Detection of bacteria in platelet concentrates: comparison of broad-range real-time 16S rDNA polymerase chain reaction and automated culturing.
BACKGROUND
Based on real-time polymerase chain reaction (PCR) technology, a broad-range 16S rDNA assay was validated and its performance was compared to that of an automated culture system to determine its usefulness for rapid routine screening of platelet concentrates (PCs).


METHODS
The presence of bacteria in pooled PCs was routinely assessed in an automated culturing system (BacT/ALERT, bioMerieux). The PCR assay was performed with DNA extracted from the same samples as used for culturing. DNA extraction was performed with a automated extraction system (MagNA Pure, Roche Diagnostics). PCR amplification was performed with a set of universal primers and probe targeting eubacterial 16S rDNA.


RESULTS
A total of 2146 PCs were tested. Eighteen (0.83%) samples were found to be contaminated. These samples were positive for the presence of bacteria by both methods. All contaminants were identified as bacteria belonging to the common human skin flora. These included Propionibacterium spp. (n = 7), Staphylococcus spp. (n = 6), Bacillus spp. (n = 2), Micrococcus spp. (n = 2), and Peptostreptococcus spp. (n = 1). Estimation of the bacterial load in PCs by real-time PCR showed that the initial levels of contamination varied between 13.6 and 9 x 10(4) colony-forming unit equivalents per PCR procedure.


CONCLUSIONS
Compared to culture in the BacT/ALERT system, the PCR assay had a sensitivity of 100 percent and a specificity of 100 percent. This real-time PCR assay has a much shorter turnaround time of 4 hours, which offers the possibility to test and obtain results on PCs before release or the day they are transfused. This would permit the withdrawal of contaminated PCs before transfusion.
Mohammadi
Tamimount
T
Sanquin Blood Bank North West Region, Amsterdam, The Netherlands.
Pietersz
Ruby N I
RN
Vandenbroucke-Grauls
Christina M J E
CM
Savelkoul
Paul H M
PH
Reesink
Henk W
HW
D001419:Bacteria / Q000235:genetics* / Q000302:isolation & purification; D001424:Bacterial Infections / Q000175:diagnosis*; D001792:Blood Platelets / Q000382:microbiology*; D004269:DNA, Bacterial / Q000032:analysis; D004275:DNA, Ribosomal / Q000032:analysis; D006801:Humans; D008828:Microbiological Techniques; D010983:Plateletpheresis; D012336:RNA, Ribosomal, 16S / Q000032:analysis*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D012336:RNA, Ribosomal, 16S