{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,3,27]],"date-time":"2026-03-27T19:23:35Z","timestamp":1774639415757,"version":"3.50.1"},"reference-count":47,"publisher":"MDPI AG","issue":"10","license":[{"start":{"date-parts":[[2022,10,6]],"date-time":"2022-10-06T00:00:00Z","timestamp":1665014400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"German Federal Ministry of Education and Research within the framework of the project Ess-B.A.R","award":["13N13983"],"award-info":[{"award-number":["13N13983"]}]},{"name":"German Federal Ministry of Education and Research within the framework of the project Ess-B.A.R","award":["13N13984"],"award-info":[{"award-number":["13N13984"]}]},{"name":"German Federal Ministry of Education and Research within the framework of the project Ess-B.A.R","award":["13N13985"],"award-info":[{"award-number":["13N13985"]}]},{"name":"Max Planck Society","award":["13N13983"],"award-info":[{"award-number":["13N13983"]}]},{"name":"Max Planck Society","award":["13N13984"],"award-info":[{"award-number":["13N13984"]}]},{"name":"Max Planck Society","award":["13N13985"],"award-info":[{"award-number":["13N13985"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Life"],"abstract":"<jats:p>Anthrax is a zoonotic infection caused by the bacterium Bacillus anthracis (BA). Specific identification of this pathogen often relies on targeting genes located on two extrachromosomal plasmids, which represent the major pathogenicity factors of BA. However, more recent findings show that these plasmids have also been found in other closely related Bacillus species. In this study, we investigated the possibility of identifying species-specific and universally applicable marker peptides for BA. For this purpose, we applied a high-resolution mass spectrometry-based approach for 42 BA isolates. Along with the genomic sequencing data and by developing a bioinformatics data evaluation pipeline, which uses a database containing most of the publicly available protein sequences worldwide (UniParc), we were able to identify eleven universal marker peptides unique to BA. These markers are located on the chromosome and therefore, might overcome known problems, such as observable loss of plasmids in environmental species, plasmid loss during cultivation in the lab, and the fact that the virulence plasmids are not necessarily a unique feature of BA. The identified chromosomally encoded markers in this study could extend the small panel of already existing chromosomal targets and along with targets for the virulence plasmids, may pave the way to an even more reliable identification of BA using genomics- as well as proteomics-based techniques.<\/jats:p>","DOI":"10.3390\/life12101549","type":"journal-article","created":{"date-parts":[[2022,10,9]],"date-time":"2022-10-09T01:43:11Z","timestamp":1665279791000},"page":"1549","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":2,"title":["Identification of Universally Applicable and Species-Specific Marker Peptides for Bacillus anthracis"],"prefix":"10.3390","volume":"12","author":[{"given":"Natalie","family":"Witt","sequence":"first","affiliation":[{"name":"Mass Spectrometry Facility, Max Planck Institute for Molecular Genetics, Ihnestra\u00dfe 63-73, 14195 Berlin, Germany"},{"name":"Bioanalytics, Institute of Biotechnology, Technische Universit\u00e4t Berlin, 13355 Berlin, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1159-2801","authenticated-orcid":false,"given":"Domenico","family":"Galante","sequence":"additional","affiliation":[{"name":"Istituto Zooprofilattico Sperimentale della Puglia e della Basilicata, Anthrax Reference Institute of Italy, Via Manfredonia 20, 71121 Foggia, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7678-9720","authenticated-orcid":false,"given":"Sandro","family":"Andreotti","sequence":"additional","affiliation":[{"name":"Bioinformatics Solution Center, Department of Mathematics and Computer Science, Freie Universit\u00e4t Berlin, 14195 Berlin, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2068-6081","authenticated-orcid":false,"given":"Mostafa","family":"Abdel Glil","sequence":"additional","affiliation":[{"name":"Friedrich-Loeffler-Institut, Institute of Bacterial Infections and Zoonoses, Naumburger Str. 96 a, 07743 Jena, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Antonio","family":"Fasanella","sequence":"additional","affiliation":[{"name":"Istituto Zooprofilattico Sperimentale della Puglia e della Basilicata, Anthrax Reference Institute of Italy, Via Manfredonia 20, 71121 Foggia, Italy"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"David","family":"Meierhofer","sequence":"additional","affiliation":[{"name":"Mass Spectrometry Facility, Max Planck Institute for Molecular Genetics, Ihnestra\u00dfe 63-73, 14195 Berlin, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6657-833X","authenticated-orcid":false,"given":"Herbert","family":"Tomaso","sequence":"additional","affiliation":[{"name":"Friedrich-Loeffler-Institut, Institute of Bacterial Infections and Zoonoses, Naumburger Str. 96 a, 07743 Jena, Germany"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2022,10,6]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"815","DOI":"10.1056\/NEJM199909093411107","article-title":"Anthrax","volume":"341","author":"Dixon","year":"1999","journal-title":"N. Engl. J. Med."