American Journal of Infectious Diseases and Microbiology
ISSN (Print): 2328-4056 ISSN (Online): 2328-4064 Website: http://www.sciepub.com/journal/ajidm Editor-in-chief: Maysaa El Sayed Zaki
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American Journal of Infectious Diseases and Microbiology. 2015, 3(3), 95-103
DOI: 10.12691/ajidm-3-3-2
Open AccessArticle

Characterization of Some Genotypic and Phenotypic Traits of Biofilm Producing Clinical Isolates of Methicillin Resistant Staphylococcus Epidermidis

Malaka Farouk El Moufti1, Manal Baddour1, Reem Abdel Hameed Harfoush1, and Hoda Mohamed Aly Owais1

1Department of Medical Microbiology and Immunology, Faculty of Medicine, Alexandria University, Egypt

Pub. Date: May 12, 2015

Cite this paper:
Malaka Farouk El Moufti, Manal Baddour, Reem Abdel Hameed Harfoush and Hoda Mohamed Aly Owais. Characterization of Some Genotypic and Phenotypic Traits of Biofilm Producing Clinical Isolates of Methicillin Resistant Staphylococcus Epidermidis. American Journal of Infectious Diseases and Microbiology. 2015; 3(3):95-103. doi: 10.12691/ajidm-3-3-2

Abstract

Staphylococcus epidermidis is a major commensal bacterium. Various strains of methicillin resistant S. epidermidis are capable of forming biofilms and it is found to be associated with many hospital-acquired infections. Bacterial biofilms, which are micro-colonies encased in extracellular polysaccharide material, mediated by gene products of the icaADBC operon, are the sources of many bacterial infections which is so difficult to respond to routine treatments. In this research, we investigated the biofilm forming capacity of a 100 methicillin resistant staphylococcus epidermidis isolates, isolated from different clinical specimens delivered to the Diagnostic Microbiology Laboratories and Surveillance Laboratory, Faculty of Medicine, Alexandria University, in relation to the icaADBC gene cluster. Also, the minimum inhibitory concentration (MIC) and minimum biofilm eradication concentration (MBEC) assays were used to evaluate the antibiotic sensitivity patterns of these MRSE isolates in their planktonic and biofilm phases to Vancomycin and Linezolid. The results showed that only 27 isolates (27%) produced detectable biofilm, and icaADBC gene was detected in only 5 of these isolates. Moreover, there was no statistical association between the presence of the gene and the biofilm status. All 27 biofilm producing isolates were susceptible to both Vancomycin and Linezolid in their planktonic state, but the MBEC values of Vancomycin were higher than those of Linezolid in almost all strains, with an agreement between both MBEC values in 15/27 (55.5%) of isolates and disagreement in 12/27 (44.5%) of isolates, and this was statistically significant (p<0.05). In conclusion, this study indicates that the presence of icaADBC gene is not always associated with in-vitro formation of biofilm. Although Vancomycin and Linezolid continued to be effective for planktonic MRSE infection, their sub-MIC concentrations can induce biofilm formation.

Keywords:
methicillin S.epidermidis biofilm planktonic vancomycin Linezolid icaADBC

Creative CommonsThis work is licensed under a Creative Commons Attribution 4.0 International License. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/

References:

[1]  Arciola, C.R., Gamberini, S., Campoccia, D., Visai, L., Speziale, P. and Baldassarri, L, “A multiplex PCR method for the detection of all five individual genes of ica locus in Staphylococcus epidermidis. A survey on 400 clinical isolates from prosthesis-associated infections”, J Biomed Mater Res A, 75(2). 408-813. 2005.
 
[2]  Von Eiff, C., Peters, G. and Heilmann, C, “Pathogenesis of infections due to coagulase negative staphylococci”, Lancet Infect Dis, 2(11). 677-685.2002.
 
[3]  Diekema, D.J., Pfaller, M.A., Schmitz, F.J., Smayevsky, J., Bell, J. and Jones, R.N, “Survey of infections due to Staphylococcus species: frequency of occurrence and antimicrobial susceptibility of isolates collected in the United States, Canada, Latin America, Europe, and the Western Pacific region for the SENTRY Antimicrobial Surveillance Program, 1997-1999”, Clin Infect Dis, 32. 114-132. 2001.
 
