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ANTIMICROBIAL AGENTS
• Fundamental principle of chemotherapy is the use of chemicals to treat
disease.
• Antimicrobials are chemotherapeutic agents with selective toxicity to the
pathogen; these substances must be harmful to microbial parasites
(virus, bacterium, fungus, protist or helminth “worm”) but not host cells
• We must distinguish antimicrobials from disinfectants or antiseptics
which can harm both parasite & host cells;
• Disinfectants and antiseptics are used to inactivate microbes on
inanimate environment and skin surface, respectively.
• Antibiotics are antimicrobials produced naturally by bacteria or fungi.
• Others are manmade (synthetic): sulfonamides, isoniazid, quinolones.
Antibiotics
• Sources of natural antimicrobial agents:
– spore-forming bacillus - produce polypeptide antibiotic
– sporulating fungi - produce penicillins & cephalosporins
– spore-forming Streptomyces - produce other major classes of
antibiotics
• In nature antibiotics are:
– 1. produced only by microorganisms that are forming spores
– 2. produced only after most cellular growth has occurred.
– 3. excreted in very small amount.
• Antibiotics have one of two effects on the growth and
viability of microorgansms:
– bacteriostatic - slows growth or prevents multiplication; not the
cure per se; cure results from combined action of drug and host's
defense mechanisms, like phagocytosis
– bacteriocidal - usually effective only for growing microbes;
ineffective dormant cells
Major Classes of Antibiotics
Penicillins
produced by mold Penicillium
Cephalosporins -
produced by mold Cephalosporium
Bacitracin -
produced by Bacillus licheniformis
Polymyxins -
produced by Bacillus polymyxa
Aminoglycosides -
produced by Streptomyces griseus
Tetracyclines
produced by Streptomyces
Chloramphenicol -
produced by Streptomyces
venezuelae
Macrolides - (Erythromycin)
produced by Streptomyces erythreus
Rifamycins -
produced by Streptomyces
mediterrani
MECHANISMS OF ANTIBIOTIC
ACTIVITY
1. inhibit cell wall synthesis
2. inhibit cell membrane function
3. inhibit protein synthesis
4. inhibit nucleic acid synthesis
5. competitive inhibition of enzyme activity
What is meant by narrow versus broad spectrum?
Inhibition of Cell Wall Synthesis
• Penicillins:
* Penicillin prevents formation of final peptide bond on
sidechains of PDG; penicillins react irreversibly with crosslinking enzymes.
* Kill growing bacterial cells; high internal osmotic pressure
causes cell lysis at defective wall sites.
* Toxicity to humans - has least of any antimicrobial drug for
host cells most serious side-effect is allergic reactions.
* Both natural and semi-synthetic forms. Common β-lactam
ring nucleus for all penicillins; side groups vary.
Natural Penicillin Structures
Naturals are very narrow spectrum and susceptible to penicillinases,
which cleave the β-lactam ring rendering the drug inactive.
Penicillin V is preferred for oral administration as it is resistant to acid
hydrolysis.
Semi-synthetic penicillin’s are designed to:
* increase range of action to include effectiveness against Gramnegative bacteria (e.g. ampicillin); or
* resistance to penicillinases (e.g. methicillin).
Inhibition of Cell Wall Synthesis
• Cephalosporins:
1. Considered the second generation of ß-lactams; i.e. works like
pennicillin
2. Possess moderate-spectrum activity against gram positive & gram
negative bacteria;
3. Resistant to penicillinases, sensitive to cephalosporinases;
4. No allergic reactions in most penicillin-allergic people.
• Bacitracin:
1. Blocks dephosphorylation of lipid carrier which transports new wall
components thru membrane; prevents peptidoglycan polymer
synthesis.
2. Toxic effect on kidney when given internally; it is used mostly as a
topical application.
Cell Membrane Injury
• Polymyxin B:
* Act as cationic detergents; integrate within & disrupt
outer membrane.
* Causes loss of osmotic function & selective membrane
permeability.
* Unique in being bacteriocidal in absence of cell growth.
* More effective against gram negatives than gram
positives (due to more LPS in gram negatives)
* Toxicity - damages kidneys; little clinical use except
topically (neomycin is the combination with bacitracin).
