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.

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 Comparative performances of Vitek-2, Disk Diffusion, and Broth Microdilution for antimicrobial susceptibility testing of canine Staphylococcus pseudintermedius

2021 ◽  
Author(s):  
Elisa Rampacci ◽  
Michele Trotta ◽  
Caterina Fani ◽  
Serenella Silvestri ◽  
Valentina Stefanetti ◽  
...  
Keyword(s):  
Antimicrobial Susceptibility ◽  
Susceptibility Testing ◽  
Antimicrobial Susceptibility Testing ◽  
Disk Diffusion ◽  
Health Concern ◽  
Broth Microdilution ◽  
Major Error ◽  
Staphylococcus Pseudintermedius ◽  
Cutaneous Infections ◽  
Vitek 2

Staphylococcus pseudintermedius is the primary cause of canine cutaneous infections and sporadically isolated as pathogen from humans. Rapidly emerging antibiotic-resistant strains are creating serious health concern so that accurate and timely antimicrobial susceptibility testing (AST) is crucial for patient care. Here, the performances of AST methods Vitek-2, Disk Diffusion (DD) and Broth Microdilution (BMD) were compared for the determination of susceptibility of 79 S. pseudintermedius isolates from canine cutaneous infections and one from human pyoderma to oxacillin (OXA), amoxicillin/clavulanate (AMC), cephalothin (CEF), gentamicin (GEN), enrofloxacin (ENR), doxycycline (DOX), clindamycin (CLI), inducible clindamycin resistance (ICR), mupirocin (MUP) and trimethoprim-sulfamethoxazole (SXT). Overall, the agreement of DD and Vitek-2 using veterinary AST-GP80 card with reference BMD was ≥ 90%, suggesting reliable AST performances. While DD generated mainly minor errors and one major error for OXA, Vitek-2 produced one very major error for GEN and it failed in identifying one ICR-positive isolate. Moreover, five bacteria were diagnosed as ICR-positive by Vitek-2 but they showed a non-induction resistance phenotype by manual methods. All S. pseudintermedius were interpreted as susceptible or intermediately susceptible to DOX using CLSI breakpoints for human staphylococci that match the DOX concentration range included in AST-GP80. However, this could lead to inappropriate antimicrobial prescription for S. pseudintermedius infections in companion animals. Considering the clinical and epidemiological importance of S. pseudintermedius , we encourage updating action by the system manufacturer to address AST for this bacterium.

scholarly journals Antimicrobial susceptibility testing of Enterobacteriaceae: determination of disk content and Kirby-Bauer breakpoint for ceftazidime/avibactam

2019 ◽  
Vol 19 (1) ◽  
Author(s):  
Xianggui Yang ◽  
Dan Wang ◽  
Qin Zhou ◽  
Fang Nie ◽  
Hongfei Du ◽  
...  
Keyword(s):  
Antibacterial Activity ◽  
Antimicrobial Susceptibility ◽  
Susceptibility Testing ◽  
Practical Method ◽  
Inhibition Zone ◽  
Antimicrobial Susceptibility Testing ◽  
Broth Microdilution ◽  
Inhibition Zone Diameter ◽  
Zone Diameter ◽  
Extended Spectrum

