No abstract

Due to drawbacks of live attenuated vaccines, much attention has been focused on screening Brucella-protective antigens as subunit vaccine candidates. Here, an immunoproteomic assay was used to identify the immunogenic soluble proteins of Brucella melitensis 16M. In the present study, 27 unique immunogenic proteins were identified from the two-dimensional electrophoresis immunoblot profiles by liquid chromatography tandem MS (LC-MS/MS). From this set, the gene encoding one immunodominant protein of interest, S-adenosyl-l-homocysteine hydrolase (AdoHcyase), was expressed in Escherichia coli. The recombinant AdoHcyase induced a strong antibody response in BALB/c mice, and the polyclonal antibody could recognize a band of approximately 52 kDa in the immunoblots of soluble protein extracts from five Brucella strains. rAdoHcyase significantly stimulated the production of interferon-γ and interleukin-2, and induced a high level of protection against B. melitensis 16M challenge at 4 weeks postchallenge. Our results indicated that rAdoHcyase could be a useful candidate for the development of subunit vaccines against B. melitensis.

Abstract. Streptococcus suis serotype 2 (SS2) is an important pathogen that affects pigs. However, neither its virulence nor its pathogenesis of infection has yet to be fully elucidated. The present study identifies a novel virulence-associated protein E gene (vapE) of SS2. To investigate the importance of vapE in SS2 infection, a vapE knock-out mutant based on SS2 wild-type strain ZY458 was designated 458ΔvapE. 458ΔvapE was generated through homologous recombination, using a combined plasmid with a vapE knock-out fragment and a pSET4s suicide vector. Additionally, the 458ΔvapE strain was transformed by a pAT18 shuttle plasmid containing the vapE gene. A functionally complemented strain for the vapE gene [termed 458ΔvapE (pvapE)] was constructed. Animal experiments demonstrated that mice infected with ZY458 and 458ΔvapE (pvapE) exhibited severe clinical symptoms, including depression, apathy, fever, anorexia, emaciation, swollen eyes and neural disorders, and died within two days of infection. All mice infected with ZY458, and 85% of mice infected with 458ΔvapE (pvapE), died within 2 days of infection. In contrast, mice inoculated with 458ΔvapE exhibited only mild clinical symptoms in the first 2 days following infection, and recovered within a week. A bacterial colonization assay demonstrated the ability of the 458ΔvapE mutant SS2 strain to colonize the heart, liver, spleen, lung and kidney of infected mice. PCR analysis of the vapE gene revealed that functional vapE was detected in virulent strains, but not in avirulent and carrier strains of S. suis SS2. These findings indicate that vapE is important for the pathogenesis of SS2.

() type 2 is an extremely important Gram-positive bacterial pathogen that can cause human or swine endocarditis, meningitis, bronchopneumonia, arthritis and sepsis. Catabolite control protein A (CcpA) is a major transcriptional regulator in type 2 that functions in catabolite control, specifically during growth on glucose or galactose. The regulation of central metabolism can affect the virulence of bacteria. In the present study, a metabolomics approach was used along with principal components analysis (PCA) and partial least-squares-discriminant analysis (PLS-DA) models and 37 metabolites were found that differed substantially between native and a mutant lacking CcpA. These results showed that CcpA is an important protein in type 2 for studying bacterial protein function.

Streptococcus suis serotype 2 (SS2) is a zoonotic pathogen that is distributed throughout the world. Virulence factors and/or markers of the virulent serotype 2 strains have not been fully identified. In this study a simple, rapid, and non-destructive method was used to extract cell wall-associated proteins from SS2 strains. Two virulent strains were compared with one avirulent strain by 2-dimensional electrophoresis (2DE). When the results of the 2DE analyses were combined with the results of mass spectrometry analyses, a total of 40 unique proteins were identified, including 26 antigens (2DE immunoblotting was used as a preliminary study). In addition to a known virulence factor, muramidase-released protein, two new proteins, catabolite control protein A and leucyl aminopeptidase, and nine potential virulence factors were also identified. The formers may be a potential virulence regulator or drug target, and the latter contains plasminogen-binding proteins and molecular chaperones. Our results complemented previous immunoproteomics studies of SS2 strains.