An integrative non-conjugative extrachromosomal genetic element, denoted as pSSVi, has been isolated from a Sulfolobus solfataricus P2 strain and was characterized. This genetic element is a double-stranded DNA of 5740 bp in size and contains eight open reading frames (ORFs). It resembles members of the pRN plasmid family in genome organization but shows only weak similarity to the latter in conserved regions. pSSVi has a copG gene similar to that of a pRN plasmid, encodes a large replication protein which, unlike a typical pRN RepA, contains no polymerase/primase domain, and lacks the plrA gene. Interestingly, pSSVi encodes an SSV-type integrase which probably catalyzes the integration of its genome into a specific site (a tRNA(Arg) gene) in the S. solfataricus P2 genome. Like pSSVx, pSSVi can be packaged into a spindle-like viral particle and spread with the help of SSV1 or SSV2. In addition, both SSV1 and SSV2 appeared to replicate more efficiently in the presence of pSSVi. Given the versatile genetic abilities, pSSVi appears to be well suited for a role in horizontal gene transfer.