Comparative Proteome Analysis ofBrucella melitensisVaccine Strain Rev 1 and a Virulent Strain, 16M

Abstract

The genus Brucella consists of bacterial pathogens that cause brucellosis, a major zoonotic disease characterized by undulant fever and neurological disorders in humans. Among the different Brucella species, Brucella melitensis is considered the most virulent. Despite successful use in animals, the vaccine strains remain infectious for humans. To understand the mechanism of virulence in B. melitensis, the proteome of vaccine strain Rev 1 was analyzed by two-dimensional gel electrophoresis and compared to that of virulent strain 16M. The two strains were grown under identical laboratory conditions. Computer-assisted analysis of the two B. melitensis proteomes revealed proteins expressed in either 16M or Rev 1, as well as up-or down-regulation of proteins specific for each of these strains. These proteins were identified by peptide mass fingerprinting. It was found that certain metabolic pathways may be deregulated in Rev 1. Expression of an immunogenic 31-kDa outer membrane protein, proteins utilized for iron acquisition, and those that play a role in sugar binding, lipid degradation, and amino acid binding was altered in Rev 1.Brucellosis is a major infectious disease afflicting humans and a wide range of domesticated animals worldwide. The disease is caused by several Brucella species, which are aerobic, nonmotile, gram-negative, facultative intracellular coccobacilli. The genus Brucella belongs to the ␣-2 subgroup of the class Proteobacteria. It is subdivided, on the basis of its pathogenicity and host preference, into six nomen species: Brucella abortus, B. canis, B. melitensis, B. neotomae, B. ovis, and B. suis (12). In addition, a new strain affecting marine mammals was recently isolated and tentatively named B. maris (32). On the basis of DNA hybridization data, it was suggested that all of these organisms be placed into a single species, B. melitensis (39). Among the various nomen species, B. abortus, B. canis, B. suis, B. maris, and B. melitensis have been reported to infect humans (21,35,13). B. melitensis is a pathogen of goats and sheep and is considered the most virulent species for humans. Human infection can result from either occupational contact or ingestion of contaminated food.Vaccination and eradication of infected hosts have been key factors in the control of brucellosis. Rev 1, an attenuated strain of virulent B. melitensis, was developed in 1957 (19). It is considered the most effective vaccine for the control of brucellosis in small ruminants and was used in comprehensive vaccination programs in many countries, including Saudi Arabia, Kuwait, Mongolia, Spain, and Turkey (8).Our laboratory has been involved in a comprehensive analysis of the B. melitensis 16M proteome, and initial results have been published recently (40). Previous proteomics studies using B. melitensis cells grown under different conditions have been reported (36, 37), and initial work on the B. abortus proteome has been described (26, 29). A comparative study was conducted with B. abortus vaccine strains S19 and RB51 and...

The mglA gene and its flanking regions in Brucella: The role of mglA in tolerance to hostile environments, Fe-metabolism and in vivo persistence

“…With respect to functional consequences of differences in this gene region, it is worth mentioning that a gene in immediate neighbourhood and functionally related to mglA, the D-galactose-binding periplasmic protein precursor (BMEII0983), has been found to be underexpressed in the attenuated vaccine strain B. melitensis Rev1 ( Eschenbrenner et al, 2002) and recently in B. abortus 2308 ( Lamontagne et al, 2009). Furthermore, the gene nnrA (BMEII0986), which is an immediate neighbour of the operon and potentially involved in its regulation ( Fig.…”

Comparative Secretome Analyses of Three Bacillus anthracis Strains with Variant Plasmid Contents

“…Such investigations often rely on the coupling of two-dimensional gel electrophoresis (2-DE) and matrix-assisted laser desorption ionization-mass spectrometry (MALDI-MS) to ensure accurate and sensitive analysis of proteins. In our laboratory, this approach was applied to a comprehensive study of the differences in protein expression between a fully virulent and a vaccinal strain of Brucella melitensis (14). Recently, focus has been on the analysis and characterization of the secretomes, i.e., the entire complement of secreted proteins, of pathogenic bacterial species.…”

Genomic analysis of the original Elberg Brucella melitensis Rev.1 vaccine strain reveals insights into virulence attenuation

“…Next, we compared our list of SNP-containing genes to a list of genes encoding for proteins that were previously reported to be under-expressed in Rev.1, as compared with 16M [12]. Out of the 25 reported under-expressed proteins, we detected five ORFs containing various SNPs (Table 3) that may be involved in the under-expression of these genes.…”

“…This protein had previously been reported to be one of the four proteins uniquely expressed by the B. melitensis 16M strain, as compared with Rev.1 [12]. A recent study determined the role of PrsA – a peptidyl-prolyl cis-trans isomerase – in the folding of Bacillus subtilis PBPs [55]; in the absence of PrsA, PBP2a, PBP2b, PBP3, and PBP4 are unstable, which results in growth arrest.…”

“…Comparative genomic analysis of Brucella vaccine and virulent strains, had been widely used in order to reveal insights into virulence attenuation [6–9]. A few comparative genomic and proteomic analyses of Rev.1 and the virulent reference strain 16M (biovar 1) have been conducted to elucidate the molecular mechanism underlying virulence attenuation [10–12]. These analyses revealed a panel of 32 genome-specific markers of the Rev.1 strain, as well as various differentially expressed proteins involved in iron metabolism, sugar transport, lipid metabolism, and protein synthesis.…”