Development and application of molecular tools in the study of IncN-related plasmids from lakewater sediments

Homology to IncN, P, Q and W inc regions was investigated amongst 114 Hg2+- resistant or antibiotic-resistant bacteria isolated from lakewater sediment s. No hybridisation signals were found with Inc P, Q and W probes, and only one plasmid, pLV1402, hybridised to the IncN probe. PCR primers designed t o conserved regions in the replicon of the IncN plasmid pCU1 and the relate d beta replicon from pGSH500 were used to amplify a 978-bp fragment from pL V1402, with sequence analysis showing a close relationship (99.2% identity) between their replication genes. A 387-bp region from the pLV1402 rep gene was used to re-screen the isolates and identified another related plasmid, pLV1403. A 3.7-kb probe containing the alpha replicon from pGSH500 hybridi sed to both pLV1402 and pLV1403, suggesting that both are multi-replicon pl asmids. The PCR primers and probes described will be useful in future studi es of plasmid diversity. (C) 2000 Federation of European Microbiological So cieties. Published by Elsevier Science B.V. All rights reserved.