Comparison of the Utility of dnaJ and 16S rDNA Sequences for Identification of Clinical Isolates of Vibrio Species

BACKGROUND: Among the many Vibrio species that can cause infections in humans, several species can cause a fatal outcome. Therefore, accurate identification of Vibrio species is very important. Since some species show atypical phenotypic features, selecting an appropriate molecular method is necessary to avoid misdiagnosis. METHODS: Vibrio clinical isolates (N=53) and reference strains (N=8) were used in this study. We analyzed the following sequences for identification: dnaJ gene, 16S rDNA, gyrase B (gyrB) V. vulnificus-specific sequence, gyrB V. navarrensis-specific sequence, and V. vulnificus hemolysin gene PCR (Vvh PCR). We performed phylogenetic analysis of the 16S rDNA, dnaJ, and gyrB sequences. Final identification was based on the combined results of all tests described above. Concordance of the 16S rDNA and dnaJ sequence analysis was measured using the Chi-square test. RESULTS: The 61 Vibrio strains were identified as follows, in descending order: V. vulnificus (78.69%), V. parahaemolyticus (6.56%), V. navarrensis (4.92%), V. mimicus (1.64%), V. cholera (1.64%), V. furnissii (1.64%), V. alginolyticus (1.64%), and Grimontia hollisae (1.64%). The accuracy rates of the dnaJ gene and 16S rDNA sequence for identification were 91.80% and 86.89%, respectively. The 16S rDNA and dnaJ sequences showed a concordance rate of 0.45, which indicates moderate agreement. CONCLUSIONS: Our results suggest that analysis of the dnaJ sequence may be a useful method for the identification of clinical isolates of Vibrio species, especially for distinguishing between closely related Vibrio species.

Cyclic AMP and Cyclic AMP-Receptor Protein are Required for Optimal Capsular Polysaccharide Expression

Vibrio vulnificus causes fatal infections in susceptible individuals. Group 1 capsular polysaccharide (CPS) operon is responsible for CPS expression, which plays an essential role in the pathogenesis of this pathogen. Cyclic AMP (cAMP) and cAMP receptor protein (crp) complex, which responds to glucose availability and functions as a global regulator, has been known to affect CPS production in this pathogen. This study was undertaken to experimentally verify whether cAMP-Crp directly or indirectly affects CPS production. A mutation in cyaA encoding adenylate cyclase, which is required for cAMP biosynthesis, inhibited V. vulnificus growth and changed opaque colonies to translucent colonies, and these changes were recovered by complementing cyaA or by adding exogenous cAMP. A mutation in crp encoding Crp also inhibited V. vulnificus growth and changed opaque colonies to translucent colonies, and these changes were recovered by complementing crp. Moreover, the crp or cyaA mutation decreased the susceptibility of V. vulnificus against NaOCl. The crp mutation reduced the transcription levels of group 1 CPS operon on a per cell basis. Glucose addition in the absence of Crp stimulated V. vulnificus growth, changed translucent colonies to opaque colonies, and increased the transcription levels of group 1 CPS operon. These results indicate that cAMP or Crp is indirectly involved in optimal CPS production by positively affecting metabolism or V. vulnificus growth rather than by directly controlling the expression of group 1 CPS operon.

Multiple High-affinity Iron-uptake Systems of Vibrio vulnificus

Vibrio vulnificus is a Gram-negative halophilic bacterium that causes necrotizing wound infections and fatal septicemia, which mainly occur in patients with elevated serum or tissue iron levels. Accumulated experimental data clearly show that V. vulnificus is a ferrophilic bacterium that requires more available iron for growth than other pathogenic bacteria, has multiple iron-uptake systems, which play important roles in the pathogenesis of the V. vulnificus infections. This review summarized the composition, regulation and significance of V. vulnificus iron-uptake systems. These iron-uptake systems may be attractive candidates for the development of V. vulnificus vaccine. Iron-chelating therapy can also be a promising modality for the prevention and treatment of V. vulnificus infections.

