scholarly journals Comparison of Vitek® 2, three different gradient strip tests and broth microdilution for detecting vanB-positive Enterococcus faecium isolates with low vancomycin MICs

2019 ◽  
Author(s):  
Ingo Klare ◽  
Jennifer K. Bender ◽  
Carola Fleige ◽  
Nancy Kriebel ◽  
Axel Hamprecht ◽  
...  
Keyword(s):  
Agar Medium ◽  
Standard Procedure ◽  
Vancomycin Resistance ◽  
Broth Microdilution ◽  
Minimal Inhibitory Concentrations ◽  
Specific Pcr ◽  
Vancomycin Mics ◽  
Confirmatory Tests ◽  
Improved Performance ◽  
Strip Test

AbstractObjectivesIn 2018, EUCAST issued a warning regarding unreliable results of gradient strip tests for confirming vancomycin resistance in enterococci. We compared the performance of various diagnostic standard and confirmatory tests to identify and determine vanB-type vancomycin resistance.MethodsWe analysed a collection of vanB-positive E. faecium isolates (n = 68) with low level vancomycin minimal inhibitory concentrations (MICs) and compared the performance of Vitek® 2 (bioMérieux), of broth microdilution and of three different gradient strip test providers (Oxoid, Liofilchem, bioMérieux). For the latter we compared the standard procedure vs. a protocol with increased inoculum, a rich agar medium and a longer incubation time (“macromethod”).ResultsThe sensitivity of Vitek® 2 was 81% of vanB-isolates compared to 72% for broth microdilution and 61 – 63% for the three gradient strip tests using standard conditions. The “macromethod” substantially improved performance of all strip tests resulting in a sensitivity of 89 – 96% at 48h readout.ConclusionsThe “macromethod” provided the overall best performance for recognition of vanB E. faecium. We therefore propose to adapt the EUCAST warning and to recommend the “macromethod” with an additional 48h readout or a confirmation by a vanB-specific PCR from culture.

scholarly journals Comparison of VITEK® 2, three different gradient strip tests and broth microdilution for detecting vanB-positive Enterococcus faecium isolates with low vancomycin MICs

2019 ◽  
Vol 74 (10) ◽  
pp. 2926-2929 ◽  
Author(s):  
Ingo Klare ◽  
Jennifer K Bender ◽  
Carola Fleige ◽  
Nancy Kriebel ◽  
Axel Hamprecht ◽  
...  
Keyword(s):  
Incubation Time ◽  
Enterococcus Faecium ◽  
Agar Medium ◽  
Standard Procedure ◽  
Vancomycin Resistance ◽  
Therapeutic Decision ◽  
Molecular Test ◽  
Broth Microdilution ◽  
Vancomycin Mics ◽  
Confirmatory Tests

Abstract Objectives In 2018, EUCAST issued a warning regarding unreliable results of gradient strip tests for confirming vancomycin resistance in enterococci. We compared the performance of various diagnostic standard and confirmatory tests to identify and determine vanB-type vancomycin resistance. Methods We analysed a collection of vanB-positive Enterococcus faecium isolates (n = 68) with low vancomycin MICs and compared the performance of VITEK® 2 (bioMérieux), broth microdilution and three gradient strip tests from different providers (Oxoid, Liofilchem and bioMérieux). For the latter we compared the standard procedure with a protocol with increased inoculum, a rich agar medium and a longer incubation time (‘macromethod’). Results The sensitivity of VITEK® 2 was 81% compared with 72% for broth microdilution and 61%–63% for the three gradient strip tests using standard conditions. The macromethod substantially improved the performance of all strip tests resulting in a sensitivity of 89%–96% after 48 h of incubation. Conclusions We recommend that EUCAST changes the present warning against the general use of MIC strips. When MIC strips are used to either exclude or confirm suspected vancomycin resistance in E. faecium, and a PCR is not available, the macromethod should be employed. For clinically relevant enterococci, where a rapid therapeutic decision is needed, a molecular test (e.g. PCR) should be favoured in order to save time and to further increase sensitivity.

