A clinical isolate of Klebsiella pneumoniae K7746 produced the extended-spectrum beta-lactamase (ESBL) SHV-12. A 6.6 kb BamHI fragment containing the blaSHV-12 gene of K7746 strain was cloned into pCRScriptCAM vector resulting in the recombinant plasmid p7746-C1. The restriction map of 3.6 kb inserted DNA and sequences immediately surrounding blaSHV-12 of p7746-C1 were homologous to plasmid pMPA2a carrying blaSHV-2a. In addition, both blaSHV-12 and blaSHV-2a were expressed from a common hybrid promoter made of the -35 region derived from the left inverted repeat of IS26 and the -10 region from the blaSHV promoter itself. The results indicate that blaSHV-12 and blaSHV-2a may have evolved from a common ancestor in the sequential order of blaSHV-2a first, followed by blaSHV-12. Furthermore, by the PCR mapping method using primers corresponding to the IS26 and blaSHV, the association between IS26 and blaSHV was studied in 12 clinical isolates carrying blaSHV-2a, 27 clinical isolates carrying blaSHV-12, and 5 reference strains carrying blaSHV-1 to blaSHV-5. All 39 strains carrying blaSHV-2a or blaSHV-12 were positive by the PCR, providing confirmative evidence that IS26 has been involved in the evolution and dissemination of blaSHV-2a and blaSHV-12. But 5 reference strains carrying blaSHV-1 to blaSHV-5 were negative by the PCR. Therefore, we concluded that the molecular evolutionary pathway of blaSHV-2a and blaSHV-12 may be different from that of other blaSHV-ESBL, e.g., blaSHV-2, blaSHV-3, blaSHV-4, and blaSHV-5.
beta-Lactamases ; Clone Cells ; DNA ; Klebsiella pneumoniae ; Plasmids ; Polymerase Chain Reaction

V. vulnificus is an estuarine bacterium which causes septicemia and shock in susceptible patients. The organism produces a hemolytic cytolysin (VvH), which has a membrane damaging effect on erythrocytes. To clarify the mechanisms by which VvH might contribute to virulence, we examined its effect on macrophages. When mouse peritoneal macrophages were harvested and co-cultured with hemolysin-positive V. vulnificus strains (100 bacteria/ cell), about 60% of the macrophages were killed; macrophages were not killed when co-cultured V. vulnificus strain CVD 707, a VvH-negative deletion mutant. Exposure of macrophages to filtered culture supernatants (2.5 HU/ml) and purified VvH (3 HU/ml) resulted in an increase in dead cells (80 and 90%, respectively), as determined by the trypan blue dye exclusion method and LDH release from macrophages was also increased (70 and 65.5%, respectively). The cytotoxic effect of VvH on macrophages was both the dose- and time-dependent. The VvH caused damage to the macrophage membrane and was blocked significantly by preincubation with cholesterol (p<0.01). Fetal bovine serum showed remarkable inhibition of VvH synthesis by V. vulnificus and inhibited VvH activity in culture supernatant. Cell viability was increased by 35% (p<0.01) and LDH release decreased by 28% (P<0.01) when macrophages were incubated with V. vulnificus (100 bacteria/ cell) in DMEM-10% FBS for 2 hr. Bacterial clearance activity of mice against V. vulnificus CVD 707 was decreased by pretreatment with 10 HU of VvH. This result suggests that the VvH can impair the membrane of macrophages and may play a role in the pathogenesis of V. vulnificus septicemia.
Animals ; Cell Survival ; Cholesterol ; Erythrocytes ; Humans ; Macrophages ; Macrophages, Peritoneal* ; Membranes ; Mice* ; Perforin ; Sepsis ; Shock ; Trypan Blue ; Vibrio vulnificus* ; Vibrio* ; Virulence

