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Biomedical subjects

D M Livermore

Publications and source records attributed to D M Livermore.

At least 19 recordsLinked to original sources
PubMed2006

Efficacy of practised screening methods for detection of cephalosporin-resistant Enterobacteriaceae.

OBJECTIVES: Enterobacteriaceae with extended-spectrum beta-lactamases (ESBLs) are now widespread and simple phenotypic tests are required to detect them in diagnostic laboratories. We investigated the performance of screening methods at 16 hospitals in South-East England. METHODS: Sixteen laboratories in South-East England submitted 1195 consecutive Enterobacteriaceae isolates found to be resistant, by their routine methods, to any or all of cefpodoxime, ceftazidime and cefotaxime. These isolates were re-tested centrally with various cephalosporin/clavulanate combinations and with multiplex PCR for bla(CTX-M) and bla(AmpC) alleles. RESULTS: Screening methods among the laboratories were the following: cefpodoxime discs alone (1 site); cefpodoxime, cefotaxime and ceftazidime discs (9 sites) or agar dilution (1 site); Phoenix (2 sites), Vitek 1 (1 site) and Vitek 2 (2 sites). A total of 8% of isolates submitted based on disc tests proved fully cephalosporin-susceptible, compared with 3% sent based on tests with automated systems and none of those sent based on agar dilution tests. Among isolates submitted solely on cefpodoxime resistance 256/372 (69%) proved cephalosporin-susceptible or had only borderline resistance with no clear mechanism demonstrable; this proportion decreased to 28/160 (18%) for those submitted on the basis of resistance to ceftazidime, 18/122 (15%) for those resistant to cefotaxime and 26/496 (5%) for those resistant to both cefotaxime and ceftazidime. The inference of ESBL production by Vitek 2 had the best agreement with reference laboratory results. CONCLUSIONS: Many isolates found resistant only to cefpodoxime at the source sites proved not to have ESBLs or AmpC; screening with cefotaxime and ceftazidime allowed better specificity for identification of mechanism-based resistance, as did the automated systems. Cefpodoxime disc tests nevertheless remain a useful primary screen for laboratories prepared only to test one agent.

Cephalosporins↗
PubMed2006

Clinical relevance of laboratory-reported antibiotic resistance in acute uncomplicated urinary tract infection in primary care.

OBJECTIVES: To determine whether patients with an uncomplicated community-acquired urinary tract infection (UTI) and an isolate resistant to trimethoprim had worse clinical outcomes following empirical treatment with trimethoprim 200 mg twice daily for 3 days than did those with a susceptible isolate. PATIENTS AND METHODS: This was a prospective cohort study of clinical outcome. We enrolled 497 women (>or=18-70 years) presenting to general practitioner surgeries in Norwich and Gloucester with at least two symptoms of acute (<7 days) uncomplicated UTI. Significant bacteriuria was defined as >or=10(4) cfu/mL from a mid-stream urine (MSU). RESULTS: Of enrolled patients 75% (334/448) had significant bacteriuria, and trimethoprim resistance was present in 13.9% (44/317) of isolates. Patients with resistant isolates had a longer median time to symptom resolution (7 versus 4 days, P=0.0002), greater reconsultation to the practice (39% versus 6% in first week, P<0.0001), more subsequent antibiotics (36% versus 4% in first week, P<0.0001) and higher rates of significant bacteriuria at 1 month (42% versus 20% with susceptible isolate, P=0.04). Half of patients reconsulting in the first week had a resistant organism. CONCLUSIONS: Patients with uncomplicated UTI caused by trimethoprim-resistant organisms had significantly worse clinical outcomes than those with trimethoprim-susceptible organisms. Nevertheless, trimethoprim resistance was rarer than predicted from routine laboratory submissions and we calculate that 23 women require microbiological investigation to prevent one reconsultation arising from resistance-based treatment failure. We therefore suggest empirical antibiotic treatment in acute, uncomplicated UTIs. If patients reconsult in the first week, we suggest a change of antibiotic treatment with urine culture and susceptibility testing then done. More generally, laboratory resources should concentrate on resistance surveillance to inform empirical antibiotic choice.

Acute Disease↗
PubMed2006

Molecular characterization of plasmids encoding CTX-M-15 beta-lactamases from Escherichia coli strains in the United Kingdom.