SmcR, the Quorum-sensing Master Regulator, Is Partially Involved in Temperature/Salinity-mediated Changes in Metalloprotease vvpE Expression in Vibrio vulnificus

Vibrio vulnificus, a gram-negative halophilic marine bacterium and opportunistic human pathogen, must withstand various environmental changes, especially simultaneous changes in temperature and salinity, from 25degrees C/2.5% to 37degrees C/0.9% (SCTS) upon entering the human body. In our previous study, SCTS stimulated vvpE expression even in the background of a mutation in luxS encoding LuxS enzyme for the biosynthesis of quorum-sensing (QS) signal molecule autoinducer-2 (AI-2), suggesting that the A1-2-mediated QS system is partially involved in the SCTS-mediated change of vvpE expression. In this study, we examined the effects of the QS master regulator SmcR on SCTS-mediated changes in vvpE expression and extracellular VvpE production. SCTS stimulated V. vulnificus growth, but with no increase in maximal growth levels. The SCTS-mediated prolongation of the stationary growth phase resulted in a significant increase in growth phase-dependent smcR and vvpE expressions. A mutation in smcR seriously repressed vvpE expression, but had no significant effect on V. vulnificus growth. However, the smcR mutation only partially attenuates SCTS-mediated changes in vvpE expression. These results indicate that SCTS stimulates the expressions of smcR and vvpE by stimulating V. vulnificus growth, and that SmcR is only partially involved in SCTS-mediated changes in vvpE expression.

Coordinate Regulation of Vibrio vulnificus Heme Receptor HupA Expression by Cyclic AMP-receptor Protein and Ferric Uptake Regulator

Vibrio vulnificus causes rapid progressing fulminant infections in susceptible individuals, especially those with elevated serum iron levels. This ferrophilic bacterium can directly acquire iron from heme-containing proteins, such as, hemoglobin, via its heme receptor protein HupA. This study was undertaken to determine the roles of cyclic AMP-receptor protein (Crp) as an activator and of ferric uptake regulator (Fur) as a repressor in regulating hupA expression at various iron and glucose concentrations. Under severely iron-deficient conditions, hupA expression in the absence of Crp was induced albeit at low levels and repressed by the addition of iron. In contrast, hupA expression in the presence of Crp was increased by the addition of iron. Under moderately iron-deficient and iron-sufficient conditions, iron addition repressed hupA expression in the presence of Fur, but not in the absence of Fur. Glucose addition repressed hupA expression in the presence of Fur but not in the absence of Fur. Furthermore, a mutation in cyaA encoding adenylate cyclase required for cAMP synthesis hupA expression, and this repression was prevented by the exogenous addition of cAMP. These results indicate that hupA expression is under the coordinate control of cAMP or Crp, which responds to glucose availability, and of Fur, which responds to iron availability, and that Crp is not essential for the constitutional expression of hupA, but is required for the optimal expression of hupA, whereas Fur is essential for the prevention of hupA over-expression.

Vibrio vulnificus Hemolysin Is Easily Inactivated in Spite of Being Produced at High Levels in Cirrhotic Ascites by a fur Mutation

Vibrio vulnificus produces Hemolysin/cytolysin (VvhA), which is one of the most potent exotoxins capable of killing mice at submicrogram levels. However, V. vulnificus growth and vvhA expression are severely repressed and extracellular VvhA produced at low levels is easily inactivated in human body fluids. This study was conducted to obtain additional unequivocal evidence of the enigmatic characteristic of VvhA. V. vulnificus growth was stimulated, vvhA expression was de-repressed, and extracellular VvhA production was increased in cirrhotic ascites, a human ex vivo experimental system, by a mutation of fur encoding ferric uptake regulator, which acts as a transcriptional repressor. However, regardless of the presence or absence of the fur mutation, extracellular VvhA activity was not detected in cirrhotic ascites. These results indicate that VvhA is easily inactivated even when vvhA expression and extracellular VvhA production are maintained at high levels in cirrhotic ascites.

Change of Vibrio vulnificus Metalloprotease VvpE Production by Temperature and Salinity

Vibrio vulnificus, a gram-negative halophilic marine bacterium and opportunistic pathogen, must withstand various environmental changes, especially the simultaneous change of temperature and salinity (SCTS) from 25degrees C/2.5% to 37degrees C/0.9% upon entering the human body. Previous studies have suggested that temperature and salinity may affect the production of metalloprotease VvpE via the LuxS-mediated autoinducer-2 quorum sensing system (AI-2-QSS). However, this hypothesis remains to be verified through coherent experiments. In this study, SCTS stimulated V. vulnificus growth with no increase in total growth levels. The SCTS-mediated prolongation of the stationary growth phase resulted in a significant increase in growth phase-dependent luxS and vvpE transcriptions; however, SCTS did not affect luxS or vvpE transcription levels during the exponential growth phase. SCTS also advanced extracellular VvpE production, which was consistent with vvpE transcription and V. vulnificus growth. SCTS-mediated modulation of vvpE expression was slightly attenuated but still observed in the background of a luxS mutation which seriously repressed vvpE expression. These results indicate that SCTS stimulates luxS and vvpE expression by stimulating V. vulnificus growth; however, the LuxS-mediated AI-2-QSS plays only a minor role, if any, in the SCTS-mediated modulation of vvpE expression.

