Objectives: A multiresistant Aeromonas bestiarum strain, shown to be persistent and spreading in a freshwater stream, was investigated for the presence, location and organization of antimicrobial resistance genes. Methods: The plasmid pAB5S9 was transferred by electroporation into Escherichia coli TG1. The resistance phenotype mediated by pAB5S9 was determined. Moreover, the plasmid was sequenced completely and analysed for its structure and organization of reading frames. Results: Plasmid pAB5S9 mediated resistances to phenicols, sulphonamides, streptomycin and tetracycline. The analysis of the 24.7 kb sequence revealed the presence of 20 predicted coding sequences (CDSs), which included the floR, sul2 and strA-strB resistance genes and a tetR-tet(Y) determinant. Approximately 7.5 kb of pAB5S9 showed 100% nucleotide sequence identity to three non-contiguous segments of the SXT element of Vibrio cholerae. Regions identical to SXT comprised the floR gene, flanked upstream by a complete and downstream by a truncated ISCR2 element, and the region of the sul2 and strA-strB genes. Other CDSs of pAB5S9 related to plasmid replication and partitioning, metabolic and gene regulation functions as well as conjugative transfer showed homology to sequences from diverse bacterial species, indicating a mosaic structure. Conclusions: This study provides the first report of a floR-carrying plasmid in the genus Aeromonas and the first description of a tetR-tet(Y) determinant. The analysis of the multiresistant A. bestiarum strain indicates that strains of this species, some of which are opportunistic pathogens for fish, might also act as a resistance gene reservoir in the freshwater environment.