Tetracycline-resistant bacteria are frequently isolated from aquaculture systems, where mobile resistance genes often transfer between bacteria associated with fish kept at high stocking densities. Bacterial isolates from an Oreochromis mossambicus (tilapia) aquaculture system (Stellenbosch, South Africa) were screened for their susceptibility to tetracycline. Genomic and plasmid DNA were used in PCR-RFLP assays employing six degenerate primer sets to identify the prevalence of nine tetracycline resistance genes. Isolates displaying a tet(A)-type tetracycline resistance gene were examined further for an association with transposon Tn1721. tet(A) was identified as the predominant tetracycline resistance determinant, followed by tet(B), -(E), and –(C) determinants. Isolates appeared to possess multiple tet genes simultaneously. Of the isolates presented with a tet(A) determinant, 73.9% appeared to be associated with Tn1721. No association between type of tetracycline resistance gene, presence on chromosome or plasmid, and MIC could be established. The Tn1721 association may explain the high frequency of isolation of tet(A). The high levels of resistance displayed by isolates from the tilapia aquaculture system not previously exposed to antimicrobial agents is of concern and will have implications for future therapeutic interventions in disease outbreak situations. Key words: Tetracycline resistance, aquaculture, tilapia, transposons.