Nucleotide sequence and transfer properties of two novel types of Actinobacillus pleuropneumoniae plasmids carrying the tetracycline resistance gene tet(H)

Abstract

To analyse the sequence and transfer properties of two tetracycline resistance plasmids found in clinical isolates of Actinobacillus pleuropneumoniae in order to assess their role in the spread of tetracycline resistance. The plasmids designated p9956 and p12494 were purified from A. pleuropneumoniae and completely sequenced. The transfer properties of both plasmids were evaluated by electroporation and/or conjugation into Pasteurella multocida and Escherichia coli. Both plasmids showed a modular structure consisting of three regions involved in mobilization, tetracycline resistance or replication. The mobilization regions included the mobA gene, encoding a relaxase, a protein involved in plasmid transfer. The tetracycline resistance regions were closely related and consisted of the tet(H) gene and its repressor gene tetR. The tetracycline resistance phenotype was transferred successfully to P. multocida and in the case of p9956 also to E. coli by electroporation of the plasmids. Moreover, plasmid p9956 could be mobilized in E. coli with the assistance of RP4 conjugal transfer functions. For the first time, the complete sequences of two tet(H)-carrying plasmids from A. pleuropneumoniae were determined. These two plasmids differed from one another and from known tet(H)-carrying plasmids from Pasteurella or Mannheimia spp. Structural analysis confirmed that these plasmids consisted of segments that have been previously detected in members of the families Pasteurellaceae and Enterobacteriaceae.