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Biomedical subjects

S G Amyes

Publications and source records attributed to S G Amyes.

At least 55 records · Page 3Linked to original sources
PubMed1995

In-vitro and in-vivo selection of Staphylococcus aureus mutants resistant to ciprofloxacin.

Staphylococcus aureus mutants resistant to ciprofloxacin were selected both in vitro and in vivo. In-vitro selection was achieved by incubating two strains of S. aureus (MICs of ciprofloxacin of 0.5 and 4 mg/L respectively) in the presence of ciprofloxacin in concentrations equivalent to 1/2 x MIC for 24 h and isolating the mutants on agar containing the quinolone at 1, 2 or 5 x MIC. Stably-resistant mutants of both strains were isolated, although the frequency of mutation of the susceptible strain was higher than that of the resistant strain. A murine subcutaneous abscess model was used for in-vivo selection. Mice which had been infected with a ciprofloxacin-susceptible strain of S. aureus were treated for 24 h with ciprofloxacin in a dosage which yielded concentrations in the abscess cavity equivalent to 1/2 or 1 x MIC for the pathogen. Additional groups of infected mice received ciprofloxacin for varying periods in a dosage which produced concentrations at the site of infection equivalent to 1/2 x MIC. Stably-resistant mutants were isolated from the abscesses, the number of mice from which mutants were isolated and the mutational frequency being inversely proportional to the dosage of ciprofloxacin administered and the duration of treatment. The results of this study confirm that, in the treatment of patients with infections caused by S. aureus, the dosage of ciprofloxacin should be adequate to ensure inhibitory concentrations at the site of infection.

Abscess↗
PubMed1995

Prevalence of trimethoprim resistant dihydrofolate reductase genes identified with oligonucleotide probes in plasmids from isolates of commensal faecal flora.

In a recent survey of trimethoprim resistance, 357 Gram-negative aerobic organisms were isolated from healthy volunteers from rural and urban populations in South Africa. Trimethoprim resistance was transferable in 184 (52%) of the isolates. A further 12 (3%) transferred in the presence of an X+ actor. The transconjugants were probed with intragenic oligonucleotide probes for the type Ia, Ib, IIIa, VIII, V, VI, VII, IX, X and XII dihydrofolate reductase genes. Contrary to all previous data, the most prevalent dihydrofolate reductase gene was the type Ib (30%) followed by the type VIII (23%), V (13%), Ia (6%), VII (3%), and XII (0.5%). None of the strains hybridised to the type IIIa, XI, X and the VI dihydrofolate reductase probes. Plasmid restriction profiles revealed that the high prevalence of the type Ib and VIII dihydrofolate reductase genes resulted from the presence of ubiquitous plasmids. These results highlight the previous problems associated with the distinction of closely related dihydrofolate reductase genes.

Base Sequence↗
PubMed1995

Back mutations to the TEM-1 beta-lactamase from TRC-1 lead to restored sensitivity to clavulanic acid.

Back mutations from the TRC-1 beta-lactamase to the TEM-1 enzyme were selected in vitro. The revertant beta-lactamase was obtained from Escherichia coli strain J62.2 carrying plasmid pUK901 which encodes the TRC-1 beta-lactamase. The revertant was obtained after repeated subculture of E. coli J62.2 (pUK901) in amoxycillin 512 mg/L for 5 days. The revertant beta-lactamase had the same pI as TEM-1 (5.4) and had restored inhibition by clavulanic acid (ID50 reduced from 4.2 microM to 0.15 microM). The prevalence of these beta-lactamases in the clinical population may be the result of a two-way flux, with mutations in both forward and backward directions.

Amoxicillin↗
PubMed1995

A piperacillin-tazobactam resistant Escherichia coli strain isolated from a faecal sample of a healthy volunteer.

As part of a surveillance programme of the prevalence of antibiotic resistance, the faecal bacteria of healthy people (n = 1348) were examined, and the antibiotic resistance of the Escherichia coli strains determined. One strain out of 142 amoxycillin-resistant isolates, E. coli strain 1662, was also resistant to piperacillin-tazobactam but susceptible to amoxycillin-clavulanic acid. The piperacillin-tazobactam resistance determinant was transferable to standard E. coli strains by conjugation. However, the strain produced a beta-lactamase with several characteristics very similar to those of the TEM-1 beta-lactamase, i.e. pI of 5.4, an M(r) value of 22,000 and a comparable substrate profile. The enzyme was as efficiently inhibited by clavulanic acid and tazobactam as the TEM-1 and TEM-2 beta-lactamases but more than the amoxycillin-clavulanic acid-resistant TRC-1 enzyme. The transferable resistance to piperacillin-tazobactam appears to be mediated by a novel resistance mechanism that has previously not been described.

Anti-Bacterial Agents↗
PubMed1994

Three beta-lactamases isolated from Aeromonas salmonicida, including a carbapenemase not detectable by conventional methods.