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"237","DOI":"10.1016\/j.mimet.2011.12.001","article-title":"Developing an Integrated Proteo-Genomic Approach for the Characterisation of Biomarkers for the Identification of Bacillus anthracis","volume":"88","author":"Misra","year":"2012","journal-title":"J. Microbiol. Methods"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"225","DOI":"10.3201\/eid0802.010164","article-title":"Public Health Assessment of Potential Biological Terrorism Agents","volume":"8","author":"Rotz","year":"2002","journal-title":"Emerg. Infect. Dis."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"451","DOI":"10.1146\/annurev.micro.091208.073255","article-title":"What Sets Bacillus anthracis Apart from Other Bacillus Species?","volume":"63","author":"Tourasse","year":"2009","journal-title":"Annu. Rev. Microbiol."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"8675","DOI":"10.1021\/ac2020992","article-title":"Sensitive Detection of Bacillus anthracis Spores by Immunocapture and Liquid Chromatography\u2013Tandem Mass Spectrometry","volume":"83","author":"Chenau","year":"2011","journal-title":"Anal. Chem."},{"key":"ref_6","doi-asserted-by":"crossref","unstructured":"Zasada, A.A. (2020). Detection and Identification of Bacillus anthracis: From Conventional to Molecular Microbiology Methods. Microorganisms, 8.","DOI":"10.3390\/microorganisms8010125"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"e00065-18","DOI":"10.1128\/mSystems.00065-18","article-title":"Genomic Characterization and Copy Number Variation of Bacillus anthracis Plasmids PXO1 and PXO2 in a Historical Collection of 412 Strains","volume":"3","author":"Marston","year":"2018","journal-title":"mSystems"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"716","DOI":"10.1074\/mcp.M113.032946","article-title":"Identification and Validation of Specific Markers of Bacillus anthracis Spores by Proteomics and Genomics Approaches*","volume":"13","author":"Chenau","year":"2014","journal-title":"Mol. Cell. Proteom."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"1411","DOI":"10.1007\/s00253-011-3845-7","article-title":"Rapid Detection Methods for Bacillus anthracis in Environmental Samples: A Review","volume":"93","author":"Irenge","year":"2012","journal-title":"Appl. Microbiol. Biotechnol."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"e00917-20","DOI":"10.1128\/MRA.00917-20","article-title":"Complete Genome Sequence of an Environmental Bacillus cereus Isolate Belonging to the Bacillus anthracis Clade","volume":"9","author":"Irenge","year":"2020","journal-title":"Microbiol. Resour. Announc."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"134","DOI":"10.1128\/JB.184.1.134-141.2002","article-title":"Bacillus anthracis PXO1 Plasmid Sequence Conservation among Closely Related Bacterial Species","volume":"184","author":"Pannucci","year":"2002","journal-title":"J. Bacteriol."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"8449","DOI":"10.1073\/pnas.0402414101","article-title":"Identification of Anthrax Toxin Genes in a Bacillus cereus Associated with an Illness Resembling Inhalation Anthrax","volume":"101","author":"Hoffmaster","year":"2004","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"3352","DOI":"10.1128\/JCM.00561-06","article-title":"Characterization of Bacillus cereus Isolates Associated with Fatal Pneumonias: Strains Are Closely Related to Bacillus anthracis and Harbor B. anthracis Virulence Genes","volume":"44","author":"Hoffmaster","year":"2006","journal-title":"J. Clin. Microbiol."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"673","DOI":"10.1111\/j.1365-2672.2006.02809.x","article-title":"Evaluation of Different Methods to Discriminate Bacillus anthracis from Other Bacteria of the Bacillus cereus Group","volume":"100","author":"Klee","year":"2006","journal-title":"J. Appl. Microbiol."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"3236","DOI":"10.1093\/bioinformatics\/bth191","article-title":"UniProt Archive","volume":"20","author":"Leinonen","year":"2004","journal-title":"Bioinformatics"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"e00219-16","DOI":"10.1128\/genomeA.00219-16","article-title":"Genome Sequence of Bacillus anthracis Strain Stendal, Isolated from an Anthrax Outbreak in Cattle in Germany","volume":"4","author":"Antwerpen","year":"2016","journal-title":"Genome Announc."