Direct Identification of 80 Percent of Bacteria from Blood Culture Bottles by Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry Using a 10-Minute Extraction Protocol

Abstract

Matrix-assisted laser desorption ionization–time of flight mass spectrometry is not widely used to identify bacteria directly from positive blood culture bottles (BCBs) because of overlong protocols. The objective of this work was to develop and evaluate a simple extraction protocol for reliable identification from BCBs. The 10-min protocol was applied over a 5-month period. Direct identifications on day 0 were compared with those obtained from colonies on day 1 [log(score) of ≥2]. We evaluated a range of seven log(score) thresholds on day 0 from 1.4 to 2.0 to find the lower confidence score that provides the higher percentage of direct identifications without loss of accuracy. With a log(score) threshold of ≥1.5 at day 0, our protocol allowed us to identify 80% of bacteria in 632 BCBs (96% of Enterobacteriaceae, 95% of Staphylococcus aureus, 92% of enterococci, and 62% of streptococci). At least one bacterial species of the mixture was identified in 77% of the polymicrobial samples. The rapidity and reliability of the protocol were factors in its adoption for routine use, allowing us to save up to 24 h in identifying 80% of the bacteria in the BCBs and, thus, to supply useful information to adapt antibiotic therapy when necessary. We currently provide reliable daily direct identifications of staphylococci, enterococci, Enterobacteriaceae, Pseudomonas aeruginosa, and beta-hemolytic streptococci.

Rapid antimicrobial susceptibility testing on positive blood cultures through an innovative light scattering technology: performances and turnaround time evaluation

“…Speeded-up positive blood culture testing is therefore an important challenge for the hospital microbiology laboratory. Many authors focused on rapid identification by MALDI-TOF MS directly on positive blood cultures [7, 20]. However, publications about direct AST methods are less prevalent and mainly address testing on AST automated systems.…”

“…This delay is now reduced through new genotypic and phenotypic technologies performing the tests directly on the blood of the positive blood culture bottle. For example direct identification with matrix-assisted laser desorption time-of-flight mass spectrometry (MALDI-TOF MS) has been extensively validated with successful identification performances beyond 80% [7, 8]. Complementary molecular-based methods also entered the market offering fast identification combined to the detection of specific resistance genes and requiring a very limited hands-on time yet requiring costly reagents [9].…”

Evaluation of Rapid Sepsityper® protocol and specific MBT-Sepsityper module (Bruker Daltonics) for the rapid diagnosis of bacteremia and fungemia by MALDI-TOF-MS

“…In particular, despite their low cost, the important handson-time of MALDI-TOF-MS assays still prevents many laboratories to perform rapid identification on positive blood cultures, which results in a loss of opportunity for the patients. In-house and commercial protocols such as the Sepsityper ® kit (Bruker Daltonics GmbH, Bremen, Germany) usually take between 20 to 40 min of turnaround time [22,28,[30][31][32][33][34][35][36][37][38][39][40][41][42][43]. A comprehensive overview of current performances and estimated hands-on time of the different rapid identification methods using MALDI-TOF-MS on positive blood cultures has been gathered in Table 1.…”

Rapid identification of respiratory bacterial pathogens from bronchoalveolar lavage fluid in cattle by MALDI-TOF MS

“…These results were obtained using a cutoff value of ≥1.7. Although a (log) score value between 1.7 and 2.0 should be interpreted as species identification with low confidence, this cutoff value or even lower is commonly used for identification procedures of positive blood cultures by MALDI-TOF MS 25,26 and was therefore also applied in the present study.…”

“…Log score values < 1.7 represent no organism identification possible, a (log) score value between 1.7 and 2.0 represents identification at species level at low confidence, and a (log) score value ≥2.0 represents identification at species level at high confidence. The threshold for correct identification was determined at a (log) score value of ≥1.7, as previously described for rapid detection protocols in positive blood cultures 25,26 .…”