Dataset Information

A rapid diagnostic workflow for cefotaxime-resistant Escherichia coli and Klebsiella pneumoniae detection from blood cultures by MALDI-TOF mass spectrometry.

ABSTRACT: Nowadays, the global spread of resistance to oxyimino-cephalosporins in Enterobacteriaceae implies the need for novel diagnostics that can rapidly target resistant organisms from these bacterial species.In this study, we developed and evaluated a Direct Mass Spectrometry assay for Beta-Lactamase (D-MSBL) that allows direct identification of (oxyimino)cephalosporin-resistant Escherichia coli or Klebsiella pneumoniae from positive blood cultures (BCs), by using the matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) technology.The D-MSBL assay was performed on 93 E. coli or K. pneumoniae growing BC samples that were shortly co-incubated with cefotaxime (CTX) as the indicator cephalosporin. Susceptibility and resistance defining peaks from the samples' mass spectra were analyzed by a novel algorithm for bacterial organism classification. The D-MSBL assay allowed discrimination between E. coli and K. pneumoniae that were resistant or susceptible to CTX with a sensitivity of 86.8% and a specificity of 98.2%.The proposed algorithm-based D-MSBL assay, if integrated in the routine laboratory diagnostic workflow, may be useful to enhance the establishment of appropriate antibiotic therapy and to control the threat of oxyimino-cephalosporin resistance in hospital.

SUBMITTER: De Carolis E

PROVIDER: S-EPMC5628900 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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A rapid diagnostic workflow for cefotaxime-resistant Escherichia coli and Klebsiella pneumoniae detection from blood cultures by MALDI-TOF mass spectrometry.

De Carolis Elena E Paoletti Silvia S Nagel Domenico D Vella Antonietta A Mello Enrica E Palucci Ivana I De Angelis Giulia G D'Inzeo Tiziana T Sanguinetti Maurizio M Posteraro Brunella B Spanu Teresa T

PloS one 20171005 10

<h4>Background</h4>Nowadays, the global spread of resistance to oxyimino-cephalosporins in Enterobacteriaceae implies the need for novel diagnostics that can rapidly target resistant organisms from these bacterial species.<h4>Methods</h4>In this study, we developed and evaluated a Direct Mass Spectrometry assay for Beta-Lactamase (D-MSBL) that allows direct identification of (oxyimino)cephalosporin-resistant Escherichia coli or Klebsiella pneumoniae from positive blood cultures (BCs), by using t ...[more]

PMID: 28982134

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Project description:BackgroundSerotyping of Streptococcus pneumoniae is important for monitoring of vaccine impact. Unfortunately, conventional and molecular serotyping is expensive and technically demanding. This study aimed to determine the ability of matrix-assisted laser desorption-ionisation time-of-flight (MALDI-TOF) mass spectrometry to discriminate between pneumococcal serotypes and genotypes (defined by global pneumococcal sequence cluster, GPSC). In this study, MALDI-TOF mass spectra were generated for a diverse panel of whole genome sequenced pneumococcal isolates using the bioMerieux VITEK MS in clinical diagnostic (IVD) mode. Discriminatory mass peaks were identified and hierarchical clustering was performed to visually assess discriminatory ability. Random forest and classification and regression tree (CART) algorithms were used to formally determine how well serotypes and genotypes were identified by MALDI-TOF mass spectrum.ResultsOne hundred and ninety-nine pneumococci, comprising 16 serotypes and non-typeable isolates from 46 GPSC, were analysed. In the primary experiment, hierarchical clustering revealed poor congruence between MALDI-TOF mass spectrum and serotype. The correct serotype was identified from MALDI-TOF mass spectrum in just 14.6% (random forest) or 35.4% (CART) of 130 isolates. Restricting the dataset to the nine dominant GPSC (61 isolates / 13 serotypes), discriminatory ability improved slightly: the correct serotype was identified in 21.3% (random forest) and 41.0% (CART). Finally, analysis of 69 isolates of three dominant serotype-genotype pairs (6B-GPSC1, 19F-GPSC23, 23F-GPSC624) resulted in the correct serotype identification in 81.1% (random forest) and 94.2% (CART) of isolates.ConclusionsThis work suggests that MALDI-TOF is not a useful technique for determination of pneumococcal serotype. MALDI-TOF mass spectra appear more associated with isolate genotype, which may still have utility for future pneumococcal surveillance activities.