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"e01002-17","DOI":"10.1128\/genomeA.01002-17","article-title":"High-Quality Genome Sequence of Bacillus anthracis Strain 14RA5914 Isolated during an Outbreak in Germany in 2014","volume":"5","author":"Elschner","year":"2017","journal-title":"Genome Announc."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"e02889-20","DOI":"10.1128\/JCM.02889-20","article-title":"A Whole-Genome-Based Gene-by-Gene Typing System for Standardized High-Resolution Strain Typing of Bacillus anthracis","volume":"59","author":"Chiaverini","year":"2021","journal-title":"J. Clin. Microbiol."},{"key":"ref_19","doi-asserted-by":"crossref","unstructured":"Van Ert, M.N., Easterday, W.R., Huynh, L.Y., Okinaka, R.T., Hugh-Jones, M.E., Ravel, J., Zanecki, S.R., Pearson, T., Simonson, T.S., and U\u2019Ren, J.M. (2007). Global Genetic Population Structure of Bacillus anthracis. PLoS ONE, 2.","DOI":"10.1371\/journal.pone.0000461"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"S137","DOI":"10.1016\/j.ijfoodmicro.2010.07.039","article-title":"A Multiplex Real-Time PCR for Identifying and Differentiating B. anthracis Virulent Types","volume":"145","author":"Wielinga","year":"2011","journal-title":"Int. J. Food Microbiol."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"455","DOI":"10.1089\/cmb.2012.0021","article-title":"SPAdes: A New Genome Assembly Algorithm and Its Applications to Single-Cell Sequencing","volume":"19","author":"Bankevich","year":"2012","journal-title":"J. Comput. Biol."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"2068","DOI":"10.1093\/bioinformatics\/btu153","article-title":"Prokka: Rapid Prokaryotic Genome Annotation","volume":"30","author":"Seemann","year":"2014","journal-title":"Bioinformatics"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"2144","DOI":"10.1021\/pr8010099","article-title":"Statistical Design of Quantitative Mass Spectrometry-Based Proteomic Experiments","volume":"8","author":"Oberg","year":"2009","journal-title":"J. Proteome Res."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"636","DOI":"10.3389\/fmicb.2020.00636","article-title":"Rapid and Culture Free Identification of Francisella in Hare Carcasses by High-Resolution Tandem Mass Spectrometry Proteotyping","volume":"11","author":"Witt","year":"2020","journal-title":"Front. Microbiol."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"14082","DOI":"10.1038\/srep14082","article-title":"Genomic Insights into the Taxonomic Status of the Bacillus cereus Group","volume":"5","author":"Liu","year":"2015","journal-title":"Sci. Rep."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"741","DOI":"10.1038\/nmeth.3959","article-title":"OpenMS: A Flexible Open-Source Software Platform for Mass Spectrometry Data Analysis","volume":"13","author":"Sachsenberg","year":"2016","journal-title":"Nat. Methods"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"5277","DOI":"10.1038\/ncomms6277","article-title":"MS-GF+ Makes Progress towards a Universal Database Search Tool for Proteomics","volume":"5","author":"Kim","year":"2014","journal-title":"Nat. Commun."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"3150","DOI":"10.1093\/bioinformatics\/bts565","article-title":"CD-HIT: Accelerated for Clustering the next-Generation Sequencing Data","volume":"28","author":"Fu","year":"2012","journal-title":"Bioinformatics"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"1731","DOI":"10.3389\/fmicb.2020.01731","article-title":"You Can\u2019t B. cereus\u2014A Review of Bacillus cereus Strains That Cause Anthrax-like Disease","volume":"11","author":"Baldwin","year":"2020","journal-title":"Front. Microbiol."},{"key":"ref_30","doi-asserted-by":"crossref","unstructured":"Rondinone, V., Serrecchia, L., Parisi, A., Fasanella, A., Manzulli, V., Cipolletta, D., and Galante, D. (2020). Genetic Characterization of Bacillus anthracis Strains Circulating in Italy from 1972 to 2018. PLoS ONE, 15.","DOI":"10.1371\/journal.pone.0227875"},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"73","DOI":"10.1007\/s00216-005-0090-x","article-title":"Bacillus anthracis: Toxicology, Epidemiology and Current Rapid-Detection Methods","volume":"384","author":"Edwards","year":"2006","journal-title":"Anal. Bioanal. Chem."},{"key":"ref_32","unstructured":"(2020). Bacillus cereus-Bakterien in Lebensmitteln k\u00f6nnen Magen-Darm-Erkrankungen verursachen: Stellungnahme Nr. 048\/2020 des BfR vom 30. Oktober 2020. BfR-Stellungnahmen, Bundesinst. f\u00fcr Risikobewertung."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"3098","DOI":"10.1021\/acs.jproteome.6b00216","article-title":"Bacillus cereus Group-Type Strain-Specific Diagnostic Peptides","volume":"15","author":"Pfrunder","year":"2016","journal-title":"J. Proteome Res."