OBJECTIVES: The UK, like other countries worldwide, has a growing problem with CTX-M beta-lactamase-producing Escherichia coli. Five major clonally related strains have been identified among CTX-M-15 producers. We characterize here the plasmids from clonal strains A and D. METHODS: Plasmids were extracted and transformed into E. coli DH5alpha; conjugative mating was attempted on agar. MICs were determined by agar dilution. beta-Lactamases were typed by isoelectric focusing; antibiotic resistance genes and integrons were identified by PCR and sequenced. Plasmid incompatibility groups were determined by replicon PCR. RESULTS: bla(CTX-M-15) was carried by a 150 kb plasmid in strain A and a 70 kb plasmid in strain D. Conjugative transfer of cefotaxime resistance was only achieved from strain D; plasmids from both strains were transferred by transformation. The plasmid from strain A additionally carried bla(TEM-1) (variably), bla(OXA-1), aac(6')-Ib-cr and tet(A), as well as a class 1 integron with the gene cassettes aadA5 and dfr(17); the plasmid from strain D carried bla(TEM-1) consistently, also bla(OXA-1), aac(6')-Ib-cr, aac3-IIa and tet(A). Both plasmids belonged to incompatibility group FII. CONCLUSIONS: bla(CTX-M-15) was plasmid-mediated in both strains A and D and was linked to other antibiotic resistance genes including aac(6')-Ib-cr, which encodes an acetyltransferase, not previously found in Europe, acting on both aminoglycosides and some fluoroquinolones. Although the plasmids from the two strains differed in size, both were related and conferred similar multi-drug resistance phenotypes, suggesting that they may share a similar genetic scaffold. Both shared features with plasmids encoding CTX-M-15 beta-lactamases in E. coli from Canada and India.

Anti-Bacterial Agents↗
PubMed2006

Comparison of four methods for detection of teicoplanin resistance in methicillin-resistant Staphylococcus aureus.

OBJECTIVES: To determine which method of determining the MIC of teicoplanin produces a result closely related to outcome in the critically ill patient. METHODS: Four methods of teicoplanin susceptibility testing-disc diffusion, Etest, VITEK (Legacy and VITEK 2) and agar incorporation-were compared for 47 methicillin-resistant Staphylococcus aureus (MRSA) isolates from invasive intensive care unit (ICU) infections and 83 isolates from ICU patients colonized with the organism. Clinical outcome was recorded prospectively for all the patients. Another 13 reference laboratory strains of MRSA with reduced susceptibility to teicoplanin were tested. RESULTS: Both VITEK systems failed to demonstrate resistance in the three isolates identified as resistant by Etest or agar incorporation, and disc testing detected only one resistant isolate. A higher MIC, as found by Etest or agar incorporation, was associated with lower survival (n = 130, 95% CI -0.082 to -0.006, P = 0.023, Etest; n = 130, 95% CI -0.156 to -0.020, P = 0.011, agar). The findings for the 13 reference strains were similar, with a > or = 4-fold reduction in MIC between agar incorporation or Etest and VITEK2 for six isolates. CONCLUSIONS: Neither disc diffusion nor the VITEK systems are reliable for detection of teicoplanin resistance in MRSA. Etest and agar incorporation remain the methods of choice.

Anti-Bacterial Agents↗
PubMed2006

Can beta-lactams be re-engineered to beat MRSA?

Methicillin-resistant Staphylococcus aureus (MRSA) strains are important nosocomial pathogens worldwide and now are also of growing importance in community-acquired infection. Their resistance depends upon a supplementary peptidoglycan transpeptidase, PBP2' (PBP-2a), which continues to function when normal PBPs have been inactivated by beta-lactams. PBP2' is encoded by the mecA gene, which is carried by the staphylococcal cassette chromosome, a large and somewhat variable DNA insert of uncertain origin. PBP2' does not wholly lack affinity for beta-lactams, but its affinity for available analogues is very weak. In principle, it should be possible to re-engineer beta-lactams to bind PBP2' strongly, and the desirability of this approach is self-evident: no other antibiotic class has a record equal to the beta-lactams for safety and efficacy. Moreover, there is consensus that beta-lactams are inherently more efficacious than vancomycin against infections due to susceptible staphylococci. In practice, finding viable PBP2'-active beta-lactams has proved difficult and the catalogue of near-misses extends back to the 1980s. At last, however, one cephalosporin with high affinity for PBP2'--ceftobiprole--is entering phase III trials. Ceftobiprole inhibits MRSA at 1-2 mg/L under standard conditions. Even when mecA/PBP2' was induced strongly, ceftobiprole MICs for MRSA only reached 4 mg/L, a clinically attainable concentration. A phase II trial in skin and skin structure infection recorded cures by ceftobiprole in 4/4 MRSA infections, and results of the phase III trials are awaited with great interest.

Anti-Bacterial Agents↗
PubMed2006

Development of high-level ceftazidime resistance via single-base substitutions of blaCTX-M-3 in hyper-mutable Escherichia coli.

