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PubMed2004

rpoB gene sequencing for identification of Corynebacterium species.

The genus Corynebacterium is a heterogeneous group of species comprising human and animal pathogens and environmental bacteria. It is defined on the basis of several phenotypic characters and the results of DNA-DNA relatedness and, more recently, 16S rRNA gene sequencing. However, the 16S rRNA gene is not polymorphic enough to ensure reliable phylogenetic studies and needs to be completely sequenced for accurate identification. The almost complete rpoB sequences of 56 Corynebacterium species were determined by both PCR and genome walking methods. In all cases the percent similarities between different species were lower than those observed by 16S rRNA gene sequencing, even for those species with degrees of high similarity. Several clusters supported by high bootstrap values were identified. In order to propose a method for strain identification which does not require sequencing of the complete rpoB sequence (approximately 3,500 bp), we identified an area with a high degree of polymorphism, bordered by conserved sequences that can be used as universal primers for PCR amplification and sequencing. The sequence of this fragment (434 to 452 bp) allows accurate species identification and may be used in the future for routine sequence-based identification of Corynebacterium species.

Base Sequence↗

PubMed2002

Cloning and expression of fructosyl-amino acid oxidase gene from Corynebacterium sp. 2-4-1 in Escherichia coli.

The gene encoding the fructosyl-amino acid oxidase (fructosyl-alpha-L-amino acid: oxygen oxidoreductase (defructosylating); EC 1.5.3) of Corynebacterium sp. 2-4-1 was cloned and expressed in Escherichia coli. The gene consists of 1,116 nucleotides and encodes a protein of 372 amino acids with a predicted molecular mass of 39,042. The open reading frame was confirmed as the gene of the fructosyl-amino acid oxidase by comparison with the N-terminal amino acid sequence of the purified fructosyl-amino acid oxidase from Corynebacterium sp. 2-4-1. The sequence of the AMP-binding motif, GXGXXG, was found in the deduced N-terminal region. The amino acid sequence of the fructosyl-amino acid oxidase showed no similarity to that of fungal fructosyl-amino acid oxidases. In addition, substrate specificities of this fructosyl-amino acid oxidase were different from those of other fructosyl-amino acid oxidases. The fructosyl-amino acid oxidase of Corynebacterium sp. 2-4-1 is an enzyme that has unique substrate specificity and primary structure in comparison with fungal fructosyl-amino acid oxidases.

Amino Acid Motifs↗

PubMed1992

[Chronic Corynebacterium endophthalmitis. Apropos of 3 cases].

Only a few isolated cases of endophthalmitis have Corynebacterium been implicated as etiology. This diphtheroid, which has been considered for a long time as a nonpathogenic contaminant from the conjunctival flora, may produce systemic diseases usually in immuno-deficient patients. Keratitis and endophthalmitis cases have been reported in the literature. We report three cases of chronic endophthalmitis after extracapsular extraction with intraocular chamber posterior lens which are characterized by many subacute iridocyclitis and vitritis attacks treated by topical steroids. These endophthalmitis are characterized by decrease of visual acuity, hypopion, white plaque on posterior capsule and vitritis. In the first case, Corynebacterium has been isolated from the culture of vitreous and in the second and third cases from the culture of aqueous humor. These bacteria are often very slow growing, 8 to 14 days in the 3 cases. Colonies may not become visible on culture plates before one week or more. Corynebacterium grow well on ordinary media (blood and chocolate agar). The major difficulty is not to discard organism frequently considered contaminants. The treatment associated systemic antibiotherapy with steroids or not, central capsulotomy and vitrectomy with intraocular injection of antibiotic with or without steroids. Antibiotics sensitivities among diphteroids vary greatly. Quinolones, penicillins, vancomycin, cyclines and aminosides are often a good choice. However, individual sensitivities determined by the antibiogram must be used for an appropriate treatment.

Aged↗

PubMed2003

[Purification and characterization of glutamate dehydrogenase. from Corynebacterium glutamicum S9114].

