Biomedical subjects

S G Amyes

Publications and source records attributed to S G Amyes.

At least 127 records · Page 7Linked to original sources

PubMed1988

Biotinylated DNA probes for trimethoprim-resistant dihydrofolate reductases types IV and V.

DNA probes for trimethoprim-resistant dihydrofolate reductases types IV and V were constructed and characterized. The type IV probe consisted of a 1.7 kb ClaI fragment of recombinant plasmid pUK1148, while the type V probe was a 0.5 kb HincII fragment of plasmid pLK09. Each probe was biotinylated and tested in non-isotopic hybridization experiments with control plasmids carrying genes for known dihydrofolate reductase types. Both probes were highly specific and will therefore be useful in epidemiological studies for monitoring the spread of these unusual plasmid-encoded enzymes.

Biotin↗

PubMed1988

Identification of a novel plasmid-encoded dihydrofolate reductase mediating high-level resistance to trimethoprim.

A Proteus mirabilis strain, isolated in South Africa, carried a 79kb transferable plasmid (pUK672) which encoded high-level resistance to trimethoprim (minimum inhibitory concentration greater than 2048 mg/l). The trimethoprim resistance was mediated by a dihydrofolate reductase which had completely different properties from any plasmid-encoded dihydrofolate reductase previously described. This enzyme has been designated type VI. The type VI enzyme is small (10,000 daltons) and is unstable in vitro. It is highly resistant to inhibition by both trimethoprim and methotrexate.

Drug Resistance, Microbial↗

PubMed1988

Trimethoprim resistance determinants encoding a dihydrofolate reductase in clinical isolates of Staphylococcus aureus and coagulase-negative staphylococci.

The molecular and biochemical basis of resistance to high concentrations (MIC greater than or equal to 1000 mg/L) of trimethoprim (Tpr(H] was examined in Australian isolates of Staphylococcus aureus and coagulase-negative staphylococci. The Tpr(H) determinant (dfr A) was located within a 2.75-Kb Bg/II fragment on the S. aureus aminoglycoside-resistance plasmids pSK1 and pSK16 as judged by comparative restriction mapping with two naturally-occurring variants, pSK9 and pSK14, which did not encode trimethoprim resistance. This was confirmed in DNA-DNA hybridisation experiments in which a 0.9-Kb sequence of pSK1 DNA was used as a specific probe for the Tpr(H) gene. Plasmid DNA from three strains of coagulase-negative staphylococci, and the chromosomal material of one other isolate, were found to share homology with the probe DNA. Dihydrofolate reductases produced by these strains were virtually identical to the type S1 enzyme encoded by the S. aureus plasmid pSK1. Interspecies transfer may have been responsible for the conservation of Tpr(H) gene sequences among staphylococci.

Chromosome Mapping↗

PubMed1988

Cephaloridine resistance in gram-negative bacteria isolated in Scotland.

The incidence of cephaloridine resistance (minimum inhibitory concentration, MIC greater than 8 mg L-1) in isolates from urinary tract infections was 45.1% in Glasgow, 22.6% in Dundee and 25.9% in Edinburgh. The incidence of ampicillin resistance (MIC greater than 8 mg L-1) was even higher:- being 45.2% in Dundee and 48.5% in Edinburgh. In Glasgow, the incidence was 71.9% which is the highest proportion of ampicillin resistance reported in the United Kingdom. The cephaloridine resistant strains were examined for beta-lactamase production. Amongst these strains 50.8% produced only a chromosomal beta-lactamase, whereas 47.9% produced beta-lactamases which were potentially plasmid-mediated on the basis of biochemical tests. Only 1% of the resistant strains produced no detectable beta-lactamase.

Ampicillin Resistance↗

PubMed1987

Plasmid-mediated trimethoprim-resistance in Staphylococcus aureus. Characterization of the first gram-positive plasmid dihydrofolate reductase (type S1).

