EUCAST technical note on Amphotericin B

The European Committee on Antimicrobial Susceptibility Testing-Subcommittee on Antifungal Susceptibility Testing (EUCAST-AFST) has determined breakpoints for amphotericin B for Candida spp. This Technical Note is based on the EUCAST amphotericin B rationale document (available on the EUCAST website: http://www.eucast.org). Species-specific breakpoints for C. albicans, C. glabrata, C. krusei, C. parapsilosis and C. tropicalis are S: MIC ≤1 mg/L, R: MIC > 1 mg/L. There are insufficient data to set breakpoints for other species. The breakpoints are based upon pharmacokinetic data, epidemiological cut-ff values and clinical experience. Breakpoints will be reviewed regularly.     

EUCAST technical note on posaconazole

The European Committee on Antimicrobial Susceptibility Testing-Subcommittee on Antifungal Susceptibility Testing (EUCAST-AFST) has determined breakpoints for posaconazole for Candida spp. This Technical Note is based on the EUCAST posaconazole rationale document (available on the EUCAST website: http://www.eucast.org). Species-specific breakpoints for C. albicans, C. parapsilosis and C. tropicalis are S: MIC ≤0.06 mg/L, R: MIC >0.06 mg/L. There are insufficient data to set breakpoints for C. glabrata and C. krusei as well as non-species-related breakpoints. The breakpoints are based upon pharmacokinetic data, epidemiological cut-off values and clinical experience. Breakpoints will be reviewed regularly.     

EUCAST Technical Note on Voriconazole and Aspergillus spp.

The European Committee on Antimicrobial Susceptibility Testing Subcommittee on Antifungal Susceptibility Testing (EUCAST-AFST) has determined breakpoints for voriconazole against Aspergillus spp. This Technical Note is based on the EUCAST rationale document for voriconazole (available on the EUCAST website: http://www.eucast.org). Voriconazole breakpoints are based on epidemiological cut-off values, pharmacokinetic/pharmacodynamic data and clinical experience. Breakpoints will be reviewed regularly or when new data emerge.     

International and multicenter comparison of EUCAST and CLSI M27-A2 broth microdilution methods for testing susceptibilities of Candida spp. to fluconazole, itraconazole, posaconazole, and voriconazole

The aim of this study was to compare MICs of fluconazole, itraconazole, posaconazole, and voriconazole obtained by the European Committee on Antibiotic Susceptibility Testing (EUCAST) and CLSI (formerly NCCLS) methods in each of six centers for 15 Candida albicans (5 fluconazole-resistant and 4 susceptible-dose-dependent [S-DD] isolates), 10 C. dubliniensis, 7 C. glabrata (2 fluconazole-resistant isolates), 5 C. guilliermondii (2 fluconazole-resistant isolates), 10 C. krusei, 9 C. lusitaniae, 10 C. parapsilosis, and 5 C. tropicalis (1 fluconazole-resistant isolate) isolates. CLSI MICs were obtained visually at 24 and 48 h and spectrophotometric EUCAST MICs at 24 h. The agreement (within a 3-dilution range) between the methods was species, drug, and incubation time dependent and due to lower EUCAST than CLSI MICs: overall, 94 to 95% with fluconazole and voriconazole and 90 to 91% with posaconazole and itraconazole when EUCAST MICs were compared against 24-h CLSI results. The agreement was lower (85 to 94%) against 48-h CLSI endpoints. The overall interlaboratory reproducibility by each method was > or =92%. When the comparison was based on CLSI breakpoint categorization, the agreement was 68 to 76% for three of the four species that included fluconazole-resistant and S-DD isolates; 9% very major discrepancies (< or =8 microg/ml versus > or =64 microg/ml) were observed among fluconazole-resistant isolates and 50% with voriconazole (< or =1 microg/ml versus > or =4 microg/ml). Similar results were observed with itraconazole for seven of the eight species evaluated (28 to 77% categorical agreement). Posaconazole EUCAST MICs were also substantially lower than CLSI MIC modes (0.008 to 1 microg/ml versus 1 to > or =8 microg/ml) for some of these isolates. Therefore, the CLSI breakpoints should not be used to interpret EUCAST MIC data.     

