{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,1,24]],"date-time":"2026-01-24T17:00:31Z","timestamp":1769274031851,"version":"3.49.0"},"reference-count":55,"publisher":"MDPI AG","issue":"3","license":[{"start":{"date-parts":[[2020,9,6]],"date-time":"2020-09-06T00:00:00Z","timestamp":1599350400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Funda\u00e7\u00e3o para a Ci\u00eancia e a Tecnologia - Portugal","award":["PTDC\/BIA-MIC\/1615\/2014"],"award-info":[{"award-number":["PTDC\/BIA-MIC\/1615\/2014"]}]},{"name":"IBB - Institute dor Bioengineering and Biosciences","award":["UIDB\/04565\/2020"],"award-info":[{"award-number":["UIDB\/04565\/2020"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Vaccines"],"abstract":"<jats:p>Infections by the Burkholderia cepacia complex (Bcc) remain seriously life threatening to cystic fibrosis (CF) patients, and no effective eradication is available. A vaccine to protect patients against Bcc infections is a highly attractive therapeutic option, but none is available. A strategy combining the bioinformatics identification of putative surface-exposed proteins with an experimental approach encompassing the \u201cshaving\u201d of surface-exposed proteins with trypsin followed by peptide identification by liquid chromatography and mass spectrometry is here reported. The methodology allowed the bioinformatics identification of 263 potentially surface-exposed proteins, 16 of them also experimentally identified by the \u201cshaving\u201d approach. Of the proteins identified, 143 have a high probability of containing B-cell epitopes that are surface-exposed. The immunogenicity of three of these proteins was demonstrated using serum samples from Bcc-infected CF patients and Western blotting, validating the usefulness of this methodology in identifying potentially immunogenic surface-exposed proteins that might be used for the development of Bcc-protective vaccines.<\/jats:p>","DOI":"10.3390\/vaccines8030509","type":"journal-article","created":{"date-parts":[[2020,9,6]],"date-time":"2020-09-06T23:12:49Z","timestamp":1599433969000},"page":"509","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":14,"title":["Characterization of the Burkholderia cenocepacia J2315 Surface-Exposed Immunoproteome"],"prefix":"10.3390","volume":"8","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-9291-9169","authenticated-orcid":false,"given":"S\u00edlvia A.","family":"Sousa","sequence":"first","affiliation":[{"name":"iBB\u2013Institute for Bioengineering and Biosciences, 1049-001 Lisbon, Portugal"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3484-3474","authenticated-orcid":false,"given":"Ant\u00f3nio M.M.","family":"Seixas","sequence":"additional","affiliation":[{"name":"iBB\u2013Institute for Bioengineering and Biosciences, 1049-001 Lisbon, Portugal"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3997-9618","authenticated-orcid":false,"given":"Manoj","family":"Mandal","sequence":"additional","affiliation":[{"name":"iBB\u2013Institute for Bioengineering and Biosciences, 1049-001 Lisbon, Portugal"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8744-7045","authenticated-orcid":false,"given":"Manuel J.","family":"Rodr\u00edguez-Ortega","sequence":"additional","affiliation":[{"name":"Departament of Biochemistry and Molecular Biology, C\u00f3rdoba University, 14071 C\u00f3rdoba, Spain"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8850-274X","authenticated-orcid":false,"given":"Jorge H.","family":"Leit\u00e3o","sequence":"additional","affiliation":[{"name":"iBB\u2013Institute for Bioengineering and Biosciences, 1049-001 Lisbon, Portugal"}]}],"member":"1968","published-online":{"date-parts":[[2020,9,6]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"2265","DOI":"10.1099\/ijs.0.000251","article-title":"Burkholderia stagnalis sp. nov. and Burkholderia territorii sp. nov., two novel Burkholderia cepacia complex species from environmental and human sources","volume":"65","author":"Mayo","year":"2015","journal-title":"Int. J. Syst. Evol. Microbiol."