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Volume 49,
Issue 5,
2000
Volume 49, Issue 5, 2000
- Editorial
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- Review Article
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Strain, clone and species: comments on three basic concepts of bacteriology
More LessDifferent aspects of the terms strain, clone and species are discussed. The term strain is commonly used to denote a pure culture – here called ‘the strain in the taxonomic sense’ – but does also refer to a natural concept closely related to the clone. The term clone on the other hand is used both in a general and in a more restricted sense, the latter indicating a low degree of genetic exchange. The important distinction between the definition of a species and the criteria for a species is emphasised and the main kinds of criteria are considered.
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- Diagnostic Microbiology
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Evaluation of a PCR-immunoassay technique for detection of Neisseria meningitidis in cerebrospinal fluid and peripheral blood
More LessA multiplex PCR-immunoassay for the rapid diagnosis of bacterial meningitis from cerebrospinal fluid (CSF) or peripheral blood was compared with conventional microbiological techniques used in the routine diagnostic laboratory. The multiplex PCR was designed to detect simultaneously a universal conserved sequence coding for bacterial 16S rRNA and the Neisseria meningitidis porA gene. The PCR-immunoassay had a detection limit of (3–5) × 102 cfu/ml (equivalent to 1–3 cfu/PCR) with spiked CSF or blood samples, compared with (3–5) × 103 cfu/ml for PCR followed by conventional agarose gel electrophoresis for detection of PCR products. Of 294 CSF samples from patients suspected on clinical grounds of suffering from meningitis, the PCR-immunoassay detected bacterial DNA in 29 CSF samples, 15 of which were also positive for N. meningitidis DNA. The 29 positive CSF samples comprised 25 samples that were also reported positive and four that were reported negative by conventional methodology; the latter four were all positive for N. meningitidis by the PCR-immunoassay. Of 173 peripheral blood samples examined, the PCR-immunoassay detected bacterial DNA in 18 samples, 14 of which were also positive for N. meningitidis DNA. In comparison, only 10 samples were reported positive for N. meningitidis by conventional methodology. All negative PCR-immunoassay results correlated with those obtained by conventional methodology for both CSF and blood samples. The sensitivity and speed of the PCR-immunoassay system indicated that it could be used as a routine diagnostic test for meningococcal meningitis, enabling a diagnosis to be made within 4 h of receipt of the specimen.
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Detection of Chlamydia trachomatis-specific antibodies in human sera by recombinant major outer-membrane protein polyantigens
More LessThis study was performed to generate and evaluate recombinant antigens for use in a species-specific Chlamydia trachomatis immunoassay. In a molecular genetic approach, fragments of the C. trachomatis major outer-membrane protein (MOMP) were produced as fusion proteins to create three different constructs encompassing the variable domains I, II and IV of selected C. trachomatis serovars. The recombinant MOMP polyantigens were affinity-purified and used in an enzyme-linked immunosorbent assay. Antibody detection was evaluated with 103 patient sera and the results were compared with titres obtained in the micro-immunofluorescence test. The results showed that the generated MOMP polyantigens detected the presence of C. trachomatis-specific human antibodies with little cross-reaction to C. pneumoniae-specific antibodies. When compared to the micro-immunofluorescence assay the MOMP polyantigen detected the presence of anti-C. trachomatis IgG antibodies with a sensitivity of 80% and a specificity of 91%.
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- Antimicrobial Resistance
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β-Lactamase expression in Yersinia enterocolitica biovars 1A, 1B and 3
More LessCharacteristic patterns of β-lactam susceptibility are associated with different biovars of Yersinia enterocolitica. In a previous study differences in β-lactam susceptibility among biovar 2, 4 and 5 strains were largely attributed to differences in expression of β-lactamase A (BlaA) and β-lactamase B (BlaB). The basis for differences in β-lactam susceptibility of strains of biovars 1A, 1B and 3 is now considered. All the strains examined had blaB; nine of 31 biovar 3 strains and two of 13 biovar 1B strains had blaA, but PCR did not amplify blaA from biovar 1A strains. Nevertheless, inhibition data indicated that the majority of uninduced biovar 1A strains expressed BlaA and BlaB in similar amounts. Strong inducibility was seen in all these strains. Biovar 1B strains (which were less inducible than strains of biovar 1A) predominantly produced BlaA without induction; ticarcillin-sensitive strains of biovar 3 produced only BlaB but were not inducible; without induction biovar 3 strains resistant to ticarcillin and amoxycillin/clavulanate produced either predominantly BlaA, predominantly BlaB or exclusively BlaB and induction was demonstrated except for strains producing BlaB alone; biovar 3 strains resistant to ticarcillin but sensitive to amoxycillin/clavulanate predominantly produced BlaA without induction and were inducible for β-lactamase activity. After induction, nearly all strains predominantly or exclusively produced BlaB. Although PCR amplification fragments with primers specific for blaA were obtained only from some strains, the induction and inhibition data suggest that all Y. enterocolitica strains possess enzymes related to BlaA- as well as BlaB. Nevertheless, expression of the β-lactamase is regulated differently in different biovars and varies within most biovars. Failure to predict β-lactamase expression profiles from MIC data indicates the presence of additional mechanisms contributing to differences in susceptibility.