How to interpret MICs of antifungal compounds according to the revised clinical breakpoints v. 10.0 European committee on antimicrobial susceptibility testing (EUCAST)

BackgroundEUCAST has revised the definition of the susceptibility category I from ‘Intermediate’ to ‘Susceptible, Increased exposure’. This implies that I can be used where the drug concentration at the site of infection is high, either because of dose escalation or through other means to ensure efficacy. Consequently, I is no longer used as a buffer zone to prevent technical factors from causing misclassifications and discrepancies in interpretations. Instead, an Area of Technical Uncertainty (ATU) has been introduced for MICs that cannot be categorized without additional information as a warning to the laboratory that decision on how to act has to be made. To implement these changes, the EUCAST-AFST (Subcommittee on Antifungal Susceptibility Testing) reviewed all, and revised some, clinical antifungal breakpoints.ObjectivesThe aim was to present an overview of the current antifungal breakpoints and supporting evidence behind the changes.SourcesThis document is based on the ten recently updated EUCAST rationale documents, clinical breakpoint and breakpoint ECOFF documents.ContentThe following breakpoints (in mg/L) have been revised or established for Candida species: micafungin against C. albicans (ATU = 0.03); amphotericin B (S ≤/> R = 1/1), fluconazole (S ≤/> R = 2/4), itraconazole (S ≤/> R = 0.06/0.06), posaconazole (S ≤/> R = 0.06/0.06) and voriconazole (S ≤/> R = 0.06/0.25) against C. dubliniensis; fluconazole against C. glabrata (S ≤/> R = 0.001/16); and anidulafungin (S ≤/> R = 4/4) and micafungin (S ≤/> R = 2/2) against C. parapsilosis. For Aspergillus, new or revised breakpoints include itraconazole (ATU = 2) and isavuconazole against A. flavus (S ≤/> R = 1/2, ATU = 2); amphotericin B (S ≤/> R = 1/1), isavuconazole (S ≤ /> R = 1/2, ATU = 2), itraconazole (S ≤/> R = 1/1, ATU = 2), posaconazole (ATU = 0.25) and voriconazole (S ≤/> R = 1/1, ATU = 2) against A. fumigatus; itraconazole (S ≤/> R = 1/1, ATU = 2) and voriconazole (S ≤/> R = 1/1, ATU = 2) against A. nidulans; amphotericin B against A. niger (S ≤/> R = 1/1); and itraconazole (S ≤/> R = 1/1, ATU = 2) and posaconazole (ATU = 0.25) against A. terreus.ImplicationsEUCAST-AFST has released ten new documents summarizing existing and new breakpoints and MIC ranges for control strains. A failure to adopt the breakpoint changes may lead to misclassifications and suboptimal or inappropriate therapy of patients with fungal infections.     

Multicentre determination of quality control strains and quality control ranges for antifungal susceptibility testing of yeasts and filamentous fungi using the methods of the Antifungal Susceptibility Testing Subcommittee of the European Committee on Antimicrobial Susceptibility Testing (AFST-EUCAST)

A multicentre study involving seven laboratories was performed using techniques recommended by the Antifungal Susceptibility Testing Subcommittee of the European Committee on Antimicrobial Susceptibility Testing (AFST-EUCAST) to evaluate and propose quality control ranges and strains for susceptibility testing of fermentative yeasts and filamentous fungi. Participating laboratories tested the susceptibilities of a panel of 12 encoded isolates to amphotericin B, flucytosine, fluconazole, itraconazole, voriconazole and posaconazole. In total, 15 lots of assay medium were tested, with one lot being common to all laboratories, and 18 144 MIC values were determined. Intra- and inter-laboratory agreements and intra-class correlation coefficients (ICCs) of the results for each drug/strain/lot combination were calculated. An average value of 85% agreement was selected for validation purposes. The average percentage of intra-laboratory agreement was 90-95%, with ICC values of 0.90-0.95 (p <0.01). Inter-laboratory reproducibility was also high, with 92% agreement and an ICC of 0.97 (p <0.01). The reproducibility was somewhat better with the common lot of assay medium (96% agreement) than with the different lots (91% agreement), but this difference was not significant. Two isolates that showed trailing growth had agreement percentages below the 85% limit selected for validation purposes and were therefore excluded from the panel of quality control strains. The recommended EUCAST methodologies were found to be highly reproducible and reliable for susceptibility testing of yeasts and filamentous fungi. Ten isolates are proposed for use as quality control strains with these EUCAST procedures.     