Evaluation of methods for detection of β-lactamase production in MSSA

2021 ◽  
Author(s):  
Robert Skov ◽  
David R Lonsway ◽  
Jesper Larsen ◽  
Anders Rhod Larsen ◽  
Jurgita Samulioniené ◽  
...  
Keyword(s):  
Susceptibility Testing ◽  
Confirmatory Test ◽  
Disc Diffusion ◽  
Broth Microdilution ◽  
Confirmatory Tests ◽  
Correct Determination ◽  
Primary Test ◽  
Zone Edge ◽  
Evaluation Of Methods

Abstract Objectives Correct determination of penicillin susceptibility is pivotal for using penicillin in the treatment of Staphylococcus aureus infections. This study examines the performance of MIC determination, disc diffusion and a range of confirmatory tests for detection of penicillin susceptibility in S. aureus. Methods A total of 286 consecutive penicillin-susceptible S. aureus blood culture isolates as well as a challenge set of 62 MSSA isolates were investigated for the presence of the blaZ gene by PCR and subjected to penicillin-susceptibility testing using broth microdilution MIC determination, disc diffusion including reading of the zone edge, two nitrocefin tests and the cloverleaf test. Results Using PCR-based detection of blaZ as the gold standard, both broth microdilution MIC testing and disc diffusion testing resulted in a relatively low accuracy (82%–93%) with a sensitivity ranging from 49%–93%. Among the confirmatory tests, the cloverleaf test performed with 100% accuracy, while zone edge interpretation and nitrocefin-based tests increased the sensitivity of β-lactamase detection to 96%–98% and 82%–96% when using MIC determination or disc diffusion as primary test, respectively. Conclusions This investigation showed that reliable and accurate detection of β-lactamase production in S. aureus can be obtained by MIC determination or penicillin disc diffusion followed by interpretation of the zone edge as a confirmatory test for apparently penicillin-susceptible isolates. The more cumbersome cloverleaf test can also be used. Nitrocefin-based tests should not be used as the only test for confirmation of a presumptive β-lactamase-negative isolate.

scholarly journals High-level vancomycin-resistant enterococci causing hospital infections

1989 ◽  
Vol 103 (1) ◽  
pp. 173-181 ◽  
Author(s):  
A. H. C. Uttley ◽  
R. C. George ◽  
J. Naidoo ◽  
N. Woodford ◽  
A. P. Johnson ◽  
...  
Keyword(s):  
Plasmid Dna ◽  
Vancomycin Resistance ◽  
Recipient Strain ◽  
Hospital Infections ◽  
Minimal Inhibitory Concentrations ◽  
Vancomycin Resistant Enterococci ◽  
Vancomycin Resistant ◽  
Many Sources ◽  
High Level ◽  
Level Resistance

SUMMARYNosocomial infection or colonization due to enterococci with high-level resistance to vancomycin (minimal inhibitory concentrations [MICs] between 64 and > 2000 mg/L) has occurred in 41 patients with renal disease. These vancomycin-resistant enterococci were cultured from many sources including blood. All but one strain contained one or more plasmids ranging in molecular weight from 1·0 to 40 Megadaltons (MDa). Vancomycin resistance was transferable by conjugation to a susceptible recipient strain ofEnterococcus faecalisbut this was not always associated with plasmid DNA. The emergence of transferable high-level vancomycin resistance in enterococci causing significant clinical infections is of particular importance since vancomycin is widely regarded as a reserve drug for the management of infections with multi-resistant Gram-positive organisms.

scholarly journals Evaluation of the Accelerate Pheno™ system to identify bacteria and determine antimicrobial susceptibility in positive blood cultures

2019 ◽  
Author(s):  
Mariana J. Fernandez-Pittol ◽  
Javier Morales ◽  
Elisa Rubio ◽  
Assumpta Fasanella ◽  
Izaskun Alejo-Cancho ◽  
...  
Keyword(s):  
Antimicrobial Susceptibility ◽  
Antibiotic Susceptibility ◽  
Susceptibility Testing ◽  
Standard Procedure ◽  
Bloodstream Infections ◽  
Blood Cultures ◽  
Bacterial Isolates ◽  
Minimal Inhibitory Concentrations ◽  
Maldi Tof ◽  
The Mean

