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W Ziebuhr

Publications and source records attributed to W Ziebuhr.

At least 19 recordsLinked to original sources
PubMed2003

Detection of Staphylococcus aureus by 16S rRNA directed in situ hybridisation in a patient with a brain abscess caused by small colony variants.

A 45 year old man was admitted to hospital with a right sided facial paralysis and three month history of seizures. Computed tomography showed a left temporal mass including both intracerebral and extracerebral structures. Ten years earlier the patient had undergone a neurosurgical intervention in the same anatomical region to treat a subarachnoid haemorrhage. In tissue samples and pus obtained during neurosurgery, Staphylococcus aureus was detected by a 16S rRNA-directed in situ hybridisation technique. Following long term cultivation, small colony variants (SCV) of methicillin resistant S aureus were identified. The patient was treated successfully with a combination of vancomycin and rifampin followed by prolonged treatment with teicoplanin, with no sign of infection on follow up nine months after discharge. This is the first report in which S aureus SCV have been identified as causative organisms in a patient with brain abscess and in which in situ hybridisation has been used to detect S aureus in a clinical specimen containing SCV. Antimicrobial agents such as rifampin which have intracellular activity should be included in treatment of infections caused by S aureus SCV.

Brain Abscess↗
PubMed2000

Evolution of microbial pathogens.

Various genetic mechanisms including point mutations, genetic rearrangements and lateral gene transfer processes contribute to the evolution of microbes. Long-term processes leading to the development of new species or subspecies are termed macroevolution, and short-term developments, which occur during days or weeks, are considered as microevolution. Both processes, macro- and microevolution need horizontal gene transfer, which is particularly important for the development of pathogenic microorganisms. Plasmids, bacteriophages and so-called pathogenicity islands (PAIs) play a crucial role in the evolution of pathogens. During microevolution, genome variability of pathogenic microbes leads to new phenotypes, which play an important role in the acute development of an infectious disease. Infections due to Staphylococcus epidermidis, Candida albicans and Escherichia coli will be described with special emphasis on processes of microevolution. In contrast, the development of PAIs is a process involved in macroevolution. PAIs are especially important in processes leading to new pathotypes or even species. In this review, particular attention will be given to the fact that the evolution of pathogenic microbes can be considered as a specific example for microbial evolution in general.

Candida albicans↗
PubMed2000

Toxin genes on pathogenicity islands: impact for microbial evolution.

Toxin-specific genes are often located on mobile genetic elements such as phages, plasmids and pathogenicity islands (PAIs). The uropathogenic E. coli strain 536 carries two alpha-hemolysin gene clusters, which are part of the pathogenicity islands I536 and II536, respectively. Using different genetic techniques, two additional PAIs were identified in the genome of the E. coli strain 536, and it is likely that further PAIs are located on the genome of this strain. Pathogenicity islands are often associated with tRNA genes. In the case of the E. coli strain 536, the PAI-associated tRNA gene leuX, which encodes a minor leucyl-tRNA, affects the expression of various virulence traits including alpha-hemolysin production. The exact mode of action of the tRNA5Leu-dependent gene expression has to be identified in the future.

Bacterial Toxins↗
PubMed2000

Chromosomal rearrangements affecting biofilm production and antibiotic resistance in a Staphylococcus epidermidis strain causing shunt-associated ventriculitis.

During two clinical courses of shunt-associated meningitis in a 3-month-old child, five multiresistant S. epidermidis isolates were obtained and analyzed with regard to biofilm production and antibiotic susceptibility. Three S. epidermidis strains, which were initially isolated from the cerebrospinal fluid, produced biofilms on polystyrene tissue culture plates. Following antibiotic treatment and subsequent exchange of the shunt system, sterilization of the CSF was achieved. However, after three weeks a relapse of the infection occurred. The two S. epidermidis isolates obtained now were biofilm negative, but showed an identical resistance pattern as those from the previous infection, except that resistance to rifampicin and increased mininal inhibitory concentrations of aminoglycoside antibiotics had emerged. DNA fingerprinting by PFGE indicated the clonal origin of all isolates. However, some DNA rearrangements and differences in the IS256-specific hybridization patterns could be identified in the isolates from the second infection period that led to altered biofilm formation and increased expression of aminoglycoside resistance traits. The data evidence that variation of biofilm expression occurs in vivo during an infection and highlight the extraordinary genome flexibility of pathogenic S. epidermidis.

