Isolation and sequencing of the replication region of plasmid pBFp1 isolated from a marine biofilm

A 24 kb plasmid, pBFp1, encoding mercury resistance was previously isolated from a marine biofilm. Isolation and sequencing of a 4280 bp DNA fragment containing the plasmid replicon (rep-pBFp1) revealed a putative open reading frame encoding a RepA protein and an oriV-like region containing an A+T rich sub-region, iterons, and DnaA boxes. Sequence comparisons showed significant similarities to the incW plasmid pSa both for the RepA amino acid sequence and in the iteron DNA sequence. Plasmid pBFp1 was also shown to be incompatible with pSa in standard incompatibility testing. A probe from the repA gene of pBFp1 was further made and tested on a collection of plasmids exogenously isolated from marine habitats in a previous study.     

Distribution of Bacillus megaterium QM B1551 plasmids among other B. megaterium strains and Bacillus species

Bacillus megaterium QM B1551 contains seven plasmids. Two are small rolling circle plasmids and five are theta-replicating plasmids with cross-hybridizing replicons that define a new family of very homologous yet compatible theta replicons. Previous sequencing of several of the plasmids has shown genes with high similarity to those on the genomes and plasmids of other Gram-positive bacteria. To test the possible distribution of these plasmids, nine other B. megaterium strains and 20 other Bacillus or related species were tested for the presence of similar replicons, and specific flanking DNA by both hybridization and PCR. The theta replicons were widespread among the B. megaterium strains, and two had one or more of the rolling circle plasmids, but none of the plasmid replicon regions were observed in the other Bacillus or related species. It appears from the data that even though some plasmids carry genes suggesting horizontal transfer, their replicons seem to be unique to B. megaterium, or rarely present in related species.     

Evidence that IncG (IncP-6) and IncU plasmids form a single incompatibility group

The Escherichia coli IncG and IncU plasmid incompatibility groups were assigned in 1980 and 1981, respectively. Complete plasmid sequences have recently been published for both these groups, and revealed that their replication proteins are related. We show that when cloned at high-copy-number, putative iterons from the previously identified IncG replicon cause strong incompatibility with IncU plasmids. Incompatibility, albeit weaker, was also demonstrated between the two replicons at their normal low-copy-number. This suggests that a single incompatibility group exists. The only known IncG plasmid, Rms149, can replicate in Pseudomonas species where it is designated IncP-6. We recommend that the combined group be known as IncU (IncP-6 in Pseudomonas spp.).