- Volume 48, Issue 11, 1999
Volume 48, Issue 11, 1999
- Editorial
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- Antimicrobial Resistance
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Expression of β-lactamases in Yersinia enterocolitica strains of biovars 2, 4 and 5
More LessCharacteristic patterns of susceptibility to β-lactam antibiotics are associated with different biovars of Yersinia enterocolitica. To elucidate the basis for these differences, the β-lactamases of strains of Y. enterocolitica biovars 4 (n = 63), 2 (n = 12) and 5 (n = 10) were characterised. PCR fragments were generated from the β-lactamase A (blaA) and B (blaB) genes; in addition, β-lactamase induction tests were performed with imipenem as the inducer and β-lactamase inhibition assays were undertaken with aztreonam and clavulanic acid. All the strains yielded PCR amplification fragments with primers to blaA and blaB. Biovar 4 strains had uniform patterns of β-lactamase induction and inhibition: uninduced biovar 4 strains predominantly expressed BlaA, but low-level expression of BlaB was also detected; after induction, biovar 4 strains predominantly produced BlaB. β-Lactamase expression varied between and within biovars 2 and 5: uninduced strains predominantly expressed either BlaA or BlaB, or exclusively BlaB; after induction BlaB was predominantly or exclusively expressed. Both the basal and induced levels of β-lactamase varied within biovars 2 and 5. Some biovar 5 strains were not inducible; these predominantly produced BlaA. The results of this study show that biovar 2, 4 and 5 strains contain both blaA and blaB, but that the expression of the enzymes is regulated differently between the biovars, and varies within biovars 2 and 5. There was some correlation between antibiogram and the clusters defined from the β-lactamase induction and inhibition tests, but it was not possible to predict β-lactamase expression profiles from MIC data.
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- Epidemiological Typing
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Analysis of Salmonella enterica serotype Typhimurium by phage typing, antimicrobial susceptibility and pulsed-field gel electrophoresis
More LessThree typing methods commonly used for bacteria - phage typing, antimicrobial susceptibility and pulsed-field gel electrophoresis (PFGE) - were used to characterise 64 Salmonella enterica serotype Typhimurium isolates from individual adult patients from Nairobi, Kenya. The isolates encompassed 11 definitive phage types (DTs), which fell into eight PFGE clusters; 31.3% of isolates were either untypable or reacted non-specifically with the phages used for typing and 26.6% were of DT 56. Plasmids of c. 100 kb were responsible for self-transferable multiresistance among the isolates. Analysis by PFGE and phage type demonstrated that multiresistant Typhimurium strains causing diarrhoea and invasive disease were multiclonal.
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- Model Of Infection
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Factors affecting the course and severity of transnasally induced Staphylococcus aureus pneumonia in mice
In order to examine several factors that may affect the course and severity of transnasally induced Staphylococcus aureus pneumonia in mice, bacteria were prepared in a free suspension or bound to fetal mouse cells. Immunosuppression was induced in five strains of mice (ICR, C57BL/6, BALB/c, C3H/He and CBA/J) by injection of cyclophosphamide (200 mg/kg body weight), 2 days before infection. Impairment of mucociliary clearance was induced by intranasal instillation of formalin. Mice were then infected with various doses and strains of the organism. Although no significant differences were observed between either form of inoculum, pretreatment with formalin plus cyclophosphamide was associated with a significant increase in lung bacterial counts. In particular, cyclophosphamide treatment was associated with a high mortality in mice infected with several strains of S. aureus irrespective of their toxin production profiles. Histopathological examination demonstrated that in mice treated with formalin plus cyclophosphamide, clusters of bacteria were observed in lung parenchyma, associated with a mild accumulation of inflammatory cells at day 2 and extensive cell infiltration at day 7. CBA/J mice represented the most susceptible strain among those examined, with 104- and 102-fold higher bacterial counts in the lungs at days 3 and 5, respectively. These results indicate that neutropenia and impaired mucociliary clearance are major factors that influence the severity of S. aureus pneumonia in mice. Analysis of the role of genetic background in enhancement of vulnerability to infection is warranted in future studies.
