Constructions of bft-k and t-3 Deficient Mutants of Bacteroides fragilis: Possible Role for Metalloprotease in Pathogenesis

We earlier reported the identification of bif-k, t-3, and a third ORF from an enterotoxigenic strain of Bacteroides fragilis 419, which was isolated from the blood of a Korean patient suffering from systemic infections. In the present study, the deleted fragments of the t-3 and the bft-k genes from B. fragilis 419 were cloned into suicide vector PJST55 and used to create a mutant with chromosomal disruption of the t-3 and bft-k genes. Structures of the selected mutants, DMP-2 and DBT-4, were found to be intermediate forms that integrated the suicide vector into the chromosome. t-3 disrupted Dmp-2 and Bft-k disrupted DBT-4 did not react with polyclonal antibodies against T-3 or BFT-K, and had no biological activity in $HT29/C_1$, cells.