Antibody response against Escherichia coli heat-stable enterotoxin expressed as fusions to flagellin

The heat-stable toxin (ST) produced by enterotoxigenic Escherichia coli str ains causes diarrhoea by altering the fluid secretion in intestinal epithel ial cells. Here, the effectiveness of a flagellin fusion protein of Salmone lla containing a 19-amino-acid sequence derived from the ST sequence (FLA-S T) in generating antibodies capable of neutralizing the toxic activity of S T was evaluated. This fusion protein, and an alternative construction where two cysteine residues in the ST sequence were substituted by alanines (ST, ), were delivered to the immune system by three distinct strategies: (i) or ally, using an attenuated Salmonella strain expressing FLA-ST; (ii) intrape ritoneally, by injection of purified FLA-ST; (iii) orally, using attenuated Salmonella carrying a eukaryotic expression plasmid (pCDNA3) with the gene encoding FLA-ST. The results showed that the flagellin system can be used as a carrier to generate ST-neutralizing antibodies. However, it should be mentioned that humoral immune response against ST was only obtained when th e mutated ST sequence was employed. FLA-ST was found to be non-immunogenic when delivered via the oral route with attenuated Salmonella strains. Howev er, a flagellin antibody response was obtained by immunizing mice with Salm onella carrying pCDNA3/FLA-STmt. Oral immunization with Salmonella carrying the eukaryotic expression plasmid (pCDNA3/FLA-STmt) seems to be a promisin g method to elicit an appropriate response against fusions to flagellin.