Mutations can increase the ceftazidimase activity of CTX-M-3 beta-lactamase, as seen with its widespread variant CTX-M-15. This study compared the frequencies of emerging ceftazidime resistance in isogenic wild-type and hyper-mutable mutS CTX-M-3-producing Escherichia coli strains, and sequenced the mutant bla(CTX-M) alleles selected. Ceftazidime resistance emerged more readily in the hyper-mutable background than in the wild-type strain. All selected CTX-M mutants, in both the wild-type and the mutS derivatives, had single amino-acid changes at position 167, including a novel Pro167Gln substitution. These data emphasise the potential for further diversification of CTX-M enzymes.

Anti-Bacterial Agents↗
PubMed2006

Development of extended-spectrum activity in TEM beta-lactamases in hyper-mutable, mutS Escherichia coli.

TEM-1 and TEM(pUC19)beta-lactamases can gain activity against ceftazidime and other expanded-spectrum cephalosporins via point mutation. The frequency of emergent resistance to ceftazidime at 4 x MIC was elevated >or= 250-fold in hyper-mutable, MutS-deficient Escherichia coli harbouring these beta-lactamase genes on high- or low-copy plasmids. Moreover, although ceftazidime-resistant mutants, or those with reduced susceptibility, were selected in both the wild-type and mutS hosts, many more mutants in the mutS host showed ceftazidimase-type extended-spectrum beta-lactamase (ESBL) activity. This correlated with a G-A point mutation at position 484 in the bla(TEM-1) and bla(TEM-pUC19) genes, conferring the Arg164His amino-acid substitution found in the TEM-29 ESBL. Non-ESBL mutants lacked changes in bla(TEM).

Ceftazidime↗
PubMed2005

Effect of medium type, age and aeration on the MICs of tigecycline and classical tetracyclines.

BACKGROUND: Tigecycline is a minocycline derivative, belonging to a chemical class prone to oxidation and to medium-related effects on susceptibility. We therefore studied the effect of medium type and age on the tigecycline MICs found in broth and on agar by the Clinical Laboratory Standards Institute and British Society for Applied Chemotherapy methods. METHODS: MICs of tigecycline, minocycline and tetracycline were determined for 96 non-fastidious bacteria and 20 streptococci using Mueller-Hinton broth (MHB), Mueller-Hinton agar (MHA), Iso-Sensitest broth (ISB) and Iso-Sensitest agar (ISA) as: (i) freshly-prepared media; (ii) fresh media but with the antibiotic added 1 day before inoculation and (iii) media stored for 7 days before antibiotic addition and use. RESULTS: MICs of tigecycline in fresh MHB were up to two doubling dilutions higher than on or in MHA, ISA or ISB. Media with tigecycline or classical tetracyclines added a day before use gave raised MICs, though rarely by more than one dilution. MICs of tigecycline (less so classical tetracyclines) were increased in 7-day-old MHB or ISB, even though the antibiotic was freshly added. This latter effect was greatest for the most susceptible strains and was absent or much reduced for organisms with MICs > or = 8 mg/L; it did not occur in the corresponding agar dilution tests. Addition of blood to MHB largely abrogated the effect, as did steaming the broth before adding the antibiotic. CONCLUSIONS: The raised MICs of tigecycline in aged broth probably reflect inactivation by dissolved oxygen. This accords with the lack of any MIC increase in newly-steamed (i.e. degassed) MHB or on aged agar (which is melted at 100 degrees C before use). Blood, which also abrogated the effect, may increase reducing capacity, protecting the tigecycline. At a practical level, broth MIC determinations for tigecycline should always employ fresh media.

Anti-Bacterial Agents↗
PubMed2005

CTX-M: changing the face of ESBLs in the UK.

The UK has experienced a sudden rise in extended-spectrum beta-lactamase (ESBL) rates, largely due to the appearance and spread of Escherichia coli producing CTX-M-15 type beta-lactamase. The British Society for Antimicrobial Chemotherapy organized two update meetings during 2004 to report and discuss the recognition, clinical diagnosis, treatment and control of bacteria producing these beta-lactamases. This paper reports the data and reviews made by contributors to the conferences. The historical distribution and emergence of ESBLs was reviewed along with the emergence of plasmid-mediated CTX-M ESBLs following their mobilization from the chromosome of Kluyvera spp. The first significant outbreak of CTX-M producers in the UK occurred in 2001 and involved Klebsiella pneumoniae with CTX-M-26 at one site, but by 2003, cloned and diverse E. coli with CTX-M-15 were widespread, with Shropshire one of the most affected regions. The specific experience in Shropshire was reported on and a comprehensive review made of the level of awareness of the need for ESBL detection in laboratories in England and Wales, together with a description of the variety of methods that may be applied, with recommendations for optimal methodology. The increased mortality associated with inappropriate treatment of infections caused by ESBL-producing strains was highlighted, together with discussion on potential control of cross-infection. The meeting concluded that the CTX-M genes have now become widespread in not only E. coli but other Enterobacteriaceae in the UK and this will represent a substantial threat to both the treatment of infections caused by these bacteria in the community and within hospitals.

