MECHANISM OF BACTERIOPHAGE SFII MEDIATED SEROTYPE CONVERSION IN SHIGELLA-FLEXNERI

We have isolated the lysogenic bacteriophage SfII, which mediates gluc osylation of Shigella flexneri O-antigen, resulting in expression of t he type II antigen. SfII belongs to group A of the Bradley classificat ion and has a genome size of 42.3 kb, DNA sequencing of a 4 kb BamHI s ubclone identified four open reading frames (ORFs), of which only two were found to be necessary for serotype conversion, These genes were n amed bgt, which encodes a putative bactoprenol glucosyl transferase, a nd gtrII, encoding the putative type II antigen determining glucosyl t ransferase. These genes are adjacent to the integrase gene (int) and a ttachment site (attP), which are highly homologous to those of Salmone lla bacteriophage P22, Another ORF encoded a highly hydrophobic protei n of 120 amino acids with homologues in Escherichia coil, Salmonella b acteriophage P22 and S. flexneri. Previous studies identified gtrX, th e glucosyl transferase gene, of bacteriophage SfX, which also glucosyl ates the O-antigen specifically, We determined that gtrX-mediated expr ession of the group 7,8 antigen also requires bgt, This allowed us to identify gtrII as being the serotype antigen II determining glucosyl t ransferase, Southern hybridization and polymerase chain reaction (PCR) analyses indicated that bgt homologues exist in the genomes of all S. flexneri serotypes and in E. coli K-12, whereas gtrII was only detect ed in strains of serotype 2. Transposon TnphoA-derived chromosomal mut ations of bgt and gtrII in S. flexneri serotype 2a were isolated and c haracterized, [S-35]-methionine labelling and the use of a T7 RNA poly merase expression system identified a protein of 34kDa corresponding t o Bgt. However, GtrII, which has a predicted molecular weight of 55 kD a, was not detected, We propose that the function of Bgt is to transfe r the glucose residues from the UDP-glucose onto bactoprenol and GtrII then transfers the glucose onto the O-antigen repeat unit at the rham nose III position, The chromosomal organization of these serotype-conv erting genes, when compared with their homologues in E. coli K-12 chro mosome and the P22 bacteriophage genome, were very similar, This sugge sts that the regions encode similar functions in these organisms and h ave a similar evolutionary origin.