[4]  Casey, A.L., Lambert, P.A. and Elliott, T.S, “Staphylococci”, Int J Antimicrob Agents, 29. 23-32. 2007.
 
[5]  Tsubakishita, S., Kuwahara-Arai, K., Sasaki, T. and Hira,matsu, K, “Origin and molecular evolution of the determinant of methicillin resistance in staphylococci”, Antimicrob Agents Chemother, 54(10). 4352-2359. 2010.
 
[6]  Ahlstrand, E., Svensson, K., Persson, L., Tidefelt, U. and Söderquist, B, “Glycopeptide resistance in coagulase-negative staphylococci isolated in blood cultures from patients with hematological malignancies during three decades”, Eur J Clin Microbiol Infect Dis, 30 (11). 1349-1354.2011.
 
[7]  Yoon, Y.K., Kim, J.Y., Park, D.W., Sohn, J.W. and Kim, M.J, “Predictors of persistent methicillin-resistant Staphylococcus aureus bacteremia in patients treated with vancomycin”, J Antimicrob Chemother, 65.1015-1018. 2010.
 
[8]  Raad, I., Hanna, H., Jiang, Y., Dvorak, T., Reitzel, R., Chaiban, J., Sherertz, R. and Hachem, R, “Comparative activities of daptomycin, linezolid, and tigecycline against catheter-related methicillin resistant Staphylococcus aureus bacteremia isolates embedded in biofilm”, Antimicrob Agents Chemother, 51. 1656-1660. 2007.
 
[9]  Hidayat, L.K., Hsu, D.I., Quist, R., Shriner, K.A. and Wong-Beringer, A, “High dose vancomycin therapy for methicillin resistant Staphylococcus aureus infections: efficacy and toxicity”, Arch Intern Med,; 166.2138-2144. 2006.
 
[10]  Klingenberg, C., Aarag, E., Ronnestad, A., Sollid, J.E., Abrahamsen, T.G., Kjeldsen, G. and Flaegstad T, ” Coagulase-negative staphylococcal sepsis in neonates. Association between antibiotic resistance, biofilm formation and the host inflammatory response”, Pediatr Infect Dis J, 24. 817-822. 2005.
 
[11]  Karatan, E. and Watnick, P,”Signals, regulatory networks, and materials that build and break bacterial biofilms”, Microbiol Mol Biol Rev, 73. 310-347. 2009.
 
[12]  Vuong, C. and Otto, M, “Staphylococcus epidermidis infections”, Microbes Infect, 4(4).481-489. 2002.
 
[13]  Mathur, T., Singhal, S., Khan, S., Upadhyay, D.J., Fatima, T. and Rattan, A, “Detection of biofilm formation among the clinical isolates of staphylococci: an evaluation of three different screening methods”, Indian J Med Microbiol, 24. 25-29. 2006.
 
[14]  De Araujo, G.L., Coelho, L.R., De Carvalho, C.B., Maciel, R.M., Coronado, A.Z., Rozebaum, R., Carvalho, B.T.F., Figueiredo, A.M.S. and Teixeira, L. A, “Commensal isolates of methicillin-resistant Staphylococcus epidermidis are also well equipped to produce biofilm on polystyrene surfaces”, J Antimicrob Chemother, 57. 855-864. 2006.
 
[15]  Stewart, P.S. and Costerton, J.W, “Antibiotic resistance of bacteria in biofilms”, Lancet, 358(9276).135-138. 2001.
 
[16]  Ceri, H., Olson, M., Morck, D., Storey, D., Read, R., Buret, A. and Olson, B, “The MBEC assay system: multiple equivalent biofilms for antibiotic and biocide susceptibility testing”, Methods Enzymol, 337.377-385. 2001.
 
[17]  Eftekhar, F. and Mirmohamadi, Z, “Evaluation of biofilm production by Staphylococcus epidermidis isolates from nosocomial infections and skin of healthy volunteers”, Int J Medicine Med Sci, 1(10).438-441. 2009.
 