Glutamate dehydrogenase (GDH) is a key enzyme in the biosynthesis of glutamate. The GDHs from Corynebacterium glutamicum S9114 the most commonly used strain in glutamate fermentation, were purified and their molecular structures and properties characterized. The coenzymes were also studied in the hope to increase glutamate production. Cells were harvested at mid-exponential phase by centrifugation and washed with Tris-HCl buffer containing DTT and EDTA (pH 7.5). The cells were then disrupted using a French pressure cell press and the supernatant was collected by centrifugation. The extract was concentrated by 70-fold using the AKTA-100 FPLC system employing a DEAE-cellulose ion exchange column, a hydrophobic interaction chromatography (HIC) and Sephadex G-200 gel filtration. The purified extracts contained NADPH-dependent GDH and NADH-dependent GDH. Both of the enzymes were highly specific for the coenzymes. The molecular masses of the NADPH-dependent GDH and its subunit were 188kD and 32kD respectively, suggesting the enzyme is a homo-hexamer. Our data reported for the first time the presence of NADH- dependent GDH in Corynebacterium glutamicum S9114, similar to other microorganisms containing both GDHs. The NADPH-dependent and NADH-dependent GDH in Corynebacterium glutamicum S9114 may participate in the assimilation and dissimilation of ammonia respectively. The absorptions of NADPH-dependent GDH was very weak at 280nm but very high at 215nm, suggesting a low phenylalanine and tyrosine content in the enzyme.

Chromatography, Gel↗

PubMed2005

[Comparison of the antibiotic sensitivity of lipophilic Corynebacterium sp. isolated from patients on the day of admission and during hospitalization].

Lipophilic species of Corynebacterium are increasing problem in hospital infections. The aim of this study was to evaluate occurrence of these microorganisms in the materials taken from patients in the day of admission and during the hospitalization as well as comparison of their antibiotic sensitivity. The investigation included 65 strains isolated from hospitalized patients and 48 strains isolated from unchanged skin. Using Api Coryne test 5 species were identified. C. urealyticum dominated, the other were C. subsp. lipophilum and C. jeikeium. Among strains isolated from unchanged diseased skin the most C. jeikeium and C. accolens occurred. All strains were sensitive to glycopeptide, quinupristin/dalphopristin. The strains isolated from hospitalized patients were usually sensitive to fuside acid, doxycycline as well as tetracycline. Strains isolated from unchanged skin were sensitive to almost all tested antibiotics. In the group of 65 strains isolated from hospitalized patients 99.0% were multiresistant. In the group of strains isolated from unchanged skin only two strains were multiresistant. Differences in antibiotic sensitivity among analysed Corynebacterium sp. were confirmed. Majority of the "hospital strains" were characterized by multiresistance. Basing on these results it is possible to suppose, that multiresistance is main factor that favours lipophilic Corynebacterium species in the process of colonization of mucous membranes, skins as well as developing infections.

Anti-Bacterial Agents↗

PubMed2006

[Cloning, sequence analysis and expression of N-acetylglutamate kinase gene in Corynebacterium crenatum].

N-Acetylglutamate kinase (EC 2.7.2.8;NAGK) genes from wild-type Corynebacterium crenatum AS 1.542 and a L-arginine-producing mutant C. crenatum 971.1 were cloned and sequenced. Analysis of argB sequences revealed that only one ORF existed, which used ATG as the initiation codon and coded a peptide of 317 amino acids with a calculated molecular weight of 33.6kDa. Only one nucleotide difference was found in the structure gene and the difference did not cause a change of amino acid by comparison of the gene sequences between the wild type C. crenatum AS 1.542 and the mutant 971.1. The ORF sequence of argB from C. crenatum AS 1.542 showed homologies of 99.89%, 76.62%, 37.94% to those from Corynebacterium glutamicum ATCC 13032, Corynebacterium efficient YS-314 and Escherichia coli k12. And the amino acid sequence deduced from ORF displayed homologies of 100%, 78.55%, 25.25% to those from microorganisms above, respectively. An internal promoter was found in the upstream of the argB gene from C. crenatum. The argB gene from C. crenatum AS 1.542 was expressed both in C. crenatum AS 1.542 and 971.1. The NAGK activity of transformed C. crenatum AS 1.542 was greatly increased by the induction of IPTG. The NAGK activity of transformed C. crenatum 971.1 was almost twice as much as that of C. crenatum 971.1 under the same induction. The amplification of the NAGK activity yielded 25% increase of L-arginine production in C. crenatum 971.1.

Amino Acid Sequence↗

PubMed1991

Transformation of Corynebacterium pseudotuberculosis by electroporation.