Anti-Bacterial Agents↗
PubMed2000

Effect of subinhibitory antibiotic concentrations on polysaccharide intercellular adhesin expression in biofilm-forming Staphylococcus epidermidis.

Biofilm production is an important step in the pathogenesis of Staphylococcus epidermidis polymer-associated infections and depends on the expression of the icaADBC operon leading to the synthesis of a polysaccharide intercellular adhesin. A chromosomally encoded reporter gene fusion between the ica promoter and the beta-galactosidase gene lacZ from Escherichia coli was constructed and used to investigate the influence of both environmental factors and subinhibitory concentrations of different antibiotics on ica expression in S. epidermidis. It was shown that S. epidermidis biofilm formation is induced by external stress (i.e., high temperature and osmolarity). Subinhibitory concentrations of tetracycline and the semisynthetic streptogramin antibiotic quinupristin-dalfopristin were found to enhance ica expression 9- to 11-fold, whereas penicillin, oxacillin, chloramphenicol, clindamycin, gentamicin, ofloxacin, vancomycin, and teicoplanin had no effect on ica expression. A weak (i.e., 2.5-fold) induction of ica expression was observed for subinhibitory concentrations of erythromycin. The results were confirmed by Northern blot analyses of ica transcription and quantitative analyses of biofilm formation in a colorimetric assay.

Adhesins, Bacterial↗
PubMed2000

Alternative transcription factor sigma(B) is involved in regulation of biofilm expression in a Staphylococcus aureus mucosal isolate.

Osmotic stress was found to induce biofilm formation in a Staphylococcus aureus mucosal isolate. Inactivation of a global regulator of the bacterial stress response, the alternative transcription factor sigma(B), resulted in a biofilm-negative phenotype and loss of salt-induced biofilm production. Complementation of the mutant strain with an expression plasmid encoding sigma(B) completely restored the wild-type phenotype. The combined data suggest a critical role of sigma(B) in S. aureus biofilm regulation under environmental stress conditions.

Bacterial Proteins↗
PubMed1999

Evolution of bacterial pathogenesis.

The evolution of bacteria is associated with continuous generation of novel genetic variants. The major driving forces in this process are point mutations, genetic rearrangements, and horizontal gene transfer. A large number of human and animal bacterial pathogens have evolved the capacity to produce virulence factors that are directly involved in infection and disease. Additionally, many bacteria express resistance traits against antibiotics. Both virulence factors and resistance determinants are subject to intrastrain genetic and phenotypic variation. They are often encoded on unstable DNA regions. Thus, they can be readily transferred to bacteria of the same species or even to non-related prokaryotes. This review article focuses on the main mechanisms of bacterial microevolution responsible for the rapid emergence of variants with novel virulence and resistance properties. In addition, processes of macroevolution are described with special emphasis on gene transfer and fixation of adaptive mutations in the genome of pathogens.

Animals↗
PubMed1999

A novel mechanism of phase variation of virulence in Staphylococcus epidermidis: evidence for control of the polysaccharide intercellular adhesin synthesis by alternating insertion and excision of the insertion sequence element IS256.

Biofilm formation of Staphylococcus epidermidis on smooth polymer surfaces has been shown to be mediated by the ica operon. Upon activation of this operon, a polysaccharide intercellular adhesin (PIA) is synthesized that supports bacterial cell-to-cell contacts and triggers the production of thick, multilayered biofilms. Thus, the ica gene cluster represents a genetic determinant that significantly contributes to the virulence of specific Staphylococcus epidermidis strains. PIA synthesis has been reported recently to undergo a phase variation process. In this study, biofilm-forming Staphylococcus epidermidis strains and their PIA-negative phase variants were analysed genetically to investigate the molecular mechanisms of phase variation. We have characterized biofilm-negative variants by Southern hybridization with ica-specific probes, polymerase chain reaction and nucleotide sequencing. The data obtained in these analyses suggested that in approximately 30% of the variants the missing biofilm formation was due to the inactivation of either the icaA or the icaC gene by the insertion of the insertion sequence element IS256. Furthermore, it was shown that the transposition of IS256 into the ica operon is a reversible process. After repeated passages of the PIA-negative insertional mutants, the biofilm-forming phenotype could be restored. Nucleotide sequence analyses of the revertants confirmed the complete excision of IS256, including the initially duplicated 8 bp target sites. These results elucidate, for the first time, a molecular mechanism mediating phase variation in staphylcocci, and they demonstrate that a naturally occurring insertion sequence element is actively involved in the modulation of expression of a Staphylococcus virulence factor.