Sequence-based identification,genotyping and EUCAST antifungal susceptibilities of Trichosporon clinical isolates from Greece

Trichosporon yeasts constitute emerging pathogens, implicated in organ-specific and systemic infections. In this first, comprehensive study of Trichosporon clinical isolates in Greece, 42 isolates were identified by sequencing the hypervariable D1/D2 domain of the Large Subunit (LSU) rDNA gene, while Trichosporon asahii were genotyped by sequencing the Intergenic Spacer 1 region, and antifungal susceptibilities were determined by the EDef 7.2 (EUCAST) method, in parallel with the CLSI standard. Trichosporon asahii was the primary species (37 isolates) followed by Trichosporon coremiiforme, Trichosporon dermatis, Trichosporon loubieri and Trichosporon mycotoxinivorans. One strain remained unidentified. Seven T. asahii genotypes were recorded. The major genotypes were: genotypes 4 (29%) and 3 (26%) followed by 1, 5 and 7 (9.5% each). Two novel genotypes were identified designated as 10 and 11. EUCAST MIC ≥2 mg/L was recorded in 58% of the isolates (amphotericin B), 41% (itraconazole), 41% (posaconazole) and 38% (voriconazole). Fluconazole MICs of ≥32 mg/L were recorded in 23.8% of the isolates. Analysis of variance performed on absolute values showed that the amphotericin B, itraconazole, posaconazole and voriconazole MICs of T. asahii were equivalent. Typically higher MIC values were displayed by fluconazole. Antifungal susceptibilities of the seven different genotypes were homogeneous. Agreements between EUCAST and CLSI ranged from 88.1 to 97.62%. Overall, the high MICs recorded among the Trichosporon isolates for all tested drugs justify routine susceptibility testing of clinical isolates.     

EUCAST technical note on anidulafungin

The European Committee on Antimicrobial Susceptibility Testing-Subcommittee on Antifungal Susceptibility Testing has determined breakpoints for anidulafungin for Candida spp. This Technical Note is based on the EUCAST anidulafungin rationale document (available at: http://www.eucast.org). Species-specific breakpoints for C. albicans are S ≤0.03 mg/L and R >0.03 mg/L and for C. glabrata, C. tropicalis and C. krusei S ≤0.06 mg/L and R >0.06 mg/L. C. parapsilosis was not regarded a good target for anidulafungin. There are insufficient data to set breakpoints for other species. The breakpoints are based upon pharmacokinetic data, epidemiological cut-off values and clinical experience. Breakpoints will be reviewed regularly.     

Unpredictable susceptibility of emerging clinical moulds to tri-azoles: review of the literature and upcoming challenges for mould identification

Tri-azoles represent the front-line drugs for the treatment of mould diseases; nevertheless, some emerging moulds, such as Fusarium spp., Scedosporium spp., Mucorales and others, may be less susceptible or resistant to these antifungals. A review of the literature was conducted on the susceptibility of rare moulds to the tri-azoles itraconazole, posaconazole and voriconazole. Particular attention was paid to isolates identified by molecular analyses. The range of susceptibility values described for the three tri-azoles was frequently large (from 0.06 to >16), and a high variability was found within each species; isolates were rarely reported as entirely susceptible to all tri-azoles. In addition, the susceptibility of 76 emerging moulds from our collection (including Hypocreales, Dothideomycetes, Scedosporium spp., Mucorales and rare Aspergillus spp.) to itraconazole and voriconazole was determined by the Clinical and Laboratory Standards Institute (CLSI) M38-A2 and European Committee for Antimicrobial Susceptibility Testing (EUCAST) methods. Susceptibility discrepancies (of two dilutions) were found comparing CLSI and EUCAST for Dothideomycetes; the values for the remaining moulds were similar. More practical, faster and inexpensive susceptibility tools are welcome for testing emerging moulds, as these tests still represent a critical tool to support clinicians on the selection of proper antifungal treatment. The susceptibility of emerging moulds to tri-azoles cannot be predicted exclusively following mould identification, as the isolates’ susceptibilities showed highly variable values. Some emerging moulds still remain very difficult to identity, even following standard molecular analyses which result in complex fungal collections. This fact limits the definition of epidemiological cut-offs and clinical breakpoints that are still imperative for emerging moulds.     

