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Epsilometer Test

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Dr. Samira Fattah
Dr. Samira Fattah

The Epsilometer test, also known as the E-test, is a quantitative method for determining the minimum inhibitory concentration (MIC) of antibiotics against bacteria. It uses plastic strips containing a continuous gradient of an antibiotic immobilized on one side to establish an antibiotic gradient when placed on an agar plate inoculated with bacteria. Elliptical zones of inhibition form where the antibiotic has inhibited bacterial growth, and the MIC is read as the intersection of this zone with the scale on the other side of the strip. The E-test allows for simple and rapid MIC testing of bacteria against various antibiotics.

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Epsilometer test Dr. Samira Fattah PhD in Medical Bacteriology College of Health Sciences-HMU
Epsilometer Test • Abbreviated as E-Test • Is a Quantitative method of antibiotic susceptibility testing. • Applies both dilution of antibiotics and diffusion of antibiotic in to the medium. • It is a very simple test to perform MIC of the bacterial isolate as compared to the other techniques like broth and agar dilution methods.
Epsilometer Test • The E-test is a plastic strip (5 x 50 mm; antibiotic carrier) with a continuous gradient of antibiotic immobilized on one side and MIC interpretative scale on the other side. • When transferred to the agar, the continuous antibiotic gradient established under the strip remains stable over a period covering the critical times of most microorganisms subjected to susceptibility testing.
Epsilometer Test E-Test can be classified to 4 categories: • Antibiotics • Antifungal • Anti-mycobacterial • Resistance phenotype testing
Purpose of E-Test 1. Determine the MIC of fastidious, slow-growing or nutritionally deficient micro-organisms ( like N. gonorrhoeae, Streptococcus pneumoniae and Haemophilius species), or for a specific type of patient or infection . 2. Detect low levels of resistance . 3. Confirm/ detect a specific antimicrobial resistance phenotype such as ESBL, MBL, and AmpC . 4. Test an antimicrobial not performed in routine use or a new, recently introduced antimicrobial agent.
Test requirements 1. Commercially available E-test strips 2. 0.5 McFarland standards, 3. sterile Mueller Hinton agar plates. In a 90 mm plate, a single antibiotic strip can be tested. In a 150 mm plate, at least 4 antibiotic strips can be tested. 4. Forceps 5. sterile swabs 6. bacterial suspension adjusted to 0.5 McFarland standards.
Method 1. The surface of agar plate inoculated with an adjusted bacterial suspension by swab in the same manner as a disk diffusion test. 2. Apply strips to agar surface using forceps (or E-test applicator if available). 3. Place the strip with the scale visible (i.e. facing upwards). Do not remove or replace a strip once it has touched the agar. 4. If strips stick together, they may be pulled apart by handling the section marked E. Do not touch any other area of the strip. 5. Incubate Plates at 37°C for 18-24 hrs. 6. After incubation, the interaction of the antimicrobial agent gradient and the test bacterial inoculum gives rise to elliptical inhibitory zones. 7. The results are read in the intersection of the ellipse with a MIC scale (in μg/ml on the strip).
Results and interpretation 1. Read plates after the recommended incubation period only if sufficient growth is seen and the inhibition eclipse is clearly visible. 2. Always read the end point at complete inhibition of all growth including hazes and isolated colonies. 3. Since E- test comprises a continuous gradient, MIC values in between two – fold dilutions can be obtained. 4. Always round up these values to the next two-fold dilution before interpretation. For example: If ampicillin breakpoints are given as S=1, I = 2, R=4 μg/ml, then an E-test MIC of 1.5 μg/ml is rounded up to 2 μg/ml and the category reported as Intermediate (I).
Problems with E-test reading Note: printed list is distributed to students

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Epsilometer Test

  • 1. Epsilometer test Dr. Samira Fattah PhD in Medical Bacteriology College of Health Sciences-HMU
  • 2. Epsilometer Test • Abbreviated as E-Test • Is a Quantitative method of antibiotic susceptibility testing. • Applies both dilution of antibiotics and diffusion of antibiotic in to the medium. • It is a very simple test to perform MIC of the bacterial isolate as compared to the other techniques like broth and agar dilution methods.
  • 3. Epsilometer Test • The E-test is a plastic strip (5 x 50 mm; antibiotic carrier) with a continuous gradient of antibiotic immobilized on one side and MIC interpretative scale on the other side. • When transferred to the agar, the continuous antibiotic gradient established under the strip remains stable over a period covering the critical times of most microorganisms subjected to susceptibility testing.
  • 4. Epsilometer Test E-Test can be classified to 4 categories: • Antibiotics • Antifungal • Anti-mycobacterial • Resistance phenotype testing
  • 5. Purpose of E-Test 1. Determine the MIC of fastidious, slow-growing or nutritionally deficient micro-organisms ( like N. gonorrhoeae, Streptococcus pneumoniae and Haemophilius species), or for a specific type of patient or infection . 2. Detect low levels of resistance . 3. Confirm/ detect a specific antimicrobial resistance phenotype such as ESBL, MBL, and AmpC . 4. Test an antimicrobial not performed in routine use or a new, recently introduced antimicrobial agent.
  • 6. Test requirements 1. Commercially available E-test strips 2. 0.5 McFarland standards, 3. sterile Mueller Hinton agar plates. In a 90 mm plate, a single antibiotic strip can be tested. In a 150 mm plate, at least 4 antibiotic strips can be tested. 4. Forceps 5. sterile swabs 6. bacterial suspension adjusted to 0.5 McFarland standards.
  • 7. Method 1. The surface of agar plate inoculated with an adjusted bacterial suspension by swab in the same manner as a disk diffusion test. 2. Apply strips to agar surface using forceps (or E-test applicator if available). 3. Place the strip with the scale visible (i.e. facing upwards). Do not remove or replace a strip once it has touched the agar. 4. If strips stick together, they may be pulled apart by handling the section marked E. Do not touch any other area of the strip. 5. Incubate Plates at 37°C for 18-24 hrs. 6. After incubation, the interaction of the antimicrobial agent gradient and the test bacterial inoculum gives rise to elliptical inhibitory zones. 7. The results are read in the intersection of the ellipse with a MIC scale (in μg/ml on the strip).
  • 8. Results and interpretation 1. Read plates after the recommended incubation period only if sufficient growth is seen and the inhibition eclipse is clearly visible. 2. Always read the end point at complete inhibition of all growth including hazes and isolated colonies. 3. Since E- test comprises a continuous gradient, MIC values in between two – fold dilutions can be obtained. 4. Always round up these values to the next two-fold dilution before interpretation. For example: If ampicillin breakpoints are given as S=1, I = 2, R=4 μg/ml, then an E-test MIC of 1.5 μg/ml is rounded up to 2 μg/ml and the category reported as Intermediate (I).
  • 9. Problems with E-test reading Note: printed list is distributed to students