Virulence Characteristics of Sucrose-fermenting Vibrio vulnificus Strains / 대한진단검사의학회지

We 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 (bioMerieux, 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.

Effect of Iron-Chelator Deferiprone on the In Vitro Growth of Staphylococci

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 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.

Evaluation of Infrequent-Restriction-Site PCR for Epidemiological Typing of Candida tropicalis / 대한임상미생물학회지

BACKGROUND: We evaluated the usefulness of a newly developed molecular typing method of infrequent restriction site polymerase chain reaction (IRS-PCR) as an epidemiological DNA fingerprinting tool for Candida tropicalis. METHODS: Thirty-two strains of C. tropicalis comprising eight sporadic strains and 24 clonal strains belonging to six clones, of which clonal type were previously confirmed by pulsed-field gel electrophoresis (PFGE), were tested by IRS-PCR to evaluate the usefulness of this technique. Twenty strains of Candida species, including C. glabrata, C. krusei, C. albicans, and C. parapsilosis, were also tested to assess the ability of IRS-PCR to discriminate among species of Candida. RESULTS: Using the IRS-PCR assay, sporadic strains of C. tropicalis could not be differentiated from clonal strains. Most strains belonging to the same clones were classified as different IRS-PCR types or clusters, and some different sporadic strains were classified as the same IRS-PCR types. When pattern variation was examined for different strains of C. tropicalis using IRS-PCR, pairwise similarity measured by the Dice coefficient was 75.4~100%. In contrast, pairwise similarity among isolates of five different species of Candida was 25~69.2%. Therefore, five different species of Candida were easily differentiated. CONCLUSION: The IRS-PCR typing assay appears to be an inadequate tool for the epidemiological typing of C. tropicalis, because the typing result of IRSPCR is not comparable to that of PFGE. To our knowledge, this is the first evaluation study for IRSPCR as an epidemiological typing tool for C. tropicalis.

Prevalence of Carbapenemase and Integrase Genes in Imipenem-resistant Acinetobacter baumannii and Pseudomonas aeruginosa Collected over Several Years in a University Hospital / 대한임상미생물학회지

BACKGROUND: The incidence of infections with imipenem- resistant Acinetobacter baumannii (IRAB) and Pseudomonas aeruginosa (IRPA) is increasing worldwide, and recent molecular studies indicate that the prevalence of carbapenemases is increasing in various parts of the world. However, few long-term longitudinal studies have assessed the prevalence of IRAB- and IRPA-derived carbapenemases and integrases in a hospital setting in Korea. METHODS: The carbapenemase genes (blaOXA-23, blaOXA-24, blaOXA-58, blaIMP-1, blaVIM-2, blaSIM-1, blaSPM-1) and integrase genes (intI1, intI2, intI3) produced by 46 IRAB strains and 51 IRPA strains collected at Chosun University Hospital between 2003 and 2006 were determined by PCR. RESULTS: The IRAB strains produced class 1 integrases more often than did the IRPA strains. However, the incidence increased steadily in both strains, reaching 100% in 2006. Carbapenemases of blaIMP-1 and blaVIM-2 types were found in 57% and 64% of the IRAB strains, respectively, in 2003. However, only one strain with blaVIM-2 was found in 2004 and another one with blaIMP-1 in 2005. The prevalence of carbapenemases was very low in the IRPA strains, just one strain with blaVIM-2 in 2005 and another one with blaoxa-23 in 2006. No other types of carbapenemase genes were detected in both strains. Rep-PCR of IRAB strains in 2003 showed different patterns. CONCLUSION: The incidence of carbapenemase varied by year but was generally low, except in 2003. The prevalence of class 1 integrases was consistently high and increased every year. The reason for the high prevalence of carbapenemases in 2003 is still unknown, but we assumed that it was not from the spread of a clone containing either blaIMP-1 or blaVIM-2 because the strains exhibited different rep-PCR patterns.

