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      Identification of Brucella by MALDI-TOF Mass Spectrometry. Fast and Reliable Identification from Agar Plates and Blood Cultures

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          Abstract

          Background

          MALDI-TOF mass spectrometry (MS) is a reliable method for bacteria identification. Some databases used for this purpose lack reference profiles for Brucella species, which is still an important pathogen in wide areas around the world. We report the creation of profiles for MALDI-TOF Biotyper 2.0 database (Bruker Daltonics, Germany) and their usefulness for identifying brucellae from culture plates and blood cultures.

          Methodology/Principal Findings

          We created MALDI Biotyper 2.0 profiles for type strains belonging to B. melitensis biotypes 1, 2 and 3; B. abortus biotypes 1, 2, 5 and 9; B. suis, B. canis, B ceti and B. pinnipedialis. Then, 131 clinical isolates grown on plate cultures were used in triplicate to check identification. Identification at genus level was always correct, although in most cases the three replicates reported different identification at species level. Simulated blood cultures were performed with type strains belonging to the main human pathogenic species ( B. melitensis, B. abortus, B. suis and B. canis), and studied by MALDI-TOF MS in triplicate. Identification at genus level was always correct.

          Conclusions/Significance

          MALDI-TOF MS is reliable for Brucella identification to the genus level from culture plates and directly from blood culture bottles.

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          Ongoing revolution in bacteriology: routine identification of bacteria by matrix-assisted laser desorption ionization time-of-flight mass spectrometry.

          Piseth Seng, Michel Drancourt, Frédérique Gouriet … (2009)
          Matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry accurately identifies both selected bacteria and bacteria in select clinical situations. It has not been evaluated for routine use in the clinic. We prospectively analyzed routine MALDI-TOF mass spectrometry identification in parallel with conventional phenotypic identification of bacteria regardless of phylum or source of isolation. Discrepancies were resolved by 16S ribosomal RNA and rpoB gene sequence-based molecular identification. Colonies (4 spots per isolate directly deposited on the MALDI-TOF plate) were analyzed using an Autoflex II Bruker Daltonik mass spectrometer. Peptidic spectra were compared with the Bruker BioTyper database, version 2.0, and the identification score was noted. Delays and costs of identification were measured. Of 1660 bacterial isolates analyzed, 95.4% were correctly identified by MALDI-TOF mass spectrometry; 84.1% were identified at the species level, and 11.3% were identified at the genus level. In most cases, absence of identification (2.8% of isolates) and erroneous identification (1.7% of isolates) were due to improper database entries. Accurate MALDI-TOF mass spectrometry identification was significantly correlated with having 10 reference spectra in the database (P=.01). The mean time required for MALDI-TOF mass spectrometry identification of 1 isolate was 6 minutes for an estimated 22%-32% cost of current methods of identification. MALDI-TOF mass spectrometry is a cost-effective, accurate method for routine identification of bacterial isolates in or =10 reference spectra per bacterial species and a 1.9 identification score (Brucker system). It may replace Gram staining and biochemical identification in the near future.
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            Complications associated with Brucella melitensis infection: a study of 530 cases.

            Rosa M. Reguera, J Colmenero, F Martos … (1996)
            We carried out a prospective study of 530 patients older than 14 years of age with brucellosis. We describe the incidence and clinical features of the focal forms of the disease, analyzing some of the possible factors associated with their appearance. One hundred sixty-nine patients (31.9%) had a focal form or complication. Osteoarticular complications were the most frequent, totaling 113 cases (66%), followed by genitourinary with 18 cases (5.1% of males), hepatic (2.5%), neurologic (1.7%), and heart (1.5%). Nine patients (1.7%) had more than 1 complication. In a multivariate analysis, diagnostic delay greater than 30 days (OR 2.0), ESR > 40 mm/hr (OR 1.9), and levels of alpha-2 globulin > 7.5 g/L (OR 6.8) were statistically significant independent variables associated with the presence of focal forms. Twenty-five patients with complications (14.8%) required surgical treatment. The relapse rate was 3.6% for those patients without complications and 4.1% for patients with focal forms (p > 0.05). However, when therapeutic failure, relapses, and mortality were considered together, the risk of an unfavorable evolution was significantly greater in patients with focal forms (10.6% versus 3.6% in patients without complications; OR 1.9, 95% CI 1.4-7.1, p < 0.005). Given the worse prognosis, knowledge and early diagnosis of the focal forms of B. melitensis infection is especially important.
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              Rapid identification of bacteria in positive blood culture broths by matrix-assisted laser desorption ionization-time of flight mass spectrometry.

              Nigel Murray, S Drake, Philip G. Stevenson (2010)
              Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry is a rapid, accurate method for identifying bacteria and fungi recovered on agar culture media. We report herein a method for the direct identification of bacteria in positive blood culture broths by MALDI-TOF mass spectrometry. A total of 212 positive cultures were examined, representing 32 genera and 60 species or groups. The identification of bacterial isolates by MALDI-TOF mass spectrometry was compared with biochemical testing, and discrepancies were resolved by gene sequencing. No identification (spectral score of or = 1.7, 162 (95.3%) of 170 isolates were correctly identified. All 8 isolates of Streptococcus mitis were misidentified as being Streptococcus pneumoniae isolates. This method provides a rapid, accurate, definitive identification of bacteria within 1 h of detection in positive blood cultures with the caveat that the identification of S. pneumoniae would have to be confirmed by an alternative test.
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                Author and article information

                Contributors
                : Role: Editor
                Journal
                Journal ID (nlm-ta): PLoS One
                Journal ID (publisher-id): plos
                Journal ID (pmc): plosone
                Title: PLoS ONE
                Publisher: Public Library of Science (San Francisco, USA )
                ISSN (Electronic): 1932-6203
                Publication date Collection: 2010
                Publication date (Electronic): 6 December 2010
                Volume: 5
                Issue: 12
                Electronic Location Identifier: e14235
                Affiliations
                [1 ]Unidad de Investigación, Hospital Universitario de Salamanca, Salamanca, Spain
                [2 ]Departamento de Microbiología, Hospital Universitario de Salamanca, Salamanca, Spain
                [3 ]Departamento de Anatomía Patológica, Microbiología, Medicina Preventiva y Salud Pública, Medicina Legal y Forense, Universidad de Valladolid, Valladolid, Spain
                [4 ]Departamento de Bioquímica y Biología Molecular, Universidad de Salamanca, Salamanca, Spain
                [5 ]Departamento de Medicina Preventiva, Salud Pública y Microbiología Médica, Universidad de Salamanca, Salamanca, Spain
                Universidad Nacional, Costa Rica
                Author notes

                Conceived and designed the experiments: LF JMGB JLMB. Performed the experiments: LF SVC FSJ SGC FM. Analyzed the data: LF FSJ JMGB JLMB. Contributed reagents/materials/analysis tools: SGC FM AOD JMGB JLMB. Wrote the paper: LF JMGB JLMB.

                Article
                Publisher ID: 10-PONE-RA-20150R1
                DOI: 10.1371/journal.pone.0014235
                PMC ID: 2997794
                PubMed ID: 21151913
                SO-VID: b41d34b2-6faa-45d2-9e74-5e990e02a3dc
                Copyright © Ferreira et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
                History
                Date received : 23 June 2010
                Date accepted : 15 November 2010
                Page count
                Pages: 8
                Categories
                Subject: Research Article
                Subject: Microbiology
                Subject: Microbiology/Medical Microbiology
                Subject: Infectious Diseases/Bacterial Infections

                ScienceOpen disciplines: Uncategorized
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                ScienceOpen disciplines: Uncategorized

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