European Committee on Antimicrobial Susceptibility Testing (EUCAST) Technical Notes on antimicrobial susceptibility testing

The main objectives of the European Committee on Antimicrobial Susceptibility Testing (EUCAST) are to harmonise breakpoints for antimicrobial agents in Europe, and to act as the breakpoint committee for the European Medicines Agency (EMEA) during the registration of new antimicrobial agents. Detailed EUCAST procedures for harmonising and setting breakpoints for antimicrobial agents are available on the EUCAST website. Beginning with the current issue, a series of EUCAST Technical Notes will be published in CMI, based on the rationale documents produced by EUCAST for each of the antimicrobial agents studied, with the aim of highlighting important background information underlying decisions on breakpoints made by EUCAST.     

How to: perform antifungal susceptibility testing of microconidia-forming dermatophytes following the new reference EUCAST method E.Def 11.0, exemplified by Trichophyton

BackgroundAntifungal drug resistance in dermatophytes was first reported shortly after the turn of the millennium and has today been reported in Trichophyton and occasionally in Microsporum, but not in Epidermophyton species. Although drug resistance in dermatophytes is not routinely investigated, resistance in Trichophyton spp. is increasingly reported worldwide. The highest rates are observed in India (36% and 68% for terbinafine (MIC ≥4 mg/L) and fluconazole (MICs ≥16 mg/L), respectively), and apparently involve the spread of a unique clade related to the Trichophyton mentagrophytes/Trichophyton interdigitale complex.ObjectivesThe European Committee on Antimicrobial Susceptibility Testing Subcommittee on Antifungal Susceptibility Testing (EUCAST-AFST) has released a new method (E.Def 11.0) for antifungal susceptibility testing against microconidia-forming dermatophytes including tentative MIC ranges for quality control strains and tentative breakpoints against Trichophyton rubrum and T. interdigitale. Here, the details of the new procedure E.Def 11.0 are described.SourcesThis technical note is based on the multicentre validation of the EUCAST dermatophyte antifungal susceptibility testing method, the mould testing method (E.Def 9.3.2) and the updated quality control tables for antifungal susceptibility testing document, v 5.0 (available on the EUCAST website).ContentsThe method is based on the EUCAST microdilution method for moulds but significant differences include: (a) an altered test medium selective for dermatophytes; (b) an altered incubation time and temperature; and (c) a different end-point criterion (spectrophotometric determination) of fungal growth. It can easily be implemented in laboratories already performing EUCAST microdilution methods and has been validated for terbinafine, voriconazole, itraconazole and amorolfine against T. rubrum and T. interdigitale.ImplicationsThis standardized procedure with automated end-point reading will allow broader implementation of susceptibility testing of dermatophytes and so facilitate earlier appropriate therapy. This is important, as resistance is rapidly emerging and largely underdiagnosed.     

European Confederation of Medical Mycology (ECMM) epidemiological survey on invasive infections due to Fusarium species in Europe

In order to better understand the epidemiology of fusariosis in Europe, a survey collecting information on the clinical characteristics of the patients infected by Fusarium as well as on the infecting isolates was launched. A total of 76 cases of invasive fusariosis occurring from January 2007 to June 2012 were collected and Fusarium isolates were identified by sequencing the translation elongation factor 1α (TEF) gene. Also, antifungal susceptibility was tested by broth microdilution according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) and the Etest. Disseminated disease was considered proven in 46 cases and probable in 17 cases. Localised infection was seen in 13 cases. Gibberella fujikuroi species complex (SC), including Fusarium verticillioides and F. proliferatum, and F. solani SC were the most frequent aetiology of disseminated and localised infections, respectively. The crude mortality rate was 46 %, the highest associated with F. solani SC (67 %) and F. proliferatum (62.5 %). A wide range of antifungal susceptibilities was observed. Amphotericin B was the most potent antifungal in vitro, and itraconazole the least effective. The azoles exhibited lower minimum inhibitory concentrations (MICs) against F. verticillioides strains, with posaconazole having a slightly better performance, while F. solani SC isolates were resistant to all three azoles tested. The essential agreement between the Etest and the EUCAST method was 100 % for itraconazole and voriconazole, and 96 % for amphotericin B and posaconazole. In conclusion, we confirm that fusariosis is a rare but severe event in Europe, that G. fujikuroi SC is the predominant cause of deep infections and that different species have different antifungal in vitro susceptibility patterns.     