The trimethoprim-resistance gene located on plasmid pSK1, originally identified in a multi-resistant Staphylococcus aureus from Australia, encodes the production of a dihydrofolate reductase (type S1), which confers a high degree of resistance to its host and is quite unlike any plasmid-encoded dihydrofolate reductase hitherto described. It has a low Mr (19,700) and has a higher specific activity than the constitutive Gram-negative plasmid dihydrofolate reductases. The type S1 enzyme is heat-stable and has a relatively low affinity for the substrate, dihydrofolate (Km 10.8 microM). It is moderately resistant to trimethoprim, and is competitively inhibited by this drug with an inhibitor-binding constant of 11.6 microM. This is the first identification and characterization of a plasmid-encoded trimethoprim-resistant dihydrofolate reductase derived from a Gram-positive species.

Drug Resistance, Microbial↗

PubMed1987

The genetics of bacterial trimethoprim resistance in tropical areas.

Resistance to trimethoprim in Gram-negative bacteria is largely manifested by two trimethoprim resistant dihydrofolate reductases (types I and II) encoded by genes originally located on resistance plasmids. Although trimethoprim resistance increased markedly after the clinical introduction of trimethoprim in the West, its spread has slowed and, in Edinburgh at least, has actually been declining. This reduction has been accompanied by the migration of a transposon, encoding the type I plasmid resistance gene, into the bacterial chromosome. In tropical areas, the incidence of trimethoprim resistance is very much higher. In Tanzania, it has spilled over into other bacteria outside the Enterobacteriaceae, but it was in India where the major problem existed. The majority (64%) of the Indian Enterobacteriaceae studied were resistant to the drug and most of the resistance genes were located on very large plasmids which also conferred resistance to many other antibacterial drugs. Some Indian plasmids carried a new trimethoprim resistance gene which is not detectable by conventional sensitivity tests and may be spreading unnoticed elsewhere. The proportion of trimethoprim resistance has been related to the volume of antibacterial drugs used.

Drug Resistance↗

PubMed1987

Trimethoprim resistance in commensal bacteria isolated from farm animals.

Trimethoprim resistance was examined in faecal bacteria obtained from chickens, sheep, cattle and pigs. The incidence of trimethoprim resistance in porcine strains was 17% (157/922) and, whereas 15.8% (146/922) of these bacteria were highly resistant, only 4% (37/922) of the isolates possessed trimethoprim resistance plasmids. Highly resistant porcine strains were obtained from 44% of the pig farms (41/93) but transferable trimethoprim resistance was found in isolates from 11% (10/93) of the farms. There was an association between the carriage of trimethoprim resistance plasmids and certain farms. Most of the resistance plasmids were not identical with those found in human clinical bacteria but one porcine plasmid was the same as the most ubiquitous trimethoprim resistance plasmid in Edinburgh.

Animals↗

PubMed1987

Ampicillin resistance in Haemophilus influenzae: identification of resistance mechanisms.

The incidence and mechanisms of ampicillin resistance (MIC greater than 1 mg/l) were investigated in 105 clinical isolates of Haemophilus influenzae collected in Edinburgh during 1983/4. Fifteen (14.3%) ampicillin-resistant strains were identified and these were non-serotypable and comprised six biotypes. Isoelectric focusing and beta-lactamase-inhibition studies demonstrated that production of the TEM-1 beta-lactamase was the principal mechanism of resistance in nine (60%) strains. Radiolabelling revealed that one beta-lactamase-positive strain also had an unusual penicillin-binding protein (PBP) profit. No beta-lactamase activity was detected in the other six (40%) ampicillin-resistant strains. Two beta-lactamase-negative ampicillin-resistant strains had atypical PBP profiles. SDS-PAGE analysis showed that four beta-lactamase-negative ampicillin-resistant strains, including one with altered PBPs, exhibited outer membrane protein profiles which differed from those of sensitive strains of the same biotype. The ampicillin-resistance mechanism of the remaining strain could not be determined. Thus, several resistance mechanisms, either acting individually or in combination, are implicated in ampicillin resistance in H. influenzae.

