Both EUCAST and CLSI recommend broth microdilution (BMD) for antimicrobial susceptibility testing of colistin, but BMD is rarely used in routine microbiology laboratories. The objective of this study ...was to evaluate five commercially available BMD products and two brands of gradient tests for colistin MIC determination using BMD according to ISO standard 20776-1 as reference.
Colistin MIC determination was performed according to the manufacturer's instructions on five commercially available BMD products (Sensititre, MICRONAUT-S, MICRONAUT MIC-Strip, SensiTest, and UMIC) and two gradient tests (Etest and MIC Test Strip). Colistin reference MICs were determined using frozen panels according to ISO standard 20776-1. An international collection of Gram-negative bacteria (n=75) with varying levels of colistin susceptibility was tested.
The colistin BMD products correlated well with reference tests, in particular for Sensititre and the two MICRONAUT products (essential agreement ≥96%: 66/69 (96%, CI 88–99%), 72/75 (96%, CI 88–99%) and 74/75 (99%, CI 92–100%)). The results were somewhat poorer for the BMD products SensiTest and UMIC: EA 88% (51/58, CI 77–95%) and 82% (61/74, CI 72–89%), respectively), and considerably poorer for the gradient tests (EA 43–71% depending on gradient test and Mueller-Hinton agar manufacturer). The gradient tests generally underestimated colistin MICs, resulting in a significant number of false susceptible results (9–18 of total 75 tests, compared with 1–3 for the BMD products).
Based on the results of this study, we advise laboratories not to trust gradient tests for colistin susceptibility testing and to use broth microdilution methods for this purpose. There are several commercial broth microdilution tests available and in principle they perform well.
Mueller–Hinton (MH) agar is recommended by EUCAST and CLSI for disc diffusion antimicrobial susceptibility testing. Using EUCAST methodology, we evaluated the performance of 21 internationally ...available brands of dehydrated MH agar from 17 manufacturers.
MH plates were prepared in-house and evaluated against four quality control (QC) strains tested in triplicate, using EUCAST disc diffusion methodology. This resulted in 30 disc–QC strain combinations and 90 readings per MH brand. All brands were tested blindly and in parallel. Results were evaluated against targets and ranges in the EUCAST QC tables. The agar depth, pH and concentration of five cations were measured for all brands.
Six brands of MH agar (Bio-Rad, Biolife, Oxoid, Sigma MH 2, BD BBL MH II and CRITERION) demonstrated excellent performance, with ≥99% of zone diameter readings within QC ranges and ≥70% on target ±1 mm. The poorest performance was seen for Biolab and Merck MH, with 10% (9/90) and 23% (21/90) of readings outside the QC ranges, respectively. Of all readings, 4.9% (93/1890) were out of range, mainly related to trimethoprim sulfamethoxazole (n = 25), aminoglycosides (n = 25) and fluoroquinolones (n = 15). The cation content differed considerably between the agars, and for four brands pH values were outside the acceptable range 7.2–7.4.
This study evaluated the performance and content of 21 brands of MH dehydrated media. Six brands showed excellent performance with all investigated antimicrobial classes. Others exhibited problems with one or more classes of agents. This could partly be explained by differences in concentration of specific chemical components and pH.
Antimicrobial discs for susceptibility testing can be obtained from many manufacturers. We evaluated the quality of discs from nine manufacturers in 2014 and 2017.
Antimicrobial discs of 16 agents ...from nine manufacturers were evaluated using EUCAST criteria. Discs were tested in triplicate on Müller–Hinton medium against EUCAST quality control (QC) strains. Mean values were compared with targets and ranges in the EUCAST QC tables.
Three manufacturers (Becton Dickinson, Mast and Oxoid) demonstrated excellent and consistent disc quality both in 2014 and 2017. Manufacturers with discs of inadequate quality improved their results between the two periods. Overall, 92% (795/861) versus 97% (1038/1071) of zone diameter readings were within QC ranges and 58% (497/861) versus 75% (806/1071) were within the QC target ± 1 mm, for the first and second studies, respectively. One manufacturer (HiMedia) had major quality problems with 33% (26/78) of readings out of range in the first study and 17% (20/120) in the second study. Discs from some manufacturers showed unexpected variation in inhibition zone diameters (4–9 mm) for discs within the same vial.
