Role of Capsule in the Pathogenesis of Fowl Cholera Caused by Pasteurella multocida Serogroup A

We have constructed a defined acapsular mutant in Pasteurella multocida X-73 (serogroup A:1) by disrupting the hexA gene through the insertion of a tetracycline resistance cassette. The genotype of the hexA::tet(M) strain was confirmed by PCR and Southern hybridization, and the acapsular phenotype of this strain was confirmed by electron microscopy. The hexA::tet(M) strain was attenuated in both mice and chickens. Complementation of the mutant with an intact hexAB fragment restored lethality in mice but not in chickens. In contrast to the results described previously for P. multocida serogroup B (J. D. Boyce and B. Adler, Infect. Immun. 68:3463–3468, 2000), the hexA::tet(M) strain was sensitive to the bactericidal action of chicken serum, whereas the wild-type and complemented strains were both resistant. Following inoculation into chicken muscle, the bacterial count of the hexA::tet(M) strain decreased significantly, while the wild-type and complemented strains both grew rapidly over 4 h. The capsule is thus an essential virulence determinant in the pathogenesis of fowl cholera

Acapsular Pasteurella multocida B:2 Can Stimulate Protective Immunity against Pasteurellosis

We have previously shown that a Pasteurella multocida cexA mutant (PBA875) was impaired in capsule export and highly attenuated in virulence for mice (J. D. Boyce and B. Adler, Infect. Immun. 68:3463–3468, 2000). In this study we show that immunization with high, but not low, doses of PBA875 can confer significant protection against wild-type challenge. We have also constructed a genetically defined acapsular P. multocida strain (AL18) by inactivation of bcbH, a gene predicted to be involved in polysaccharide biosynthesis. AL18 failed to produce immunoreactive polysaccharide as determined by immunofluorescence and Western immunoblot. Immunization of mice with live AL18 conferred significant protection against wild-type challenge, while immunization with similar doses of either killed wild-type or killed AL18 failed to confer protection

Preliminary analysis and annotation of the partial genome sequence of Francisella tularensis strain Schu 4.

Francisella tularensis, the aetiological agent of tularemia, is an important pathogen throughout much of the Northern hemisphere. We have carried out sample sequencing of its genome in order to gain a greater insight into this organism about which very little is known, especially at the genetic level. Nucleotide sequence data from a genomic DNA shotgun library of the virulent F. tularensis strain Schu 4 has been partially assembled to provide 1.83 Mb of the genome sequence. A preliminary analysis of the F. tularensis genome sequence has been performed and the data compared with 20 fully sequenced and annotated bacterial genomes. Plasmid-encoded genes, previously isolated from low virulence strains of F. tularensis, were not identified. A total of 1289 potential coding ORFs were identified in the data set., An analysis of this data revealed 413 ORFs which would encode proteins with no homology to known proteins. ORFs which could encode proteins involved in amino acid and purine biosynthesis were also identified. These biosynthetic pathways provide targets for the construction of a defined attenuated mutant of F. tularensis for use as a vaccine against tularemia

Genetic and Immunologic Analyses of PlpE, a Lipoprotein Important in Complement-Mediated Killing of Pasteurella haemolytica Serotype 1

Pasteurella haemolytica serotype 1 is the bacterium most commonly associated with bovine shipping fever. The presence of antibodies against P. haemolytica outer membrane proteins (OMPs) correlates statistically with resistance to experimental P. haemolytica challenge in cattle. Until now, specific P. haemolytica OMPs which elicit antibodies that function in host defense mechanisms have not been identified. In this study, we have cloned and sequenced the gene encoding one such protein, PlpE. Analysis of the deduced amino acid sequence revealed that PlpE is a lipoprotein and that it is similar to an Actinobacillus pleuropneumoniae lipoprotein, OmlA. Affinity-purified, anti-PlpE antibodies recognize a protein in all serotypes of P. haemolytica except serotype 11. We found that intact P. haemolytica and recombinant E. coli expressing PlpE are capable of absorbing anti-PlpE antibodies from bovine immune serum, indicating that PlpE is surface exposed in P. haemolytica and assumes a similar surface-exposed conformation in E. coli. In complement-mediated killing assays, we observed a significant reduction in killing of P. haemolytica when bovine immune serum that was depleted of anti-PlpE antibodies was used as the source of antibody. Our data suggest that PlpE is surface exposed and immunogenic in cattle and that antibodies against PlpE contribute to host defense against P. haemolytica