Cloning and sequence of a region encoding a surface polysaccharide of Vibrio cholerae O139 and characterization of the insertion site in the chromosome of Vibrio cholerae O1

Vibrio cholerae serogroup O139 Bengal is the first documented serogroup other than O1 to cause epidemic cholera. The O139 Bengal strains are very similar to V. cholerae serogroup O1 biotype El Tor strains. The major differences between the two serogroups are that O139 Bengal contains a distinct O antigen and produces a polysaccharide capsule. We previously described three Tn phoA mutants of O139 strain AI1837 which abolish both O antigen and capsule production. These Tn phoA insertions were mapped to a 21.5 kb Eco RI fragment of the O139 chromosome. We describe here the cloning and mapping of this 21.5 kb Eco RI fragment and it was shown to complement each of the mutants in trans to produce O antigen and capsule. The Eco RI fragment contains 13 kb of DNA that is specific to O139 and 8.5 kb of DNA that is common to O1 and O139. Sequence analysis of the 13 kb of O139-specific DNA revealed that it contains 11 open reading frames all of which are transcribed in the same direction. Eight of the 11 open reading frames are homologous to sugar biosynthesis genes from other organisms. Using extended polymerase chain reactions, we show that the extent of the DNA region in O139 that is not present in O1 is approximately 35kb. The site of insertion of this O139-specific DNA in the O1 chromosome was mapped to the rfb _O1 region. We also demonstrate that O139 Bengal strain AI1837 contains a deletion of 22 kb that in serogroup O1 strains contains the rfb region. Therefore, O139 Bengal probably arose from an O1 strain that had undergone genetic rearrangements including deletion of the O1 rfb region and acquisition of a 35 kb region of DNA which encodes O139 surface polysaccharide.