Simple Sample Preparation with SDS for Direct Identification from Positive Blood Culture Using MALDI-TOF MS.
Keywords:
Direct identification, Blood culture, MALDI-TOF MSAbstract
Septicemia is a life-threatening condition. Accurate and rapid pathogen diagnosis is essential for providing appropriate and expedient care to patients, thereby reducing the severity and complications of the disease. The study aimed to evaluate the sample preparation method for direct bacterial identification by the matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF MS) technique. One hundred fifty-seven positive blood culture samples from the BACTEX FX automated culture system were prepared for direct identification by MALDI-TOF MS. The preparation was done by using 10% sodium dodecyl sulfate (SDS) as a lysis buffer and 70% formic acid and acetonitrile for protein extraction. Then, the samples were determined by MALDI-TOF MS and the results were compared to results from the standard method using colony from solid media. The results showed that by using the direct method, the identification results were correctly identified at species and genus levels at 81.53% (128/157) and 14.01% (22/157), respectively. There were unreliable identification results at 4.46% (7/157). Those that were identified consistently but the score represented lower than 1.70 were 3.82% (6/157) and no peak 0.64% (1/157). There was no misidentification by using the direct identification method. The direct identification results correctly identified at species levels were 75.68% (56/74), 88.24% (60/68) and 80.00% (12/15) for gram negative bacteria, gram positive bacteria and others (yeast and mycobacteria), respectively and at genus levels were 17.58% (13/74), 10.29% (7/68), and 13.33% (2/15), respectively. There was no difference of the spectral score level obtained from the direct identification compared to using colonies with the Wilcoxon Signed Rank test at a confidence level of 95% in the gram-negative bacteria (p = 0.142) and the others (yeast and mycobacteria) (p = 0.530). However, using the standard method with MALDI-TOF in the gram-positive bacteria yielded higher score compared to using the direct method, which are statistically significant (p < 0.05). This study revealed that the preparation of positive blood cultures with 10% SDS for direct identification by MALDI-TOF MS showed excellent performance for pathogenic identification and shorter turn-around time by several hours or even days (< 30 minutes to 2 hours). This rapid direct identification allows a much faster optimization of antibiotic therapy in patients with sepsis compared to conventional workflows (only the gram stain result was known), for effective treatment thus reduced mortality and preventing drug resistance.
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