title: Comprehensive Identification of Immunodominant Proteins of Brucella abortus and Brucella melitensis Using Antibodies in the Sera from Naturally Infected Hosts.
abstract: Brucellosis is a debilitating zoonotic disease that affects humans and animals. The diagnosis of brucellosis is challenging, as accurate species level identification is not possible with any of the currently available serology-based diagnostic methods. The present study aimed at identifying Brucella (B.) species-specific proteins from the closely related species B. abortus and B. melitensis using sera collected from naturally infected host species. Unlike earlier reported investigations with either laboratory-grown species or vaccine strains, in the present study, field strains were utilized for analysis. The label-free quantitative proteomic analysis of the naturally isolated strains of these two closely related species revealed 402 differentially expressed proteins, among which 63 and 103 proteins were found exclusively in the whole cell extracts of B. abortus and B. melitensis field strains, respectively. The sera from four different naturally infected host species, i.e., cattle, buffalo, sheep, and goat were applied to identify the immune-binding protein spots present in the whole protein extracts from the isolated B. abortus and B. melitensis field strains and resolved on two-dimensional gel electrophoresis. Comprehensive analysis revealed that 25 proteins of B. abortus and 20 proteins of B. melitensis were distinctly immunoreactive. Dihydrodipicolinate synthase, glyceraldehyde-3-phosphate dehydrogenase and lactate/malate dehydrogenase from B. abortus, amino acid ABC transporter substrate-binding protein from B. melitensis and fumarylacetoacetate hydrolase from both species were reactive with the sera of all the tested naturally infected host species. The identified proteins could be used for the design of serological assays capable of detecting pan-Brucella, B. abortus- and B. melitensis-specific antibodies.
authors.affiliation: Institute of Animal Hygiene and Environmental Health, Centre for Infectious Medicine, Freie Universität Berlin, Robert-von-Ostertag-Str. 7-13, Berlin 14163, Germany. Gamal.Wareth@fli.bund.de.|Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Institute of Bacterial Infections and Zoonoses, Naumburger Str. 96a, Jena 07743, Germany. Gamal.Wareth@fli.bund.de.|Faculty of Veterinary Medicine, Benha University, Moshtohor, Toukh 13736, Egypt. Gamal.Wareth@fli.bund.de.
authors.forename: Gamal
authors.initials: G
authors.lastname: Wareth
authors.affiliation: Institute of Chemistry and Biochemistry, Freie Universität Berlin, Thielallee 63, Berlin 14195, Germany. eravci@zedat.fu-berlin.de.
authors.forename: Murat
authors.initials: M
authors.lastname: Eravci
authors.affiliation: Institute of Chemistry and Biochemistry, Freie Universität Berlin, Thielallee 63, Berlin 14195, Germany. chris.weise@biochemie.fu-berlin-de.
authors.forename: Christoph
authors.initials: C
authors.lastname: Weise
authors.affiliation: Institute of Animal Hygiene and Environmental Health, Centre for Infectious Medicine, Freie Universität Berlin, Robert-von-Ostertag-Str. 7-13, Berlin 14163, Germany. Uwe.Roesler@fu-berlin.de.
authors.forename: Uwe
authors.initials: U
authors.lastname: Roesler
authors.affiliation: Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Institute of Bacterial Infections and Zoonoses, Naumburger Str. 96a, Jena 07743, Germany. Falk.Melzer@fli.bund.de.
authors.forename: Falk
authors.initials: F
authors.lastname: Melzer
authors.affiliation: Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Institute of Bacterial Infections and Zoonoses, Naumburger Str. 96a, Jena 07743, Germany. Lisa.Sprague@fli.bund.de.
authors.forename: Lisa D
authors.initials: LD
authors.lastname: Sprague
authors.affiliation: Friedrich-Loeffler-Institut, Federal Research Institute for Animal Health, Institute of Bacterial Infections and Zoonoses, Naumburger Str. 96a, Jena 07743, Germany. Heinrich.Neubauer@fli.bund.de.
authors.forename: Heinrich
authors.initials: H
authors.lastname: Neubauer
authors.affiliation: Institute of Animal Hygiene and Environmental Health, Centre for Infectious Medicine, Freie Universität Berlin, Robert-von-Ostertag-Str. 7-13, Berlin 14163, Germany. jayaseelan.murugaiyan@fu-berlin.de.
authors.forename: Jayaseelan
authors.initials: J
authors.lastname: Murugaiyan
mesh_terms: D000818:Animals; D000906:Antibodies / Q000097:blood / Q000276:immunology*; D000942:Antigens, Bacterial / Q000276:immunology / Q000378:metabolism*; D001426:Bacterial Proteins / Q000276:immunology / Q000378:metabolism*; D015153:Blotting, Western; D002003:Brucella abortus / Q000302:isolation & purification / Q000378:metabolism*; D017347:Brucella melitensis / Q000302:isolation & purification / Q000378:metabolism*; D002007:Brucellosis, Bovine / Q000382:microbiology* / Q000473:pathology; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D015180:Electrophoresis, Gel, Two-Dimensional; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000276:immunology / Q000378:metabolism; D006836:Hydro-Lyases / Q000276:immunology / Q000378:metabolism; D020543:Proteome / Q000032:analysis; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D000906:Antibodies; D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D020543:Proteome; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D006836:Hydro-Lyases; C020040:4-hydroxy-tetrahydrodipicolinate synthase
keywords: 2D-PAGE; Brucella; Liquid chromatography–mass spectrometry (LC-MS); Matrix-assisted laser desorption/ionization-Tine of Flight-Mass Spectrometry MALDI-TOF MS; Western blot; host specificity; mass spectrometry; proteomics; two dimensional electrophoresis