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"83","DOI":"10.1016\/j.biochi.2021.10.003","article-title":"Bacillus thuringiensis: From Biopesticides to Anticancer Agents","volume":"192","author":"Santos","year":"2022","journal-title":"Biochimie"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"82","DOI":"10.1038\/nature23309","article-title":"Persistent Anthrax as a Major Driver of Wildlife Mortality in a Tropical Rainforest","volume":"548","author":"Hoffmann","year":"2017","journal-title":"Nature"},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"5333","DOI":"10.1128\/JB.00303-06","article-title":"Characterization of Bacillus anthracis-Like Bacteria Isolated from Wild Great Apes from C\u00f4te d\u2019Ivoire and Cameroon","volume":"188","author":"Klee","year":"2006","journal-title":"J. Bacteriol."},{"key":"ref_37","doi-asserted-by":"crossref","unstructured":"Marston, C.K., Ibrahim, H., Lee, P., Churchwell, G., Gumke, M., Stanek, D., Gee, J.E., Boyer, A.E., Gallegos-Candela, M., and Barr, J.R. (2016). Anthrax Toxin-Expressing Bacillus cereus Isolated from an Anthrax-like Eschar. PLoS ONE, 11.","DOI":"10.1371\/journal.pone.0156987"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"671","DOI":"10.4161\/viru.26288","article-title":"In Silico and in Vitro Evaluation of PCR-Based Assays for the Detection of Bacillus anthracis Chromosomal Signature Sequences","volume":"4","author":"Hamidjaja","year":"2013","journal-title":"Virulence"},{"key":"ref_39","doi-asserted-by":"crossref","unstructured":"Braun, P., Nguyen, M.D.-T., Walter, M.C., and Grass, G. (2021). Ultrasensitive Detection of Bacillus anthracis by Real-Time PCR Targeting a Polymorphism in Multi-Copy 16S RRNA Genes and Their Transcripts. Int. J. Mol. Sci., 22.","DOI":"10.1101\/2021.09.20.21263746"},{"key":"ref_40","doi-asserted-by":"crossref","unstructured":"Bremer, H., and Dennis, P.P. (2008). Modulation of Chemical Composition and Other Parameters of the Cell at Different Exponential Growth Rates. EcoSal Plus, 3.","DOI":"10.1128\/ecosal.5.2.3"},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"2205","DOI":"10.1038\/s41598-020-59156-3","article-title":"Elucidation of Protein Biomarkers for Verification of Selected Biological Warfare Agents Using Tandem Mass Spectrometry","volume":"10","author":"Rajoria","year":"2020","journal-title":"Sci. Rep."},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"5199","DOI":"10.1002\/pmic.200600314","article-title":"Early Events of Bacillus anthracis Germination Identified by Time-Course Quantitative Proteomics","volume":"6","author":"Jagtap","year":"2006","journal-title":"Proteomics"},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"7601","DOI":"10.1128\/JB.01054-08","article-title":"The Bacillus anthracis SleL (YaaH) Protein Is an N-Acetylglucosaminidase Involved in Spore Cortex Depolymerization","volume":"190","author":"Lambert","year":"2008","journal-title":"J. Bacteriol."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"1359","DOI":"10.1099\/mic.0.056630-0","article-title":"In Vitro and in Vivo Analyses of the Bacillus anthracis Spore Cortex Lytic Protein SleL","volume":"158","author":"Lambert","year":"2012","journal-title":"Microbiology"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"313","DOI":"10.1016\/j.mcp.2008.06.001","article-title":"Real-Time PCR System Targeting a Chromosomal Marker Specific for Bacillus anthracis","volume":"22","author":"Antwerpen","year":"2008","journal-title":"Mol. Cell. Probes"},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"301","DOI":"10.1016\/j.femsim.2004.10.005","article-title":"Rapid Genotypic Detection of Bacillus anthracis and the Bacillus cereus Group by Multiplex Real-Time PCR Melting Curve Analysis","volume":"43","author":"Kim","year":"2005","journal-title":"FEMS Immunol. Med. Microbiol."},{"key":"ref_47","doi-asserted-by":"crossref","unstructured":"Janse, I., Hamidjaja, R.A., Bok, J.M., and van Rotterdam, B.J. (2010). Reliable Detection of Bacillus anthracis, Francisella Tularensis and Yersinia Pestis by Using Multiplex QPCR Including Internal Controls for Nucleic Acid Extraction and Amplification. BMC Microbiol., 10.","DOI":"10.1186\/1471-2180-10-314"}],"container-title":["Life"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2075-1729\/12\/10\/1549\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T00:47:14Z","timestamp":1760143634000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2075-1729\/12\/10\/1549"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2022,10,6]]},"references-count":47,"journal-issue":{"issue":"10","published-online":{"date-parts":[[2022,10]]}},"alternative-id":["life12101549"],"URL":"https:\/\/doi.org\/10.3390\/life12101549","relation":{},"ISSN":["2075-1729"],"issn-type":[{"value":"2075-1729","type":"electronic"}],"subject":[],"published":{"date-parts":[[2022,10,6]]}}}