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"1","DOI":"10.3389\/fmicb.2017.00207","article-title":"Volcanic soils as sources of novel CO-oxidizing Paraburkholderia and Burkholderia: Paraburkholderia hiiakae sp. nov., Paraburkholderia metrosideri sp. nov., Paraburkholderia paradisi sp. nov., Paraburkholderia peleae","volume":"8","author":"Weber","year":"2017","journal-title":"Front. Microbiol."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"14","DOI":"10.1099\/ijsem.0.002293","article-title":"Burkholderia puraquae sp. nov., a novel species of the Burkholderia cepacia complex isolated from hospital settings and agricultural soils","volume":"68","author":"Martina","year":"2018","journal-title":"Int. J. Syst. Evol. Microbiol."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"e00139-19","DOI":"10.1128\/CMR.00139-19","article-title":"Burkholderia cepacia complex bacteria: A feared contamination risk in water-based pharmaceutical products","volume":"33","author":"Tavares","year":"2020","journal-title":"Clin. Microbiol. Rev."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"31","DOI":"10.1007\/s00253-010-2528-0","article-title":"Pathogenicity, virulence factors, and strategies to fight against Burkholderia cepacia complex pathogens and related species","volume":"87","author":"Sousa","year":"2010","journal-title":"Appl. Microbiol. Biotechnol."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"206","DOI":"10.1016\/S0022-3476(84)80993-2","article-title":"Pseudomonas cepacia infection in cystic fibrosis: An emerging problem","volume":"104","author":"Isles","year":"1984","journal-title":"J. Pediatr."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"1101","DOI":"10.1007\/s10096-008-0552-0","article-title":"Variation of the antimicrobial susceptibility profiles of Burkholderia cepacia complex clonal isolates obtained from chronically infected cystic fibrosis patients: A five-year survey in the major Portuguese treatment center","volume":"27","author":"Sousa","year":"2008","journal-title":"Eur. J. Clin. Microbiol. Infect. Dis."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"1592","DOI":"10.3389\/fmicb.2017.01592","article-title":"Burkholderia cenocepacia infections in cystic fibrosis patients: Drug resistance and therapeutic approaches","volume":"8","author":"Scoffone","year":"2017","journal-title":"Front. Microbiol."},{"key":"ref_9","doi-asserted-by":"crossref","unstructured":"Sousa, S.A., Feliciano, J.R., Pita, T., Guerreiro, S.I., and Leit\u00e3o, J.H. (2017). Burkholderia cepacia Complex Regulation of Virulence Gene Expression: A Review. Genes, 8.","DOI":"10.3390\/genes8010043"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"194","DOI":"10.1128\/CMR.15.2.194-222.2002","article-title":"Lung Infections Associated with Cystic Fibrosis Lung Infections Associated with Cystic Fibrosis","volume":"15","author":"Lyczak","year":"2002","journal-title":"Clin. Microbiol. Rev."},{"key":"ref_11","first-page":"CD009876","article-title":"Eradication therapy for Burkholderia cepacia complex in people with cystic fibrosis","volume":"11","author":"Regan","year":"2016","journal-title":"Cochrane Database Syst. Rev."},{"key":"ref_12","doi-asserted-by":"crossref","unstructured":"Sousa, S., Seixas, A., and Leit\u00e3o, J. (2018). Postgenomic Approaches and Bioinformatics Tools to Advance the Development of Vaccines against Bacteria of the Burkholderia cepacia Complex. Vaccines, 6.","DOI":"10.3390\/vaccines6020034"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"315","DOI":"10.1016\/j.tim.2019.12.005","article-title":"Vaccines to Overcome Antibiotic Resistance: The Challenge of Burkholderia cenocepacia","volume":"28","author":"Scoffone","year":"2020","journal-title":"Trends Microbiol."