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- Correspondence
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- Bacterial Characterisation And Pathogenicity
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Expression in Escherichia coli of flaB, the gene coding for a periplasmic flagellin of Leptospira interrogans serovar pomona
More LessA periplasmic flagellin gene, flaB, of Leptospira interrogans serovar pomona (strain Pomona) was expressed in Escherichia coli for the production and antigenic characterisation of the protein. The flaB structural gene, which was previously cloned into pUC118, was derived by PCR from the recombinant plasmid and used to generate an expression construct with the trc promoter-driven pProEx HT system. Under the conditions employed, the flaB was expressed as inclusion bodies formed within E. coli, yielding c. 120 mg of the recombinant protein/L of culture. A polyhistidine tag introduced at the amino-acid terminus of the FlaB protein allowed for the purification of the protein by nickel-chelate affinity chromatography. The expressed protein reacted with both mouse and bovine antisera to L. interrogans on Western blots, indicating that it could be of use in the diagnosis of leptospirosis. The recombinant leptospiral flagellin may also be of value in studying its role in the pathogenesis of leptospirosis.
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Virulence properties of type VII Streptococcus agalactiae (group B streptococci) and immunochemical analysis of capsular type polysaccharide
Strains of a new polysaccharide type of group B streptococci (GBS), type VII, have been isolated from human carriers and invasive infections. Some of these strains bear the protein antigen c or R, as do other GBS serotypes. The capsular type polysaccharide is sialylated and this residue is involved in the immunodeterminant structure. All type VII strains examined were virulent in CD-1 mice; the LD50 after intraperitoneal (i.p.) challenge was 4.57 (SD 0.12) x 107 cfu for the reference strain and 5.49 (SD 1.5) x 107 cfu for clinical isolates. A particular feature of this serotype was the ability to induce septic arthritis not only when injected intravenously (i.v.), but also when injected i.p. Rabbit antiserum against the capsular type VII polysaccharide exhibited opsonic activity in a phagocytosis assay and protective activity against infection.
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Fine structural characterisation of a Rickettsia-like organism in human platelets from patients with symptoms of ehrlichiosis
More LessSince 1982, Ehrlichia platys infection has been diagnosed in canines from Venezuela by the use of buffy coat smears. In 1992, ehrlichia-like bodies were observed in platelets from a severely ill girl by light microscopy. The patient was seropositive to E. chaffeensis by the indirect fluorescent antibody test (IFAT). Tetracycline was administered and the patient recovered. More than 400 cases with such intra-platelet organisms have been studied at this laboratory over the past 6 years, and all the patients had a good response to the treatment. To determine whether the organisms in human blood platelets were truly platelet ehrlichiae, IFAT and transmission electron microscopy (TEM) studies were undertaken in four patients. Light microscopic examination of blood samples revealed the dense organism inside platelets, and a great reactivity of the blood cells. Sera from the four patients were seronegative against E. chaffeensis and E. platys antigens. Three of four samples contained the intra-platelet organisms when examined by TEM. Electron microscopy showed platelets with vacuoles containing pleomorphic organisms. These organisms had a thickened membrane, an electron-translucent inner area and an electron-dense granular component in the periphery. An abundant electron-dense material was observed surrounding them. The ultrastructure of such micro-organisms has not been reported previously. Based on the similarity of many of their characteristics with rickettsiae, we suggest that the microorganisms found in the present study might belong to the family Rickettsiaceae.