The beta-lactamases of seven strains of Aeromonas salmonicida subsp. achromogenes resistant to amoxicillin (MIC > 1024 mg/l) and responsible for furunculosis in farmed Atlantic salmon in Scotland were examined to establish the mechanisms of beta-lactam resistance. Separation of a cell-free extract on an isoelectric focusing gel stained with the chromogenic cephalosporin nitrocefin showed the presence of two beta-lactamases, one with a pI of 7.9 and the other with a pI of 6.0. Hydrolysis assays of cell-free extracts of these strains demonstrated carbapenemase, penicillinase and cephalosporinase activity. However, when the beta-lactamases were separated by anion exchange chromatography, the carbapenemase activity could not be retrieved in either of the peak fractions containing the separated enzymes that had been visualised by nitrocefin. Consequently, a novel carbapenemase was discovered which cannot be detected with nitrocefin.

Aeromonas↗
PubMed1994

Beta-lactam resistance in aerobic faecal flora from general practice patients in the UK.

One hundred faecal specimens submitted to a diagnostic laboratory in Edinburgh and found to be negative for gastrointestinal pathogens were examined for the presence of antibiotic-resistant bacteria. The results were compared with findings in the healthy population in the same area. The highest incidence of resistance was observed to cefuroxime (65%) and ampicillin (60%). Of the ampicillin-resistant isolates, 62% could transfer their resistance determinants to a standard Escherichia coli host strain. In 100% of these transconjugants ampicillin resistance was shown to result from the presence of the TEM-1 beta-lactamase which was identified in a heterogeneity of plasmid profiles. These plasmids commonly mediated resistance to streptomycin and tetracycline in addition to ampicillin.

Ampicillin Resistance↗
PubMed1994

Trimethoprim resistant dihydrofolate reductases in normal faecal flora isolated in India.

A high incidence of resistance to trimethoprim has been shown in the normal faecal flora in a population in south India. The dihydrofolate reductase (dhfr) genes mediating transferable resistance to trimethoprim have been identified. Unusually, in this study, the dhfrV was shown to be the predominant resistance gene (dhfrV 50% of transconjugants, dhfrIa 30%), the dhfrIb was also detected being distinguished from the dhfrV by an oligo-probe. However, when non-transferable resistance was considered, the dhfrIa was the most prevalent of the dhfrs identified. All those plasmids harbouring the dhfrIa were shown to possess Tn7. All the plasmids that probed positive for the dhfrV and the dhfrIb were shown to be associated with the integrase of the Tn21-like transposons, but 8 of the dhfrV genes were not associated with the Tn21 resolvase. The dhfrIV was shown to be present in all seven plasmids that produced low level trimethoprim-resistance. The dhfrV, first characterized in Sri Lanka, would seem to have a local distribution in this region of Asia but is distinguishable from the dhfrIb only by the use of an oligo-probe.

Enterobacteriaceae↗
PubMed1994

A multicentre study of the in-vitro activity of cefotaxime, cefuroxime, ceftazidime, ofloxacin and ciprofloxacin against blood and urinary pathogens.

The in-vitro susceptibilities of aerobic bacteria isolated from 1804 blood and 4529 urine specimens collected at nine hospitals in the UK were examined. An agar dilution method was used to determine the MICs of each isolate to three cephalosporins, cefotaxime, cefuroxime and ceftazidime, and to two fluoroquinolones, ofloxacin and ciprofloxacin. Sensitivities were then calculated using British Society for Antimicrobial Chemotherapy recommended breakpoints. Of the cephalosporins tested cefotaxime was the most active against the Enterobacteriaceae. All the systemic staphylococcus isolates collected were sensitive to both cefotaxime and cefuroxime. As expected, ceftazidime was the only cephalosporin active against the Pseudomonas isolates. Both quinolones were highly active against the Enterobacteriaceae and Pseudomonas spp. They also demonstrated good Gram-positive activity, particularly against Staphylococcus aureus and Enterococcus spp.

Anti-Infective Agents↗
PubMed1994

Biochemical properties of inducible beta-lactamases produced from Xanthomonas maltophilia.

Four different beta-lactamases have been found in several strains of Xanthomonas maltophilia isolated from blood cultures during 1984 to 1991 at the Edinburgh Royal Infirmary. One was a metallo-beta-lactamase with predominantly penicillinase activity and an isoelectric point of 6.8. Its molecular size as determined by gel filtration was 96 kDa but was only 26 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), suggesting a tetramer of four equal subunits. The enzyme hydrolyzed all classes of beta-lactams except the monobactam aztreonam. This enzyme was not inhibited by potassium clavulanate or BRL 42715 but was inhibited by p-chloromercuribenzoate, mercuric chloride, and EDTA. The beta-lactamase was unstable in 50 mM sodium phosphate buffer (pH 8.0) but stable in 50 mM Tris HCl (pH 8.0). The other beta-lactamases focused as a series of different isoelectric points, ranging from pI 5.2 to 6.6. Together, these enzymes exhibited a broad spectrum of activity, hydrolyzing most classes of beta-lactams but not imipenem or aztreonam. Their molecular size was 48 kDa by Sephadex gel filtration and 24 kDa by SDS-PAGE, indicating that they were enzymes consisting of two equal subunits. They were inhibited by p-chloromercuribenzoate, mercuric chloride, potassium clavulanate, and BRL 42715 but not EDTA. This study demonstrated that X. maltophilia produces more than just the L1 and L2 beta-lactamases.