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"1424","DOI":"10.1128\/IAI.01248-15","article-title":"Linocin and OmpW Are Involved in Attachment of the Cystic Fibrosis-Associated Pathogen Burkholderia cepacia Complex to Lung Epithelial Cells and Protect Mice against Infection","volume":"84","author":"McClean","year":"2016","journal-title":"Infect. Immun."},{"key":"ref_15","doi-asserted-by":"crossref","unstructured":"Pradenas, G., Ross, B., and Torres, A. (2016). Burkholderia cepacia Complex Vaccines: Where Do We Go from here?. Vaccines, 4.","DOI":"10.3390\/vaccines4020010"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"181","DOI":"10.1016\/S0167-7799(01)01600-6","article-title":"Antibacterial vaccine design using genomics and proteomics","volume":"19","author":"Grandi","year":"2001","journal-title":"Trends Biotechnol."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"164","DOI":"10.1016\/j.jprot.2013.03.035","article-title":"Surfomics: Shaving live organisms for a fast proteomic identification of surface proteins","volume":"97","year":"2014","journal-title":"J. Proteomics"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"174","DOI":"10.1002\/elps.200900050","article-title":"Membrane proteins and proteomics: Love is possible, but so difficult","volume":"30","author":"Rabilloud","year":"2009","journal-title":"Electrophoresis"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"62","DOI":"10.1016\/j.nbt.2019.08.006","article-title":"New insights into the immunoproteome of B. cenocepacia J2315 using serum samples from cystic fibrosis patients","volume":"54","author":"Sousa","year":"2020","journal-title":"N. Biotechnol."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"131","DOI":"10.1016\/j.mimet.2009.04.013","article-title":"Identification of uropathogenic Escherichia coli surface proteins by shotgun proteomics","volume":"78","author":"Walters","year":"2009","journal-title":"J. Microbiol. Methods"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"1220","DOI":"10.1016\/j.fm.2011.04.009","article-title":"Surfome analysis of a wild-type wine Saccharomyces cerevisiae strain","volume":"28","author":"Braconi","year":"2011","journal-title":"Food Microbiol."},{"key":"ref_22","doi-asserted-by":"crossref","unstructured":"Castro-Borges, W., Dowle, A., Curwen, R.S., Thomas-Oates, J., and Wilson, R.A. (2011). Enzymatic shaving of the tegument surface of live schistosomes for proteomic analysis: A rational approach to select vaccine candidates. PLoS Negl. Trop. Dis., 5.","DOI":"10.1371\/journal.pntd.0000993"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"317","DOI":"10.1172\/JCI0213870","article-title":"Effects of reduced mucus oxygen concentration in airway Pseudomonas infections of cystic fibrosis patients","volume":"109","author":"Worlitzsch","year":"2002","journal-title":"J. Clin. Investig."},{"key":"ref_24","first-page":"1","article-title":"The unexpected discovery of a novel low-oxygen-activated locus for the anoxic persistence of Burkholderia cenocepacia","volume":"14","author":"Sass","year":"2013","journal-title":"ISME J."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"539","DOI":"10.1128\/mr.60.3.539-574.1996","article-title":"Microbial pathogenesis in cystic fibrosis: Mucoid Pseudomonas aeruginosa and Burkholderia cepacia","volume":"60","author":"Govan","year":"1996","journal-title":"Microbiol. Rev."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1186\/s13568-016-0212-1","article-title":"The Burkholderia cenocepacia OmpA-like protein BCAL2958: Identification, characterization, and detection of anti-BCAL2958 antibodies in serum from B. cepacia complex-infected Cystic Fibrosis patients","volume":"6","author":"Sousa","year":"2016","journal-title":"AMB Express"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"21","DOI":"10.1007\/978-1-4939-7553-2_2","article-title":"\u2018Shaving\u2019 Live Bacterial Cells with Proteases for Proteomic Analysis of Surface Proteins","volume":"Volume 1722","year":"2018","journal-title":"Methods in Molecular Biology (Clifton, N.J.)"