Phenotypic and genotypic characterization of biofilm formation among Staphylococcus aureus isolates from clinical specimens, an Atomic Force Microscopic (AFM) study

Abstract

Introduction: Staphylococcus aureus is a major cause of nosocomial infections. Biofilm formation is an important factor for bacterial pathogenesis. Its mechanisms are complex and include of many genes depends on expression of icaADBC operon involved in the synthesis of a polysaccharide intercellular adhesion.

Aim: The aim of study was to investigate biofilm forming ability of Staphylococcus aureus strains by phenotypic and genotypic methods. Also Atomic Force microscope (AFM) was used to visualize biofilm formation.

Methods: 140 Isolates were collected from clinical specimens of patients in Milad Hospital, Tehran and diagnosed by biochemical tests. The ability of strains to produce slime was evaluated by CRA method. For diagnosing of bacterial EPS, Indian ink staining were used and finally biofilm surface of 3 isolates observed by AFM. The prevalence of icaA and icaD genes was determined by PCR.

Results: By CRA method 15% of samples considered as positive slime producers, 44.28% as intermediate and 40.71% indicative as negative slime producers. 118 staphylococcus aureus strains showed a distinct halo transparent zone but 22 strains showed no halo zone. AFM analysis of Slime positive isolates showed a distinct and complete biofilm formation. In slime negative strain, there was not observed biofilm. The prevalence of icaA, icaD genes was 44.2% and 10% of the isolates had both genes simultaneously.

Conclusion: There is a relationship between exopolysaccharide layer and biofilm formation of Staphylococcus aureus isolates. The presence of icaAD genes among isolates is not associated with in vitro formation of biofilm. AFM is a useful tool for observation of bacterial biofilm formation.

Keywords: AFM; Biofilm formation; EPS; Staphylococcus aureus; icaA; icaD.