Does biofilm formation have different pathways in Staphylococcus aureus?

Document Type : Original Article


Microbial Biotechnology Research Center, Department of Microbiology, School of Medicine, Iran University of Medical Sciences, Tehran, Iran


Objective(s): Biofilm formation is one of the most important factors in the development of infections caused by Staphylococcus aureus. In this study, the expression levels of genes responsible for biofilm formation were studied in methicillin sensitive and methicillin resistant S. aureus.
Materials and Methods: A total of 100 meticillin-resistant s.aureus (MRSA) and meticillin-sensetive s.aureus (MSSA) isolates were studied. Bacterial biofilm formation was evaluated phenotypically using microtiter plate method. Real-time PCR tests were conducted to determine the expression levels of genes involved in biofilm formation.
Results: Quantitative biofilm formation test was repeated three times for each specimen. The prevalence of weak, medium, and strong biofilm producers were 16%, 49%, and 35%, respectively. In MSSA isolates, expression levels of ica genes increased compared to the fnbA, fnbB, clfA and clfB genes. These results were different in MRSA isolates, and ica genes showed a decreased gene expression levels compared to the aforementioned genes.  
Conclusion: Considering the results of this study, clf genes probably contribute to the same extent in both MRSA and MSSA isolates, and there is probably no significant difference in the role of these genes in these isolates. In addition, the results of this study indicated that MRSA may not use the conventional route for biofilm formation and may use independent pathways through Polysaccharide intercellular adhesion (PIA).


