Matrix-assisted laser desorption ionization-time of flight mass spectrometry has emerged as a rapid, cost-effective alternative for bacterial species identification. Identifying 60 blind-coded nonfermenting bacteria samples, this international study (using eight laboratories) achieved 98.75% interlaboratory reproducibility. Only 6 of the 480 samples were misidentified due to interchanges (4 samples) or contamination (1 sample) or not identified because of insufficient signal intensity (1 sample).Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) has emerged as a fast and costeffective alternative for bacterial species identification in microbiology. On the basis of mass analysis of the protein composition of a bacterial cell, which is assumed to be characteristic for each bacterial species, it is possible to determine the species within few minutes, starting from whole cells, cell lysates, or crude bacterial extracts (2, 3, 5, 6). The proof of principle of MALDI-TOF MS for bacterial species identification was shown a decade ago (2, 5, 6); however, due to low reproducibility, it has not been widely adopted in clinical microbiology. We have recently shown that use of a larger mass range for detection (2,000 to 20,000 Da), dedicated analysis software for spectral pattern matching, and a highquality reference database of spectra generated from qualitycontrolled culture collection strains resulted in accurate species identifications, with high intralaboratory reproducibility (7). For interlaboratory reproducibility, there are only very limited data available (8, 10). We therefore evaluated the interlaboratory reproducibility for MALDI-TOF MS-based species identification in a multicenter study, applying the above-described MALDI-TOF MS improvements.(

The incidence of vancomycin-resistant Enterococcus faecium isolation was low (

(17,23,24,25). Although the species are closely related genetically, they differ in host and geographic range, certain phenotypes, and pathogenicity. M. tuberculosis is the most significant pathogen for humans in Europe and America, whereas M. africanum is widely distributed among African patients (4, 15). Both subspecies of M. bovis are reported to infect humans, yet they have a broad host range, including wildlife and domestic livestock like bovines and goats (12,26). Of particular interest is the intrinsic resistance of M. bovis subsp. bovis against pyrazinamide, one of the first-line antituberculous drugs (11,17). The vaccine strain M. bovis BCG is more frequently used for bladder cancer immunotherapy and can be detected in human urine specimens from bladder cancer patients (2, 6). Thus, for those specimens, the isolation of M. tuberculosis complex is not necessarily an indication for antituberculous treatment. Therefore, rapid differentiation to the species and subspecies levels should be obtained not only for epidemiological purposes but also for adequate treatment of each patient.So far, rapid identification of the M. tuberculosis complex has been achieved by using AccuProbe gene probes (Gen-Probe, San Diego, Calif.), which target the 16S rRNA. However, species identification still relies on the determination of a series of phenotypic and growth characteristics and on biochemical tests, which all require sufficient growth, are time-consuming, and may not be performed in every laboratory (5, 25). Recently, the GenoType MTBC (Hain Lifescience GmbH, Nehren, Germany)-a new commercially available DNA strip assay for the rapid identification of the members of the M. tuberculosis complex-was evaluated by use of a well-characterized collection of M. tuberculosis complex isolates and was proven to be useful for species differentiation (20). Moreover, due to the inclusion of an M. tuberculosis complex-specific oligonucleotide, this test may replace the AccuProbe tests of cultures from clinical specimens as a method for identification of the M. tuberculosis complex, giving the additional information provided for the species involved.The aim of this work was to evaluate the performance of the GenoType MTBC with positive liquid cultures from clinical specimens received for mycobacterial culture in comparison to species identification by classical methods. Furthermore, the GenoType MTBC results were compared to results obtained by AccuProbe tests.Specimens. Two German microbiology laboratories were involved in this study: (i) the National Reference Center for Mycobacteria, Borstel, Germany, and (ii) the Department of Microbiology of the Laboratory Group, Heidelberg, Germany. All samples used in this study originated from human specimens and a few veterinary specimens sent to the laboratories for routine mycobacterial culture. Specimens (pulmonary and extrapulmonary) were processed according to national and international guidelines (5, 10) by using an N-acetyl-L-cysteineNaOH decontamination procedure, inoculated into B...