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"667","DOI":"10.1099\/jmm.0.45969-0","article-title":"Microcolony formation: A novel biofilm model of Pseudomonas aeruginosa for the cystic fibrosis lung","volume":"54","author":"Sriramulu","year":"2005","journal-title":"J. Med. Microbiol."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"1852","DOI":"10.1038\/nprot.2006.288","article-title":"Silver staining of proteins in polyacrylamide gels","volume":"1","author":"Chevallet","year":"2006","journal-title":"Nat. Protoc."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"2803","DOI":"10.1093\/bioinformatics\/btn524","article-title":"The Burkholderia Genome Database: Facilitating flexible queries and comparative analyses","volume":"24","author":"Winsor","year":"2008","journal-title":"Bioinformatics"},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"D427","DOI":"10.1093\/nar\/gky995","article-title":"The Pfam protein families database in 2019","volume":"47","author":"Mistry","year":"2019","journal-title":"Nucleic Acids Res."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"1608","DOI":"10.1093\/bioinformatics\/btq249","article-title":"PSORTb 3.0: Improved protein subcellular localization prediction with refined localization subcategories and predictive capabilities for all prokaryotes","volume":"26","author":"Yu","year":"2010","journal-title":"Bioinformatics"},{"key":"ref_33","unstructured":"Green, M.R., and Sambrook, J. (2012). Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press."},{"key":"ref_34","unstructured":"(2020, July 30). Burkholderia cenocepacia\u2014Wellcome Sanger Institute. Available online: https:\/\/www.sanger.ac.uk\/resources\/downloads\/bacteria\/burkholderia-cenocepacia.html."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"W24","DOI":"10.1093\/nar\/gkx346","article-title":"BepiPred-2.0: Improving sequence-based B-cell epitope prediction using conformational epitopes","volume":"45","author":"Jespersen","year":"2017","journal-title":"Nucleic Acids Res."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1111\/imm.12213","article-title":"Discovery of protective B-cell epitopes for development of antimicrobial vaccines and antibody therapeutics","volume":"142","author":"Sharon","year":"2014","journal-title":"Immunology"},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"3733","DOI":"10.1016\/j.jprot.2012.04.037","article-title":"Another turn of the screw in shaving Gram-positive bacteria: Optimization of proteomics surface protein identification in Streptococcus pneumoniae","volume":"75","author":"Obando","year":"2012","journal-title":"J. Proteomics"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"277","DOI":"10.1093\/bioinformatics\/btl595","article-title":"A predictive model for identifying proteins by a single peptide match","volume":"23","author":"Higdon","year":"2007","journal-title":"Bioinformatics"},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"277","DOI":"10.1093\/nar\/gku954","article-title":"MoonProt: A database for proteins that are known to moonlight","volume":"43","author":"Mani","year":"2014","journal-title":"Nucleic Acids Res."},{"key":"ref_40","unstructured":"(2020, July 30). The Burkholderia Genome Database. Available online: https:\/\/www.burkholderia.com\/."},{"key":"ref_41","unstructured":"(2020, July 30). Pfam Database. Available online: http:\/\/pfam.xfam.org\/."},{"key":"ref_42","unstructured":"(2020, July 30). BepiPred 2.0 Server. Available online: http:\/\/tools.iedb.org\/bcell\/."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"261","DOI":"10.1128\/JB.01230-08","article-title":"The genome of Burkholderia cenocepacia J2315, an epidemic pathogen of Cystic Fibrosis patients","volume":"191","author":"Holden","year":"2009","journal-title":"J. Bacteriol."