Sixty-eight clinical isolates of Stenotrophomonas maltophilia from inpatients of 2 university hospitals in Taegu were epidemiologically analyzed by using the minimum inhibitory concentrations of 25 antimicrobial drugs, biochemical reaction, pulsed-field gel elctropgoresis (PFGE), and PCR with enterobacterial repetitive intergenic consensus sequences as primer (ERIC-PCR). 1. All the strains were susceptible to minocycline. More than 57% were susceptible to sulfisomidine (Su), ciprofloxacin (Ci), Ofloploxacin (Of), nalidixic acid (Na), and chloramphenicol (Cm), and 19apprx35% to ceftazidime (Cd), trimethoprim (Tp), Ticacillin-clavulanic acid, and cefoperazone-sulbactam. Most isolates were resistant to beta-lactam antibiotics such as ampicillin (Ap), carbenicillin (Cb), cefotaxim (Ct), cefoxitin (Cx), and aminoglycosides including gentamicin (Gm), tobramycin (Tb), amikacin (Ak). 2. All the isolates were multiply resistant of 5 to 17 drugs and showed 40 different resistance pattern types. 3. All the strains showed very similar biochemical reactions except beta-galactosidase and nitrate reduction test. Fourteen strains selected randomly were classified 10 different pattern type by PFGE and ERIC-PCR. These two methods showed identical result. Four strains isolated from wound in 1994 showed similar MIC pattern and identical API 20NE profile, PFGE, and ERIC-PCR pattern indicating episodes of cross-infection among patients. These results indicate that PFGE or ERIC-PCR profile has comparable discriminatory power for epidemiological typing of S. maltophilia.
Amikacin ; Aminoglycosides ; Ampicillin ; Anti-Bacterial Agents ; beta-Galactosidase ; Carbenicillin ; Cefotaxime ; Cefoxitin ; Ceftazidime ; Chloramphenicol ; Ciprofloxacin ; Consensus Sequence ; Daegu ; Gentamicins ; Hospitals, University ; Humans ; Inpatients ; Microbial Sensitivity Tests ; Minocycline ; Nalidixic Acid ; Polymerase Chain Reaction ; Stenotrophomonas maltophilia* ; Stenotrophomonas* ; Sulfisomidine ; Tobramycin ; Trimethoprim ; Wounds and Injuries

Vibrio vulnificus, a halophilic vibrio is an estuarine gram-negative bacteria that is associated with severe and frequently fatal wound infections and life-threatening septicemia. Bacteriocins are defined as antibacterial substance produced by various species of bacteria which are usually active against closely related organisms. Bacteriocins have found widespread application in epidemiological studies as specific markers of bacteria. It was proposed by Ha et al. (1990. J. Korean. Soc. Microbiol. 25: 586.) to give the bacteriocins produced by V. vulnificus the name "vulnificins". In the present study, a total of 72 strains of V. vulnificus isolated from patients and oysters were subjected to screen potential producers and indicators of vulnificin, applying ultraviolet induction method. Sensitivity of several strains of Serratia marcesans, Pseudomonas aeruginosa, Shigella flexneri, Salmonella typhi and Yersinia enterocolitica to vulnificins were also examined out. All the tested strains of V. vulnificus produced vulnificins active against indicator strains with various different inhibitory patterns. The spectrum of vulnificin activity and sensitive spectrum of indicator strains were considerably broad. Interestingly, almost all strains of S. marcescens, P. aeruginosa, Salmonella sp., Shigella sp. and Y. enterocolitica tested were sensitive to 1-7 vulnificin(s). Taken together, the present study demonstrated that all of the isolates of V. vulnificus produced vulnificins and that 8 good vulnificin producers and 10 good indicators were detected. These strains can be employed efficiently for establishing vulnificin typing scheme of V. vulnificus and for the detection of bacteriocinogeny and sensitivity in V. vulnificus. Biological role of vulnificin remains to be further elucidated.
Bacteria ; Bacteriocins ; Gram-Negative Bacteria ; Humans ; Ostreidae ; Pseudomonas aeruginosa ; Salmonella ; Salmonella typhi ; Sepsis ; Serratia ; Shigella ; Shigella flexneri ; Vibrio vulnificus* ; Vibrio* ; Wound Infection ; Yersinia enterocolitica

No Abstract Available.
Helicobacter pylori* ; Helicobacter*

No Abstract Available.
Clone Cells* ; Cloning, Organism* ; Mycobacterium tuberculosis* ; Mycobacterium*

No Abstract Available.
Astrocytes* ; Humans*

No Abstract Available.
Heparitin Sulfate* ; Orientia tsutsugamushi*

No Abstract Available.
Anti-Infective Agents* ; Enterohemorrhagic Escherichia coli* ; Quinolones* ; Shiga Toxins*

No Abstract Available.
Vibrio parahaemolyticus* ; Vibrio*