Molecular typing of nosocomial Staphylococcus aureus strains associated to biofilm based on the coagulase and protein A gene polymorphisms

Document Type : Original Article


1 Department of Biology, Rasht Branch, Islamic Azad University, Rasht, Iran

2 Department of Biology, Faculty of Science, University of Guilan, Rasht, Iran

3 Young Researchers and Elite Club, East Tehran Branch, Islamic Azad University, Tehran, Iran


Objective(s): Staphylococcus aureus is an important bacterial pathogen responsible for a variety numbers of nosocomial and community acquired infections. Biofilm formation is regarded as an important factor in the establishment of S. aureus infection. The contribution of the genetic background of S. aureus to biofilm formation is poorly understood. The aim of the present work was to genotype S. aureus strains associated to biofilm based on the coagulase and protein A genes and to evaluate the association between the genetic background and the biofilm forming ability of clinical S. aureus isolates.
Materials and Methods: A total number of 100 S. aureus were isolated from nosocomial infections and biofilm formation capability was investigated using phenotypic assay and molecular detection of biofilm associated genes. The strains were genotyped based on coagulase (coa) and protein A (spa) gene polymorphisms using restriction fragments length polymorphism-polymerase chain reaction (RFLP-PCR).
Results: RFLP-PCR of coa gene generated two types and three subtypes. Amplification of spa gene resulted in two banding patterns and their restriction digestion generated three subtypes. The combined coa and spa RFLP patterns generated nine genotypes (G1-G9). The genotypes G4 and G1 were the most prevalent (32.1% and 24.3%, respectively).
Conclusion: High clonal diversity of S. aureus strains able to produce biofilm was observed. Biofilm formation correlates with the spa and coa clonal lineage in our population and testing for multiple gene polymorphisms could be employed for local epidemiologic purposes.


1.Naicker PR, Karayem K, Hoek KG, Harvey J, Wasserman E. Biofilm formation in invasive Staphylococcus aureus isolates is associated with the clonal lineage. Microb Pathog 2016; 90:41-49.
2.Cosgrove SE, Fowler VG. Management of methicillin-resistant Staphylococcus aureus bacteremia. Clin Infect Dis 2008; 46:S386-S393.
3.Kwiecinski J, Kahlmeter G, Jin T. Biofilm formation by Staphylococcus aureus isolates from skin and soft tissue infections. Currmicrobial 2015; 70:698-703.
4.Khoramian B, Jabalameli F, Niasari-Naslaji A, Taherikalani M, Emaneini M. Comparison of virulence factors and biofilm formation among Staphylococcus aureus strains isolated from human and bovine infections. Microb Pathog 2015; 88:73-77.
5.Arciola CR, Baldassarri L, Montanaro L. Presence of icaA and icaDGenes and slime production in a collection of Staphylococcal strains from catheter-associated infections. J Clin Microbial 2001; 39:2151-2156.
6.Khameneh B, Diab R, Ghazvini K, Fazly Bazzaz BS. Breakthroughs in bacterial resistance mechanisms and the potential ways to combat them. Microb Pathog 2016; 95:32-42.
7.Atshan SS, Shamsudin MN, Lung T, Than L, Sekawi Z, Ghaznavi-Rad E, Pei Pei C. Comparative characterisation of genotypically different clones of MRSA in the production of biofilms. J Biomed Biotecnol 2012; 2012:417247.
8.Cosandey A, Boss R, Luini M, Artursson K, Bardiau M, Breitenwieser F, et al. Staphylococcus aureus genotype B and other genotypes isolated from cow milk in European countries. J Dairy Sci 2016; 99:529-540.
9.Stranden A, Frei R, Widmer AF. Molecular typing of methicillin-resistant Staphylococcus aureus: can PCR replace pulsed-field gel electrophoresis?. J Clinlmicrobial 2003; 41:3181-3186.
10.Mahon CR, Lehman DC, Manuselis G. Text Book of Diagnostic Microbiology. 3th ed. Philadelphia, PA, USA: 2007.
11.Das S, Dash HR. Microbial Biotechnology-A Laboratory Manual for Bacterial Systems. Springer; 2014.p.1-5.
12.Freeman DJ, Falkiner FR, Keane, CT. New method for detecting slime production by coagulase negative staphylococci. J Clin Pathol 1989; 42:872–874.
13.Dhanawade NB, Kalorey DR, Srinivasan R, Barbuddhe SB, Kurkure NV. Detection of intercellular adhesion genes and biofilm production in Staphylococcus aureus isolated from bovine subclinical mastitis. Vet Res Commun 2010; 34:81-89.
14.Omar NY, Ali HA, Harfoush RA, El Khayat EH. Molecular typing of methicillin resistant staphylococcus aureus clinical isolates on the basis of protein A and coagulase gene polymorphisms. Int J Microbiol 2014; 2014:650328.
15.Vasudevan P, Nair MK, Annamalai T,Venkitanarayanan KS. Phenotypic and genotypic characterization of bovine mastitis isolates of Staphylococcus aureus for biofilm formation. Vet Microbiol 2003; 92:179-185.
16.Yazdani R, Oshaghi M, Havayi A, Pishva E, Salehi R,Sadeghizadeh M, et al. Detection of icaAD gene and biofilm formation in Staphylococcus aureus isolates from wound infections. Iran J Publ Health 2006; 35:25-28.
17.Hookey JV, Richardson JF and Cookson BD. Molecular typing of Staphylococcus aureus based on PCR restriction fragment length polymorphism and DNA sequence analysis of the coagulase gene. J Clin Microbiol 1998; 36:1083-1089.
18.Shopsin B, Gomez M, Waddington M, Riehman M, Kreiswirth BN. Use of coagulase gene (coa) repeat region nucleotide sequences for typing of methicillin-resistant Staphylococcus aureus strains. J Clin Microbiol 2000; 38:3453-3456.
19.Frenay HM, Theelen JP, Schouls LM, Vandenbroucke-Grauls CM, Verhoef J, Van Leeuwen WJ, et al. Discrimination of epidemic and nonepidemic methicillin-resistant Staphylococcus aureus strains on the basis of protein A gene polymorphism. J Clin Microbiol 1994; 32:846-847.
20.Janwithayanuchit I, Ngam-Ululert S, Paungmoung P, Rangsipanuratn W. Epidemiologic study of methicillin-resistant Staphylococcus aureus by coagulase gene polymorphism. Sci Asia 2006; 32: 127-132.
21.Saei HD, Ahmadi M. Discrimination of staphylococcus aureusisolates on the basis of gene coding protein A using PCR-restriction enzyme analysis. Comp Clin Path 2012; 21:645-652.
22.Mitani N, Koizumi A, Sano R, Masutani T, Murakawa K, Okamoto Y, et al. Molecular typing of methicillin-resistant Staphylococcus aureus by PCR-RFLP and its usefulness in an epidemiological study of an outbreak. Jpn J Infect Dis 2005; 58:250.