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1.  Streptococcus pneumoniae DivIVA: Localization and Interactions in a MinCD-Free Context▿ †  
Journal of Bacteriology  2006;189(4):1288-1298.
To clarify the function of DivIVA in Streptococcus pneumoniae, we localized this protein in exponentially growing cells by both immunofluorescence microscopy and immunoelectron microscopy and found that S. pneumoniae DivIVA (DivIVASPN) had a unique localization profile: it was present simultaneously both as a ring at the division septum and as dots at the cell poles. Double-immunofluorescence analysis suggested that DivIVA is recruited to the septum at a later stage than FtsZ and is retained at the poles after cell separation. All the other cell division proteins that we tested were localized in the divIVA null mutant, although the percentage of cells having constricted Z rings was significantly reduced. In agreement with its localization profile and consistent with its coiled-coil nature, DivIVA interacted with itself and with a number of known or putative S. pneumoniae cell division proteins. Finally, a missense divIVA mutant, obtained by allelic replacement, allowed us to correlate, at the molecular level, the specific interactions and some of the facets of the divIVA mutant phenotype. Taken together, the results suggest that although the possibility of a direct role in chromosome segregation cannot be ruled out, DivIVA in S. pneumoniae seems to be primarily involved in the formation and maturation of the cell poles. The localization and the interaction properties of DivIVASPN raise the intriguing possibility that a common, MinCD-independent function evolved differently in the various host backgrounds.
PMCID: PMC1797354  PMID: 17098892
2.  Effects of crp deletion in Salmonella enterica serotype Gallinarum 
Salmonella enterica serotype Gallinarum (S. Gallinarum) remains an important pathogen of poultry, especially in developing countries. There is a need to develop effective and safe vaccines. In the current study, the effect of crp deletion was investigated with respect to virulence and biochemical properties and the possible use of a deletion mutant as vaccine candidate was preliminarily tested.
Mutants were constructed in S. Gallinarum by P22 transduction from Salmonella Typhimurium (S. Typhimurium) with deletion of the crp gene. The effect was characterized by measuring biochemical properties and by testing of invasion in a chicken loop model and by challenge of six-day-old chickens. Further, birds were immunized with the deleted strain and challenged with the wild type isolate.
The crp deletions caused complete attenuation of S. Gallinarum. This was shown by ileal loop experiments not to be due to significantly reduced invasion. Strains with such deletions may have vaccine potential, since oral inoculatoin with S. Gallinarum Δcrp completely protected against challenge with the same dose of wild type S. Gallinarum ten days post immunization. Interestingly, the mutations did not cause the same biochemical and growth changes to the two biotypes of S. Gallinarum. All biochemical effects but not virulence could be complemented by providing an intact crp-gene from S. Typhimurium on the plasmid pSD110.
Transduction of a Tn10 disrupted crp gene from S. Typhimurium caused attenuation in S. Gallinarum and mutated strains are possible candidates for live vaccines against fowl typhoid.
PMCID: PMC1885444  PMID: 17488512
3.  Mapping Antigenic Sites of an Immunodominant Surface Lipoprotein of Mycoplasma agalactiae, AvgC, with the Use of Synthetic Peptides  
Infection and Immunity  2002;70(1):171-176.
As a first step toward the design of an epitope vaccine to prevent contagious agalactia, the strongly immunogenic 55-kDa protein of Mycoplasma agalactiae was studied and found to correspond to the AvgC protein encoded by the avgC gene. The avg genes of M. agalactiae, which encode four variable surface lipoproteins, display a significant homology to the vsp (variable membrane surface lipoproteins) genes of the bovine pathogen Mycoplasma bovis at their promoter region as well as their N-terminus-encoding regions. Some members of the Vsp family are known to be involved in cytoadhesion to host cells. In order to localize immunogenic peptides in the AvgC antigen, the protein sequence was submitted to epitope prediction analysis, and five sets of overlapping peptides, corresponding to five selected regions, were synthesized by Spot synthesis. Reactive peptides were selected by immunobinding assay with sera from infected sheep. The three most immunogenic epitopes were shown to be surface exposed by immunoprecipitation assays, and one of these was specifically recognized by all tested sera. Our study indicates that selected epitopes of the AvgC lipoprotein may be used to develop a peptide-based vaccine which is effective against M. agalactiae infection.
PMCID: PMC127643  PMID: 11748179

Results 1-3 (3)