Choon-Mee Kim

Publications of Choon-Mee Kim

• Cyclic AMP-receptor protein activates aerobactin receptor IutA expression in Vibrio vulnificus.

  Authors: Choon-Mee Kim, Seong-Jung Kim, Sung-Heui Shin

  Journal of microbiology (Seoul, Korea). 04/2012; 50(2):320-5.

  The ferrophilic bacterium Vibrio vulnificus can utilize the siderophore aerobactin of Escherichia coli for iron acquisition via its specific receptor IutA. This siderophore piracy by V. vulnificusThe ferrophilic bacterium Vibrio vulnificus can utilize the siderophore aerobactin of Escherichia coli for iron acquisition via its specific receptor IutA. This siderophore piracy by V. vulnificus may contribute to its survival and proliferation, especially in mixed bacterial environments. In this study, we examined the effects of glucose, cyclic AMP (cAMP), and cAMP-receptor protein (Crp) on iutA expression in V. vulnificus. Glucose dose-dependently repressed iutA expression. A mutation in cya encoding adenylate cyclase required for cAMP synthesis severely repressed iutA expression, and this change was recovered by in trans complementing cya or the addition of exogenous cAMP. Furthermore, a mutation in crp encoding Crp severely repressed iutA expression, and this change was recovered by complementing crp. Accordingly, glucose deprivation under iron-limited conditions is an environmental signal for iutA expression, and Crp functions as an activator that regulates iutA expression in response to glucose availability.

• Modulation of iron-uptake systems by a mutation of luxS encoding an autoinducer-2 synthase in Vibrio vulnificus.

  Authors: Choon Mee Kim, Sung Heui Shin

  Biological & pharmaceutical bulletin. 01/2011; 34(5):632-7.

  Vibrio vulnificus possesses multiple iron-uptake systems which are mediated by VuuA (vulnibactin receptor), IutA (aerobactin receptor) and HupA (heme receptor). In this study, we determined theVibrio vulnificus possesses multiple iron-uptake systems which are mediated by VuuA (vulnibactin receptor), IutA (aerobactin receptor) and HupA (heme receptor). In this study, we determined the effect of a mutation of luxS encoding autoinducer-2 (AI-2) synthase on the expressions of the three receptors. A mutation and an in trans complementation of luxS did not affect the growing ability of V. vulnificus in iron-deficient conditions. Nevertheless, the luxS mutation slightly decreased vuuA expression, but slightly increased iutA and hupA expressions in the transcriptional reporter assay or Western blot analysis. These changes were all recovered by the luxS complementation. These results suggest that AI-2-mediated quorum sensing system may be involved in the fine modulation of V. vulnificus iron-uptake systems, positively affecting vuuA expression but negatively affecting iutA and hupA expressions.

• Regulation of the Vibrio vulnificus vvpE expression by cyclic AMP-receptor protein and quorum-sensing regulator SmcR.

  Authors: Choon-Mee Kim, Sung-Heui Shin

  Microbial pathogenesis. 12/2010; 49(6):348-53.

  In Vibrio vulnificus, cAMP-receptor protein (CRP) and the quorum-sensing regulator SmcR are simultaneously and cooperatively required for the metalloprotease vvpE gene expression, rather thanIn Vibrio vulnificus, cAMP-receptor protein (CRP) and the quorum-sensing regulator SmcR are simultaneously and cooperatively required for the metalloprotease vvpE gene expression, rather than sequentially in a regulatory cascade. However, this study shows a new temporal and functional sequence between the two factors in regulating vvpE expression. A crp mutation inhibited vvpE expression with growth impairment from early stage. In contrast, a smcR mutation inhibited vvpE expression only at the late stage with no effect on growth. A crp-smcR double mutation severely inhibited vvpE expression with growth impairment from early stage. The inhibited vvpE expression was restored only at the early stage by a crp single complementation, but not at all by a smcR complementation. These results indicate that CRP functions as an essential activator, whereas SmcR functions in the presence of CRP for full vvpE expression.

• Virulence characteristics of sucrose-fermenting Vibrio vulnificus strains.

  Authors: Seong-Jung Kim, Choon-Mee Kim, Sung-Heui Shin

  The Korean journal of laboratory medicine. 10/2010; 30(5):507-10.