Clinical breakpoint changes and their impact on surveillance of antimicrobial resistance in Escherichia coli causing bacteraemia

Dutch laboratories are currently changing their breakpoint criteria from mostly Clinical Laboratory and Standards Institute (CLSI) breakpoints to European Committee on Antimicrobial Susceptibility Testing (EUCAST) breakpoints. To evaluate the impact of these changes, we studied antimicrobial resistance trends of Escherichia coli in blood specimens from January 2008 to January 2012 using CLSI and EUCAST breakpoints and compared them with the antimicrobial susceptibility test (AST) interpretations reported by Dutch laboratories participating in the Infectious Disease Surveillance Information System for Antibiotic Resistance (ISIS-AR). ISIS-AR collects AST interpretations, including underlying minimal inhibitory concentrations (MICs) of routinely cultured bacterial species on a monthly basis from Dutch laboratories. MICs of Etests or automated systems were reinterpreted according to the CLSI 2009 and EUCAST 2010 guidelines. Trends in non-susceptibility (i.e. intermediate resistant and resistant) over time were analysed by the Cochran–Armitage test for trend. The effects of the change from CLSI to EUCAST breakpoints on non-susceptibility were small. There were no differences in non-susceptibility to amoxicillin, amoxicillin/clavulanic acid, cefuroxim, gentamicin and co-trimoxazol and only small differences (1–1.5%) for ciprofloxacin between AST interpretations by CLSI or EUCAST. However, for ceftazidime, and cefotaxime/ceftriaxone the proportion of non-susceptibility was substantially higher when EUCAST breakpoints were used (2–3%). The effects on time trends of the change in guidelines were limited, with only substantial differences for the oxymino-cephalosporins. Our study shows that the implementation of EUCAST breakpoints has a limited effect on the proportion of non-susceptible isolates and time trends in E. coli for most, but not all, antimicrobial agents.     

Characterization of a New Clinical Yeast Species,Candida tunisiensis sp. nov., Isolated from a Strain Collection from Tunisian Hospitals

From a collection of yeast isolates isolated from patients in Tunisian hospitals between September 2006 and July 2010, the yeast strain JEY63 (CBS 12513), isolated from a 50-year-old male that suffered from oral thrush, could not be identified to the species level using conventional methods used in clinical laboratories. These methods include matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS), germ tube formation, and the use of CHROMagar Candida and metabolic galleries. Sequence analysis of the nuclear rRNA (18S rRNA, 5.8S rRNA, and 26S rRNA) and internal transcribed spacer regions (ITS1 and ITS2) indicated that the ribosomal DNA sequences of this species were not yet reported. Multiple gene phylogenic analyses suggested that this isolate clustered at the base of the Dipodascaceae (Saccharomycetales, Saccharomycetes, and Ascomycota). JEY63 was named Candida tunisiensis sp. nov. according to several phenotypic criteria and its geographical origin. C. tunisiensis was able to grow at 42°C and does not form chlamydospores and hyphae but could grow as yeast and pseudohyphal forms. C. tunisiensis exhibited most probably a haploid genome with an estimated size of 10 Mb on at least three chromosomes. Using European Committee for Antimicrobial Susceptibility Testing (EUCAST) and Clinical and Laboratory Standards Institute (CLSI) Candida albicans susceptibility breakpoints as a reference, C. tunisiensis was resistant to fluconazole (MIC = 8 μg/ml), voriconazole (MIC = 0.5 μg/ml), itraconazole (MIC = 16 μg/ml), and amphotericin B (MIC = 4 μg/ml) but still susceptible to posaconazole (MIC = 0.008 μg/ml) and caspofungin (MIC = 0.5 μg/ml). In conclusion, MALDI-TOF MS permitted the early selection of an unusual isolate, which was still unreported in molecular databases but could not be unambiguously classified based on phylogenetic approaches.     

