REPLICATION AND INTEGRATION OF A VIBRIO-CHOLERAE CRYPTIC PLASMID LINKED TO THE CTX PROPHAGE

We identified a 4.7 kb cryptic plasmid in all ctxAB(+) Vibrio cholerae strains we tested. An isolate of the V. cholerae classical biotype st rain O395 that harbours the cryptic plasmid at high copy number was fo und. Hybridization analysis demonstrated that sequences highly related or identical to this plasmid exist in all toxigenic strains of V.chol erae but were notably absent in all non-toxigenic environmental isolat es that lacked the genes for toxin-co-regulated pill and the filamento us CTX prophage. Accordingly, we have named the cryptic plasmid pTLC f or toxin-linked cryptic. The complete nucleotide sequence of pTLC from the high-copy-number isolate was determined. The largest open reading frame in the plasmid is predicted to encode a protein similar to the replication initiation protein (pll) of Escherichia coli F-specific fi lamentous phages. The nucleotide sequence of pTLC also facilitated the structural characterization of the DNA homologous to pTLC in other st rains of V.cholerae. pTLC-related DNA exists in these strains as both low-copy-number, covalently closed circular DNA and tandemly duplicate d, chromosomally integrated DNA. Remarkably, the chromosomally integra ted form of pTLC is adjacent to the CTX prophage. The strain distribut ion, chromosomal location and DNA sequence of pTLC suggests that it ma y be a genetic element that plays some role in the biology of CTX phi, perhaps facilitating either its acquisition or its replication.