Zhang Z, Chen F, Shen H, et al.. Distribution of the optrA gene in Enterococcus isolates at a tertiary care hospital in China. J Glob Antimicrob Re 2019; 17: 1806.

    • Crossref
    • Search Google Scholar
    • Export Citation
  • [49]

    Chen H, Wang X, Yin Y, Li S, Zhang Y, Wang Q, et al.. Molecular characteristics of oxazolidinone resistance in enterococci from a multicenter study in China. BMC Med 2019; 19: 162.

    • Search Google Scholar
    • Export Citation

 

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Senior editors

Editor-in-Chief: Prof. Dóra Szabó (Institute of Medical Microbiology, Semmelweis University, Budapest, Hungary)

Managing Editor: Dr. Béla Kocsis (Institute of Medical Microbiology, Semmelweis University, Budapest, Hungary)

Co-editor: Dr. Andrea Horváth (Institute of Medical Microbiology, Semmelweis University, Budapest, Hungary)

Editorial Board

  • Prof. Éva ÁDÁM (Institute of Medical Microbiology, Semmelweis University, Budapest, Hungary)
  • Prof. Sebastian AMYES (Department of Medical Microbiology, University of Edinburgh, Edinburgh, UK.)
  • Dr. Katalin BURIÁN (Institute of Clinical Microbiology University of Szeged, Szeged, Hungary; Department of Medical Microbiology and Immunobiology, University of Szeged, Szeged, Hungary.)
  • Dr. Orsolya DOBAY (Institute of Medical Microbiology, Semmelweis University, Budapest, Hungary)
  • Prof. Ildikó Rita DUNAY (Institute of Inflammation and Neurodegeneration, Medical Faculty, Otto-von-Guericke University, Magdeburg, Germany; Center for Behavioral Brain Sciences (CBBS), Magdeburg, Germany)
  • Prof. Levente EMŐDY(Department of Medical Microbiology and Immunology, University of Pécs, Pécs, Hungary.)
  • Prof. Anna ERDEI (Department of Immunology, Eötvös Loránd University, Budapest, Hungary, MTA-ELTE Immunology Research Group, Eötvös Loránd University, Budapest, Hungary.)
  • Prof. Éva Mária FENYŐ (Division of Medical Microbiology, University of Lund, Lund, Sweden)
  • Prof. László FODOR (Department of Microbiology and Infectious Diseases, University of Veterinary Medicine, Budapest, Hungary)
  • Prof. József KÓNYA (Department of Medical Microbiology, University of Debrecen, Debrecen, Hungary)
  • Prof. Yvette MÁNDI (Department of Medical Microbiology and Immunobiology, University of Szeged, Szeged, Hungary)
  • Prof. Károly MÁRIALIGETI (Department of Microbiology, Eötvös Loránd University, Budapest, Hungary)
  • Prof. János MINÁROVITS (Department of Oral Biology and Experimental Dental Research, University of Szeged, Szeged, Hungary)
  • Prof. Béla NAGY (Centre for Agricultural Research, Institute for Veterinary Medical Research, Budapest, Hungary.)
  • Prof. István NÁSZ (Institute of Medical Microbiology, Semmelweis University, Budapest, Hungary)
  • Prof. Kristóf NÉKÁM (Hospital of the Hospitaller Brothers in Buda, Budapest, Hungary.)
  • Dr. Eszter OSTORHÁZI (Institute of Medical Microbiology, Semmelweis University, Budapest, Hungary)
  • Prof. Rozália PUSZTAI (Department of Medical Microbiology and Immunobiology, University of Szeged, Szeged, Hungary)
  • Prof. Peter L. RÁDY (Department of Dermatology, University of Texas, Houston, Texas, USA)
  • Prof. Éva RAJNAVÖLGYI (Department of Immunology, Faculty of Medicine, University of Debrecen, Debrecen, Hungary)
  • Prof. Ferenc ROZGONYI (Institute of Laboratory Medicine, Semmelweis University, Budapest, Hungary)
  • Prof. Zsuzsanna SCHAFF (2nd Department of Pathology, Semmelweis University, Budapest, Hungary)
  • Prof. Joseph G. SINKOVICS (The Cancer Institute, St. Joseph’s Hospital, Tampa, Florida, USA)
  • Prof. Júlia SZEKERES (Department of Medical Biology, University of Pécs, Pécs, Hungary.)
  • Prof. Mária TAKÁCS (National Reference Laboratory for Viral Zoonoses, National Public Health Center, Budapest, Hungary.)
  • Prof. Edit URBÁN (Department of Medical Microbiology and Immunology University of Pécs, Pécs, Hungary; Institute of Translational Medicine, University of Pécs, Pécs, Hungary.)

 

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2020  
Total Cites 662
WoS
Journal
Impact Factor
2,048
Rank by Immunology 145/162(Q4)
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Impact Factor 1,904
without
Journal Self Cites
5 Year 0,671
Impact Factor
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Rank by Journal  Immunology 146/174 (Q4)
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Citable 42
Items
Total 40
Articles
Total 2
Reviews
Scimago 28
H-index
Scimago 0,439
Journal Rank
Scimago Immunology and Microbiology (miscellaneous) Q4
Quartile Score Medicine (miscellaneous) Q3
Scopus 438/167=2,6
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Scopus General Immunology and Microbiology 31/45 (Q3)
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Scopus 0,760
SNIP
Days from  225
submission
to acceptance
Days from  118
acceptance
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Acceptance 19%
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2019  
Total Cites
WoS
485
Impact Factor 1,086
Impact Factor
without
Journal Self Cites
0,864
5 Year
Impact Factor
1,233
Immediacy
Index
0,286
Citable
Items
42
Total
Articles
40
Total
Reviews
2
Cited
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Citing
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7,7
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0,00059
Article Influence
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0,246
% Articles
in
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95,24
Normalized
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0,07317
Average
IF
Percentile
7,690
Scimago
H-index
27
Scimago
Journal Rank
0,352
Scopus
Scite Score
320/161=2
Scopus
Scite Score Rank
General Immunology and Microbiology 35/45 (Q4)
Scopus
SNIP
0,492
Acceptance
Rate
16%

 

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Acta Microbiologica et Immunologica Hungarica
Language English
Size A4
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1954
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2021 Volume 68
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Founder Magyar Tudományos Akadémia
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