Abstract Background Detection of ceftazidime/avibactam (CAZ/AVI) antibacterial activity is absolutely vital with the rapid growth of carbapenem resistant Enterobacteriaceae (CRE). But now, there is no available automated antimicrobial susceptibility testing card for CAZ/AVI, so Kirby-Bauer has become an economical and practical method for detecting CAZ/AVI antibacterial activity against Enterobacteriaceae. Result In this study, antimicrobial susceptibility testing of CAZ/AVI against 386 Enterobacteriaceae (188 Klebsiella pneumoniae, 122 Escherichia coli, 76 Enterobacter cloacae) isolated from clinical patients was performed by broth microdilution. Of the 386 strains, 54 extended spectrum β lactamases negative (ESBL(−)), 104 extended spectrum β lactamases positive (ESBL(+)), 228 CRE. 287 isolates were susceptible to CAZ/AVI and 99 isolates were resistant to CAZ/AVI. At the same time, to obtain optimal content avibactam (AVI) disk containing ceftazidime (30 μg), inhibition zone diameter of four kinds of ceftazidime (30 μg) disk containing different AVI content (0 μg, 10 μg, 25 μg, 50 μg) were tested by Kirby-Bauer method. The microdilution broth method interpretation was used as the standard to estimate susceptible or resistance and then coherence analysis was carried out between Kirby-Bauer and broth microdilution. The result shows the inhibition zone diameter of 30 μg/50 μg disk, susceptible isolates: 20.5 mm–31.5 mm, resistance isolates: 8.25 mm–21.5 mm. The inhibition zone diameter of 30 μg/25 μg disk, susceptible isolates: 19.7 mm–31.3 mm, resistance isolates: 6.5 mm–19.2 mm. The inhibition zone diameter of 30 μg/10 μg disk, susceptible isolates: 19.5 mm–31 mm, resistance isolates: 6.5 mm–11 mm. The inhibition zone diameter of ceftazidime (30 μg), susceptible isolates: 6.5 mm–27.5 mm, resistance isolates 6.5 mm. Conclusion Our results show that 30 μg/50 μg, 30 μg/25 μg, 30 μg/10 μg CAZ/AVI disk have significant statistical differences to determinate CAZ/AVI antibacterial activity, but for 30 μg/50 μg disk, there has a cross section between susceptible isolates (minimum 20.5 mm) and resistance isolates (maximum 21.5 mm). For 30 μg/25 μg disk, it is hard to distinguish the difference between susceptible isolates (minimum 19.7 mm) and resistance isolates (maximum 19.2 mm), so 30 μg/10 μg CAZ/AVI disk is more conducive to determinate antibacterial activity.

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 Comparison of Agar Dilution, Disk Diffusion, MicroScan, and Vitek Antimicrobial Susceptibility Testing Methods to Broth Microdilution for Detection of Fluoroquinolone-Resistant Isolates of the FamilyEnterobacteriaceae

1999 ◽  
Vol 37 (3) ◽  
pp. 544-547 ◽  
Author(s):  
Christine D. Steward ◽  
Sheila A. Stocker ◽  
Jana M. Swenson ◽  
Caroline M. O’Hara ◽  
Jonathan R. Edwards ◽  
...  
Keyword(s):  
Antimicrobial Susceptibility ◽  
Susceptibility Testing ◽  
Error Rates ◽  
Disk Diffusion ◽  
Fluoroquinolone Resistance ◽  
Broth Microdilution ◽  
Major Error ◽  
Testing Methods ◽  
Minor Error ◽  
Agar Dilution

Fluoroquinolone resistance appears to be increasing in many species of bacteria, particularly in those causing nosocomial infections. However, the accuracy of some antimicrobial susceptibility testing methods for detecting fluoroquinolone resistance remains uncertain. Therefore, we compared the accuracy of the results of agar dilution, disk diffusion, MicroScan Walk Away Neg Combo 15 conventional panels, and Vitek GNS-F7 cards to the accuracy of the results of the broth microdilution reference method for detection of ciprofloxacin and ofloxacin resistance in 195 clinical isolates of the familyEnterobacteriaceae collected from six U.S. hospitals for a national surveillance project (Project ICARE [Intensive Care Antimicrobial Resistance Epidemiology]). For ciprofloxacin, very major error rates were 0% (disk diffusion and MicroScan), 0.9% (agar dilution), and 2.7% (Vitek), while major error rates ranged from 0% (agar dilution) to 3.7% (MicroScan and Vitek). Minor error rates ranged from 12.3% (agar dilution) to 20.5% (MicroScan). For ofloxacin, no very major errors were observed, and major errors were noted only with MicroScan (3.7% major error rate). Minor error rates ranged from 8.2% (agar dilution) to 18.5% (Vitek). Minor errors for all methods were substantially reduced when results with MICs within ±1 dilution of the broth microdilution reference MIC were excluded from analysis. However, the high number of minor errors by all test systems remains a concern.

Evaluation of standardized automated rapid antimicrobial susceptibility testing of Enterobacterales-containing blood cultures: a proof-of-principle study

2020 ◽  
Vol 75 (11) ◽  
pp. 3218-3229
Author(s):  
Stefano Mancini ◽  
Elias Bodendoerfer ◽  
Natalia Kolensnik-Goldmann ◽  
Sebastian Herren ◽  
Kim Röthlin ◽  
...  
Keyword(s):  
Standard Method ◽  
Antimicrobial Susceptibility ◽  
Susceptibility Testing ◽  
Bloodstream Infections ◽  
Inhibition Zone ◽  
Blood Cultures ◽  
Antimicrobial Susceptibility Testing ◽  
Disc Diffusion ◽  
Inhibition Zones ◽  
Reading System

