Plasmid-like replication of a minimal streptococcal integrative and conjugative element

Integrative and conjugative elements (ICEs) are mobile genetic elements encoding their own excision from a replicon of their bacterial host, transfer by conjugation to a recipient bacterium and reintegration for maintenance. The conjugation, recombination and regulation modules of ICEs of the ICESt3 family are grouped together in a region called the ICE 'core region'. In addition to this core region, elements belonging to this family carry a highly variable region including cargo genes that could be involved in bacterial adaptation or in the maintenance of the element. Although ICEs are a major class of mobile elements through bacterial genomes, the functionality of an element encoding only its excision, transfer, integration and regulation has never been demonstrated experimentally. We engineered MiniICESt3, an artificial ICE derived from ICESt3, devoid of its cargo genes and thus only harbouring the core region. The functionality of this minimal element was assessed. MiniICESt3 was found to be able to excise at a rate of 3.1 %, transfer with a frequency of 1.0 × 10^- 5 transconjugants per donor cell and stably maintain by site-specific integration into the 3' end of the fda gene, the same as ICESt3. Furthermore, MiniICESt3 was found in ∼10 copies per chromosome, this multicopy state likely contributing to its stability for >100 generations even in the absence of selection. Therefore, although ICEs were primarily assumed to only replicate along with the chromosome, our results uncovered extrachromosomal rolling-circle replicating plasmid-like forms of MiniICESt3.