  We identified 6 sucrose-fermenting Vibrio vulnificus strains and examined their virulence characteristics. They were all encapsulated, motile, capable of producing toxins and utilizingWe identified 6 sucrose-fermenting Vibrio vulnificus strains and examined their virulence characteristics. They were all encapsulated, motile, capable of producing toxins and utilizing transferrin-bound iron, cytotoxic to cultured cells, and virulent enough to kill mice. They could be definitely identified only by genetic identification methods such as PCR, and not by conventional culture-based identification methods such as API 20E (bioMérieux, France). These results indicate that it is essential to adopt genetic approaches as early as possible in order to avoid misdiagnosis of such strains, especially in clinical situations.

• Outer membrane vesicles of Vibrio vulnificus deliver cytolysin-hemolysin VvhA into epithelial cells to induce cytotoxicity.

  Authors: Young Ran Kim, Bang Ul Kim, Soo Young Kim, Choon Mee Kim, Hee Sam Na, Jeong Tae Koh, Hyon E Choy, Joon Haeng Rhee, Shee Eun Lee

  Biochemical and biophysical research communications. 09/2010; 399(4):607-12.

  The Gram-negative bacterium Vibrio vulnificus produces cytotoxins that induce the acute death of host cells. However, the secretory mechanisms of such cytotoxins have not been extensively studied.The Gram-negative bacterium Vibrio vulnificus produces cytotoxins that induce the acute death of host cells. However, the secretory mechanisms of such cytotoxins have not been extensively studied. Previously, we reported that substantial amounts of V. vulnificus cytolysin-hemolysin (VvhA) are produced in vivo during the bacterial infection in mice and that this cytotoxin, in conjunction with RtxA1, mediates cytotoxicity. In this study, we investigated whether V. vulnificus cells release outer membrane vesicles (OMVs), which are used by some Gram-negative bacteria to deliver virulence factors into host cells. We found that V. vulnificus produce OMVs and that these vesicles can induce host cell death. This process appears to be mediated by VvhA, as evidenced by the finding that OMVs isolated from VvhA-null mutants do not induce cytotoxicity. In addition, cholesterol sequestration in the host cells prevents OMV-mediated VvhA delivery, indicating that VvhA-bearing OMVs interact with cholesterol on the host cell surface. Furthermore, intracellular expression experiments revealed that VvhA-mediated cytotoxicity is driven by its N-terminal leukocidin domain.

• Iron Differentially Regulates Gene Expression and Extracellular Secretion of Vibrio vulnificus Cytolysin-Hemolysin.

  Authors: Choon-Mee Kim, Yoon-Young Chung, Sung-Heui Shin

  The Journal of infectious diseases. 09/2009; 200(4):582-9.

  In a previous study, we showed that Vibrio vulnificus is a ferrophilic bacterium that requires high levels of available iron for growth. In the present study, we show that iron stimulates, in anIn a previous study, we showed that Vibrio vulnificus is a ferrophilic bacterium that requires high levels of available iron for growth. In the present study, we show that iron stimulates, in an unusual manner, the production of cytolysin-hemolysin (VvhA), the most potent exotoxin produced by V. vulnificus. The vvhA gene possesses a putative ferric uptake regulator (Fur)-binding box in the regulatory region, andvvhA transcription was repressed by iron and de-repressed by fur mutation. However, extracellular secretion of VvhA was conversely increased by iron. Iron increased transcription of pilD, which encodes PilD, a component of the type II general secretion system responsible for extracellular VvhA secretion. These results indicate that iron increases extracellular VvhA secretion via the type II general secretion system, although it can repress vvhA transcription via Fur.

• Effect of iron-chelator deferiprone on the in vitro growth of staphylococci.

  Authors: Choon-Mee Kim, Sung-Heui Shin

  Journal of Korean medical science. 05/2009; 24(2):289-95.

