MALDI-Biotyper und Milchanalytik: Spezielle Datenbanken für spezielle Aufgaben. Dr. Gerold Schwarz Product Manager MALDI
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1 MALDI-Biotyper und Milchanalytik: Spezielle Datenbanken für spezielle Aufgaben Dr. Gerold Schwarz Product Manager MALDI
2 2 BIOTECON Diagnostics GmbH Location and History Company Founded 1998 Headquarter in Potsdam CEO: Dr. Kornelia Berghof-Jäger CBO: Alois Schneiderbauer Certificates ISO 9001:2008 ISO GMP
3 3 Industrial Microbiology Food Market Pathogens Bacteria and viruses Spoilage Organisms Animal Identification GMOs Allergens Hygiene Parameters DNA Extraction Kits Detection Kits Real-Time PCR Cyclers Automation, Service and MALDI-Biotyper solutions
4 4 25 Years MALDI-TOF MS MALDI = Matrix-assisted Laser Desorption Ionisation TOF MS = Time of Flight Mass Spectrometry Laser beam Biomolecules embedded in matrix-molecules Protonation (Almost) intact biomolecules can be measured after being embedded in matrix molecules and ablated via a laser beam. Nobel Prize for Chemistry 2002 Laser desorption ionization of proteins with molecular masses exceeding 10,000 daltons. Karas M., Hillenkamp F. Anal Chem Oct 15; 60 (20):
5 5 MALDI-TOF Dairy Database D-MASS New Entries D-MASS Microorganisms in dairy products Farm environment: Soil, water, air Animal environment: Feed, bedding material Animal: Skin (udder, mastitis), faeces Milk collection: Milking equipment, transfer, storage Post-Pasteurization: Employees, air Very broad spectrum Useful for other applications
6 6 D-MASS Details MALDI-Biotyper D-MASS Extension of the database with e.g.: Bacillus spp. Paenibacillus spp. Geobacillus spp. Clostridium ~ 125 spp. Bacillus, Geobacillus, Paenibacillus spp., Lactobacillus ~ 20 Clostridium spp. spp. Lactococcus spp. Enterobacter ~ 40 Cronobacter spp. spp. Enterococcus ~ 20 Enterobacter spp. spp. and former Enterobacter spp. Cronobacter ~ 15 Listeria spp. and Campylobacter spp. Campylobacter spp. Listeria ~ 25 spp. Lactobacillus spp. Pseudomonas ~ 35 other spp. Lactic Acid Bacteria Staphylococcus ~ 15 Micrococcus, spp. Kocuria, Staphylococcus spp. ~ 5 Pseudomonas, Aeromonas spp. ~ 300 database entries of different strains
7 7 Classification result before and after High confidence identification Low confidence identification No identification
8 8 Classification result before and after High confidence identification Low confidence identification No identification
9 9 Stringent QM policy for new entries Start with relevant and representative strains (DSMZ, ATCC, BCD, etc.; sequence information available) 1. Extraction 3. Quality control of spectra and generation of main spectra (MSP) Verification and validation 2. Recording of protein spectra
10 10 Validation and Evaluation Control measurements & Dendrograms All requirements are met Correct identification High score values High rank in result table Good integration to other entries Entry remains in database Requirements are not met, e.g. Wrong or doubtful identification Low scores Low rank in results table Bad integration to other entries Entry is being replaced or deleted
11 11 Validation data report D-MASS Specificity Interference with previous entries Exclusion tests (no false positive IDs) Reproducibility Created and approved by A. Thomas (Final approval and release by C. Grönewald and K. Berghof-Jäger)
