1. Antibiotics Biotechnology II

2. Univ S. Carolina Antibiotics Disrupt Cell Wall Synthesis, Protein Synthesis, Nucleic Acid Synthesis and Metabolism

3. Principles and Definitions Selectivity Selectivity –Selectivity vs toxicity Therapeutic index Therapeutic index –Toxic dose/ Effective dose Categories of antibiotics Categories of antibiotics –Bacteriostatic  Reversibly inhibit growth  Duration of treatment sufficient for host defenses to eradicate infection –Bactericidal-  Kill bacteria  Usually antibiotic of choice for infections in sites such as endocardium or the meninges where host defenses are ineffective.

4. Principles and Definitions Selectivity Selectivity Therapeutic index Therapeutic index Categories of antibiotics Categories of antibiotics –Use of bacteriostatic vs bactericidal antibiotic  Therapeutic index better for bacteriostatic antibiotic  Resistance to bactericidal antibiotic  Protein toxin mediates disease – use bacteriostatic protein synthesis inhibitor to immediately block synthesis of toxin.

5. Principles and Definitions Antibiotic susceptibility testing (in vitro) Antibiotic susceptibility testing (in vitro) –Bacteriostatic Antibiotics  Minimum inhibitory concentration (MIC)  Lowest concentration that results in inhibition of visible growth (colonies on a plate or turbidity of liquid culture) –Bactericidal Antibiotics  Minimum bactericidal concentration (MBC)  Lowest concentration that kills 99.9% of the original inoculum

6. Antibiotic Susceptibility Testing-MIC 8 4021 Tetracycline (  g/ml) MIC = 2  g/ml Determination of MIC ChlAmp Ery Str Tet Disk Diffusion Test Size of zone of inhibition depends on sensitivity, solubility, rate of diffusion. Compare results to MIC tables generated using standards.

7. Principles and Definitions Combination therapy Combination therapy –Prevent emergence of resistant strains –Temporary treatment until diagnosis is made –Take advantage of antibiotic synergism  Penicillins and aminoglycosides inhibit cell wall synthesis and allow aminoglycosides to enter the bacterium and inhibit protein synthesis.  CAUTION: Antibiotic antagonism –Penicillins and bacteriostatic antibiotics. Cell wall synthesis is not occurring in cells that are not growing. Antibiotics vs chemotherapeutic agents vs antimicrobials Antibiotics vs chemotherapeutic agents vs antimicrobials –Antibiotics-naturally occurring materials –Chemotherapeutic-synthesized in the lab (most antibiotics are now synthesized and are therefore actually chemotherapeutic agents.

8. Mechanisms of Antibiotics Inhibition of Protein Synthesis Mostly bacteriostatic Mostly bacteriostatic Selectivity due to differences in prokaryotic and Selectivity due to differences in prokaryotic and eukaryotic ribosomes eukaryotic ribosomes Some toxicity - 70S ribosomes eukaryotic in mitochondria Some toxicity - 70S ribosomes eukaryotic in mitochondria Inhibitors of INITATION 30S Ribosomal Subunit (Aminoglycosides, Tetracyclines, Spectinomycin) 50S Ribosomal Subunit (Chloramphenicol, Macrolides) Inhibitors of ELONGATION Elongation Factor G (Fusidic acid)

9. Mechanisms of Antibiotics Inhibitors of Nucleic Acid Synthesis Inhibitors of RNA Synthesis: Selectivity due to differences between prokaryotic and eukaryotic RNA polymerase: (Rifampcin) Quinolones)Inhibitors of DNA Synthesis: Selectivity due to differences between prokaryotic and eukaryotic enzymes: (Quinolones) Inhibitors of Folic Acid Synthesis :bacteria make it, we get it from diet (Sulfonamides)

10. Antimicrobial Drug Resistance Principles and Definitions Clinical resistance vs actual resistance Clinical resistance vs actual resistance Resistance can arise by new mutation or by gene transfer (e.g. acquisition of a plasmid) Resistance can arise by new mutation or by gene transfer (e.g. acquisition of a plasmid) Resistance provides a selective advantage. Resistance provides a selective advantage. Resistance can result from single or multiple steps Resistance can result from single or multiple steps Cross resistance vs multiple resistance Cross resistance vs multiple resistance –Cross resistance -- Single mechanism-- closely related antibiotics are rendered ineffective –Multiple resistance -- Multiple mechanisms -- unrelated antibiotics. Acquire multiple plasmids. Big clinical problem.

11. Antimicrobial Drug Resistance Mechanisms Altered permeability Altered permeability –Altered influx  Mutation in a transporter necessary to import antibiotic can lead to resistance. –Altered efflux  Acquire transporter gene that will pump the antibiotic out (Tetracycline) Inactivation of the antibiotic Inactivation of the antibiotic  -lactamase Chloramphenicol Acetyl Transferase Mutation in the target site. Mutation in the target site. –Penicillin binding proteins (penicillins) –RNA polymerase (rifampin) –30S ribosome (streptomycin) Replacement of a sensitive enzyme with a resistant enzyme Replacement of a sensitive enzyme with a resistant enzyme –Plasmid mediated acquisition of a resistant enzyme (sulfonamides, trimethoprim)