Protein Synthesis Inhibition:
Macrolides
Aminoglycosides
Aminoglycosides (streptomycin):
• Binds to the 30S subunit causing a conformation change.
• Broad-spectrum; Bactericidal;
• Results in a misreading of the mRNA;
- Wrong amino acids are inserted in peptide chain & faulty
proteins are produced;
- Faulty membrane proteins may create membrane channels
that permit influx of more antibiotic.
Tetracyclines:
• Binds to 30S; blocks binding of tRNA to mRNA-30S complex.
• New amino acids cannot be added to peptide so protein synthesis
stops.
• Broad-spectrum; inhibits only rapidly multiplying bacteria (bacteristatic).
• Problems with tetracyclines are:
1. inhibition of normal flora leads to "superinfection"
2. deposition in calcified tissues (bones & teeth) causing staining,
3. weakening of bone structure (esp. true in growing children);
4. photosensitivity in some hosts.
Chloramphenicol:
• Attaches to 50S; inhibits the enzyme that catalyzes
peptide bond formation. New amino acids cannot be
added and protein synthesis stops.
• Broad-spectrum; Bacteristatic
• Toxicity - high doses tend to cause abnormalities of early
RBC development leading to anemia.
Macrolides (Erythromycin):
• Attaches to 50S; Blocks release of uncharged tRNA from
P site preventing ribosome movement (= translocation)
• Primarily bacteristatic;
• Less effective than penicillins, yet good alternative in
cases of penicillin allergy.
• Problem - numbers of resistant mutants arise with use.
Inhibition of Nucleic Acid Synthesis
Rifamycins:
• Inhibit transcription; binds to prokaryote RNA polymerase.
Rifampin blocks RNA chain elongation.
Inactive on RNA polymerase from eucaryotes.
• Toxicity - occasional rashes, platelet decrease & some decline in
liver function may occur.
• Imparts an orange color to urine and sweat.
Quinolones:
• Binds and inhibits activity of DNA gyrase (enzyme that nicks,
untwists & reseals dsDNA at the replication fork)
• Quinolones prevent resealing of nicked DNA allowing degradation
by DNAses.
• Quinolones are used primarily for urinary tract infections.
Competitive Inhibitors of Enzymes
“Metabolic Antagonists”
Sulfonamides:
• p-aminobenzoic acid (PABA) is the substrate in
the essential synthesis of folic acid in most
bacteria.
• Sulfonamides are structural analogs of PABA; act
as competitive inhibitors.
• No effect on human cells; we cannot synthesize
folic acid, but rather obtain folic acid in diet.
ANTIMICROBIAL RESISTANCE
Natural Resistance:
• Many bacteria will be naturally resistant to certain narrow
spectrum antimicrobials:
– They simply do not possess the target, or
– the drug does not have access to the target of its action.
• For unknown infections it may be a good initial strategy
to use a broad spectrum drug.
– However, this will also cause death of normal microbiota.
– This could lead to secondary infection by another naturally
resistant microbe.
• E.g.: fungal yeast infections (e.g. Candida albicans). A
drug that may be broad spectrum for bacteria my have
no effect on fungal yeast, hence the yeast is considered
naturally resistant.
Acquired Resistance Mechanisms:
1. Acquire an enzyme that destroys the drug (e.g. β-lactamase a
penicillinase).
2. Changing the target compound the drug acts on so that no longer
happens (e.g. Streptococcus pnuemoniae mutated penicillin binding
protein)
3. Actively eliminating the drug so it does not accumulate, via membrane
pumps or change in the drugs permeability through a porin. (e.g.
Neisseria gonorrhoeae porin change prevents penicillin entry).
Genetics:
• Slowly, mutations that increase resistance are selected and the numbers
of these strains becomes greater than sensitive strains in the presence
of antibiotic.
• Rapid transfer via a conjugative plasmid or transduction of a transposon
carrying resistance genes.
Slowing Resistance:
• cut unnecessary use in medicine and agriculture.
• use high dose for long duration to prevent re-growth of
resistant mutants.
• combined therapy (two is better than one).
Testing for Resistance/Sensitivity
• Kirby Bauer Assay (KBA); easy and rapid.
• KBA interpretation is based on the Minimal
Inhibitory Concentration (MIC).