Molecular Detection of Virulence Factors of Enterococcus Faecalis Isolated From Urine Samples in Duhok City, Kurdistan Region/Iraq.
Keywords:
Enterococcus faecalis, Urinary tract infection, PCR, Virulence genesAbstract
Enterococcus faecalis is one of the leading causes of many infections and mainly urinary tract infections. This pathogen developed high resistance to multiple antibiotics and it harbor many virulence factors genes. This study aimed to determine the antibiotic resistance patterns and screening for some virulence factor genes of E. faecalis isolated from urinary tract infection. Urine samples were collected from 788 outpatient’s clinic having clinical signs of UTI that visited Azadi Teaching Hospital in Duhok city. Urine samples were cultured on bacteriological media and isolated colonies identified using standard bacteriological methods. Antibiotic susceptibility was performed by Kirby Bauer test. All isolates were subjected to species-specific PCR assay for confirmatory identification followed by targeting virulence genes. Twenty five isolates of E. faecalis were detected and confirmed by species-specific PCR assay that expressed high antibiotic-resistance to many selected drugs except norfloxacin, penicillin and ampicillin. The most prevalent genes among all isolates were cpd genes followed by asa1, ace, esp, and gelE. Bearing of virulence genes combination were more frequent among multiple-antibiotic resistant strains. This study highlighted on E. faecalis as causes of UTI in Duhok city that showed multiple resistances to common antibiotics and harboring more than one virulence gene.
References
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Kariyama, R.; Mitsuhata, R.; Chow, J.W.; Clewell, D.B. and Kumon, H. (2000). Simple and reliable multiplex PCR assay for surveillance isolates of vancomycin-resistant enterococci. J. Clin. Microbiol., 38(8):3092-5.
Klibi, N.; Ben Slama, K.; Saenz, Y.; Masmoudi, A.; Zanetti, S.; Sechi, L.A.; Boudabous, A. and Torres, C. (2007). Detection of virulence factors in high-level gentamicin-resistant Enterococcus faecalis and Enterococcus faecium isolates from a Tunisian hospital. Can. J. Microbiol., 53(3):372–379.
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Lindenstrau, A.G.; Pavlovic, M.; Bringmann, A.; Behr, J.; Ehrmann, M.A. and Vogel, R.F. (2011). Comparison of genotypic and phenotypic cluster analyses of virulence determinants and possible role of CRISPR elements towards their incidence in Enterococcus faecalis and Enterococcus faecium. Syst. Appl. Microbiol., 34(8): 553-560.
Manero, A. and Blanch, A.R. (1999). Identification of Enterococcus spp. with a biochemical key. Appl .Environ. Microbiol., 65(10):4425-30.
Medeiros, A.W; Pereira, R.I.; Oliveira, D.V.; Martins, P.D.; d'Azevedo, P.A.; Van der Sand, S.; Frazzon, J. and Frazzon, A.P. (2014). Molecular detection of virulence factors among food and clinical Enterococcus faecalis strains in South Brazil. Braz. J. Microbiol., 45(1):327-32.
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Murray, B.E. (1990). The life and times of the Enterococcus. Clin.Microbiol., Rev., 3(1):46-65.
Salah, R.; Dar-Odeh, N.; Abu Hammad, O. andShehabi, A.A. (2008). Prevalence of putative virulence factors and antimicrobial susceptibility of Enterococcus faecalis isolates from patients with dental diseases. BMC. Oral Health, 8(17) :1-7
Schlievert, P.M.; Chuang-Smith, O.N.; Peterson, M.L.; Cook, L.C. and Dunny, G.M. (2010). Enterococcus faecalis endocarditis severity in rabbits is reduced by IgG Fabs interfering with aggregation substance. PLoS One, 5(10): e13194.