Anti-Bacterial Agents↗
PubMed1993

Dissemination of the TEM-I beta-lactamase gene into disparate plasmids in faecal strains of Escherichia coli isolated in Vellore, south India.

The genetic basis for ampicillin resistance in commensal strains of Escherichia coli isolated in Vellore, south India has been examined. Of the 58 strains tested, 41% could transfer ampicillin resistance to a standard E. coli host strain. With the exception of one isolate, transferable ampicillin resistance was shown to result from the presence of the TEM-I beta-lactamase which was found on a wide variety of plasmid types.

Ampicillin Resistance↗
PubMed1993

Molecular epidemiology of the plasmid-encoded TEM-1 beta-lactamase in Scotland.

A survey of the beta-lactamases responsible for ampicillin resistance in urinary Escherichia coli isolated in central Scotland has been performed. The TEM-1 beta-lactamase was found to be most prevalent occurring in 88.2% of ampicillin-resistant isolates tested. Forty-six percent of the TEM-1 producing strains were able to transfer this resistance to E. coli J62-2 at 37 degrees C. Analysis of the resulting transconjugants revealed that the degree of resistance to amoxycillin and amoxycillin in combination with clavulanic acid was related to the specific activity of the TEM-1 beta-lactamase. The variation in specific activity was shown to be related to plasmid type as determined by restriction analysis. No obvious relationship between beta-lactamase specific activity and resistance to amoxycillin and amoxycillin plus clavulanic acid could be demonstrated in the original plasmid donor strains.

Amoxicillin↗
PubMed1993

The prevalence of antimicrobial resistance in human faecal flora in South Africa.

Between January and March 1992, 361 faecal specimens were collected from the healthy black population in the Transvaal Province of South Africa. Each specimen was examined for the prevalence of antimicrobial resistance in commensal bacteria. Volunteers, from both rural and urban dwellings, were divided into four age groups. The overall carriage rate of resistance varied from 88.6% for ampicillin, 74.2% for trimethoprim, 52.6% for chloramphenicol, 10.2% for nalidixic acid to 7.5% for gentamicin. The carriage of resistance found to each individual antimicrobial agent was slightly higher in the rural population rather than the urban population but there was no correlation between the prevalence of antimicrobial resistance and the age group.

Adolescent↗
PubMed1993

Selection of variants of the TEM-1 beta-lactamase, encoded by a plasmid of clinical origin, with increased resistance to beta-lactamase inhibitors.

A TEM-1 derived variant beta-lactamase with increased resistance to beta-lactamase inhibitors was selected in vitro. The variant beta-lactamase was obtained from the parent strain Escherichia coli J62-2 containing resistance plasmid R1 which encodes the TEM-1 beta-lactamase. The variants were obtained by repeated subculture of E. coli J62-2 (R1) for five days in the presence of sub-inhibitory concentrations of amoxycillin plus clavulanic acid. Comparison of this variant beta-lactamase with a clinically isolated TEM derived beta-lactamase, with increased resistance to beta-lactamase inhibitors, suggests that they are the same enzyme. Both beta-lactamases have the same pI at 5.25 and have similar ID50 values for the beta-lactamase inhibitors clavulanic acid, sulbactam and tazobactam.

Amoxicillin↗
PubMed1993

Stability of cefdinir (C1-983, FK482) to extended-spectrum plasmid-mediated beta-lactamases.

Fourteen plasmid-encoded extended-spectrum beta-lactamases were purified from Escherichia coli transconjugants of original clinical isolates. The Vmax, Km and Vmax/Km were each determined for ampicillin, carbenicillin, cephaloridine, cephalexin, cefuroxime, cefixime, cefdinir, ceftazidime and cefotaxime as substrates with eight of these enzymes and with the narrow-spectrum beta-lactamase, TEM-1. The relative rates of hydrolysis of ampicillin, cephaloridine, cephalexin, cefuroxime, cefixime, cefdinir, ceftazidime and cefotaxime were also determined for the remaining six enzymes. Cefdinir had Vmax/Km or relative rates of hydrolysis values either equal to or lower than ampicillin, cephaloridine, cephalexin and cefotaxime for all the enzymes tested. Overall, cefdinir was more stable to the 15 beta-lactamases tested than either cefuroxime or cefixime; however, ceftazidime was more stable than cefdinir to hydrolysis by eight of the enzymes tested.

Cefdinir↗