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Altered expression of oligopeptide-binding protein (OppA) and aminoglycoside resistance in laboratory and clinical Escherichia coli strains
More LessOligopeptide-binding protein (OppA) is the periplasmic component of the major oligopeptide transport system of enteric bacteria. Genetic and biochemical evidence suggests that OppA plays a role in the uptake of aminoglycoside antibiotics in Escherichia coli K-12. Forty-six (82%) of 56 aminoglycoside-resistant mutants of E. coli K-12 selected in vitro had reduced or undetectable OppA levels, as compared with their parent strain. Moreover, nine (36%) of 25 aminoglycoside-resistant clinical isolates of E. coli expressed reduced or undetectable levels of OppA. No decrease in OppA expression was observed among aminoglycoside-sensitive E. coli strains from patients. Twenty-three (42%) of 56 aminoglycoside-resistant mutants of E. coli K-12 and six (24%) of 25 clinical isolates also were deficient for expression of ornithine or arginine decarboxylases, or both, and these deficiencies might negatively affect OppA expression by reducing polyamine synthesis. These results support the view that reduced OppA expression is associated with aminoglycoside resistance in E. coli strains.
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Primary and combined resistance to four antimicrobial agents in Helicobacter pylori in Sofia, Bulgaria
More LessThe aim of this study was to evaluate the primary and combined resistance of Helicobacter pylori against four antimicrobial agents by a screening agar method (SAM) and a modified disk diffusion method (MDDM) alone and in combination. Pre-treatment H. pylori isolates from 192 consecutive H. pylori-positive patients at three hospitals in Sofia were investigated. MDDM was performed with disks containing metronidazole (5 μg), clarithromycin (15 μg) or erythromycin (15 μg), ciprofloxacin (5 μg) and tetracycline (30 μg). Resistance was determined by an inhibitory zone of <16 mm for metronidazole and ≤30 mm for other agents tested. The cut-off concentrations used to define resistance by SAM were: metronidazole >8 mg/L, clarithromycin >2 mg/L, tetracycline >4 mg/L and ciprofloxacin >1 mg/L. Primary resistance rates in H. pylori were: metronidazole 28.6%, clarithromycin 9.7%, metronidazole+clarithromycin 2.8%, ciprofloxacin 3.9%, metronidazole+ciprofloxacin 2.3%, tetracycline 1.9% and metronidazole+tetracycline 1.2%. Among metronidazole-resistant isolates, combined resistance to clarithromycin, ciprofloxacin and tetracycline was present in 11.4% (5 of 44 strains), 8.3% (3 of 36) and 4.9% (2 of 41), respectively. Two strains exhibited triple resistance to macrolides, metronidazole and either ciprofloxacin or tetracycline. Three tetracycline-resistant strains were detected in 1999; however, resistance rates to other agents were relatively stable during the 6 years. Primary H. pylori resistance to metronidazole is moderate and resistance to clarithromycin and to ciprofloxacin is considerable in comparison with results in most other countries. The alarming appearance of strains harbouring combined resistance or multiresistance provides the motivation for continued surveillance of H. pylori at global, national and regional levels.