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Semedo, T., Almeida Santos, M.; Martins, P.; Silva Lopes, M.F.; Figueiredo Marques, J.J.; Tenreiro, R. and Barreto Crespo, M.T. (2003). Comparative study using type strains and clinical and food isolates to examine hemolytic activity and occurrence of the cyl operon in enterococci. J ClinMicrobiol., 41(6):2569-2576.
Shankar, N.; Lockatell, C.V.; Baghdayan, A.S.; Drachenberg, C.; Gilmore, M.S. and Johnson, D.E. (2001). Role of Enterococcus faecalis surface protein Esp in the pathogenesis of ascending urinary tract infection. Infect. Immun., 69(7) 4366-72.
Sharifi, Y.; Hasani, A.; Ghotaslou, R.; Naghili, B.; Aghazadeh, M.; Milani, M. and Bazmani, A. (2013). Virulence and antimicrobial resistance in enterococci isolated from urinary tract infections. Adv. Pharm. Bull., 3(1):197-201.
Sharifi, Y.; Hasani, A.; Ghotaslou, R.; Varshochi, M.; Hasani, A.; Aghaza¬deh, M. and Milani, M.(2012). Survey of virulence determinants among vancomycin resistant Enterococcus faecalis and Enterococcus faecium isolated from clinical specimens of hospitalized patients of northwest of Iran. Open Microbiol. J. 6: 34-39.
Singh, K.V.; Nallapareddy, S.R.; Sillanpää, J.; Murray, B.E. (2010). Importance of the collagen adhesin Ace in pathogenesis and protection against Enterococcus faecalis experimental endocarditis. PLo. S. Pathog., 6(1):e1000716
Thouverez, M. and Talon, D. (2004). Microbiological and epideralological studies of Enterococcus faecium resistant to amoxicillin a university hospital in eastern France. Clin. Microbiol. Infect., 10(5):441-7.
Toledo-Arana, A.; Valle, J.; Solano, C.; Arrizubieta, M. J.; Cucarella, C.; Lamata, M.; Amorena, B.; Leiva, J.; Penades, J. R. and Lasa, I. (2001). The Enterococcal surface protein, Esp, is involved in Enterococcus faecalis biofilm formation. Appl. Environ. Microbiol., 67(10):4538–4545.
Vankerckhoven, V.; Van Autgaerden, T.; Vael, C.; Lammens, C.; Chapelle, S.; Rossi, R.; Jabes, D. and Goossens, H. (2004). Development of a multiplex PCR for the detection of asa1, gelE, cylA, esp and hyl genes in enterococci and survey for virulence determinants among European hospital isolates of Enterococcus faecium. J. Clin. Microbiol., 42(10): 4473–4479.
Waar, K.; Muscholl-Silberhorn, A.B.; Willems, R.J.; Slooff, M.J.; Harmsen, H.J. and Degener, J.E. (2002).Genogrouping and incidence of virulence factors of Enterococcus faecalis in liver transplant patients differ from blood culture and fecal isolates. J. Infect. Dis., 185(8):1121-1127.
Wierzchowska, W.C.; Zadernowska, A.; Nalepa, B. and Laniewska, T.L. (2012). Occurrence and antibiotic resistance of enterococci in ready-to-eat food of animal origin. Afr .J. Microbiol. Res., 6(39):6773-80.
Xia, Z.; Xichu, W.; Bingjiao, G. and Xiaolan, W. (2013). Isolation and identification of Enterococcus faecalis and detection of its virulence factor genes in lambs presenting with encephalitis in Xinjiang Province, China. Afr. J. Microbiol. Res., 7(20):2238–2244.
Yazgi, H.; Ertek, M.; Erol, S.; and Ayyildiz, A. (2002). A comparison of high-level aminoglycoside resistance in vancomycin-sensitive and vancomycin-resistant Enterococcus species. J. Int. Med. Res., 30(5): 529-534.