Antimicrobial discs from three of nine manufacturers exhibited excellent and reproducible quality. The discs of the other six manufacturers demonstrated various quality issues, some of which were severe. After presenting the results to manufacturers and users, all managed to improve the quality. Our study points to the need for more stringent criteria for disc manufacturing. Criteria should not only address the nominal potency of discs but also define the end result.
Increasing use of improved culture techniques and sensitive nucleic acid amplification assays have resulted in recognition of Kingella kingae as an important cause of invasive infections in young ...children, especially in septic arthritis, osteomyelitis, bacteraemia, and endocarditis. In 2016, EUCAST established clinical MIC breakpoints for K. kingae (published in EUCAST Clinical Breakpoint Tables v 7.0, 2017). The present study was carried out to produce MIC-zone diameter correlations for K. kingae on an international collection of isolates, with the aim of suggesting zone diameter breakpoints corresponding to the clinical MIC breakpoints.
Antimicrobial susceptibility testing was performed for 18 clinically relevant agents on a collection of 159 clinical isolates of K. kingae. Broth microdilution MIC determination and disk diffusion were performed according to EUCAST recommendations for fastidious organisms.
The correlation between MICs and zone diameters was good for all agents with EUCAST breakpoints for K. kingae. β-lactamase was detected in 41 isolates (26%) and these isolates were resistant to aminopenicillins. These isolates were also resistant to trimethoprim-sulfamethoxazole. Resistance to tetracyclines was detected in 8% of all isolates. All resistant isolates were correctly categorized for these agents with the proposed zone diameter breakpoints. One isolate, resistant to erythromycin but susceptible to other macrolides, was categorized as susceptible with erythromycin disk diffusion. No resistance was detected for the cephalosporins, carbapenems, and fluoroquinolones tested.
Based on the results in this study, zone diameter breakpoints for K. kingae calibrated to EUCAST clinical MIC breakpoints were proposed and approved by EUCAST.
Melioidosis, caused by Burkholderia pseudomallei, requires intensive antimicrobial treatment. However, standardized antimicrobial susceptibility testing (AST) methodology based on modern principles ...for determining breakpoints and ascertaining performance of methods are lacking for B. pseudomallei. This study aimed to establish MIC and zone diameter distributions on which to set epidemiological cut-off (ECOFF) values for B. pseudomallei using standard EUCAST methodology for non-fastidious organisms.
Non-consecutive, non-duplicate clinical B. pseudomallei isolates (9–70 per centre) were tested at eight study centres against eight antimicrobials by broth microdilution (BMD) and the EUCAST disc diffusion method. Isolates without and with suspected resistance mechanisms were deliberately selected. The EUCAST Development Laboratory ensured the quality of study materials, and provided guidance on performance of the tests and interpretation of results. Aggregated results were analysed according to EUCAST recommendations to determine ECOFFs.
MIC and zone diameter distributions were generated using BMD and disc diffusion results obtained for 361 B. pseudomallei isolates. MIC and zone diameter ECOFFs (mg/L; mm) were determined for amoxicillin-clavulanic acid (8; 22), ceftazidime (8; 22), imipenem (2; 29), meropenem (2; 26), doxycycline (2; none), tetracycline (8; 23), chloramphenicol (8; 22) and trimethoprim-sulfamethoxazole (4; 28).
We have validated the use of standard BMD and disc diffusion methodology for AST of B. pseudomallei. The MIC and zone diameter distributions generated in this study allowed us to establish MIC and zone diameter ECOFFs for the antimicrobials studied. These ECOFFs served as background data for EUCAST to set clinical MIC and zone diameter breakpoints for B. pseudomallei.
Until recently, the European Committee on Antimicrobial Susceptibility Testing (EUCAST) recommended the cefoxitin disc to screen for mecA-mediated β-lactam resistance in Staphylococcus ...pseudintermedius. A recent study indicated that cefoxitin was inferior to oxacillin in this respect. We have re-evaluated cefoxitin and oxacillin discs for screening for methicillin resistance in S. pseudintermedius.
