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.

Antimicrobial Susceptibility Testing for Staphylococcus lugdunensis

2021 ◽  
Author(s):  
Joanne S.K. Teh ◽  
Ioanna Pantelis ◽  
Xiao Chen ◽  
Tania Sadlon ◽  
Kelly Papanaoum ◽  
...  
Keyword(s):  
Antimicrobial Susceptibility ◽  
Susceptibility Testing ◽  
Resistant Isolate ◽  
Disc Diffusion ◽  
Broth Microdilution ◽  
Major Error ◽  
Staphylococcus Lugdunensis ◽  
Zone Diameter ◽  
Oxacillin Resistance ◽  
Zone Edge

Evaluation of penicillin and oxacillin susceptibility testing was conducted on two hundred Staphylococcus lugdunensis isolates. Disc diffusion with penicillin 1 IU (P1, EUCAST) and penicillin 10 IU (P10, CLSI) was compared with nitrocefin discs (Cefinase®) and automated broth microdilution (Vitek2®). Oxacillin susceptibility was extrapolated from cefoxitin 30μg disc diffusion (FOX) and compared with Vitek2®. Reference methods were blaZ and mecA PCR. Penicillin zone diameter and zone edge correlated with blaZ in all except two P10 susceptible isolates (VME; very major error) and one P1 resistant isolate (ME). One hundred and forty-eight isolates were blaZ -negative of which one hundred and forty-six and one hundred and forty-nine isolates were susceptible by P1 and P10 respectively. One hundred and twenty-seven isolates were penicillin susceptible by Vitek2®. Vitek2® overcalled resistance in twenty-one blaZ -negative, twenty P1 and twenty-two P10 susceptible isolates (Vitek2® ME rate, 14.2%). Two mecA -positive isolates were oxacillin resistant by FOX and Vitek2® (categorical agreement). However, eighteen FOX susceptible, mecA -negative isolates tested resistant by Vitek2®. In conclusion, Vitek2® over-estimated penicillin and oxacillin resistance compared with disc diffusion and PCR. Disc diffusion with zone edge interpretation was more accurate and specific than automated broth microdilution for S. lugdunensis in our study.

scholarly journals Comparison of disc diffusion, E-Test and a modified CLSI broth microdilution method for in vitro susceptibility testing of itraconazole, posaconazole and voriconazole against Madurella mycetomatis

2021 ◽  
Author(s):  
Bertrand Nyuykonge ◽  
Lukas van Amelsvoort ◽  
Kimberly Eadie ◽  
Ahmed H. Fahal ◽  
Annelies Verbon ◽  
...  
Keyword(s):  
Low Income ◽  
Susceptibility Testing ◽  
Fungal Infections ◽  
Disc Diffusion ◽  
Broth Microdilution ◽  
In Vitro Susceptibility ◽  
Bead Beating ◽  
Broth Microdilution Method ◽  
Madurella Mycetomatis

For many fungal infections, in vitro susceptibility testing is used to predict if an isolate is resistant or susceptible to the antifungal agent used to treat the fungal infection. For Madurella mycetomatis , the main causative agent of mycetoma, in vitro susceptibility testing currently is not performed on a routine basis. The current in vitro susceptibility testing method is labor intensive and sonication must be done to generate a hyphal inoculum. For endpoint visualization, expensive viability dyes are needed. Here we investigated if the currently used in vitro susceptibility method could be adapted to make it amendable for use in a routine setting which can be used in low income countries, where mycetoma is endemic. First, we developed a methodology in which hyphal fragments can be generated without the need for sonication, by comparing different bead beating methodologies. Next, in vitro susceptibility was assessed using standard broth microdilution assays as well as disc diffusion, E-testing and VIPcheck™ methodologies. We demonstrate that after a hyphal suspension is generated by glass bead beating, disc diffusion, E-testing and VIPcheck™ can be used to determine susceptibility towards itraconazole, posaconazole and voriconazole of Madurella mycetomatis . The MICs found with the E-test were comparable to those obtained with our modified CLSI-based broth microdilution in vitro susceptibility assay for itraconazole and posaconazole. Furthermore, we found an inverse relationship between the zone of inhibition and MIC obtained with E-test and the modified CLSI broth microdilution technique.