Frequency of Mutation of Codon 249, Overexpression of p53, and Hepatitis B Virus DNA Positivity in Hepatocellular Carcinoma / 대한임상미생물학회지

BACKGROUND: In hepatocellular carcinoma (HCC), the frequency of p53 mutation and the association with hepatitis B virus (HBV) infection varies with geographic locations and risk factors. The aim of this study was to determine the frequency of codon 249 mutation of p53, p53 overexpression, and HBV DNA positivity and to observe the relationship between them in Korean HCC. METHODS: We analyzed overexpression of p53 in hepatoma tissue from 17 HCC patients by immunohistochemistry (IHC), specific mutations at the third base position of codon 249 by PCR-restriction fragment length polymorphism (PCR-RFLP) method, and presence of HBV by nested PCR. RESULTS: Although a point mutation at codon 250 was seen in one (5.8%) of 17 patients, no codon 249 mutations were found in the patient cohort. The p53 protein was overexpressed in 4 (23.5%) of 17 HCCs. PCR for HBV DNA from HCCs showed a positivity rate of 82.4% (14 of 17 specimens). CONCLUSION: In HCC of this study, HBV infection was not associated with either 249 mutation or overexpression of p53, and overexpression of p53 protein seemed to be related to other than this mutation.

Effect of Maternal Thyroxine Treatment on the Postnatal Development of Brain-derived Neurotrophic Factor-containing Neuron in the Brain of Pups of Alcohol Abused Mother / 대한해부학회지

Maternal alcohol abuse is thought to be the common cause of mental retardation. Especially, continuous alcohol consumption during critical period of brain development induce fetal alcohol effects. In this study, the authors investigated the effects of maternal alcohol drinking on the postnatal changes of BDNF contents and patterns of BDNF-containing neuron in neonatal rat brain, and, the influence of maternal thyroxine treatment on the brain of pups of alcohol abused mother. Pregnant rats were divided into three groups. Alcohol-fed group (n=4) received 35 calories of liquid alcohol diet daily from gestation day 6; control pair-fed group (n=4) was fed a liquid diet in dextrin replaced alcohol isocalorically; alcohol+T4 group (n=4) received 35 calories liquid alcohol diet and exogenous thyroxine (5 microgram/kg/day) subcutaneously. The amount of BDNF was significantly higher in the alcohol+T4 group as compared to the alcohol group at P7, P14 and P21, especially, alcohol+T4-exposed pups showed a significant increase of BDNF at P7. The decrease in BDNF was found in alcohol group compared to control pair-fed group at all ages. In alcohol+T4 group, BDNF-containing Purkinje cells exhibited mature pattern and monolayer arrangement at P14. Alcohol+T4 group showed mature pattern and numerical increase of BDNF-containing cells in cerebral cortex, hypothalamus and hippocampus at P7. The BDNF immunoreactivity of hippocampus continued to show prominent configuration in alcohol+T4 group at P28. These results indicate that the increase of the BDNF-containing neurons and BDNF amount in pups of thyroxinesupplemented alcohol-exposed dams as compared to control pair-fed and alcohol-exposed pups at P7, presumably suggest the early postnatal growth stimulatory effect of the exogenously supplemented thyroxine. Therefore, the increase of BDNF synthesis caused by maternal administration of exogenous thyroxine may ameliorate fetal alcohol effects, one of the ill effects as a result of the dysthyroid state following maternal alcohol abuse.

Growth of Streptonigrin-Resistant Staphylococcus epidermidis with Defective Siderophore-Mediated Iron-Uptake System in Human Peritoneal Dialysate Solution / 감염과화학요법

BACKGROUND: Staphylococcus epidermidis is the most common pathogen of chronic ambulatory peritoneal dialysis peritonitis. It has been believed that the activity of iron-uptake system (IUS) may play an important role in the growth of S. epidermidis in human peritoneal dialysate (HPD) solution, but there is no report using mutants with defective IUS. A streptonigrin-resistant S. epidermidis (SRSE) strain was isolated from S. epidermidis KCTC 1917 and functionally characterized. MATERIALS AND METHODS: Bacterial growth was monitored by measuring the optical densities of culture fluids obtained at appropriate intervals at a wavelength of 600 nm. CAS agar diffusion assay was used for the comparison of siderophore production, 6 M urea-gel electrophoresis for the comparison of the ability to capture iron from transferrin, and bioassay for the observation of the ability to utilize iron-siderophore complexes. RESULTS: The SRSE strain ineffectively utilized transferrin-bound iron for growth despite its ability to produce considerably larger amount of siderophores than its parental strain. The growth of the parental strain, but not the SRSE strain, was stimulated on transferrin-bound iron by its own siderophores each. The growth of the SRSE strain in the HPD solution was retarded compared to that of the parental strain. CONCLUSION: These results indicate that the SRSE strain is defective in its ability to utilize the iron-siderophore complexes, rather than its ability to produce siderophores, and that the siderophore-mediated IUS plays an important role in the growth of S. epidermidis in HPD solution.

