Studies of aerolysin promoters from different Aeromonas spp

Aeromonas spp. have been related to food and waterborne diseases. The pore forming toxin aerolysin is regarded as the most important virulence factor in Aeromonas food poisoning. In this work the aerolysin promoters from several Aeromonas spp. strains have been sequenced, and divided into two sequence groups. Further analyses of the promoters were carried out in a reporter-plasmid, pSTINA-II. This plasmid was constructed as a hybrid of pUC4K, pACYC184 and pKK232-8. We could conclude that our constructed reporter-gene plasmid was functional and was used to compare different promoters in an aerolysin negative Aeromonas spp. This construct made it possible to study the expression of the reporter gene using different aerolysin promoters under several conditions. We were able to show that the two obtained sequence groups of aerolysin promoters gave different expression of the reporter gene, and that this expression was dependent of temperature and osmolarity. Reducing the size of one promoter sequence from 254 to 148 bp and 102 bp gave a gradual reduction of the reporter gene expression under all conditions. According to our assay there seems to be more than one functional promoter upstream of the aerolysin gene, although the RT-PCR indicated one transcription starting point, under all test conditions.