scholarly journals Evaluation of penicillin G susceptibility testing methods for Staphylococcus lugdunensis

2020 ◽  
Vol 75 (5) ◽  
pp. 1206-1211
Author(s):  
Malin Hagstrand Aldman ◽  
Lisa I Påhlman
Keyword(s):  
Susceptibility Testing ◽  
Reference Method ◽  
Penicillin G ◽  
Diffusion Method ◽  
Disc Diffusion ◽  
Diffusion Test ◽  
Staphylococcus Lugdunensis ◽  
Susceptible Isolate ◽  
Focus Of Infection ◽  
Zone Edge

Abstract Background Staphylococcus lugdunensis belongs to the CoNS group, but is regarded to be more virulent than most other CoNS. It is also remarkably susceptible to antibiotics, including penicillin G. Objectives To evaluate different methods for penicillin susceptibility testing, to assess penicillin susceptibility rates among S. lugdunensis and to describe the clinical presentation including antibiotic treatment. Methods Clinical isolates of S. lugdunensis were tested for penicillin susceptibility using disc diffusion according to CLSI (10 U disc) and EUCAST (1 U disc), assessment of zone-edge appearance, nitrocefin test and Etest for MIC determination. PCR of the blaZ gene was used as a reference method. Results Of the 112 isolates included in the study, 67% were susceptible to penicillin G according to blaZ PCR. The EUCAST disc diffusion test had 100% sensitivity, whereas the CLSI method had one very major error with a false-susceptible isolate. When zone-edge appearance was included in the assessment, the false-susceptible isolate was correctly classified as resistant. Foreign-body infection was the most common focus of infection, affecting 49% of the participants. Only 4% of the patients were treated with penicillin G. Conclusions Penicillin susceptibility is common in S. lugdunensis and the disc diffusion method according to EUCAST had a higher sensitivity than that of CLSI. Assessment of zone-edge appearance could increase the sensitivity of the disc diffusion test. Penicillin susceptibility testing and treatment should be considered in S. lugdunensis infections.

scholarly journals Comprehensive Statistical Evaluation of Etest®, UMIC®, MicroScan and Disc Diffusion versus Standard Broth Microdilution: Workflow for an Accurate Detection of Colistin-Resistant and Mcr-Positive E. coli

Antibiotics ◽  
2020 ◽  
Vol 9 (12) ◽  
pp. 861
Author(s):  
Isidro García-Meniño ◽  
Pilar Lumbreras ◽  
Pablo Valledor ◽  
Dafne Díaz-Jiménez ◽  
Luz Lestón ◽  
...  
Keyword(s):  
Susceptibility Testing ◽  
Statistical Evaluation ◽  
Disc Diffusion ◽  
Broth Microdilution ◽  
Colistin Resistance ◽  
Testing Methods ◽  
Disc Diffusion Assay ◽  
E Coli ◽  
Diffusion Assay ◽  
Microscan System

Four colistin susceptibility testing methods were compared with the standard broth microdilution (BMD) in a collection of 75 colistin-susceptible and 75 mcr-positive E. coli, including ST131 isolates. Taking BMD as reference, all methods showed similar categorical agreement rates (CA) of circa 90%, and a low number of very major errors (VME) (0% for the MicroScan system and Etest®, 0.7% for UMIC®), except for the disc diffusion assay (breakpoint ≤ 11 mm), which yielded false-susceptible results for 8% of isolates. Of note is the number of mcr-positive isolates (17.3%) categorized as susceptible (≤2 mg/L) by the BMD method, but as resistant by the MicroScan system. ST131 mcr-positive E. coli were identified as colistin-resistant by all MIC-based methods. Our results show that applying the current clinical cut-off (>2 mg/L), many mcr-positive E. coli remain undetected, while applying a threshold of >1 mg/L the sensitivity of detection increases significantly without loss of specificity. We propose two possible workflows, both starting with the MicroScan system, since it is automated and, importantly, it categorized all mcr-positive isolates as colistin-resistant. MicroScan should be followed by either BMD or MIC-based commercial methods for colistin resistance detection; or, alternatively, MicroScan, followed by PCR for the mcr screening.

