Deletion in the C-terminal domain of ClpX delayed entry of Salmonella enterica into a viable but non-culturable state.

Under stressful conditions, bacteria enter a viable but non-culturable (VBNC) state in which they are alive but fail to grow on conventional media. The molecular basis underlying this state is unknown. To identify the key gene responsible for the VBNC state in Salmonella spp., we examined a S. Typhimurium LT2 VBNC mutant, which shows a characteristic delay in entering the VBNC state. The mutant showed a higher level of expression of general stress sigma factor RpoS than wild-type LT2. The mutant carried a 99-bp in-frame deletion in the clpX gene (clpXΔ323-355). ClpX is known to form a ClpXP protease complex with ClpP, which plays a role in the degradation of RpoS. To investigate the effect of clpXΔ323-355 on VBNC induction, ΔclpX and clpXΔ323-355 strains were generated from LT2 cells. Compared to LT2, the ΔclpX and clpXΔ323-355 strains showed greater amounts of RpoS and required a longer incubation time for induction into the VBNC state. These results suggest that residues 323-355 of ClpX play a major role in the hexameric formation or function of ClpX and in the rate of induction of the VBNC state.

Endoscopic submucosal dissection with sheath-assisted counter traction for early gastric cancers.

AIM: Endoscopic submucosal dissection (ESD) is associated with frequent complications, such as bleeding and perforation. The procedure is technically difficult, requires considerable skill and is longer than conventional endoscopic mucosal resection (EMR). Thus, non-invasive tools and methods are needed to facilitate direct visualization of the submucosal layer during ESD. METHODS: An insulation-tipped (IT) knife was inserted into one channel, and a conventional injection sheath was inserted to the other channel of a double-channel fiberscope. The submucosal layer was lifted via a circumferential incision using the conventional injection sheath for visualizing cutting lines, and the submucosal layer under the lesion was directly dissected from the underlying muscularis propria. RESULTS: Sheath-assisted traction resulted in successful ESD with en bloc resection in all 25 patients. Hemostasis and manipulation of blood vessels were uncomplicated and dissection was completed safely, without either bleeding or perforation. The movement of the sheath was not suppressed while lifting the submucosal layer and the IT knife could be moved freely, which allowed submucosal dissection independently of the sheath movement. CONCLUSION: Sheath-assisted traction ESD, using simple materials and methods, has several advantages over other standard traction methods. Our procedure is straightforward, safe, non-invasive, cost-effective and uses readily available instruments to enhance visualization of cutting lines.

Immune responses in mice to intranasal and intracutaneous administration of a DNA vaccine encoding Helicobacter pylori-catalase.

We previously reported that the intracutaneous injection of DNA vaccines encoding Helicobacter pylori heat shock proteins elicited specific immune responses, and led to reduced infection in mice. In this study, we constructed DNA vaccine encoding H. pylori-catalase (pcDNA3.1-kat) and investigated the immune responses to intranasal and intracutaneous administration of pcDNA3.1-kat. C57/BL6 mice were immunized intracutaneously with 10 microg of pcDNA3.1-kat or intranasally with 50 microg of pcDNA3.1-kat. Catalase-specific IgG antibody was detected in the sera of intranasal and intracutaneous immunized mice. Both intranasal and intracutaneous immunized mice were significantly protected from colonization by H. pylori and had significantly reduced degrees of gastritis. These results demonstrate that DNA vaccine encoding H. pylori-catalase can induce an immune response against H. pylori, and that intranasal immunization works as well as intracutaneous immunization.