Bad Bug,No Test: Tigecycline Susceptibility Testing Challenges and Way Forward

Tigecycline is a reserve antibiotic increasingly used for the treatment of multidrug-resistant bacteria, especially Klebsiella pneumoniae and Acinetobacter baumannii. At present, there are concerns regarding the testing and interpretation of tigecycline susceptibility to bugs such as K. pneumoniae and A. baumannii, which limit clinicians in appropriate usage. Use of appropriate method for testing such as broth microdilution is essential. In addition, tigecycline susceptibility testing is a challenge due to inconsistent results from various antimicrobial susceptibility testing automated platforms. There is a great need to define a suitable methodology along with interpretive criteria, especially for K. pneumoniae and A. baumannii. The European Committee on Antimicrobial Susceptibility Testing (EUCAST) and the Food and Drug Administration (FDA) breakpoints show wide variation and are defined for different set of organisms. Non-species-related pharmacokinetic/pharmacodynamic (PK/PD) breakpoints defined by the EUCAST can be used for organisms such as K. pneumoniae and A. baumannii.     

Susceptibility profile of echinocandins,azoles and amphotericin B against yeast phase of <Emphasis Type="Italic">Talaromyces marneffei</Emphasis> isolated from HIV-infected patients in Guangdong,China

Talaromyces marneffei (T. marneffei) can cause talaromycosis, a fatal systemic mycosis, in patients with AIDS. With the increasing number of talaromycosis cases in Guangdong, China, we aimed to investigate the susceptibility of 189 T. marneffei clinical strains to eight antifungal agents, including three echinocandins (anidulafungin, micafungin, and caspofungin), four azoles (posaconazole, itraconazole, voriconazole, and fluconazole), and amphotericin B, with determining minimal inhibition concentrations (MIC) by Sensititre YeastOne? YO10 assay in the yeast phase. The MICs of anidulafungin, micafungin, caspofungin, posaconazole, itraconazole, voriconazole, fluconazole, and amphotericin B were 2 to >?8 μg/ml, >8 μg/ml, 2 to >?8 μg/ml, ≤?0.008 to 0.06 μg/ml, ≤?0.015 to 0.03 μg/ml, ≤?0.008 to 0.06 μg/ml, 1 to 32 μg/ml, and ≤?0.12 to 1 μg/ml, respectively. The MICs of all echinocandins were very high, while the MICs of posaconazole, itraconazole, and voriconazole, as well as amphotericin B were comparatively low. Notably, fluconazole was found to have a higher MIC than other azoles, and exhibited particularly weak activity against some isolates with MICs over 8 μg/ml. Our data in vitro support the use of amphotericin B, itraconazole, voriconazole, and posaconazole in management of talaromycosis and suggest potential resistance to fluconazole.     

In vitro synergistic effect of minocycline combined with antifungals against Cryptococcus neoformans

BackgroundCryptococcus neoformans infections occur in immunocompromised patients, especially those with HIV infection, chemoradiotherapy after cancer, and organ transplantation. Infection can cause pneumonia and meningoencephalitis in severe cases with a high mortality rate if not treated. Although fluconazole and amphotericin B are the first-line treatments for cryptococcosis, the rate of fluconazole resistance has increased significantly due to long-term use. Minocycline is a derivative of tetracycline that exerts its antibacterial effect through inhibition of bacterial protein synthesis. It is also able to pass the blood-brain barrier to act on the central nervous system. The present study investigates the effects of minocycline in combination with antifungals in treating C. neoformans.ObjectiveTo determine in vitro interactions of minocycline combined with itraconazole, voriconazole, posaconazole, fluconazole and amphotericin B against C. neoformans.MethodsThe minimum inhibitory concentrations (MIC) of the antifungals were determined by the CLSI Clinical and Laboratory Standards Institute M27-A3 microdilution method. The in vitro synergistic effects of minocycline combined with itraconazole, voriconazole, posaconazole, fluconazole, and amphotericin B on C. neoformans were detected by the broth microdilution checkerboard technique and disk diffusion testing.Results and ConclusionThe working concentration ranges were 0.125–4 µg/mL for itraconazole, 0.03–0.125 µg/ml for voriconazole, 0.03–1 µg/ml for posaconazole, 0.25–16 µg/ml for fluconazole, and 0.125–2 µg/ml for amphotericin B. The synergistic rates of minocycline combinations against C. neoformans were 55% with itraconazole, 10% with voriconazole, 85% with posaconazole, 20% with fluconazole, and 70% with amphotericin B. The effective MIC value of minocycline in the synergistic combination decreased to 2–32 µg/ml, while the MIC of itraconazole decreased to 0.03–0.125 µg/ml, voriconazole 0.03–0.125 µg/ml, posaconazole 0.03–0.125 µg/ml, 0.125–4 µg/ml fluconazole, and 0.06–0.50 µg/ml amphotericin B. The disk diffusion assay showed that the plates containing minocycline and antifungal drugs produced inhibition zones with diameters larger than the single drug plates. Minocycline showed no antagonistic effect in the combinations. In conclusion, the combination of minocycline and azoles or amphotericin B has synergistic effects against C. neoformans in vitro.     

