| Objectives: Platelets are stored aerobically for up to 5 days at 20°C to 24°C, allowing a wide variety of bacteria to grow. Transfusion of platelet concentrates (PCs) contaminated with bacteria may cause serious septic reactions that can be fatal to the recipient. Prevention or reduction of the occurrence of these septic reactions is therefore a major challenge faced in blood banking and transfusion medicine. Up to now various techniques, like pretransfusion bacterial screening of PCs, have been adopted by blood centers to reduce the risk of transfusion-associated septic reactions. The blood culture system is regarded by many as the'gold standard'for bacterial screening of platelet concentrates. This is a culture technology and is used by the majority of blood services that are currently undertaking platelet concentrate screening. It has proven to be useful for identification of bacterially contaminated PCs but has also proven to be inconvenient because of the time they require to indicate the presence of bacteria. To be useful as an optimal tool for routine screening of PCs, a dependable assay is desirable. The assay should be simple, sensitive enough to detect all clinically significant levels of bacteria, highly specific, and rapid to allow the release of PCs for clinical use within hours. The aim of this study was a test that would meet all these criteria. As reported in a previous study, a broad-range 16SrDNA polymerase chain reaction (PCR) assay was developed。To assess the applicability of this method for detecting the bacteria contamination of platelet concentrations, the assay were compared with a blood culture system.Methods: The presence of bacteria in pooled PCs was routinely assessed ina culturing system (BacT/ALERT, bioMérieux). The PCR assay was performed with DNA extracted from the same samples as used for culturing. PCR amplification was performed with a set of universal primers targeting eubacterial 16S rDNA and 16S to 23S spacer region. The PCR product was then digested by restriction endonuclease HaeⅢfor 30 minutes. The mixture solutions were diluted for 20 times before taking capillary electrophoresis, and the Restriction-Fragment-length polymorphism(RFLP) were detected by ABI prism 310 Genetic Analyzer. To determine the detection limit of the assay, 10 ml of DW was spiked with 1 ml of serial dilutions of Escherichia coli, Staphylococcus aureaus and Staphylococcus epidermidis. The number of bacteria ranged from 1 to 108 CFU/ml. DNA was subsequently extracted from these spiked samples. The sensitivity of this assay can be hampered by contaminating DNA in the PCR reagents. Digestion of the PCR reagents with Sau3AI prior to PCR amplification was effective in eliminating this contaminating DNA without affecting the sensitivity of the assay.Results: A total of 276 PCs were tested. 144 PCs were stored after 24 hours, the others were stored after 48 hours. 2 samples(0.7246%) were positive for the presence of bacteria by both methods. contaminants were identified as Staphylococcus aureaus and Bacillus subtilis. The turnaround time of this PCR assay were about 20 hours and 58 hours respectively. Staphylococcus epidermidis were detected in the withdrawal of contaminated PCs in about 35 hours with the PCR assay. The detection limit of the assay is about 10CFU/ml of Escherichia coli and 100CFU/ml of Staphylococcus.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 PCR assay has a much shorter turnaround time of about 4 hours, which offers the possibility to test and obtain results on PCs before release or the day they were transfused. This would permit the withdrawal of contaminated PCs before transfusion.
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