Main Subjects

1. Lowy FD. Staphylococcus aureus infections. New England J Med 1998; 339:520-532.
2. Chambers HF, DeLeo FR. Waves of resistance: Staphylococcus aureus in the antibiotic era. Nat Rev Microbiol 2009; 7:629.
3. Agarwal A, Singh KP, Jain A. Medical significance and management of Staphylococcal biofilm. FEMS Immunol Med Microbiol 2010; 58:147-160.
4. Maki DG, Kluger DM, Crnich CJ, editors. The risk of bloodstream infection in adults with different intravascular devices: a systematic review of 200 published prospective studies. Mayo Clin Proc Innov; 2006: Elsevier.
5. Periasamy S, Joo HS, Duong AC, Bach T-HL, Tan VY, Chatterjee SS, et al. How Staphylococcus aureus biofilms develop their characteristic structure. Proc Natl Acad Sci 2012; 109:1281-1286.
6. Otto M. Looking toward basic science for potential drug discovery targets against community‐associated MRSA. Med Res Rev 2010; 30:1-22.
7. Alp E, Klaassen CH, Doganay M, Altoparlak U, Aydin K, Engin A, et al. MRSA genotypes in Turkey: persistence over 10 years of a single clone of ST239. J Infect 2009; 58:433-438.
8. Evans ME, Kralovic SM, Simbartl LA, Freyberg RW, Obrosky DS, Roselle GA, et al. Nationwide reduction of health care–associated methicillin-resistant Staphylococcus aureus infections in Veterans Affairs long-term care facilities. Am J Infect Control 2014; 42:60-62.
9. Kardaś-Słoma L, Boëlle P, Opatowski L, Brun-Buisson C, Guillemot D, Temime L. Impact of antibiotic exposure patterns on the selection of community-associated MRSA in hospital settings. Antimicrob Agents Chemother 2011:AAC. 01626-01610.
10. Tenover FC, McDougal LK, Goering RV, Killgore G, Projan SJ, Patel JB, et al. Characterization of a isolate of community-associated methicillin-resistant Staphylococcus aureus widely disseminated in the United States. J Clin Microbiol 2006; 44:108-118.
11. Donlan RM. Biofilms and device-associated infections. Emerg Infect Dis 2001; 7:277.
12. Ghaznavi-Rad E, Shamsudin MN, Sekawi Z, Khoon LY, Aziz MN, Hamat RA, et al. Predominance and emergence of clones of hospital-acquired methicillin-resistant Staphylococcus aureus in Malaysia. J Clin Microbiol 2010; 48:867-872.
13. Archer NK, Mazaitis MJ, Costerton JW, Leid JG, Powers ME, Shirtliff ME. Staphylococcus aureus biofilms: Properties, regulation and roles in human disease. Virulence 2011; 2: 445–459.
14. Tormo MA, Knecht E, Götz F, Lasa I, Penades JR. Bap-dependent biofilm formation by pathogenic species of Staphylococcus: evidence of horizontal gene transfer? Microbiology 2005; 151:2465-2475.
15. Liberto MC, Matera G, Quirino A, Lamberti AG, Capicotto R, Puccio R, et al. Phenotypic and genotypic evaluation of slime production by conventional and molecular microbiological techniques. Microbiol Res 2009; 164:522-528.
16. Rohde H, Knobloch JK, Horstkotte MA, Mack D. Correlation of Staphylococcus aureus icaADBCgenotype and biofilm expression phenotype. J Clin Microbiol 2001; 39:4595-4596.
17. Rohde H, Frankenberger S, Zähringer U, Mack D. Structure, function and contribution of polysaccharide intercellular adhesin (PIA) to Staphylococcus epidermidis biofilm formation and pathogenesis of biomaterial-associated infections. Eur J Cell Biol 2010; 89:103-111.
18. Cucarella C, Solano C, Valle J, Amorena B, Lasa Í, Penadés JR. Bap, a Staphylococcus aureus surface protein involved in biofilm formation. J Bacteriol 2001; 183:2888-2896.
19. Beenken KE, Dunman PM, McAleese F, Macapagal D, Murphy E, Projan SJ, et al. Global gene expression in Staphylococcus aureus biofilms. J Bacteriol 2004; 186:4665-4684.
20. Houston P, Rowe SE, Pozzi C, Waters EM, O’Gara JP. Essential role for the major autolysin in the fibronectin-binding protein-mediated Staphylococcus aureus biofilm phenotype. Infect Immun 2011; 79:1153-1165.
21. Seno Y, Kariyama R, Mitsuhata R, Monden K, Kumon H. Clinical implications of biofilm formation by Enterococcus faecalis in the urinary tract. Acta Med Okayama 2005; 59:79-87.
22. Ando E, Monden K, Mitsuhata R, Kariyama R, Kumon H. Biofilm formation among methicillin-resistant Staphylococcus aureus isolates from patients with urinary tract infection. Acta Med Okayama 2004; 58:207-214.
23. Atshan SS, Shamsudin MN, Karunanidhi A, van Belkum A, Lung LTT, Sekawi Z, et al. Quantitative PCR analysis of genes expressed during biofilm development of methicillin resistant Staphylococcus aureus (MRSA). Infection, Genetics and Evolution 2013; 18:106-112.
24. Shrestha L, Kayama S, Sasaki M, Kato F, Hisatsune J, Tsuruda K, et al. Inhibitory effects of antibiofilm compound 1 against Staphylococcus aureus biofilms. Microbiology and immunology 2016; 60:148-159.
25. Moghadam SO, Pourmand MR, Aminharati F. Biofilm formation and antimicrobial resistance in methicillin-resistant Staphylococcus aureus isolated from burn patients, Iran.  J  Infect Dev Ctries 2014; 8:1511-1517.
26. Vasudevan P, Nair MKM, Annamalai T, Venkitanarayanan KS. Phenotypic and genotypic characterization of bovine mastitis isolates of Staphylococcus aureus for biofilm formation. Veterinary microbiology 2003; 92:179-185.
27. Rohde H, Burandt EC, Siemssen N, Frommelt L, Burdelski C, Wurster S, et al. Polysaccharide intercellular adhesin or protein factors in biofilm accumulation of Staphylococcus epidermidis and Staphylococcus aureus isolated from prosthetic hip and knee joint infections. Biomaterials 2007; 28:1711-1720.
28. Oliveira A, Maria de Lourdes R. Comparison of methods for the detection of biofilm production in coagulase-negative staphylococci. BMC research notes 2010; 3:260.
29. Fowler Jr VG, Fey PD, Reller LB, Chamis AL, Corey GR, Rupp ME. The intercellular adhesin locus ica is present in clinical isolates of Staphylococcus aureus from bacteremic patients with infected and uninfected prosthetic joints. Medical microbiology and immunology 2001; 189:127-131.
30. Knobloch JK-M, Horstkotte MA, Rohde H, Mack D. Evaluation of different detection methods of biofilm formation in Staphylococcus aureus. Medical microbiology and immunology 2002; 191:101-106.
31. Kwon AS, Park GC, Ryu SY, Lim DH, Lim DY, Choi CH, et al. Higher biofilm formation in multidrug-resistant clinical isolates of Staphylococcus aureus. Int J Antimicrob Agents 2008; 32:68-72.
32. Boles BR, Thoendel M, Roth AJ, Horswill AR. Identification of genes involved in polysaccharide-independent Staphylococcus aureus biofilm formation. PloS one 2010; 5:e10146.
33. O’Neill E, Pozzi C, Houston P, Smyth D, Humphreys H, Robinson DA, et al. Association between methicillin susceptibility and biofilm regulation in Staphylococcus aureus isolates from device-related infections. J Clin Microbiol 2007; 45:1379-1388.
34. O’Neill E, Pozzi C, Houston P, Humphreys H, Robinson DA, Loughman A, et al. A novel Staphylococcus aureus biofilm phenotype mediated by the fibronectin-binding proteins, FnBPA and FnBPB. Journal of bacteriology 2008; 190:3835-3850.
35. Shanks RM, Meehl MA, Brothers KM, Martinez RM, Donegan NP, Graber ML, et al. Genetic evidence for an alternative citrate-dependent biofilm formation pathway in Staphylococcus aureus that is dependent on fibronectin binding proteins and the GraRS two-component regulatory system. Infection and immunity 2008; 76:2469-2477.
36. Dastgheyb S, Parvizi J, Shapiro IM, Hickok NJ, Otto M. Effect of biofilms on recalcitrance of Staphylococcal joint infection to antibiotic treatment. J Infec Dis 2014; 211:641-650.