[18]  Clinical and Laboratory Standards Institute. Performance standards for Antimicrobial Susceptibility Testing; twenty-third informational supplement. CLSI document M100- S23Vol 31 No.1. Clinical and Laboratory Standards Institute, Wayne, PA, USA, 2013.
 
[19]  Clinical and Laboratory Standards Institute, “Performance standards for Antimicrobial Susceptibility Testing”, 21th informational supplement, CLSI document M100- S21Vol 31 No.1. Clinical and Laboratory Standards Institute, Wayne, PA, USA, 2011.
 
[20]  Schmitz, F.J., Steiert, M., Tichy, H.V., Hofmann, B., Verhoef, J., Heinz, H.P, “Typing of methicillin resistant Staphylococcus aureus isolates from Dusseldorf by six genotypic methods”, J Med Microbiol, 47. 341-351, 1998.
 
[21]  Eftekhar, F. and Speert, D.P, ‘Biofilm formation by persistent and nonpersistent isolates of Staphylococcus epidermidis from a neonatal intensive care unit”, J Hosp Infect, 71. 112-116. 2009.
 
[22]  Qu, Y., Daley, A.J., Istivan, T.S., Garland, S.M. and Deighton, M.A, “Antibiotic susceptibility of coagulase-negative staphylococci isolated from very low birth weight babies: comprehensive comparisons of bacteria at different stages of biofilm formation”, Ann Clin Microbiol Antimicrob, 9.16. 2010.
 
[23]  Nasr, R.A., Abu Shady, H.M. and Hussein, H.S, “Biofilm formation and presence of icaAD gene in clinical isolates of staphylococci, Egyp J Med Human Genet, 13. 269-74. 2012.
 
[24]  Abd El Hafez, M., Khalaf, N.G., El Ahmady, M., Abd El Aziz, A. and Hashim, A, “An outbreak of methicillin resistant Staphylococcus epidermidis among neonates in a hospital in Saudi Arabia”, J Infect Dev Ctries, 5(10).692-699. 2011.
 
[25]  Iorio, N.L., Caboclo, R.F., Azevedo, M.B., Barcellos, A.G., Neves, F.P., Domingues, R.M., Dos Santos, K.R.N, “Characteristics related to antimicrobial resistance and biofilm formation of widespread methicillin-resistant Staphylococcus epidermidis ST2 and ST23 lineages in Rioe Janeiro hospitals, Brazil”, Diagn Microbiol Infect Dis, 72. 32-40. 2012.
 
[26]  Hassan, A., Usman, J., Kaleem, F., Omair, M., Khalid, A., Iqbal, M, “Evaluation of different detection methods of biofilm formation in the clinical isolates”, Braz J Infect Dis, 15(4). 305-311. 2011.
 
[27]  Terki, I.K., Hassaine, H., Oufrid, S., Bellifa, S., Mhamedi, I., Lachachi, M., Timinouni, M, “Detection of icaA and icaD genes and biofilm formation in Staphylococcus spp.isolated from urinary catheters at the University Hospital of Tlemcen (Algeria)”, Afr J Microbiol Res, 7 (47). 5350-2357. 2013.
 
[28]  Seif El-Din, S.S., El-Rehewy, M.S., Ghazaly, M.M. and Abd-Elhamid, M.H, “Biofilm formation by blood stream staphylococcal isolates from febrile pediatric cancer patients at south Egypt cancer institute”, J Am Sci, 7(1). 674-686. 2011.
 
[29]  Arciola, C.R., Baldassarri, L. and Montanaro, L, “Presence of icaA and icaD genes and slime production in a collection of staphylococcal strains from catheter-associated infections” J Clin Microbiol, 39. 2151-2156. 2001.
 
[30]  Votava, M. and Woznicova, V, “Production of slime by staphylococcal isolates from blood cultures”, Cent Eur J Public Health, 8. 18-20. 2000.
 
[31]  Fluckiger, U., Ulrich, M., Steinhuber, A., Doring, G., Mack, D., Landmann, R., Goerke, C. and Wolz, C, “Biofilm formation, icaADBC transcription, and polysaccharide intercellular adhesin synthesis by staphylococci in a device-related infection model”, Infect Immun, 73. 1811-1819. 2005.
 