Ampicillin↗

PubMed1986

A new mechanism of plasmid trimethoprim resistance. Characterization of an inducible dihydrofolate reductase.

The dihydrofolate reductase encoded by plasmid pUK1123, which confers only a moderate level of trimethoprim resistance on its host, has been isolated and characterized. This enzyme, designated type IV, differs markedly from all previously described plasmid dihydrofolate reductases. It has a relatively high molecular weight of 46,700 as measured by gel filtration and, unlike previous plasmid dihydrofolate reductases, its synthesis is induced in the presence of increasing concentrations of trimethoprim. It is only slightly resistant to trimethoprim but is competitively inhibited by this drug with an inhibitor binding constant of 63 nM. In addition, the enzyme has a relatively low affinity for the substrate, dihydrofolate (Km = 37 microM). This is the first report of a plasmid trimethoprim resistance mechanism resulting from the induced synthesis of a large molecular weight dihydrofolate reductase which is only slightly resistant to trimethoprim. The possible origins of the type IV enzyme are discussed.

Chromatography, Gel↗

PubMed1986

High-level trimethoprim resistance in urinary bacteria.

The results of a three year evaluation of the incidence and type of trimethoprim resistance in pathogens responsible for significant bacteriuria in a general hospital in Edinburgh UK, are presented and compared to results of a previous study. In the present study, trimethoprim resistance was 50% more frequent in bacteria isolated from men and nearly twice as frequent in bacteria from elderly patients. However, the proportion of trimethoprim resistant strains fell annually when resistance was measured at trimethoprim concentrations of both 10 mg/l and 1000 mg/l. The proportion of strains able to transfer trimethoprim resistance also fell by half, and there was some movement of trimethoprim resistance transposons into the bacterial chromosome. These results suggest that migration of high-level trimethoprim resistance genes into the permanent location of the bacterial chromosome is occurring.

Adolescent↗

PubMed1986

Trimethoprim resistance amongst urinary pathogens in south India.

Two hundred and eighty four strains of Enterobacteriaceae, responsible for significant bacteriuria, were isolated, over a three month period, in Vellore, India. Sixty-four per cent of these strains were resistant to 10 mg/l of trimethoprim. Moreover, this population was dominated by high level resistance (minimum inhibitory concentration greater than 1000 mg/l) and these accounted for 57.3% of all strains studied. Over half of the resistant strains were able to transfer trimethoprim resistance to standard Escherichia coli strains. However, the high incidence of transferable resistance did not result from the spread of one plasmid type as 58 different plasmid types were identified. These results are in marked contrast to recent findings in Europe where the incidence of high level transferable trimethoprim resistance is falling.

Ampicillin↗

PubMed1986

Plasmid trimethoprim resistance in Vibrio cholerae: migration of the type I dihydrofolate reductase gene out of the Enterobacteriaceae.

Two strains of Vibrio cholerae biotype el tor were isolated in Tanzania. Each possessed a single resistance plasmid of 113 kbases belonging to the C incompatibility group. Both plasmids conferred resistance to sulphamethoxazole, ampicillin, streptomycin, chloramphenicol, tetracycline and trimethoprim. They are the first plasmids derived from V. cholerae strains, isolated in Southern Africa, to confer trimethoprim resistance. The trimethoprim resistance resulted from the plasmid-mediated production of a trimethoprim resistant dihydrofolate reductase which was very similar to the type I plasmid enzyme. This is the first example of movement of the gene encoding the type I dihydrofolate reductase into genera of other families besides the Enterobacteriaceae.

Drug Resistance, Microbial↗

PubMed1986

Epidemiology of trimethoprim resistance.

