Premium
Evaluation of an ethidium monoazide–enhanced 16 S r DNA real‐time polymerase chain reaction assay for bacterial screening of platelet concentrates and comparison with automated culture
Author(s) -
Garson Jeremy A.,
Patel Poorvi,
McDonald Carl,
Ball Joanne,
Rosenberg Gillian,
Tettmar Kate I.,
Brailsford Susan R.,
Pitt Tyrone,
Tedder Richard S.
Publication year - 2014
Publication title -
transfusion
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.045
H-Index - 132
eISSN - 1537-2995
pISSN - 0041-1132
DOI - 10.1111/trf.12256
Subject(s) - taqman , microbiology and biotechnology , real time polymerase chain reaction , polymerase chain reaction , biology , listeria monocytogenes , microbiological culture , bacteria , biochemistry , genetics , gene
Background Culture‐based systems are currently the preferred means for bacterial screening of platelet (PLT) concentrates. Alternative bacterial detection techniques based on nucleic acid amplification have also been developed but these have yet to be fully evaluated. In this study we evaluate a novel 16 S rDNA polymerase chain reaction ( PCR ) assay and compare its performance with automated culture. Study Design and Methods A total of 2050 time‐expired, 176 fresh, and 400 initial‐reactive PLT packs were tested by real‐time PCR using broadly reactive 16S primers and a “universal” probe ( TaqMan , Invitrogen). PLTs were also tested using a microbial detection system ( BacT / ALERT , bioMérieux) under aerobic and anaerobic conditions. Results Seven of 2050 (0.34%) time‐expired PLTs were found repeat reactive by PCR on the initial nucleic acid extract but none of these was confirmed positive on testing frozen second aliquots. BacT / ALERT testing also failed to confirm any time‐expired PLTs positive on repeat testing, although 0.24% were reactive on the first test. Three of the 400 “initial‐reactive” PLT packs were found by both PCR and BacT / ALERT to be contaminated ( Escherichia coli , Listeria monocytogenes , and Streptococcus vestibularis identified) and 14 additional packs were confirmed positive by BacT / ALERT only. In 13 of these cases the contaminating organisms were identified as anaerobic skin or oral commensals and the remaining pack was contaminated with Streptococcus pneumoniae . Conclusion These results demonstrate that the 16 S PCR assay is less sensitive than BacT / ALERT and inappropriate for early testing of concentrates. However, rapid PCR assays such as this may be suitable for a strategy of late or prerelease testing.