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Microbiological Assays of Antimicrobial Drugs

Learn about the assessment of antimicrobial sensitivity in vitro through microbiological assays. Explore different methods such as broth and agar dilution, disk diffusion, and microdilution. Understand the concept of Minimum Inhibitory Concentration (MIC) and its significance in determining antimicrobial effectiveness.

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Microbiological Assays of Antimicrobial Drugs

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  1. Lecture 6 Antimicrobial Chemotherapy MICROBIOLOGICAL ASSAYS of antimicrobial drugs (Assessment of anti-microbial Sensitivity in-vitro) Dr.: Abeer El-Sherbiny

  2. MICROBIOLOGICAL ASSAYS Microbiological Assay • refers to measurement of the activity of antibiotics on suitable test microorganism e.g. bacteria, yeast and molds under standard condition. • Microbiological assays are used to evaluate the in vitro activity of drug • The specific strains of test organism are used to evaluate the activity of the specific anti -microbial agent.

  3. Specific mediums are used for specific microbial testing, selecting the specific condition of the test. • Microbiological assay may be used to evaluate the agent quantitatively or qualitatively. • In qualitative evaluation, we consider test organism as sensitive, intermediate or resistant to the agent (disc diffusion test) • In quantitative evaluation in terms of concentration of the agent which inhibit the growth of micro organism the minimum inhibitory concentration (MIC) or that which kill microbes, the minimum bactericidal concentration (MBC). MEDIA: - • A substance use to provide nutrients for the growth and multiplication of micro Organism. • There are several types of media

  4. METHOD OF MICROBIOLOGICAL ASSAY: - • Following are the four micro biological methods: - • Macro dilution broth susceptibility test (MIC). • Agar dilution susceptibility test (MIC in agar plates). A standardized suspension of bacteria is inoculated onto a series of agar plates each containing a different concentration of antibiotic, encompassing the therapeutic range of the drug. • Disk diffusion susceptibility test • Micro dilution susceptibility test The microtube dilution procedure is similar in principle to the macrotube method, except that the susceptibility of microorganisms to antibiotics is determined in a series of microtube wells in a plastic plate, each plate may contain 80, 96, or more wells, depending on the number and concentration of antibiotics that are to be included in the susceptibility test panel. • The advantages of the microtube method are that small volumes of bacteria can be tested and inexpensively against a panel of antibiotics

  5. Common features of microbiological assayes a. The compound (antibiotic) being assayed must influence the growth of the test organism. b. A varying response in growth must be produced by addition of varying quantities of the test material. c. The growth medium must contain an excess of the compound required by the test organism for growth. d. The compound being assayed is the only growth promoting or inhibiting compound present.

  6. Minimum inhibitory concentration (MIC)assayAntibiotic sensitivity expressed in terms of MIC and or MBC gives quantitative data not obtainable with the disc diffusion method. These quantitative results are useful in certain purposes ( e.g. dosage calculation). In Broth Tube Dilution Method: Suspension of the test bacteria are incubated overnight with dilutions of antibiotics; the lowest concentration that inhibits growth is MIC.

  7. Minimal Inhibitory Concentration: is the lowest concentration of antimicrobial agent (antibiotic) that inhibits the growth of bacterium (No turbidity= no visible growth )Lowest concentration of subcultured broth with no growth on a plate after overnight incubation= Minimum Bactericidal Concentration

  8. Measuring Antimicrobial Sensitivity • Epsilomenter or ‘E’ test: • New techniques for direct determination of MIC • A gradual increasing concentration of antibiotic is fixed along plastic strip which applied to the surface of inoculated agar plate. • after overnight incubation  tear drop shaped inhibition zone is seen . • the zone edge intersects the graded test strip at the MIC of the antibiotics

  9. Phenol Coefficient • You will quantify the effectiveness of a disinfectant compared to the activity of a standard amount of phenol (Phenol Coefficient). The phenol coefficient test compares the antimicrobial activity of an antiseptic or disinfectant to that of phenol under standardized conditions. Phenol's ability to inhibit bacterial growth is assigned an arbitrary value of 1. Antiseptics and disinfectants are then assigned numbers relative to 1 and to each other. For example, a disinfectant with a phenol coefficient of 0.8 would be "better" than a disinfectant with a coefficient of 0.6, and neither would be as good as phenol.

  10. Measuring Antimicrobial Sensitivity Kirby-Bauer method for determining drug susceptibility (Disk Diffusion) • Bacteria spread on surface of agar plate • Disks, each with different antimicrobial drug, placed on agar plate • Incubated- drugs diffuse outward and kill susceptible bacteria • Zone of inhibition around each disk • Measure diameter of clear zones, then Compare size of zone to chart

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