Corynebacterium pseudotuberculosis was transformed by electroporation, using pNG2, an erythromycin-resistance plasmid from C diphtheriae. Corynebacterium pseudotuberculosis cultivated in brain-heart infusion broth was washed 3 times with water, and resuspended to a final concentration of about 5 x 10(13) colony-forming units/ml. An electroporator constructed in our laboratory incorporated an electrode with 0.8-mm interelectrode gap, using disposable spectrophotometer cuvettes as containers for electroporation. The pNG2 was prepared in Escherichia coli and 4 to 16 micrograms of pNG2 DNA was mixed with 400-microliters amounts of cell suspension in prechilled cuvettes. After incubation on ice for 5 to 10 minutes, the mixture was electroporated at field strengths of up to 18 kV/cm, mixed with 1.5 ml of brain-heart infusion broth, and incubated at 37 C for 2 hours with agitation. Aliquots were then plated on brain-heart infusion blood agar with 15 micrograms of erythromycin/ml. Corynebacterium pseudotuberculosis was transformed at a maximal efficiency of approximately 4 x 10(4) transformants/micrograms of pNG2 DNA. Most total transformants and most transformants per microgram of pNG2 were generated at a field strength of 18 kV/cm. When the concentration of pNG2 DNA was varied, the average total number of transformants increased through a concentration of 30 micrograms/ml, but the efficiency of transformation was highest at the lowest DNA concentration. Transformants contained unmodified pNG2.

Animals↗

PubMed1990

[Classical pseudomembranous diphtheria caused by Corynebacterium ulcerans].

In a 63-year-old patient presenting with signs of classical diphtheria including membranous pharyngotracheobronchitis and myocarditis, Corynebacterium ulcerans was isolated in pure culture from the membranes. The strain produced a toxin specific for C. ulcerans but no diphtheria toxin. The patient was discharged after a complicated hospital course including respiratory and cardiac failure. Besides Corynebacterium diphtheriae, which has very rarely been isolated in Switzerland in recent years, Corynebacterium ulcerans should be considered as a cause of classical diphtheria.

Bacterial Toxins↗

PubMed1979

Corynebacterium pseudogenitalium sp. nov. Commensals of the human male and female urogenital tracts.

Antisera to Corynebacterium genitalium Types C-1 to C-6 were prepared in rabbits and the titers of complement fixing antibodies to the homologous strains, to the heterologous strains, to C. genitalium Types I to V, and to the reference species Corynebacterium xerosis and Corynebacterium minutissimum ascertained. Five Types stimulated low levels of cross-reacting antibodies to all corynebacteria tested including Type C-3. In contrast the antiserum to Type C-3 had antibodies to only two heterologous strains suggesting that these corynebacteria usually shared more than one minor cell wall antigen. The biologic reactions and serotypes of C. genitalium Types C-1 to C-6 have been compared with those of Types I to V. It is considered that C. genitalium should be retained for corynebacteria having the properties of Types I to V whereas corynebacteria having the characteristics of Types C-1 to C-6 that are commensals of the male and female urogenital tracts should be incorporated in a new species Corynebacteria pseudogenitalium sp. nov. The differences in the biologic characteristics of the two species have been discussed and summarized.

Antibodies, Bacterial↗

PubMed1980

Diagnostic utilization of hemolytically active exosubstances of certain Gram-positive bacteria. II. Detection of staphylococcal hemolysins by means of a pre-purified hemolytically active exosubstance of Corynebacterium pseudotuberculosis.

The use of Corynebacterium pseudotuberculosis or of its hemolytically active exosubstance for the detection of staphylococcal hemolysins represents a suitable alterative to the use of the beta-toxin producing strain of Staphylococcus aureus, or of isolated beta-toxin. An increased capture of beta-toxin production was recorded when the CAMP-factor of Streptococcus agalactiae was used. A total of 446 Staphylococcus aureus strains and 48 coagulase-negative staphylococal strains were tested against the prepurified form of hemolytically active exosubstance of Corynebacterium pseudotuberculosis and against the CAMP-factor. The results obtained are in agreement with the testing by means of staphylococcal beta-toxin and of the CAMP-factor. The two modifications of the method for the detection of staphylococcal hemolysins using one plate as proposed by the authors give good results. Their ability of assessment, however, is higher by one degree when the exocubstance of Corynebacterium pseudotuberculosis is employed.

Bacterial Toxins↗

PubMed1993

[Corynebacterium pseudodiphtheriticum pneumonia in a patient diagnosed with HIV infection].

Corynebacterium pseudodiphteriticum is a diphteromorphic bacterium, previously described as pathogenic in very few cases. We present the case of a patient diagnosed of infection by the human immunodeficiency virus (HIV), who developed pneumonia probably caused by Corynebacterium pseudodiphtheriticum and who had a good response to the initial empirical therapy with cephotaxime. We reviewed the literature on patients with diagnosis of HIV infection, acquired immunodeficiency syndrome (AIDS) and AIDS-related complex and we found just one case of this corynebacterium acting as pathogenic and causing a pulmonary abscess.