AbstractIntroductionNew platforms have recently been developed to reduce response time of identification and antimicrobial susceptibility of bacterial isolates in positive blood cultures from patients with bloodstream infections. The Accelerate Pheno™ system (Accelerate Diagnostics, Inc.) provides information on pathogen identification and antibiotic susceptibility in approximately 1.5 and 7 hours, respectively.MethodsIn this study we compared the Accelerate Pheno™ system with the standard procedure used in our laboratory. A total of 41 blood cultures were prospectively analysed with the Accelerate Pheno™ system and our standard methods, which include identification by MALDI-TOF mass spectrometry and antibiotic susceptibility testing (AST) by BD Phoenix system and E-test.ResultsThe correlation between the two methods using Cohen’s kappa coefficient was 0.82; mean (sd) time of identification for MALDI-TOF MS was 0.7 (0.22) hours and 1.43 (0.14) hours for the Accelerate Pheno™ system. The mean (sd) time of AST with the BD Phoenix system was 15.85 (2.57) hours and with the Accelerate Pheno™ system 6.7 (0.12) hours. AST results showed an overall essential agreement of 92% for the minimal inhibitory concentrations (MIC) and an overall category agreement of 96%. Among Gram positive isolates, essential and category agreements of 100% were observed. In Gram negative isolates 10 discrepancies were detected, which were classified as 7 major and 3 minor errors. Discrepancies in the Accelerate Pheno™ system were observed particularly for P. aeruginosa.ConclusionThe Accelerate Pheno™ system can improve turn-around time in the management of patients with bloodstream infections.

scholarly journals Manual Reading of Sensititre™ Broth Microdilution System Panels Improves Accuracy of Susceptibility Reporting for Polymyxin Antibiotics

2021 ◽  
Author(s):  
Michelle M. Bellerose ◽  
Andrew E. Clark ◽  
Jung-Ho Youn ◽  
Rebecca A. Weingarten ◽  
Chelsea M. Crooks ◽  
...  
Keyword(s):  
Bacterial Infections ◽  
Microbial Growth ◽  
Susceptibility Testing ◽  
Polymyxin B ◽  
Multidrug Resistant ◽  
Error Rates ◽  
Broth Microdilution ◽  
Minimal Inhibitory Concentrations ◽  
Manual Inspection ◽  
Agar Dilution

Accurate and reproducible antimicrobial susceptibility testing (AST) of polymyxin antibiotics is critical, as these drugs are last-line therapeutic options for the treatment of multidrug-resistant Gram-negative bacterial infections. However, polymyxin AST in the routine laboratory remains challenging. In this study, we evaluated the performance of an automated broth microdilution (BMD) system (Sensititre™, ThermoFisher) compared to agar dilution (AD) for colistin and polymyxin B AST of 129 Enterobacterales, Pseudomonas aeruginosa and Acinetobacter baumannii complex clinical isolates. Minimal inhibitory concentrations (MICs) derived from the Sensititre™ instrument based on two operator comparisons demonstrated overall categorical agreement (CA) of 86% and 89% compared to AD for colistin and 89% and 92% compared to AD for polymyxin B. However, error rates were higher than recommended by CLSI. Manual inspection of microdilution wells revealed microbial growth and skip wells which were erroneously interpreted by the Aris™2X instrument. Using manually interpreted BMD MICs read by two operators increased the overall categorical agreements to 88% and 95% compared to AD for colistin and 92% and 96% compared to AD for polymyxin B. Laboratories choosing to use the Sensititre™ platform for polymyxin AST should consider manual evaluation of wells as part of their algorithm.

scholarly journals Colistin resistance in Gram-negative bacteria analysed by five phenotypic assays and inference of the underlying genomic mechanisms

2021 ◽  
Author(s):  
Diana Albertos Torres ◽  
Helena M.B. Seth-Smith ◽  
Nicole Jossee ◽  
Claudia Lang ◽  
Oliver Dubius ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Test Strip ◽  
Clinical Samples ◽  
Broth Microdilution ◽  
Colistin Resistance ◽  
Minimal Inhibitory Concentrations ◽  
Chromosomal Mutation ◽  
Susceptibility Data ◽  
Antimicrobial Resistance Genes ◽  
Vitek 2