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Expression of Bacteroides fragilis virulence markers in vitro
Bacteroides fragilis isolates from intestinal and non-intestinal infections, normal flora and the environment were examined for properties linked with interactions among cells in vitro. Different adhesion molecules were detected in agglutination assays with human erythrocytes and tests for auto-agglutination and adherence to human colon carcinoma cells (HT29). There was no correlation between these properties, indicating that independent molecules are involved. Treatment with trypsin, heat or EDTA inhibited agglutination and adherence, suggesting that these molecules are proteins. The lack of correlation with the origin of the strains did not permit any of these activities to be recognised as virulence markers. The expression of fragilysin, a protease associated with damage to intestinal cells and bacterial translocation, was examined. Only those strains from patients with diarrhoea expressed this protease activity in assays with HT29 cells and this was confirmed by specific PCR for the bft gene. The activity of fragilysin as an enterotoxin was confirmed in the rabbit intestinal ligated loop assay. The association of this property only with strains from intestinal infections indicates that it is too early to suggest this protease as a determinant factor of B. fragilis invasiveness.
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- Immunodiagnosis
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Oral fluid antibody detection in the diagnosis of Helicobacter pylori infection
More LessThe aim of this study was to evaluate an enzyme-linked immunosorbent assay (ELISA) for the detection of anti-Helicobacter pylori specific IgG antibodies in specimens of oral fluid. Antral biopsy specimens, serum and oral fluid samples were collected from 81 patients attending for upper gastrointestinal endoscopy. The presence or absence of current H. pylori infection was determined by culture, histology and urease detection. Anti-H. pylori specific IgG was detected in serum by an established in-house ELISA and in oral fluid by an ELISA developed for this study. In all, 34 (42%) of 81 patients were positive for H. pylori by one or more of the ‘gold standard’ tests (culture, histology and urease detection). The oral fluid ELISA had a sensitivity of 94% and specificity of 85% with regard to current H. pylori infection. The serum ELISA had a sensitivity and specificity of 91%. There was an overall agreement of 88% between serum and oral fluid antibody detection. The detection of anti-H. pylori specific IgG in oral fluid by ELISA is comparable in sensitivity and specificity with serum-based methods. Oral fluid-based ELISA could provide a reliable, non-invasive method for the diagnosis of H. pylori infection, and may be of particular benefit for population surveys.
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- Molecular Diagnosis
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Use of PCR to identify enteroaggregative Escherichia coli as an important cause of acute diarrhoea among children living in Calcutta, India
More LessThe importance of enteroaggregative Escherichia coli (EAggEC) as a possible aetiological agent of acute diarrhoea among children in Calcutta, India, was investigated. Simultaneously the use of a previously described PCR diagnostic system was assessed for its ability to identify EAggEC infection. E. coli strains isolated during a 1-year case-control study from faecal samples of 388 children aged <5 years, with or without diarrhoea, were examined for EAggEC by HeLa cell adherence assay in parallel with a PCR assay with primers generated from an EAggEC DNA probe. A blind comparison was made between the two methods to determine their diagnostic potential. E. coli isolates that adhered to HeLa cells in an aggregative pattern were the sole isolates significantly more often in 254 cases (9%) than in 134 control (2%) children. Age stratification showed that EAggEC were isolated more frequently from children aged <36 months. The EAggEC isolates belonged to several O serogroups and showed multiple drug resistance. Both methods were positive for 26 samples, nine samples were positive by PCR alone and seven samples were positive by culture alone, thus indicating a 78% sensitivity and 97% specificity for the PCR assay. EAggEC is an important aetiological agent of acute diarrhoea among infants in and around Calcutta, and the PCR diagnostic system may be useful to identify such infection in epidemiological studies.
