The 3’ end of the insert (module E) is homologous to Tn1806
of S. pneumoniae which confers erythromycin resistance. Although this element has not been shown to transfer via conjugation, transfer via transformation was shown . In C. difficile strain M120 this element appears to be the backbone into which several other elements have been inserted (see Figure 1 top panel). The first 7.3 kb on the 5’ end of the insert (module A) has only moderate homology (60–70% maximum sequence identity) to known sequences. Interestingly, this part of the insert contains 2 putative modification DNA methylases and a putative endonuclease, possibly enabling a form of molecular vaccination as described by Kobayashi et al. . During this process methylation protects the incoming
FG-4592 mouse element from host endonucleases and, following integration, will protect the host chromosome from endonucleases present on other mobile genetic elements. This sequence is followed by a complete prophage of approximately 39.5 kb (module B), which shows 92% sequence identity to a Thermoanaerobacter sp. prophage (Genbank accession no. CP002210). The next 4.5 kb stretch (module C) is 99% identical to part of the Enterococcus faecalis plasmid pEF418 containing, amongst others, a putative methyltransferase and a putative spectinomycin adenyltransferase (ant(9)Ia) . It is also described to be part of a pathogenicity island in Streptococcus suis. Finally, an insertion of approximately www.selleckchem.com/products/elafibranor.html 4.5 kb (module D) with 90% sequence identity to the transferable pathogenicity island of Campylobacter fetus subsp fetus is present within the sequence of Tn1806. This sequence contains, amongst others, putative tet(44) and ant(6)-Ib genes, which could respectively confer tetracycline and selleck chemicals streptomycin resistance. The G + C content of the entire insert (34%) was significantly higher than that of the Forskolin purchase entire genome (29%), clearly indicating that the insert was of foreign origin (see Additional file 1). In addition, within the insert the different modules could be distinguished by their G + C contents. The G + C content
of module A, B, C, D and E was 31%, 41%, 35%, 28% and 31%, respectively. The 100 kb insert is a transposon Based on the bioinformatic comparison of the insert described above, the possible excision of 3 (independent) elements was predicted. Primers were designed (primers 14–20, see Table 3) to amplify the circular intermediates of the complete insert (primers 14 and 15), the putative Thermoanaerobacter sp. phage (module B, primers 15 and 16) and the C. fetus pathogenicity island (module D, primers 17 and 18) of the element. PCR confirmed only the excision and circularisation of the entire insert (results not shown). It is expected that the serine recombinase at the 3’ end of the element is responsible for excision (see Table 1).