  The standard iron-chelator deferoxamine is known to prevent the growth of coagulase-negative staphylococci (CoNS) which are major pathogens in iron-overloaded patients. However, we found thatThe standard iron-chelator deferoxamine is known to prevent the growth of coagulase-negative staphylococci (CoNS) which are major pathogens in iron-overloaded patients. However, we found that deferoxamine rather promotes the growth of coagulase-positive Staphylococcus aureus. Accordingly, we tested whether deferiprone, a new clinically-available iron-chelator, can prevent the growth of S. aureus strains as well as CoNS. Deferiprone did not at least promote the growth of all S. aureus strains (n=26) and CoNS (n=27) at relatively low doses; moreover, it could significantly inhibit the growth of all staphylococci on non-transferrin-bound-iron and the growth of all CoNS on transferrin-bound iron at relatively high doses. At the same doses, it did not at least promote the growth of all S. aureus strains on transferrin-bound-iron. These findings indicate that deferiprone can be useful to prevent staphylococcal infections, as well as to improve iron overload, in iron-overloaded patients.

• Two forms of Vibrio vulnificus metalloprotease VvpE are secreted via the type II general secretion system.

  Authors: Jong Park, So Yeon Ryu, Choon-Mee Kim, Sung-Heui Shin

  Journal of microbiology (Seoul, Korea). 07/2008; 46(3):338-43.

  Vibrio vulnificus has been known to secrete one form of metalloprotease VvpE (45 kDa) that is cleaved to 34 kDa-VvpE and 11 kDa-C-terminal propeptide via extracellular autoproteolysis. However, weVibrio vulnificus has been known to secrete one form of metalloprotease VvpE (45 kDa) that is cleaved to 34 kDa-VvpE and 11 kDa-C-terminal propeptide via extracellular autoproteolysis. However, we found that extracellular secretion of both the 34 and 45 kDa forms of VvpE began in the early growth phase; moreover, 34 kDa-VvpE existed as the major form in V. vulnificus cell lysates and culture supernatants. In addition, extracellular secretion of both 34 and 45 kDa-VvpE was blocked by mutation of the pilD gene, which encodes for the type IV leader peptidase/N-methyltransferase of the type II general secretion system, and the blocked VvpE secretion was recovered by in trans-complementation of the wild-type pilD gene. These results indicate that 34 kDa-VvpE is the major form secreted along with 45 kDa-VvpE from the early growth phase via the PilD-mediated type II general secretion system.

• A widespread deferoxamine-mediated iron-uptake system in Vibrio vulnificus.

  Authors: Choon-Mee Kim, Yong-Jin Park, Sung-Heui Shin

  The Journal of infectious diseases. 12/2007; 196(10):1537-45.

  Vibrio vulnificus can use the standard iron chelator deferoxamine (Desferal) for efficient iron-uptake via the specific receptor DesA, which is encoded by desA. We investigated the ubiquity of theVibrio vulnificus can use the standard iron chelator deferoxamine (Desferal) for efficient iron-uptake via the specific receptor DesA, which is encoded by desA. We investigated the ubiquity of the deferoxamine-mediated iron-uptake system in V. vulnificus strains and the potential risk of the system. By polymerase chain reaction (PCR), desA was found in 10 of 10 clinical strains and in 9 of 10 environmental strains, and their growth was stimulated by deferoxamine. By reverse-transcriptase PCR, desA was expressed only under iron-limited conditions containing deferoxamine. V. vulnificus growth in the presence of deferoxamine was suppressed by desA mutation, and the suppressed growth was recovered by desA complementation. Deferoxamine stimulated V. vulnificus growth in iron-limited in vitro and ex vivo backgrounds containing transferrin-bound iron. Overall, V. vulnificus can use transferrin-bound iron via the widespread deferoxamine-mediated iron-uptake system; this cautions that deferoxamine therapy in patients with iron overload may increase the risk of fatal infections caused by V. vulnificus.

• Production of Vibrio vulnificus metalloprotease VvpE begins during the early growth phase: usefulness of gelatin-zymography.

  Authors: Choon-Mee Kim, Sang-Mee Kang, Ho-Jong Jeon, Sung-Heui Shin

  Journal of microbiological methods. 08/2007; 70(1):96-102.

  Recent studies have demonstrated that expression of the vvpE gene begins during the early growth phase albeit at low levels. However, we found that the traditional protease assay method that is usedRecent studies have demonstrated that expression of the vvpE gene begins during the early growth phase albeit at low levels. However, we found that the traditional protease assay method that is used to measure caseinolytic activity in culture supernatants is not suitable for the measurement of extracellular VvpE that is produced at low levels during the early growth phase. By using gelatin-zymography in place of the protease assay, we could specifically detect only VvpE of several proteases produced by Vibrio vulnificus. Moreover, we could sensitively measure VvpE produced at low levels during the early growth phase, which was consistent with transcription of the vvpE gene. The extracellular production of VvpE was reduced or delayed by mutation of the pilD gene which encodes for the type IV leader peptidase/N-methyltransferase associated with the type II general secretion system; the delayed production of VvpE was recovered by in trans complementation of the wild-type pilD gene. These results indicate that VvpE begins to be produced during the early growth phase via the PilD-mediated type II general secretion system, and that the use of gelatin-zymography is recommended as a simple method for the sensitive and specific detection of VvpE production.