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Direct detection of Prevotella intermedia and P. nigrescens in suppurative oral infection by amplification of 16S rRNA gene
More LessA specific 16S rDNA PCR and subsequent hybridisation reaction was designed to discriminate between strains of Prevotella intermedia (n = 15) and P. nigrescens (n = 15). This technique was then used to detect the presence of these two bacterial species in acute suppurative oral infection. A total of 36 pus samples aspirated from 26 peri-apical abscesses, three root canals, three periodontal abscesses, two cases of refractory periodontitis, one cyst and one haematoma was examined. A portion of the pus sample was processed by PCR and the remainder of the specimen was subjected to routine culture. The PCR-based technique gave an identical pattern of detection of P. intermedia or P. nigrescens to that obtained by culture for 30 of the 36 specimens. Either P. intermedia or P. nigrescens was present in 14 samples and neither species was detected in 16 samples. In the remaining six samples the PCR method indicated the presence of one (n = 3) or both (n = 3) of the Prevotella species but neither or only one species was isolated by culture. It is concluded that the presence of P. intermedia and P. nigrescens in pus can be detected rapidly and specifically by direct PCR amplification of 16S rDNA. P. nigrescens was detected more frequently than P. intermedia in suppurative peri-apical infection both by culture and PCR.
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- Virology
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Comparison of three PCR techniques for detecting cytomegalovirus (CMV) DNA in serum, detection of early antigen fluorescent foci and culture for the diagnosis of CMV infection
More LessThree PCR assays were developed for detection of cytomegalovirus (CMV) DNA in serum and were evaluated with samples from organ transplant recipients. The Qiamp Blood Kit was efficient for extraction of DNA from sera. Single-round PCR of a 293-bp region of CMV DNA was sensitive and highly specific for CMV targets and was more sensitive than detection of early antigen fluorescent foci (DEAFF) testing or isolation of CMV from buffy coat by cell culture. The results of a significant proportion of buffy coat samples were not interpretable because of toxicity in conventional culture or DEAFF tests. A non-competitve quantitative PCR test and semi-quantitative PCR test for the detection of CMV DNA in serum yielded comparable results for samples taken serially from three bone marrow transplant recipients. Single-round PCR was superior to conventional techniques for the diagnosis of CMV infection, was simple to perform and was completed rapidly. The semi-quantitative technique has added advantages where quantification is important.
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- Book Reviews
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Volumes and issues
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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 70 (2021)
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Volume 69 (2020)
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Volume 68 (2019)
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Volume 67 (2018)
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Volume 66 (2017)
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Volume 65 (2016)
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Volume 64 (2015)
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Volume 63 (2014)
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Volume 62 (2013)
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Volume 61 (2012)
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Volume 60 (2011)
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Volume 59 (2010)
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Volume 58 (2009)
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Volume 57 (2008)
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Volume 56 (2007)
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Volume 55 (2006)
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Volume 54 (2005)
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Volume 53 (2004)
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Volume 52 (2003)
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Volume 51 (2002)
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Volume 50 (2001)
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Volume 49 (2000)
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Volume 48 (1999)
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Volume 47 (1998)
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Volume 46 (1997)
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Volume 45 (1996)
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Volume 44 (1996)
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Volume 43 (1995)
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Volume 42 (1995)
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Volume 41 (1994)
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Volume 40 (1994)
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Volume 39 (1993)
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Volume 38 (1993)
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Volume 37 (1992)
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Volume 36 (1992)
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Volume 35 (1991)
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Volume 34 (1991)
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Volume 33 (1990)
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Volume 32 (1990)
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Volume 31 (1990)
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Volume 30 (1989)
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Volume 29 (1989)
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Volume 28 (1989)
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Volume 27 (1988)
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Volume 26 (1988)
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Volume 25 (1988)
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Volume 24 (1987)
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Volume 23 (1987)
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Volume 22 (1986)
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Volume 21 (1986)
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Volume 20 (1985)
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Volume 19 (1985)
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Volume 18 (1984)
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Volume 17 (1984)
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Volume 16 (1983)
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Volume 15 (1982)
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Volume 14 (1981)
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Volume 13 (1980)
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Volume 12 (1979)
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Volume 11 (1978)
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Volume 10 (1977)
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Volume 9 (1976)
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Volume 8 (1975)
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Volume 7 (1974)
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Volume 6 (1973)
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Volume 5 (1972)
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Volume 4 (1971)
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Volume 3 (1970)
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Volume 2 (1969)
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Volume 1 (1968)