Abstract Background Colistin is used against multi-drug resistant pathogens, yet resistance emerges either through dissemination of plasmid-mediated genes (mcr) or chromosomal mutation of genes involved in lipopolysaccharide synthesis (i.e. mgrB, phoPQ, pmrCAB). Phenotypic susceptibility testing is challenging due to poor diffusion of colistin in agar media, leading to an underestimation of resistance. We aimed to compare the performance of four different phenotypic approaches in the context of different molecular mechanism of resistance. Methods We compared the performance of Vitek 2® (bioMérieux, AST N242), Colistin MIC Test Strip (Liofilchem Diagnostici), UMIC (Biocentric), and Rapid Polymyxin NP Test (ELITechGroup) against the standard broth microdilution (BDM) method. We used whole genome sequencing (WGS) to infer the molecular mechanisms of resistance. A total of 97 Enterobacterales and non-fermenting bacterial isolates were collected from clinical samples during 2016–2017 and tested for colistin susceptibility. Data was analysed by comparing the susceptibility category (susceptible or resistant) and minimal inhibitory concentrations (MIC). We determined diversity of isolates by core genome multi locus sequencing typing (cgMLST) and identified antimicrobial resistance genes using NCBI and CARD databases. Results Of the 97 clinical isolates, 54 (56%) were resistant by the standard broth microdilution. The highest susceptibility category concordance was achieved by the Rapid Polymyxin NP test (98.8%) followed by UMIC (97.9%), E-test MIC strip (96.9%) and Vitek 2 (95.6%). The highest MIC concordance was achieved by UMIC (80.4%), followed by Vitek 2 (72.5%) and E-test MIC strip (62.9%). Among the resistant isolates, 23/54 (43%) were intrinsically resistant to colistin, whereas 31/54 (57%) isolates had acquired colistin resistance. Of these, mcr-1 was detected in four isolates and mcr-2 in one isolate. Mutations in mgrB, phoQ, pmrA, pmrB, and pmrC genes that led to amino acid changes were encountered in Klebsiella pneumoniae, Escherichia coli, and Acinetobacter bereziniae resistant isolates. Conclusions The Rapid Polymyxin NP test showed the highest categorical concordance and the UMIC test provided MIC values with a high concordance to the standard method. We found colistin resistance in diverse species occurred predominantly through spontaneous chromosomal mutation rather than plasmid-mediated resistance.

scholarly journals Glycopeptide Susceptibility Profiles ofStaphylococcus haemolyticus Bloodstream Isolates

2000 ◽  
Vol 44 (11) ◽  
pp. 3122-3126 ◽  
Author(s):  
Francesca Biavasco ◽  
Carla Vignaroli ◽  
Raffaella Lazzarini ◽  
Pietro E. Varaldo
Keyword(s):  
Vancomycin Resistance ◽  
Blood Cultures ◽  
Methicillin Resistant ◽  
Staphylococcal Species ◽  
Clinical Strains ◽  
Staphylococcus Haemolyticus ◽  
Vancomycin Mics ◽  
Susceptibility Profiles

ABSTRACT Twelve clinical strains of Staphylococcus haemolyticus(eight methicillin resistant and three methicillin susceptible), isolated from blood cultures between 1982 and 1997, were investigated for teicoplanin and vancomycin susceptibility profiles. On the basis of conventional MIC tests and breakpoints, four isolates were susceptible (MICs, 1 to 8 μg/ml) and eight were resistant (MICs, 32 to 64 μg/ml) to teicoplanin while all were susceptible to vancomycin (MICs, 1 to 2 μg/ml). All four strains for which the conventional teicoplanin MICs were within the range of susceptibility expressed heterogeneous resistance to teicoplanin and homogeneous vancomycin susceptibility. Of the eight strains for which the conventional teicoplanin MICs were within the range of resistance, six expressed heterogeneous and two expressed homogeneous teicoplanin resistance while seven showed heterogeneous vancomycin resistance profiles (with subpopulations growing on 8 μg of the drug per ml at frequencies of ≥10−6 for six strains and 10−7 for one) and one demonstrated homogeneous vancomycin susceptibility. Of six bloodstream isolates of other staphylococcal species (S. aureus, S. epidermidis, and S. simulans), for all of which the conventional teicoplanin MICs were ≥4 μg/ml and the vancomycin MICs were ≤2 μg/ml, none exhibited heterogeneous susceptibility profiles for teicoplanin while three showed homogeneous and three showed heterogeneous susceptibility profiles for vancomycin (with subpopulations growing on 8 μg of the drug per ml found for only one strain). The results of this study indicate that a heterogeneous response to glycopeptides is a common feature of S. haemolyticus isolates and suggest that susceptibility to glycopeptides as determined by conventional MIC tests may not be predictive of the outcome of glycopeptide therapy.