• The pyrH gene of Vibrio vulnificus is an essential in vivo survival factor.

  Authors: Shee Eun Lee, Soo Young Kim, Choon-Mee Kim, Mi-Kwang Kim, Young Ran Kim, Kwangjoon Jeong, Hwa-Ja Ryu, Youn Suhk Lee, Sun Sik Chung, Hyon E Choy, Joon Haeng Rhee

  Infection and immunity. 07/2007; 75(6):2795-801.

  We have suggested an important role of the pyrH gene during the infectious process of Vibrio vulnificus. Previously, we have identified 12 genes expressed preferentially during human infections byWe have suggested an important role of the pyrH gene during the infectious process of Vibrio vulnificus. Previously, we have identified 12 genes expressed preferentially during human infections by using in vivo-induced antigen technology. Among the in vivo-expressed genes, pyrH encodes UMP kinase catalyzing UMP phosphorylation. Introduction of a deletion mutation to the pyrH gene was lethal to V. vulnificus, and an insertional mutant showed a high frequency of curing. We constructed a site-directed mutant strain (R62H/D77N) on Arg-62 and Asp-77, both predicted to be involved in UMP binding, and characterized the R62H/D77N strain compared with the previously reported insertional mutant. We further investigated the essential role of the pyrH gene in the establishment of infection using the R62H/D77N strain. Cytotoxicity was decreased in the R62H/D77N strain, and the defect was restored by an in trans complementation. The intraperitoneal 50% lethal dose of the R62H/D77N strain increased by 26- and 238,000-fold in normal and iron-overloaded mice, respectively. The growth of the R62H/D77N strain in 50% HeLa cell lysate, 100% human ascitic fluid, and 50% human serum was significantly retarded compared to that of the isogenic wild-type strain. The R62H/D77N mutant also had a critical defect in the ability to survive and replicate even in iron-overloaded mice. These results demonstrate that pyrH is essential for the in vivo survival and growth of V. vulnificus and should be an attractive new target for the development of antibacterial drugs and replication-controllable live attenuated vaccines.

• Vibrio vulnificus metalloprotease VvpE is essentially required for swarming.

  Authors: Choon-Mee Kim, Ra-Young Park, Ho-Jong Chun, Soo Young Kim, Joon Haeng Rhee, Sung-Heui Shin

  FEMS microbiology letters. 05/2007; 269(1):170-9.

  Bacterial swarming constitutes a good in vitro model for surface adherence and colonization, and is accompanied by expressions of virulence factors related to invasiveness. In this study, it wasBacterial swarming constitutes a good in vitro model for surface adherence and colonization, and is accompanied by expressions of virulence factors related to invasiveness. In this study, it was determined that Vibrio vulnificus swarming was abolished by mutation of the vvpE gene encoding a metalloprotease VvpE and this swarming defect was recovered by complementation of the vvpE gene. Expression of the vvpE gene began simultaneously with the beginning of swarming and increased along with expression of the luxS gene encoding the synthase of the precursor of quorum-sensing signal molecule autoinducer 2, and this increased vvpE expression was decreased by mutation of the luxS gene. Moreover, VvpE destroyed IgA and lactoferrins, which are responsible for mucosal immunity. These results suggest that VvpE may play important roles in the surface adherence and colonization of V. vulnificus by facilitating swarming and in the mucosal invasion of V. vulnificus by destroying IgA and lactoferrin.

• Ferrophilic characteristics of Vibrio vulnificus and potential usefulness of iron chelation therapy.

  Authors: Choon-Mee Kim, Ra-Young Park, Mi-Hwa Choi, Hui-Yu Sun, Sung-Heui Shin

  The Journal of infectious diseases. 02/2007; 195(1):90-8.