[32]  Arciola, C.R., Campoccia, D., Baldassarri, L., Donati, M.E., Pirini, V., Gamberini, S. and Montanaro, L, “Detection of biofilm formation in Staphylococcus epidermidis from implant infections. Comparison of a PCR-method that recognizes the presence of ica genes with two classic phenotypic methods”, J Biomed Mater Res A, 76. 425-430. 2006.
 
[33]  Chokr, A., Leterme, D., Watier, D. and Jabbouri, S, “Neither the presence of ica locus, nor in vitro-biofilm formation ability is a crucial parameter for some Staphylococcus epidermidis strains to maintain an infection in a guinea pig tissue cage model”, Microb Pathog, 42. 94-97. 2007.
 
[34]  Li, M., Wang, X., Gao, Q. and Lu, Y, “Molecular characterization of Staphylococcus epidermidis strains isolated from a teaching hospital in Shanghai, China”, J Med Microbiol, 58. 456-461. 2009.
 
[35]  Kloos, W.E. and Bannerman, T.L, “Update on clinical significance of coagulase-negative staphylococci”, Clin Microbiol Rev, 7. 117-140. 1994.
 
[36]  Bradford, R., Abdul Manan, R. and Daley, A.J, “Coagulase-negative staphylococci in very-low-birth-weight infants: inability of genetic markers to distinguish invasive strains from blood culture contaminants”, Eur J Clin Microbiol Infect Dis, 25, 283-290. 2006.
 
[37]  Li, H., Xu, L., Wang, J., Wen, Y., Vuong, C., Otto, M. and Gao, Q, “ Conversion of Staphylococcus epidermidis strains from commensal to invasive by expression of the ica locus encoding production of biofilm exopolysaccharide”, Infect Immun, 73. 3188-3191. 2005.
 
[38]  Nemati, M., Hermans, K., Devriese, L.A., Maes, D. and Haesebrouck, F, “Screening of genes encoding adhesion factors and biofilm formation in Staphylococcus aureus isolates from poultry”, “Avian Pathol” 38.513-517. 2009.
 
[39]  Francois, P., Tu Quoc, P.H., Bisognano, C., Kelley, W.L., Lew, D.P., Schrenzel, J., Cramton, S.E., Götz, F., and Vaudaux, P, “Lack of biofilm contribution to bacterial colonisation in an experimental model of foreign body infection by Staphylococcus aureus and Staphylococcus epidermidis”, FEMS Immunol Med Microbiol, 35. 135-140. 2003.
 
[40]  Yazdani, R., Oshaghi, M., Havayi, A., Pishva, E., Salehi, R. and Sadeghizadeh, M, “Detection of icaAD gene and biofilm formation in Staphylococcus aureus isolates from wound infections” Iranian J Publ Health, 35(2).25-28. 2006.
 
[41]  De Silva, G.D., Kantzanou, M., Justice, A., Massey, R.C., Wilkinson, A.R., Day, N.P. and. Peacock, S. J, “The ica operon and biofilm production in coagulase-negative Staphylococci associated with carriage and disease in a neonatal intensive care unit”, J Clin Microbiol, 40. 382-388. 2002.
 
[42]  Gad, G.F., El-Feky, M.A., El-Rehewy, M.S., Hassan, M.A., Abolella, H. and El-Baky, R.M, “Detection of icaA, icaD genes and biofilm production by Staphylococcus aureus and Staphylococcus epidermidis isolated from urinary tract catheterized patients”, J Infect Dev Ctries, 3(5).342-51. 2009.
 
[43]  Rohde, H., Burandt, E.C. and Siemssen, N, “Polysaccharide intercellular adhesin or protein factors in biofilm accumulation of Staphylococcus epidermidis and Staphylococcus aureus isolated from prosthetic hip and knee joint infections”, Biomaterials, 28. 1711-1720. 2007.
 