Growth of Streptonigrin-Resistant Staphylococcus epidermidis with Defective Siderophore-Mediated Iron-Uptake System in Human Peritoneal Dialysate Solution / 감염과화학요법

BACKGROUND: Staphylococcus epidermidis is the most common pathogen of chronic ambulatory peritoneal dialysis peritonitis. It has been believed that the activity of iron-uptake system (IUS) may play an important role in the growth of S. epidermidis in human peritoneal dialysate (HPD) solution, but there is no report using mutants with defective IUS. A streptonigrin-resistant S. epidermidis (SRSE) strain was isolated from S. epidermidis KCTC 1917 and functionally characterized. MATERIALS AND METHODS: Bacterial growth was monitored by measuring the optical densities of culture fluids obtained at appropriate intervals at a wavelength of 600 nm. CAS agar diffusion assay was used for the comparison of siderophore production, 6 M urea-gel electrophoresis for the comparison of the ability to capture iron from transferrin, and bioassay for the observation of the ability to utilize iron-siderophore complexes. RESULTS: The SRSE strain ineffectively utilized transferrin-bound iron for growth despite its ability to produce considerably larger amount of siderophores than its parental strain. The growth of the parental strain, but not the SRSE strain, was stimulated on transferrin-bound iron by its own siderophores each. The growth of the SRSE strain in the HPD solution was retarded compared to that of the parental strain. CONCLUSION: These results indicate that the SRSE strain is defective in its ability to utilize the iron-siderophore complexes, rather than its ability to produce siderophores, and that the siderophore-mediated IUS plays an important role in the growth of S. epidermidis in HPD solution.

Utilization of Transferrin-Bound Iron by Medically Important Staphylococcal Species

Staphylococcus aureus is able to utilize efficiently transferrin-bound iron as an iron source, whereas other staphylococci are not. The reason for this difference remains unclear. We compared the activity of siderophore-mediated iron-uptake systems among S. aureus, S. epidermidis, and S. saprophyticus. S. aureus was more susceptible to streptonigrin than the other two staphylococci. S. aureus was able to utilize efficiently transferrin-bound iron in proportion to the level of iron-saturation and produced siderophores in an inverse relation to iron-saturation. In contrast to S. aureus, S. epidermidis and S. saprophyticus were able to utilize only holotransferrin (HT; about 80% iron- saturated) and produced siderophores only in media containing HT. Moreover, they utilized HT less efficiently than S. aureus, though they produced greater amount of siderophores than S. aureus in media containing HT. The ability of the equivalent siderophores per se to capture iron from HT was not significantly different among the three species. Nevertheless, the siderophores from S. aureus stimulated the growth of the staphylococci to a greater degree than did the siderophores from S. epidermidis and S. saprophyticus. The siderophores from S. epidermidis and S. saprophyticus also stimulated the growth of S. aureus to a greater degree than those of the original bacteria which produced them. This indicates that S. aureus possesses a greater ability to produce more-efficient siderophores responding to very low iron-availability, as well as a greater ability to utilize iron-siderophore complexes, than the other two staphylococci. This explains in part the higher virulence of S. aureus compared to other staphylococci.

Immunoreactivity of Iron-Repressible Cell Wall Proteins and Exoproteins of Staphylococcus aureus / 감염과화학요법

In the present study, we tried to investigate immunoreactivity of iron-repressible cell wall proteins and exoproteins of Staphylococcus aureus when the convalescent serum obtained from the patient with septicemia was applied. On SDS-PAGE and western blot analysis, several iron-repressible cell wall proteins expressed in iron-deficient BHI showed strong immunoreactivity, whereas no or relatively weak immunoreactivity was shown in iron-sufficient BHI. Several exoproteins were expressed only in iron-deficient BHI and these exoproteins showed strong immunoreactivity. These results indicate that several iron-repressible cell wall proteins and exoproteins are expressed and are immunogenic in vivo. Since in vivo state is an relatively iron-restricted condition, we recommend the use of iron-deficient media for studies concerning pathogenicity of staphylococcal in human.