Al-Zu'bi, E.; Bdour, S. and Shehabi, A.A. (2004). Antibiotic resistance patterns of mecA-positive Staphylococcus aureus isolates from clinical specimens and nasal carriage. Microb Drug Resist, 10(4):321-324.
Archimbaud, C.; Shankar, N.; Forestier, C.; Baghdayan, A.; Gilmore, M.S.; Charbonné, F.; and Joly, B. (2002). In vitro adhesive properties and virulence factors of Enterococcus faecalis strains. Res. Microbiol., 153(2):75-80.
Arularasi Aberna, R. and Prabakaran, K. (2011). Evaluation for the association of virulence determinants among E. faecalis with its clinical outcome. Int. J. Biol. Med. Res., 2(2):523-527.
Ballering, K.S.; Kristich, C.J.; Grindle, S.M.; Oromendia, A.; Beattie, D.T. and Dunny, G.M. (2009). Functional genomics of Enterococcus faecalis: multiple novel genetic determinants for biofilm formation in the core ge¬nome. J. Bacteriol., 191(8): 2806-2814.
Bhardwaj, S.; Bhamre, K.; Dhawale, J.; Patil, M. and Divase, S. (2013). Enterococcus faecium and Enterococcus faecalis, the nosocomial pathogens with special reference to multi-drug resistance and phenotypic characterization. International Journal of Pharmaceutical Science and Practice, 2(1):1-10.
Cariolato, D.; Andrighetto, C. and Lombardi, A. (2008). Occurrence of virulence factors and antibiotic resistances in Enterococcus faecalis and Enterococcus faecium collected from dairy and human samples in North Italy. Food Control, 19(9):886-892.
Chuang, O.N.; Schlievert, P.M.; Wells, C.L.; Manias, D.A.; Tripp, T.J. and Dunny, G.M. (2009). Multiple functional domains of Enterococcus faecalis aggrega¬tion substance Asc10 contribute to endocarditis virulence. Infect. Immun., 77(1): 539-548.
Chuang-Smith, O.N.; Wells, C.L.; Henry-Stanley, M.J. and Dunny, G.M. (2010). Acceleration of Enterococcus faecalis biofilm formation by ag-gregation substance expression in an ex vivo model of cardiac valve colonization. PLoS ONE, 5(12): e15798.
Clinical and Laboratory Standards Institute (2006). Performance standards for antimicrobial susceptibility testing; 16th informational supplement. M100-S16. Wayne, PA: Clinical and Laboratory Standards Institute.
Cosentino, S.; Podda, G.S.; Corda, A.; Fadda, M.E.; Deplano, M. and Pisano, M.B. (2010). Molecular detection of virulence factors and antibiotic resistance pattern in clinical Enterococcus faecalis strains in Sardinia. J Prev Med Hyg., 51(1):31-6.
Creti, R.; Imperi, M.; Bertuccini, L.; Fabretti, F.; Orefici, G.; Di Rosa, R. and Baldassarri, L. (2004). Survey for virulence determinants among Enterococcus faecalis isolated from different sources. J Med Microbiol, 53(Pt 1):13-20.
De Marques, E.B. and Suzart, S. (2004). Occurrence of virulence-associated genes in clinical Enterococcus faecalis strains isolated in Londrina. Brazil J Med Microbiol., 53(11):1069-1073.
Dupont, H.; Vael, C.; Muller-Serieys, C.; Chosidow, D.; Mantz, J., Marmuse, J.P.; Andremont, A.; Goossens, H. and Desmonts, J.M. (2008). Prospective evaluation of virulence factors of enterococci isolated from patients with peritonitis: Impact on outcome. DiagMicrobiol Infect. Dis., 60(3):247-253.
Eaton, T. J. and Gasson, M. J. (2001). Molecular screening of Enterococcus virulence determinants and potential for genetic exchange between food and medical isolates. Appl. Environ. Microbiol., 67(4):1628–1635.