Anti-Bacterial Agents↗
PubMed2004

Community and hospital spread of Escherichia coli producing CTX-M extended-spectrum beta-lactamases in the UK.

OBJECTIVES: During 2003, the Health Protection Agency's Antibiotic Resistance Monitoring and Reference Laboratory began to receive isolates of Escherichia coli for confirmation of extended-spectrum beta-lactamase production with a phenotype implying a CTX-M-type beta-lactamase, i.e. MICs of cefotaxime > or = 8-fold higher than MICs of ceftazidime. Many were referred as being from community patients. We examined 291 CTX-M-producing isolates from the UK and investigated the genetic basis of their phenotype. METHODS: PCR was used to detect alleles encoding CTX-M enzymes and to assign these to their blaCTX-M phylogenetic groups. Selected alleles were sequenced. Producers were compared by analysis of banding patterns generated by pulsed-field gel electrophoresis of XbaI-digested genomic DNA. MICs were determined by an agar dilution method or by Etest. RESULTS: Of 291 CTX-M-producing E. coli isolates studied from 42 UK centres, 70 (24%) were reportedly from community patients, many of whom had only limited recent hospital contact. Community isolates were referred by 12 centres. Two hundred and seventy-nine (95.9%) producers contained genes encoding group 1 CTX-M enzymes and 12 contained blaCTX-M-9-like alleles. An epidemic CTX-M-15-producing strain was identified, with 110 community and inpatient isolates referred from six centres. Representatives of four other major strains also produced CTX-M-15, as did several sporadic isolates examined. Most producers were multi-resistant to fluoroquinolones, trimethoprim, tetracycline and aminoglycosides as well as to non-carbapenem beta-lactams. CONCLUSIONS: CTX-M-producing E. coli are a rapidly developing problem in the UK, with CTX-M-15 particularly common. The diversity of producers and geographical scatter of referring laboratories indicates wide dissemination of blaCTX-M genes. Because of the public health implications, including for the treatment of community-acquired urinary tract infections, the spread of these strains--and CTX-M-15 beta-lactamase in particular--merits close monitoring.

Alleles↗
PubMed2004

Reliability of routine disc susceptibility testing by the British Society for Antimicrobial Chemotherapy (BSAC) method.

OBJECTIVES: To ascertain the agreement between MICs determined at a central laboratory, and susceptible, intermediate and resistant categorizations based on zone diameters recorded at diagnostic laboratories using the BSAC standardized method. METHODS: Standardized disc susceptibility tests were performed at sentinel laboratories in three surveys, with MIC tests performed on the collected isolates at a reference laboratory. The organisms comprised over 3300 Enterobacteriaceae, Acinetobacter spp., pseudomonads, staphylococci and enterococci, with over 29 000 antibiotic/organism tests in total. RESULTS: More than 90% of the antibiotic/organism combinations classed as susceptible by disc tests in the sentinel laboratories were confirmed by MIC testing. Disagreements were more frequent where disc tests indicated resistance, with half of the piperacillin/tazobactam resistance and one-third of the cephalosporin resistance found in Enterobacteriaceae by disc tests not being confirmed, and with three-quarters of teicoplanin resistance in enterococci not confirmed. None of the few apparent cases of meropenem resistance in Enterobacteriaceae or linezolid, quinupristin/dalfopristin or vancomycin resistance in staphylococci were confirmed by MIC testing. When disagreements were found between disc- and MIC-based categorization, MICs were commonly, although not invariably, one to three doubling dilutions above or below the breakpoint. However, many of the disagreements where MICs were three or more dilutions from the breakpoint were not seen when disc tests were repeated in the central laboratory. CONCLUSIONS: The BSAC disc method seems adequate for confirming susceptibility to guide therapy and to monitor resistance trends. Nevertheless, there must be concern about the over-estimation of many resistances, and frequent zone:MIC disagreements for isolates with borderline susceptibility.

Bacteria↗
PubMed2004

Activity of daptomycin against multi-resistant Gram-positive bacteria including enterococci and Staphylococcus aureus resistant to linezolid.

The activity of daptomycin was assessed against 545 isolates of Gram-positive bacteria including enterococci, coagulase-positive and -negative staphylococci, beta-haemolytic streptococci, viridans streptococci and coryneforms. The panel of isolates was biased towards those with resistance to first-line antimicrobial agents and included linezolid-resistant enterococci and Staphylococcus aureus. MICs of daptomycin were < or =1mg/l for all the staphylococci, streptococci, and coryneforms, except for two isolates of Corynebacterium aquaticum, for which MICs were 8 mg/l. Daptomycin MICs were < or =2 and < or =4 mg/l for Enterococcus faecalis and Enterococcus faecium, respectively.

Acetamides↗