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K C Jeong

Publications and source records attributed to K C Jeong.

6 recordsLinked to original sources

Isolation of Escherichia coli O157:H7 from the gall bladder of inoculated and naturally-infected cattle.

To determine if Escherichia coli O157:H7 is capable of residing in the gall bladder of cattle, inoculation studies were conducted with O157:H7 strain 86-24 in weaned Holstein calves. Strain 86-24 was isolated from the gall bladders of five calves 36 days after inoculation. Two other calves contained the inoculation strain in the distal colon but the organism was absent in their gall bladders. A second trial in which the calves were euthanized 15 days after inoculation found strain 86-24 in six of seven inoculated calves but only in colon and/or rumen samples. In a third trial that inoculated eight calves with a four-strain cocktail of O157:H7 strains, the gall bladders from all eight animals were positive 9 days after inoculation. The colon and rumen samples from these calves were also positive. E. coli O157:H7 isolates recovered from bile samples and subtyped by pulsed field gel electrophoresis found that three of the four inoculation strains were present in one or more of the calves. Thus, residence in the gall bladder is not restricted to a single strain. Additional evidence of the ability to localize in the gall bladder of cattle was provided by testing the bile from 150 gall bladders (five collection dates, 30 samples each) obtained at an abbatoir and the isolation of E. coli O157:H7 from four samples (2.7%). This study establishes that E. coli O157:H7 can reside transiently or permanently at a low level in the gall bladder of cattle.

Animals↗

Differential expression of Vibrio vulnificus elastase gene in a growth phase-dependent manner by two different types of promoters.

Elastase activity of Vibrio vulnificus was highly dependent on growth phase, reached a maximum during the stationary phase, and was regulated at the level of transcription. The stationary phase production of elastase in crp or rpoS mutants, which were constructed by allelic exchanges, decreased about 3- and 10-fold, respectively. However, the promoter activity of vvpE encoding elastase was unaffected by those mutations in the log phase when analyzed using a vvpE-lux fusion. A primer extension analysis revealed that the transcription of vvpE begins at two different sites, consisting of putative promoter L (PL) and promoter S (PS). The PL activity was constitutive through the log and stationary phases, lower than the PS activity, and unaffected by the crp or rpoS mutations. The transcription of PS, induced only in the stationary phase, was dependent on RpoS. The mutation in crp reduced the activity of PS; however, the additional inactivation of crp did not influence the PS activity in the rpoS mutant, indicating that CRP exerted its effects through PS requiring RpoS. These results demonstrate that vvpE expression is differentially directed by PL and PS depending on the growth phase and elevated by RpoS and CRP in the stationary phase.

Alleles↗

Construction and phenotypic evaluation of a Vibrio vulnificus vvpE mutant for elastolytic protease.

Vibrio vulnificus is an opportunistic gram-negative pathogen that commonly contaminates oysters. Predisposed individuals who consume raw oysters can die within days from sepsis, and even otherwise healthy people are susceptible to serious wound infection after contact with contaminated seafood or seawater. Numerous secreted and cell-associated virulence factors have been proposed to account for the fulminating and destructive nature of V. vulnificus infections. Among the putative virulence factors is an elastolytic metalloprotease. We cloned and sequenced the vvpE gene encoding an elastase of V. vulnificus ATCC 29307. The functions of the elastase were assessed by constructing vvpE insertional knockout mutants and evaluating phenotypic changes in vitro and in mice. Although other types of protease activity were still observed in vvpE mutants, elastase activity was completely absent in the mutants and was restored by reintroducing the recombinant vvpE gene. In contrast to previous characterization of elastase as a potential virulence factor, which was demonstrated by injecting the purified protein into animals, inactivation of the V. vulnificus vvpE gene did not affect the ability of the bacteria to infect mice and cause damage, either locally in subcutaneous tissues or systemically in the liver, in both iron-treated and normal mice. Furthermore, a vvpE mutant was not affected with regard to cytolytic activity toward INT407 epithelial cells or detachment of INT407 cells from culture dishes in vitro. Therefore, it appears that elastase is less important in the pathogenesis of V. vulnificus than would have been predicted by examining the effects of administering purified proteins to animals. However, V. vulnificus utilizes a variety of virulence factors; hence, the effects of inactivation of elastase alone could be masked by other compensatory virulence factors.

Animals↗

Vibrio vulnificus has the transmembrane transcription activator ToxRS stimulating the expression of the hemolysin gene vvhA.

In an attempt to dissect the virulence regulatory mechanism in Vibrio vulnificus, we tried to identify the V. cholerae transmembrane virulence regulator toxRS (toxRS(Vc)) homologs in V. vulnificus. By comparing the sequences of toxRS of V. cholerae and V. parahaemolyticus (toxRS(Vp)), we designed a degenerate primer set targeting well-conserved sequences. Using the PCR product as an authentic probe for Southern blot hybridization, a 1.6-kb BglII-HindIII fragment and a 1.2-kb HindIII fragment containing two complete open reading frames and one partial open reading frame attributable to toxR(Vv), toxS(Vv), and htpG(Vv) were cloned. ToxR(Vv) shared 55.0 and 63.0% sequence homology with ToxR(Vc) and ToxR(Vp), respectively. ToxS(Vv) was 71.5 and 65.7% homologous to ToxS(Vc) and ToxS(Vp), respectively. The amino acid sequences of ToxRS(Vv) showed transmembrane and activity domains similar to those observed in ToxRS(Vc) and ToxRS(Vp). Western blot analysis proved the expression of ToxR(Vv) in V. vulnificus. ToxRS(Vv) enhanced, in an Escherichia coli background, the expression of the V. vulnificus hemolysin gene (vvhA) fivefold. ToxRS(Vv) also activated the ToxR(Vc)-regulated ctx promoter incorporated into an E. coli chromosome. A toxR(Vv) null mutation decreased hemolysin production. The defect in hemolysin production could be complemented by a plasmid harboring the wild-type gene. The toxR(Vv) mutation also showed a reversed outer membrane protein expression profile in comparison to the isogenic wild-type strain. These results demonstrate that ToxR(Vv) may regulate the virulence expression of V. vulnificus.

Amino Acid Sequence↗

Thermal-death times of opaque and translucent morphotypes of Vibrio vulnificus.

Thermal-death times were determined for Vibrio vulnificus strains with different morphotypes. Opaque strains showed higher D values (times required to reduce the viable population of a given strain by 90%) than translucent strains. Z values (absolute values of the temperature required to reduce 1 log scale of D values) were also significantly higher in opaque morphotypes (2.4 to 2.5 degrees C) than in translucent ones (1.7 to 2.1 degrees C). These results indicate that the morphotype is related to the organism's susceptibility to heat.

Bacteriological Techniques↗