  We determined the ferrophilic characteristics of Vibrio vulnificus to evaluate the potential usefulness of iron chelation therapy for the prevention of V. vulnificus infection. Readily availableWe determined the ferrophilic characteristics of Vibrio vulnificus to evaluate the potential usefulness of iron chelation therapy for the prevention of V. vulnificus infection. Readily available non-transferrin-bound iron (NTBI) is required for the initiation of V. vulnificus growth under in vitro iron-limited conditions and human ex vivo conditions. NTBI aided efficient transferrin-bound iron (TBI) use by V. vulnificus, and the vulnibactin-mediated iron-uptake system was expressed after bacterial growth had been started by NTBI. V. vulnificus required higher NTBI levels for the initiation of growth, produced siderophores at lower levels, and used TBI less efficiently than other bacteria. In addition, the growth of V. vulnificus was inhibited by deferiprone, a clinically available iron chelator. These results show that V. vulnificus is a ferrophilic bacterium that requires higher NTBI levels than other pathogens and that iron chelation therapy might be an effective means of preventing the in vivo growth of V. vulnificus in susceptible patients.

• Pseudomonas aeruginosa alkaline protease can facilitate siderophore-mediated iron-uptake via the proteolytic cleavage of transferrins.

  Authors: Seong-Jung Kim, Ra-Young Park, Sang-Mi Kang, Mi-Hwa Choi, Choon-Mee Kim, Sung-Heui Shin

  Biological & pharmaceutical bulletin. 12/2006; 29(11):2295-300.

  In order to determine whether Pseudomonas aeruginosa alkaline protease AprA is involved in facilitating siderophore-mediated iron-acquisition from human transferrins, we measured bacterial growth,In order to determine whether Pseudomonas aeruginosa alkaline protease AprA is involved in facilitating siderophore-mediated iron-acquisition from human transferrins, we measured bacterial growth, the production of siderophore and AprA, iron-acquisition from transferrins, and the proteolytic cleavage of transferrins in an alkaline minimal medium (pH 8.3) containing human transferrins as an iron source and compared these on a time scale. The growth of P. aeruginosa was found to be stimulated in proportion to the iron-saturation levels of transferrins. AprA production and the proteolytic cleavage of transferrins began concomitantly with siderophore production from the early growth phase when P. aeruginosa was actively growing and consuming most iron for growth. However, the AprA-free, but siderophore-containing, culture ultrafiltrates could also remove iron from transferrin. These results indicate that alkaline protease AprA can facilitate the siderophore-mediated iron-uptake of P. aeruginosa via the proteolytic cleavage of transferrins. However, the proteolytic cleavage by AprA is not essentially required for iron-acquisition from transferrins.

• Vibrio vulnificus metalloprotease VvpE has no direct effect on iron-uptake from human hemoglobin.

  Authors: Hui-Yu Sun, Song Iy Han, Mi-Hwa Choi, Seong-Jung Kim, Choon-Mee Kim, Sung-Heui Shin

  Journal of microbiology (Seoul, Korea). 11/2006; 44(5):537-47.

  This study was designed to determine whether or not Vibrio vulnificus metalloprotease VvpE can promote iron uptake via the proteolytic cleavage of human hemoglobin. We found that V. vulnificusThis study was designed to determine whether or not Vibrio vulnificus metalloprotease VvpE can promote iron uptake via the proteolytic cleavage of human hemoglobin. We found that V. vulnificus utilized hemoglobin as an iron source more efficiently via the vulnibactin-mediated iron-uptake system than via the HupA-mediated iron-uptake system and, of the proteases produced by V. vulnificus, VvpE was found to be the only protease capable of destroying hemoglobin. However, VvpE expression, on both the transcriptional and protein levels, was suppressed in iron-limited media. However, vvpE transcription, but not extracellular VvpE production, was reactivated by the addition of hemoglobin or inorganic iron into iron-limited media. Moreover, vvpE transcription began only in the late growth phase when V. vulnificus had already consumed most of the iron for growth. In addition, neither vvpE mutation nor in trans vvpE complementation affected the ability of V. vulnificus to acquire iron or to grow in iron-limited media or in cirrhotic ascites containing hemoglobin. Hemoglobin added into iron-limited media was not destroyed, but gradually formed an insoluble aggregate during culture; this aggregation of hemoglobin occurred regardless of vvpE mutation or complementation. These results indicate that VvpE is not required for efficient iron uptake from hemoglobin. On the contrary, hemoglobin or iron is required for efficient vvpE transcription. In addition, a discrepancy exists between vvpE transcription and extracellular VvpE production in iron-limited media containing inorganic iron or hemoglobin, which suggests that additional unknown posttranscriptional events may be involved in the extracellular production of VvpE.

