Project Leader:

Dr. Wolfgang Koester

Project Leader:

Dr. Andrew Potter

Background: Bacterial pathogens, including Salmonella species, C. jejuni and E. coli O157:H7 in food products and water are responsible for a significant number of illnesses and deaths each year internationally. In recent years, the rate of enteric disease caused by Campylobacter species has surpassed the rate of diarrhea caused by Salmonella. In addition, parasites such as Cryptosporidium parvum have been associated with several disease outbreaks including one in North Battleford, Saskatchewan. Many of these organisms exist naturally as part of the flora of animals and thus pose a risk of environmental contamination or contamination of food.

Each of the bacterial species possesses an assortment of virulence factors that are relatively well understood in relation to human disease. However, the mechanism by which they colonize their animal hosts is not well understood. Both enterohemorrhagic E. coli strains and Salmonella strains produce a novel set of virulence determinants which are delivered to host cells via type III secretion systems. These proteins have been shown to form an effective vaccine for the reduction of E. coli O157:H7 levels in cattle following vaccination, but their role in Salmonella pathogenesis is unclear. Likewise, C. jejuni produces a number of secreted proteins, but their role in colonization of poultry is not known.

Animals are generally well-adapted to the organic dusts in their environments and show little response to dust control measures. In contrast, both acute and chronic adverse health effects occur in people who are exposed to these dusts which are widespread in contemporary livestock production settings. There is increasing evidence that the amount of endotoxin on these particles is a primary risk factor associated with adverse effects.

Objectives: The long-term objective of this program is to develop intervention methods for the control of human pathogens in their animal hosts.

• Characterization of non-O157 EHEC serotypes and C. jejuni virulence determinants
• Characterization of Salmonella enterica serotype Enteritidis (S. Enteritidis) SPI-1, SPI-2 and construction of mutants
• Cross-protective capacity of EHEC vaccine
• Virulence testing of C.jejuni mutants, S. Enteritidis SPI-1, SPI-2 mutants
• EHEC formulations for mucosal immunity and efficacy of mucosal vaccine formulations
• S. Enteritidis, C. jejuni vaccine testing
• Optimize delivery of poultry vaccines


Progress:

Recent publication

Characterization of non-O157 EHEC isolates. Our hypothesis was that vaccination with Type III-secreted proteins from E. coli O157 would also block colonization by other EHEC serotypes. Our results suggest that it will not be possible to protect against heterologous EHEC serotypes by vaccinating with proteins from a single serotype. However, given the relatively high degree of amino acid homology between the Tir and the EspA proteins, we believe that it will be possible to construct chimeric antigens containing epitopes representative of each of these strains.

Optimization of mucosal immunity to EHEC vaccines. Animals that are immunized intranasally with formulations containing mucosal adjuvants show a significant increase in antibody relative to control animals which received only the antigen in the absence of a mucosal adjuvant. However, all mice showed a secondary (anamnestic) response upon re-exposure at seven weeks. The best responses when animals were challenged with E. coli O157 were seen in groups receiving the cholera toxin or CpG as adjuvants.

Roles of S. Enteritidis molecules in colonization of chickens. Chromosomal deletion mutants were constructed in Salmonella enterica serotype Enteritidis isolated from chickens. The resulting mutant strains carrying single or double gene knock-outs are suitable candidates for testing of colonization abilities in birds. Surface-exposed and secreted components which are essential for colonization will be targets for vaccine trials.

Characterization of C. jejuni antigens. We are evaluating C. jejuni flagella for potential as an antigen. In order to characterize the immune response against flagellar protein, we synthesized synthetic peptides covering the entire sequence of the protein and used these to monitor immune responses from birds that were exposed to a variety of C. jejuni strains. The results indicated that most of the response was targeted against epitopes within the hypervariable region, making them unsuitable for use in vaccine antigens. However, we believe cross-serotype protection may be possible through an immunodominant epitope portion of the protein. We are currently testing this hypothesis using synthetic peptide-based vaccines.


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