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Characterisation of methicillin-resistant Staphylococcus aureus from Kuwait hospitals with high-level fusidic acid resistance
E.E. UDO and L.E. JACOBForty-seven fusidic acid- and methicillin-resistant Staphylococcus aureus isolates from clinical samples in four hospitals in Kuwait were studied for their relatedness by biotyping and pulsed-field gel electrophoresis (PFGE) and for the genetic location of their resistance determinants. Forty-four isolates were resistant to gentamicin, kanamycin and neomycin. Forty-one isolates were resistant to erythromycin and trimethoprim, 10 were resistant to chloramphenicol and four were resistant to ciprofloxacin. They contained two or three plasmids of c. 28, 2.8 and 1.8 kb. Genetic studies demonstrated that resistance to cadmium, propamidine isethionate and ethidium bromide were linked and were carried on the c. 28-kb plasmid. Chloramphenicol resistance was encoded by the 2.8-kb plasmid in resistant isolates. No resistance was associated with the 1.8-kb plasmid and this was considered to be a cryptic plasmid. Resistance to fusidic acid, methicillin, benzylpenicillin, gentamicin, kanamycin, neomycin, tetracycline, trimethoprim, erythromycin and ciprofloxacin were located on the chromosome. All the isolates produced urease, but varied in the production of haemolysins, pigments, lipase and lecithinase and were classified into nine biotypes. In contrast, PFGE divided the isolates into two major patterns with one PFGE type constituting the majority of isolates in all four hospitals. The presence of the dominant PFGE pattern in all four hospitals suggests that it is an epidemic MRSA clone with the capacity to spread. Infection control measures should be directed towards restricting the further spread of this clone.
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Prevalence of the carbapenemase gene (cfiA) among clinical and normal flora isolates of Bacteroides species in Hungary
More LessThe carbapenemase gene (cfiA) was detected in 4 (5.7%) of 70 clinical isolates of Bacteroides fragilis from different parts of Hungary. Among 24 other Bacteroides species isolated from infectious processes or from normal faecal flora, none was cfiA-positive. The MIC of imipenem and meropenem for all cfiA-positive B. fragilis isolates was ≤0.25 mg/L, but 17% of the B. fragilis and 46% of the non-fragilis Bacteroides isolates exhibited reduced susceptibility to imipenem (MICs 0.5–2 mg/L). Only one of these isolates produced increased levels of β-lactamase. No difference was observed in the outer-membrane proteins of B. fragilis isolates that harboured the cfiA gene and those with reduced susceptibility to imipenem.
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- Models Of Infection
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Translocation of indigenous microflora in an experimental model of sepsis
More LessTranslocation of viable bacteria from gut to bloodstream and other sterile body sites during shock has been demonstrated in several experimental and clinical studies. The factors causing translocation and its incidence at different stages of shock are not known. The aim of the study was to evaluate the importance of several factors causing translocation of indigenous microflora in an experimental model of septic shock based on intraperitoneal Escherichia coli sepsis in rats. Counts of inoculated E. coli and translocated bacteria in different locations, gut morphology and haematological values were evaluated at different stages of sepsis. Sepsis developed in all animals and E. coli achieved the highest counts in blood 6 h after inoculation. Translocation was commonest at 6 and 12 h after inoculation. Frequently translocating bacteria were lactobacilli, bifidobacteria, bacteroides and peptostreptococci. In early sepsis, translocation was associated with high E. coli counts in blood, yet in late sepsis the opposite correlation was present. Low infiltration by neutrophils in the ileum and decreased mitotic activity in the colon were associated with a high translocation rate. In early sepsis, translocation was associated with low lymphocyte counts, but in late sepsis, with low neutrophil counts. Translocation of bacteria (including anaerobes) that colonise the gut in high counts takes place during sepsis. Putative influencing factors such as activity of the primary disease (bacterial counts in blood), gut morphology or haematological values seem to have different impacts on translocation, depending on the stage of the disease.
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- Bacterial Pathogenicity
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Molecular characterisation of group A streptococci from invasive and non-invasive disease episodes in Belgium during 1993–1994
More LessFive hundred clinical group A streptococcal (GAS) isolates were collected in Belgium during the period 1 Nov. 1993 to 31 Oct. 1994. Clinical and laboratory data were recorded and isolates were characterised. The presence of the genes encoding streptococcal pyrogenic exotoxin types A (speA), B (speB), C (speC), F (speF) and streptococcal superantigen (ssa) were determined by PCR to target specific sequences. These isolates were also emm-typed and analysed by pulsed-field gel electrophoresis (PFGE) of genomic macrorestriction fragments with the enzyme SmaI. In total, 136 unrelated GAS PFGE types were identified and genetic diversity was clearly demonstrated. Two GAS PFGE types predominated; a first PFGE type comprised 66 (13.2%) emm1 isolates characterised by speA +, speB +, speC −, speF + and ssa −; the second PFGE type comprised 44 (8.8%) emm12 isolates characterised by speA −, speB +, speC + (or speC −), speF + and ssa −. Indistinguishable PFGE types were observed among both invasive and non-invasive isolates. Ten different PFGE types were found among 11 streptococcal toxic shock syndrome (STSS) isolates, and five of these lacked speA. Twenty-five (34.7%) of 72 invasive isolates gave negative results for speA, speC and ssa. This retrospective study confirmed the observation that the dissemination of one specific clone cannot be associated with invasive GAS disease and posed a question regarding the role of SPE A as a major virulence factor. Other streptococcal virulence factors in conjunction with host factors may determine the outcome of invasive GAS infection.
