Infection with the human pathogen Vibrio vulnificus leads to the generation of reactive oxygen species (ROS) via NAD(P)H oxidase (Nox) in host cells. In the present study, we employed mutant V. vulnificus strains to identify an essential virulence factor responsible for this ROS generation. We found that repeats-in-toxin A1 (RtxA1) expressed by V. vulnificus acts via Nox1 to induce significant ROS generation in the intestine epithelial cells, which ultimately results in cell death. Furthermore, RtxA1 modulates the small GTPase Rac2, which is known to play an important role in the activation of Nox. When mice were infected by the oral method, in contrast with the wild-type bacteria, an RtxA1-deficient V. vulnificus mutant was unable to induce ROS generation within the intestine and failed to cause death. These findings strongly suggest that RtxA1-induced Rac2 expression is a critical step underlying the pathogenicity of V. vulnificus.

A simple assay for the rapid screening of bacterial species-or subspecies-specific DNA probes for the random cloning method is presented, involving the use of genomic DNAs as probes and recombinant plasmid DNAs containing genomic DNA digested with HindIII as targets. The optimal amount of target DNAs and the concentration of digoxigenin-labeled genomic DNA probes were 20 ng and 100 ng ml 31 (or 10 ng and 200 ng ml 31 ), respectively. The method was applied to the development of Fusobacterium nucleatum subspecies-specific probes. Our results showed that four out of 96 probes were F. nucleatum subspecies-specific, which was confirmed by Southern blot analysis. Our results indicate that the new method can be used for the rapid screening of species-or subspecies-specific probes. ß

The aim of this study was to identify the bacteria isolated from acute endodontic lesions by cell culture and 16S rDNA sequencing. The necrotic pulpal tissue was collected from 17 infected root canals, which were diagnosed as being either an acute pulpitis or acute periapical abscess. Samples were collected aseptically from the infected pulpal tissue of the infected root canals using a barbed broach and a paper point. The cut barbed broaches and paper points were transferred to an eppendorf tube containing 500 ul of 1 X PBS. The sample solution was briefly mixed and plated onto a BHI-agar plate containing 5% sheep blood. The agar plates were incubated in a 37℃ anaerobic chamber for 7 days. The bacteria growing on the agar plate were identified by 16S rRNA coding gene (rDNA) cloning and sequencing at the species level. Among the 71 colonies grown on the agar plates, 56 strains survived and were identified. In dental caries involving the root canals, Streptococcus spp. were mainly isolated. Actinomyces, Clostridia, Bacteroides and Fusobacteria were isolated in the periapical lesion without dental caries. Interestingly, two new Actinomyces spp. (ChDC B639 and ChDC B631) were isolated in this study. These results showed that there was diversity among the species in endodontic lesions. This suggests that an endodontic infection is a mixed infection with a polymicrobial etiology. These results may offer the bacterial strains for pathogenesis studies related to an endodontic infection. [J Kor Acad Cons Dent 30(5):409-422, 2005]