12 12 D-MASS Details Entries D-MASS (156species) Species Species Species Species Acinetobacter pittii Clostridium tetani Kocuria salsicia Paenibacillus jamilae Aeribacillus pallidus Clostridium tyrobutyricum Kosakonia arachidis Paenibacillus lactis Aeromonas schubertii Comamonas testosteroni Kosakonia cowanii Paenibacillus terrae Aeromonas sobria Cronobacter condimenti Kosakonia sacchari Pediococcus acidilactici Aneurinibacillus aneurinilyticus Cronobacter dublinensis Lactobacillus acetotolerans Pediococcus cellicola Aneurinibacillus migulanus Cronobacter malonaticus Lactobacillus buchneri Pediococcus claussenii Aneurinibacillus thermoaerophilus Cronobacter muytjensii Lactobacillus gasseri Pediococcus damnosus Anoxybacillus flavithermus Cronobacter sakazakii Lactobacillus harbinensis Pediococcus ethanolidurans Bacillus amyloliquefaciens Cronobacter turicensis Lactobacillus helveticus Pediococcus inopinatus Bacillus aryabhattai Cronobacter universalis Lactobacillus hilgardii Pediococcus parvulus Bacillus cereus Enterobacter asburiae Lactobacillus parabuchneri Pediococcus stilesii Bacillus circulans Enterobacter cancerogenus Lactobacillus plantarum Pluralibacter gergoviae Bacillus clausii Enterobacter hormaechei Lactobacillus rhamnosus Propionibacterium acidifaciens Bacillus coagulans Enterobacter kobei Lactobacillus rossiae Propionibacterium australiense Bacillus farraginis Enterobacter ludwigii Lactobacillus sakei Pseudomonas anguilliseptica Bacillus flexus Enterobacter massiliensis Lactobacillus spicheri Pseudomonas brassicacearum Bacillus fordii Enterobacter mori Lactobacillus zymae Pseudomonas brenneri Bacillus ginsengihumi Enterobacter xiangfangensis Lactococcus lactis Pseudomonas fragi Bacillus licheniformis Enterococcus casseliflavus Lactococcus plantarum Pseudomonas jessenii Bacillus mycoides Enterococcus durans Leclercia adecarboxylata Pseudomonas mosselii Bacillus oleronius Enterococcus faecium Lelliottia amnigena Pseudomonas taetrolens Bacillus pumilus Enterococcus raffinosus Lelliottia nimipressuralis Pseudopropionibacterium propionicum Bacillus smithii Falsibacillus pallidus Leuconostoc gelidum Psychrobacter cryohalolentis Bacillus sporothermodurans Fictibacillus barbaricus Leuconostoc mesenteroides Psychrobacter immobilis Bacillus subtilis Fictibacillus nanhaiensis Leuconostoc miyukkimchii Shewanella putrefaciens Bacillus thermoamylovorans Fictibacillus phosphorivorans Leuconostoc pseudomesenteroides Siccibacter turicensis Bacillus thuringiensis Franconibacter helveticus Listeria aquatica Staphylococcus equorum Bacillus weihenstephanensis Franconibacter pulveris Listeria booriae Staphylococcus hyicus Brevibacillus agri Geobacillus kaustophilus Listeria cornellensis Staphylococcus pseudointermedius Brevibacillus borstelensi Geobacillus stearothermophilus Listeria floridensis Staphylococcus warneri Brevibacillus centrosporus Geobacillus subterraneus Listeria grandensis Streptococcus acidominimus Campylobacter coli Geobacillus thermoglucosidasius Listeria newyorkensis Streptococcus moroccensis Campylobacter lari Geobacillus thermoleovorans Listeria riparia Streptococcus pluranimalium Campylobacter peloridis Kluyvera intermedia Lysinibacillus sphaericus Terrisporobacter glycolicus Candida sorbophila Kocuria carniphila Micrococcus cohnii Virgibacillus proomi Clostridium beijerinckii Kocuria flava Micrococcus flavus Weissella cibaria Clostridium butyricum Kocuria koreensis Micrococcus lylae Weissella confusa Clostridium chromiireducens Kocuria palustris Micrococcus terreus Weissella hellenica Clostridium estertheticum ssp. estertheticum Kocuria rhizophila Neomicrococcus lactis Clostridium pasteurianum Kocuria rosea Paenibacillus amylolyticus
13 12 Conclusions 302 new dairy relevant entries Straightforward and convenient (open library concept) Stringent Quality control Product description with species entries details Validation report on request Robust, reproducible and easy to use Excellent Fit for Industrial Microbiology
14 Thank you for your attention! BIOTECON Diagnostics GmbH Hermannswerder Potsdam For safer food BIOTECON Diagnostics: simply builds up trust Tel.: +49 (0) Fax: +49 (0)
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