In vitro activity of ampicillin and ceftriaxone against ampicillin-susceptible Enterococcus faecium

2019 ◽  
Vol 74 (8) ◽  
pp. 2269-2273 ◽  
Author(s):  
Michael P Lorenzo ◽  
James M Kidd ◽  
Stephen G Jenkins ◽  
David P Nicolau ◽  
Seth T Housman
Keyword(s):  
Enterococcus Faecium ◽  
Susceptibility Testing ◽  
Diffusion Time ◽  
Disc Diffusion ◽  
In Vitro Activity ◽  
Broth Microdilution ◽  
Gradient Diffusion ◽  
The Usa ◽  
Time Kill

AbstractObjectivesTo assess activity of the combination of ceftriaxone and ampicillin against clinical isolates of ampicillin-susceptible Enterococcus faecium.MethodsAmpicillin-susceptible E. faecium (n = 29) and Enterococcus faecalis (n = 10) collected from locations in the USA and France were used for this analysis. Susceptibility testing was performed by gradient diffusion strip (GDS) and broth microdilution (BMD). Synergy with the combination of ceftriaxone and ampicillin was assessed in all isolates using GDS crossing and double disc diffusion methods. Selected isolates (nine E. faecium and three E. faecalis) were assessed for synergy in time–kill studies using ampicillin alone and in combination with ceftriaxone.ResultsIn isolates of E. faecium, the median (range) ampicillin MIC by BMD was 0.5 (0.25–4) mg/L and by GDS it was 2 (1–8)  mg/L. In E. faecalis, the median (range) ampicillin MIC by BMD was 0.5 (0.5–1) mg/L and by GDS it was 2 (0.75–3) mg/L. A total of 24/29 (82.8%) isolates of E. faecium displayed synergy by GDS and 22/29 (75.9%) by double disc diffusion. Seven of 10 (70%) isolates of E. faecalis displayed synergy by GDS and 4/10 (40%) by double disc diffusion. Time–kill studies found synergy in 3/9 (33.3%) E. faecium and 3/3 (100%) E. faecalis.ConclusionsIn contrast to the demonstrated synergy in time–kill models of ceftriaxone and ampicillin for E. faecalis, this combination does not appear to provide uniform synergy in E. faecium. Antagonism was not observed. Clinical correlation is necessary and caution should be used when considering ampicillin and ceftriaxone for the treatment of infections caused by ampicillin-susceptible E. faecium.

scholarly journals Determination of adrenal hypersecretion in primary Aldosteronism without aldosterone-production adenomas

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Fang Sun ◽  
Yangning Hong ◽  
Hexuan Zhang ◽  
Xiaoli Liu ◽  
Zhigang Zhao ◽  
...  
Keyword(s):  
Primary Aldosteronism ◽  
Metabolic Diseases ◽  
Heart Diseases ◽  
Confirmatory Test ◽  
Plasma Aldosterone Concentration ◽  
Aldosterone Production ◽  
Confirmatory Tests ◽  
Biochemical Evaluation ◽  
Aldosterone To Renin Ratio

Abstract Background Primary aldosteronism (PA) is highly prevalent in hypertensive population. Adrenal vein sampling (AVS) is the only procedure to assess adrenal aldosterone hypersecretion in PA. PA patients without aldosterone-producing adenomas (APA) frequently have unilateral aldosterone hypersecretion (UAH). These patients could bear inappropriate adrenalectomy without AVS. This study aims to identify which clinical characteristics should be recommended to perform AVS in these PA patients. Methods This study was performed from January 2018 to July 2019 at a center for hypertension and metabolic diseases. Adrenal computed tomography (CT) scan, biochemical evaluation, and AVS were performed. Results Total 141 patients were included in this study. Aldosterone to renin ratio (ARR) after confirmatory test is highly associated with adrenal laterality. The specificity of ARR > 10 (ng/dL)/(mU/L) after confirmatory test is 100%. After confirmatory test, patients with ARR > 10 (ng/dL)/(mU/L) had higher plasma aldosterone concentration and incidences of ischemic heart diseases and renal damage(p < 0.05). Conclusions After confirmatory tests, ARR > 10 (ng/dL)/(mU/L) indicates adrenal laterality, with increasingly cardiorenal damage in PA patients without APA. Thus, AVS should be recommended in these patients before surgery. Trial registration NCT03398785, Date of Registration: December 24, 2017.

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