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Case Reports
. 2008 Aug;90(8):1751-8.
doi: 10.2106/JBJS.G.00838.

Direct demonstration of viable Staphylococcus aureus biofilms in an infected total joint arthroplasty. A case report

Paul Stoodley  1 , Laura Nistico, Sandra Johnson, Leslie-Ann Lasko, Mark Baratz, Vikram Gahlot, Garth D Ehrlich, Sandeep Kathju
Affiliations
Case Reports

Direct demonstration of viable Staphylococcus aureus biofilms in an infected total joint arthroplasty. A case report

Paul Stoodley et al. J Bone Joint Surg Am. 2008 Aug.
No abstract available

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Figures

Fig. 1
Fig. 1
Radiographs documenting the total elbow arthroplasty before and after revision. A: Radiograph demonstrating nonunion of the distal part of the humerus and loose hardware. There is no evidence of periosteal reaction, which would suggest osteomyelitis. B: Lateral radiograph of elbow following total elbow arthroplasty with placement of an intercalary tibial allograft. C: Lateral radiograph of elbow thirteen months after the total elbow arthroplasty. An area of lucency (arrowheads) surrounds the ulnar component.
Fig. 2
Fig. 2
Large aggregate of viable (greenish-yellow) cocci in the aspirate stained with use of Molecular Probes LIVE/DEAD viability kit. The largest detected clumps were up to 100 μm in diameter and had a heterogeneous morphology consistent with that of “in vitro” grown Staphylococcus aureus biofilms. The aggregates may represent clumps of bacteria that had shed naturally from the biofilm, possibly contributing to systemic symptoms (e.g., fever). Sagittal sections through the clumps along the x and y axes are shown in the vertical planes, labeled “x-z” and “y-z.” The human nuclei (arrow) and associated tissue stained red. Scale: major divisions = 10 μm.
Fig. 3
Fig. 3
Single cells and clumps of viable cocci (stained green with use of Molecular Probes LIVE/DEAD viability kit) attached to tissue displaying characteristic staphylococcal morphology. Human tissue stained red with propidium iodide. A: Low-power image showing bacterial cells (arrow) attached to fibrous material (red striation). The nuclei of human cells were also visible (black arrow). Scale bar = 20 μm. B: Higher-power magnification of a group of cocci attached to fibrous material. Some cells were in the process of division (arrow), indicating that they were viable, which was consistent with the results of viability staining and culturing. Scale bar = 10 μm. C: Three-dimensional orthogonal projection of panel “A” showing that the biofilm clumps (arrow) were attached and protruding from the fibrous material. Scale: major divisions = 10 μm.
Fig. 4
Fig. 4
Viable Staphylococcus aureus biofilm cocci attached to a piece of bone cement that was removed during the surgical revision. A: Macroscopic view of the cement immersed in buffer in a 10-cm-diameter Petri plate. The specimen was oriented for subsequent confocal microscopic observation with use of a water-immersion objective. B: Microscopic three-dimensional orthogonal view showing the clumps of biofilm (greenish-yellow and indicated by arrows) attached to the surface of the bone cement (blue), as constructed from a confocal microscopy stack. Scale: major divisions = 10 μm.
Fig. 5
Fig. 5
Agarose gel electrophoresis of amplimers following polymerase chain reaction and reverse transcriptase-polymerase chain reaction. The upper panel shows the results for reverse transcriptase-polymerase chain reaction; the lower panel, for polymerase chain reaction with no reverse transcriptase. The primer sets are noted at the top. The nucleic acid templates, and whether they have been treated with DNase, are noted above each sample lane. “EF” signifies the clinical sample obtained from elbow-fluid aspirate. In the reverse transcriptase (+RT) lanes, all three Staphylococcus aureus primer sets (first three columns) yielded amplimer in both the aspirate and positive controls. In the lanes with no reverse transcriptase (-RT), only the non-DNase-treated specimens yielded amplimer, as expected. No amplimer was obtained with the Staphylococcus epidermidis primers (last column) in either the clinical specimen or the Staphylococcus aureus negative control, demonstrating that the Staphylococcus epidermidis primers used were species-specific and that no Staphylococcus epidermidis was detected in our clinical sample. Lanes 1, 8, and 15 were “1 kilobase (kb) plus” molecular weight standards. RT = reverse transcriptase; Sau = Staphylococcus aureus primer set; Staph GAPDH = Staphylococcal primer set (GF-1/GR-2) directed against glyceraldehyde-3-phosphate dehydrogenase gene; and Sepi = Staphylococcus epidermidis primer set.
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