• X-gal inhibits the swarming of Vibrio species.

  Authors: Moon-Young Kim, Ra-Young Park, Young-Hoon Bai, Yoon-Young Chung, Choon-Mee Kim, Soo Young Kim, Joon Haeng Rhee, Sung-Heui Shin

  Journal of microbiological methods. 10/2006; 66(3):552-5.

  The expressional levels of genes in swarmer cells can be determined by a simple method using X-gal-containing semisolid agars and lacZ-fusion transcription reporter strains of the genes concerned.The expressional levels of genes in swarmer cells can be determined by a simple method using X-gal-containing semisolid agars and lacZ-fusion transcription reporter strains of the genes concerned. However, X-gal alone inhibited the swarming of Vibrio, regardless of their ability to digest X-gal. Moreover, X-gal inhibited the growth of V. vulnificus containing functional lacZ. These effects of X-gal itself should be carefully considered when trying to determine the expression levels of genes in swarming cells using X-gal-containing semisolid agar.

• Suppression and inactivation of Vibrio vulnificus hemolysin in cirrhotic ascites, a human ex vivo experimental system.

  Authors: Mi-Hwa Choi, Ra-Young Park, Hui-Yu Sun, Choon-Mee Kim, Young-Hoon Bai, Shee Eun Lee, Soo Young Kim, Young Ran Kim, Joon Haeng Rhee, Sung-Heui Shin

  FEMS immunology and medical microbiology. 08/2006; 47(2):226-32.

  To elucidate the mechanisms underlying the in vivo suppression and inactivation of Vibrio vulnificus hemolysin (VvhA), we used cirrhotic ascites fluid as a human ex vivo experimental system. VvhATo elucidate the mechanisms underlying the in vivo suppression and inactivation of Vibrio vulnificus hemolysin (VvhA), we used cirrhotic ascites fluid as a human ex vivo experimental system. VvhA expression was suppressed in proportion to the amount of cirrhotic ascites. The expression of vvhA in undiluted cirrhotic ascites could be suppressed further by the addition of glucose, a constituent of cirrhotic ascites. VvhA was readily inactivated in the presence of cirrhotic ascites by a cholesterol-mediated oligomerization and interaction with an undefined constituent(s) of cirrhotic ascites. These results indicate that the expression of vvhA can be suppressed and that any VvhA produced is inactivated by the constituents of cirrhotic ascites. Our results suggest that only a very small portion of the VvhA that is produced in human body fluids may actually contribute to the pathogenesis of V. vulnificus septicemia. It is suggested that cirrhotic ascites could be used as a human ex vivo experimental system for the studies on the in vivo expression and the significance of V. vulnificus virulence factors.

• Vibrio vulnificus vulnibactin, but not metalloprotease VvpE, is essentially required for iron-uptake from human holotransferrin.

  Authors: Choon-Mee Kim, Ra-Young Park, Jeong-Hee Park, Hui-Yu Sun, Young-Hoon Bai, Phil-Yeol Ryu, Soo Young Kim, Joon Haeng Rhee, Sung-Heui Shin

  Biological & pharmaceutical bulletin. 06/2006; 29(5):911-8.

  The roles of metalloprotease (VvpE) and catechol-siderophore (vulnibactin) in the uptake of iron from human transferrins by Vibrio vulnificus have been determined using different experimentalThe roles of metalloprotease (VvpE) and catechol-siderophore (vulnibactin) in the uptake of iron from human transferrins by Vibrio vulnificus have been determined using different experimental conditions and methods. Therefore, in this study, we attempted to elucidate the roles of VvpE and vulnibactin using the same methods and experimental conditions, in an in vitro and a human ex vivo system, and in accordance with the molecular version of Koch's postulates. Neither vvpE mutation nor in trans vvpE complementation affected vulnibactin production, iron-assimilation from human holotransferrin (HT), and bacterial growth in a HT-containing deferrated Heart-Infusion medium (HT-DF-HI) or a HT-containing cirrhotic ascites (HT-CA). In contrast, the mutation of fur gene encoding Fur, a repressor regulating expression of the vulnibactin-mediated iron-uptake system, derepressed vulnibactin production, and facilitated iron-assimilation from HT and bacterial growth in HT-DF-HI or HT-CA. The mutation of vis gene encoding isochorismate synthase required for vulnibactin synthesis abolished vulnibactin production, iron-assimilation from HT and bacterial growth in HT-DF-HI or HT-CA. These results demonstrate that vulnibactin is essentially required for iron-assimilation from transferrin, and that VvpE has no direct effect on facilitating vulnibactin-mediated iron-assimilation from transferrin in vitro or in a human ex vivo system.