scholarly journals Performance of Vitek 2 for Antimicrobial Susceptibility Testing of Acinetobacter baumannii, Pseudomonas aeruginosa, and Stenotrophomonas maltophilia with Vitek 2 (2009 FDA) and CLSI M100S 26th Edition Breakpoints

2016 ◽  
Vol 55 (2) ◽  
pp. 450-456 ◽  
Author(s):  
April M. Bobenchik ◽  
Eszter Deak ◽  
Janet A. Hindler ◽  
Carmen L. Charlton ◽  
Romney M. Humphries
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Acinetobacter Baumannii ◽  
Antimicrobial Susceptibility ◽  
Stenotrophomonas Maltophilia ◽  
Susceptibility Testing ◽  
Antimicrobial Susceptibility Testing ◽  
Broth Microdilution ◽  
Content Type ◽  
Vitek 2 ◽  
Improved Performance

ABSTRACTThe performances of Vitek 2 AST-GN69 and AST-XN06 cards were compared to Clinical and Laboratory Standards Institute (CLSI) reference broth microdilution (BMD) for 99 isolates ofPseudomonas aeruginosa, 26Acinetobacter baumanniiisolates, and 11Stenotrophomonas maltophiliaisolates. In total, 15 antimicrobials were evaluated, with 11 forP. aeruginosa, 14 forA. baumannii, and 2 forS. maltophilia. Categorical agreement (CA) was assessed using both Vitek 2 breakpoints and 2016 CLSI M100S 26th edition breakpoints. The essential agreement values forP. aeruginosa,A. baumannii, andS. maltophiliawere 99.5%, 99.2%, and 100%, respectively. The CA values forP. aeruginosa,A. baumannii, andS. maltophiliawere 94.1%, 92.7%, and 95.5%, respectively, by the Vitek 2 breakpoints, and 93.4%, 92.3%, and 95.5%, respectively, by the CLSI breakpoints. Overall, the Vitek 2 performance was comparable to that of BMD using both Vitek 2 breakpoints and 2016 CLSI M100S 26th edition breakpoints. Improved performance was noted for the reformulated piperacillin-tazobactam and imipenem found on the AST-GN69 card, with no very major or major errors noted when using the CLSI breakpoints.

scholarly journals Comparison of Two Commercial Colorimetric Broth Microdilution Tests for Candida Susceptibility Testing: Sensititre YeastOne versus MICRONAUT-AM

2021 ◽  
Vol 7 (5) ◽  
pp. 356
Author(s):  
Sophie Philips ◽  
Frederik Van Van Hoecke ◽  
Emmanuel De De Laere ◽  
Steven Vervaeke ◽  
Roos De De Smedt ◽  
...  
Keyword(s):  
Amphotericin B ◽  
Antifungal Agents ◽  
Susceptibility Testing ◽  
Antifungal Susceptibility ◽  
Reference Method ◽  
Daily Practice ◽  
Broth Microdilution ◽  
Minimal Inhibitory Concentrations ◽  
Clinical Breakpoints ◽  
Susceptibility Tests

