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The complete sequences of plasmids pB2 and pB3 provide evidence for a recent ancestor of the IncP-1 group without any accessory genes

¹ Department of Biological Sciences, University of Idaho, Moscow, ID 83844-3051, USA
² Fakultät für Biologie, Lehrstuhl für Genetik, Universität Bielefeld, Postfach 100131, D-33501 Bielefeld, Germany

Correspondence
E. M. Top
evatop{at}uidaho.edu

The nucleotide sequences of the broad-host-range antibiotic resistance plasmids pB2 (61 kb) and pB3 (56 kb), which were isolated from a wastewater treatment plant, were determined and analysed. Both have a nearly identical IncP-1 backbone, which diverged early from the sequenced IncP-1 plasmids R751, pB10, pJP4, pADP1 and pUO1. In contrast to the latter plasmids, the pB2 and pB3 backbone does not seem to have undergone any deletions. The complete partition gene parA is located downstream of the mating pair formation (trb) module. A 14·4 kb or 19·0 kb mobile genetic element is present between traC and parA of pB3 and pB2, respectively. This region is typical for insertions in IncP-1 plasmids, but the insertion site is unique. Both elements differ only by a duplication in pB2 of a tetA(C)–tetR–tnpA_IS26 fragment. The 5 bp target site duplication and the 26 bp inverted repeats flanking the mobile genetic elements are still intact, indicating that the insertion occurred recently. The element consists of three nested transposable elements: (i) a relict of a Tn402-like transposon with a gene for a new class D -lactamase (bla_NPS-2); (ii) within that, another Tn402-like element with a class 1 integron harbouring the gene cassettes cmlA1 for a chloramphenicol efflux protein and aadA2 encoding a streptomycin/spectinomycin adenylyltransferase, and a copy of IS6100; (iii) into the integrase gene intI1 a tetracycline resistance module tetA(C)–tetR flanked by copies of IS26 is inserted. Interestingly, in contrast to all other IncP-1 plasmids analysed so far, the oriV region between trfA and klcA is not interrupted by accessory genes, and there is no indication that previously inserted accessory genes have subsequently been deleted. The genes kluAB are also missing in that region and should thus be considered acquired genes. These findings, together with the fact that IncP-1 plasmids acquired accessory elements at various positions in the backbone, suggest that IncP-1 plasmids without any accessory genes exist in microbial communities. They must occasionally acquire accessory genes by transposition events, resulting in those plasmids that have been found based on selectable phenotypic traits.

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