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Antibiotics: Protein Synthesis, Nucleic Acid Synthesis and
Metabolism
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Principles and Definitions
• Selectivity– Selectivtytoxicity
• Therapeutic index– Toxic dose/ Effective dose
• Categories of antibiotics– Bacteriostatic
• Duration of treatment sufficient for host defenses
– Bactericidal• Usually antibiotic of choice
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Principles and Definitions
• Selectivity
• Therapeutic index• 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
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Principles and Definitions
• Antibiotic susceptibility testing (in vitro)– Minimum inhibitory concentration (MIC)
• Lowest concentration that results in inhibition of visible growth
– Minimum bactericidal concentration (MBC)• Lowest concentration that kills 99.9% of the original
inoculum
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Antibiotic Susceptibility Testing
8 4 02 1 Tetracycline (g/ml)
MIC = 2 g/ml
Determination of MIC
Chl Amp
Ery
Str
Tet
Disk Diffusion Test
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Zone diameter (mm) Approx. MIC( g/ml) for:Antimicrobial agent
(amt. per disk)and organism R I MS S R S
Ampicillin (10 g)
Enerobacteriacae 11 12-13 14 32 8
Haemophilus spp. 19 20 4 2
Enterococci 16 17 16
Tetracycline (30 g) 14 15-18 19 16 4
Zone Diameter Standards for Disk Diffusion Tests
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Principles and Definitions
• Combination therapy– Prevent emergence of resistant strains– Temporary treatment until diagnosis is made– Antibiotic synergism
• Penicillins and aminoglycosides
• CAUTION: Antibiotic antagonism– Penicillins and bacteriostatic antibiotics
• Antibiotics vs chemotherapeutic agents vs antimicrobials
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Antibiotics that Inhibit Protein Synthesis
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Review of Initiation of Protein Synthesis
30S1 32 GTP
1 2 3 GTP
Initiation Factors
mRNA
3
1
2 GTP
30S Initiation Complex
f-met-tRNA
Spectinomycin
Aminoglycosides
12
GDP + Pi 50S
70S Initiation Complex
AP
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Review of Elongation of Protein Synthesis
GTP
AP
Tu GTP Tu GDP
Ts
TsTu
+
GDPTs
Pi
P ATetracycline
AP
Erythromycin
Fusidic Acid
Chloramphenicol
G GTPG GDP + Pi
G
GDP
AP
+GTP
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Protein Synthesis
Microbe Library -American Society for Microbiology
www.microbelibrary.org
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Survey of Antibiotics
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Protein Synthesis Inhibitors
• Mostly bacteriostatic
• Selectivity due to differences in prokaryotic and eukaryotic ribosomes
• Some toxicity - eukaryotic 70S ribosomes
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Antimicrobials that Bind to the 30S Ribosomal Subunit
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Aminoglycosides (bactericidal)streptomycin, kanamycin, gentamicin, tobramycin,
amikacin, netilmicin, neomycin (topical)
• Mode of action - The aminoglycosides irreversibly bind to the 16S ribosomal RNA and freeze the 30S initiation complex (30S-mRNA-tRNA) so that no further initiation can occur. They also slow down protein synthesis that has already initiated and induce misreading of the mRNA. By binding to the 16 S r-RNA the aminoglycosides increase the affinity of the A site for t-RNA regardless of the anticodon specificity. May also destabilize bacterial membranes.
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Microbe Library
American Society for Microbiology
www.microbelibrary.org
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Aminoglycosides (bactericidal)streptomycin, kanamycin, gentamicin, tobramycin,
amikacin, netilmicin, neomycin (topical)
• Spectrum of Activity -Many gram-negative and some gram-positive bacteria; Not useful for anaerobic (oxygen required for uptake of antibiotic) or intracellular bacteria.
• Resistance - Common
• Synergy - The aminoglycosides synergize with -lactam antibiotics. The -lactams inhibit cell wall synthesis and thereby increase the permeability of the aminoglycosides.
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Tetracyclines (bacteriostatic)tetracycline, minocycline and doxycycline
• Mode of action - The tetracyclines reversibly bind to the 30S ribosome and inhibit binding of aminoacyl-t-RNA to the acceptor site on the 70S ribosome.
• Spectrum of activity - Broad spectrum; Useful against intracellular bacteria
• Resistance - Common
• Adverse effects - Destruction of normal intestinal flora resulting in increased secondary infections; staining and impairment of the structure of bone and teeth.
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Spectinomycin (bacteriostatic)
• Mode of action - Spectinomycin reversibly interferes with m-RNA interaction with the 30S ribosome. It is structurally similar to the aminoglycosides but does not cause misreading of mRNA.
• Spectrum of activity - Used in the treatment of penicillin-resistant Neisseria gonorrhoeae
• Resistance - Rare in Neisseria gonorrhoeae
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Antimicrobials that Bind to the 50S Ribosomal Subunit
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Chloramphenicol, Lincomycin, Clindamycin (bacteriostatic)
• Mode of action - These antimicrobials bind to the 50S ribosome and inhibit peptidyl transferase activity.
• Spectrum of activity - Chloramphenicol - Broad range;Lincomycin and clindamycin -
Restricted range
• Resistance - Common
• Adverse effects - Chloramphenicol is toxic (bone marrow suppression) but is used in the treatment of bacterial meningitis.