Abstract Background Rapid antimicrobial susceptibility testing (RAST) of bacteria causing bloodstream infections is critical for implementation of appropriate antibiotic regimens. Objectives We have established a procedure to prepare standardized bacterial inocula for Enterobacterales-containing clinical blood cultures and assessed antimicrobial susceptibility testing (AST) data generated with the WASPLabTM automated reading system. Methods A total of 258 blood cultures containing Enterobacterales were examined. Bacteria were enumerated by flow cytometry using the UF-4000 system and adjusted to an inoculum of 106 cfu/mL. Disc diffusion plates were automatically streaked, incubated for 6, 8 and 18 h and imaged using the fully automated WASPLabTM system. Growth inhibition zones were compared with those obtained with inocula prepared from primary subcultures following the EUCAST standard method. Due to time-dependent variations of the inhibition zone diameters, early AST readings were interpreted using time-adjusted tentative breakpoints and areas of technical uncertainty. Results and conclusions Inhibition zones obtained after 18 h incubation using an inoculum of 106 cfu/mL prepared directly from blood cultures were highly concordant with those of the EUCAST standard method based on primary subcultures, with categorical agreement (CA) of 95.8%. After 6 and 8 h incubation, 89.5% and 93.0% of the isolates produced interpretable results, respectively, with CA of >98.5% and very low numbers of clinical categorization errors for both the 6 h and 8 h readings. Overall, with the standardized and automated RAST method, consistent AST data from blood cultures containing Enterobacterales can be generated after 6–8 h of incubation and subsequently confirmed by standard reading of the same plate after 18 h.

scholarly journals Detection of Colistin Resistance in Carbapenem Resistant Enterobacteriaceae by Reference Broth Microdilution and Comparative Evaluation of Three Other Methods

2021 ◽  
Author(s):  
Punyatoya Kar ◽  
Bijayini Behera ◽  
Srujana Mohanty ◽  
Jayanti Jena ◽  
Ashoka Mahapatra
Keyword(s):  
Susceptibility Testing ◽  
Good Alternative ◽  
Broth Microdilution ◽  
Major Error ◽  
Colistin Resistance ◽  
Carbapenem Resistant ◽  
Agar Dilution ◽  
Moderate Sensitivity ◽  
Phenotypic Methods ◽  
User Friendly

Abstract Objective Challenges in susceptibility testing of colistin along with increase in the prevalence of colistin-resistant carbapenemase-producing Enterobacteriaceae (CRE) pathogens needs addressal. Evaluation of user-friendly methods is necessary as an alternative to broth microdilution (BMD), the reference susceptibility testing method, for routine implementation in diagnostic clinical microbiology laboratories. Genotypic detection of the plasmid-mediated colistin resistance is also needed for infection control purposes. Materials and Methods Colistin susceptibility of 200 nonduplicate clinical CRE isolates from December 2017 to June 2019 was determined by BMD, agar dilution (AD), E test, and rapid polymyxin NP test and interpreted as per the European Committee on Antimicrobial Susceptibility Testing. The results of AD, E test, and NP test were compared with that of BMD, considering minimal inhibitory concentration (MIC) ≤ 2 µg/mL as susceptible and > 2 µg/mL as resistant. Presence of any plasmid-mediated colistin resistance (mcr-1 and 2) was evaluated in 27 colistin-resistant CRE isolates by polymerase chain reaction. Statistical Analysis Performance of different phenotypic methods was analyzed by comparing MIC results of AD and E test with that of reference BMD method. Agreement between BMD and the other two methods was expressed in terms of categorical agreement and essential agreement. Errors were expressed as very major error (VME: false-susceptible) and major error (ME: false-resistance) by AD/E test. VME and ME of 3% disagreement were considered unacceptable. Results Colistin resistance was found in 27 (13.5%) isolates by BMD method. The VME rates of both AD (11%) and E test (37%) could not meet the Clinical and Laboratory Standards Institute recommendation (< 3% VME rate is acceptable) as alternative tests to the reference BMD. Colistin NP test showed sensitivity and specificity of 85% and 98%, respectively. The percentage discordant result in NP test was highest in Enterobacter spp. (17%). None of the 27 colistin resistant isolates showed presence of mcr-1 and mcr-2 genes. Conclusion High VME rate in AD and E tests precludes their use as alternatives to BMD for colistin susceptibility testing. NP test with moderate sensitivity but excellent specificity can be a good alternative for testing colistin susceptibility in CRE isolates, except in Enterobacter spp. Absence of mcr-1 and mcr-2 gene necessitates the exploration of other mechanisms of colistin resistance.

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