Acquired Immunodeficiency Syndrome↗

PubMed1995

Evaluation of the applicability of amplified rDNA-restriction analysis (ARDRA) to identification of species of the genus Corynebacterium.

The 16S rRNA genes (rDNA) of 50 strains belonging to 26 different coryneform bacterial species and genomospecies and of the type strain of Rhodococcus equi were enzymatically amplified. Amplified rDNA restriction analysis (ARDRA) with the enzymes AluI, CfoI and RsaI was carried out. The combination of the ARDRA patterns obtained after restriction with these three different enzymes enabled the differentiation between the following species: Corynebacterium accolens (number of strains = 2), C. afermentans subsp. afermentans (2), C. afermentans subsp. lipophilum (2), C. amycolatum (3), CDC coryneform group ANF-1-like (1), CDC coryneform group ANF-3-like (1), C. cystitidis (1), C. diphtheriae (4), C. jeikeium (3), C. macginleyi (2), C. minutissimum (1), C. pilosum (1), C. pseudotuberculosis (2), C. renale (2), C. striatum (2), C. urealyticum (3), C. xerosis (1), CDC coryneform groups B-1 (2), B-3 (2), F-1, genomospecies 1 and 2 (6), G, genomospecies 1 (1) and G, genomospecies 2 (2). The following strains or species could not be differentiated from each other: C. pseudodiphtheriticum (2) from C. propinquum (former CDC coryneform group ANF-3) (2), CDC coryneform group F-1, genomospecies 1 (4) from genomospecies 2 (2) and C. jeikeium genomospecies A (1) from genomospecies C (2). ARDRA may represent a possible alternative for identification of coryneforms, since this technique enabled the identification of most coryneforms tested and since DNA extraction (i.e. cell lysis by boiling), amplification, restriction and electrophoresis can be carried out within 8 hours. This might allow quick identification of C. diphtheriae and other possible pathogens of the genus Corynebacterium.

Corynebacterium↗

PubMedDate not supplied

Exudative pharyngitis possibly due to Corynebacterium pseudodiphtheriticum, a new challenge in the differential diagnosis of diphtheria.

Corynebacterium pseudodiphtheriticum has rarely been reported to cause disease in humans, despite its common presence in the flora of the upper respiratory tract. We report here a case of exudative pharyngitis with pseudomembrane possibly caused by C. pseudodiphtheriticum in a 4-year-old girl. The case initially triggered clinical and laboratory suspicion of diphtheria. Because C. pseudodiphtheriticum can be easily confused with Corynebacterium diphtheriae in Gram stain, clarification of its role in the pathogenesis of exudative pharyngitis in otherwise healthy persons is of public health importance. Simple and rapid screening tests to differentiate C. pseudodiphtheriticum from C. diphtheriae should be performed to prevent unnecessary concern in the community and unnecessary outbreak control measures.

Child, Preschool↗

PubMed1979

[Adriamycin-combination chemotherapy with or without immune stimulation by corynebacterium parvum in metastasizing carcinoma of the breast (author's transl)].

In a randomised, multicentre study the effect of changing the combined cytostatic treatment with VAC (vincristin, adriamycin, cyclophosphamide) to FMC (fluorouracil, methotrexate, cyclophosphamide) was tested, as well as the effect of active non-specific immune stimulation with Corynebacterium parvum, and compared with a control group. In 59 of 120 patients (49%) in whom the results could be analysed there was a measurable significant regression of tumour size to less than 50% of its initial value. The course was stabilized in a further 43 (36%) women. Further tumour-growth progression or death in the early phase of treatment occurred in only 18 women (15%). Average remission was 16 months. Median survival time from start of the study in all analysable patients averaged 22 months. Patients with an ulcer at the site of application of Corynebacterium parvum had a clearly prolonged survival time. In all, immune stimulation with Corynebacterium parvum was no better, either with regard to the remission rate, remission duration or survival time.

Breast Neoplasms↗

PubMed1965

Time-Lapse Photomicrography of Lashing, Flexing, and Snapping Movements in Escherichia coli and Corynebacterium Microcultures.

Hoffman, Heiner (New York University, New York, N.Y.), and Michael E. Frank. Time-lapse photomicrography of lashing, flexing, and snapping movements in Escherichia coli and Corynebacterium microcultures. J. Bacteriol. 90:789-795. 1965.-A series of four consecutive photomicrographs of snapping by an Escherichia coli cell pair indicates that this maneuver consists initially of lashing movements of one cell unattached to the underlying surface and consequent flexion of the second cell, which is strongly attached to the underlying surface at its distal end. This is followed by a final unidirectional movement of the unattached cell which carries the attached cell with it. Lashing movements were also observed in microcultures of fusobacteria and a Corynebacterium, and photomicrographs were obtained of the latter. The observations cannot be interpreted to support the conventional hypothesis that snapping is a response to partial rupture of the capsule during cell division. A new hypothesis is suggested which is based upon motile intracellular granules that have been observed to break out of E. coli, corynebacteria, and fusobacteria cells, typically from the cell ends. Photomicrographs of the minute granules were obtained for both E. coli and the Corynebacterium.