Two colorimetric broth microdilution antifungal susceptibility tests were compared, Sensititre YeastOne and MICRONAUT-AM for nine antifungal agents. One hundred clinical Candida isolates were tested, representing a realistic population for susceptibility testing in daily practice. The reproducibility characteristics were comparable. Only for fluconazole, caspofungin, 5-flucytosine and amphotericin B, an essential agreement of ≥90% could be demonstrated. Sensititre minimal inhibitory concentrations (MICs) were systematically higher than MICRONAUT MICs for all antifungals, except for itraconazole. CLSI clinical breakpoints (CBPs) and epidemiological cut-off values (ECVs) were used for Sensititre MICs while for MICRONAUT the EUCAST CBPs and ECVs were used. Only fluconazole, micafungin, and amphotericin B had a categorical agreement of ≥90%. For fluconazole, micafungin, and amphotericin B the susceptibility proportions were comparable. Susceptibility proportion of posaconazole and voriconazole was higher using the MICRONAUT system. For itraconazole and anidulafungin, the susceptibility proportion was higher using Sensititre. It was not possible to determine the true MIC values or the correctness of a S/I/R result since both commercial systems were validated against a different reference method. These findings show that there is a significant variability in susceptibility pattern and consequently on use of antifungals in daily practice, depending on the choice of commercial system.

scholarly journals First Report of Mango Malformation Disease Caused by Fusarium mangiferae in Spain

Plant Disease ◽  
2012 ◽  
Vol 96 (2) ◽  
pp. 286-286 ◽  
Author(s):  
M. Crespo ◽  
F. M. Cazorla ◽  
J. M. Hermoso ◽  
E. Guirado ◽  
M. Maymon ◽  
...  
Keyword(s):  
Agar Medium ◽  
Economic Losses ◽  
Conidial Suspension ◽  
Control Strain ◽  
Potato Dextrose Agar Medium ◽  
Floral Tissue ◽  
First Report ◽  
Specific Pcr ◽  
Mango Malformation ◽  
Fusarium Mangiferae

Mango (Mangifera indica L.) malformation disease (MMD) is one of the most important diseases affecting this crop worldwide, which causes severe economic losses because of the reduction of productivity. Symptoms of MMD in Spain were observed for the first time in April of 2006 in three mango orchards in the Axarquia Region (southern Spain). Symptoms included an abnormal development of vegetative shoots with shortened internodes and dwarfed leaves and hypertrophied short and thickened panicles. In the years of 2006, 2009, and 2010, isolates of Fusarium were obtained from vegetative shoots and floral tissue of symptomatic mango trees from 21 different orchards of cvs. Keitt, Kent, Osteen, Tommy Atkins, and a variety of minor commercial cultivars, all showing typical symptoms of MMD. Different Fusarium-like strains were isolated from infected tissues. Colonies from single-spored isolates possessed dark purple-to-salmon-colored mycelium when grown on potato dextrose agar medium. On fresh carnation leaf agar medium, mycelium contained aerial conidiophores possessing three- to five-celled macroconidia and abundant microconidia in false heads from mono- and polyphialides; while cream-orange-colored sporodochia were produced on the surface of the medium, typical for Fusarium mangiferae. The identification of 37 isolates was confirmed as F. mangiferae by species-specific PCR analysis with the primer pair 1-3 F/R that amplified a 608-bp DNA fragment from all Spanish isolates as well as a representative Israeli control strain, Fus 34, also designated as MRC7560 (2). Pathogenicity using four representative isolates, UMAF F02, UMAF F10, UMAF F17, and UMAF F38 of F. mangiferae from Spain as well as isolate MRC7560, was tested on 2-year-old healthy mango seedlings cv. Keitt by inoculating 15 buds from three different trees with a 20-μl conidial suspension (5 × 107 conidia per ml) per isolate (1). This experiment was conducted twice with two independent sets of plants and at different times (March and November 2010). Typical mango malformation symptoms were detected after bud break in March 2011, 5 and 12 months after inoculation. Symptoms were observed for 60% of the inoculated buds with the four F. mangiferae Spanish isolates and 75% with the MRC7560 control strain, but not with water-inoculated control plants. Recovered isolates from the infected floral and vegetative malformed buds were identical morphologically to those inoculated, and the specific 608-bp fragment described for F. mangiferae was amplified with specific-PCR, thus fulfilling Koch's postulates. To our knowledge, this is the first report of mango malformation disease caused by F. mangiferae in Spain and Europe. References: (1) S. Freeman et al. Phytopathology 89:456, 1999. (2) Q. I. Zheng and R. C. Ploetz. Plant Pathol. 51:208, 2002.

Sign in / Sign up

Export Citation Format

Share Document