FIG. 4.

(A) Planktonic growth of S. epidermidis O-47, the hemB mutant (O-47hemB), and the complemented mutant (O-47hemB comp.) in TSB containing 10 μg/ml hexosaminidase DspB and in TSB containing 100 μg/ml trypsin (21). Bacteria were grown overnight in 24-well tissue culture plates in a shaker (120 rpm) at 37°C. In an independent experiment, the enzymes were added to the medium after overnight growth. This measure also led to the disintegration of cell clusters by DspB but not by trypsin (data not shown), giving further evidence that expression of PIA is essential for cluster formation by O-47hemB. (B) Semiquantitative detection of cell wall-associated PIA in S. epidermidis O-47, the hemB mutant, and the complemented mutant. Whereas PIA is detected in S. epidermidis O-47 and the complemented mutant at dilutions of up to 1:8 and 1:4, respectively, a much stronger signal was found in the hemB mutant at dilutions of up to 1:16. PIA-positive reference strain S. epidermidis 1457 served as a control.