Molecular organization and expression of the tetracycline resistance (TetB1) determinant specified by IncHI1 plasmids

Abstract

R27, a plasmid of the HI1 incompatibility group, carries a tetracycline resistance (TcR) determinant previously designated class B1. A 4.5 kbHindIII fragment of R27 specifying TcR was cloned into pACYC177 and the recombinant plasmid designated pDT1539. The minimal inhibitory concentration (MIC) of tetracycline for R27 and pDT1539 was 64 μ/ml, whereas the MIC for the TetB determinant from Tn10 was 128 μg/ml. The restriction maps of the TetB and TetB1 determinants appeared identical, and DNA-DNA heteroduplex analysis of the 4.5 kbHindIII fragment from R27 and pRT11 derived from Tn10, which contained thetetR andtetB genes, was consistent with extended homology between the two DNA molecules. The TetB and TetB1 polypeptides produced in anEscherichia coli in vitro transcription/translation system had the same apparent molecular size of 36 kilodaltons. Thus, apart from the minor variation in MIC levels, there appeared to be little difference between the TetB and TetB1 determinants. DNA homology was demonstrated between a TetB1 probe and ten other incompatibility group HI1 plasmids from various geographic sources. Moreover, all specified a level of tetracycline resistance similar to that of R27. These data are consistent with clonal spread of a progenitor IncHI1 plasmid carrying the TetB1 determinant.