Immunoreactivity of Iron-Repressible Cell Wall Proteins and Exoproteins of Staphylococcus aureus / 감염과화학요법

In the present study, we tried to investigate immunoreactivity of iron-repressible cell wall proteins and exoproteins of Staphylococcus aureus when the convalescent serum obtained from the patient with septicemia was applied. On SDS-PAGE and western blot analysis, several iron-repressible cell wall proteins expressed in iron-deficient BHI showed strong immunoreactivity, whereas no or relatively weak immunoreactivity was shown in iron-sufficient BHI. Several exoproteins were expressed only in iron-deficient BHI and these exoproteins showed strong immunoreactivity. These results indicate that several iron-repressible cell wall proteins and exoproteins are expressed and are immunogenic in vivo. Since in vivo state is an relatively iron-restricted condition, we recommend the use of iron-deficient media for studies concerning pathogenicity of staphylococcal in human.

Bacterial Growth in Amniotic Fluid Is Dependent on the Iron-Availability and the Activity of Bacterial Iron-Uptake System

In the present study, the relationship among iron-availability, antibacterial activity, role of meconium as an iron source and the activity of bacterial iron-uptake system (IUS) for bacterial growth in amniotic fluid (AF) were investigated. Staphylococcus aureus ATCC 6538 and its streptonigrin-resistant (SR) mutant with defective IUS were used as the test strains. The growth of S. aureus in AF was stimulated dosedependently by addition of meconium. Bacterial growth stimulated by meconium was re-inhibited dose-dependently by addition of iron-chelator, dipyridyl and apotransferrin. Iron concentration was correlated with the meconium content in AF (r(2)= 0.989, p=0.001). High-affinity IUS of S. aureus was expressed only in AF but not in AF with meconium. The growth of SR strain was more retarded than that of the parental strain in the iron-deficient brain heart infusion (ID-BHI), clear AF and AF containing apotransferrin. The retarded growth of both strains in the ID-BHI and AF was recovered by addition of holotransferrin, hemoglobin and FeCl3. Taken together, the antibacterial activity of AF is closely related with low iron-availability. Bacterial growth in AF considerably depends on the activity of bacterial IUS. Meconium acts as one of the exogenous iron-sources and thus can stimulate bacterial growth in AF.

Transferrin Binding Protein of Staphylococcus aureus is not Essential for Iron-uptake from Transferrin in vitro / 감염과화학요법

BACKGROUND: Staphylococcus aureus produces siderophore and expresses transferring-binding protein for uptake of iron essentially required for multiplication under the iron-restricted conditions. We investigated which of the two iron-uptake systems is dominant or essential for S. aureus growth. METHODS: Streptonigrin-resistant mutant (SR-1) of S. aureus, isolated by repeated exposure to streptonigrin, was used in this study. SR-1 strain produced less amount of siderophore than the parent strain and did not express the transferring-binding protein. Growth and iron consumption of the two bacteria were compared during cultivation in the media containing various concentrations of iron, by measuring optical densities of the culture fluids at 600 nm and residual iron concentrations of the culture supernatants. RESULTS: Growth of the two bacteria in the iron-sufficient media containing various sources of iron was more active than in the iron-deficient media, in which iron concentration was 0.2 microgram/dL. Growth of SR-1 strain was markedly inhibited in the iron-deficient media with/without apotransferrin (0.5 mg/mL). However, growth of SR-1 strain did not show any difference in the iron-sufficient media containing FeCl3 (0.2 micrometer) and hemoglobin (0.5 mg/mL) as compared with that of the parental strain. Moreover, growth of the two bacteria did not show any difference in the media containing holotransferrin (0.5 mg/mL). There was no difference in iron consumption between the parental and mutant strains, evidenced by the residual iron concentration of the culture supernatants. CONCLUSION: Siderophore production is thought to be dominant or essential iron-uptake system for in vitro growth of S. aureus. The pathogenetic role of multifunctional transferring-binding protein, including transferring-binding activity and glycolytic enzyme activity, is expected to be established through further continued studies.