Download Antibiotic Placement

“Chance Favors The Prepared Mind”
-Alexander Fleming (1928)
Penicillium notatum
Cephalosporin
Streptomyacin
Bacitracin
Cell Wall Disruption
Antibiotic
Producer organism
Activity
Site or mode of action
Penicillin
Penicillium chrysogenum
Gram-positive bacteria
Wall synthesis
Cephalosporin
Cephalosporium acremonium
Broad spectrum
Wall synthesis
Griseofulvin
Penicillium griseofulvum
Dermatophytic fungi
Microtubules
Bacitracin
Bacillus subtilis
Gram-positive bacteria
Wall synthesis
Polymyxin B
Bacillus polymyxa
Gram-negative bacteria
Cell membrane
Amphotericin B
Streptomyces nodosus
Fungi
Cell membrane
Erythromycin
Streptomyces erythreus
Gram-positive bacteria
Protein synthesis
Neomycin
Streptomyces fradiae
Broad spectrum
Protein synthesis
Streptomycin
Streptomyces griseus
Gram-negative bacteria
Protein synthesis
Tetracycline
Streptomyces rimosus
Broad spectrum
Protein synthesis
Vancomycin
Streptomyces orientalis
Gram-positive bacteria
Protein synthesis
Gentamicin
Micromonospora purpurea
Broad spectrum
Protein synthesis
Rifamycin
Streptomyces mediterranei
Tuberculosis
Protein synthesis
Antibiotic Explosion or Antibiotic Exploitation?
• In last 10 years, use
of antibiotics has
risen from 6 million
lbs/yr to 50 million
• 90% goes to raise the
U.S. 1.3 billion cattle
Ubiquity of Antibiotic Use
How Do Drug Resistant
Bacteria Evolve?
The Solution Is The Problem
• A stronger antibiotic
without a change in
its use will continue to
promote growing
antibiotic resistance
How Do We Determine Which Strains
Are Susceptible To Which Antibiotics?
Kirby-Bauer Antibiotic Sensitivity Test
Step 1: Bacterial Lawn
• Petri dish w/nutrient
agar is streaked with
bacterial culture to
produce a “lawn” of
growth
• Make sure that the
entire dish gets
streaked so that an
even lawn is
produced (not
isolated plaques)
Step 2: Antibiotic Placement
• Various discs
impregnated with
antibiotics are placed
on the bacterial “lawn
• The antibiotics will
diffuse out of the
discs and onto the
agar/”lawn”
Step 3: Incubation
• Incubate the bacteria
for 24 hrs at 37 C
• If the bacteria are
resistant, they will grow
over the antibiotic discs
• If the bacterial are
susceptible to the
antibiotics diffusing out
of the discs, a zone of
inhibition (no growth)
will be produced
Measuring Zones of Inhibition
• Measure the diameter
of the zone of
inhibition
• Subtract the diameter
of the disc from the
total distance
• If zone of inhibition is
irregularly shaped,
use average size or
radius x 2
Interpreting Sensitivity
Sensitivity
Most = Largest Zone of Inhibition
Least = Smallest Zone of Inhibition
Mutation & Resistance
Antibiotic Spectrum
• Broad Spectrum
– Has zones of
inhibition for many
bacterial strains
• Narrow Spectrum
– Has large zone of
inhibition but for
only 1-2 bacterial
strains
Lab Protocol
• 1) Each group will receive
– 2 LB Agar Plates
– 1 Strain of Monera
– Cotton Swabs
– 10 Antibiotic Discs
• 2) Dip a cotton swab into the bacterial
broth/gel and swirl.
• 3) Innoculate the first plate with your
assigned Moneran strain. Repeat step 2
with a fresh swab and the second plate
Protocol: Placing The Discs
• Once both plates have been inoculated
with your assigned Monera, add 4 to 5
antibiotic discs from the dispensers to
each dish. Use a tweezers or probe to
transfer them to the plate.
• Making sure that the discs have adhered,
invert the plate
• Label your plates with your group, your
period AND which strain of Monera it is
• Incubate at 37 C