Project description:BackgroundNeuropeptides are a diverse category of signaling molecules in the nervous system regulating a variety of processes including food intake, social behavior, circadian rhythms, learning, and memory. Both the identification and functional characterization of specific neuropeptides are ongoing fields of research. Matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) analysis of nervous tissues from a variety of organisms allows direct detection and identification of neuropeptides. Here, we demonstrate an analysis workflow that allows for the detection of differences in specific neuropeptides amongst a variety of neuropeptides being simultaneously measured. For sample preparation, we describe a straight-forward and rapid (minutes) method where individual adult Drosophila melanogaster brains are analyzed. Using a MATLAB-based data analysis workflow, also compatible with MALDI-TOF mass spectra obtained from other sample preparations and instrumentation, we demonstrate how changes in neuropeptides levels can be detected with this method.ResultsOver fifty isotopically resolved ion signals in the peptide mass range are reproducibly observed across experiments. MALDI-TOF MS profile spectra were used to statistically identify distinct relative differences in organ-wide endogenous levels of detected neuropeptides between biological conditions. In particular, three distinct levels of a particular neuropeptide, pigment dispersing factor, were detected by comparing groups of preprocessed spectra obtained from individual brains across three different D. melanogaster strains, each of which express different amounts of this neuropeptide. Using the same sample preparation, MALDI-TOF/TOF tandem mass spectrometry confirmed that at least 14 ion signals observed across experiments are indeed neuropeptides. Among the identified neuropeptides were three products of the neuropeptide-like precursor 1 gene previously not identified in the literature.ConclusionsUsing MALDI-TOF MS and preprocessing/statistical analysis, changes in relative levels of a particular neuropeptide in D. melanogaster tissue can be statistically detected amongst a variety of neuropeptides. While the data analysis methods should be compatible with other sample preparations, the presented sample preparation method was sufficient to identify previously unconfirmed D. melanogaster neuropeptides.

Project description:Bacillus cereus plays an often unrecognized role in food borne diseases. Food poisoning caused by this pathogen is manifested by either diarrhea or emesis. Due to the relatively high prevalence of emetic toxin cereulide associated food poisoning, methods for simple and reliable detection of cereulide producing strains are of utmost importance. Recently, two different studies reported on the application of MALDI-ToF MS for either the differentiation of emetic and non-emetic strains of B. cereus or for direct detection of cereulide from bacterial colony smears. However, for implementation of cereulide detection using MALDI-ToF MS in routine microbiological diagnostics additional investigations on the sensitivity and specificity as well as on the fitting into common workflows for bacterial identification are needed. These aspects prompted us to investigate open issues and to test sample preparation methods, commonly used for microbial identification for their suitability to detect the emetic toxin from bacteria. Based on our experimental findings we propose a workflow that allows identification of B. cereus and sensitive detection of cereulide in parallel, using linear-mode MALDI-ToF MS equipment. The protocol was validated in a blinded study and is based on the well-established ethanol/formic acid extraction method. Cereulide is detected in the ethanol wash solution of samples identified as B. cereus as peaks at m/z 1175 and 1191. Peak position difference of 16 Th (Thomson) indicates detection of the sodium and potassium adducts of cereulide. This sample treatment offers possibilities for further characterization by more sophisticated LC-MS-based methods. In summary, the ease of use and the achieved level of analytical sensitivity as well as the wide-spread availability of MALDI-ToF MS equipment in clinical microbiological laboratories provides a promising tool to improve and to facilitate routine diagnostics of B. cereus associated food intoxications.

Project description:BackgroundMatrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry (MS) has become a widely used technique for the rapid and accurate identification of bacteria, mycobacteria and certain fungal pathogens in the clinical microbiology laboratory. Thus far, only few attempts have been made to apply the technique in clinical parasitology, particularly regarding helminth identification.MethodsWe systematically reviewed the scientific literature on studies pertaining to MALDI-TOF MS as a diagnostic technique for helminths (cestodes, nematodes and trematodes) of medical and veterinary importance. Readily available electronic databases (i.e. PubMed/MEDLINE, ScienceDirect, Cochrane Library, Web of Science and Google Scholar) were searched from inception to 10 October 2018, without restriction on year of publication or language. The titles and abstracts of studies were screened for eligibility by two independent reviewers. Relevant articles were read in full and included in the systematic review.ResultsA total of 84 peer-reviewed articles were considered for the final analysis. Most papers reported on the application of MALDI-TOF for the study of Caenorhabditis elegans, and the technique was primarily used for identification of specific proteins rather than entire pathogens. Since 2015, a small number of studies documented the successful use of MALDI-TOF MS for species-specific identification of nematodes of human and veterinary importance, such as Trichinella spp. and Dirofilaria spp. However, the quality of available data and the number of examined helminth samples was low.ConclusionsData on the use of MALDI-TOF MS for the diagnosis of helminths are scarce, but recent evidence suggests a potential role for a reliable identification of nematodes. Future research should explore the diagnostic accuracy of MALDI-TOF MS for identification of (i) adult helminths, larvae and eggs shed in faecal samples; and (ii) helminth-related proteins that are detectable in serum or body fluids of infected individuals.

Dataset Information

A rapid diagnostic workflow for cefotaxime-resistant Escherichia coli and Klebsiella pneumoniae detection from blood cultures by MALDI-TOF mass spectrometry.

Publications

A rapid diagnostic workflow for cefotaxime-resistant Escherichia coli and Klebsiella pneumoniae detection from blood cultures by MALDI-TOF mass spectrometry.

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