Fontana, R.; Cerini, R.; Longoni, P.; Grossato, A. and Canepari P. (1983). Identification of a streptococcal penicillin-binding protein that reacts very slowly with penicillin. J. Bacteriol., 155:1343-50.
Kaçmaz, B. and Aksoy, A. (2005). Antimicrobial resistance of enterococci in Turkey. Int. J. Antimicrob. Agents, 25(6): 535-538.
Kariyama, R.; Mitsuhata, R.; Chow, J.W.; Clewell, D.B. and Kumon, H. (2000). Simple and reliable multiplex PCR assay for surveillance isolates of vancomycin-resistant enterococci. J. Clin. Microbiol., 38(8):3092-5.
Klibi, N.; Ben Slama, K.; Saenz, Y.; Masmoudi, A.; Zanetti, S.; Sechi, L.A.; Boudabous, A. and Torres, C. (2007). Detection of virulence factors in high-level gentamicin-resistant Enterococcus faecalis and Enterococcus faecium isolates from a Tunisian hospital. Can. J. Microbiol., 53(3):372–379.
Lebreton, F.; Riboulet-Bisson, E.; Serror, P.; Sanguinetti M.; Posteraro, B.; Torelli, R.; Hartke, A.; Auffray, Y. and Giard, J.C. (2009). ace, which encodes an adhesin in Enterococcus faecalis, is regulated by Ers and is involved in virulence. Infect. Immun., 77(7):2832–2839.
Lindenstrau, A.G.; Pavlovic, M.; Bringmann, A.; Behr, J.; Ehrmann, M.A. and Vogel, R.F. (2011). Comparison of genotypic and phenotypic cluster analyses of virulence determinants and possible role of CRISPR elements towards their incidence in Enterococcus faecalis and Enterococcus faecium. Syst. Appl. Microbiol., 34(8): 553-560.
Manero, A. and Blanch, A.R. (1999). Identification of Enterococcus spp. with a biochemical key. Appl .Environ. Microbiol., 65(10):4425-30.
Medeiros, A.W; Pereira, R.I.; Oliveira, D.V.; Martins, P.D.; d'Azevedo, P.A.; Van der Sand, S.; Frazzon, J. and Frazzon, A.P. (2014). Molecular detection of virulence factors among food and clinical Enterococcus faecalis strains in South Brazil. Braz. J. Microbiol., 45(1):327-32.
Mengeloğlu, F.Z.; Çakır, D. and Terzi, H.A. (2011). Comparison of resistance in isolates of Enterococcus faecalis and Enterococcus faecium. J. Clin.Microbiol. Infect. Dis., 1(1): 10-13.
Murray, B.E. (1990). The life and times of the Enterococcus. Clin.Microbiol., Rev., 3(1):46-65.
Salah, R.; Dar-Odeh, N.; Abu Hammad, O. andShehabi, A.A. (2008). Prevalence of putative virulence factors and antimicrobial susceptibility of Enterococcus faecalis isolates from patients with dental diseases. BMC. Oral Health, 8(17) :1-7
Schlievert, P.M.; Chuang-Smith, O.N.; Peterson, M.L.; Cook, L.C. and Dunny, G.M. (2010). Enterococcus faecalis endocarditis severity in rabbits is reduced by IgG Fabs interfering with aggregation substance. PLoS One, 5(10): e13194.
Schouten, M.A.; Hoogkamp-Korstanje, J.A.; Meis, J.F. and Voss, A. (2000). Prevalence of vancomycin-resistant enterococci in Europe. Eur. J. Clin. Microbiol. Infect. Dis., 19(11): 816-822.
Semedo, T., Almeida Santos, M.; Martins, P.; Silva Lopes, M.F.; Figueiredo Marques, J.J.; Tenreiro, R. and Barreto Crespo, M.T. (2003). Comparative study using type strains and clinical and food isolates to examine hemolytic activity and occurrence of the cyl operon in enterococci. J ClinMicrobiol., 41(6):2569-2576.