Bacterial Adhesion↗
PubMed1999

[Adherence and invasion-two pathogenicity factors in bacterial and fungal pathogens].

Pathogenicity factors such as adhesins, toxins, capsules, and other microbial gene products are involved as causative agents for infectious diseases. Therefore, the pathogenicity of organisms is increasingly studied on a molecular level. In bacteriology, unspecific adherence mechanisms and receptor-specific adhesins have to be distinguished. An adhesin-mediated invasion of pathogenic organisms in eukaryotic host cells could be relevant for pathogenesis. In mycology, various specific adhesins are involved in colonization of the host. Aspartyl proteases and phospholipases are relevant for adherence and invasion of host structures by pathogenic yeasts. Resistance factors have a central function in the distribution of infectious organisms. Gene-transfer, point mutations and efflux mechanisms are involved in the development of antibiotic drug resistance. Antifungal drug resistance does occur predominantly in Candida albicans against azole drugs. As underlying mechanisms point mutations in the ERG11 gene, encoding for the target enzyme of azoles, as well as energy-dependent efflux mechanisms were identified. Whether these mycotic factors are specific virulence factors or "fitness-factors" for a better survival of these organisms in the host, and if a possible alternating effect exists between resistance and virulence mechanisms is currently under investigation.

Adhesins, Bacterial↗
PubMed1999

Detection of Staphylococcus aureus and Staphylococcus epidermidis in clinical samples by 16S rRNA-directed in situ hybridization.

Staphylococcus epidermidis and Staphylococcus aureus are the most common causes of medical device-associated infections, including septicemic loosenings of orthopedic implants. Frequently, the microbiological diagnosis of these infections remains ambiguous, since at least some staphylococci have the capacity to reduce their growth rate considerably. These strains exhibit a small-colony phenotype, and often they are not detectable by conventional microbiological techniques. Moreover, clinical isolates of S. aureus and S. epidermidis adhere to polymer and metal surfaces by the generation of thick, multilayered biofilms consisting of bacteria and extracellular polysaccharides. This study reports improved detection and identification of S. aureus and S. epidermidis by an in situ hybridization method with fluorescence-labeled oligonucleotide probes specific for staphylococcal 16S rRNA. The technique has proven to be suitable for the in situ detection of staphylococci, which is illustrated by the identification of S. epidermidis in a connective tissue sample obtained from a patient with septicemic loosening of a hip arthroplasty. We also show that this technique allows the detection of intracellularly persisting bacteria, including small-colony variants of S. aureus, and the differentiation of S. epidermidis from other clinically relevant staphylococci even when they are embedded in biofilms. These results suggest that the 16S rRNA in situ hybridization technique could represent a powerful diagnostic tool for the detection and differentiation of many other fastidious microorganisms.

Genes, rRNA↗
PubMed1998

Adherence of Staphylococcus aureus isolated in peritoneal dialysis-related exit-site infections to HEp-2 cells and silicone peritoneal catheter materials.

BACKGROUND: Peritoneal catheter exit-site infections cause a relevant morbidity in peritoneal dialysis patients and are frequently caused by Staphylococcus aureus. We tested the hypothesis that adherence of exit-site-derived S. aureus to epithelial cells and peritoneal catheter silicone tubes discriminates virulent and less virulent strains. METHODS: The binding of isolated S. aureus to an epithelial cell line (HEp-2) and to silicone tubes was analyzed using light-microscopy or radioactive labeling of bacteria. RESULTS: Of 378 exit-site swabs, 99 (26%) were positive for microbial growth. S. aureus was cultured in 25 of 99 positive swabs; three of 13 swabs taken in exit-site infections grade 3 and 4 that had tested positive for S. aureus. Adherence of S. aureus from exit-site infections grade 2, 3 and 4 to Hep-2 cells did not differ from adherence of bacteria isolated from asymptomatic or moderately inflamed catheter exit sites (grade 0-2). However, binding of S. aureus to silicone tubes was enhanced in grade 0/1 compared with grade 2-4 exit-site isolates. CONCLUSIONS: Staphylococcus aureus is an important pathogen in CAPD-related exit-site infection being isolated in about 6.6% of all exit-site swabs (and in 25% of all positive swabs). Silicone-adhesive strains may be of more clinical significance in peritoneal dialysis patients since adhesion to silicone was increased in S. aureus strains isolated in more severe exit-site infections.