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Macrolides (bacteriostatic)erythromycin, clarithromycin, azithromycin, spiramycin
• Mode of action - The macrolides inhibit translocation.
• Spectrum of activity - Gram-positive bacteria, Mycoplasma, Legionella
• Resistance - Common
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Microbe Library
American Society for Microbiology
www.microbelibrary.org
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Antimicrobials that Interfere with Elongation Factors
Selectivity due to differences in prokaryotic and eukaryotic elongation factors
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Fusidic acid (bacteriostatic)
• Mode of action - Fusidic acid binds to elongation factor G (EF-G) and inhibits release of EF-G from the EF-G/GDP complex.
• Spectrum of activity - Gram-positive cocci
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Inhibitors of Nucleic Acid Synthesis
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Inhibitors of RNA Synthesis
Selectivity due to differences between prokaryotic and eukaryotic
RNA polymerase
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Rifampin, Rifamycin, Rifampicin, Rifabutin (bactericidal)
• Mode of action - These antimicrobials bind to DNA-dependent RNA polymerase and inhibit initiation of mRNA synthesis.
• Spectrum of activity - Broad spectrum but is used most commonly in the treatment of tuberculosis
• Resistance - Common
• Combination therapy - Since resistance is common, rifampin is usually used in combination therapy.
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Inhibitors of DNA Synthesis
Selectivity due to differences between prokaryotic and eukaryotic enzymes
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Quinolones (bactericidal)nalidixic acid, ciprofloxacin, ofloxacin, norfloxacin,
levofloxacin, lomefloxacin, sparfloxacin
• Mode of action - These antimicrobials bind to the A subunit of DNA gyrase (topoisomerase) and prevent supercoiling of DNA, thereby inhibiting DNA synthesis.
• Spectrum of activity - Gram-positive cocci and urinary tract infections
• Resistance - Common for nalidixic acid; developing for ciprofloxacin
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Antimetabolite Antimicrobials
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Inhibitors of Folic Acid Synthesis
• Basis of Selectivity
• Review of Folic Acid Metabolism
p-aminobenzoic acid + Pteridine
Dihydropteroic acid
Dihydrofolic acid
Tetrahydrofolic acid
Pteridine synthetase
Dihydrofolate synthetase
Dihydrofolate reductase
ThymidinePurines
Methionine
Trimethoprim
Sulfonamides
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Sulfonamides, Sulfones (bacteriostatic)
• Mode of action - These antimicrobials are analogues of para-aminobenzoic acid and competitively inhibit formation of dihydropteroic acid.
• Spectrum of activity - Broad range activity against gram-positive and gram-negative bacteria; used primarily in urinary tract and Nocardia infections.
• Resistance - Common
• Combination therapy - The sulfonamides are used in combination with trimethoprim; this combination blocks two distinct steps in folic acid metabolism and prevents the emergence of resistant strains.
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Trimethoprim, Methotrexate, Pyrimethamine (bacteriostatic)
• Mode of action - These antimicrobials binds to dihydrofolate reductase and inhibit formation of tetrahydrofolic acid.
• Spectrum of activity - Broad range activity against gram-positive and gram-negative bacteria; used primarily in urinary tract and Nocardia infections.
• Resistance - Common
• Combination therapy - These antimicrobials are used in combination with the sulfonamides; this combination blocks two distinct steps in folic acid metabolism and prevents the emergence of resistant strains.
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Anti-Mycobacterial Antibiotics
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Para-aminosalicylic acid (PSA) (bacteriostatic)
• Mode of action - Similar to sulfonamides
• Spectrum of activity - Specific for Mycobacterium tuberculosis
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Dapsone (bacteriostatic)
• Mode of action - Similar to sulfonamides
• Spectrum of activity - Used in treatment of leprosy (Mycobacterium leprae)
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Antimicrobial Drug ResistancePrinciples and Definitions
• Clinical resistance vs actual resistance
• Resistance can arise by mutation or by gene transfer (e.g. acquisition of a plasmid)
• Resistance provides a selective advantage
• Resistance can result from single or multiple steps
• Cross resistance vs multiple resistance– Cross resistance -- Single mechanism-- closely related
antibiotics– Multiple resistance -- Multiple mechanisms -- unrelated
antibiotics
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Antimicrobial Drug ResistanceMechanisms
• Altered permeability– Altered influx
• Gram negative bacteria
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Microbe Library
American Society for Microbiology
www.microbelibrary.org
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Antimicrobial Drug ResistanceMechanisms
• Altered permeability– Altered efflux
• tetracycline
Microbe Library
American Society for Microbiology
www.microbelibrary.org
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Antimicrobial Drug ResistanceMechanisms
• Inactivation -lactamase
– Chloramphenicol acetyl transferase
Microbe Library
American Society for Microbiology
www.microbelibrary.org
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Antimicrobial Drug ResistanceMechanisms
• Altered target site– Penicillin binding
proteins (penicillins)
– RNA polymerase (rifampin)
– 30S ribosome (streptomycin)
Microbe Library
American Society for Microbiology
www.microbelibrary.org
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Antimicrobial Drug ResistanceMechanisms
• Replacement of a sensitive pathway– Acquisition of a resistant
enzyme (sulfonamides, trimethoprim)