Trimethoprim resistance in the Enterobacteriaceae has been dominated by the spread of two genes encoding different dihydrofolate reductases (types IA and II). The original carriage of these genes was by transferable bacterial plasmids. Now, however, one of these genes is often found to be located on the bacterial chromosome. Two new plasmid-mediated dihydrofolate reductases have been identified. One of these enzymes (type IV) confers a low degree of insusceptibility but is inducible, to give levels 600 times higher than the bacterial host enzyme. The technique used to demonstrate the plasmids that carry the gene for this enzyme suggests that they would not be identified by the plasmid transfer methods in current use.

Animals↗

PubMed1986

The transfer of genes encoding production of mannose-resistant haemagglutinating fimbriae from uropathogenic enterobacteria.

Two hundred and thirty-seven bacterial strains, isolated from patients with urinary tract infections, were examined for the presence of plasmid-determined fimbrial adhesins. Ninety-nine strains were capable of producing mannose-resistant haemagglutination. Seventeen of these strains possessed transferable resistance plasmids and 11 of these were also able to transfer the mannose-resistant haemagglutination gene, suggesting that it was carried on the R plasmid or had been mobilized by it.

Adhesins, Escherichia coli↗

PubMed1986

Acquisition of a gene encoding mannose-resistant haemagglutinating fimbriae by a resistance plasmid during long-term urinary infection.

Urine was collected twice weekly from one patient during a 25-month period. Escherichia coli harbouring a resistance plasmid, pUK28, which confers trimethoprim, ampicillin, sulphamethoxazole, spectinomycin and streptomycin resistance was identified in the urine. Carriage of strains containing plasmid pUK28 was observed during three separate periods which totalled 16 months, even though the patient did not receive antibacterial drug therapy for most of that time. The plasmid was able to acquire the genes responsible for mannose-resistant haemagglutination and these genes were increasingly associated with the plasmid towards the end of the study period.

Aged↗

PubMed1986

Plasmid penicillin resistance in Vibrio cholerae: identification of new beta-lactamase SAR-1.

Two strains of Vibrio cholerae biotype El Tor, isolated in Tanzania, possessed a single IncC resistance plasmid of 113 kilobases. Both plasmids encoded the production of a novel beta-lactamase, SAR-1, which was 33,700 daltons in size and was able to hydrolyze carbenicillin as well as penicillin G. The SAR-1 beta-lactamase was quite distinct from all other plasmid beta-lactamases by virtue of its unusually low isoelectric point and a combination of its size, substrate profile, and inhibition properties. This enzyme is only the second beta-lactamase identified in V. cholerae species and the first to be reported in V. cholerae strains isolated in Southern Africa.

Ampicillin↗

PubMed1986

Unusual expression of new low-level-trimethoprim-resistance plasmids.

In a survey, 42% of trimethoprim-resistant clinical members of the family Enterobacteriaceae were able to transfer trimethoprim resistance to standard Escherichia coli strains when selection was made on complex bacteriological media. When transfer experiments were performed with minimal medium, another 16% of the clinical strains were shown to have transferred trimethoprim resistance. Twelve transconjugants produced negligible trimethoprim resistance in complex media but were resistant in minimal medium. The methionine, glycine, and purine components of complex media appeared to be responsible for the reduced expression of trimethoprim resistance in these strains.

Conjugation, Genetic↗

PubMed1986

Trimethoprim and co-trimoxazole: a comparison of their use in respiratory tract infections.

A single blind prospective study was undertaken with 74 patients suffering from acute bronchitis, taken from general practice and one geriatric ward. Half were randomly allocated to treatment with 200 mg trimethoprim twice a day and the other half with 160 mg trimethoprim plus 800 mg sulphamethoxazole twice a day; both therapies were used for 7 days. We found little difference in the clinical or bacteriological responses to the different regimens although the higher concentration of trimethoprim in the single therapy gave a slightly more successful eradication of Haemophilus spp. Resistant bacteria appeared during and after therapy in a few cases but this was a greater problem with the sulphamethoxazole-containing preparation.

Acute Disease↗