Journal Article↗

PubMed1980

Intrapleural immunotherapy with Corynebacterium parvum in recurrent malignant pleural effusions.

Twenty-one patients with proven recurrent malignant pleural effusions were randomly allocated to treatment groups receiving either intrapleural Corynebacterium parvum in a dose of 7 mg or intrapleural mustine (20 mg). The designated intrapleural therapy was repeated on one occasion if further pleural aspiration was required. Corynebacterium parvum (nine patients) proved superior to mustine (12 patients) in suppressing the reaccumulation of pleural fluid, and was associated with only minimal side-effects of fever and nausea in two patients. Mustine caused marked nausea and vomiting in almost all patients. Three of the four patients who were deemed "failures" after mustine therapy had complete suppression of pleural fluid reaccumulation after a single dose of C parvum, the survival of the fourth being too short to assess a response adequately. There were no failures in the C parvum treated group. Corynebacterium parvum appears to be an effective, well-tolerated agent in the management of recurrent pleural effusions. The relative contribution of its potent immunological stimulant effect to its mode of action remains uncertain.

Aged↗

PubMed2004

Prediction of DtxR regulon: identification of binding sites and operons controlled by Diphtheria toxin repressor in Corynebacterium diphtheriae.

BACKGROUND: The diphtheria toxin repressor, DtxR, of Corynebacterium diphtheriae has been shown to be an iron-activated transcription regulator that controls not only the expression of diphtheria toxin but also of iron uptake genes. This study aims to identify putative binding sites and operons controlled by DtxR to understand the role of DtxR in patho-physiology of Corynebacterium diphtheriae. RESULT: Positional Shannon relative entropy method was used to build the DtxR-binding site recognition profile and the later was used to identify putative regulatory sites of DtxR within C. diphtheriae genome. In addition, DtxR-regulated operons were also identified taking into account the predicted DtxR regulatory sites and genome annotation. Few of the predicted motifs were experimentally validated by electrophoretic mobility shift assay. The analysis identifies motifs upstream to the novel iron-regulated genes that code for Formamidopyrimidine-DNA glycosylase (FpG), an enzyme involved in DNA-repair and starvation inducible DNA-binding protein (Dps) which is involved in iron storage and oxidative stress defense. In addition, we have found the DtxR motifs upstream to the genes that code for sortase which catalyzes anchoring of host-interacting proteins to the cell wall of pathogenic bacteria and the proteins of secretory system which could be involved in translocation of various iron-regulated virulence factors including diphtheria toxin. CONCLUSIONS: We have used an in silico approach to identify the putative binding sites and genes controlled by DtxR in Corynebacterium diphtheriae. Our analysis shows that DtxR could provide a molecular link between Fe+2-induced Fenton's reaction and protection of DNA from oxidative damage. DtxR-regulated Dps prevents lethal combination of Fe+2 and H2O2 and also protects DNA by nonspecific DNA-binding. In addition DtxR could play an important role in host interaction and virulence by regulating the levels of sortase, a potential vaccine candidate and proteins of secretory system.

Bacterial Proteins↗

PubMed1995

Pleurodesis with doxycycline or Corynebacterium parvum in malignant pleural effusion.

Pleurodesis with doxycycline (100 mg and 600 mg) and Corynebacterium parvum (1 mg and 7 mg) were compared in 41 patients with malignant effusion. To evaluate the mechanisms, pleural fluid pH, leukocytes, granulocytes, interleukin-6 (IL-6) and serum IL-6, as well as C-reactive protein (CRP) were measured before and on 2 consecutive days after treatment. Corynebacterium parvum produced a greater acute-phase response measured with fever, serum CRP and IL-6 than doxycycline. However, no change in pleural fluid IL-6 was demonstrated. Among the 35 assessed patients, 26 had objective response, similar in all four treatment groups. Side-effects were more common with Corynebacterium parvum. Based on this preliminary study we conclude that doxycycline, even in low doses, is a highly effective and well tolerated agent for palliative treatment of malignant pleural effusion. As the responses were similar despite different inflammatory reactions, the two agents probably induce pleural obliteration through different mechanisms.

Acute-Phase Reaction↗