We included 224 animal and human S. pseudintermedius isolates from Europe (n = 108) and North America (n = 116), of which 109 were mecA-positive. Disc diffusion was performed per EUCAST recommendations using 30-μg cefoxitin and 1-μg oxacillin discs from three manufacturers and Mueller–Hinton agar from two manufacturers.
Cefoxitin inhibition zones ranged from 6 to 33 mm for mecA-positive S. pseudintermedius (MRSP) and from 29 to 41 mm for mecA-negative S. pseudintermedius (MSSP). The corresponding oxacillin zone intervals were 6–20 mm and 19–30 mm. For cefoxitin 16% (95% CI 14.8–18.0%) of the isolates were in the area where positive and negative results overlapped. For oxacillin the corresponding number was 2% (1.6–2.9%). For oxacillin a breakpoint of susceptible (S) ≥ 20 mm and resistant (R) <20 mm resulted in only 0.4% and 1.1% very major error and major error rates respectively.
This investigation confirms that the 1-μg oxacillin disc predicts mecA-mediated methicillin resistance in S. pseudintermedius better than the 30-μg cefoxitin disc. For a 1-μg oxacillin disc we propose that 20 mm should be used as cut off for resistance, i.e. isolates with a zone diameter <20 mm are resistant to all β-lactam antibiotics except those with activity against methicillin-resistant staphylococci.
Objective:
Women with a newly diagnosed hormone receptor-positive breast cancer are offered adjuvant endocrine therapy (AET). Although the treatment reduces the risk of relapse and death not all ...women are adherent to it. Many factors, including the therapy’s menopausal side effects, can adversely affect adherence to the treatment. This study explores the extent to which women treated with AET perceived that health care providers addressed their side effects.
Methods:
Ten focus groups were set up, containing between four to nine women. In total, 58 women participated in the study—45 from the Stockholm metropolitan region and 13 from the scarcely populated Norrbotten region. The interviews were analyzed using qualitative content analysis with an inductive approach.
Results:
The women were usually satisfied with the care they received from the health care providers. However, their experiences were more complex when it came to their satisfaction with the care in terms of the menopausal side effects of therapy, sexuality in particular. The participants reported that their healthcare providers rarely asked about sex life-related side effects of the treatment.
Conclusions:
Health care providers need to communicate and consult about issues related to their patients’ sex lives following their breast cancer diagnosis and during their treatment.
The nematode-trapping fungus Arthrobotrys oligospora was transformed to hygromycin resistance using the hygromycin-B phosphotransferase gene from Escherichia coli under the control of various ...heterologous fungal promoters. Plasmid DNA was introduced into fungal protoplasts by polyethylene glycol/CaCl2 treatment. Transformation frequencies varied between 1-6 transformants per microgram DNA. Seven out of 13 integration events analyzed from transformants were single copy integrations, whereas the remaining were multiple and more complex integrations. The addition of restriction enzymes during transformations increased the frequency of single copy integrations. Co-transformation, using the E. coli uidA gene encoding the beta-glucuronidase reporter gene under the control of an Aspergillus nidulans promoter, occurred at frequencies of up to 63%.
The nematode-trapping fungus
Arthrobotrys oligospora was transformed to hygromycin resistance using the hygromycin-B phosphotransferase gene from
Escherichia coli under the control of various ...heterologous fungal promoters. Plasmid DNA was introduced into fungal protoplasts by polyethylene glycol/CaCl
2 treatment. Transformation frequencies varied between 1–6 transformants per μg DNA. Seven out of 13 integration events analyzed from transformants were single copy integrations, whereas the remaining were multiple and more complex integrations. The addition of restriction enzymes during transformations increased the frequency of single copy integrations. Co-transformation, using the
E. coli uidA gene encoding the β-glucuronidase reporter gene under the control of an
Aspergillus nidulans promoter, occurred at frequencies of up to 63%.
Asymmetric Arylation of Ketone Enolates Åhman, Jens; Wolfe, John P; Troutman, Malisa V ...
Journal of the American Chemical Society,
03/1998, Letnik:
120, Številka:
8
Journal Article
PDF