Impact of EUCAST ceftaroline breakpoint change on the susceptibility of methicillin-resistant Staphylococcus aureus isolates collected from patients with complicated skin and soft-tissue infections

ObjectivesIn 2018, the European Committee on Antimicrobial Susceptibility Testing (EUCAST) introduced an intermediate breakpoint for ceftaroline against Staphylococcus aureus. The objective of this study was to compare data on resistance to ceftaroline among methicillin-resistant S. aureus (MRSA) isolates using versions 7.1 (March 2017) and 8.0 (January 2018) of the EUCAST breakpoints.MethodsParticipating centers were located in Africa, Asia, Europe, Oceania and South America. Isolates were collected from patients with complicated skin and soft-tissue infections and were cultured from integumentary sources. Methicillin resistance among S. aureus was confirmed locally using the oxacillin method. The CLSI broth microdilution method was used to measure ceftaroline MICs at the central laboratory. Versions 7.1 and 8.0 of the EUCAST breakpoints were used to interpret MIC data.ResultsBetween 2015 and 2016, 9559 isolates of S. aureus were collected, of which 5566 (58.2%) isolates were MRSA. Overall, the lowest rate of MRSA was in Asia (56.5%; 705/1247) and the highest rate was in Oceania (62.7%; 299/477). Using version 7.1 of the EUCAST breakpoints, 4.5% (250/5566) of all MRSA isolates were resistant to ceftaroline and when version 8.0 of the breakpoints was applied, 4.2% (235/5566) of MRSA were in the intermediate category and 0.3% (15/5566) of all isolates were considered resistant.ConclusionsBy applying version 8.0 of the EUCAST breakpoints, the majority of MRSA isolates that were resistant are now in the intermediate category for ceftaroline. Ceftaroline resistance among MRSA now appears rare.     

Harmonization of antimicrobial breakpoints in Europe — Can it be achieved?

The European Committee on Antimicrobial Susceptibility Testing (EUCAST) isconvened by the European Society of Clinical Microbiology and Infectious Diseases (ESCMID) and supported by representatives of almost all European countries. It is financed by ESCMID, the European Union, and the national breakpoint committees of France, Germany, Norway, Sweden, the Netherlands, and the United Kingdom. The Committee has recently published harmonized European breakpoints for aminoglycosides, fluoroquinolones, glycopeptides, and linezolid and is currently addressing aztreonam, carbapenems and cephalosporins. EUCAST has recognized the inconsistencies between clinical breakpoints primarily aimed at predicting better (susceptible) versus worse (resistant) outcome and epidemiological cutoff values for early detection of antimicrobial resistance development. EUCAST clinical breakpoints are based primarily on pharmacokinetic-pharmacodynamic relationships but do take into account other factors, such as differences in dosing regimens, toxicology, resistance mechanisms, clinical outcome data, and wild-type MIC distributions. EUCAST has devised a system for collecting MIC distributions of wild-type bacteria and for setting epidemiological cutoff values. The output of EUCAST is freely available via the EUCAST website ().     

Comparison of the broth microdilution methods of the European Committee on Antimicrobial Susceptibility Testing and the Clinical and Laboratory Standards Institute for testing itraconazole, posaconazole, and voriconazole against Aspergillus isolates

We compared EUCAST and CLSI antifungal susceptibility testing methods for itraconazole, posaconazole, and voriconazole by testing 245 Aspergillus clinical isolates. The essential agreement (EA) between methods was excellent: 100% (itraconazole), 98.4% (posaconazole), and 99.6% (voriconazole) assessing EA at ±2 dilutions and 99.6% (itraconazole), 87.7% (posaconazole), and 96.3% (voriconazole) at ±1 dilution.