Rapid and robust analytical protocol for E. coli STEC bacteria subspecies differentiation using whole cell MALDI mass spectrometry

McLean, Kevin; Palarea‐Albaladejo, Javier; Currie, Colin; Imrie, Lisa; Manson, Erin; Fraser-Pitt, D.; Wright, Frank; Alexander, Colin James; Pollock, Kevin G; Allison, Lesley; Hanson, Mary; Smith, David George Emslie

doi:10.1016/j.talanta.2018.01.055

Cited by 10 publications

(9 citation statements)

References 29 publications

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“…The different percentage of acid used in other studies [50% ACN and 2.5% TFA] [16,19,20] may alter the pattern of those extracted proteins. Nevertheless, the ability to tailor a database expands the application of MALDI Biotyper as an identification tool for bacterial species specific to a host or location, and at below species-levels, such as subspecies, strain, or serotype [21,22].…”

Section: Discussionmentioning

confidence: 99%

Identification of bacterial pathogens in cultured fish with a custom peptide database constructed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS)

et al. 2020

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Background: The majority of infectious diseases of cultured fish is caused by bacteria. Rapid identification of bacterial pathogens is necessary for immediate management. The present study developed a custom Main Spectra Profile (MSP) database and validate the method using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) for rapid identification of fish bacterial pathogens. Streptococcus agalactiae, Streptococcus iniae, Aeromonas hydrophila, Aeromonas veronii, and Edwardsiella tarda obtained from diseased fish were used as representative bacterial pathogens in this study. Bacterial peptides were extracted to create a Main Spectra Profile (MSP), and the MSPs of each bacterial species was added into the MALDI Biotyper database. Fifteen additional isolates of each bacterial species were tested to validate the utilized technique. Results: The MSPs of all field isolates were clearly distinguishable, and the MSPs of the same species were clustered together. The identification methodology was validated with 75 bacterial isolates. The reliability and specificity of the method were determined with MALDI Biotyper log score values and matching results with 16 s rDNA sequencing. The species identification using the public MALDI Biotyper library (Bruker MALDI Biotyper) showed unreliable results (log score < 2.000) with 42.67% matching result with the reference method. In contrast, accurate identification was obtained when using the custom-made database, giving log score > 2.115, and a 100% matching result. Conclusion: This study demonstrates an effective identification of fish bacterial pathogens when a complete custom-made MSP database is applied. Further applications require a broad, well-established database to accommodate prudent identification of many fish bacterial pathogens by MALDI-TOF MS.

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Section: Discussionmentioning

confidence: 99%

Identification of bacterial pathogens in cultured fish with a custom peptide database constructed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS)

et al. 2020

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“…We used the detected lipid features to cluster individual cells using t-SNE with subsequent Louvain–Jaccard clustering. , Overall, 101 distinct clusters were identified using the Louvain–Jaccard clustering criteria (Figure A). To our knowledge, this is the largest number of clusters determined using any single-cell MS approach ,− and rivals that of single-cell transcriptomic measurements, demonstrating the rich information garnered using our high-throughput lipid profiling approach. Cluster 1 was the largest and contained 882 cells; clusters 100 and 101 were the smallest with 31 cells each.…”

Section: Resultsmentioning

confidence: 89%

Lipid Analysis of 30 000 Individual Rodent Cerebellar Cells Using High-Resolution Mass Spectrometry

et al. 2019

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Single cell measurements aid our understanding of chemically heterogeneous systems such as the brain. Lipids are one of the least studied chemical classes and their cell-to-cell heterogeneity remains largely unexplored. We adapted microscopy guided single cell profiling using matrix assisted laser desorption / ionization ion cyclotron resonance mass spectrometry to profile the lipid composition of over 30,000 individual rat cerebellar cells. We detected 520 lipid features, many of which were found in subsets of cells; Louvain clustering identified 101 distinct groups that can be correlated to neuronal and astrocytic classifications and lipid classes. Overall, the two most common lipids found were [PC(32:0)+H] + and [PC(34:1)+H] + , which were present within 98.9% and 89.5% of cells, respectively; lipid signals present in <1% of cells were also detected, including [PC(34:1)+K] + and [PG(40:2(OH))+Na] + . These results illustrate the vast lipid heterogeneity found within rodent cerebellar cells and hint at the distinct functional consequences of this heterogeneity.

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“…(Demirci et al, 2019); Listeria spp. (Thouvenot et al, 2018); Escherichia coli STEC (Mclean et al, 2018); Campylobacter (Lawton et al, 2018); Staphylococcus enterotoxin (Tonacini et al, 2019); Bacillus cereus cereulide (Ulrich et al, 2019); epidemiological studies (Camarasa and Cobo, 2018;Welker et al, 2019), detection of Legionella pneumophila in water system (Trnková et al, 2018), resistance to antibiotics (Carvalho et al, 2017;Cordovana et al, 2019) and pathogens in blood and urinary tract (Opota, 2018;Sánchez-Juanes et al, 2018), among other research studies. The limitation of the technology is that the identification of new isolates is possible only if the spectral database contains fingerprints of peptide mass of the strain type of specific genera, species or subspecies.…”

Section: Resultsmentioning

confidence: 99%

Recent trends for detection, quantification and identification of microorganisms in food industry

Fröder¹,

Ruis²,

Drebes³

et al. 2020

Microbiol Res Int

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Phenotypic characterization of microorganisms was originally performed using culture and physiological/biochemical methods. Throughout the 90s, traditional PCR was being replaced by the secondgeneration PCR, the real-time PCR (qPCR). This method is very effective for negative screening and also to confirm an isolate. However, limitations of qPCR are the inability to distinguish viable from dead cells and the sensitivity of amplification reactions to inhibitors. Sensitivity, specificity, robustness, time of response and the cost per analysis are often the criteria used for choosing a bacteria identification protocol. In this context, bacterial identification based on MALDI-TOF MS is becoming a method of choice for determining the genus, species and even subspecies of bacterial and fungal isolates in food analysis. MALDI-TOF MS is a promising method to identify bacteria and yeast in a few minutes directly from colonies grown in culture dishes. However, the identification of new isolates is possible only if the spectral database contains fingerprints of peptide mass from the strain type of specific microorganism. All current methods have their own guarantees, and so far, no method meets all attributes desidered by the laboratory; therefore, recent research studies are associating two or more methods with different technologies.

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Rapid and robust analytical protocol for E. coli STEC bacteria subspecies differentiation using whole cell MALDI mass spectrometry

Cited by 10 publications

References 29 publications

Identification of bacterial pathogens in cultured fish with a custom peptide database constructed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS)

Identification of bacterial pathogens in cultured fish with a custom peptide database constructed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS)

Lipid Analysis of 30 000 Individual Rodent Cerebellar Cells Using High-Resolution Mass Spectrometry

Recent trends for detection, quantification and identification of microorganisms in food industry

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Rapid and robust analytical protocol for E. coli STEC bacteria subspecies differentiation using whole cell MALDI mass spectrometry

Cited by 10 publications

References 29 publications

Identification of bacterial pathogens in cultured fish with a custom peptide database constructed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS)

Identification of bacterial pathogens in cultured fish with a custom peptide database constructed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS)

Lipid Analysis of 30 000 Individual Rodent Cerebellar Cells Using High-Resolution Mass Spectrometry

Recent trends for detection, quantification and identification of microorganisms in food industry

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Product

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Lipid Analysis of 30 000 Individual Rodent Cerebellar Cells Using High-Resolution Mass Spectrometry