[44]  Stapper, A.P., Narasimhan, G., Ohman, D.E., Barakat, J., Hentzer, M., Molin, S., Kharazmi, A., Høiby, N. and Mathee, K, “Alginate production affects Pseudomonas aeruginosa biofilm development and architecture, but is not essential for biofilm formation”, J Med Microbiol, 53. 679-690. 2004.
 
[45]  Conrady, D.G., Brescia, C.C., Horii, K., Weiss, A.A., Hassett, D.J., Herr, A.B, “A zinc-dependent adhesion module is responsible for intercellular adhesion in staphylococcal biofilms”, Proc Natl Acad Sci USA, 105. 19456-19461. 2008.
 
[46]  Ninin, E., Caroff, N., Espaze, E., Maraillac, J., Lepelletier, D., Milpied, N. and Richet, H, “Assessment of ica operon carriage and biofilm production in Staphylococcus epidermidis isolates causing bacteraemia in bone marrow transplant recipients”, CMI, 12. 446-452. 2006.
 
[47]  Hennig, S., Wai, S.N. and Zibuhr, W, “Spontaneous switch to PIA-independent biofilm formation in an ica-positive Staphylococcus epidermidis isolate”, Int J Med Microbiol, 297: 117-122. 2007.
 
[48]  Dobinsky, S., Kiel, K., Rohde, H., Bartscht, K., Knobloch, J.K.M. and Horstkotte, M.A, “Glucose-related dissociation between icaADBC transcription and biofilm expression by Staphylococcus epidermidis: evidence for an additional factor required for polysaccharide intercellular adhesin synthesis”, J Bacteriol, 185. 2879-2886. 2003.
 
[49]  Ceri, H., Olson, M.E., Stremick, C., Read, R.R., Morck, D. and Buret, A, “The Calgary Biofilm Device: New technology for rapid determination of antibiotic susceptibilities in bacterial biofilms”, J Clin Microbiol, 37. 1771-1776. 2001.
 
[50]  Antunes, A.L., Bonfanti, J.W., Perez, L.R., Pinto, C.C., Freitas, A.L., Macedo, A.J. and Barth, A.L, “High vancomycin resistance among biofilms produced by Staphylococcus species isolated from central venous catheters”, Mem Inst Oswaldo Cruz, 106(1).51-55. 2011.
 
[51]  El-Sheikh, N.A., Ayoub, A.M., El-Hendawy, H.H., Abada, E.A. and Khalifa, S.Y, “In-vitro Activity of some Antimicrobial Agents against Intact and Disrupted Biofilms of Staphylococci in the indwelling vascular catheter patients”, World Appl Sci J, 108-120. 2010.
 
[52]  Lee, J.Y., Ko, K.S., Peck, K.R., Oh, W.S. and Song, J.H, “In-vitro evaluation of the antibiotic lock technique (ALT) for the treatment of catheter-related infections caused by staphylococci”, J Antimicrob Chemother “57. 1110-1115. 2006.
 
[53]  Frank, K.L., Reichert, E.J., Piper, K.E. and Patel, R, In-vitro effects of antimicrobial agents on planktonic and biofilm forms of Staphylococcus lugdunensis clinical isolates”, Antimicrob Agents Chemother, 51 (3). 888-895. 2007.
 
[54]  Wilcox, M.H., Kite, P., Mills, K. and Sudden, S, “Ampicillin, trimethoprim-sulfamethoxazole and In situ measurement of linezolid and vancomycin gentamicin as treatments of catheter-associated concentrations in intravascular catheter-associated urinary tract infections in a rabbit model”, Int J biofilm J Antimicrob Chemother, 47. 171-175. 2001.
 
[55]  Curtin, J., Cormican, M., Fleming, G., Keelehan, J. and Colleran, E, “Linezolid compared with Eperezolid, Vancomycin, and Gentamicin in an in-vitro model of antimicrobial lock therapy for Staphylococcus epidermidis central venous catheter-related biofilm infections”, Antimicrob Agents Chemother, 47 (10). 3145-3148. 2003.
 
[56]  Uckay, I., Pittet, D., Vaudaux, P., Sax, H., Lew, D. and Waldvogel, F, “Foreign body infections due to Staphylococcus epidermidis”, Ann Med, 41.109-119. 2009.