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Characterisation of cytolethal distending toxin (CDT) mutants of Campylobacter jejuni
More LessIn order to assess the contribution of cytolethal distending toxin (CDT) to the toxigenicity and pathogenicity of Campylobacter jejuni, the C. jejuni 81-176 and C. jejuni NCTC 11168 CDTs were inactivated by insertional mutation of the cdtB toxin subunit. Cell-free sonicates from isogenic C. jejuni 81-176 cdtB − strains were found to be greatly attenuated in HeLa cytotoxicity assays, whilst still retaining some toxigenicity. Sonicates from a C. jejuni NCTC 11168 cdtB − strain produced no detectable cytotoxicity. When orally administered to adult severe combined immunodeficient (SCID) mice, C. jejuni cdtB mutant strains were unaffected in enteric colonisation abilities but demonstrated impaired invasiveness into blood, spleen and liver tissues. These data suggest that CDT may be the principal toxin produced by this species and that some C. jejuni strains may generate additional toxigenic factor(s) distinct from CDT.
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The role of SEF14 and SEF17 fimbriae in the adherence of Salmonella enterica serotype Enteritidis to inanimate surfaces
More LessTo gain an understanding of the role of fimbriae and flagella in the adherence of Salmonella enterica serotype Enteritidis to inanimate surfaces, the extent of adherence of viable wild-type strains to a polystyrene microtitration plate was determined by a crystal violet staining assay. Elaboration of surface antigens by adherent bacteria was assayed by fimbriae- and flagella-specific ELISAs. Wild-type Enteritidis strains adhered well at 37°C and 25°C when grown in microtitration wells in Colonisation Factor Antigen broth, but not in other media tested. At 37°C, adherent bacteria elaborated copious quantities of SEF14 fimbrial antigen, whereas at 25°C adherent bacteria elaborated copious quantities of SEF17 fimbrial antigen. Non-fimbriate and non-flagellate knock-out mutant strains were also assessed in the adherence assay. Mutant strains unable to elaborate SEF14 and SEF17 fimbriae adhered poorly at 37°C and 25°C, respectively, but adherence was not abolished. Non-motile mutant strains showed reduced adherence whilst type-1, PEF and LPF fimbriae appeared not to contribute to adherence in this assay. These data indicate that SEF17 and SEF14 fimbriae mediate bacterial cell aggregation on inanimate surfaces under appropriate growth conditions.
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- Prevention Of Infection
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Use of scanning electron microscopy to investigate the prophylactic efficacy of rifampin-impregnated CSF shunt catheters
More LessInfection continues to be one of the major complications of cerebrospinal fluid (CSF) shunting procedures, and is caused mainly by skin-derived bacteria. Production of an extracellular biofilm plays an important role in the pathogenesis of shunt-associated infections by protecting bacteria from immune mechanisms and antibiotics. So far, removal of the original shunt and implantation of a new shunting device has been the only successful treatment for most patients. As an alternative strategy to prevent CSF infections, a rifampin-impregnated silicone catheter was designed to provide high initial and long-lasting (>60 days) release of bactericidal drug. To investigate the pathophysiological mechanism of its function, this new device was investigated both in vitro and in a rodent model of CSF infection by scanning electron microscopy (SEM) and bacterial culture. Staphylococcus epidermidis (108 cfu/ml) and S. aureus (104 cfu/ml) served as test strains. SEM demonstrated that, in contrast to the unloaded catheters, initial bacterial adherence on the catheter surface could be reduced to a few single cells, which did not show visible signs of proliferation. Bacterial cultures obtained simultaneously were all sterile, showing that adherent bacteria were killed immediately by the rifampin released from the catheter. Although rifampin incorporation into silicone polymers was not able to prevent initial bacterial adhesion completely, subsequent colonisation could be prevented.
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Volumes and issues
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Volume 74 (2025)
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Volume 73 (2024)
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Volume 72 (2023 - 2024)
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Volume 71 (2022)
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Volume 69 (2020)
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Volume 68 (2019)
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