Transfer Of Tn916 Research Articles

Conjugative transposition of Tn916 and Tn925 in Bacillus popilliae

Interspecies transfer of the conjugative transposons Tn916 and Tn925 into B. popilliae Pj1 occurred using Enterococcus faecalis and Bacillus subtilis CU4049 as transposon donors. Tn916 was stably maintained in B. popilliae Pj1 following growth without selective pressure and was successfully introduced into the plasmid-containing B. popilliae strains NRRL B-2524, Ch1, and KLN4 using E. faecalis CG110. In B. popilliae, expression of the tetracycline resistant determinants on Tn916 and Tn925 provided resistance to 25 μg/mL and 50 μg/mL tetracycline, respectively. An erythromycin resistant determinant, present in Tn916ΔE, was also functional in B. popilliae Pj1 and provided resistance to 1 mg/mL erythromycin. Transfer of Tn916 into E. faecalis, B. subtilis, and between B. popilliae strains was accomplished using a transposon-containing strain of B. popilliae as donor. Efforts to transfer Tn916 between E. coli and B. popilliae were unsuccessful. Key words: Bacillus popilliae, milky disease, Tn916, conjugative transposon.

Broad host range gene transfer: plasmids and conjugative transposons

Conjugation is the primary route of broad host range DNA transfer between different genera of bacteria. Plasmids are the most familiar conjugative elements, but there are also self-transmissible integrated elements called conjugative transposons. Conjugative transposons have been found in many genera of gram-positive bacteria, in mycoplasmas and in gram negative bacteria such as Bacteriodes spp. and Moraxella spp., and they have a very broad host range. The best-studied conjugative transposons are: the ones related to Tn916, a 16 kb conjugative transposon found originally in Gram-positive bacteria; Tn5276, a 70 kb conjugative transposon from Lactococcus lactis; and a group of large (> 70 kb) conjugative transposons found in Bacteroides spp. Transfer of conjugative transposons takes place in three steps: excision to form a circular intermediate, transfer of one strand of the circular intermediate to a recipient, and integration into the recipient genome. Some conjugative transposons integrate almost randomly, whereas other integrate site-specifically. Conjugative transposons not only transfer themselves but also mobilize co-resident plasmids, either by providing transfer functions in trans or by inserting themselves into the plasmid. In addition, the conjugative transposons found in Bacteroides spp. can excise and mobilize unlinked integrated elements, called NBUs. Transfer of many of the Bacteroides conjugative transposons is regulated by tetracycline, whereas transfer of Tn916 and other conjugative transposons appears to be constitutive. The conjugative transposons are clearly widespread in clinical isolates, but their distribution in environmental isolates remains to be determined.

Transfer of Tn916 and Tn916 delta E into Clostridium difficile: demonstration of a hot-spot for these elements in the C. difficile genome.

The conjugative transposon Tn916 and a derivative Tn916 delta E was transferred from Bacillus subtilis into Clostridium difficile CD37 by filter mating. All the C. difficile transconjugants appeared to contain one copy of the transposon integrated into the same position in the genome. Transposition from the original site of integration was not observed. Like Tn916 the transferable tetracycline resistance determinant (Tc-CD) of C. difficile has a preferred site of integration in C. difficile and is homologous with Tn916 along the whole length of Tn916. However comparisons of the distribution of TaqI and Sau3AI sites in the homologous regions of the two elements did not demonstrate any hybridizing fragments in common.