Bacterial Adhesion↗
PubMed1998

Effects of subinhibitory concentrations of antibiotics on alpha-toxin (hla) gene expression of methicillin-sensitive and methicillin-resistant Staphylococcus aureus isolates.

Concentrations of antibiotics below the MIC are able to modulate the expression of virulence-associated genes. In this study, the influence of subinhibitory doses of 31 antibiotics on the expression of the gene encoding the staphylococcal alpha-toxin (hla), a major virulence factor of Staphylococcus aureus, was investigated with a novel gene fusion protocol. The most striking observation was a strong induction of hla expression by subinhibitory concentrations of beta-lactams and an almost complete inhibition of alpha-toxin expression by clindamycin. Whereas glycopeptide antibiotics had no effect, the macrolide erythromycin and several aminoglycosides reduced and fluoroquinolones slightly stimulated hla expression. Furthermore, Northern blot analysis of hla mRNA and Western blot (immunoblot) analysis of culture supernatants of both methicillin-sensitive and methicillin-resistant S. aureus strains revealed that methicillin-induced alpha-toxin expression is a common phenomenon of alpha-toxin-producing strains. Some methicillin-resistant S. aureus isolates produced up to 30-fold more alpha-toxin in the presence of 10 microg of methicillin per ml than in its absence. The results indicate that the novel gene fusion technique is a useful tool for studying the modulation of virulence gene expression by antibiotics. Moreover, the results suggest that the effects of certain antibiotics on virulence properties may be relevant for the management of S. aureus infections.

Anti-Bacterial Agents↗
PubMed1997

Detection of the intercellular adhesion gene cluster (ica) and phase variation in Staphylococcus epidermidis blood culture strains and mucosal isolates.

Staphylococcus epidermidis is a common cause of catheter-associated infections and septicemia in immunocompromised patients. To answer the question whether S. epidermidis skin isolates differ from isolates causing septicemic diseases, 51 strains obtained from blood cultures, 1 strain from shunt-associated meningitis, and 36 saprophytic isolates were characterized. The study demonstrates that most of the blood culture strains formed a multilayered biofilm on plastic material, whereas skin and mucosal isolates did not. Moreover, biofilm-producing strains were found to generate large bacterial autoaggregates in liquid culture. Autoaggregation and biofilm formation on polymer surfaces was associated with the presence of a DNA sequence encoding an intercellular adhesion gene cluster (ica) that mediates the production of a polysaccharide intercellular adhesin. The presence of the intercellular adhesion genes in blood culture isolates was also found to be correlated with the exhibition of black colonies on Congo red agar, whereas the adhesin-negative strains formed red colonies. Upon subcultivation on Congo red agar, the black colony forms of the blood culture strains exhibited red colony variants which were biofilm and autoaggregation negative and occurred at a frequency of 10(-5). The DNA analysis of these S. epidermidis variants by pulsed-field gel electrophoresis and Southern hybridization with an ica-specific gene probe revealed no detectable difference between the black and red colony types. Moreover, after repeated passage, the phenotype of the parent strain could be restored. Therefore, these colony forms were regarded as phase variants. This phenotypic change was observed exclusively in adhesin-positive clinical isolates and not in adhesin-negative saprophytic strains of S. epidermidis.

Bacteremia↗
PubMed1994

Lack of mecA transcription in slime-negative phase variants of methicillin-resistant Staphylococcus epidermidis.

Five phase variants (PV1 to PV5) of the well-characterized, slime-producing, methicillin-resistant, pathogenic strain of Staphylococcus epidermidis sensu strictu RP62A (ATCC 35984) were isolated by the Congo red agar method. In comparison with the parent strain, the phase variants showed a different colonial morphology on Congo red agar, a strongly reduced adherence capacity, and decreased levels of resistance to methicillin, oxacillin, and penicillin. All phase variants yielded biochemical reaction patterns and profiles in pulsed-field gel electrophoresis identical to those of parent strain RP62A, indicating a common origin. All phase variants proved to have the capacity to shift back to the original phenotype of parent strain RP62A. A search for the resistance mechanisms of strain RP62A revealed beta-lactamase production and the presence of mecA in PV1 to PV5 as well as parent strain RP62A. In Northern blots of total staphylococcal RNA, the phase variants showed no detectable mecA-specific transcription product, whereas parent strain RP62A revealed a strong signal, indicating that mecA transcription is not the mechanism responsible for the decreased methicillin resistance phenotype of phase variants PV1 to PV5.

Anti-Bacterial Agents↗