The performance of the BD Phoenix Automated Microbiology System (BD Diagnostic Systems, Sparks, Md.) was assessed for identification (ID) and antimicrobial susceptibility testing (AST) for the majority of clinically encountered bacterial isolates in a European collaborative two-center trial. A total of 469 bacterial isolates of the genera Staphylococcus (275 isolates), Enterococcus (179 isolates), and Streptococcus (15 isolates, for ID only) were investigated; of these, 367 were single patient isolates, and 102 were challenge strains tested at one center. Sixty-four antimicrobial drugs were tested, including the following drug classes: aminoglycosides, beta-lactam antibiotics, beta-lactam-beta-lactamase inhibitors, carbapenems, cephems, folate antagonists, quinolones, glycopeptides, macrolides-lincosamides-streptogramin B (MLS), and others. Phoenix ID results were compared to those of the laboratories' routine ID systems (API 32 Staph, API 32 Strep, and VITEK 2 [bioMérieux, Marcy l'Etoile, France]); Phoenix AST results were compared to those of frozen standard broth microdilution (SBM) panels according to NCCLS guidelines (NCCLS document M 100-S 9, approved standard M 7-A 4). Discrepant results were repeated in duplicate. Concordant IDs of 97.1, 98.9, and 100% were observed for staphylococci, enterococci, and streptococci, respectively. For AST results the overall essential agreement was 93.3%; the category agreement was 97.3%; and the very major error rate, major error rate, and minor error rate were 1.2, 1.9, and 1.3%, respectively. In conclusion, the Phoenix ID results showed high agreement with results of the systems to which they were being compared; the AST performance was highly equivalent to that of the SBM reference method.The clinical microbiology laboratory is confronted with an alarming increase of antimicrobial resistance on a global scale (7,8,9,13). Furthermore, the emergence of bacterial isolates with special resistance mechanisms such as oxacillin-resistant staphylococci or vancomycin-resistant enterococci constitutes a major problem, especially in intensive care units (1, 4). Both accurate and rapid diagnosis of oxacillin-resistant staphylococci and vancomycin-resistant enterococci has therefore become essential in the current health care environment.The BD Phoenix Automated Microbiology System (BD Diagnostic Systems, Sparks, Md.[BD]) is a newly developed instrument for the reliable and accurate identification and susceptibility testing for the majority of clinically encountered strains. The system is comprised of disposable panels, which combine both identification testing (ID) and antimicrobial susceptibility testing (AST), and an instrument which performs automatic reading at 20-min intervals during incubation. The system claims to provide accurate and rapid susceptibility results with easy workflow for the laboratory worker.We report on the ability of the Phoenix system to accurately perform ID and AST of clinical and challenge isolates in a large collaborative two-center trial involving th...

At present, the rapid diagnosis of pulmonary tuberculosis rests with microscopy. However, this technique is insensitive and many cases of pulmonary tuberculosis cannot be initially confirmed. Nucleic acid amplification techniques are extremely sensitive, but when they are applied to tuberculosis diagnosis, they have given variable results. Investigators at six centers in Europe compared a standardized PCR system (Amplicor; Roche) against conventional culture methods. Defined clinical information was collected. Discrepant samples were retested, and inhibition assays and backup amplification with a separate primer pair were performed. Mycobacterium tuberculosis complex organisms were recovered from 654 (9.1%) of 7,194 samples and 293 (7.8%) of 3,738 patients. Four hundred fifty-two of the M. tuberculosis isolates from 204 patients were smear positive and culture positive. Among the culture-positive specimens, PCR had a sensitivity of 91.4% for smear-positive specimens and 60.9% for smear-negative specimens, with a specificity of 96.1%. Analysis of 254 PCR-positive, culture-negative specimens with discrepant results revealed that 130 were from patients with recently diagnosed tuberculosis and 94 represented a presumed laboratory error. Similar analysis of 118 PCR-negative, culture-positive specimens demonstrated that 27 discrepancies were due to presumed uneven aliquot distribution and 11 were due to presumed laboratory error; PCR inhibitors were detected in 8 specimens. Amplicor enables laboratories with little previous experience with nucleic acid amplification to perform PCR. Disease in more than 60% of the patients with tuberculosis with smear-negative, culture-positive specimens can be diagnosed at the time of admission, and potentially all patients with smear-positive specimens can immediately be confirmed as being infected with M. tuberculosis, leading to improved clinical management.