ᰔ Bizzini et al. described the use of a commercial matrixassisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) system as a reliable, fast, and efficient method for the identification (ID) of bacteria and yeasts (1). Among 1,371 isolates identified by conventional methods, 1,037 (75.6%) isolates were identified by MALDI-TOF to either the species or genus level by direct analysis of a colony. A small amount of colony was smeared onto a gridded steel plate, allowed to dry briefly, and overlaid with 1 l of HCCA (␣-cyano-4-hydroxycinnamic acid) matrix before analysis in a Bruker Microflex MS system. For isolates which are not identified by this direct method, Bizzini et al. used an extraction method in which colonies were suspended in water and alcohol and centrifuged and bacterial proteins in the deposit were extracted with formic acid and acetonitrile before analysis (1).We have been using a similar approach in the diagnostic laboratory at the Royal London Hospital, a tertiary referral hospital which serves a population of 2 million people in East London. Here we show that by a minor modification of the direct analysis method, it is possible to increase the number of successfully identified isolates without recourse to the alcohol extraction protocol, which, although simple, significantly increases the processing time.We analyzed 2,020 isolates drawn from a routine diagnostic microbiology laboratory by the recommended direct smear method on a Bruker Microflex MS with the Biotyper software (version 2.0, database 3,740) and automation control. Approximately 42% of these isolates were Gram-negative bacilli, 35% staphylococci, 16% streptococci, 2% Gram-positive bacilli, 2% Gram-negative cocci, and 3% yeasts. Of the 2,020 isolates analyzed directly, 1,784 (88.3%) were acceptably identified to at least the genus level by the direct smear method. We accepted all scores of Ͼ1.7 as a valid identification to the genus level, based on the manufacturer's recommendations and our experience during the initial validation of this technology. The 236 isolates which did not give an acceptable ID (with scores of Ͻ1.7), mainly Gram-positive bacilli, coagulase-negative staphylococci, yeasts, and anaerobes, were then retested, with the additional step of adding 1 l of neat formic acid to the air-dried bacterial spot. This was then allowed to air dry before the addition of the matrix and analyzed in the usual way. This approach yielded a further 220 (10.9%) reliable identifications, leaving only 16 (0.8%) without a reliable ID, and these were then analyzed by alcohol extraction. Seven isolates did not give a satisfactory identification by alcohol extraction; two of these were regarded as being of little clinical significance, and no attempt was made to identify them further. The remaining five isolates were identified as Fusobacterium nucleatum, Prevotella oris, Paenibacillus barengoltzii, Actinomyces lignae, Actinomyces odontolyticus, and Pseudomonas aeruginosa by sequencing 1,250-to 1,300-bp fragments of ...
Several studies have reported poor results when trying to identify microorganisms directly from the bioMérieux BacT/ALERT blood culture system using matrix-assisted laser desorption/ionisation-time of flight (MALDI-TOF) mass spectrometry. The aim of this study is to evaluate two new methods, Sepsityper and an enrichment method for direct identification of microorganisms from this system. For both methods the samples were processed using the Bruker Microflex LT mass spectrometer (Biotyper) using the Microflex Control software to obtain spectra. The results from direct analysis were compared with those obtained by subculture and subsequent identification. A total of 350 positive blood cultures were processed simultaneously by the two methods. Fifty-three cultures were polymocrobial or failed to grow any organism on subculture, and these results were not included as there was either no subculture result, or for polymicrobial cultures it was known that the Biotyper would not be able to distinguish the constituent organisms correctly. Overall, the results showed that, contrary to previous reports, it is possible to identify bacteria directly from bioMérieux blood culture bottles, as 219/297 (74%) correct identifications were obtained using the Bruker Sepsityper method and 228/297 (77%) were obtained for the enrichment method when there is only one organism was present. Although the enrichment method was simpler, the reagent costs for the Sepsityper method were approximately pound 4.00 per sample compared to pound 0.50. An even simpler and cheaper method, which was less labour-intensive and did not require further reagents, was investigated. Seventy-seven specimens from positive signalled blood cultures were analysed by inoculating prewarmed blood agar plates and analysing any growth after 1-, 2- and 4-h periods of incubation at 37 degrees C, by either direct transfer or alcohol extraction. This method gave the highest number of correct identifications, 66/77 (86%), and was cheaper and less labour-intensive than either of the two above methods.
Background and Purpose-Neurocognitive dysfunction complicates coronary artery bypass surgery. Erythropoietin may be neuroprotective. We sought to determine whether human recombinant erythropoietin would reduce the incidence of neurocognitive dysfunction after surgery. Methods-We randomly assigned 32 elective first-time coronary artery bypass graft patients to receive placebo or 375 U/kg, 750 U/kg, or 1500 U/kg of recombinant human erythropoietin divided in 3 daily doses, starting the day before surgery. Primary outcomes were feasibility and safety, and secondary outcomes were neurocognitive dysfunction at discharge and 2 months. Results-All subjects were male, mean age 60 years (range 46 to 73). No significant differences were found in pump time, cross-clamp time, or hospital length of stay. Mortality and pure red cell aplasia were not observed. One patient in the 375 U/kg group had ST changes compatible with myocardial injury immediately postoperative, but no other thrombotic complications were observed. Neurocognitive dysfunction occurred in 21/32 (66%) of patients at discharge and 5/32 (16%) at 2 months. Neurocognitive dysfunction at discharge by group was: placebo 6/8 (75%), 375 U/kg 4/8 (50%), 750 U/kg 6/8 (75%), and 1500 U/kg 5/8 (63%). Neurocognitive dysfunction at 2 months by group was: placebo 3/8 (38%), 375 U/kg 1/8 (13%), 750 U/kg 1/8 (13%), and 1500 U/kg 0/8 (0%). Neurocognitive dysfunction at 2 months for erythropoietin at any dose was 2/24 (8.3%) versus 3/8 (38%) for placebo (Pϭ0.085). Conclusions-This
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