Role of Fur and transferrin binding protein in Staphylococcus epidermidis during in vitro and in vivo biofilm formation

Abstract number: 1134_03_413

Massonet C., Van Eldere J.

Objectives:  

To determine whether GADPH (transferrin binding protein-TBP) and its regulator (Fur) play a role in the biofilm formation by Staphylococcus epidermidis.

Methods:  

A biofilm forming strain of S. epidermidis, isolated from a patient with proven catheter-related infection was used. RNA and DNA isolations were performed as described by S. Vandecasteele et al. (Biochem. Biophys. Res. Commun. 2002. 291:528–534). For In vitro studies bacteria were grown overnight in BHI or RPMI, without iron (fRPMI) and re-incubated in fRPMI or fRPMI with 1 mm FeCl₃ or fRPMI with apotransferrin (iron free; 20 mg/ml) or fRPMI with holotransferrin (iron- saturated; 20 mg/ml). For in vivo studies a subcutaneous rat model was used. Samples were quantified through real-time PCR described by S. Vandecasteele et al. (Biochem. Biophys. Res. Commun. 2002. 291:528–534).

Results:  

During the growth of the bacteria, the expression of fur and tbp for planktonic bacteria was higher in an iron depleted medium and lower in an iron rich medium. In vitro studies in an iron depleted medium show a statistically significant (2-way ANOVA, Bonferonni, N = 12) up regulation of fur expression in sessile bacteria in comparison with planktonic bacteria. In an iron rich environment there is no variation in the expression of fur between sessile and planktonic bacteria. In fRPMI with holotransferrin or apotransferrin the expression of fur for sessile bacteria in comparison with that of planktonic bacteria is significantly up regulated in fRPMI with holotransferrin and down regulated in medium with apotransferrin. The results obtained for tbp are similar.

In vivo, the expression of fur is down regulated in the beginning, followed by an increase of expression after one week. The expression of tbp is very low and stays constant over a period of two weeks. (Statistically significant: 1-way ANOVA, Bonferroni, N = 16).

Conclusion:  

According to in vitro results, the lack of iron has a greater impact on sessile bacteria than on planktonic bacteria. Holotransferrin and apotransferrin also affect the expression of fur and tbp in sessile bacteria, but further investigation is required. In vivo results show that fur plays a role in the late stages of the biofilm.