Shankar, N.; Lockatell, C.V.; Baghdayan, A.S.; Drachenberg, C.; Gilmore, M.S. and Johnson, D.E. (2001). Role of Enterococcus faecalis surface protein Esp in the pathogenesis of ascending urinary tract infection. Infect. Immun., 69(7) 4366-72.
Sharifi, Y.; Hasani, A.; Ghotaslou, R.; Naghili, B.; Aghazadeh, M.; Milani, M. and Bazmani, A. (2013). Virulence and antimicrobial resistance in enterococci isolated from urinary tract infections. Adv. Pharm. Bull., 3(1):197-201.
Sharifi, Y.; Hasani, A.; Ghotaslou, R.; Varshochi, M.; Hasani, A.; Aghaza¬deh, M. and Milani, M.(2012). Survey of virulence determinants among vancomycin resistant Enterococcus faecalis and Enterococcus faecium isolated from clinical specimens of hospitalized patients of northwest of Iran. Open Microbiol. J. 6: 34-39.
Singh, K.V.; Nallapareddy, S.R.; Sillanpää, J.; Murray, B.E. (2010). Importance of the collagen adhesin Ace in pathogenesis and protection against Enterococcus faecalis experimental endocarditis. PLo. S. Pathog., 6(1):e1000716
Thouverez, M. and Talon, D. (2004). Microbiological and epideralological studies of Enterococcus faecium resistant to amoxicillin a university hospital in eastern France. Clin. Microbiol. Infect., 10(5):441-7.
Toledo-Arana, A.; Valle, J.; Solano, C.; Arrizubieta, M. J.; Cucarella, C.; Lamata, M.; Amorena, B.; Leiva, J.; Penades, J. R. and Lasa, I. (2001). The Enterococcal surface protein, Esp, is involved in Enterococcus faecalis biofilm formation. Appl. Environ. Microbiol., 67(10):4538–4545.
Vankerckhoven, V.; Van Autgaerden, T.; Vael, C.; Lammens, C.; Chapelle, S.; Rossi, R.; Jabes, D. and Goossens, H. (2004). Development of a multiplex PCR for the detection of asa1, gelE, cylA, esp and hyl genes in enterococci and survey for virulence determinants among European hospital isolates of Enterococcus faecium. J. Clin. Microbiol., 42(10): 4473–4479.
Waar, K.; Muscholl-Silberhorn, A.B.; Willems, R.J.; Slooff, M.J.; Harmsen, H.J. and Degener, J.E. (2002).Genogrouping and incidence of virulence factors of Enterococcus faecalis in liver transplant patients differ from blood culture and fecal isolates. J. Infect. Dis., 185(8):1121-1127.
Wierzchowska, W.C.; Zadernowska, A.; Nalepa, B. and Laniewska, T.L. (2012). Occurrence and antibiotic resistance of enterococci in ready-to-eat food of animal origin. Afr .J. Microbiol. Res., 6(39):6773-80.
Xia, Z.; Xichu, W.; Bingjiao, G. and Xiaolan, W. (2013). Isolation and identification of Enterococcus faecalis and detection of its virulence factor genes in lambs presenting with encephalitis in Xinjiang Province, China. Afr. J. Microbiol. Res., 7(20):2238–2244.
Yazgi, H.; Ertek, M.; Erol, S.; and Ayyildiz, A. (2002). A comparison of high-level aminoglycoside resistance in vancomycin-sensitive and vancomycin-resistant Enterococcus species. J. Int. Med. Res., 30(5): 529-534.
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2016-06-30
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Khalid, H. M. (2016). Molecular Detection of Virulence Factors of Enterococcus Faecalis Isolated From Urine Samples in Duhok City, Kurdistan Region/Iraq. Science Journal of University of Zakho, 4(1), 63–72. Retrieved from https://sjuoz.uoz.edu.krd/index.php/sjuoz/article/view/306
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