},{"key":"ref_44","doi-asserted-by":"crossref","unstructured":"Spencer, H.K., Spitznogle, S.L., Borjan, J., and Aitken, S.L. (2020). An Overview of the Treatment of Less Common Non-Lactose Fermenting Gram Negative Bacteria. Pharmacother. J. Hum. Pharmacol. Drug Ther., accepted.","DOI":"10.1002\/phar.2447"},{"key":"ref_45","doi-asserted-by":"crossref","unstructured":"Pradenas, G., Myers, J., and Torres, A. (2017). Characterization of the Burkholderia cenocepacia TonB Mutant as a Potential Live Attenuated Vaccine. Vaccines, 5.","DOI":"10.3390\/vaccines5040033"},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"6041","DOI":"10.4049\/jimmunol.1402273","article-title":"CD4 + T Cell Epitopes of FliC Conserved between Strains of Burkholderia: Implications for Vaccines against Melioidosis and Cepacia Complex in Cystic Fibrosis","volume":"193","author":"Musson","year":"2014","journal-title":"J. Immunol."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"8","DOI":"10.2174\/1574893608999140109121908","article-title":"Bioinformatics: A Molecular Microbiologist\u2019s Perspective","volume":"9","author":"Sousa","year":"2014","journal-title":"Curr. Bioinform."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"75","DOI":"10.1007\/s00248-014-0460-z","article-title":"Protein Profiling Analyses of the Outer Membrane of Burkholderia cenocepacia Reveal a Niche-Specific Proteome","volume":"69","author":"Liu","year":"2015","journal-title":"Microb. Ecol."},{"key":"ref_49","doi-asserted-by":"crossref","unstructured":"Wolden, R., Pain, M., Karlsson, R., Karlsson, A., Aarag Fredheim, E.G., and Cavanagh, J.P. (2020). Identification of surface proteins in a clinical Staphylococcus haemolyticus isolate by bacterial surface shaving. BMC Microbiol., 20.","DOI":"10.1186\/s12866-020-01778-8"},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"47","DOI":"10.1007\/978-1-4939-3676-2_4","article-title":"Cell shaving and False-Positive control strategies coupled to novel statistical tools to profile Gram-Positive bacterial surface proteomes","volume":"Volume 1440","author":"Solis","year":"2016","journal-title":"Methods in Molecular Biology"},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"3476","DOI":"10.1128\/IAI.00179-11","article-title":"Bacterial virulence in the moonlight: Multitasking bacterial moonlighting proteins are virulence determinants in infectious disease","volume":"79","author":"Henderson","year":"2011","journal-title":"Infect. Immun."},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"3644","DOI":"10.1016\/j.vaccine.2013.05.100","article-title":"Outer membrane protein A (OmpA) from Shigella flexneri 2a: A promising subunit vaccine candidate","volume":"31","author":"Pore","year":"2013","journal-title":"Vaccine"},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"207","DOI":"10.1016\/j.vetmic.2012.08.019","article-title":"The OmpA family of proteins: Roles in bacterial pathogenesis and immunity","volume":"163","author":"Confer","year":"2013","journal-title":"Vet. Microbiol."},{"key":"ref_54","doi-asserted-by":"crossref","unstructured":"Darwin, A.J. (2013). Stress relief during host infection: The phage shock protein response supports bacterial virulence in various ways. PLoS Pathog., 9.","DOI":"10.1371\/journal.ppat.1003388"},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"2192","DOI":"10.1099\/mic.0.000175","article-title":"Evaluating the role of phage-shock protein A in Burkholderia pseudomallei","volume":"161","author":"Southern","year":"2015","journal-title":"Microbiology"}],"container-title":["Vaccines"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2076-393X\/8\/3\/509\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T10:07:22Z","timestamp":1760177242000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2076-393X\/8\/3\/509"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2020,9,6]]},"references-count":55,"journal-issue":{"issue":"3","published-online":{"date-parts":[[2020,9]]}},"alternative-id":["vaccines8030509"],"URL":"https:\/\/doi.org\/10.3390\/vaccines8030509","relation":{},"ISSN":["2076-393X"],"issn-type":[{"value":"2076-393X","type":"electronic"}],"subject":[],"published":{"date-parts":[[2020,9,6]]}}}