• Swarming differentiation of vibrio vulnificus downregulates the expression of the vvhBA hemolysin gene via the LuxS quorum-sensing system.

  Authors: Moon-Young Kim, Ra-Young Park, Mi-Hwa Choi, Hui-Yu Sun, Choon-Mee Kim, Soo Young Kim, Joon Haeng Rhee, Sung-Heui Shin

  Journal of microbiology (Seoul, Korea). 05/2006; 44(2):226-32.

  Swarming has proven to be a good in vitro model for bacterial surface adherence and colonization, and the swarming differentiation of a bacterium has been shown to be coupled with changes in theSwarming has proven to be a good in vitro model for bacterial surface adherence and colonization, and the swarming differentiation of a bacterium has been shown to be coupled with changes in the expression of virulence factors associated with its invasiveness, particularly in the early stages of infection. In this study, we attempted to determine whether the expression of vvhA, which encodes for hemolysin/cytolysin (VvhA), is either upregulated or downregulated during the swarming differentiation of V. vulnificus. The insertional inactivation of vvhA itself exerted no detectable effect on the expression of V. vulnificus swarming motility. However, in our lacZ-fused vvhA transcriptional reporter assay, vvhA expression decreased in swarming V. vulnificus as compared to non-swarming or planktonic V. vulnificus. The reduced expression of vvhA in swarming V. vulnificus increased as a result of the deletional inactivation of luxS, a gene associated with quorum sensing. These results show that vvhA expression in swarming V. vulnificus is downregulated via the activity of the LuxS quorum-sensing system, suggesting that VvhA performs no essential role in the invasiveness of V. vulnificus via the adherence to and colonization on the body surfaces required in the early stages of the infection. However, VvhA may play a significant role in the pathophysiological deterioration occurring after swarming V. vulnificus is differentiated into planktonic V. vulnificus.

• Effect of the crp mutation on the utilization of transferrin-bound iron by Vibrio vulnificus.

  Authors: Mi-Hwa Choi, Hui-Yu Sun, Ra-Young Park, Choon-Mee Kim, Young-Hoon Bai, Young Ran Kim, Joon Haeng Rhee, Sung-Heui Shin

  FEMS microbiology letters. 05/2006; 257(2):285-92.

  Cyclic AMP-cAMP receptor protein (CRP) complex plays an essential role in the global regulation of Vibrio vulnificus virulence. We found that growth retardation of V. vulnificus caused by mutation ofCyclic AMP-cAMP receptor protein (CRP) complex plays an essential role in the global regulation of Vibrio vulnificus virulence. We found that growth retardation of V. vulnificus caused by mutation of the crp gene encoding CRP was exacerbated under iron-limited conditions. Accordingly, we investigated the effect of crp mutation on the expression of the vulnibactin-mediated iron-uptake system and the ability of V. vulnificus to utilize transferrin-bound iron, and thus to grow in cirrhotic ascites, a human ex vivo system. The production of vulnibactin was suppressed, and the transcription of the vis and vuuA genes, which encode an enzyme required for vulnibactin synthesis and vulnibactin receptor protein, was also suppressed in the crp mutant. Moreover, the crp mutant could not utilize transferrin-bound iron, and its growth was severely suppressed both on transferrin-bound iron and in cirrhotic ascites. All the defects in the crp mutant were recovered by the in trans complementation of the wild-type crp gene. Putative CRP-binding sequences were found in the regulatory regions of the fur, vis and vuuA genes. These results indicate that crp mutation attenuates the ability to grow on transferrin-bound iron and in a human body fluid by down-regulating the vulnibactin-mediated iron-uptake system.